Journal articles on the topic 'CML, TKI, AOEs, cytokines'
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1
Sicuranza, Anna, Ilaria Ferrigno, Elisabetta Abruzzese, Alessandra Iurlo, Sara Galimberti, Antonella Gozzini, Luigiana Luciano, et al. "Pro-Inflammatory and Pro-Oxidative Changes during Nilotinib Treatment in CML Patients: Results of a Prospective Multicenter Front-Line TKIs Study (KIARO Study)." Blood 138, Supplement 1 (November 5, 2021): 1479. http://dx.doi.org/10.1182/blood-2021-152530.
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Abstract Background: Tyrosine kinase inhibitors (TKI) may offer a normal life expectancy to Chronic Myeloid Leukemia (CML) patients. However, during treatment with nilotinib, a higher than expected incidence of arterial occlusive events (AOEs) was observed. We retrospectively showed an "inflammatory status" during nilotinib treatment that may explain this increased incidence of AOEs. Here, we report results of a prospective multicenter (KIARO) study including 186 CML patients (89 imatinib, 59 nilotinib, 38 dasatinib) in which pro/anti-inflammatory cytokines were measured at diagnosis and during treatment, with the aim to investigate potential changes in each patient's inflammatory status possibly favoring AOEs. Aims: The aims of this study are: 1) to analyze prospectively inflammation status during TKI treatment; 2) to record AOEs; 3) to calculate the SCORE and evaluate its predictive role for AOEs; 4) to analyze possible associations of AOEs with altered inflammation status. Methods: Inflammatory status was evaluated by measuring IL6, IL10, TNFα, oxLDL and hs-CRP plasma levels at diagnosis and during treatment (+1, +3, +6, +12 months); additionally, clinical and biochemical pro-atherothrombotic profiles and the 10-year SCORE chart were also evaluated. Results: 186 newly-diagnosed CML patients starting either imatinib, nilotinib or dasatinib treatment, entered this study. Regarding the inflammation status, we observed that TNFα and IL6 levels were high at diagnosis and decreased during the first 12 months of treatment regardless the type of TKI; instead, IL10 levels were comparable among the 3 TKI cohorts at baseline, but showed a remarkably different evolution during treatment. In fact, IL10 levels were significantly higher after 6 and 12 months of imatinib (p=0.012 and p=0.009, respectively) and dasatinib (p=0.032 and p=0.014, respectively) compared to nilotinib. Consequently, TNFα/IL10 ratio was significantly higher in nilotinib cohort at 6 and 12 months respect to imatinib (p=0.044 at 6 months and p=0.041 at 12 months) and dasatinib (p=0.040 at 6 months and p=0.044 at 12 months). As well, IL6/IL10 ratio was significantly higher in nilotinib cohort compared to imatinib and dasatinib both at 6 (p=0.042 and p=0.049, respectively) and 12 months (p=0.040 and p=0.041, respectively) (Figure 1). OxLDL levels were similar in the 3 groups for the first 6 months. At 12 months we detected a significant increase of oxLDL levels in the nilotinib cohort (p=0.041), respect to imatinib and dasatinib. We did not find significant differences in hs-CRP levels across the 3 TKIs, although a trend for higher levels was observed in nilotinib cohort. Overall, these results suggest a TKI-driven pro-inflammatory status in nilotinib treated patients. After a median follow-up of 23.3 months of TKI treatment, 10 patients (5.4%) suffered an AOE, specifically: 6 ACS, 2 PAOD, 1 TIA and 1 stroke. 5 events (50%) occurred in patients treated with nilotinib, either in first line (4 patients) or in third line (1 patient, after failure following brief treatment with imatinib and dasatinib). In this subgroup of 10 patients experiencing an AOE, we observed a trend of increased IL6 and TNFα median values both at diagnosis and at each time point, compared with the remaining no-AOE patients. IL10 and oxLDL had similar median concentrations in both AOE and no-AOE cohorts, except for oxLDL at 12 months which resulted higher in patients who experienced AOEs. Moreover, regarding AOEs, nilotinib treatment showed a 3.1 times increased risk compared to other TKIs (HR 3.1, 95% CI 2.6-4.4 p< 0.001), whereas 10-year SCORE was not predictive in the whole cohort (HR 0.6, 95% CI 0.33-0.94 p= 0.094) or in any subgroup (imatinib HR 0.8, 95% CI 0.49-1.03 p= 0.067; nilotinib HR 0.4, 95% CI 0.29-0.76 p= 0.112, dasatinib HR 0.6, 95% CI 0.37-0.92 p= 0.082). Conclusions: Our results showed a pro-inflammatory/oxidative milieu increasing along treatment with nilotinib compared with imatinib or dasatinib, as demonstrated by higher IL6/IL10 and TNFα/IL10 ratios, higher levels of oxLDL and a trend for higher hs-CRP only in nilotinib cohort. However, due to the low number of observed events, a formal statistical analysis for any association between AOEs and pro/anti-inflammatory cytokines levels was not possible. Therefore, a longer follow-up is needed to further confirm the active role of nilotinib in AOEs pathogenesis. Figure 1 Figure 1. Disclosures Abruzzese: Incyte: Consultancy, Honoraria; Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Bristol Myers Squibb: Consultancy, Honoraria. Galimberti: Incyte: Speakers Bureau; AbbVie, Janssen: Honoraria, Other: Travel grants. Stagno: Pfizer: Consultancy, Honoraria, Other: Support for attending meetings and/or travel; InCyte: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Other: Support for attending meetings and/or travel, Research Funding.
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Levy, Moshe Yair, Lin Xie, Yuexi Wang, Frank Neumann, Shouryadeep Srivastava, Daniel Naranjo, Qisu Zhang, and Mehul Dalal. "Real-World Comparisons of Cardiovascular Events between Different Tyrosine Kinase Inhibitors Among Patients with Chronic Myeloid Leukemia." Blood 132, Supplement 1 (November 29, 2018): 3567. http://dx.doi.org/10.1182/blood-2018-99-113490.
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Abstract INTRODUCTION: Chronic myeloid leukemia (CML) is a bone marrow and blood disorder accounting for 15% of adult leukemia. A shift in CML management has occurred over the past decade with the introduction of tyrosine kinase inhibitors (TKIs), changing CML status from fatal to a chronic, lifelong illness. However, an association between TKI use and cardiovascular events has been observed. This study aimed to compare major adverse cardiac events (MACE), arterial occlusive events (AOEs), and venous thrombotic events (VTEs) among CML patients in chronic phase (CP-CML) treated with different TKIs. METHODS: A retrospective observational study of adult (aged ≥18 years) CP-CML patients prescribed a TKI was conducted using the IBM® MarketScan® Research Databases from July 2012-June 2017. The index date was defined as the index drug prescription date, identified based on TKI use during the identification period (January 2013-December 2016) in hierarchical order: ponatinib, bosutinib without ponatinib, and other TKIs (imatinib, dasatinib, nilotinib) excluding ponatinib and bosutinib. Patients were required to have continuous health plan enrollment for ≥6 months pre-index date (baseline period) and at least 6 months post-index date (follow-up period). Patients with use of one or two previous TKI(s) before the index date were examined separately. Cardiovascular events occurring through the earliest of discontinuation of index TKI, switch to another TKI, or end of follow-up period using ICD-9/10-CM diagnosis codes were calculated as the number of events per 100 person-years. MACE was defined as a composite of stroke (hemorrhagic stroke and ischemic stroke), myocardial infarction, and inpatient death; AOEs included cardiovascular, cerebrovascular, and peripheral vascular events; VTEs included pulmonary embolism and deep vein thrombosis. RESULTS: After applying the selection criteria, 161 patients had one previous use of a TKI, with 50 ponatinib, 80 bosutinib and 31 other TKI patients. Mean ages were 54, 57, and 58 years for ponatinib, bosutinib, and other TKI cohorts, respectively. Most ponatinib patients initiated treatment with a dose of 45 mg (60%); most bosutinib patients initiated with a dose of 500 mg (53%) . For patients with use of one previous TKI, the average Charlson Comorbidity Index score was 1.4 for ponatinib, 1.8 for bosutinib and 0.8 for other TKI patients. Common baseline comorbid conditions by drug included anemia (ponatinib: 50%; bosutinib 31%; other TKI: 19%), hypertension (ponatinib: 32%, bosutinib: 43%, other TKI: 29%), and diabetes (ponatinib: 16%; bosutinib: 28%; other TKI: 10%). CP-CML patients were observed to have cardiovascular events prior to index TKI use, especially MACE (ponatinib: 4%, bosutinib: 16%, other TKI: 3%), and AOEs (ponatinib: 12%; bosutinib: 25%; other TKI: 19%). During the follow-up period, no significant differences were found for cardiovascular events across patients with TKI use (Table 1); the incidence of MACE was 4.7-8.3, AOEs: 25.8-33.3, and VTE: 2.3-9.1 (in 100 person-years). For those with use of two types of TKIs before the index date, 29 ponatinib, 29 bosutinib, and 4 other TKI patients were identified, with an average age of 51, 59, and 65 years, respectively. A similar trend was observed for patients with use of two prior TKIs. CONCLUSION: CP-CML patients treated with different TKIs (ponatinib, bosutinib, imatinib, dasatinib, and nilotinib) did not have different incidence of cardiovascular events (MACE, AOEs, VTEs) in this small cohort of real-world patients with ≥6-month of follow-up. The results were consistent among patients with prior use of one and two TKI types. Disclosures Levy: Takeda (Millennium Pharmaceuticals, Inc.): Consultancy. Xie:STATinMED Research: Employment. Wang:STATinMED Research: Employment. Neumann:Takeda (Millennium Pharmaceuticals, Inc.): Employment. Srivastava:Takeda (Millennium Pharmaceuticals, Inc.): Employment. Naranjo:Takeda (Millennium Pharmaceuticals, Inc.): Employment. Zhang:STATinMED Research: Employment. Dalal:Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd, Cambridge, MA, USA: Employment, Equity Ownership.
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Kantarjian, Hagop M., Javier Pinilla-Ibarz, Philipp D. Le Coutre, Ronald Paquette, Charles Chuah, Franck E. Nicolini, Jane Apperley, et al. "Five-year results of the ponatinib phase II PACE trial in heavily pretreated CP-CML patients (pts)." Journal of Clinical Oncology 35, no. 15_suppl (May 20, 2017): 7012. http://dx.doi.org/10.1200/jco.2017.35.15_suppl.7012.
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7012 Background: The tyrosine kinase inhibitor (TKI) ponatinib has potent activity against native and mutant BCR-ABL1 and is approved for use in pts with relapsed/intolerant CML or Ph+ ALL, or with BCR-ABL1/T315I. Methods: In the pivotal PACE study (NCT01207440), ponatinib (starting dose 45 mg/d) was assessed in pts with CML or Ph+ ALL resistant/intolerant to dasatinib or nilotinib, or with T315I. In Oct ’13, dose reductions were implemented due to observed arterial occlusive events (AOEs). Efficacy and safety at 5 yrs (data as of 3 Oct ’16) for CP-CML pts are reported. Results: Of 270 CP-CML pts in the safety population, 60% received ≥3 prior TKIs. At initiation of study closure, 99 pts were ongoing; among these pts, minimum follow-up was 52 months, and most (78%) had 15 mg/d as their last dose. In all CP-CML pts (n = 267, efficacy evaluable), cumulative response rates were: MCyR, 60%; CCyR, 54%; MMR, 40%; and MR4.5, 24%. Among pts who achieved MCyR (n = 148) or MMR (n = 108), the Kaplan-Meier (KM) estimated probability of remaining in response at 5 yrs was 74% (95% CI, 62 – 83) and 61% (95% CI, 51 – 70), respectively. Regardless of dose reduction in Oct ’13, maintenance of response was high (Table). KM estimated 5-yr rate for PFS/OS was 49%/77%. TEAEs in ≥45% of CP-CML pts were rash 47%, abdominal pain 46%, and thrombocytopenia 46%. Most newly occurring AEs were observed within the first year. The incidence of any AOEs/serious AOEs for CP-CML pts was 29%/23%. Among CP-CML pts with no prior AOEs who had a prospective dose reduction, 17% (11/63) had a first AOE occurring after Oct ‘13. Conclusions: Long-term (5-yr) results from PACE demonstrate that ponatinib continues to show clinical benefit, irrespective of dose reductions, with deep and lasting responses in heavily pretreated CP-CML pts. Safety results were consistent with the safety profile across the ponatinib clinical program. Clinical trial information: NCT01207440. [Table: see text]
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Levy, Moshe Yair, Lin Xie, Yuexi Wang, Frank Neumann, Shouryadeep Srivastava, Daniel Naranjo, Jing Xu, Qisu Zhang, and Mehul Dalal. "Major Adverse Cardiac, Arterial Occlusive, and Venous Occlusive Events Among Chronic Myeloid Leukemia Patients Prescribed Ponatinib Vs Bosutinib." Blood 134, Supplement_1 (November 13, 2019): 4751. http://dx.doi.org/10.1182/blood-2019-129053.
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INTRODUCTION: Chronic myeloid leukemia (CML) is a bone marrow and blood disorder accounting for 15% of adult leukemia. Tyrosine kinase inhibitors (TKIs) have been the standard of care for CML treatment. However, an association between TKI use and cardiovascular events has been observed. Ponatinib and bosutinib are introduced to provide more options for patients who failed their first-line treatment. The incidence of major adverse cardiac events (MACEs), arterial occlusive events (AOEs), and venous occlusive events (VOEs) were assessed among CML patients who were prescribed ponatinib vs bosutinib. METHODS: A retrospective observational study was conducted among adult CML patients aged ≥18 years with use of 1 or 2 prior TKIs who were prescribed bosutinib or ponatinib. Study patients were selected from the IBM® MarketScan® Research database from July 2012-June 2017. The index date was defined as the index drug prescription date, identified based on TKI use during the identification period (January 2013-December 2016) in a hierarchical order based on the sequence of treatment lines: ponatinib and bosutinib without ponatinib. Continuous health plan enrollment for ≥6 months pre-index date (baseline period) and at least 6 months post-index date (follow-up period) was required. Cardiovascular (CV) events (MACEs, AOEs, VOEs) using ICD-9/10-CM diagnosis codes occurring through the earliest of index TKI discontinuation, switch to another TKI, or end of follow-up period, were calculated as the number of events per 100 person-years (PYs). Inverse probability of treatment weighting (IPTW) was applied to adjust for differences in baseline characteristics between the treatment cohorts. Kaplan-Meier (KM) and Cox proportional hazard model analyses were conducted on the adjusted sample to examine any difference in CV event risk. RESULTS: After applying the selection criteria, 79 and 109 patients were included in the ponatinib and bosutinib cohorts, respectively. Mean ages were 53 years (ponatinib cohort) and 58 years (bosutinib cohort). The average Charlson Comorbidity Index (CCI) scores - defined by categorizing comorbidities using diagnosis codes - were 1.23 for ponatinib and 1.81 for the bosutinib cohort. Common baseline comorbid conditions included anemia (ponatinib: 49%; bosutinib 34%), hypertension (ponatinib: 33%, bosutinib: 46%), and diabetes (ponatinib: 15%; bosutinib: 29%). Some patients were observed to have CV events, specifically MACEs (ponatinib 8%; bosutinib 16%) and AOEs (ponatinib 15%; bosutinib 28%), before index ponatinib or bosutinib use. In the follow-up period, ponatinib patients were associated with a similar incidence of MACEs (14.70 vs 10.46 per 100 PYs; p=0.464), AOEs (29.56 vs 34.50 per 100 PYs; p=0.632), and VOEs (36.21 vs 34.70 per 100 PYs; p=0.890) compared to bosutinib patients. After applying IPTW, similar risks of the CV events (MACE, AOEs, VTEs) were observed in the KM analysis (Figure 1) expressed as time to CV event. After adjusting for additional confounders using Cox models, compared to those with bosutinib use, ponatinib patients were associated with similar rate of MACEs (Hazard Ratio [HR]: 1.02; 95% CI: 0.34, 3.01), AOEs (HR: 0.90; 95% CI: 0.43, 1.85), and VOEs (HR: 0.92; 95% CI: 0.44, 1.94). CONCLUSION: Among CML patients treated with ponatinib or bosutinib in second or third line, similar risks of cardiovascular events (MACE, AOEs, VTEs) were observed in the follow-up in this study in a community setting. Figure 1 Disclosures Levy: Takeda (Millennium Pharmaceuticals): Consultancy. Xie:STATinMED Research: Other: I am a paid employee of STATinMED Research which is a paid consultant to Takeda Pharmaceutical Corporation. . Wang:STATinMED Research: Other: I am a paid employee of STATinMED Research which is a paid consultant to Takeda Pharmaceutical Company. . Neumann:Millennium Pharmaceuticals (Takeda): Employment, Other: I am a paid employee of Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd. Srivastava:Millennium Pharmaceuticals (Takeda): Other: I am a paid employee of Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd. Naranjo:Millennium Pharmaceuticals (Takeda): Other: I am a paid employee of Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd. Xu:Millennium Pharmaceuticals (Takeda): Other: I am a paid employee of Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd. Zhang:STATinMED Research: Other: I am a paid employee of STATinMED Research which is a paid consultant to Takeda Pharmaceutical Corporation.. Dalal:Millennium Pharmaceuticals (Takeda): Other: I am a paid employee of Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Ltd.
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Cortes, Jorge E., Jane Apperley, Elza Lomaia, Beatriz Moiraghi, Maria Undurraga Sutton, Carolina Pavlovsky, Charles Chuah, et al. "OPTIC primary analysis: A dose-optimization study of 3 starting doses of ponatinib (PON)." Journal of Clinical Oncology 39, no. 15_suppl (May 20, 2021): 7000. http://dx.doi.org/10.1200/jco.2021.39.15_suppl.7000.
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7000 Background: PON, a third-generation tyrosine kinase inhibitor (TKI), demonstrated deep and long-lasting responses and survival in patients (pts) with chronic-phase chronic myeloid leukemia (CP-CML) resistant/intolerant to second-generation TKI therapy (PACE; NCT01207440); post hoc analysis suggested a relationship between dose and both adverse events and response. Here we present the primary analysis of OPTIC (NCT02467270), an ongoing, randomized, phase 2 trial with a novel response-based dosing regimen of PON in pts with resistant/intolerant CP-CML. Methods: Pts with CP-CML resistant/intolerant to ≥2 TKIs or with the BCR-ABL1 T315I mutation were randomized to PON starting doses of 45 mg (cohort A; 45 mg → 15 mg), 30 mg (B; 30 mg →15 mg), and 15 mg (C) once daily. Doses were reduced to 15 mg with achievement of ≤1% BCR-ABL1IS in cohorts A and B. The primary endpoint is ≤1% BCR-ABL1IS at 12 mo; secondary endpoints include cytogenetic and molecular responses and safety outcomes. AOEs were adjudicated prospectively by an independent review committee. Results: 283 pts were randomized (A/B/C: n=94/95/94) and had the following baseline characteristics: median age 48 y (18‒81 y); 98% received ≥2 (55% ≥3) TKIs; 99% had resistant disease; 40% had ≥1 baseline mutations (23% T315I). At the primary analysis with 32 mo median follow-up, 134 pts (47%; n=50/41/43) remained on treatment and 204 pts (72%) had PON exposure ≥12 mo. At 12 mo, 44% (41/93) in A, 29% (27/93) in B, and 23% (21/91) in C achieved ≤1% BCR-ABL1IS (Table); primary endpoint was met by cohort A. Dose reductions to 15 mg after achieving response (A/B) were 48/29%. Most common grades ≥3 TEAEs were thrombocytopenia, 27%; neutropenia, 17%; and anemia, 7%. AOEs/serious AOEs were reported in cohorts A (10%/4%), B (5%/4%), and C (3%/3%). Dose reductions or discontinuations for TEAEs (A/B/C) were 46/35/32% and 19/16/14%, respectively. Conclusions: The OPTIC primary analysis demonstrates the optimal benefit:risk profile for PON was achieved with a response-based dosing regimen starting with 45 mg/d, followed by dose reduction to 15 mg/d upon achieving ≤ 1% BCR-ABL1IS; 30 mg→15 mg and 15 mg cohorts may provide benefit, especially in pts without T315I mutation (Table). The observed ≤1% BCR-ABL1IS responses are supported by robust survival outcomes in pts with CP-CML resistant to second-generation BCR-ABL1 TKI therapy, both with and without BCR-ABL1 mutations. Clinical trial information: NCT02467270. [Table: see text]
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Caocci, Giovanni, Olga Mulas, Elisabetta Abruzzese, Alessandra Iurlo, Imma Attolico, Sara Galimberti, Luigia Luciano, et al. "Arterial Occlusive Events in Chronic Myeloid Leukemia Patients Treated with Ponatinib in the Real-Life Practice: Prophylaxis and Identification of Risk Factors." Blood 132, Supplement 1 (November 29, 2018): 3006. http://dx.doi.org/10.1182/blood-2018-99-111502.
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Abstract Background . Arterial occlusive events (AOEs) represent emerging complications in chronic myeloid leukemia (CML) patients treated with ponatinib, with a cumulative incidence correlated with the higher dose of the drug and longer treatment duration. Current recommendations highlight the importance of a careful evaluation of cardiovascular (CV) risk factors at baseline.Moreover, a preventive strategy with primary prophylaxis based on aspirin still remains under discussion and no data have been reported on secondary prophylaxis. Methods. We investigated a consecutive series of adult CML patients (mean age 50 years, range 24-81) who initiated ponatinib, between January 2012 and December 2016 at 15 Italian centers. Patients were stratified according to the Systematic Coronary Risk Evaluation (SCORE) assessment, based on gender, age, smoking habits, systolic blood pressure, and total cholesterol levels. Additional risk factors were considered the presence of diabetes, body mass index > 24.5 kg/m2, mild or severe renal insufficiency, and dyslipidemia. CML patients were also evaluated for both comorbidities and a positive anamnesis of CV diseases.Primary and secondary CV prophylaxis before starting ponatinib was also reported. We evaluated the cumulative incidence of AOEs (myocardial infarction, angina, ischemic cerebrovascular events and peripheral vascular disease) after initiating treatment with ponatinib, and their management. Results. A total of 71 patients were retrospectively identified. The reasons for treatment with ponatinib were inefficacy of previous tyrosine kinase inhibitors (TKIs) in 80.2% and intolerance in 19.8%. The median time of exposure to ponatinib was 16 months (range 3-69).The 60-month cumulative incidence of AOEs was 30.9±11.5%.Patients aged ≥60 years showed a higher incidence of AOEs (61.8±19.5% vs 19.5±12.0%, p=0.001) (Figure 1). The majority of patients (95%) were classified as at low-intermediate SCORE risk and 5% as at high-very high SCORE risk. Patients with a high-very high SCORE showed a significantly higher incidence of AOEs (100% vs. 25.8±11.5%; p<0.002). In multivariate analysis, no association was found with positive anamnesis of CV diseases, CV risk factors, dose of ponatinib (15, 30, 45 mg), exposure time to ponatinib, and number of TKI treatments; only age ≥60 years showed a significative association (p=0.016) . Overall, 13 patients underwent aspirin 100 mg primary prophylaxis before starting ponatinib and showed a a lower albeit not statistically significant incidence of AOEs (14.3±13.2% vs 33.4±12.8%, p=NS). In 6 of these patients with age < 60 years, no AOEs was reported. Conversely, the cumulative incidence of AOEs in the 6 patients undergoing secondary prophylaxis (anticoagulant, cardioaspirin) for previous events was 100% at 30 months. Overall, 23 CV AEs were registered; 52.1% of CV AEs were graded as 3/4 according to common toxicity criteria. In 43.4% of cases, ponatinib treatment did not require dose modification, 34.8% of patients reduced the dose and 21.8% discontinued the treatment. The majority of patients required additional diagnostic tests as ECG/cardiac ultrasound, peripheral vascular Doppler or cranial/CT; 4 patients underwent a coronarography procedure and 1 patient required invasive procedures as percutaneous transluminal angioplasty and application of coronary stents. Finally, the 5-year cumulative incidence of MR4 following ponatinib treatment was 45.0±12.3% and it was not influenced significantly by AOEs occurrence. Conclusions. This study confirms the increased risk of AOEs in CML patients treated with ponatinib in the real-life, particularly in patients aged ≥60 years. Our findings emphasize the need of personalized prevention strategies based on CV risk factors, in close collaboration with cardio-oncologists, angiologists and vascular surgeons. We suggest that patients treated with ponatinib should undergo prophylaxis with aspirin 100 mg. Data on the efficacy of primary prophylaxis need to be confirmed in larger cohorts of patients and in prospective randomised trials. Figure 1.Arterial Occlusive Events (AOEs) cumulative incidence according to age ≥ 60 years in 71 CML patients treated with ponatinib Figure. Figure. Disclosures Abruzzese: BMS: Consultancy; Novartis: Consultancy; Ariad: Consultancy; Pfizer: Consultancy. Bonifacio:Incyte: Consultancy; Pfizer: Consultancy; Amgen: Consultancy; Novartis: Research Funding; Bristol Myers Squibb: Consultancy. Foà:CELTRION: Other: ADVISORY BOARD; GILEAD: Speakers Bureau; ABBVIE: Other: ADVISORY BOARD, Speakers Bureau; CELGENE: Other: ADVISORY BOARD, Speakers Bureau; NOVARTIS: Speakers Bureau; ROCHE: Other: ADVISORY BOARD, Speakers Bureau; INCYTE: Other: ADVISORY BOARD; AMGEN: Other: ADVISORY BOARD; JANSSEN: Other: ADVISORY BOARD, Speakers Bureau. Breccia:Incyte: Honoraria; BMS: Honoraria; Novartis: Honoraria; Pfizer: Honoraria.
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Ito, Tomoki, Nobuhiko Uoshima, Yasuhiro Maeda, Masaaki Hotta, Hideaki Yoshomura, Shinya Fujita, Atsushi Satake, et al. "Evaluation Of Large Granular Lymphocytes and Endothelial-Cell-Related Biomarkers In Patients With Chronic Myeloblastic Leukemia: Comparison Among 3 TKIs." Blood 122, no. 21 (November 15, 2013): 5167. http://dx.doi.org/10.1182/blood.v122.21.5167.5167.
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Abstract Background Tyrosine kinase inhibitors (TKIs) currently represent the main therapy for chronic myeloblastic leukemia (CML). Although they are therapeutically effective, some TKI-related events such as pleural effusion and elevation of large granular lymphocytes (LGL) have been reported. In addition, these events itself may affect the therapeutic response to TKIs. In the present study, we measured the levels of some cytokines, chemokines, soluble factors and coagulation markers in patients with CML, and investigated the relationship between these markers and TKI-related events. Methods The subjects were 48 patients with CML. Blood samples were collected and levels of cytokines (interleukin (IL)-6, tumor necrosis factor-α, high-mobility group box 1), chemokines (monocyte chemotactic protein (MCP)-1, RANTES), soluble molecules (soluble vascular cell adhesion molecule-1, soluble E-selectin, angiopoietin-2, vascular endothelial growth factor, and soluble thrombomodulin) and coagulation markers (plasminogen activator inhibitor (PAI)-1, platelet-derived microparticles (PDMP)) were measured by ELISA. Results Levels of all markers except IL-6 and MCP-1 were significantly increased in patients with de novo CML. Level of LGL/lymphocyte was significantly higher in dasatinib-treated CML patients compared to other TKI-treated patients. LGL/lymphocyte and many biomarkers exhibit the time-dependent increase after dasatinib treatment. In addition, LGL/lymphocyte was already increasing in patients with pleural effusion before dasatinib treatment. Conclusion These results suggest that LGL and endothelial-cell-related factors play an important role in the pathophysiology of CML and its response to TKIs. Disclosures: No relevant conflicts of interest to declare.
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Jedema, Inge, Linda van Dreunen, Roelof Willemze, and J. H. Frederik Falkenburg. "Treatment with Tyrosine Kinase Inhibitors May Impair the Potential Curative Effect of Allogeneic Stem Cell Transplantation." Blood 114, no. 22 (November 20, 2009): 857. http://dx.doi.org/10.1182/blood.v114.22.857.857.
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Abstract Abstract 857 Tyrosine kinase inhibitors (TKI) like imatinib and dasatinib are the current treatment of choice for patients with chronic myeloid leukemia (CML). Although most patients enter a complete remission during treatment, cure of the disease is usually not achieved since recurrence of the disease is seen in the majority of patients upon discontinuation of the treatment, indicating that the leukemic stem cell is not efficiently targeted. Furthermore, in accelerated phase and blast crisis of CML TKI treatment only results in temporary control of the disease. In these situations allogeneic stem cell transplantation (allo-SCT) and application of donor T cells may be the only curative treatment. Besides the direct anti-leukemic effect of allo-SCT, alloreactive T cells recognizing CML (progenitor) cells, and the formation of immunological memory may lead to effective lifelong immune surveillance. Therefore, we investigated whether the leukemic cells persisting during TKI treatment are susceptible targets for the anti-leukemic effect mediated by donor T cells after allo-SCT and whether continuous TKI treatment may have an additive effect during the immunological intervention. To investigate the anti-leukemic effect of the two strategies, CD34+ positive CML cells were isolated from bone marrow, and labeled with the fluorescent dyes CFSE or PKH to allow monitoring of single cell proliferation. CML cells were exposed to imatinib (1-100μM) or dasatinib (0.01-50nM), and/or to CD8+ alloreactive cytotoxic T lymphocyte (CTL) clones in the presence of proliferation-inducing cytokines. The number, phenotype, and proliferative status of the CML cells persisting after single and combined interventions were measured by quantitative flowcytometric analysis. In the absence of therapeutic interventions the majority of CD34+ CML cells entered proliferation. However, a small population of CD34+ CML stem cells residing in the non-dividing peak could be identified despite the addition of cytokines. Addition of imatinib or dasatinib resulted in efficient dose-dependent induction of cell death of the leukemic cells (99% lysis by 25μM imatinib or 10nM dasatinib). However, the population of quiescent CD34+ CML stem cells was not affected. Moreover, the number of cells present in the non-dividing population increased 2-fold compared to the non-treated controls at the highest TKI concentrations, indicating additional growth arrest of a population of proliferating CML precursor cells. We next tested the capacity of different HLA-A2-restricted CD8+ CTL clones to kill non-treated or imatinib or dasatinib treated CML cells. Whereas the proliferating CD34+ CML precursors were efficiently lysed, the population of quiescent stem cells was capable of withstanding CTL exposure. Detailed phenotypic analysis revealed significant downregulation of HLA-A2 and the adhesion molecules CD49d and CD58 on these quiescent cells, probably resulting in the impaired ability of these target cells to form a high avidity interaction with the T cells. The increased population of non-dividing cells as a result of the TKI pretreatment showed similar resistance to T cell induced cell death, indicating that TKI treatment may even diminish the anti-leukemic effect of allo-SCT. In the absence of therapeutic control, as mimicked by the removal of T cells and TKI from the cultures, outgrowth of the leukemic cells re-occurred, illustrating their capacity to give rise to a relapse of the disease. Next, we analyzed the effect of TKI treatment on T cell survival and functionality. Whereas resting primary T cells were insensitive to TKI treatment, T cells activated by either polyclonal stimulation with anti-CD3/CD28 beads or stimulation with allogeneic stimulator cells died after TKI exposure at similar concentrations as the leukemic cells. In conclusion, TKI treatment results in selection of a population of quiescent leukemic stem cells showing cross-resistance to CTL-induced cell death, most likely due to their inability to form a high avidity interaction. Moreover, T cells actively participating in the anti-leukemic immune response after allo-SCT are suppressed by TKI. These data indicate that continuous TKI treatment may potentially hamper the curative effect of allo-SCT. Disclosures: No relevant conflicts of interest to declare.
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Stemhagen, Annette, Deyaa Adib, Stephanie Lustgarten, Lisa McGarry, Ruth Du Moulin, and Sergio Santillana. "The OMNI patient registry: A prospective observational registry to assess vascular safety in patients with CML and Ph+ ALL treated with ponatinib." Journal of Clinical Oncology 35, no. 15_suppl (May 20, 2017): TPS7073. http://dx.doi.org/10.1200/jco.2017.35.15_suppl.tps7073.
Full textAbstract:
TPS7073 Background: Ponatinib is an oral TKI with potent activity against BCR-ABL1. The pivotal PACE study (NCT01207440) formed the basis for approval of ponatinib in the US for the treatment of patients with resistant/intolerant CML or Ph+ ALL, or those with the T315I mutation. Long-term follow-up of PACE showed a higher cumulative incidence of vascular occlusive events (VOEs) than reported at the time of approval — dose reductions were later implemented to mitigate VOEs. VOEs comprise arterial occlusive events (AOE) and venous thromboembolic events. The exposure-adjusted incidence of AOEs has not increased over time in PACE; in patients with a history of ischemic disease, the relative risk of serious AOEs was 2.6 in those with ≥2 vs 0 risk factors. The primary objective of this patient registry is to assess VOEs occurring during ponatinib use in routine clinical practice in the US. Methods: OMNI (NCT02455024) is a prospective observational registry of eligible patients (Table) with CML or Ph+ ALL for whom the decision to initiate treatment with ponatinib has already been made for the approved US indications. Patients voluntarily enroll into the registry, which is non-interventional with no protocol-mandated tests/procedures — all treatment decisions are made at the discretion of the health care practitioner in consultation with their patient. Study duration is anticipated as ~30 mo (~18-mo enrollment followed by 12 mo of data collection, which will occur every 3 mo). The primary analysis will be performed 12 mo after last patient enrolled and will estimate the incidence of VOEs by duration of ponatinib exposure. To understand differences between those with and without VOEs, exploratory analyses will be performed, considering factors such as patient demographics, risk factors for developing VOEs, dose and duration of ponatinib treatment, and concomitant medications. VOE outcomes also will be assessed. Enrollment will begin in 2017, with a target of ≥300 patients. Clinical trial information: NCT02455024. [Table: see text]
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Truitt, Luke, Catherine Hutchinson, Karen Mochoruk, John F. DeCoteau, and C. Ronald Geyer. "Chaetocin Anti-Leukemia Activity Against Chronic Myelogenous Leukemia Stem Cells Is Potentiated By Bone Marrow Stromal Factors and Overcomes Innate Imatinib Resistance." Blood 124, no. 21 (December 6, 2014): 4517. http://dx.doi.org/10.1182/blood.v124.21.4517.4517.
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Abstract Chronic myelogenous leukemia (CML) is maintained by a minor population of leukemic stem cells (LSCs) that exhibit innate resistance to tyrosine kinase inhibitors (TKIs) targeting BCR-ABL. Innate resistance can be induced by cytokines and growth factors secreted by bone marrow stromal cells (BMSFs) that protect CML-LSCs from TKIs, resulting in minimal residual disease. Developing therapies that can be combined with TKIs to eradicate TKI-insensitive CML-LSCs, is critical for disrupting innate TKI resistance and preventing disease relapse. Cancer cells balance reactive oxygen species (ROS) and antioxidants at higher than normal levels, which promotes their proliferation and survival, but also makes them susceptible to damage by ROS-generating agents. BCR-ABL expression increases cellular ROS levels, whereas, TKI inhibition of BCR–ABL reduces ROS. Furthermore, BMSFs, which are implicated in innate TKI resistance, can increase ROS levels in CML cells. Thus, we postulated that BMSF mediated increases in ROS might enhance triggering of ROS-mediated damage in TKI treated CML-LSCs by chaetocin, a mycotoxin with anticancer properties that imposes oxidative stress by inhibiting thioredoxin reductase-1. To investigate chaetocin effects on innate TKI resistance, we first compared its activity with imatinib against TonB210, a murine hematopoietic cell line with inducible BCR-ABL expression, in response to BMSFs. Imatinib did not affect the growth of BCR-ABL(-) TonB210 cells but suppressed BCR-ABL(+) Ton-B210 growth, and BMSFs protected against imatinib growth suppression. In contrast, chaetocin significantly suppressed the growth of both BCR-ABL(-) and BCR-ABL(+) TonB210 cells, and these effects were potentiated by BMSFs. We then compared the effects of chaetocin as a single agent, and in combination with imatinib, on the growth of CML-LSCs derived from an established murine retroviral transduction/transplantation model of CML blast crisis, in response to BMSFs. The presence of BMSFs reduced cytotoxicity and apoptosis induction by imatinib, but potentiated these effects in chaetocin treated CML-LSCs. Colony formation by CML-LSCs was significantly inhibited by treatment with either imatinib or chaetocin. However, BMSFs conferred significant protection from colony inhibition by imatinib, whereas, no colony formation was observed in cells exposed to chaetocin and BMSFs. Both BMSFs and chaetocin increased ROS in CML-LSCs and the addition of BMSFs and chaetocin resulted in significantly higher levels compared to chaetocin or BMSFs alone. Pretreatment of CML-LSCs with the anti-oxidant N-acetyl-cysteine blocked chaetocin cytotoxicity, even in the presence of BMSFs. Chaetocin effects on CML-LSC self-renewal in vivo were assessed by transplanting CML-LSCs into secondary recipients following in vitro exposure to chaetocin, in the presence or absence of BMSFs. Disease latency in mice transplanted with CML-LSCs following chaetocin treatment more than doubled compared to mice transplanted with untreated CML-LSCs or CML-LSCs exposed to BMSFs. Mice transplanted with CML-LSCs following chaetocin treatment in the presence of BMSFs had significantly extended survival time compared to mice transplanted with CML-LSCs treated with chaetocin alone. Our findings indicate that chaetocin activity against leukemia initiating cells is significantly enhanced in the presence of BMSFs and suggest that chaetocin may be effective as a co-drug to complement TKIs in CML treatment by disrupting the innate resistance of CML-LSCs through an ROS dependent mechanism. Disclosures No relevant conflicts of interest to declare.
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Pulte, E. Dianne, Haiyan Chen, Lauren S. L. Price, Ramadevi Gudi, Hongshan Li, Olanrewaju O. Okusanya, Lian Ma, et al. "FDA Approval Summary: Revised Indication and Dosing Regimen for Ponatinib Based on the Results of the OPTIC Trial." Oncologist 27, no. 2 (February 1, 2022): 149–57. http://dx.doi.org/10.1093/oncolo/oyab040.
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Abstract On December 18, 2020, US Food and Drug Administration (FDA) approved a supplemental application for ponatinib extending the indication in patients with chronic-phase chronic myeloid leukemia (CP-CML) to patients with resistance or intolerance of at least 2 prior kinase inhibitors. Ponatinib was initially approved in December 2012 but was briefly voluntarily withdrawn due to serious safety concerns including the risk of arterial occlusive events (AOE). It returned to the market in December 2013 with an indication limited to patients with T315I mutation or for whom no other tyrosine kinase inhibitor (TKI) therapy was indicated with revised warnings and precautions. A post-marketing requirement was issued to identify the optimal safe and effective dose for CP-CML. Thus, the OPTIC trial was performed, which randomized patients to 1 of 3 doses, 45 mg, 30 mg, or 15 mg, with a dose reduction to 15 mg on achievement of MR2 (BCR-ABLIS ≤1%). Patients enrolled were treated with at least 2 prior TKIs or had a T315I mutation. Patients with a history of clinically significant, uncontrolled, or active cardiovascular disease were excluded. Efficacy was established on an interim analysis based on the rate of MR2 at 12 months in the modified intent-to-treat population of 261 patients, with 88, 86, and 87 patients in the 45, 30, and 15 mg cohorts, respectively. With a median follow-up of 28 months, the rate of achievement of MR2 at 12 months was 42%, 28%, and 24% in the respective cohorts. The safety profile was consistent with that observed in prior evaluations of ponatinib with notable adverse reactions including pancreatitis, hypertension, hyperlipidemia, liver dysfunction, and AOE. Of patients treated at the 45/15 mg dose, AOEs were seen in 13%, with a higher rate being observed in patients age 65 or older compared to younger patients. A readjudication of AOEs seen on the prior pivotal phase 2 study resulted in a rate of 26%. Overall, the results supported a modification of the recommended dose for patients with CP-CML to 45 mg until the achievement of MR2 followed by a reduction to 15 mg. The expansion of the indication to patients with exposure to 2 prior TKIs was approved given data showing that ponatinib could be successfully used for the treatment of this population with appropriate monitoring and screening for risk factors.
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Jalkanen, Sari, Satu Mustjoki, Kimmo Porkka, and Jukka Vakkila. "Cytokine-Mediated Signaling Is Suppressed in Myeloid Cells and Enhanced in Lymphatic Cells in Patients with Chronic Myeloid Leukemia (CML) — Partial Normalization with Tyrosine Kinase Inhibitors." Blood 114, no. 22 (November 20, 2009): 2180. http://dx.doi.org/10.1182/blood.v114.22.2180.2180.
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Abstract Abstract 2180 Poster Board II-157 Introduction. Aberrant phosphorylation of the BCR-ABL1 tyrosine kinase (TK) is characteristic of chronic myeloid leukemia (CML). This oncoprotein interacts directly with intracellular signaling proteins, alters the responsiveness of cytokine receptors and regulates secretion of autocrine cytokines. Targeted inhibition of BCR-ABL1 with TK inhibitor (TKI) imatinib mesylate (IM) is the current standard treatment of CML. For overcoming IM resistance or intolerance, 2nd generation TKIs (nilotinib, dasatinib) with broader kinase inhibition profile have been approved for clinical use. Although in vitro results suggest that TKIs are immunosuppressive, no increases in opportunistic infections or secondary malignancies have been observed to date. In contrast, in some TKI-treated patients immunoactivation in the form of chronic lymphocytosis linked to excellent therapy responses has recently been shown. Dynamic monitoring of aberrant cytokine signaling pathways would aid in understanding and predicting the development of TKI-resistance or adverse/off-target effects. The aim of this study was to analyze the responsiveness of leukocytes to cytokine stimuli in CML patients at diagnosis and during TKI therapy using single-cell profiling of phosphoprotein networks by multiparameter flow cytometry. Patients and methods. The study consisted of 4 healthy controls, 6 CML patients at diagnosis, 6 IM patients and 5 dasatinib patients. Stimuli included GM-CSF, IL-2+IL-10+IFNα and IL-4+IL-6+IFNγ and they were added immeadately to freshly drawn whole blood ex vivo. The readout phosphoproteins were pERK1/2, pSTAT1, pSTAT3, pSTAT5a and pSTAT6 (with isotype controls), and were analyzed separately from granulocytes, monocytes, CD4+ CD25neg T helper cells (Th), CD4neg lymphocytes and CD4+CD25+ T cells including regulatory T-cells (Treg). Analysis was performed with heatmap function of Cytobank software (http://cytobank.stanford.edu/public/). Results. Unstimulated phosphoprotein levels reflecting the activation state of leukocytes in vivo did not differ between healthy controls and CML patients at diagnosis or during dasatinib therapy. Strikingly, in IM patients, baseline levels of pSTAT3 were relatively high indicating in vivo occurring activation of leukocytes in this patient group. We next studied ex vivo responsiveness of immune effector cells with cytokines and found clear differences between healthy controls and CML patients. At CML diagnosis. GM-CSF/pERK1+pSTAT5a, IFNa/pSTAT1,and IL-4/pSTAT6 (stimulus/readout) as well as pSTAT3 responses with all stimuli were suppressed in monocytes. In granulocytes, GM-CSF/pSTAT1 levels were diminished. In Th and Treg lymphocytes, IL-6/pSTAT3 responses were markedly pronounced, while IL-10/pSTAT3 responses were not affected when compared to healthy controls. Such difference was not observed in CD4neg lymphocytes. During TKI therapy. Most patients (9/11) were in cytogenetic remission at the time of analysis. The unresponsiveness of myeloid cells at diagnosis was restored by IM or dasatinib therapy in most, but not all patients. Similarly, in Th and Treg lymphocytes TKI-therapy normalized the enhanced IL-6/pSTAT3 responses that were evident at diagnosis. However, in Th and Treg cells pSTAT3 responses provoked by IL-10 were particularly prominent. Interestingly, one dasatinib patient with aberrant constant blood NK-lymphocytosis and monocytosis had uniquely strong IFNg/pSTAT1 and IL-4/pSTAT6 responses in monocytes. Furthermore, one patient who have stayed in persistent remission after IM discontinuation had exceptionally high pSTAT3 responses with all of stimuli used. Similar kind of signaling profile was unseen with the other patients and could reflect immunoactivation related to leukemia control. Conclusions. Dynamic single-cell profiling of signaling networks is feasible in CML patients and can be used to study mechanisms of aberrant immune reactivity in TKI-treated patients. The method could be particularly suitable for assessing candidate patients for TKI discontinuation. Although in vitro results suggest immunosuppressive effects of TKIs on lymphocytes, leukocytes ex vivo from patients were able to respond similarly to cytokine stimuli as in healthy controls. Disclosures: Mustjoki: BMS: Honoraria. Porkka:BMS: Honoraria, Research Funding; Novartis: Honoraria, Research Funding.
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Rouhimoghadam, Milad, Anthony D. Pomicter, Alexandria Van Scoyk, Greg Poffenberger, Ivaylo Kirov, Lyubomir G. Nashev, Helong Zhao, Martin Martinov, and Michael W. Deininger. "Exploiting LY3009120 and Asciminib Combination to Target TKI-Resistant CML." Blood 138, Supplement 1 (November 5, 2021): 3600. http://dx.doi.org/10.1182/blood-2021-153353.
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Abstract The oncogenic BCR-ABL1 tyrosine kinase is the driver of chronic myeloid leukemia (CML) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL). Tyrosine kinase inhibitors (TKIs) targeting ABL kinase are generally effective, but subsets of patients treated with single-agent TKIs develop resistance due to mutations in BCR-ABL1 that impair TKI binding. We have previously reported that BCR-ABL1 compound mutants (exhibiting two mutations within the same BCR-ABL molecule) that include the T315I gatekeeper mutation confer a high degree of resistance to all clinical ABL TKIs used as single agents, including ponatinib and the allosteric inhibitor asciminib. However, combining asciminib with ponatinib provides an effective strategy for overcoming compound mutation-based resistance (Eide et al. Cancer Cell 2019). As the clinical utility of ponatinib is limited by cardiovascular toxicity, including arterial occlusive events (AOEs), we decided to search for alternative molecules for use in combination with asciminib. To identify functional ponatinib analogs, we performed Quantum Similarity Modeling (QSM) on the reported crystal structure of T315I mutant ABL1 kinase in complex with nilotinib and asciminib (5MO4) (Wylie et al. Nature 2017) to search for other molecules. Compared to conventional computational modeling, QSM identifies novel classes of structurally distinct compounds that are comparable on a quantum level by precisely defining their interaction with the target. Affinity inferred by close complementarity with the shared ligand-protein surface in the region of the surveyed binding site is mapped, using multiple weak local associations. Our in silico QSM platform combines quantum methods with machine learning to investigate extensive chemical spaces. We screened several million compounds against BCR-ABL1 and identified 51 potential candidates predicted effectively to block T315I mutant BCR-ABL1 when combined with asciminib. To prioritize potent and non-toxic drug combinations for further development against compound mutants, we initially profiled all 51 compounds for their efficacy against Ba/F3 BCR-ABL T315I cells, alone and in combination with asciminib (1 nM). Of 51 compounds, LY3009120, a pan-RAF inhibitor that is currently in phase I clinical development for advanced solid malignancies (Sullivan et al. Mol Cancer Ther 2020), showed strong activity against BCR-ABL T315I when combined with asciminib. These data provided proof of principle for the QSM approach. We next tested the efficacy of all 51 candidates ± asciminib against Ba/F3 cells harboring T315I-inclusive BCR-ABL1 compound mutants, including Y253H/T315I, E255V/T315I, H396R/T315I, G250E/T315I, and T315L as the most resistant mutants. Neither single agent showed any effect. However, LY3009120 strongly inhibited BCR-ABL1 compound mutants when combined with asciminib. No toxicity was observed against Ba/F3 parental cells, confirming that the effects of the combinations are mediated by inhibition of BCR-ABL1. Synergy quantification of the dose-response matrix for the LY3009120/asciminib combination using the Zero Interaction Potency model demonstrated highly synergistic interactions (Synergy score > 10) between the two inhibitors. To directly assess the binding affinity of LY3009120 to the ABL1 kinase domain, we used the cell-based NanoBRET intracellular ABL1 kinase assay on HEK-293 cells expressing luciferase-tagged ABL1. The NanoBRET assay uses energy transfer to quantify the affinity of test compounds by competitive displacement of a cell-permeable fluorescent tracer that is reversibly bound to an ABL1-luciferase fusion protein. We found that LY3009120 competes off the fluorescent tracer at a low micromolar range (EC 50 = 0.75 μM), confirming direct binding of LY3009120 to the kinase domain of ABL1. We hypothesize that the binding of LY3009120 to the ABL1 kinase domain induces a conformational change that re-establishes asciminib binding to the myristoyl binding pocket, allowing for synergy. Studies to quantify the binding affinity of LY3009120 and asciminib to BCR-ABL1 mutants are underway, and data will be presented. In summary, our findings validate QSM as a novel in silico approach to identify TKI combinations. Combining LY3009120 with asciminib may be an effective, low-risk strategy to target BCR-ABL1 compound mutants in patients with clinical TKI resistance. Disclosures Deininger: SPARC, DisperSol, Leukemia & Lymphoma Society: Research Funding; Sangamo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Honoraria, Research Funding; Fusion Pharma, Medscape, DisperSol: Consultancy; Novartis: Consultancy, Research Funding; Blueprint Medicines Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding.
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Kantarjian, Hagop M., Michael W. Deininger, Elisabetta Abruzzese, Jane Apperley, Jorge E. Cortes, Charles Chuah, Daniel J. DeAngelo, et al. "Efficacy and Safety of Ponatinib (PON) in Patients with Chronic-Phase Chronic Myeloid Leukemia (CP-CML) Who Failed One or More Second-Generation (2G) Tyrosine Kinase Inhibitors (TKIs): Analyses Based on PACE and Optic." Blood 136, Supplement 1 (November 5, 2020): 43–44. http://dx.doi.org/10.1182/blood-2020-133922.
Full textAbstract:
Introduction: The use of 2G TKIs in patients with CP-CML who have failed ≥1 2G TKIs is not associated with durable responses; there is limited clinical evidence to support that switching to alternate 2G TKI therapy improves long-term clinical outcomes for these patients. Patients with resistant and intolerant CP-CML with substantial prior 2G treatment demonstrated deep, lasting responses to PON in the pivotal PACE trial. A post hoc modeling analysis of the data from PACE suggested a relationship between dose and safety events (including arterial occlusive events [AOEs]). The OPTIC trial was designed to prospectively evaluate response-based PON dosing regimens with the aim of optimizing its efficacy and safety in patients with CP-CML; the interim analysis (IA) demonstrated clinically manageable safety and AOE profiles with response-based PON dosing regimens. The combined PACE and OPTIC trials comprise the largest patient population in a post-2G TKI setting. We present the efficacy and safety outcomes of these patients over time. Methods: PACE (NCT01207440) evaluated PON in patients with refractory CML or Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL). OPTIC (NCT02467270) is a multicenter, randomized Phase 2 trial characterizing the safety and efficacy of PON over a range of 3 starting doses (45, 30, or 15 mg/day); patients with CP-CML receiving 45 or 30 mg/day reduced their doses to 15 mg/day upon achieving ≤1% BCR-ABL1IS. Data from patients with CP-CML in PACE (n=254) and the 45-mg starting dose cohort (45 mg→15 mg) in OPTIC (n=92) who have been treated with ≥1 2G TKI are presented; OPTIC data are from the IA. Efficacy data includes molecular responses (measured using polymerase chain reaction and performed at the same central lab for both studies) and survival outcomes over time. Safety data, including treatment-emergent AOE rates following adjudication, are also presented. Results: A combined 350 PON-treated patients from the PACE and OPTIC trials who have received ≥1 prior 2G TKI were analyzed; efficacy results are summarized in Table 1. The ≤1% BCR-ABL1IS response rates increased over time and ranged from 42% to 52% in the OPTIC IA 45-mg starting dose cohort and in PACE. Progression-free survival and overall survival were 52% and 73%, respectively, in PACE (up to 5 years) and 81% and 93%, respectively, at the OPTIC IA (up to 2 years) (Table 1). Serious treatment-emergent adverse event (AE) and AOE rates (including exposure-adjusted AOE rates) were lower in OPTIC IA with a response-adjusted dosing regimen compared with PACE (Table 2). Propensity score analyses comparing AOE incidence among all patients in OPTIC vs PACE demonstrate that the relative risk for adjudicated AOEs is 64% lower in OPTIC when compared with PACE after adjusting for baseline differences, duration of exposure, and total PON dose received. Analyses on responses by mutation status also will be presented. Conclusions: In this analysis, comprising the largest patient population of CP-CML patients in a post-2G TKI setting, ponatinib shows high response rates and robust survival outcomes in patients who have failed 2G TKIs. With the response-adjusted dosing regimen in OPTIC (starting at 45 mg and reducing to 15 mg upon response), efficacy outcomes were consistent with that of PACE, while the overall incidences of AOEs and serious treatment emergent-AEs were lower; exposure-adjusted AOEs during the first 2 years were also lower. The ongoing OPTIC study is also evaluating lower starting doses of ponatinib (30 and 15 mg) and primary analysis of this study will provide a refined understanding of the benefit:risk profile of the 3 starting doses of ponatinib in CP-CML patients. Disclosures Kantarjian: Adaptive biotechnologies: Honoraria; Ascentage: Research Funding; Actinium: Honoraria, Membership on an entity's Board of Directors or advisory committees; Janssen: Honoraria; Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Sanofi: Research Funding; Immunogen: Research Funding; BMS: Research Funding; Daiichi-Sankyo: Honoraria, Research Funding; Aptitute Health: Honoraria; BioAscend: Honoraria; Delta Fly: Honoraria; Amgen: Honoraria, Research Funding; Abbvie: Honoraria, Research Funding; Oxford Biomedical: Honoraria; Jazz: Research Funding. Deininger:Fusion Pharma: Consultancy; Blueprint Medicines Corporation: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: part of a study management committee, Research Funding; SPARC: Research Funding; DisperSol: Consultancy; Leukemia & Lymphoma Society: Research Funding; Novartis: Consultancy, Other, Research Funding; Medscape: Consultancy; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Other, Research Funding; Galena: Consultancy, Honoraria, Other; Pfizer: Honoraria, Other, Research Funding; Ariad: Consultancy, Honoraria, Other; Celgene: Research Funding; Gilead Sciences: Research Funding; Sangamo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Honoraria, Other, Research Funding. Abruzzese:Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Bms: Honoraria. Apperley:Pfizer: Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Incyte: Honoraria, Research Funding, Speakers Bureau; Bristol Myers Squibb: Honoraria, Speakers Bureau. Cortes:Pfizer: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Jazz Pharmaceuticals: Consultancy, Research Funding; Sun Pharma: Research Funding; BiolineRx: Consultancy, Research Funding; Amphivena Therapeutics: Research Funding; Telios: Research Funding; BioPath Holdings: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Bristol-Myers Squibb: Research Funding; Astellas: Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Merus: Research Funding; Immunogen: Research Funding; Arog: Research Funding. Chuah:Pfizer: Other: Travel, Research Funding; Korea Otsuka Pharmaceutical: Honoraria; Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria. DeAngelo:Jazz: Consultancy; Autolos: Consultancy; Novartis: Consultancy, Research Funding; Abbvie: Research Funding; Forty-Seven: Consultancy; Amgen: Consultancy; Agios: Consultancy; Blueprint Medicines Corporation: Consultancy, Research Funding; Takeda: Consultancy; Glycomimetics: Research Funding; Pfizer: Consultancy; Shire: Consultancy; Incyte Corporation: Consultancy. DiPersio:Magenta Therapeutics: Membership on an entity's Board of Directors or advisory committees. Hochhaus:Bristol-Myers Squibb: Honoraria, Research Funding; Incyte: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Takeda: Honoraria; MSD: Research Funding. Lipton:Takeda: Consultancy, Honoraria, Research Funding; Ariad: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Bristol-Myers Squibb: Honoraria; BMS: Consultancy, Research Funding; Pfizer: Consultancy, Honoraria, Research Funding. Nicolini:Sun Pharma Ltd: Consultancy; Incyte: Research Funding, Speakers Bureau; Novartis: Research Funding, Speakers Bureau. Pinilla Ibarz:Takeda: Consultancy, Speakers Bureau; Sanofi: Consultancy; TG Therapeutics: Consultancy; Sunesis Pharmaceuticals: Consultancy; Abbvie: Consultancy, Speakers Bureau; Janssen: Consultancy, Speakers Bureau; Novartis: Consultancy; Pharmacyclics: Consultancy, Speakers Bureau; AstraZeneca: Consultancy, Speakers Bureau. Rea:BMS: Membership on an entity's Board of Directors or advisory committees; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees. Rosti:Pfizer: Research Funding, Speakers Bureau; Bristol-Myers Squibb: Speakers Bureau; Incyte: Speakers Bureau; Novartis: Speakers Bureau. Rousselot:Pfizer: Consultancy, Research Funding; Incyte: Consultancy, Research Funding; Takeda: Consultancy; Bristol-Myers Squibb: Consultancy; Novartis: Consultancy. Shah:Bristol-Myers Squibb: Research Funding. Talpaz:IMAGO: Consultancy; BMS: Membership on an entity's Board of Directors or advisory committees; Takeda: Research Funding; Novartis: Research Funding; Constellation Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees. Srivastava:Takeda: Current Employment. Lu:Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Current Employment. Mauro:Bristol-Myers Squibb: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Pfizer: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Sun Pharma/SPARC: Research Funding; Takeda: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Novartis: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding.
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Kuznetsova, Valeriya, Virginia Camacho, Sweta Patel, Victoria Matkins, and Robert S. Welner. "Perturbed function of natural killer cells by inflammatory cytokines in chronic myeloid leukemia." Journal of Immunology 206, no. 1_Supplement (May 1, 2021): 57.06. http://dx.doi.org/10.4049/jimmunol.206.supp.57.06.
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Abstract Natural killer (NK) cells’ mature phenotype and counts positively correlate with prolonged treatment-free survival in chronic myeloid leukemia (CML), while dysfunctional NK cells are predictive of relapse in leukemic patients. However, the molecular drivers of NK cell alterations during leukemia remain unclear. Given an established role of inflammatory cytokines in CML progression, we define the leukemic environmental cues impacting NK maturation and function in a BCR-ABL chimeric mouse model of CML where NK cells do not carry the CML oncogene. Reflecting clinical observations, NK cell frequencies and numbers are severely decreased in leukemic mice. Moreover, terminally mature NK subsets are the most affected, implying preferential loss of cells with potent anti-leukemic activity. In addition, this phenotype suggests a shift toward the immature cytokine-producing state of NK development, which could favor the survival of leukemia-initiating cells. We map the transcriptional landscape of NK cells using single-cell RNA-sequencing from multiple tissues. NK cells exposed to CML microenvironment possess a lowered expression of surface activating receptors such as Nkp46 while upregulating inhibitory molecules Tim-3 and TIGIT. BCR-ABL reversion for six weeks partially restored these defects, consistent with findings in patients before and after TKI initiation. Finally, we found that leukemia-associated soluble factors dampen target-specific degranulation of NK cells by applying serum samples from healthy or leukemic mice. These data indicate that the altered cytokine milieu contributes to NK cell dysfunction and represents an optimal target for NK-boosting CML immunotherapies of leukemia.
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Hiwase, Devendra K., Deborah L. White, Jason A. Powell, Verity A. Saunders, Stephanie Zrim, Amity Frede, Mark Guthridge, et al. "Blocking of Cytokine Survival Signals along with Intense Bcr-Abl Kinase Inhibition May Eradicate CML Progenitor Cells." Blood 114, no. 22 (November 20, 2009): 3250. http://dx.doi.org/10.1182/blood.v114.22.3250.3250.
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Abstract Abstract 3250 Poster Board III-1 Preclinical studies of imatinib set the paradigm of continuous Bcr-Abl kinase inhibition for optimal response in chronic myeloid leukemia (CML). However, the clinical success of once daily dasatinib, despite its short serum half life, implies that intermittent inhibition of Bcr-Abl kinase activity is sufficient for clinical response. In vitro studies also demonstrated that short-term intense (≥90%) Bcr-Abl kinase inhibition triggers cell death in BCR-ABL + cell lines, demonstrating their oncogene addiction. However, the effect of short-term intense kinase inhibition on CD34+ CML progenitors is not studied. Clinical, mathematical modelling and in vitro studies suggest that leukemic stem cells (LSC) are difficult to eradicate and hence the majority of CML patients may not be cured with tyrosine kinase inhibitors (TKI). Inadequate Bcr-Abl kinase inhibition has been postulated to cause refractoriness of LSC to TKI's. This may be due to increased expression of ABCB1 and ABCG2 efflux proteins, or the quiescent state of LSC. However, the phenomenon could be independent of Bcr-Abl kinase activity. In vivo leukemic progenitors live in a cytokine rich environment which may be providing a mechanism for Bcr-Abl independent resistance. We have assessed the impact of short-term intense Bcr-Abl kinase inhibition on CML cell lines and CML CD34+ primary cells in the presence and absence of cytokines. In CML cell lines, short-term (cells were cultured with dasatinib for 30 min and following thorough drug washout, cells were recultured in drug free media for 72 hr) intense Bcr-Abl kinase inhibition with 100 nM dasatinib triggers cell death. In CML-CD34+ cells 30 min of culture with 100 nM dasatinib (n=13) or 30 μM IM (n=7) reduced the level of p-Crkl (surrogate marker of Bcr-Abl kinase activity) by 97±3% and 96±4% respectively. In the presence of either a six growth factors cocktail (6-GF; n=10) or GM-CSF (n=11) or G-CSF (n=4) alone, despite 97% inhibition of p-Crkl, short-term culture with 100 nM dasatinib (D100ST) reduced colony forming cells (CFC) by only 24%, 32% or 5%, respectively. However without cytokines, D100ST reduced CML-CD34+ CFCs by 70%. Consistent with the results observed with dasatinib, short-term culture with 30 μM imatinib (IM) (n=3) also reduced 90% CFC in the absence of cytokines but by only 38% in the presence of 6-GF. These results suggest that in CML-CD34+ cells, GM-CSF, G-CSF or 6-GF mediate Bcr-Abl independent TKI resistance. It is possible that cytokines may be promoting cell survival via signalling pathways that are refractory to dasatinib. To examine this possibility, we assessed the effect of D100ST on p-STAT5 signalling in CML-CD34+ cells, in the presence and absence of GM-CSF, G-CSF or 6-GF. STAT5 was constitutively phosphorylated in CML-CD34+ cells, and in the absence of cytokines, D100ST reduced the p-STAT 5. STAT5 phosphorylation was not inhibited by D100ST when cells were cultured with 6-GFs or GM-CSF however, the combination of D100ST and a Janus kinase (Jak) inhibitor dramatically reduced p-STAT5. Similarly, in the presence of GM-CSF (32.35±5.16% vs. 68.33±14.90%) or G-CSF (58.13±13 vs. 94.68±21.12) combination of D100ST and JAK inhibitor significantly reduced CFC compared to D100ST only. Thus our data suggest that in contrast to CML cell lines, primary CML progenitors may not be completely dependent on the BCR-ABL oncogene and that activation of the cytokine mediated JAK-2/STAT-5 pathway may circumvent the need for BCR-ABL signalling for maintenance of survival. Thus a therapeutic strategy based on short-term intense kinase inhibition may have limited success unless critical redundant cytokine-induced survival pathways are also inhibited. We postulate that blockade of cytokine signalling along with short-term intense Bcr-Abl kinase inhibition with a potent second generation TKI may provide a novel strategy to eradicate primitive CML cells. Fig 1 In CML-CD34+ cells, Jak kinase inhibition abrogates the rescuing effect of cytokines on cell death induced by BCR-ABL blockade: In the absence of cytokines (No GF, n=11) short-term culture with 100 nM dasatinib (D100ST) reduced CFCs by 67% of control, however in the presence of 6-GFs (n=10), GM-CSF (n=10) or G-CSF (n=4) it could reduce CFCs by only 24%, 32% or 5% of control respectively (B) In the presence of GM-CSF (n= 4) or G-CSF (n= 4), combination of Jak inhibition and D100ST reduced CFC compared to dasatinib alone. Fig 1. In CML-CD34+ cells, Jak kinase inhibition abrogates the rescuing effect of cytokines on cell death induced by BCR-ABL blockade: In the absence of cytokines (No GF, n=11) short-term culture with 100 nM dasatinib (D100ST) reduced CFCs by 67% of control, however in the presence of 6-GFs (n=10), GM-CSF (n=10) or G-CSF (n=4) it could reduce CFCs by only 24%, 32% or 5% of control respectively (B) In the presence of GM-CSF (n= 4) or G-CSF (n= 4), combination of Jak inhibition and D100ST reduced CFC compared to dasatinib alone. Disclosures: White: Novartis and Britol-Myers Squibb: Research Funding. Hughes:BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.
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Busch, Caroline, and Helen Wheadon. "Bone marrow niche crosses paths with BMPs: a road to protection and persistence in CML." Biochemical Society Transactions 47, no. 5 (September 24, 2019): 1307–25. http://dx.doi.org/10.1042/bst20190221.
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Abstract Chronic myeloid leukaemia (CML) is a paradigm of precision medicine, being one of the first cancers to be treated with targeted therapy. This has revolutionised CML therapy and patient outcome, with high survival rates. However, this now means an ever-increasing number of patients are living with the disease on life-long tyrosine kinase inhibitor (TKI) therapy, with most patients anticipated to have near normal life expectancy. Unfortunately, in a significant number of patients, TKIs are not curative. This low-level disease persistence suggests that despite a molecularly targeted therapeutic approach, there are BCR-ABL1-independent mechanisms exploited to sustain the survival of a small cell population of leukaemic stem cells (LSCs). In CML, LSCs display many features akin to haemopoietic stem cells, namely quiescence, self-renewal and the ability to produce mature progeny, this all occurs through intrinsic and extrinsic signals within the specialised microenvironment of the bone marrow (BM) niche. One important avenue of investigation in CML is how the disease highjacks the BM, thereby remodelling this microenvironment to create a niche, which enables LSC persistence and resistance to TKI treatment. In this review, we explore how changes in growth factor levels, in particular, the bone morphogenetic proteins (BMPs) and pro-inflammatory cytokines, impact on cell behaviour, extracellular matrix deposition and bone remodelling in CML. We also discuss the challenges in targeting LSCs and the potential of dual targeting using combination therapies against BMP receptors and BCR-ABL1.
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Zhao, Helong, Anthony D. Pomicter, Anna M. Eiring, Anca Franzini, Jonathan Ahmann, Jae-Yeon Hwang, Anna Senina, et al. "MS4A3 promotes differentiation in chronic myeloid leukemia by enhancing common β-chain cytokine receptor endocytosis." Blood 139, no. 5 (February 3, 2022): 761–78. http://dx.doi.org/10.1182/blood.2021011802.
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Abstract The chronic phase of chronic myeloid leukemia (CP-CML) is characterized by the excessive production of maturating myeloid cells. As CML stem/progenitor cells (LSPCs) are poised to cycle and differentiate, LSPCs must balance conservation and differentiation to avoid exhaustion, similar to normal hematopoiesis under stress. Since BCR-ABL1 tyrosine kinase inhibitors (TKIs) eliminate differentiating cells but spare BCR-ABL1-independent LSPCs, understanding the mechanisms that regulate LSPC differentiation may inform strategies to eliminate LSPCs. Upon performing a meta-analysis of published CML transcriptomes, we discovered that low expression of the MS4A3 transmembrane protein is a universal characteristic of LSPC quiescence, BCR-ABL1 independence, and transformation to blast phase (BP). Several mechanisms are involved in suppressing MS4A3, including aberrant methylation and a MECOM-C/EBPε axis. Contrary to previous reports, we find that MS4A3 does not function as a G1/S phase inhibitor but promotes endocytosis of common β-chain (βc) cytokine receptors upon GM-CSF/IL-3 stimulation, enhancing downstream signaling and cellular differentiation. This suggests that LSPCs downregulate MS4A3 to evade βc cytokine-induced differentiation and maintain a more primitive, TKI-insensitive state. Accordingly, knockdown (KD) or deletion of MS4A3/Ms4a3 promotes TKI resistance and survival of CML cells ex vivo and enhances leukemogenesis in vivo, while targeted delivery of exogenous MS4A3 protein promotes differentiation. These data support a model in which MS4A3 governs response to differentiating myeloid cytokines, providing a unifying mechanism for the differentiation block characteristic of CML quiescence and BP-CML. Promoting MS4A3 reexpression or delivery of ectopic MS4A3 may help eliminate LSPCs in vivo.
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Cortes, Jorge E., Dong-Wook Kim, Javier Pinilla-Ibarz, Philipp D. le Coutre, Ronald Paquette, Charles Chuah, Franck E. Nicolini, et al. "Evaluation of the Benefit/Risk Profile of Ponatinib in CP-CML Patients over Time: 4-Year Follow-up of the Phase 2 PACE Study." Blood 126, no. 23 (December 3, 2015): 5142. http://dx.doi.org/10.1182/blood.v126.23.5142.5142.
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Abstract Introduction: Ponatinib, a potent oral tyrosine kinase inhibitor (TKI), is approved for use in patients with refractory CML and Ph+ ALL, including patients with the BCR-ABLT315I mutation. This post hoc analysis was undertaken to assess the evolution of the efficacy and safety of ponatinib over time in CP-CML patients in the ongoing phase 2 PACE trial (NCT01207440). Updated data with 4 years of follow-up, including an analysis of the management of select adverse events (AEs), will be presented. Methods: Patients with CML/Ph+ ALL who were resistant or intolerant to dasatinib or nilotinib or who had the T315I mutation (N=449) were enrolled and treated with ponatinib (starting dose 45 mg once daily). Accumulation of arterial occlusive events (AOEs) in the ponatinib program led to recommendations for dose reduction in PACE in October 2013. Efficacy and safety among CP-CML patients are reported as of February 2, 2015 in this abstract. Results: Included in the analysis were 270 CP-CML patients who received treatment, with a median follow-up of 42 (0.1-52.5) months. At the time of analysis, 42% of CP-CML patients (114/270) continued to receive ponatinib. Primary reasons for discontinuation were AEs (18%), withdrawal by patient (11%), and disease progression (10%). Characteristics of CP-CML patients continuing to receive ponatinib were similar to those of the overall cohort (Table 1). Of 267 CP-CML patients analyzed for efficacy, 59% and 53% achieved MCyR and CCyR at any time, and 39% and 23% achieved at least MMR and MR4.5, respectively; despite dose reductions from 45 mg/d at baseline, responses continued to deepen after the first year (Table 2). The probability of maintaining MCyR at 3 years was 83%, and the estimated PFS/OS rates were 60%/81% at 3 years. AEs in ≥20% of CP-CML patients were rash (46%), abdominal pain (46%), thrombocytopenia (45%), headache (43%), dry skin (41%), constipation (41%), hypertension (33%), arthralgia (32%), fatigue (30%), nausea (27%), increased lipase (26%), pyrexia (26%), myalgia (24%), pain in an extremity (21%), and back pain (20%). Most newly occurring AEs were observed within the first year. Rates of AOEs (any/serious) were 28%/23%, including cardiovascular (14%/11%), cerebrovascular (11%/9%), and peripheral vascular (11%/8%) events; median time to onset for AOEs was 14.1 (0.3-44.0) months. Exposure-adjusted incidence rates of new AOEs (events per 100 patient-years) have appeared to remain approximately stable or decrease over time; rates were 14.3 in Year 1, 15.2 in Year 2, 11.7 in Year 3, and 9.9 in Year 4 (analysis for Year 4 does not yet cover a full fourth year for all CP-CML patients). Conclusions: With longer follow-up in the PACE trial, the benefit/risk profile of ponatinib continues to evolve and remains favorable for many patients in this trial. In this analysis, the baseline characteristics of patients remaining on study did not differ substantially across the time points examined. Despite a decline from initial dose intensity, deep and lasting responses have continued to be observed after the first year in this trial of heavily pretreated patients. Most AEs have occurred early in treatment; AOEs have typically had a later onset-exposure-adjusted incidence rates of AOEs have not increased over time. Table 1. Characteristics of CP-CML Patients Continuing to Receive Ponatinib Over Time Baseline (n=270) Patients Receiving Ponatinib at 1 Year (n=179) Patients Receiving Ponatinib at 2 Years (n=156) Patients Receiving Ponatinib at 3 Years (n=125) Median age at baseline, years 60 58 58 58 ≥2 prior TKIs at baseline, % 93 91 92 92 ≥3 prior TKIs at baseline, % 60 55 54 54 Median time from diagnosis to first dose, years 7.0 6.9 7.0 6.7 Median dose intensity, mg/d 45.0 29.1 29.0 28.8 Mutations detected at baseline, % 49 49 48 47 T315I mutation detected at baseline, % 24 25 23 22 Table 2. Cumulative Response Rates in CP-CML Patients Receiving Ponatinib Over Time (n=267) By 1 Year, % By 2 Years, % By 3 Years, % Total, % MCyR 55 58 59 59 CCyR 50 53 53 53 MMR 30 36 39 39 MR4.5 9 16 22 23 Disclosures Cortes: Pfizer: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; ARIAD Pharmaceuticals Inc.: Consultancy, Research Funding; Teva: Consultancy, Research Funding. Pinilla-Ibarz:Pfizer: Consultancy, Other: Consulting & Advisory Role, Research Funding, Speakers Bureau; Novartis: Consultancy, Other: Consulting & Advisory Role, Research Funding; Teva: Consultancy, Speakers Bureau; ARIAD Pharmaceuticals, Inc.: Consultancy, Other: Consulting & Advisory Role, Research Funding; BMS: Consultancy, Honoraria, Other: Consulting & Advisory Role, Speakers Bureau. le Coutre:Novartis: Honoraria; ARIAD Pharmaceuticals Inc.: Honoraria; BMS: Honoraria; Pfizer: Honoraria. Paquette:ARIAD Pharmaceuticals, Inc.: Honoraria, Speakers Bureau; BMS: Honoraria, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Incyte: Honoraria, Speakers Bureau; Astra-Zeneca: Consultancy. Chuah:Children International: Honoraria; Novartis: Honoraria; Bristol Meyers Squibb: Honoraria. Nicolini:Ariad Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Apperley:BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ARIAD: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Talpaz:Pfizer: Consultancy, Other: Travel/Expenses, Research Funding; Novartis: Consultancy, Other: Travel/Expenses, Research Funding; ARIAD Pharmaceuticals, Inc.: Consultancy, Other: Travel/Expenses, Research Funding; Sanofi: Research Funding; Incyte: Other: Travel/Expenses, Research Funding. DeAngelo:Incyte: Consultancy; Novartis: Consultancy; Bristol Myers Squibb: Consultancy; Pfizer: Consultancy; Celgene: Consultancy; Ariad: Consultancy; Agios: Consultancy; Amgen: Consultancy. Abruzzese:Pfizer: Other: Consulting or Advisory Role; Novartis: Other: Consulting or Advisory Role; BMS: Other: Consulting or Advisory Role; ARIAD Pharmaceuticals Inc.: Other: Consulting & Advisory Role. Rea:Bristol-Myers Squibb: Honoraria; Novartis: Honoraria; Pfizer: Honoraria; Ariad: Honoraria. Baccarani:NOVARTIS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ARIAD Pharmaceuticals, Inc.: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; PFIZER: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Müller:BMS: Honoraria, Other: Consulting or Advisory Role, Research Funding; Novartis: Honoraria, Other: CONSULTING OR ADVISORY ROLE, Research Funding; ARIAD Pharmaceuticals Inc.: Honoraria, Other: Consulting & Advisory Role, Research Funding. Lustgarten:ARIAD Pharmaceuticals Inc.: Employment, Equity Ownership, Other: Stock. Conlan:ARIAD Pharmaceuticals Inc.: Other: Stock. Guilhot:Novartis: Honoraria, Other: TRAVEL, ACCOMODATIONS, EXPENSES; Pfizer: Honoraria; Celgene: Consultancy, Other: CONSULTING OR ADVISORY ROLE. Deininger:BMS: Consultancy, Honoraria, Other: Consulting & Advisory Role, Research Funding; Novartis: Consultancy, Honoraria, Other: Consulting or Advisory Role, Research Funding; Celgene: Research Funding; Gilead: Research Funding; ARIAD Pharmaceutical Inc.: Consultancy, Honoraria, Other: Consulting or Advisory Role; Pfizer: Consultancy, Honoraria, Other: Consulting or Advisory Role; Incyte: Consultancy, Honoraria, Other: Consulting or Advisory Role. Hochhaus:Bristol-Myers Squibb: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding. Hughes:ARIAD: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Kantarjian:Novartis: Research Funding; BMS: Research Funding; Pfizer: Research Funding; Amgen: Research Funding. Shah:ARIAD Pharmaceuticals, Inc.: Research Funding; Bristol-Myers Squibb: Research Funding; Pfizer: Research Funding; Plexxikon: Research Funding; Daiichi-Sankyo: Research Funding.
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Jabbour, Elias J., Jorge E. Cortes, Moshe Talpaz, Michele Baccarani, Michael J. Mauro, Andreas Hochhaus, Timothy P. Hughes, et al. "Long-Term Follow-up of the Efficacy and Safety of Ponatinib in Philadelphia Chromosome-Positive Leukemia Patients with the T315I Mutation." Blood 128, no. 22 (December 2, 2016): 3067. http://dx.doi.org/10.1182/blood.v128.22.3067.3067.
Full textAbstract:
Abstract Background: Ponatinib is a tyrosine kinase inhibitor (TKI) approved for adult patients (pts) with relapsed/refractory CML or Ph+ ALL and those with the BCR-ABL T315I mutation, which is uniformly resistant to other TKIs approved for the treatment of CML and Ph+ ALL. Prior to the availability of ponatinib, resistant pts with the T315I mutation (T315I+) had worse outcomes than those without the mutation; in a matched pair analysis in chronic phase (CP)-CML pts, median overall survival (OS) was 48.4 mos after development of resistance in T315I+ pts versus not reached in those without the mutation (Nicolini FE, et al. Haematologica 2013). Methods: We evaluated the efficacy and safety of ponatinib in a pooled analysis of a subgroup of CP-CML pts with the T315I mutation (detected in a central laboratory by Sanger sequencing at baseline) from the phase 1 (NCT00660920) and pivotal phase 2 PACE (NCT01207440) trials. In addition, we evaluated the impact of continuation of ponatinib treatment at a 2-yr landmark time point on OS at 1-yr past the landmark in T315I+ CP-CML pts in PACE. The phase 1 trial is an open-label, dose escalation study of ponatinib (starting dose 2-60 mg once daily) in 81 adults with relapsed/refractory hematologic malignancies. PACE is an open-label, single-arm trial of ponatinib (starting dose 45 mg daily) in 449 adults with CML or Ph+ ALL resistant or intolerant to dasatinib or nilotinib or with the T315I mutation. Dose reductions were instructed in Oct 2013 in response to an accumulation of arterial occlusive events (AOEs) reported with longer follow-up across the ponatinib clinical program. Response assessments included major cytogenetic response (MCyR), complete cytogenetic response (CCyR), major molecular response (MMR; assessed in a central laboratory), and molecular response 4.5 (MR4.5). OS and progression-free survival (PFS) data were only collected in PACE; 3-yr outcomes were examined for all evaluable T315I+ CP-CML pts, along with a log-rank test for OS by treatment status as of the 2-yr landmark time point. Exposure-adjusted incidence rates of new AOEs are reported as number of events/100 pt-yrs. Pooled data as of February 2, 2015 is reported here. Results: There were 76 T315I+ CP-CML pts included in this analysis (phase 1, n=12; PACE, n=64). At the time of analysis, median duration of follow-up in T315I+ CP-CML pts was 40 (range: 1.5-74) mos; 37 pts (49%) remain on study. Median baseline ponatinib dose intensity was 33 (range: 5-56) mg daily; 25/37 (68%) ongoing pts were receiving 15 mg daily as their current dose as of the data cut-off. Primary reasons for discontinuation in T315I+ CP-CML pts were disease progression [10/76 (13%)], adverse events [AEs; 9/76 (12%)], consent withdrawn [5/76 (7%)], physician/administrative decision [4/76 (5%)], death [3/76 (4%)], lack of efficacy [2/76 (3%)], and other [6/76 (8%)]; criteria for disease progression included death, development of advanced phase CML, loss of CHR (in absence of cytogenetic response) and loss of MCyR. Cumulative response rates in T315I+ CP-CML pts (n=76) were: MCyR, 75%; CCyR, 72%; MMR, 61%, and MR4.5, 37%. In PACE, estimated 3-yr PFS/OS rates for 64 T315I+ CP-CML pts were 60%/78% (medians not reached). One yr post-landmark outcomes for T315I+ CP-CML pts by treatment status at the 2-yr landmark time point are displayed in the table. Most common treatment-emergent AEs (≥40%) in the pooled group of T315I+ CP-CML pts (n=76 phase 1 and PACE) were: rash, 55%; dry skin, 49%; headache, 46%; abdominal pain, 43%; nausea, 41%; and fatigue, 41%. Among these pts, the cumulative incidence of any AOE (grade 3/4) was 32% (20%); by subcategory: cardiovascular 20% (15%), cerebrovascular 12% (5%), and peripheral vascular 13% (8%) events. Two T315I+ CP-CML pts had grade 5 AOEs. The exposure-adjusted incidence rate of new AOEs in T315I+ CP-CML pts was 12/100 pt-yrs. Conclusions: Ponatinib continues to provide deep and durable responses with >3 yrs median follow-up in T315I+ CP-CML pts. Survival outcomes with ponatinib treatment in these highly refractory pts were high overall and compare favorably with those observed in T315I+ CP-CML pt populations prior to the availability of ponatinib. Although pt numbers are limited for the landmark analysis, continuation of ponatinib treatment at 2 yrs was associated with a trend for improved OS, where data continue to mature. Updated data in all T315I+ pts will be presented. Study sponsor: ARIAD Pharmaceuticals, Inc. Disclosures Jabbour: ARIAD: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Novartis: Research Funding; BMS: Consultancy. Cortes:ARIAD: Consultancy, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding. Talpaz:Incyte Corporation: Other: Travel expense reimbursement, Research Funding; Novartis: Research Funding; Ariad: Other: Expense reimbursement, travel accomodation expenses, Research Funding; Pfizer: Consultancy, Other: travel accomodation expenses, Research Funding. Baccarani:ARIAD: Consultancy, Honoraria, Other: travel, accommodations, expenses , Speakers Bureau; BMS: Honoraria, Speakers Bureau; Novartis: Honoraria, Other: travel, accommodations, expenses , Speakers Bureau; Pfizer: Honoraria, Speakers Bureau. Mauro:BMS: Consultancy, Honoraria; ARIAD: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria. Hochhaus:Pfizer: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding. Hughes:Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Ariad: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Consultancy, Honoraria. Guilhot:ARIAD: Honoraria. Deininger:BMS: Consultancy, Research Funding; Gilead: Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; CTI BioPharma Corp.: Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Ariad: Consultancy, Membership on an entity's Board of Directors or advisory committees. Shah:ARIAD: Research Funding; Daiichi-Sankyo: Research Funding; BMS: Research Funding; Pfizer: Research Funding; Plexxikon: Research Funding. Flinn:Janssen: Research Funding; Gilead Sciences: Research Funding; Pharmacyclics LLC, an AbbVie Company: Research Funding; ARIAD: Research Funding; RainTree Oncology Services: Equity Ownership. Lustgarten:ARIAD: Employment, Equity Ownership. Rivera:ARIAD: Employment, Equity Ownership. Santillana:ARIAD: Employment, Equity Ownership. Kantarjian:Amgen: Research Funding; ARIAD: Research Funding; Bristol-Myers Squibb: Research Funding; Pfizer Inc: Research Funding; Delta-Fly Pharma: Research Funding; Novartis: Research Funding.
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Nievergall, Eva, Deborah L. White, Agnes S. M. Yong, Hayley S. Ramshaw, Samantha J. Busfield, Gino Vairo, Angel F. Lopez, Timothy P. Hughes, and Devendra K. Hiwase. "Effective Elimination of CML Progenitor and Stem Cells Through Combination of α-CD123 Antibody-Dependent Cell-Mediated Cytotoxicity and Tyrosine Kinase Inhibitor Treatment." Blood 120, no. 21 (November 16, 2012): 32. http://dx.doi.org/10.1182/blood.v120.21.32.32.
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Abstract Abstract 32 Since the introduction of tyrosine kinase inhibitor (TKI) therapy overall survival and complete molecular response rates in chronic phase chronic myeloid leukemia (CP-CML) patients have significantly improved. However, leukemic stem cells (LSCs) and progenitor cells persist and are thought to be responsible for disease progression, development of TKI resistance and disease recurrence after stopping TKI therapy. Protection by cytokines, such as IL-3 and GM-CSF, provides a potential mechanism of LSC resistance. While in acute myeloid leukemia (AML) monoclonal antibody (mAb) targeting of IL-3 receptor α (CD123), a recognized marker for AML LSCs, has been studied in vitro and in vivo, similar investigations have not been undertaken in CML to date. CSL362 is a genetically-engineered form of the specific blocking mAb 7G3 optimized for Fc receptor binding to achieve maximal antibody-dependent cell-mediated cytotoxicity (ADCC) capacity. Here we investigate the expression of CD123 in CD34+ progenitors and CD34+CD38− LSCs, isolated from CP- and blast crisis (BC) - CML patients, and study the benefits of targeting those cells by CSL362 alone and in combination with TKIs. Flow cytometry analysis established significantly elevated expression of CD123 on CD34+CD38− cells from CP-CML (53.0 ± 5.8 %, n=16, p=0.003) and BC-CML (73.2 ± 6.7 %, n=18, p<0.001) patients compared to normal donors (20.3 ± 4.2 %, n=8), with clear increases in CD123 expression with disease progression in matched samples (n=2). Subsequent assessment of apoptosis, colony forming unit (CFU-GM) and long-term culture-initiating cell (LTC-IC) potential confirmed the ability of CSL362 to block IL-3-mediated rescue of TKI-induced cell death. However, in the presence of other cytokines, likely found in the physiological bone marrow microenvironment, this effect was lost. We also demonstrate by lactate dehyrogenase release and clonogenic assays that CML CD34+ cell numbers are significantly reduced, in a dose-dependent manner, by CSL362-induced ADCC employing NK cells from healthy donors (42.4 ± 8.1 % lysis, n=3, and CFU-GM decreased to 30 ± 10.8 % of controls, n=5, p=0.003). In keeping with this, flow cytometry analysis revealed specific elimination of CP- and BC-CML CD123+ CD34+CD38− cells (from 42.9 % to 18.6 %, n=5, p=0.004, and from 71 % to 35.3 %, n=3, p=0.044, respectively). Importantly, autologous CML patient NK cells, collected after achievement of major molecular response, also mediate CSL362-dependent cytotoxicity similar to allogeneic healthy donor NK cells as indicated by equivalent numbers of remaining CFUs (28 ± 6.7 % vs. 34.9 ± 3.4 %, n=5, Fig. A). We further have evidence to suggest preferential elimination of CML over normal LTC-ICs (30.3 ± 9.9 % vs. 62.6 ± 11.2 % remaining, n=3, p=0.096) in the autologous setting. Of clinical importance, the combination of Nilotinib and CSL362 resulted in a significantly greater reduction in CFUs (additive effect) when compared to either agent alone (Fig. B). Taken together these data suggest that selective ADCC-mediated lysis, likely the major mode of action of CSL362 in vivo, efficiently eliminates CML progenitor and stem cells. Promising results evaluating CSL362/TKI combination treatments, with the expectation to further enhance specificity for leukemic while sparing normal progenitor and stem cells as indicated from preliminary experiments, warrant further studies. A: Autologous NK cells are able to confer CSL362-induced ADCC against CML CD34+ cells. Cells were co-cultured at an effector to target cell ratio (E:T) of 10:1 in the absence and presence of CSL362 as indicated for 4 h and remaining CFU-GM were enumerated. Data is normalized to target cells alone (*** p<0.001). B: CSL362-mediated ADCC and TKI treatment show additive effects. CP-CML CD34+ cells were cultured with nilotinib at varying concentrations as indicated for 48 h before overnight exposure to CSL362 (1 μg/ml) with or without allogeneic NK cells (E:T 1:1). Mean ± SE of CFU-GM colony numbers is shown (n=3, * p<0.05, ** p<0.01). A: Autologous NK cells are able to confer CSL362-induced ADCC against CML CD34+ cells. Cells were co-cultured at an effector to target cell ratio (E:T) of 10:1 in the absence and presence of CSL362 as indicated for 4 h and remaining CFU-GM were enumerated. Data is normalized to target cells alone (*** p<0.001). . / B: CSL362-mediated ADCC and TKI treatment show additive effects. CP-CML CD34+ cells were cultured with nilotinib at varying concentrations as indicated for 48 h before overnight exposure to CSL362 (1 μg/ml) with or without allogeneic NK cells (E:T 1:1). Mean ± SE of CFU-GM colony numbers is shown (n=3, * p<0.05, ** p<0.01). Disclosures: Nievergall: CSL Ltd: Research Funding. White:BMS: Research Funding; CSL Ltd: Research Funding; Novartis Oncology: Honoraria, Research Funding. Ramshaw:CSL Ltd: Research Funding. Busfield:CSL Ltd: Employment. Vairo:CSL Ltd: Employment. Lopez:CSL Ltd: Research Funding. Hughes:Ariad: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis Oncology: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; CSL Ltd: Research Funding. Hiwase:CSL Ltd: Research Funding.
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Hernesniemi, Sari, Jukka Vakkila, Kimmo Porkka, and Satu Mustjoki. "Single-Cell Profiling of Aberrant Cytokine Signaling in Patients with Chronic Myeloid Leukemia (CML) at Diagnosis and during Dasatinib Therapy." Blood 112, no. 11 (November 16, 2008): 4214. http://dx.doi.org/10.1182/blood.v112.11.4214.4214.
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Abstract Aberrant cytokine and growth factor signaling is the hallmark of CML and results from constitutive oligomerization of the oncogenic BCR-ABL tyrosine kinase (TK). Inhibition of BCR-ABL by imatinib mesylate is the current standard of care of CML and results in durable responses in majority of patients. However, a proportion of patients shows primary or secondary resistance to imatinib, which can be attributed either to selection of clones harboring mutations in the kinase domain of BCR-ABL or activation of a BCR-ABL independent pathway. Dasatinib, a potent multikinase inhibitor, can rescue some imatinib-resistant patients, but carries an increased risk of adverse effects due to inhibition of off-target wild-type kinases, particularly in immune effector cells. In concord, recent in vitro data indicate a profound immunosuppressive effect of dasatinib. The aim of this study was to analyze and predict TK inhibitor (TKI) resistance and off-target effects using single-cell profiling of aberrant phosphoprotein networks upon cytokine stimulus by multiparameter flow cytometry. The study cohort consisted of 5 healthy controls, 4 non-treated CML patients at diagnosis and 5 CML patients on dasatinib therapy and in cytogenetic remission. Stimuli included GM-CSF, IL-4+IL-6+IFNγ and IL-2+IL-10+IFNα and they were added to freshly drawn whole blood or bone marrow. The readout phosphoproteins were pERK1/2, pSTAT1, pSTAT3, pSTAT5a and pSTAT6 (with isotype controls), and were analyzed separately from granulocytes, monocytes, CD3+, CD4+ and CD8+ lymphocytes and regulatory T-cells. In unstimulated blood samples from healthy controls the phosphoproteins were essentially unphosphorylated. The responses to cytokines were consistent among individuals resulting in phosphorylation of ERK1/2, STAT3 and STAT5a on GM-CSF stimulus, STAT-1, STAT-3 and STAT-5a on IL-2+IL10+IFNα and STAT-1, STAT-3 and STAT-6 on IL4+IL6+IFN-γ. Compared to healthy controls, increased baseline phosphorylation of STAT-1, STAT-3 and STAT5a, but not ERK1/2 was seen in CML patients at diagnosis, especially in myeloid cell lineages (neutrophils/monocytes), but also in lymphocyte subgroups. The responses to cytokine stimulation were modest overall, in particular the ERK1/2 responses to GM-CSF were absent. This indicated the inactivation of the Ras/MEK/MAPK pathway and saturation of other BCR-ABL downstream pathways. Already at diagnosis, the phosphorylation pattern of a TKI primary resistant patient differed profoundly from the responding patients. Marked activation of STAT-1 and STAT-3 was seen in granulocytes and monocytes stimulated either by GM-CSF or by combination of IL2+IL10+IFN-α, suggesting activation by a pathway circumventing BCR-ABL. In dasatinib treated patients, the baseline activation status was similar in granulocytes and monocytes and slightly diminished in lymphocytes when compared to healthy controls. Similarly, the responses to cytokines resembled those seen in healthy controls, in contrast to published in vitro data. Remarkably, in some of the dasatinib treated patients, STAT1 and STAT3 responses were even more pronounced than in healthy controls. This underlines the importance of studying the in vivo/ex vivo effects of TKIs on off-target kinases, in particular of drugs with a short half-life such as dasatinib. In conclusion, inter-individual differences in TKI response and immunomodulatory effects of pan-TKI dasatinib can readily be discerned by analyzing key intracellular phosphoprotein responses to cytokine and growth factor stimuli ex vivo. The method allows profiling of aberrant signaling pathways in different subsets of leukocytes in CML patients and can be used to predict TKI resistance and spectrum of potential adverse effects due to inhibition of wildtype targets. Similar analyses of signaling pathways at the stem cell level are ongoing and may aid in understanding TKI resistance of CML stem cells.
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Held, Stefanie AE, Anita Bringmann, Annkristin Heine, Grethe Kock, Ruth-Miriam Körber, Nina Manon Seltmann, and Peter Brossart. "Interferon Gamma (IFNγ) Interferes with the Effects of Tyrosine Kinase Inhibitors (TKI) In CML Cells." Blood 116, no. 21 (November 19, 2010): 3393. http://dx.doi.org/10.1182/blood.v116.21.3393.3393.
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Abstract Abstract 3393 Introduction: Chronic myeloid leukemia (CML) is a myeloproliferative disease which resolves in the constitutive activity of the BCR-ABL protein. Since the introduction of imatinib as first line therapy for CML, the overall survival of patients could be significantly extended. However, mutations of the BCR-ABL protein and environmental effects mediated by adhesion molecules, cytokines and growth factors were shown to induce resistance to TKIs. For the development of immunotherapeutic approaches that combine TKIs with vaccination strategies or donor lymphocyte infusions it is important to understand the mechanisms of possible interactions and inhibition. Therefore, we thought to determine the role of activated T cells and to analyze a possible antiapoptotic effect of released IFNγ on the CML cells. Methods: CML cell lines and primary cells from patients with CML were treated with imatinib or nilotinib and incubated with activated (PMA/ionomycin) or resting T cells in transwell experiments. Induction of apoptosis was analyzed by flow cytometry. In addition, IFNγ was used together with TKIs and apoptotic cell death and proliferation was determined. Immunoblotting was performed to analyze the involved pathways. Result: Incubation of CML cells in transwell assays with activated but not resting T lymphocytes resulted in significant inhibition of the apoptosis induction by imatinib and nilotinib. To further determine the mechanisms mediating these effects, CML cells were treated with IFNγ in addition to the TKIs. In line with the results from previous experiments, IFNγ reduced the rate of cell death and proliferation inhibition mediated by imatinib and nilotinib. Interestingly, in contrast to IFNα, IFNγ inhibited the TKIs induced downregulation of MHC-class I molecules on CML cells. In line with these results, Western blot analysis of K-562 cells incubated with imatinib or nilotinib showed an increased caspase-3 activation and PARP cleavage as well as reduced down regulation of anti-apoptotic molecules such as xIAP as compared to IFNγ treated cells. Furthermore, we observed a diminished TKI induced downregulation of Jak-2 and STAT-5 phosphorylation as well as increased nuclear expression of RUNX-1 in cells treated with IFNγ. Conclusion: Our results demonstrate that IFNγ released by activated T-cells can interfere with the action of TKIs in CML patients and might have important implications in the development of immunotherapeutic approaches. Disclosures: No relevant conflicts of interest to declare.
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Okabe, Seiichi, Tetsuzo Tauchi, Yuko Tanaka, and Kazuma Ohyashiki. "Interleukin 1 Beta As a Promising Therapeutic Target in ABL Tyrosine Kinase Inhibitor Resistant Chronic Myeloid Leukemia Cells." Blood 132, Supplement 1 (November 29, 2018): 4249. http://dx.doi.org/10.1182/blood-2018-99-114030.
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Abstract Introduction: Although ABL tyrosine kinase inhibitor (TKI), imatinib, nilotinib and dasatinib have demonstrated the potency against chronic myeloid leukemia (CML) and Philadelphia chromosome (Ph) positive acute lymphoblastic leukemia (Ph+ALL) patients, resistance to ABL TKI can develop in the many patients. It has already reported that ABL kinase domain mutations have been implicated in the pathogenesis of ABL TKI resistance, however, it is fully not known the molecular mechanism of drug resistance ABL TKIs. Therefore, new approach against ABL TKI resistant cells may improve the outcome of Ph-positive leukemia patients. Interleukin-1 (IL-1) is a proinflammatory cytokine and central mediator of innate immunity. IL-1β also controls essential cell responses. Because enhanced IL-1β signaling is reported in patients of hematological malignancies, IL-1β may be the promising therapeutic value in ABL TKI resistant CML patients. Materials and methods: In this study, we established ABL TKI-resistant in vitro cell line models (K562 imatinib-R, K562 nilotinib-R, K562 dasatinib-R, K562 ponatinib-R). We also investigated whether IL-1β was involved in ABL TKI resistant Ph-positive leukemia cells and cytokines were induced by IL-1β in human umbilical vein endothelial cells (HUVEC). Results: We analyzed the relationship of IL-1β signaling pathways and ABL TKI sensitivity by microarray gene expression data from the online Gene Expression Omnibus (GEO). IL-1β is related to imatinib sensitivity and resistant in CML patients from the public microarray datasets of GSE14671. We next examined ABL TKI resistant cell lines (K562 imatinib-R, K562 nilotinib-R, K562 dasatinib-R, K562 ponatinib-R) in this study. BCR-ABL point mutation was not found in ABL TKI resistant cells. BCR-ABL expression levels were not increased in ABL TKI resistant K562 cells. These cells were highly resistant to ABL TKIs compare to K562 cells (K562 imatinib-R: imatinib 2μM, nilotinib-R: nilotinib 2μM, dasatinib-R: dasatinib 100nM, ponatinib-R: ponatinib 50nM). We investigated gene expression profiles in cultured ABL TKI resistant K562 cells by DNA microarray. We found gene expression of IL-1β and IL-1β mediated signaling pathway was increased ABL TKI resistant K562 cells. IL-1β gene amplification was confirmed by RT-PCR analysis. Nuclear Factor kappa-light-chain-enhancer of activated B cells (NF-kB) acts as a central mediator of inflammatory responses. Because bortezomib is a proteasome inhibitor whose anti-cancer action is partly mediated through inhibition of NF-κB, we examined the bortezomib in ABL TKI resistant cells. Combined treatment of ABL TKI resistant cells with ponatinib or imatinib and bortezomib caused more cytotoxicity than each drug alone. Caspase 3/7 activity and cellular cytotoxicity was also increased. ABL TKIs are also associated with vascular adverse events (VAEs) in CML, we next investigated the in vitro effects of ABL TKIs on cultured HUVEC. We found gene expression of IL-1β was increased after ABL TKI especially dasatinib and ponatinib treatment. IL-1β was increased in the cell culture supernatant after ABL TKIs treatment. In the immunoblot analysis, p38 mitogen-activated protein kinase (MAPK) and c-Jun N-terminal kinase (JNK) phosphorylation was increased by IL-1β in the time dependent manner. We also found that gene expression of IL-6, IL-8, intercellular adhesion molecule 1 (ICAM1) and monocyte chemotactic protein-1 (MCP-1) was enhanced by IL-1β stimulation. Conclusion: The IL-1β signaling pathway is involved in ABL TKI sensitivity and drug resistant in CML cells and plays a key role in cytokine production of the HUVEC. We also provide the promising clinical relevance as a candidate drug for treatment of ABL TKI resistant leukemia patients. Disclosures Ohyashiki: Asahikase: Research Funding; Taiho Pharmaceutical KK: Honoraria, Research Funding; Dainippon Sumitomo KK,: Honoraria, Research Funding; MSD,: Honoraria, Research Funding; Bristol Meyer Squibb KK,: Honoraria, Research Funding; Ono Pharmaceutical KK,: Honoraria, Research Funding; Celegene KK,: Honoraria, Research Funding; Pfizer KK,: Honoraria, Research Funding; Kyowakko Kirin KK,: Research Funding; Nihon-Seiyaku,: Research Funding; Eizai,: Research Funding; Chugai KK,: Honoraria, Research Funding; Takeda Pharmaceutical KK,: Honoraria, Research Funding; Asteras KK,: Research Funding; Jansen Pharma KK,: Research Funding; Nippon-shinyaku,: Honoraria, Research Funding; Novartis KK,: Honoraria, Research Funding.
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Jabbour, Elias J., Michael W. Deininger, Elisabetta Abruzzese, Jane F. Apperley, Jorge E. Cortes, Charles Chuah, Daniel J. DeAngelo, et al. "Dose Modification Dynamics of Ponatinib in Patients with Chronic-Phase Chronic Myeloid Leukemia (CP-CML) from the PACE and Optic Trials." Blood 138, Supplement 1 (November 5, 2021): 2550. http://dx.doi.org/10.1182/blood-2021-146175.
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Abstract Introduction: Ponatinib is a potent, oral, third-generation tyrosine kinase inhibitor (TKI) that is FDA approved for treatment of patients with relapsed/refractory CML. Patients with resistant or intolerant CP-CML demonstrated deep, lasting responses to ponatinib 45 mg once daily in the pivotal phase 2 PACE trial (Ponatinib Ph+ ALL and CML Evaluation, NCT01207440; completed). The phase 2 OPTIC trial (Optimizing Ponatinib Treatment in CP-CML, NCT02467270; ongoing) prospectively evaluated a response-based dose-reduction strategy in an attempt to optimize the dose schedule of ponatinib in patients with CP-CML resistant to second-generation (2G) BCR-ABL1 TKI therapy or with a T315I mutation. Unique dosing strategies in the 2 trials (PACE and OPTIC) provide the opportunity to closely evaluate the dose and schedule of ponatinib. Here, we conduct an in-depth analysis of dosing dynamics between the 2 trials and compare efficacy and safety outcomes. Methods: In this analysis, adults with resistant or intolerant CP-CML from the PACE and OPTIC trials were enrolled and received an initial dose of ponatinib 45 mg once daily (PACE) or were randomly assigned (1:1:1) to an initial oral dose of ponatinib 45 mg, 30 mg, or 15 mg once daily (OPTIC). In PACE, proactive dose reductions were mandated in ≈2 years from initiation of first patient (in 2013) as arterial occlusive events (AOEs) emerged as notable adverse events (AEs); patients who achieved major cytogenetic response (MCyR) had doses reduced to 15 mg once daily and those without MCyR reduced to 30 mg once daily, unless benefit-risk analysis justified treatment with a higher dose. OPTIC was designed to incorporate a mandatory response-based dose-reduction strategy; patients in the 45-mg and 30-mg cohorts in OPTIC achieving ≤1% BCR-ABL1 IS reduced their dose to 15 mg once daily; doses also were reduced to manage AEs. Data from patients with CP-CML in PACE and from the 45-mg starting dose cohort in OPTIC are included in this analysis. Efficacy outcomes included ≤1% BCR-ABL1 IS, progression-free survival (PFS), and overall survival (OS). Safety and dosing data are also presented, including treatment-emergent adverse events (TEAEs) and treatment-emergent AOE (TE-AOE) rates. Results: Overall, 364 patients with CP-CML had at least 1 prior 2G TKI or had a T315I mutation and received a starting dose of ponatinib 45 mg (PACE, n=270; OPTIC, n=94). Median follow-up, 57 months (PACE) and 32 months (OPTIC). Efficacy outcomes were generally comparable or better in OPTIC when compared with PACE, including ≤1% BCR-ABL1 IS response by 24 months (PACE, 52%; OPTIC, 56%), 2-year PFS (68%; 80%), and 2-year OS (86%; 91%). Median time to ≤1% BCR-ABL1 IS response, 5.6 months (PACE) and 6 months (OPTIC). Median duration of response was not reached in either trial. Overall, median relative dose intensity (Table 1) was 27 mg/d in PACE and 15 mg/d in OPTIC, and dose reduction occurred more rapidly compared with PACE median (Figure). Dose reductions due to AEs occurred in 82% of patients in PACE and 46% in OPTIC. Median time to dose reduction for AEs was 2.85 months in PACE and 3.64 months in OPTIC. Median time on therapy was 12.6 months in PACE and 19.5 months in OPTIC. Exposure-adjusted TE-AOEs were 15.8 events per 100-patient years at 0 to <1 year in PACE and 7.6 events per 100-patient years at 0 to <1 year in OPTIC (Table 2). There were differences in baseline characteristics between the 2 trials, including differences in cardiovascular baseline status; however, these were accounted for in the propensity score analyses comparing AOE incidence, which after adjusting for baseline differences, showed a 60% reduction in relative risk for AOEs in OPTIC vs PACE. In-depth individual dosing dynamics by safety and efficacy will be presented. Conclusions: The response-based dose-reduction strategy in the OPTIC trial resulted in more rapid dose reductions, lower overall median relative dose intensity, fewer dose reductions related to AEs, and longer median time on therapy in OPTIC compared with PACE, further demonstrating the benefit of the response-based dosing regimen used in OPTIC. These data from the PACE and OPTIC trials suggest that treatment with a response-based dose-reduction strategy may provide comparable or better efficacy while mitigating risk of AEs/AOEs with ponatinib. Furthermore, this abstract supports the rationale to explore response-based dose-modification strategies for other BCR-ABL1 TKIs. Figure 1 Figure 1. Disclosures Jabbour: Amgen, AbbVie, Spectrum, BMS, Takeda, Pfizer, Adaptive, Genentech: Research Funding. Deininger: SPARC, DisperSol, Leukemia & Lymphoma Society: Research Funding; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding; Novartis: Consultancy, Research Funding; Fusion Pharma, Medscape, DisperSol: Consultancy; Sangamo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Blueprint Medicines Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding; Incyte: Consultancy, Honoraria, Research Funding. Abruzzese: Novartis: Consultancy, Honoraria; Pfizer: Consultancy, Honoraria; Incyte: Consultancy, Honoraria; Bristol Myers Squibb: Consultancy, Honoraria. Apperley: Bristol Myers Squibb, Novartis: Honoraria, Speakers Bureau; Incyte, Pfizer: Honoraria, Research Funding, Speakers Bureau. Cortes: Bristol Myers Squibb, Daiichi Sankyo, Jazz Pharmaceuticals, Astellas, Novartis, Pfizer, Takeda, BioPath Holdings, Incyte: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Sun Pharma: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Bio-Path Holdings, Inc.: Consultancy, Membership on an entity's Board of Directors or advisory committees. Chuah: Pfizer: Other: Travel, Research Funding; Steward Cross: Honoraria; Bristol Myers Squibb: Honoraria, Research Funding; Novartis, Korea Otsuka Pharmaceutical: Honoraria. DeAngelo: Abbvie: Research Funding; Takeda: Consultancy; Servier: Consultancy; Pfizer: Consultancy; Novartis: Consultancy, Research Funding; Jazz: Consultancy; Incyte: Consultancy; Forty-Seven: Consultancy; Autolus: Consultancy; Amgen: Consultancy; Agios: Consultancy; Blueprint: Research Funding; Glycomimetrics: Research Funding. Hochhaus: Bristol-Myers Squibb: Research Funding; Novartis: Membership on an entity's Board of Directors or advisory committees, Research Funding; Pfizer: Research Funding; Incyte: Research Funding. Lipton: Bristol Myers Squibb, Ariad, Pfizer, Novartis: Consultancy, Research Funding. Nicolini: BMS: Honoraria; Sun Pharma Ltd.: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte Biosciences: Honoraria, Other: travel, accommodations, expenses, Research Funding, Speakers Bureau; Kartos Therapeutics: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: travel, accommodations, expenses, Research Funding. Pinilla Ibarz: AbbVie, Janssen, AstraZeneca, Takeda: Speakers Bureau; MEI, Sunesis: Research Funding; Sellas: Other: ), patents/royalties/other intellectual property; AbbVie, Janssen, AstraZeneca, Novartis, TG Therapeutics, Takeda: Consultancy, Other: Advisory. Rea: Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees. Rosti: Pfizer: Research Funding, Speakers Bureau; Bristol Myers Squibb, Incyte, Novartis: Speakers Bureau. Rousselot: Incyte, Pfizer: Consultancy, Research Funding. Mauro: Novartis: Consultancy, Research Funding; Takeda: Consultancy; Bristol Myers Squibb: Consultancy, Research Funding; Pfizer: Consultancy; Sun Pharma / SPARC: Research Funding. Shah: Bristol-Myers Squibb: Research Funding. Talpaz: Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Other: Grant/research support ; Constellation: Membership on an entity's Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity's Board of Directors or advisory committees; Imago: Consultancy; Celgene: Consultancy. Vorog: Takeda: Current Employment. Lu: Takeda: Current Employment. Kantarjian: Daiichi-Sankyo: Research Funding; AbbVie: Honoraria, Research Funding; Amgen: Honoraria, Research Funding; BMS: Research Funding; Astra Zeneca: Honoraria; Ipsen Pharmaceuticals: Honoraria; KAHR Medical Ltd: Honoraria; Pfizer: Honoraria, Research Funding; Jazz: Research Funding; Novartis: Honoraria, Research Funding; Astellas Health: Honoraria; NOVA Research: Honoraria; Ascentage: Research Funding; Aptitude Health: Honoraria; Immunogen: Research Funding; Precision Biosciences: Honoraria; Taiho Pharmaceutical Canada: Honoraria.
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Zhang, Bin, Yin Wei Ho, Tessa L. Holyoake, and Ravi Bhatia. "Inhibition Of Microenvironmental Interleukin-1 Signaling Enhances TKI-Mediated Targeting Of Chronic Myelogenous Leukemia Stem Cells." Blood 122, no. 21 (November 15, 2013): 512. http://dx.doi.org/10.1182/blood.v122.21.512.512.
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Abstract BCR-ABL tyrosine kinase inhibitors (TKI), although highly effective in inducing remission and improving survival in chronic myelogenous leukemia (CML) patients, fail to eliminate leukemia stem cells (LSC), which remain a potential source of relapse. Most CML patients need continued TKI treatment to prevent disease relapse, and new strategies to eliminate residual leukemia stem cells are required to enhance possibility of achieving treatment-free remission. In previous studies we have shown that increased several cytokines expressed by leukemia cells may provide a selective growth advantage to CML compared with normal long term hematopoietic stem cells (LTHSC) within the CML BM microenvironment. Studies evaluating the effects of individual factors indicated that exposure to Interleukin-1α/β (IL-1α/β) at concentrations similar to those observed in CML BM resulted in significantly increased growth of CML compared with normal LTHSC (Cancer Cell 2012, 21:577). Consistent with previous reports (PNAS 2010, 107:16280), we observed that expression of the IL-1 receptor-associated protein (IL-1RAP), an important IL-1 signaling component, was increased in primitive CML cells, potentially explaining enhanced IL-1 sensitivity. To further evaluate the role of microenvironmental IL-1 in maintenance of CML LTHSC, we used recombinant IL-1 receptor antagonist (IL-1RA) to block IL-1 receptor signaling. IL-1RA is clinically approved for the treatment of rheumatoid arthritis. Purified LTHSC (Lin-Sca-1+Kit+Flt3-CD150+CD48- cells) from the SCL-tTA/BCR-ABL inducible mouse model of CML (CD45.1) and from congenic FVBN mice (CD45.2) were mixed in a 1:1 ratio and cultured with CML BM plasma, with and without IL-1RA. Culture with CML BM plasma for 7 days results in significantly increased growth of CML compared to normal LTHSC. The ratio of CML to normal cells was significantly reduced in the presence of IL-1RA (2.5μg/ml) (3.6:1 without IL-1RA, 1.7:1 with IL-1RA, p=0.0002), indicating that inhibition of IL-1 signaling reduced the growth advantage of CML LTHSC cultured in CML BM plasma. We next investigated the effect of IL-1RA on CML hematopoiesis in vivo. BM cells from CML mice (CD45.1) were transplanted into congenic FVBN mice (CD45.2) to generate CML-like disease in recipient mice. Four weeks after transplantation mice were treated with Nilotinib (NIL, 50mg/kg/d, gavage), IL-1RA (150mg/kg/d s.c.), the combination of NIL and IL-1RA, or vehicle (control) for 3 weeks. Treatment with NIL plus IL-1RA resulted in significantly greater reduction in CD45.1+ CML cells in blood, and in CML LTHSC, MPP, CMP and GMP in BM, compared with NIL alone (CML LTHSC/2 femurs: control 738±122, NIL 486±94, IL-1RA 525±49, combination 360±33, P=0.01 combination vs. Nilotinib). Mice treated with NIL plus IL-1RA also showed significantly prolonged survival after completion of treatment compared to mice treated with NIL alone (median survival 6 days for NIL alone versus 45 days for combination, p=0.02). Following transplantation of BM cells from treated mice into 2nd recipients (CD45.2), significantly lower CML cell engraftment in BM and reduced development of leukemia was seen after transplantation of cells from mice treated with the combination compared with NIL or untreated controls (8 out of 8 mice developed leukemia for control, 6 out of 8 for NIL, 5 out of 8 for IL-1RA, 3 out of 8 for the combination). We also studied the effect of treatment with NIL (5μm), IL-1RA (5μg/ml), NIL+IL-1RA, or vehicle for 72 hours on human CML and normal CD34+CD38+ and CD34+CD38- cells cultured with CML BM conditioned medium (CM). The combination of NIL and IL-1RA significantly reduced CML CD34+CD38+ and CD34+CD38- cell growth compared to Nilotinib alone (CD38- cells: NIL 23.7±10.1%, combination 13.1±8.9% of control, p<0.05), cell division (measured by CFSE labeling) (CD38- proliferation index: NIL 3.3±1.0, combination 2.4±0.6, p=0.06) and CFC frequency in methylcellulose progenitor assays (CD38- cells: NIL 67±22 per 1000 cells, combination 39±26, p<0.05); and moderately increased apoptosis of CML CD34+CD38- cells. We conclude that inhibition of microenvironmental IL-1 signaling using IL-1RA significantly increases inhibition of self-renewing murine and human CML stem cells in combination with NIL. Our results support further evaluation of IL-1 inhibition as a strategy to enhance elimination of CML LSC in TKI-treated patients. Disclosures: Holyoake: Novartis: Membership on an entity’s Board of Directors or advisory committees; Bristol Myers Squibb: Membership on an entity’s Board of Directors or advisory committees; Ariad: Membership on an entity’s Board of Directors or advisory committees.
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Zhang, Bin, YinWei Ho, Tinisha McDonald, Allen Lin, David S. Snyder, Vu N. Ngo, Tessa L. Holyoake, and Ravi Bhatia. "Role of Enhanced Microenvironmental Interleukin-1 (IL-1) Expression and Increased IL-1 Responsiveness in Persistence of Leukemia Stem Cells in TKI Treated CML Patients." Blood 124, no. 21 (December 6, 2014): 4357. http://dx.doi.org/10.1182/blood.v124.21.4357.4357.
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Abstract BCR-ABL tyrosine kinase inhibitors (TKI), although highly effective in the treatment of chronic myelogenous leukemia (CML) patients, fail to eliminate leukemia stem cells (LSC), which remain a potential source of relapse. In previous studies we have shown that altered expression of inflammatory cytokines in the CML bone marrow (BM) microenvironment provides a selective growth advantage to CML compared with normal long term hematopoietic stem cells (LTHSC) (Cancer Cell 2012, 21:577). Our studies suggest an important role for the pivotal pro-inflammatory cytokine Interleukin-1α/β (IL-1α/β) in selectively promoting growth of CML LTHSC. Using a transgenic BCR-ABL mouse model of CML and human CML and normal CD34+CD38- cells, we showed that inhibition of IL-1 signaling using recombinant IL-1 receptor antagonist (IL-1RA) in combination with nilotinib (NIL) resulted in significantly greater inhibition of CML LSC, compared with NIL alone (Blood 2013, abstract 512). To further investigate the mechanisms underlying increased IL-1 sensitivity of CML stem cells, we evaluated expression of the IL-1 receptor components, IL-1 receptor-associated protein (IL-1RAP) and IL-1R1, on CML and normal stem cells using flow cytometry. Expression of both IL-1RAP and IL-1R1 were increased on primary CML CD34+CD38-CD90+ cells compared to their normal counterparts (n=5, p<0.05). Exposure to IL-1α (10ng/ml) resulted in increased expression of p-NF-kB (p65), p-p38 MAPK and p-JNK in CML compared to normal CD34+CD38-CD90+ cells as evaluated by flow cytometry, indicating enhanced sensitivity to IL-1 induced signaling (n=5, p<0.05). The expression of p-NF-kB(p65), p-p38 MAPK and p-JNK in CML CD34+CD38-CD90+ cells cultured in CML BM conditioned medium (CM) was reduced after treatment with NIL or IL-1RA, and further reduced by the combination of NIL and IL-1RA (n=4, p<0.05). Immunohistochemistry (IHC) analysis showed that nuclear NF-κB p65 protein was reduced in NIL and IL-1RA treated CML CD34+CD38-CD90+ cells compared with controls. Treatment with NIL and IL-1RA also significantly reduced expression of the NF-κB target genes NFκB1A, BCL2L1, BIRC3 and CD83 (n=6, p<0.001), and of the inflammatory cytokines IL6, CXCL1, CXCL2, CCL2, CCL3, CCL4 and TNF-α (n=6, p<0.05), as assessed by Q-RT-PCR. We evaluated IL-1 expression in BM samples from CML patients with undetectable minimal residual disease (UMRD) using Q-RT-PCR. Interestingly IL-1α, but not IL-1β, expression was increased in BM samples from CML patients with UMRD compared to normal BM samples (n=12, p<0.05). To evaluate the source of increased IL-1α expression we analyzed selected monocyte (CD45+CD14+), non-monocytic myeloid cell (CD45+CD14-CD33+), T cell (CD45+CD14-CD33-CD3+), B cell (CD45+CD14-CD33-CD19+), endothelial cell (CD45-GPA-CD31+) and mesenchymal cell (CD45-GPA-CD31-) populations from BM samples obtained from CML patients with UMRD and from normal healthy controls. These studies revealed significantly elevated IL-1α expression in BM CD14+ monocytic and CD31+ endothelial cells from CML patients with UMRD compared to normal controls (n=12, p<0.05). Our studies indicate that CML LSC demonstrate increased IL-1 receptor expression and IL-1 induced NF-kB, p38 MAPK and JNK signaling. We also observe enhanced IL-1α expression in BM endothelial and monocytic cells from CML patients achieving UMRD, indicating persistence of an inflammatory microenvironment that may contribute to persistence of residual LSC. Our studies provide a strong rationale for the application of anti-IL-1 directed strategies to inhibit inflammatory signaling and enhance LSC elimination in TKI treated CML patients. Disclosures No relevant conflicts of interest to declare.
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Zhao, Helong, Anthony D. Pomicter, Anna M. Eiring, Anca Franzini, Jonathan Ahmann, Jae-Yeon Hwang, Anna V. Senina, et al. "MS4A3 Promotes Differentiation in Chronic Myeloid Leukemia By Enhancing Common β Chain Cytokine Receptor Endocytosis." Blood 138, Supplement 1 (November 5, 2021): 59. http://dx.doi.org/10.1182/blood-2021-151559.
Full textAbstract:
Abstract Background Chronic phase chronic myeloid leukemia (CP-CML) is characterized by overproduction of differentiating myeloid cells, while blast phase CML (BP-CML) cells exhibit a differentiation block. Tyrosine kinase inhibitors (TKIs) are effective in CP-CML, but resistance is common in BP-CML and can occur without explanatory BCR-ABL1 kinase mutations (BCR-ABL1-independent resistance). Similarly, CML stem/progenitor cells (LSPCs) are insensitive to TKIs, and residual leukemia persists in the majority of CML patients on TKI therapy. We previously reported overlap between the transcriptomes of CD34 + cells from BP-CML and TKI-naïve CP-CML patients with primary TKI resistance, pointing to commonalities between LSPC quiescence, BCR-ABL1-independent TKI resistance, and BP-CML. Results To identify common mechanisms, we performed a meta-analysis of published CML transcriptomes. We identified a small set of genes with consistently low expression in LSPC quiescence, BCR-ABL1-independent TKI resistance, and BP-CML, including Membrane Spanning 4-Domains A3 (MS4A3), a signaling protein previously reported to inhibit hematopoietic cell cycle progression. Low MS4A3 in CD34 + cells from TKI-naïve CP-CML patients was associated with shorter survival on subsequent TKI therapy, suggesting that MS4A3 governs TKI response. To understand the function of MS4A3, we lentivirally introduced MS4A3 shRNA or an MS4A3 expression vector into CML CD34 + LSPCs. MS4A3 knockdown increased clonogenicity and imatinib resistance, while ectopic MS4A3 expression had opposite effects. MS4A3 KD also increased LSPC persistence ex vivo in LTC-IC assays, and in vivo in NSG mice xenografts, while modulating MS4A3 expression had no effect on normal CD34 + cells. We next generated Ms4a3+/+│-/-; Scl-tTA+; TRE-BCR-ABL1+ compound transgenic mice. Upon BCR-ABL1 induction, Ms4a3-/-; Scl-tTA+; TRE-BCR-ABL1+ mice developed leukocytosis comparable to Ms4a3+/+ controls. However, BM of Ms4a3-/-; Scl-tTA+; TRE-BCR-ABL1+ mice showed increased short-term HSCs and multipotent progenitor cells, and reduced granulocyte-macrophage progenitors. When Lin - BM cells from leukemic mice were transplanted into irradiated recipients, Ms4a3-/-; Scl-tTA+; TRE-BCR-ABL1+ cells showed increased engraftment and myeloid leukocytosis, validating our observations in human cells. To determine how MS4A3 is downregulated in CML, we expressed BCR-ABL1in 32D-cl3 cells. p210 BCR-ABL1 drastically reduced Ms4a3 expression, while kinase-inactive p210 BCR-ABL1-K271R had no effect. Moreover, we found that suppression of C/EBPε by MECOM reduces MS4A3, consistent with previous reports of MECOM as a driver of TKI resistance and progression to BP. Treatment of CML CD34 + cells with a library of epigenetic pathway inhibitors revealed that MS4A3 is suppressed by both DNA methylation and PRC2/EZH2-mediated H3K27 trimethylation, which was confirmed by patch-PCR and ChIPseq. These data indicate that multi-levelled mechanisms cooperate in the suppression of MS4A3 in CML. To determine how MS4A3 regulates clonogenicity and TKI response, we expressed MS4A3-EGFP fusion protein in LAMA-84 CML cells. We found that MS4A3 resides on the plasma membrane and in endosomes. Surface protein biotin labelling and tandem mass spectrometry ± MS4A3 KD showed that MS4A3 controls endocytosis of membrane proteins, including common β chain (βc) cytokine receptors. Specifically, MS4A3 promotes endocytosis of βc cytokine receptors upon GM-CSF/IL-3 stimulation of primary LSPCs and enhances downstream signaling and differentiation, suggesting that restoring MS4A3 expression has therapeutic efficacy. To test this, we manufactured a prototype MS4A3 protein-loaded liposomal nanoparticle (NP) using coating with the CD34 CD62L for targeted delivery to CD34 + cells. Compared to MS4A3-free NPs, MS4A3 NPs increased CD34 +CD38 + and CD34 -CD38 + at the expense of CD34 +CD38 - cells, reduced clonogenicity, and increased sensitivity to TKIs, mimicking ectopic MS4A3 expression. Conclusion MS4A3 governs response to differentiating myeloid cytokines, providing a unifying mechanism for the differentiation block characteristic of primitive LSPCs and BP-CML cells. We posit that LSPCs downregulate MS4A3 to evade βc cytokine-induced differentiation to maintain a primitive, TKI-insensitive state. MS4A3 re-expression or delivery of ectopic MS4A3 may eliminate LSPCs. Figure 1 Figure 1. Disclosures Druker: Aptose Therapeutics: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Cepheid: Consultancy, Membership on an entity's Board of Directors or advisory committees; EnLiven: Consultancy, Research Funding; Blueprint Medicines: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Aileron: Membership on an entity's Board of Directors or advisory committees; Amgen: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Bristol-Myers Squibb: Research Funding; ALLCRON: Consultancy, Membership on an entity's Board of Directors or advisory committees; GRAIL: Current equity holder in publicly-traded company; Iterion Therapeutics: Membership on an entity's Board of Directors or advisory committees; Merck & Co: Patents & Royalties; Nemucore Medical Innovations, Inc.: Consultancy; Novartis Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees, Patents & Royalties; Pfizer: Research Funding; Recludix Pharma, Inc.: Consultancy; The RUNX1 Research Program: Membership on an entity's Board of Directors or advisory committees; Third Coast Therapeutics: Membership on an entity's Board of Directors or advisory committees; VB Therapeutics: Membership on an entity's Board of Directors or advisory committees; Vincerx Pharma: Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees; Vivid Biosciences: Membership on an entity's Board of Directors or advisory committees. Tyner: Agios: Research Funding; Astrazeneca: Research Funding; Array: Research Funding; Genentech: Research Funding; Janssen: Research Funding; Takeda: Research Funding; Gilead: Research Funding; Incyte: Research Funding; Petra: Research Funding; Seattle Genetics: Research Funding; Constellation: Research Funding; Schrodinger: Research Funding. Oehler: BMS: Consultancy; OncLive: Honoraria; Pfizer: Research Funding; Takeda: Consultancy; Blueprint Medicines: Consultancy. Radich: BMS: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees; Amgen: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees. Deininger: Sangamo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding; Incyte: Consultancy, Honoraria, Research Funding; Fusion Pharma, Medscape, DisperSol: Consultancy; Novartis: Consultancy, Research Funding; SPARC, DisperSol, Leukemia & Lymphoma Society: Research Funding; Blueprint Medicines Corporation: Consultancy, Membership on an entity's Board of Directors or advisory committees, Other: Part of a Study Management Committee, Research Funding.
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Mustjoki, Satu, Peter Rohon, Katrin Rapakko, Sari Hernesniemi, Perttu Koskenvesa, Tuija Lundán, and Kimmo Porkka. "Low or Undetectable Numbers of Philadelphia Chromosome Positive (Ph+) Leukemia Cells in the Primitive (CD34posCD38neg) Stem Cell Fraction in Chronic Myeloid Leukemia (CML) Patients during Tyrosine Kinase Inhibitor Therapy." Blood 112, no. 11 (November 16, 2008): 1079. http://dx.doi.org/10.1182/blood.v112.11.1079.1079.
Full textAbstract:
Abstract Targeted tyrosine kinase inhibitors (TKIs) efficiently induce rapid hematologic and cytogenetic remission in most chronic myeloid leukemia (CML) patients. However, in vitro experiments have suggested that the most primitive CML stem cells residing in the CD34posCD38neg fraction are relatively resistant to TKIs. The prevalence of these stem cells in vivo in patients under TKI therapy is unclear. The aim of this project was to analyze the effect of TKI therapy on Ph+ leukemia stem cell pool in patients and to analyze the proportion of Ph+ cells in different stem cell fractions. A total of 26 chronic phase CML patients were included in the study. 18 patients were treated with imatinib, 5 with dasatinib, and 3 with bosutinib. The median time of TKI treatment was 20 months (range 3–72 months). Large volume (median 30 ml, range 5–55 ml) of bone marrow (BM) aspirate was collected and mononuclear cells (MNC) were isolated. CD34pos cells were separated with paramagnetic beads and further sorted into CD34posCD38pos and CD34posCD38neg cell populations with multicolor flow cytometry in order to analyze progenitor cell fractions of different maturation stage. Proportion of Ph+ cells was determined with interphase FISH by counting 1000 cells in each fraction. The median yield of MNCs from 30 ml of BM aspirate was 280x106 cells resulting in a median of 32 000 CD34posCD38neg cells (range 1000–91000). High-sensitivity counting of the proportion of Ph+ cells was feasible with a median number of counted interphase nuclei of 1005. During TKI therapy the CD34pos cells expressing highest CD38 antigen level were already mostly differentiated into B-cell lineage (CD19 positive). The CD34pos cells expressing low CD38 antigen levels expressed markers of more primitive cells such as C-kit (CD117) and CD133. Of 26 patients with CML, 19 were in complete cytogenetic remission (CCyR) when assessed by metaphase FISH of non-fractionated BM cells (1000 cells analyzed). Only 3 patients had single Ph+ cells in CD34pos cell fractions (less than 1%). In remainder of patients, all progenitor cell fractions, including the most primitive CD34posCD38neg cells, were negative for Ph+ cells. 3 patients had 0–1% of Ph+ cells in non-fractionated BM sample. One of them had 0.2% of Ph+ cells in CD34posCD38neg fraction, but the other 2 patients had 0/1000 Ph+ stem cells. 4 patients had a partial cytogenetic response (5–20% of Ph+ cells in non-fractionated BM sample). Again, the proportion of Ph+ cells was not increased in the most primitive CD34posCD38neg cell fraction. Interestingly, patients who had discontinued imatinib treatment had lower level of Ph+ cells in different CD34pos fractions (median 0.1%) when compared to non-fractionated BM (median 9.3%). Based on our data, in chronic phase CML patients, TKI therapy eradicates most Ph+ CD34pos progenitor cells. Unexpectedly, leukemic stem cells were not enriched in the most primitive CD34posCD38neg cell fraction in vivo. These results differ from the in vitro studies, where CD34posCD38neg cells have been shown to be resistant to TKIs. This could be due to non-physiological conditions (growth factor sensitivity, other cytokines) in cell culture assays. In addition, leukemic stem cells in vivo may be located in the subcortical hypoxic stem cell niche in the BM and are less likely to be aspirated. Our data underline the tremendous proliferative potential of very rare stem cells in CML patients in CCyR, as is evident after discontinuation of TKI therapy. Future studies evaluating the kinetics of disappearance of Ph+ cells from stem cell fractions during TKI therapy and the location of residual Ph+ stem cells in the BM are warranted and may give important information on the depth of the therapy response. Furthermore, this knowledge may aid in targeting therapy to these cells and finding curative treatment strategies in CML.
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Kuznetsova, Valeriya, Sweta Patel, Francesca Luca, Virginia Camacho, Victoria Matkins, and Robert S. Welner. "Perturbed function of natural killer cells by inflammatory cytokines in acute (AML) and chronic (CML) myeloid leukemias." Journal of Immunology 208, no. 1_Supplement (May 1, 2022): 62.04. http://dx.doi.org/10.4049/jimmunol.208.supp.62.04.
Full textAbstract:
Abstract Natural killer (NK) cells have the capacity to eliminate malignant cells by releasing cytotoxic granules. Indeed, mature NK cell counts are associated with favorable prognosis in AML and prolonged TKI-free remission in CML. However, NK cell maturation and anti-leukemic activity are suppressed during the disease, and the environmental drivers of this impairment remain unclear. Given an established role of inflammatory cytokines (particularly IL-1β, IL-6, and TNFα) in the progression of myeloid malignancies, we define their impact on NK cell function in chimeric mouse models of leukemia – Flt3-ITD/TET2, AML/ETO9a, Meis/Hox (AML), and BCR-ABL1 (CML), where NK cells do not carry the oncogene. Reflecting clinical observations, NK cell frequencies and numbers are reduced, implying oncogene-independent mechanisms of NK dysfunction in different leukemic subtypes. Moreover, both AML- and CML-exposed NK cells display immature phenotypes, diminished expression of activating receptors, and impaired ex vivo target-specific degranulation. The single cell RNA-seq analysis of CML-exposed NK cells confirmed decreased expression of NK maturation and cytotoxicity markers and upregulation of inhibitory receptors and genes associated with inflammatory cytokine response. Furthermore, IL-6-STAT3 and IL-1β/TNFα transcriptional signatures are enriched during the disease, which is likely to be triggered by leukemic cytokines. Thus, we next found that leukemic serum dampens NK cell degranulation ex vivo, compared to serum from control mice. These data suggest that soluble leukemic factors contribute to NK cell dysfunction and represent an optimal target for NK-boosting immunotherapies across different types of myeloid malignancies.
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Hochhaus, Andreas, Jorge E. Cortes, Dong-Wook Kim, Javier Pinilla-Ibarz, Philipp D. le Coutre, Ronald Paquette, Charles Chuah, et al. "Efficacy and Safety of Ponatinib in CP-CML Patients By Number of Prior Tyrosine Kinase Inhibitors: 4-Year Follow-up of the Phase 2 PACE Trial." Blood 126, no. 23 (December 3, 2015): 4025. http://dx.doi.org/10.1182/blood.v126.23.4025.4025.
Full textAbstract:
Abstract Introduction: Ponatinib is a potent oral tyrosine kinase inhibitor (TKI) approved for use in patients with refractory CML and Ph+ ALL, including patients with the resistant BCR-ABLT315I mutation. To evaluate whether patient characteristics and outcomes with ponatinib differed by extent of pretreatment with other TKIs, this post hoc analysis examines results among CP-CML patients enrolled in the phase 2 PACE trial (NCT01207440) according to the number of TKIs received prior to study entry. Updated data with 4 years of follow-up will be presented. Methods: Patients with CML or Ph+ ALL who were resistant or intolerant to dasatinib or nilotinib or who had the T315I mutation were enrolled (N=449). Ponatinib was initiated at 45 mg once daily. CP-CML patients were evaluated based on previous treatment with 1, 2, 3, or 4 prior TKIs approved for use in CP-CML (ie, imatinib, dasatinib, nilotinib, and bosutinib). Data reported in this abstract are as of February 2, 2015. Results: Overall, 270 CP-CML patients were enrolled and treated in PACE. Patient characteristics and disposition varied by number of prior TKIs (Table 1). Both median age and median time from diagnosis increased with number of prior TKIs; median dose intensity was highest in patients who had received only 1 prior TKI. The most common reasons for discontinuation across groups were adverse events (AEs) and withdrawal by patient request. Responses by number of prior TKIs are shown in Table 2. Rates of cytogenetic and molecular response to ponatinib were higher with fewer prior TKIs. While the frequency of individual AEs did not follow a consistent trend, the incidence of grade ≥3 AEs appeared to increase with the number of prior TKIs received (68%, 86%, 89%, and 100%, respectively); grade ≥3 AEs in ≥10% of CP-CML patients overall were thrombocytopenia (35%), neutropenia (17%), hypertension (13%), increased lipase (12%), and abdominal pain (10%). A similar frequency pattern was observed for serious AEs, which occurred in 58%, 53%, 62%, and 92% of patients who had previously received 1, 2, 3, and 4 approved TKIs, respectively. Serious AEs in ≥5% of CP-CML patients overall were pancreatitis (7%), angina pectoris (5%), and pneumonia (5%). The frequency of arterial occlusive events (AOEs) was 32% (6/19), 26% (25/98), 28% (39/141), and 42% (5/12) by increasing number of prior TKIs; exposure-adjusted incidence rates of new AOEs were 11.75, 10.4, 12.6, and 33.3 events per 100 patient-years, respectively. Conclusions: With 4 years of follow-up, ponatinib continues to provide benefit to ongoing CP-CML patients in the PACE trial. Analysis by treatment history indicates that patients who had received fewer TKIs prior to study entry appear to exhibit better efficacy and safety profiles. However, treatment decisions should be primarily guided by individual patient and disease factors, including mutation status, and physicians should weigh both the benefits and risks of prescribing ponatinib. Table 1. Patient Characteristics and Disposition by Number of Prior TKIs 1 TKI (n=19) 2 TKIs (n=98) 3 TKIs (n=141) 4 TKIs (n=12) Median age at baseline, years 52 58 63 67 Median time from diagnosis to first dose, years 2.8 5.2 7.9 12.4 Median dose intensity, mg/d 34.0 28.7 29.9 31.0 Mutations detected at baseline, % 68 51 43 75 T315I mutation detected at baseline, % 63 31 16 0 Prior TKI exposure, %, imatinib/dasatinib/nilotinib/bosutinib 68/21/5/5 97/66/36/1 100/96/96/7 100/100/100/100 Remain on study, % 53 48 40 8 Discontinued, % 47 52 60 92 Primary reason for discontinuation, % AE 16 18 17 33 Withdrawal by patient request 5 11 11 25 Disease progression 16 5 13 0 Lack of efficacy 0 2 9 8 Death 0 2 3 17 Othera 11 13 8 8 Median follow-up, months 42.3 42.9 42.1 28.2 aIncludes noncompliance, physician decision, protocol violation, and other reasons Table 2 Responses to Ponatinib by Number of Prior TKIs 1 TKI (n=16a) 2 TKIs (n=98) 3 TKIs (n=141) 4 TKIs (n=12) MCyR 12 (75) 69 (70) 69 (49) 7 (58) CCyR 12 (75) 63 (64) 63 (45) 4 (33) MMR 10 (63) 41 (42) 51 (36) 1 (8) MR4 6 (38) 30 (31) 38 (27) 1 (8) MR4.5 4 (25) 22 (22) 34 (24) 1 (8) All responses are n (%) a16/19 patients were evaluable for efficacy Disclosures Hochhaus: Bristol-Myers Squibb: Honoraria, Research Funding; Novartis: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding. Cortes:ARIAD Pharmaceuticals Inc.: Other: Consulting & Advisory Role, Research Funding. Pinilla-Ibarz:Novartis: Consultancy, Other: Consulting & Advisory Role, Research Funding; ARIAD Pharmaceuticals, Inc.: Consultancy, Other: Consulting & Advisory Role, Research Funding; Teva: Consultancy, Speakers Bureau; Pfizer: Consultancy, Other: Consulting & Advisory Role, Research Funding, Speakers Bureau; BMS: Consultancy, Honoraria, Other: Consulting & Advisory Role, Speakers Bureau. le Coutre:Novartis: Honoraria; BMS: Honoraria; ARIAD Pharmaceuticals Inc.: Honoraria; Pfizer: Honoraria. Paquette:ARIAD Pharmaceuticals Inc.: Honoraria; BMS: Honoraria; Novartis: Honoraria. Chuah:Children International: Honoraria; Novartis: Honoraria; Bristol Meyers Squibb: Honoraria. Nicolini:Ariad Pharmaceuticals: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Bristol-Myers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding, Speakers Bureau. Apperley:BMS: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; ARIAD: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Novartis: Membership on an entity's Board of Directors or advisory committees, Speakers Bureau. Talpaz:Pfizer: Other: CONSULTING OR ADVISORY ROLE; Novartis: Other: CONSULTING OR ADVISORY ROLE; ARIAD Pharmaceutical Inc.: Other: CONSULTING OR ADVISORY ROLE; Pfizer: Other: TRAVEL, ACCOMODATIONS, EXPENSES, Research Funding; Novartis: Other: TRAVEL, ACCOMODATIONS, EXPENSES, Research Funding; ARIAD Pharmaceutical Inc.: Other: TRAVEL, ACCOMODATIONS, EXPENSES, Research Funding; Incyte: Other: TRAVEL, ACCOMODATIONS, EXPENSES, Research Funding; Sanofi: Research Funding. DeAngelo:Incyte: Other: Consulting or Advisory Role; Pfizer: Other: Consulting or Advisory Role; Novartis: Other: Consulting or Advisory Role; BMS: Other: Consulting or Advisory Role; ARIAD Pharmaceuticals Inc.: Other: Consulting & Advisory Role; Amgen: Other: Consulting or Advisory Role. Abruzzese:BMS: Other: Consulting or Advisory Role; ARIAD Pharmaceuticals Inc.: Other: Consulting & Advisory Role; Novartis: Other: Consulting or Advisory Role; Pfizer: Other: Consulting or Advisory Role. Rea:Bristol-Myers Squibb: Honoraria; Novartis: Honoraria; Pfizer: Honoraria; Ariad: Honoraria. Baccarani:Pfizer: Other: Travel, Accommodations, Expenses; BMS: Other: Travel, Accommodations, Expenses; Novartis: Other: Travel, Accommodations, Expenses; ARAID Pharmaceutical Inc.: Other: Consulting or Advisory Role, Speakers Bureau; Pfizer: Honoraria, Other: Consulting or Advisory Role, Speakers Bureau; Novartis: Honoraria, Other: Consulting or Advisory Role, Speakers Bureau; BMS: Honoraria, Speakers Bureau; ARIAD Pharmaceutical Inc.: Other: Travel, Accommodations, Expenses. Muller:ARIAD Pharmaceuticals Inc.: Honoraria, Other: Consulting & Advisory Role, Research Funding; Novartis: Honoraria, Other: Consulting or Advisory Role, Research Funding; BMS: Honoraria, Other: Consulting or Advisory Role, Research Funding. Lustgarten:ARIAD Pharmaceuticals Inc.: Employment, Equity Ownership, Other: Stock. Conlan:ARIAD Pharmaceuticals Inc.: Other: Stock. Rivera:ARIAD Pharmaceuticals Inc.: Employment, Other: Full-time Employee & Shareholder (self-managed). Guilhot:Celgene: Consultancy, Other: CONSULTING OR ADVISORY ROLE; Pfizer: Honoraria; Novartis: Honoraria, Other: TRAVEL, ACCOMODATIONS, EXPENSES. Deininger:Incyte: Consultancy, Honoraria, Other: Consulting or Advisory Role; Pfizer: Consultancy, Honoraria, Other: Consulting or Advisory Role; ARIAD Pharmaceutical Inc.: Consultancy, Honoraria, Other: Consulting or Advisory Role; Gilead: Research Funding; Celgene: Research Funding; Novartis: Consultancy, Honoraria, Other: Consulting or Advisory Role, Research Funding; BMS: Consultancy, Honoraria, Other: Consulting & Advisory Role, Research Funding. Hughes:Bristol-Myers Squibb: Honoraria, Research Funding; ARIAD: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Shah:Pfizer: Research Funding; Bristol-Myers Squibb: Research Funding; Plexxikon Inc.: Research Funding. Kantarjian:Novartis: Research Funding; BMS: Research Funding; Pfizer: Research Funding; Amgen: Research Funding.
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de Lavallade, Hugues, David Marin, Melanie Hart, Takuya Sekine, Ian Gabriel, Abdullah Alsuliman, Alexandra Bazeos, et al. "Chronic Myeloid Leukemia Patients on Tyrosine Kinase Inhibitor Have Normal T Cell Responses to Vaccination but An Impaired IgM Humoral Response Associated with Loss of Discrete Memory B Cell Subsets,." Blood 118, no. 21 (November 18, 2011): 3753. http://dx.doi.org/10.1182/blood.v118.21.3753.3753.
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Abstract Abstract 3753 The tyrosine kinase inhibitors (TKIs) imatinib (IM), nilotinib (NIL) and dasatinib (DAS) are remarkably effective as single-agent therapy for chronic myeloid leukemia (CML) in chronic phase (CP). However little is known of their potential impact on the immune response. No human in vivo studies to assess how these molecular-targeted drugs affect immune function in patients are available and data from in vitro and animal studies with imatinib have been contradictory, ranging from impaired antigen-specific T-cell response to enhanced stimulation of tolerant T cells. Furthermore, although the immunomodulatory effects of TKIs on T cells, NK cells and dendritic cells have been explored in vitro, little is known of their potential impact on B cells. To characterize the in vivo immunomodulatory effects of TKIs, 51 patients with CP-CML in complete cytogenetic response (CCyR) on standard dose IM (n=26), DAS (n=14) or NIL (n=12) and 28 adult controls were recruited during two influenza seasons (2008 and 2009). Patients and controls were concomitantly immunized with an influenza vaccine (Ph. Eur. 2008/2009 or Ph. Eur. 2009/2010, CSL Biotherapies) and with the 23-valent polysaccharide pneumococcal vaccine (Pneumovax II; Sanofi Pasteur MSD). Peripheral blood mononuclear cells (PBMCs) and serum samples were collected from patients and donors prior to vaccination and T and B responses to vaccination were assessed at 4 weeks and at 2–3 months post-immunization. T-cell responses to influenza vaccine were analyzed quantitatively and qualitatively using flow cytometry and intracellular cytokine assay for TNF-α, IFN-γ, IL-2 and the cytotoxicity marker CD107a. Serum titers of IgM and IgG pneumococcal antibodies were determined by ELISA. Analysis of B cell subsets was performed using flow cytometry and correlated with the pneumococcal IgM and IgG humoral response. Following vaccination, Flu-specific T cells were detected in 24/51 (47.0%) patients on TKI and 15/24 (62.5%) healthy controls (p=0.16). Polyfunctional T-cell responses (defined as the production of 2 or more cytokines or one cytokine and the cytotoxic marker CD107a) were induced in 6/10 evaluable patients and 4/8 normal controls (p=1.0). T-cell independent humoral responses to vaccination were assessed in 45 patients and 12 healthy controls by measuring pneumococcal IgM titers. Four weeks postimmunization, 11/12 (92%) controls achieved IgM pneumococcal Ab titers >80 U/ml compared to only 23/45 (53%) CML patients on TKI (p=0.010). The pneumococcal IgM titers were significantly lower in patients with CML on TKI compared to healthy controls (median, 89.0 U/ml, range 5–200 vs 200 U/ml, range 58–200, p=0.0006), suggesting that CML patients on TKI have impaired IgM responses to vaccination. To further characterize the humoral immune response to Pneumovax, we stratified CML patients based on their pneumococcal IgM titers. We found a significantly lower percentage of IgM memory B cell subset in CML patients who failed to mount a significant pneumococcal IgM response compared to patients who achieved a pneumococcal IgM response (median, 6.25% vs 16.4%, p=0.0059) and healthy controls (median, 6.25% vs 14.3%, p=0.0086). Furthermore, we found a significant correlation between anti-pneumococcal IgM titers and IgM memory B cell percentage (Spearman rank correlation test, r=0.61, p<.0001). To investigate a putative role of TKIs for the loss of IgM memory B cell subsets in CML patients, we determined the frequencies of IgM memory B cells in paired samples collected from 15 CML-CP patients at diagnosis (i.e. prior to initiating IM) and once CCyR was achieved. We found a significant decrease in the percentage of IgM memory B cells in CML-CP patients treated with IM compared to the pre-treatment sample (median 9.4%, vs. 15.2% respectively, p=0.0023). In summary, patients with CML on TKIs can mount effective T-cell immune responses to influenza vaccination. Our data suggest that TKIs (IM, DAS and NIL) impair T-cell independent humoral immune responses, namely IgM responses to vaccination. This is associated with a loss of IgM memory B cell subsets. Further investigations to understand the mechanisms by which TKIs may impact B-cell subsets are underway. These results are of particular interest in terms of the long-term effects of TKI on tumor immune surveillance and susceptibility to infections and may have implication for vaccination strategies in CML patients. Disclosures: No relevant conflicts of interest to declare.
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Thiant, Stephanie, Moutuaata M.Moutuou, Philippe Laflamme, Radia Sidi Boumedine, Fanny Larochelle, Dominique Leboeuf, Denis-Claude Roy, and Martin Guimond. "Disruption of the Peripheral Lymphoid Niche Contributes to Lymphopenia in CML Patients Undergoing Imatinib Treatments." Blood 126, no. 23 (December 3, 2015): 5165. http://dx.doi.org/10.1182/blood.v126.23.5165.5165.
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Abstract PURPOSE: Chronic myelogenous leukemia (CML) is a disorder affecting early hematopoietic stem cells (HSC) and is characterized by excessive proliferation and accumulation of myeloid progenitors and progeny in the periphery. During the chronic phase of the disease, CML patients are normally at low risk of developing infections but such complications tend to rise during the progression of the disease. Gleevec (imatinib mesylate) is currently administered as first line therapy for patients with Philadelphia chromosome-positive CML. Despite the relative high specificity of tyrosine kinase inhibitor (TKI) treatment towards the BCR-ABL fusion protein, off-target multikinase inhibitory effects occur and can interfere with normal hematopoiesis. This study was conducted to evaluate how myeloid and lymphoid immune homeostasis are affected by Imatinib mesylate. METHODS: Healthy volunteer donors (n=25) and CML patients were recruited during their first visit at our CML clinic. Seven CML patients were treated with Imatinib (400mg). The median time of Gleevec treatment was 2.9 years (range: 0.5-10.9). The median time of remission post TKI was 1.1 years (range: 0.3-3). Phenotypic analysis of dendritic cell (DCs) subsets: plasmacytoid (pDCs) and myeloid type 1, 2 and 3 (mDC1, mDC2, mDC3) were evaluated by flow cytometry. Percentage and absolute numbers of naive and memory CD4+ and CD8+ T cells, NK cells and B cells were also evaluated. DCs were differentiated from purified CD34+ cells culturedwith GM-CSF (800 U/ml) or Flt3-L (50ng/ml), IL-4 (10 U/ml) and TNFa (50 U/ml), in the presence of varying concentrations of Imatinib mesylate (0 to 5µM/mL). TCR and IL-7 signaling were evaluated based on ERK-phosphorylation (-p) and STAT5-p after incubation with 3µM of Imatinib. RESULTS AND CONCLUSION: At diagnosis, several CML patients have a deficit in DCs resulting from a severe skewing affecting BM progenitor cells. After initiating Gleevec therapy, normalization of stem cell progenitors occurs but DC counts remain well below normal levels in all CML patients. We demonstrated a direct and dose dependent interference of Imatinib on GM-CSF and Flt3-L pathways for DC differentiation from CD34+ stem cells. For T lymphocytes, Imatinib interfered with TCR and IL-7 signaling through the inhibition of ERK and STAT5 phosphorylation respectively. The failure to maintain adequate numbers of DCs combined to diminished homeostatic response to cytokines and TCR stimuli explains T cell lymphopenia in these patients. Such immune dysfunction is at least in part responsible for infectious complications that are often increased in patients treated with Imatinib. Disclosures No relevant conflicts of interest to declare.
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Kalota, Anna, and Alan M. Gewirtz. "Computationally Designed, Small Molecule Inhibitors of Tubulin Inhibit the Growth of Gleevec Resistant Chronic Myelogenous Leukemia Cells." Blood 108, no. 11 (November 1, 2006): 2173. http://dx.doi.org/10.1182/blood.v108.11.2173.2173.
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Abstract The abl tyrosine kinase inhibitor (TKI) imatinib mesylate has revolutionized treatment of Chronic Myelogenous Leukemia (CML). Nonetheless, for the small percentage of chronic phase patients in whom resistance to imatinib develops, and for patients in accelerated phase/blast crisis, disease management is problematic. Second generation TKI may address the resistance issue, but serious cardiotoxicity might be a concern for all TKIs. We are therefore investigating other candidates for rationally directed CML therapy. Herein we report that targeting tubulin with computationally designed small molecules may prove useful for the treatment of TKI resistant CML cells. Tubulin inhibitors were obtained from Locus Pharmaceuticals, Blue Bell, PA. They were designed using a method that computes virtual inhibitor molecule binding efficiency after integrating free energy calculations from all chemically possible combinations of molecule fragment poses. Molecules that perform well in in silico screens are then synthesized for biological testing. We evaluated 3 such molecules (LP-261, LOC-011294, and LOC-011423), and 2 control compounds (LOC-007708, LP-590), on K562 human leukemia cells, murine BaF3 cells expressing wild type bcr-abl, or the Y253F, T315I, E255K, H296P and M351T kinase domain mutations (gift from B. Druker, Portland, Oregon), and on consenting donors of normal, and CML, bone marrow cells. LP-261, LOC-011294, and LOC-011423 all had significant activity in K562 cells and BaF3 cells expressing wild type bcr-abl. Inhibition of cell growth in these lines was ~90% when employed at concentrations of > 100nM. More importantly in BaF3 cells expressing each of the mutant abl kinases, including T315I, growth inhibition was also ~90%. Moreover, LP-261, and LOC-011294 were also highly effective against primary cells obtained from patients with chronic phase and blast crisis CML. Treatment of primary CML cells with LP-261 resulted in >80% inhibition of proliferation in all five CML patient samples when compared to control cells. LOC-011294 inhibited cell proliferation by >80% in 3 out of 5 primary patient samples, and by ~50% in one other. Additional testing revealed that LP-261 was not a substrate for the p-glycoprotein multi-drug resistance porter, and that it is orally bioavailable. Neither LOC-007708, (a selective inhibitor of p38 kinase), nor LP-590 (an inhibitor of p38, Flt-3 and tie-2 kinases) inhibited proliferation of the cells expressing mutated bcr-abl. Since these kinases are not thought to influence bcr-abl driven cell growth, these results were expected. In contrast, MOLM14 cells, which overexpress Flt-3, were profoundly inhibited by LP-590. Disease specificity is suggested by the fact that none of these compounds had any effect on growth of acute lymphoid leukemia (ALL) patient samples. To be clinically useful, it was important to demonstrate that normal CD34+ were less sensitive to the growth inhibitory effects of these compounds. For this purpose, normal CD34+ cells were exposed to LP-261, LOC-011294, and LOC-011423 for 24 hours at concentrations shown to inhibit CML cell growth and then plated in methylcellulose with cytokines. Under these conditions, no significant inhibition of CFU-GM, CFU-E, or BFU-E in comparison to control cells was shown. These results suggests that rationally designed anti-tubulin small molecules, alone, or in combination with other active agents, may prove quite useful for treating kinase inhibitor resistant, as well as de novo, CML. This hypothesis may be tested in the near future as an IND has been filed for LP-261.
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Gutknecht, Michael, Lisa Güttler, Mark-Alexander Schwarzbich, Julia Salih, Lothar Kanz, Helmut R. Salih, Susanne Malaika Rittig, and Frank Grünebach. "Upregulation of Osteoactivin Upon Exposure to Tyrosine Kinase Inhibitors Impairs the T Cell Stimulatory Capacity of Monocyte-Derived Dendritic Cells." Blood 118, no. 21 (November 18, 2011): 1112. http://dx.doi.org/10.1182/blood.v118.21.1112.1112.
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Abstract Abstract 1112 Targeted therapies using tyrosine kinase (TK) inhibitors have significantly improved the treatment of cancer patients. Imatinib (Glivec, Gleevec, STI 571) was the first TK inhibitor (TKI) established for the treatment of cancer and efficiently blocks the activity of c-ABL, a non-receptor TK which is pathologically activated in philadelphia chromosome-positive (Ph+) chronic myelogenous leukemia (CML). Nilotinib (Tasigna) and dasatinib (Sprycel) are second-generation TKI that have shown efficacy in the treatment of Ph+ CML resistant or intolerant to imatinib. However, molecularly detectable disease persists in the majority of patients treated with TKI, causing relapse after discontinuation of TKI treatment in many cases. Thus, multiple approaches presently aim to combine TKI treatment with immunotherapy. As TKI, besides targeting their eponymous enzyme, influence multiple other signaling pathways involved in cellular functions, analysis of potential effects of TKI on immune effector cells may be key to develop successful combinatorial strategies. Due to their unique ability to initiate powerful anti-tumor T cell responses, dendritic cells (DC) are employed in many immunotherapeutic strategies aiming to eradicate the malignant cell population. Upon activation they change their expression pattern of cell surface molecules and secreted cytokines/chemokines, a process called DC maturation. Osteoactivin, also known as transmembrane glycoprotein NMB (GPNMB) and dendritic cell-associated transmembrane protein (DC-HIL), is a type I transmembrane glycoprotein that is detected abundantly in DC but not or substantially less in monocytes. Its expression can inhibit T cell activation by binding the type 1 transmembrane proteoglycan syndecan-4 (SD-4) on T cells. Here we extend our findings that the exposure of human peripheral blood monocytes to the immunosuppressive and anti-inflammatory cytokine IL-10 or to therapeutic concentrations of TKI during differentiation into monocyte-derived DC (moDC) leads to significant upregulation of osteoactivin at the transcript and protein level in vitro (Blood 2010 116: abstract 1733). We analyzed the expression of other inhibitory receptors, such as PD-L1, PD-L2, CD80, or CD86 and observed no significant differences of the expression under TKI treatment. Furthermore, we thoroughly examined the expression of osteoactivin in the presence of relevant maturation signals such as TLR ligands, IFN-γ or TNF. LPS, Poly I:C, Pam3Cys or R848 nearly abolished osteoactivin expression compared to untreated control cells. In contrast, IFN-γ or TNF did not significantly reduce osteoactivin expression below the basal level. To evaluate the involvement of osteoactivin in TKI-triggered effects on moDC function we performed mixed lymphocyte reactions with allogenic T cells. Osteoactivin upregulation upon exposure to imatinib, dasatinib and nilotinib resulted in significantly reduced T cell stimulatory capacity of moDC. This was not due to IL-10 upregulation but rather due to direct inhibitory effects of osteoactivin on T cell proliferation which could be overcome by addition of blocking anti-osteoactivin antibody. Our data demonstrate that upregulation of osteoactivin upon exposure of immature moDC to TKI is critically involved in the inhibition of DC function. These findings indicate that inhibition of osteoactivin expression or function may serve as a novel strategy in combinatory approaches using TKI and DC-based immunotherapy and may enhance the efficacy of immunotherapeutic interventions in cancer patients. Disclosures: No relevant conflicts of interest to declare.
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Ågerstam, Helena, Nils Hansen, Sofia Von Palffy, Carl Sandén, Kristian Reckzeh, Christine Karlsson, Henrik Lilljebjörn, et al. "IL1RAP Antibodies Block IL1-Induced Expansion of Primitive CML Cells and Display Therapeutic Effects in Xenograft Models." Blood 128, no. 22 (December 2, 2016): 1118. http://dx.doi.org/10.1182/blood.v128.22.1118.1118.
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Abstract Chronic myeloid leukemia (CML) is currently treated with tyrosine kinase inhibitors (TKIs) but these do not effectively eliminate the CML stem cells. As a consequence, CML stem cells persist and cause relapse in most patients upon drug discontinuation. Furthermore, no effective therapy exists for the advanced stages of the disease. Thus, there is still a need for novel treatment strategies in CML. We have previously shown that Interleukin 1 receptor accessory protein (IL1RAP), a co-receptor of IL1R1, is highly expressed on primitive CML cells and that a polyclonal IL1RAP antibody can direct natural killer (NK) cells to specifically target and destroy CD34+CD38- CML cells in an in vitro-based antibody dependent cell-mediated cytotoxicity (ADCC) assay (Järås et al, PNAS, 2010). The aim of the present study was to investigate the consequences of IL1RAP expression on primitive CML cells and the in vivo therapeutic efficacy of monoclonal IL1RAP antibodies against CML cells. Primary chronic phase (CP) CD34+ CML cells were cultured in medium supplemented with cytokines known to signal through receptor complexes involving IL1RAP. The addition of IL1 to the cultures resulted in a marked cellular expansion specifically for the primitive CD34+CD38- CML cells. Moreover, the CD34+CD38- cells showed phosphorylation of the downstream mediator of IL1-signaling NFKB. RNA-sequencing confirmed the activation of NFKB and of genes involved in cell cycling, indicating that IL1 stimulation of CD34+CD38- CML cells induced proliferation. Upon addition of an IL1RAP antibody capable of blocking IL1-signaling to the suspension cultures, the IL1-induced expansion and NFKB phosphorylation of CD34+CD38- CML cells was suppressed. Interestingly, both the IL1RAP expression and the response to IL1 as measured by NFKB phosphorylation was retained during TKI treatment of the cells. To assess the in vivo effects of IL1RAP antibodies in CML models, we first engrafted NOD/SCID mice with BCR/ABL1 expressing BV173 cells and treated the mice with the monoclonal IL1RAP antibody mAb81.2. Mice receiving treatment with mAb81.2 displayed a prolonged survival compared to controls, accompanied by reduced levels of leukemic cells in the BM. In vitro studies showed that mAb81.2 lacked a direct effect on cellular expansion or apoptosis. Instead, the IL1RAP antibody could direct NK cells to elicit killing of the leukemic cells, thereby suggesting effector cell mediated mechanisms to be an important in vivo mode-of-action. To validate the in vivo effects on primary CML cells, we next engrafted CP or blast phase (BP) CML cells into immunodeficient mice. Following engraftment of CP CD34+ CML cells into NSG mice and subsequent treatment with mAb81.2, a reduction of human myeloid cells was observed, suggesting that the treatment targeted the leukemic graft. Importantly, mAb81.2 treatment also reduced the levels of candidate CD34+CD38-IL1RAP+ CML stem cells. Finally, BP CML cells were engrafted into NOD/SCID mice that have a more intact effector cell function compared to NSG mice. Following treatment with mAb81.2 a significant reduction of leukemic cells in the BM as well as in the periphery was observed compared to control mice. Importantly, secondary transplantations revealed a therapeutic effect also on the BP CML stem cells. In vitro ADCC assays confirmed that CML BP cells, including a sample with the highly TKI-resistant T315I mutation, could be targeted and killed using mAb81.2. We conclude that IL1RAP antibodies can suppress IL1-induced expansion of primitive CML cells and that in vivo administration of IL1RAP antibodies in CML xenograft models has anti-leukemic effects that extend to the CML stem cells. These results show that an antibody-based therapy against IL1RAP can be used to efficiently target CML stem cells. Disclosures Richter: BMS: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Ariad: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Järås:Cantargia AB: Equity Ownership. Fioretos:Cantargia AB: Equity Ownership.
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Rein, Lindsay A. M., and David A. Rizzieri. "A Phase I Trial of Incorporating Natural Killer (K-NK) Cells for Patients with Chronic Myeloid Leukemia (CML) and Molecular Residual Disease after Tyrosine Kinase Inhibitor (TKI) Therapy." Blood 136, Supplement 1 (November 5, 2020): 5. http://dx.doi.org/10.1182/blood-2020-142062.
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Tyrosine kinase inhibitors (TKIs) have revolutionized the treatment of patients with chronic myeloid leukemia (CML) ushering in an era where, in select patient populations, treatment planning goals have shifted towards the achievement of treatment free remission (TFR) after TKI cessation. Both duration and depth of response to TKI therapy are predictors of future success in achieving a lasting TFR and with improved outcomes independent of TKI cessation. Unfortunately, molecular residual disease (MRD) persists in many patients despite optimal therapy and predicts for worse outcomes over time and decreased ability to maintain a TFR after TKI cessation. Achievement of a major molecular response (MMR) and probability of TFR have been associated with increased numbers of NK cells, particularly mature cytolytic NK cells. Kiadis K-NK003 cells are off-the-shelf NK cells from a universal donor expanded using PM21, proprietary membrane particles modified to express membrane bound IL-21 and 4-1bb ligand. The resulting expanded K-NK003 cells have a hyperfunctional phenotype that are simultaneously highly cytotoxic with high release of perforin and Granzyme B, and potent producers of the cytokines IFN-γ, TNF-α and IL-2. This is an open label, non-randomized, prospective phase I pilot study designed to evaluate safety and to examine whether the addition of K-NK003 to ongoing TKI therapy for CML patients with persistent MRD will allow patients to achieve MRD negative status. Patients will be treated with K-NK003 on day 1 of each 14 day cycle, for a total of 6 cycles, in conjunction with their ongoing TKI therapy. The primary endpoint is safety. The efficacy objective is to estimate the rate of optimal molecular responses (negative to at least MR4.5). Secondary and exploratory endpoints include the proportion of patients with a reduction in BCR-ABL transcripts and NK cell number and function. Adult patients with chronic phase CML who have been on TKI therapy for at least 1 year prior to enrollment in the study will be eligible. Patients must have been on their most recent TKI consistently for at least 6 months prior to enrollment on study and must be expected to remain on current TKI for the duration of the study. Patients with current accelerated or blast crisis phase disease will be excluded. Disclosures Rein: Celgene: Consultancy; Blueprint Medicine: Consultancy; Novartis: Consultancy; Clinical Care Options: Consultancy, Other: Speaker. Rizzieri:Bayer: Membership on an entity's Board of Directors or advisory committees; AROG: Membership on an entity's Board of Directors or advisory committees; Celltrion: Membership on an entity's Board of Directors or advisory committees; abbvie: Membership on an entity's Board of Directors or advisory committees; Mustang: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Sanofi: Membership on an entity's Board of Directors or advisory committees; Teva: Membership on an entity's Board of Directors or advisory committees; Acrobiotech: Membership on an entity's Board of Directors or advisory committees; Celgene: Membership on an entity's Board of Directors or advisory committees; Gilead: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Pfizer: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Incyte: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Stemline: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Seattle Genetics: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Jazz: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Kite: Honoraria, Speakers Bureau; Karyopharm: Honoraria, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau.
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Ilander, Mette Matilda, Ulla Olsson-Strömberg, Hanna Lähteenmäki, Tiina Kasanen, Perttu Koskenvesa, Stina Söderlund, Martin Hoglund, et al. "Disease Relapse After TKI Discontinuation In CML Is Related Both To Low Number and Impaired Function Of NK-Cells:Data From Euro-SKI." Blood 122, no. 21 (November 15, 2013): 379. http://dx.doi.org/10.1182/blood.v122.21.379.379.
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Abstract Background The inhibition of oncogenic BCR-ABL1 kinase with tyrosine kinase inhibitors (TKIs) has significantly improved the prognosis of CML. Recent reports suggest that approximately 40 % of CML patients who have achieved optimal therapy response (complete molecular remission, CMR) can stop imatinib treatment without recurrence of detectable BCR-ABL1 transcripts. However, no predictive prognostic factors for successful therapy discontinuation have yet been identified. We therefore set up an immunological substudy in the ongoing pan-European EURO-SKI stopping study. We aimed to identify predictive biomarkers for relapse and non-relapse after TKI discontinuation. In addition, we aimed to understand more on the mechanisms of immune surveillance in CML and to study the effects of TKI treatment on the immune system. Materials and methods Patients in deep molecular remission (MR4, BCR-ABL < 0,01% IS) for at least one year and with TKI treatment for at least 3 years were eligible for the clinical study. Basic lymphocyte immunophenotyping (the proportions and absolute numbers of NK-, T- and B-cells) was performed at the university hospital laboratories at the time of therapy discontinuation, and 1, 6, and 12 months after the TKI discontinuation. In a proportion of patients a more detailed immunophenotypic (analysis of CD45RA, CD57, CD27 and CD62L expressions) and functional analyses were done from fresh blood samples in a central immunology laboratory (Helsinki) at the same time points. The cytotoxicity of NK-cells was studied by measuring the direct killing of target cells (K562) and by the degranulation assay (CD107a/b expression). The secretion of Th1 type of cytokines IFN-γ/TNF-α was studied from both T- and NK-cells. Results Thus far the basic lymphocyte subclass measurement has been analyzed from 62 patients who have discontinued TKI treatment within the EURO-SKI study. Functional analyses have been performed from 30 patients. 60 patients have used imatinib before treatment discontinuation and 2 patients dasatinib. At baseline, before the treatment discontinuation both CD4+ and CD8+ T-cell counts were within the normal range (median CD4+ 0.73x 109/L, range 0.11-2.4x 109/L; CD8+ 0.35x 109/L, 0.07-1.92 x 109/L). The TKI stop had no significant numerical or functional effect on T-cells, and at 1 month time-point the median T-cell counts were unchanged (CD4+ 0.73x 109/L; CD8+ 0.35x 109/L). Similarly, at the baseline, the median NK-cell count was within a normal range (0.26 x 109/L, range 0.04-1.04 x 109/L), and no significant change was observed 1 month after stopping the treatment (median 0.29 x 109/L). Furthermore, at the baseline and at the 1-month time-point the cytotoxicity of NK-cells and the cytokine secretion of T- and NK-cells did not significantly differ from the healthy controls when all patients were considered as a one group. However, when patients were divided in two groups based on the relapse status, the patients who eventually relapsed had significantly fewer NK-cells already at the baseline (Figure A; absolute count 0.18x 109/L vs. 0.32 109/L, p=0.008; proportions 11% vs. 21%, p=0.001). The phenotype of NK-cells also differed between the two groups, and the patients who relapsed had less NK-cells expressing CD57 (median 58% vs. 69%, p=0.046) and CD16 (median 67% vs. 83%, p=0.018) on the cell surface. Furthermore, the cytotoxicity of NK-cells was impaired in patients who failed to discontinue the TKI treatment successfully and no killing activity was observed in their samples (Figure B; alive K652 cells after co-incubation with effector cells 100% vs. 88%, p=0.07). No clear differences were observed in the function or the numbers of T-cells between relapsing and non-relapsing patients. Conclusions The NK-cell numbers and their function may predict disease relapse after TKI discontinuation. This may have impact on the future stopping trials. In addition, it further illustrates the importance of the immune system in the successful long-term treatment of CML. Disclosures: Ekblom: Novartis: Honoraria; Bristol-Myers Squibb: Honoraria. Hjorth-Hansen:Pfizer, BMS: Honoraria, Travel expenses Other. Porkka:BMS: Honoraria, Research Funding; Novartis: Honoraria, Research Funding. Richter:Bristol-Myers Squibb: Consultancy, Speakers Bureau; Novartis: Consultancy, Research Funding, Speakers Bureau. Mustjoki:Novartis: Honoraria; BMS: Honoraria, Research Funding.
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Sweet, Kendra, Ehab L. Atallah, Jerry P. Radich, Mei-Jie Zhang, Eva Sahakian, Melanie Mediavilla-Varela, Alexis Vistocky, et al. "Second Treatment Free Remission after Combination Therapy with Ruxolitinib Plus Tyrosine Kinase Inhibitors in Chronic Phase Chronic Myeloid Leukemia (CML)." Blood 138, Supplement 1 (November 5, 2021): 2555. http://dx.doi.org/10.1182/blood-2021-147954.
Full textAbstract:
Abstract Background: Discontinuation of tyrosine kinase inhibitors (TKIs) is feasible in a subset of CML patients who have maintained a deep molecular response for at least two years. Numerous discontinuation trials have been performed and consistently show approximately 50% of patients relapse after stopping TKIs. A recent study examining rates of treatment free remission (TFR) after a second attempt at stopping TKIs found, with a median follow up time of 38.3 months, 64.3% of patients had a molecular relapse (defined as a loss of major molecular response (MMR)). At 12, 24 and 36 months, TFR rates were 48%, 42% and 35%, respectively. These data suggest some patients with a history of molecular relapse upon TKI cessation could successfully stop treatment on a subsequent attempt, yet the majority will relapse a second time. 'Complete eradication' of CML remains elusive in most patients likely as a result of minimal residual disease (MRD), which is the result of BCR-ABL independent drug resistance. More specifically, CML cells that reside in sanctuary sites such as the bone marrow adhere to fibronectin and demonstrate cell adhesion mediated drug resistance (CAM-DR). The bone marrow microenvironment contains many cytokines and growth factors capable of inducing STAT3-Y705 phosphorylation via the JAK-STAT pathway leading to protection against TKI-induced cell death. Inhibiting JAK2 and TYK2 leads to complete inhibition of pSTAT3-Y705, thereby implicating the role of activation of JAK2 and TYK2 in STAT3-Y705 phosphorylation and resistance towards BCR-ABL TKI-induced cell death. A phase I clinical trial combined ruxolitinib, which inhibits JAK2 and TYK2, plus nilotinib in chronic phase (CP) CML patients and found that ruxolitinib 15mg PO BID was safe and well tolerated with 4/10 patients achieving undetectable BCR-ABL1 transcripts by PCR. Study Design and Methods: This single arm phase II study (NCT03610971) will enroll 41 subjects from the H Jean Khoury Cure CML Consortium. Eligible subjects must have a confirmed diagnosis of CP-CML and have previously attempted to discontinue TKI therapy per NCCN guidelines and had molecular recurrence, defined as loss of MMR, and were restarted on TKI. This trial combines ruxolitinib 15mg BID plus BCR-ABL TKI (imatinib, dasatinib, nilotinib or bosutinib) for 12 28-day cycles in the combination treatment phase (CTP). RQ-PCR to measure BCR-ABL transcripts will be checked at screening and every three months during the CTP. In the event that a subject experiences intolerance to a TKI, has confirmed loss of MMR, or loss of MR4.5 (>0.0032% IS) on two central PCR results, or discontinues ruxolitinib, the subject will be removed from CTP and enter into long term follow-up (LTFU). CTP phase will be followed by further RQ-PCR screening for the concurrent TFR phase. At this time ruxolitinib will be discontinued and any subject who has met the criteria for the TFR phase will be enrolled. During the TFR phase, subjects will discontinue their TKI and be monitored off treatment with RQ-PCR checked monthly for the first year, every six weeks for year two, and every 12 weeks during year three. Upon molecular recurrence, defined as loss of MMR, TKIs will be restarted. The primary endpoint is the 12-month TFR rate subsequent to completion of 12 cycles of combination therapy; however, subjects will remain in the TFR phase for three years. Therefore, the total duration of the trial will be approximately five years (one year on CTP + three years in the TFR phase + one-year LTFU). Study statistical design was calculated to yield a one-sided type I error rate of 0.025 and power of 65% when the true one-year relapse rate is 35%. This study will additionally assess patient-reported outcomes in conjunction with RQ-PCR testing. PROMIS and other measures will be self-administered through REDCap. Correlative studies will include comparing changes in pSTAT3 in K562 and KU812 cell lines using plasma from CML patients being treated with TKIs plus ruxolitinib, using the plasma inhibitory assay technique. Changes in pSTAT3 and pSTAT5 will be correlated with clinical response and rate of TFR. Additional correlatives include multiparameter flow-based assessment of the T-cell compartment (activity/polarization) as well as natural killer cell fractions in CML patients at various time points (TKIs alone, TKIs plus ruxolitinib and during TFR). Thus far, 14 patients have been enrolled. Disclosures Sweet: Astellas: Consultancy, Membership on an entity's Board of Directors or advisory committees; Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees; AROG: Membership on an entity's Board of Directors or advisory committees; Gilead: Membership on an entity's Board of Directors or advisory committees; Bristol Meyers Squibb: Honoraria, Membership on an entity's Board of Directors or advisory committees. Atallah: Amgen: Consultancy; BMS: Honoraria, Speakers Bureau; Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Research Funding; Takeda: Consultancy, Research Funding; Abbvie: Consultancy, Speakers Bureau. Radich: Amgen: Membership on an entity's Board of Directors or advisory committees; Novartis: Membership on an entity's Board of Directors or advisory committees; Genentech: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees. Thompson: Novartis/ Bristol-Myers Squibb: Research Funding. Mauro: Pfizer: Consultancy; Takeda: Consultancy; Bristol Myers Squibb: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Sun Pharma / SPARC: Research Funding. Pinilla Ibarz: AbbVie, Janssen, AstraZeneca, Novartis, TG Therapeutics, Takeda: Consultancy, Other: Advisory; Sellas: Other: ), patents/royalties/other intellectual property; MEI, Sunesis: Research Funding; AbbVie, Janssen, AstraZeneca, Takeda: Speakers Bureau. OffLabel Disclosure: Ruxolitinib is being used off-label in chronic myeloid leukemia
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Abrishami, Mahsa, Landon Pastushok, Karen Mochoruk, John F. DeCoteau, and C. Ronald Geyer. "Synthetic Anti-IL3 Receptor Antibodies As Therapeutics to Block Innate Imatinib Resistance in Chronic Myelogenous Leukemia." Blood 124, no. 21 (December 6, 2014): 4519. http://dx.doi.org/10.1182/blood.v124.21.4519.4519.
Full textAbstract:
Abstract Chronic myelogenous leukemia (CML) is maintained by a minor population of leukemic stem cells (LSCs) that exhibit innate resistance to tyrosine kinase inhibitors (TKIs) targeting BCR-ABL. Innate resistance can be induced by cytokines and growth factors secreted by bone marrow stromal cells that protect CML-LSCs from TKIs, resulting in minimal residual disease. Developing therapies that can be combined with TKIs to eradicate TKI-insensitive CML-LSCs, is critical for disrupting innate TKI resistance and preventing disease relapse. We previously showed that interleukin-3 (IL-3) reduces the sensitivity of CML cells to BCR-ABL TKIs. Based on this, we hypothesized that inhibiting IL-3 receptor (IL-3R) would reduce the innate resistance of CML cells to BCR-ABL TKIs. We generated IL-3R inhibitors using synthetic human antibody technology. We constructed combinatorial libraries of human antibody antigen binding fragments (Fabs) using a single-framework synthetic Fab based on the optimized human 4D5 Fab framework that is derived from one of the most frequently used light chain (Vκ1-1) and heavy chain (VH3-23) germline sequences. We generated the Fab library by randomizing amino acids in the Fab heavy-chain complementarity regions (CDRs) and the L3 CDR of the light chain. In CDRs H1 and H2, solvent-accessible positions were softly randomized with binary degenerate codons that encode equal proportions of two amino acids, mostly tyrosine and serine. CDRs L3 and H3 were randomized using codon mixes that encoded pre-defined proportions of nine amino acids (Y (25%), S (20%), G (20%), A (10%), F (5%), W (5%), H (5%), P (5%) or V (5%)). CDR H3 contain different lengths ranging form 1 to 17 amino acids and CDR L3 contained different lengths ranging form 3 to 7 amino acids. Oligonucleotide directed mutagenesis was used to generate a combinatorial Fab library of 1010 members, where CDRs H1-3 and CDR L3 of the 4D5 Fab were replaced with the above amino acid combinations. The Fab library was expressed in E. coli as a fusion to the PIII coat protein of M13 filamentous phage. The Fab phage library was screened for interactions against the ectodomain of murine IL-3R (aa17-331). After four rounds of phage display selections, we analyzed CDR-H3 sequences in the Fab phage pool using the Ion Torrent Next Generation sequencing (NGS) platform. We chose the most abundant Fab to further mature by “softly” randomizing CDRs H1-3 and L3, where each codon was mutated to contain 70% of the wild-type nucleotide and 10% of each of the other three nucleotides. After three rounds of phage display selection, we analyzed the Fab phage pools by NGS and chose the most abundant Fab to convert to the IgG1 format. Dissociation constants for the IgGs generated from the original selection and the affinity maturation were 1.3x10-8 nM and 1.1x10-9 nM, respectively. The specificity of IL-3R antibodies was demonstrated by showing that they bound to HEK293 cells transiently transfected with IL-3R receptor and not to untransfected HEK293 cells. To investigate IL-3R antibody effects on IL-3 mediated innate TKI resistance, we compared its activity with imatinib against TonB210, a murine hematopoietic cell line with inducible BCR-ABL expression, in response to IL-3. Imatinib did not suppressed BCR-ABL(+) Ton-B210 growth, and IL-3 protected against imatinib growth suppression. IL-3R antibody suppressed the ability of IL-3 to block imatinib activity of BCR-ABL(+) TonB210 cells. We then compared effects of IL-3R antibody as a single agent, and in combination with imatinib, on the growth of CML-LSCs derived from an established murine retroviraltransduction/transplantation model of CML blast crisis, in response to IL-3. The presence of IL-3 reduced cytotoxicity, apoptosis induction, and colony formation by imatinib. Treatment with IL-3R antibody reduced the protective effect of IL-3 in these assays. In summary, our findings demonstrate that synthetic human antibodies against IL-3R can effectively block IL-3 mediated innate TKI resistance in in vitro assays and suggest that IL-3R antibodies may be effective as a co-drug to complement TKIs in CML treatment by disrupting the innate resistance of CML-LSCs. Interestingly, the human IL3-R antibodies isolated against murine IL-3R by phage display also recognize human IL-3R, suggesting they recognize a conserved epitope and that they may also be useful in developing human therapies. Disclosures No relevant conflicts of interest to declare.
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Nievergall, Eva, John Reynolds, Chung H. Kok, Dale Watkins, Mark Biondo, Samantha J. Busfield, Gino Vairo, et al. "High Plasma Levels of TGF-α and IL-6 at Diagnosis Predict Early Molecular Response Failure and Transformation in CML." Blood 124, no. 21 (December 6, 2014): 1788. http://dx.doi.org/10.1182/blood.v124.21.1788.1788.
Full textAbstract:
Abstract Introduction: Early molecular response (EMR, BCR-ABL (IS) ≤ 10% at 3 months) is a strong predictor of outcome in imatinib-treated chronic phase chronic myeloid leukemia (CP-CML) patients, but for patients who transform early 3 months may be too late for effective therapeutic intervention. Thus, alternative approaches are required to identify poor responders at the time of diagnosis. The aim of this study was to identify plasma biomarkers at diagnosis that will predict for subsequent EMR failure, early transformation or the development of BCR-ABL1 kinase domain mutations. Cytokine profiling has proven valuable in identifying prognostic factors in myelofibrosis and myelodysplastic syndromes; however, similar comprehensive studies are lacking to date in CML. Methods: Plasma samples from CP-CML patients enrolled to the TIDEL II trial were collected prior to starting imatinib treatment (n=186) and after 6 months on TKI (n=17); and compared to those of healthy donors (n=19). The levels of 39 cytokines, chemokines and growth factors (CC&GF) were measured using a Luminex multiplex assay. To identify potential biomarkers to predict EMR failure, random forest analysis and recursive partitioning techniques in R were applied as statistical methods. Results: Plasma concentrations of 13/39 CC&GF were significantly elevated at CP-CML diagnosis compared to healthy donor samples. Most (EGF, bFGF, VEGF, TGF-α, CXCL1, CCL4, sCD40L and IL-4) werenormalized after 6 months of TKI treatment while others (TNF-α, sIL-2Ra, IL-8, IL-10, IL-1a) remained at higher levels, possibly reflecting persistent disease-induced alterations within the microenvironment. A third subset of CC&GF, such as CCL2, CCL3 and CCL22, showed higher circulation levels only in TKI-treated patients but not at diagnosis, suggesting that changes in these CC&GF could be treatment-related. 183/186 patients had BCR-ABL1 assessments available at 3 months, and 23/183 (13%) did not achieve EMR. Random forest analysis identified TGF-α, IL-6 and IFN-α as the most important CC&GF associated with EMR failure. Recursive partitioning incorporating these three variables produced a classification tree based only on TGF-α and IL-6, and demonstrated that 12/20 (60%) of patients who were TGF-αhi/IL-6hi failed to achieve EMR (Table 1). Importantly, this group contained 3/3 (100%) patients who transformed within the first 12 months of TKI treatment. Both TGF-α (7.99 vs. 60.57 pg/ml, p<0.001) and IL-6 (0.26 vs. 8.35 pg/ml, p=0.004) were increased in plasma samples of EMR failure patients compared to those who achieved EMR, and this was independent of white cell count (both for TGF-α and IL-6) and Sokal Score (TGF-α only). While less compelling, high TGF-α alone was associated with EMR failure in 15/39 (38%) patients compared to 8/144 (6%) in the TGF-αlo group (p<0.001) and reduced transformation-free survival (92% vs. 99%, p=0.029). Interestingly, the TGF-αlo group also had higher rates of complete molecular response (38% vs. 8%, p=0.005). While the relevance of IL-6 in CML pathogenesis has been previously proposed the association of the EGFR ligand TGF-α with CML treatment outcomes is novel. Conclusion: These data highlight for the first time the prognostic value of cytokine profiling in CML patients. The combination of TGF-α and IL-6 plasma levels at CML diagnosis is strongly predictive for EMR, transformation and CMR. While TGF-α alone delineates a poor response group of patients, the addition of IL-6 provides significant additional power and strongly selects for high-risk patients who may benefit significantly from very early proactive intervention. Thus, incorporation of these simple measurements to the diagnostic work-up of CP-CML patients may enable therapy intensity to be individualized according to the risk profile of the patient. Abstract 1788. Table 1: Treatment outcomes in patients grouped according to predictive plasma biomarkers N(of 183 total patients) EMR failure Trans-formation Mutation MMR by 24 months CMR by 24 months + TGF-αhi/IL-6hi 20 12 (60%) 3# 5 9 (45%) 1 (5%) TGF-αhi/IL-6lo 19 3 (16%) 0 0 12 (63%) 2 (11%) TGF-αlo 144 8 (6%) 2## 6 117 (81%) 55 (38%) Overall p value (log-rank) < 0.001* <0.001 <0.001 0.004 0.004 MMR – major molecular response CMR – complete molecular response (undetectable BCR-ABL1) * p value is only indicative as the groups were formed based on EMR # all early transformations (< 1 year of TKI treatment) ## all late transformations Disclosures Reynolds: Novartis: Shareholder Other. Biondo:CSL Limited: Employment. Busfield:CSL Limited: Employment. Vairo:CSL Limited: Employment. Yeung:Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding. White:Novartis: Honoraria, Research Funding; BMS: Honoraria, Research Funding; Ariad: Research Funding. Hughes:Novartis: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; BMS: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding; Ariad: Honoraria, Membership on an entity's Board of Directors or advisory committees; Pfizer: Membership on an entity's Board of Directors or advisory committees.
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Hayashi, Yoshiki, Hirohisa Nakamae, Takako Katayama, Takahiko Nakane, Hideo Koh, Mitsutaka Nishimoto, Mika Nakamae, et al. "Immunoprofile of Patients with Chronic Myeloid Leukemia Treated with Imatinib, Nilotinib or Dasatinib,." Blood 118, no. 21 (November 18, 2011): 3764. http://dx.doi.org/10.1182/blood.v118.21.3764.3764.
Full textAbstract:
Abstract Abstract 3764 Recent reports showed that dasatinib induces significant immunostimulation with clonal expansion of large granular lymphocytes (LGL) which, in chronic myeloid leukemia (CML), is related to both better prognosis and to autoimmune-like side effects. It is speculated that lower levels of circulating T-regulatory cells play a partial role in LGL proliferation in patients receiving dasatinib. The immunoprofile was studied using flow cytometry to evaluate lymphocyte subsets and NK-cell reactivity in the peripheral blood of 61 patients in the chronic phase of CML during treatment with a tyrosine kinase inhibitor (TKI) (Median age: 58 years; imatinib 36, nilotinib 9, dasatinib 16). Furthermore, we measured plasma levels of 27 types of cytokines or chemokines in 58 patients in the chronic phase of CML so that a comprehensive comparison could be made of the differences in immunoprofile among the patients receiving these three TKIs. There were no significant differences between the three TKI-treated groups in terms of CD4/8 ratios or the number of T-cells (CD3+CD8+ or CD4+) and NK cells (CD3-CD56+). However, the number of NK-LGL (CD56+CD57+) and T-LGL (CD3+CD57+) increased significantly in the group that received dasatinib. Furthermore, dasatinib significantly enhanced NK-cell reactivity as compared to imatinib and nilotinib. In contrast nilotinib significantly suppressed NK-cell reactivity (E/T ratio =10:1: Median (interquartile range), 8.7% (5.0–16.2), 5.2% (4.8–11.4), 20.8% (13.4–33.3), for imatinib, nilotinib and dasatinib, respectively). In addition, the number of regulatory T-cells (CD4+CD25int-hiCD127low) was similar among the three groups (Median (interquartile range), 36/mm3 (27–53), 48/mm3 (34–60), 39/mm3 (26–53), for imatinib, nilotinib and dasatinib, respectively). Furthermore, in the analysis of cytokines and chemokines, plasma levels of IL-8, IP-10, and MCP-1 were significantly elevated while the level of PDGF-bb was significantly decreased in all three groups compared to those of healthy control. Plasma levels of IL-1 beta, IFN-gamma, and FGF-basic were significantly decreased in only the dasatinib group compared to those of control (P=.02, P=.04, P=.03, respectively). In addition, plasma levels of GM-CSF were significantly elevated in the imatinib and dasatinib groups (Median (interquartile range), 6.1 pg/ml (2.7–11.7) and 7.9 pg/ml (4.5–8.2), P=.02, and P=.03, respectively) but not in the nilotinib group (P=.34) when compared to those of control. In the multiple regression analysis that evaluated the relationship between NK-reactivity and cytokines or chemokines in the patients receiving dasatinib, only plasma levels of GM-CSF were significantly associated with NK-reactivity (P=.03). Notably, in our data, dasatinib and nilotinib exerted opposite effects on NK-cell reactivity, expansion of LGL, and showed different cytokine and chemokine profiles. Based on our results, the activation of NK-cell reactivity induced by dasatinib might be caused by a mechanism other than a decrease in the number of regulatory T-cells. Additionally, in an unphysiological immunological status mediated by dasatinib, GM-CSF might make some contribution to NK-cell reactivity. Disclosures: Nakamae: BMS: Research Funding, Speakers Bureau; Novartis: Research Funding, Speakers Bureau. Hino:BMS: Research Funding, Speakers Bureau; Novartis: Research Funding, Speakers Bureau.
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Mauro, Michael J., Jorge E. Cortes, Hagop M. Kantarjian, Neil P. Shah, Dale L. Bixby, Ian W. Flinn, Thomas O'Hare, et al. "Ponatinib in Chronic-Phase Chronic Myeloid Leukemia Patients: Final Report from a Phase 1 Trial." Blood 128, no. 22 (December 2, 2016): 3063. http://dx.doi.org/10.1182/blood.v128.22.3063.3063.
Full textAbstract:
Abstract Background: Ponatinib, an oral tyrosine kinase inhibitor with potent activity against native and mutant BCR-ABL1, is approved for patients with refractory chronic myeloid leukemia (CML) or Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL) for whom no other tyrosine kinase inhibitor (TKI) therapy is indicated, or for patients with the T315I mutation. The efficacy and safety of ponatinib in patients with resistant/refractory hematologic malignancies were evaluated in a phase 1 trial (NCT00660920). Here, we report 4-year follow-up data from chronic-phase (CP)-CML patients; final data (approximately 5-year follow-up) will be presented. Methods: In this open-label, dose-escalation, phase 1 trial, 81 patients with resistant/refractory hematologic malignancies (CP-CML, 43 patients; accelerated-phase CML, 9 patients; blast-phase CML, 8 patients; Ph+ ALL, 5 patients) were enrolled. Patients were treated with ponatinib at a starting dose of 2 mg/d - 60 mg/d; intra-patient dose escalation was permitted. In Oct 2013, dose reduction instructions were provided in response to an observed accumulation of arterial occlusive events (AOEs) with longer follow-up across the ponatinib clinical program. For data presented herein, the data cutoff date is 2 Feb 2015, with median follow-up of 53.1 months (range, 1.7 - 69.9 months) for CP-CML patients. Results: Among CP-CML patients, at baseline, median age was 55 years and median time since diagnosis was 6.6 years; BCR-ABL1 kinase domain mutations were reported in 63% of patients, with T315I confirmed at a central laboratory in 28% of patients. Patients were heavily pretreated, with 37% having received 2 prior TKIs and 60% having received ≥3 prior TKIs. Of 43 CP-CML patients, 22 (51%) remained on ponatinib treatment at data cutoff. Adverse events (AEs; 26%) and disease progression (9%) were the most common reasons for discontinuation of treatment. Cumulative response rates were: major cytogenetic response (MCyR), 72%; complete cytogenetic response (CCyR), 65%; major molecular response (MMR; assessed at a central laboratory), 56%; molecular response 4 (MR4), 42%; MR4.5, 28%. Responses were durable (Table), with median durations of response not reached for MCyR, CCyR, and MMR. Among patients who received ponatinib at starting doses of ≤30 mg/d (n = 15), MCyR was achieved by 67%, CCyR by 53%, and MMR by 47%; ponatinib dose was ≤30 mg/d in all but one of these patients at the time of response. Of 19 patients who were ongoing and in MCyR as of Oct 2013, 13 had their dose reduced; all 13 dose-reduced patients maintained MCyR at data cutoff. Of the 22 ongoing patients at the time of the present analysis, 18 (82%) had CCyR and 17 (77%) had MMR or better (MMR, 6 patients; MR4, 1 patient; MR4.5, 9 patients; MR5, 1 patient) as their response at the data cutoff; 14/22 (64%) ongoing patients were receiving 15 mg/d as their current dose as of the data cutoff. Rash (65%), fatigue (63%), abdominal pain (58%), headache (58%) and arthralgia (53%) were the most common treatment-emergent AEs. The incidence of AOEs (any/serious) was 40%/30% (by subcategory: cardiovascular, 30%/21%; cerebrovascular, 9%/7%; peripheral vascular, 14%/9%). Conclusions: With median follow-up of over 4 years in this phase 1 study, ponatinib continues to provide clinical benefit to heavily pre-treated CP-CML patients, approximately half of whom continue to receive ponatinib, with a majority in deep response that has been long-lasting; final study data will be presented. The most common treatment-emergent AEs were consistent with the AE profile across the clinical program. Potential for long-term benefit, demonstrated herein, versus risk should be considered when using ponatinib in this patient population. Study sponsor: ARIAD Pharmaceuticals, Inc. Disclosures Mauro: BMS: Consultancy, Honoraria; ARIAD: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Research Funding; Pfizer: Consultancy, Honoraria. Cortes:ARIAD: Consultancy, Research Funding; Bristol-Myers Squib: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Teva: Research Funding. Kantarjian:Bristol-Myers Squibb: Research Funding; Amgen: Research Funding; ARIAD: Research Funding; Pfizer Inc: Research Funding; Delta-Fly Pharma: Research Funding; Novartis: Research Funding. Shah:ARIAD: Research Funding; BMS: Research Funding; Daiichi-Sankyo: Research Funding; Pfizer: Research Funding; Plexxikon: Research Funding. Flinn:Janssen: Research Funding; Pharmacyclics LLC, an AbbVie Company: Research Funding; Gilead Sciences: Research Funding; ARIAD: Research Funding; RainTree Oncology Services: Equity Ownership. Rivera:ARIAD: Employment, Equity Ownership. Lustgarten:ARIAD: Employment, Equity Ownership. Santillana:ARIAD: Employment, Equity Ownership. Heinrich:Novartis: Consultancy, Patents & Royalties, Research Funding; Pfizer: Consultancy; Bayer: Research Funding; BMS: Research Funding; Blueprint Medicines: Consultancy; MolecularMD: Consultancy, Equity Ownership; ARIAD: Consultancy, Research Funding; Onyx: Consultancy. Druker:Agios: Honoraria; Ambit BioSciences: Consultancy; ARIAD: Patents & Royalties, Research Funding; Array: Patents & Royalties; AstraZeneca: Consultancy; Blueprint Medicines: Consultancy, Equity Ownership, Other: travel, accommodations, expenses ; BMS: Research Funding; CTI: Equity Ownership; Curis: Patents & Royalties; Cylene: Consultancy, Equity Ownership; D3 Oncology Solutions: Consultancy; Gilead Sciences: Consultancy, Other: travel, accommodations, expenses ; Lorus: Consultancy, Equity Ownership; MolecularMD: Consultancy, Equity Ownership, Patents & Royalties; Novartis: Research Funding; Oncotide Pharmaceuticals: Research Funding; Pfizer: Patents & Royalties; Roche: Consultancy. Deininger:Pfizer: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Novartis: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Gilead: Research Funding; Incyte: Consultancy, Membership on an entity's Board of Directors or advisory committees; BMS: Consultancy, Research Funding; CTI BioPharma Corp.: Membership on an entity's Board of Directors or advisory committees; Celgene: Research Funding; Bristol Myers Squibb: Consultancy, Research Funding; Ariad: Consultancy, Membership on an entity's Board of Directors or advisory committees. Talpaz:Novartis: Research Funding; Incyte Corporation: Other: Travel expense reimbursement, Research Funding; Ariad: Other: Expense reimbursement, travel accomodation expenses, Research Funding; Pfizer: Consultancy, Other: travel accomodation expenses, Research Funding.
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Toda, Jun, Michiko Ichii, Hirohiko Shibayama, Hideaki Saito, Yuichi Kitai, Ryuta Muromoto, Jun-ichi Kashiwakura, et al. "Role of Signal Transducing Adaptor Protein-1 (STAP-1) in Chronic Myelogenous Leukemia Stem Cells." Blood 132, Supplement 1 (November 29, 2018): 4245. http://dx.doi.org/10.1182/blood-2018-99-113725.
Full textAbstract:
Abstract Chronic myelogenous leukemia (CML) is a clonal myeloproliferative disorder caused by hematopoietic stem cells expressing the BCR-ABL fusion oncoprotein, which constitutively activates multiple signal transduction pathways such as mitogen-activated protein kinase, phosphatidylinositol 3-kinase/Akt, and Janus kinase/signal transducer and activator of transcription (JAK/STAT). Although tyrosine kinase inhibitor (TKI) therapy results in dramatic clinical success, studies have shown that TKIs are unable to eradicate leukemic stem cells (LSCs). Several key signaling molecules and pathways have been proposed to regulate the survival of CML LSCs in the presence of TKI; however, the details remain unclear. It is necessary to elucidate the mechanisms that maintain LSCs to better understand the pathogenesis of CML and develop new treatment approaches. The family of signal-transducing adaptor proteins (STAPs), which includes STAP-1 and STAP-2, has been implicated in various intracellular signaling pathways. In 2003, we cloned STAP-2 as a c-fms interacting protein and reported that STAP-2 binds to BCR-ABL and enhances activity, leading to the activation of downstream molecules such as ERK, STAT5, BCL-xL, and BCL2. STAP-1 was cloned as a c-kit interacting protein from a hematopoietic stem cell library, but it is unknown whether STAP-1 plays a role in CML. Given the structural homology between STAP-1 and STAP-2 and the hematopoietic expression of STAP-1, we hypothesized that STAP-1 might contribute to the leukemogenesis of CML. A STAP-1-deficient (KO) CML mouse model was developed. To generate this model, lineage (Lin)− Sca-1+ c-Kithigh (LSK) fraction isolated from bone marrow (BM) cells was infected with a retrovirus carrying BCR-ABL1 and GFP and subsequently transplanted into congeneric recipients. STAP-1 KO CML mice showed significantly longer survival than WT CML mice and displayed less severe splenomegaly and lung hemorrhages compared with WT mice. In recipient BM, absolute numbers of STAP-1 KO LSCs (GFP+ LSK cells) were significantly lower than WT LSCs. In the colony-forming assay, STAP-1 KO LSCs generated fewer colonies compared to WT LSCs. Using flow cytometric analysis, we found that STAP-1 KO LSCs had a higher apoptotic rate than WT LSCs. These findings suggest that the suppression of apoptosis induced by STAP-1 mediates longer survival of LSCs. To further understand the effects of STAP-1, we performed a gene expression analysis using RNA-sequence (RNA-seq) and compared WT and STAP-1 KO CML LSCs. When canonical pathways were analyzed with Ingenuity Pathway Analysis, various pathways associated with inflammatory cytokines were observed to be regulated in STAP-1 KO CML LSCs. Changes in mRNA expression, including that of SOS1, SOS2, FOXO3, FASLG, NFKB2, and BCL-xL, indicated that the PTEN signaling pathway, known to play a tumor suppressive role in CML, was significantly activated by STAP-1 KO (p=1.096E-3, activation Z-score=2.611). The pathway related to JAK/STAT signaling was also affected (p=2.04E-5, activation Z-score=-3.286). Downstream genes in the JAK/STAT signaling pathway, such as STAT5B and BCL-xL, were downregulated more than 2-fold in STAP-1 KO LSCs, suggesting that the deletion of STAP-1 inhibits the expression of STAT5-targeted anti-apoptotic protein and induced apoptosis of CML LSCs. To confirm the results of the RNA-seq experiment, an intracellular flow cytometric assay with CML Lin− cells was conducted. The frequency of cells positive for phosphorylated STAT5 was reduced for STAP-1 KO compared with that for WT. Quantitative PCR with CML LSCs confirmed the downregulation of BCL2 and BCL-xL, which are STAT5-targeted anti-apoptotic genes, in STAP-1 KO CML LSCs. In conclusion, we show that STAP-1 plays a crucial role in the maintenance of CML LSCs using a murine model of CML. STAP-1 deficiency results in the reduction of phosphorylated STAT5, downregulation of anti-apoptotic genes BCL-2 and BCL-xL, and induced apoptosis of CML LSCs. These findings suggest that STAP-1 and related signaling pathways could be potential therapeutic targets for CML LSCs. Disclosures Ichii: Celgene K.K.: Speakers Bureau; Kowa Pharmaceutical Co.,LTD.: Speakers Bureau; Novartis Pharma K.K.: Speakers Bureau. Shibayama:Fujimoto Pharmaceutical: Honoraria, Research Funding; Takeda Pharmaceutical Co.,LTD.: Honoraria, Research Funding; Celgene K.K.: Honoraria, Research Funding; Jansen Pharmaceutical K.K: Honoraria; Ono Pharmaceutical Co.,LTD: Honoraria, Research Funding; Novartis Pharma K.K.: Honoraria, Research Funding; Mundipharma K.K.: Honoraria, Research Funding; Bristol-Meyer Squibb K.K: Honoraria, Research Funding. Oritani:Novartis Pharma: Speakers Bureau. Kanakura:Alexion Pharmaceuticals, Inc.: Consultancy, Honoraria, Research Funding.
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Cortes, Jorge E., Jane Apperley, Andreas Hochhaus, Michael J. Mauro, Philippe Rousselot, Tomasz Sacha, Moshe Talpaz, et al. "Outcome By Mutation Status and Line of Treatment in Optic, a Dose-Ranging Study of 3 Starting Doses of Ponatinib in Patients with CP-CML." Blood 136, Supplement 1 (November 5, 2020): 44–45. http://dx.doi.org/10.1182/blood-2020-135883.
Full textAbstract:
Introduction: In PACE (NCT01207440), patients with refractory chronic-phase chronic myeloid leukemia (CP-CML) with substantial prior second-generation tyrosine kinase inhibitor (TKI) treatment demonstrated deep, lasting responses to ponatinib. However, long-term follow-up identified rates of arterial occlusive events (AOEs) as a risk. OPTIC (NCT02467270) is a randomized Phase 2 trial evaluating ponatinib at 3 starting doses: 45 mg, 30 mg, and 15 mg daily in patients with CP-CML resistant/intolerant to ≥2 TKIs or with a T315I mutation. The interim analysis showed that the 45-mg (vs 30 mg or 15 mg) starting dose (with reduction to 15 mg upon response) provided the best clinical outcomes and responses were maintained in >75% of patients who dose reduced. Here, we present efficacy and safety outcomes by baseline mutation status and line of treatment for the 3 dose cohorts. Methods: Patients with CP-CML resistant/intolerant to ≥2 TKIs or with T315I mutation were randomized to ponatinib starting doses of 45 mg (Cohort A; 45 mg → 15 mg), 30 mg (B; 30 mg → 15 mg), and 15 mg (C) once daily (qd). Doses were reduced to 15 mg on achievement of ≤1% BCR-ABL1ISin Cohorts A and B. Doses also could be reduced for safety. The primary endpoint is ≤1% BCR-ABL1IS at 12 months. In this analysis, the outcome was analyzed by baseline mutation status (none, any, T315I, and non-T315I) and number of prior TKIs (≤2 or ≥3) in the intent-to-treat (ITT) population. Treatment-emergent adverse events (TEAEs), serious TEAEs, and AOEs by adjudication were summarized by number of prior TKIs (≤2 or ≥3). Interim analysis results are descriptive. Results: Patients (N=283) were randomized: A/B/C n=94/95/94; median age was 48 y (18‒81 y). Seven patients were excluded from the intent-to-treat population (N=276) because they had atypical transcripts. Mutation status was well balanced between cohorts; 59% had no mutation, 41% had ≥1 baseline mutation, 24% had T315I, and 17% had a non-T315I mutation. In all categories of mutation status, the rate of ≤1% BCR-ABL1IS by 12 months was highest in Cohort A, with the most notable differences seen in patients with T315I (A: 60%, B: 25%, C: 6%) (Table 1). Patients with no mutations or other mutations had smaller differences but the outcomes all still favored 45 mg. Patients in all cohorts were treated with multiple TKIs, with 54% (A), 60% (B), and 53% (C) having 3 or more prior TKIs. The rate of ≤1% BCR-ABL1IS by 12 months was highest in Cohort A, both in patients treated with ≤2 or ≥3 prior TKIs (43% and 49%, respectively) (Table 1). Table 2 shows rates of TEAEs and TE-AOEs by cohort and number or prior TKIs. There was a trend toward higher event rates in Cohort A and for patients treated with ≥3 TKIs. Rates of adjudicated AOEs were low (≤6%) in all 3 cohorts irrespective of the number of prior TKIs. Conclusions: At this interim analysis with a median follow-up of ~21 months, the maximum benefit:risk, regardless of mutation status or number of prior TKIs, was observed in patients treated with a 45-mg starting dose, with a reduction to 15 mg upon achievement of response. Patients with the T315I mutation who initiated ponatinib at 45 mg experienced better response rates than those who initiated ponatinib at 30-mg or 15-mg starting doses. Primary analysis will provide a refined understanding of the benefit:risk profile of 3 different starting doses of ponatinib. Disclosures Cortes: BioPath Holdings: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Takeda: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Pfizer: Consultancy, Research Funding; Sun Pharma: Research Funding; Telios: Research Funding; Amphivena Therapeutics: Research Funding; Astellas: Research Funding; Bristol-Myers Squibb: Research Funding; BiolineRx: Consultancy, Research Funding; Arog: Research Funding; Jazz Pharmaceuticals: Consultancy, Research Funding; Daiichi Sankyo: Consultancy, Research Funding; Immunogen: Research Funding; Merus: Research Funding. Apperley:Bristol Myers Squibb: Honoraria, Speakers Bureau; Incyte: Honoraria, Research Funding, Speakers Bureau; Novartis: Honoraria, Speakers Bureau; Pfizer: Honoraria, Research Funding, Speakers Bureau. Hochhaus:Novartis: Honoraria, Research Funding; Incyte: Honoraria, Research Funding; Bristol-Myers Squibb: Honoraria, Research Funding; Pfizer: Honoraria, Research Funding; Takeda: Honoraria; MSD: Research Funding. Mauro:Sun Pharma/SPARC: Research Funding; Pfizer: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Takeda: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Novartis: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding; Bristol-Myers Squibb: Consultancy, Honoraria, Other: Travel, Accommodation, Expenses, Research Funding. Rousselot:Bristol-Myers Squibb: Consultancy; Pfizer: Consultancy, Research Funding; Novartis: Consultancy; Takeda: Consultancy; Incyte: Consultancy, Research Funding. Sacha:Incyte: Consultancy, Honoraria, Speakers Bureau; Bristol-Myers Squibb Company: Consultancy, Honoraria, Speakers Bureau; Adamed: Consultancy, Honoraria; Novartis: Consultancy, Honoraria, Speakers Bureau; Pfizer: Consultancy, Honoraria, Speakers Bureau. Talpaz:Novartis: Research Funding; IMAGO: Consultancy; Constellation Pharmaceuticals: Membership on an entity's Board of Directors or advisory committees; BMS: Membership on an entity's Board of Directors or advisory committees; Takeda: Research Funding. Chuah:Korea Otsuka Pharmaceutical: Honoraria; Pfizer: Other: Travel, Research Funding; Novartis: Honoraria; Bristol-Myers Squibb: Honoraria, Research Funding. Lipton:Bristol-Myers Squibb: Honoraria; Pfizer: Consultancy, Honoraria, Research Funding; BMS: Consultancy, Research Funding; Novartis: Consultancy, Research Funding; Ariad: Consultancy, Research Funding; Takeda: Consultancy, Honoraria, Research Funding. Deininger:SPARC: Research Funding; Novartis: Consultancy, Other, Research Funding; Sangamo: Consultancy, Membership on an entity's Board of Directors or advisory committees; Incyte: Consultancy, Honoraria, Other, Research Funding; Blueprint Medicines Corporation: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Other: part of a study management committee, Research Funding; Fusion Pharma: Consultancy; Takeda: Consultancy, Membership on an entity's Board of Directors or advisory committees, Research Funding; Medscape: Consultancy; DisperSol: Consultancy; Leukemia & Lymphoma Society: Research Funding; Gilead Sciences: Research Funding; Ariad: Consultancy, Honoraria, Other; Bristol-Myers Squibb: Consultancy, Honoraria, Other, Research Funding; Galena: Consultancy, Honoraria, Other; Celgene: Research Funding; Pfizer: Honoraria, Other, Research Funding. Schiffer:BMS: Consultancy; Novartis: Consultancy; Takeda: Research Funding. García Gutiérrez:Pfizer: Honoraria, Other: travel/accommodations/expenses, Research Funding; Novartis Pharma AG: Consultancy, Honoraria, Other: travel/accommodations/expenses, Research Funding; Incyte: Consultancy, Honoraria, Other: travel/accommodations/expenses, Research Funding. de Lavallade:Novartis: Honoraria; Pfizer: Honoraria; BMS: Honoraria, Research Funding; Incyte: Honoraria, Research Funding. Etienne:Bristol-Myers Squibb: Consultancy, Research Funding, Speakers Bureau; Novartis: Consultancy, Research Funding, Speakers Bureau; Pfizer: Consultancy, Speakers Bureau; Incyte: Consultancy, Speakers Bureau. Lu:Millennium Pharmaceuticals, Inc., a wholly owned subsidiary of Takeda Pharmaceutical Company Limited: Current Employment. Srivastava:Takeda: Current Employment. Rosti:Novartis: Speakers Bureau; Incyte: Speakers Bureau; Bristol-Myers Squibb: Speakers Bureau; Pfizer: Research Funding, Speakers Bureau.
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Mackenzie, Ryan, Jennifer Snead, Jonathan VanDyke, and Michael Deininger. "Defining Extrinsic Survival Pathways in CML Progenitor Cells." Blood 114, no. 22 (November 20, 2009): 941. http://dx.doi.org/10.1182/blood.v114.22.941.941.
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Abstract Abstract 941 Defining extrinsic survival pathways in CML progenitor cells Ryan MacKenzie1, Jennifer Snead1, Jonathan VanDyke1, Brian Druker1,2, Michael Deininger1 1Center for Hematologic Malignancies, Oregon Health & Science University Cancer Institute, 2Howard Hughes Medical Institute, 3181 S.W. Sam Jackson Park Rd., Portland, Oregon 97239-3098, USA Background: The stem cell niche is thought to provide a protective microenvironment that allows for the persistence of malignant progenitors in various types of leukemia treated with chemotherapy. Tyrosine kinase inhibitor (TKI) treatment of chronic myeloid leukemia (CML) does not completely eliminate residual BCR/ABL positive progenitors, despite complete cytogenetic responses in the majority of patients, a phenomenon referred to as disease persistence. We hypothesize that niche factors may contribute to CML progenitor survival in the presence of imatinib and that the identification of defined survival factors may lead to therapeutic approaches to eliminate residual disease. Approach and results: In an initial series of experiments CD34+ CML progenitor cells were cultured in (a) serum free media supplemented with low concentrations of cytokines (=control), (b) in direct contact with M210B4 murine stroma or (c) separated from M210B4 murine stroma through a transwell membrane, in the presence or absence of TKIs (dasatinib, imatinib). After 96h, cells were plated in semisolid media and CFU-GM scored. Murine stroma consistently increased CFU-GM survival 4-5-fold compared to controls, with most of the effect attributable to soluble factors. Similar observations were made with human stroma (HS-5), although direct contact accounted for approximately 40% of the protective effect. Since integrin-mediated adhesion has been shown to protect CML cells lines from imatinib and chemotherapy, we hypothesized that direct contact protection of CML CD34+ cells may be related to integrin engagement. To test this hypothesis we cultured CML CD34+ cells on plates coated or not with fibronectin and in the presence or absence of integrin activating (B44) or blocking antibodies (6S6), with imatinib added at 1.5 and 5 uM or dasatinib added at 5 or 50 nM. However, in several experiments (N=3), we found that integrin-mediated adhesion to fibronectin failed to promote CFU-GM survival in the presence of TKIs. To identify soluble factors responsible for progenitor cell survival in the presence of TKIs we measured cytokine and chemokine concentrations in HS-5 stroma conditioned media (CM) and found measurable levels of several cytokines with JAK2-dependent signal transduction, including IL3, IL6, GM-CSF and G-CSF. Based on this we asked whether a JAK2 inhibitor (CYT387) would synergize with imatinib to inhibit CML CFU-GM survival. Thus, CD34+ cells were cultured for 96h with imatinib and CYT387 HS-5 stroma cells. Imatinib (5uM) reduced CFU-GM survival by 79.4%. Addition of 5 uM of CYT387, a JAK2 inhibitor (IC50 for growth inhibition of BAF3 cells expressing JAK2V617F: 1.5uM) reduced CFU-GM survival by 92.9%. Combination of imatinib with graded concentrations of CYT387 led to a further reduction of CFU-GM in a dose-dependent manner (Table 1). Conclusions: (a) Contact to and soluble factors produced by stromal cells appear to protect CD34+ CML progenitor cells in the presence of TKIs. (b) We find no evidence that integrin-mediated adhesion to fibronectin is involved in the protection of primary cells (unlike BCR-ABL expressing cell lines). (c) Combination of a JAK2 inhibitor with imatinib synergistically reduces colony formation, suggesting that this combination may be effective in targeting residual leukemia cells in vivo. Disclosures: Deininger: Novartis: Consultancy; Bristol-Myers Squibb: Consultancy; Calistoga: Research Funding; Genzyme: Research Funding.
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Hi Wase, Devendra K., Deborah L. White, Verity A. Saunders, Junia V. Melo, Sharad Kumar, and Timothy P. Hughes. "Short –Term Intense Bcr-Abl Kinase Inhibition Is Adequate to Trigger Cell Death in CML Cell Lines but Not in CML-CD34+ Cells Unless They Are Growth Factor Deprived." Blood 112, no. 11 (November 16, 2008): 1086. http://dx.doi.org/10.1182/blood.v112.11.1086.1086.
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Abstract After 5 years of imatinib treatment, only a minority of newly diagnosed chronic myeloid leukemia chronic phase (CML-CP) patients achieve complete molecular response. Imatinib has antiproliferative effects, but may not be able to eradicate CML-stem cells. Preclinical studies of imatinib suggested that sustained BCR-ABL kinase inhibition was required to block proliferation and induce apoptosis in CML cells. This formed the rationale for treatment regimens that maintain continuous kinase inhibition. Clinical studies with dasatinib suggested that daily dosing achieves equivalent response to twice daily even though ABL kinase inhibition only persists for 4–6 hours. We have demonstrated that 30 minutes of exposure to 100 nM dasatinib or 30 μ M imatinib (equipotent) inhibit p-Crkl (surrogate marker of Bcr-Abl kinase activity) by 80 to 90% in Bcr-Abl +ve cell lines and CML-CD34+ cells (n=8). We then sought to compare antiproliferative and pro-apoptotic effects of short term (ST; cells were cultured with dasatinib/imatinib for 30 minutes and after thorough wash, were recultured without dasatinib/imatinib for 72 hours) and continuous (CT, cells were cultured with drugs continuously) dasatinib or imatinib in BCRABL +ve cell lines (K562, Meg 01) and CD34+ cells of CML-CP patients. Although Bcr- Abl kinase reactivated within 30 minutes of drug removal, ST 100 nM dasatinib (D100ST) or 30μ M of imatinib (IM30ST) induced apoptosis (~80%) and blocked cell proliferation equivalent to continuous dasatinib (10 nM; D10CT) or imatinib (2μ M, IM2CT) in Bcr-Abl +ve cell lines. The kinetics of cell death and caspase-3 activation over 72 hours of culture were similar in D100ST and D10CT. In the presence of 6-growth factors (GFs; IL-3, IL- 6, G-CSF, SCF, TPO, Flt-3) D100ST and IM30ST reduced cell viability and CFU-GM colonies of CML-CD34+ cells by only 25 to 30% of no drug control. Moreover in the presence of GFs, 30 to 40% CD34+ve cells were viable and retained CFU-GM potential in spite of continuous dasatinib 100 nM (D100CT) or 30 μ M of imatinib (IM30CT). However, in the absence of GFs, D100ST and IM30ST reduced viability by 60 to 70%, and CFU-GM by 95% of control (with GFs, no TKI control; Fig 1). Figure 1: Survival of CFU-GM according to growth factor and dasatinib exposure: Figure 1:. Survival of CFU-GM according to growth factor and dasatinib exposure: Conclusion: Short term intense inhibition of BCR-ABL kinase activity triggers apoptosis in CML cell lines, which demonstrate their Bcr-Abl oncogene dependence. However, in spite of >80% kinase inhibition, D100ST and D100CT did not eliminate the majority of CML-CD34+ cells in the presence of GFs. In the absence of GFs, D100ST and IM30ST were able to inhibit cell proliferation, induce cell death and eliminate 95% of CFU-GM. This data suggests that oncogene dependence of CML CD34+ cells can be overcome by cytokines. Unlike CML cell lines where transient intense kinase inhibition leads to cell death, primary CML cells are only sensitive to this short term kinase inhibition in the absence of cytokines. Strategies that block cytokine pathways in combination with Bcr-Abl kinase inhibition may eliminate leukemic stem cells in-vivo even if only applied intermittently. CFU-GM colonies expressed as % of control. CML-CD34+ cells (n=3) were cultured with dasatinib in the presence (With GFs) or absence (No GFs) of 6-growth factors (GF) and CFU-GM colonies were plated on D3, using Methocult 4230 (Invitrogen) along with growth factors in all cases. Colonies were read after 14 days. In each patient values were normalised to cells cultured with GFs and no dasatinib. Short term (ST) and continuous (CT), Dasatinib 10 nM (D10), 100 nM (D100).
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Gutknecht, Michael, Simone Joas, Lisa Güttler, Lothar Kanz, Helmut R. Salih, Frank Grünebach, and Susanne M. Rittig. "Monocyte-Derived Dendritic Cells Induce Functionally Active Regulatory T Cells Upon Exposure to BCR-ABL Tyrosine Kinase Inhibitors." Blood 120, no. 21 (November 16, 2012): 2156. http://dx.doi.org/10.1182/blood.v120.21.2156.2156.
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Abstract Abstract 2156 Multiple approaches for treatment of malignant disease presently aim to combine targeted therapy with tyrosine kinase inhibitors (TKI) with immunotherapy. Dendritic cells (DC) are frequently used in such strategies due to their unique ability to initiate potent T cell anti-tumor immunity. Unfortunately, DC may also activate suppressive CD25+FOXP3+ regulatory T cells (Treg), which depends on the stimuli that influence DC in immature state and/or during development from precursor cells. High frequencies of Treg have been described in several types of tumors within the tumor microenvironment, which is associated with poor prognosis and reduced survival. DC development and function are moreover governed by various tyrosine kinases of which some are also inhibited by clinically used TKI. TKI thus may cause immunoinhibitory side effects, and we previously demonstrated that exposure of monocyte-derived DC to the BCR-ABL inhibitor imatinib causes up-regulation of the immunosuppressive type I transmembrane glycoprotein osteoactivin (GPNMB, DC-HIL) and reduces expression of activating surface antigens as well as T cell-stimulatory capacity of DC in vitro (Schwarzbich et al., 2012). Other investigators reported that imatinib induces functionally Treg in CML patients, but the underlying mechanisms are so far unknown. (Bachy et al., 2011). On the other hand, TKI may inhibit proliferation and suppressive capacity of regulatory T cells in vitro (Chen et al., 2007). Here we tried to solve this apparent discrepancy by analyzing the influence of TKI on DC-Treg interaction. Monocyte-derived DC (moDC) were generated over 7 days by exposing blood monocytes to GM-CSF and IL-4. TNF was added on day 6 of culture in case of maturation, and imatinib or nilotinib (3μM each) were added to the culture medium every second day starting from the first day of culture. Induction and functionality of Treg was determined by FACS and so called effector T cell suppression assays upon culture of moDC with autologous PBMC. We found that exposure of moDC to imatinib or nilotinib only slightly increased the frequency of Treg as compared to controls. However, these Treg strongly inhibited autologous T cell proliferation as assessed by T cell suppression assays. This was mediated by direct cellular interaction, as culture supernatants of TKI-treated DC did not alter Treg function and also did not contain elevated levels of the immunosuppressive (and Treg inducing) cytokines TGF-β and IL-10. Thus, our data indicate that the seemingly contradictory results of the in vivo and in vitro studies described above may be explained by the effects caused by exposure of moDC to BCR-ABL TKI which results in the induction of functionally active Treg. These findings are of special importance for future combinatory approaches using TKI and DC-based immunotherapy. Disclosures: No relevant conflicts of interest to declare.
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Kreutzman, Anna, Peter Rohon, Edgar Faber, Karel Indrak, Vesa Juvonen, Veli Kairisto, Jaroslava Voglova, et al. "Interferon Alpha Treated Patients with Chronic Myeloid Leukemia (CML) In Prolonged Complete Remission Have Increased Numbers of NK-Cells and Clonal Gamma-Delta T-Cells, and a Distinct Plasma Cytokine Profile." Blood 116, no. 21 (November 19, 2010): 1201. http://dx.doi.org/10.1182/blood.v116.21.1201.1201.
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Abstract Abstract 1201 Background. Before the era of tyrosine kinase inhibitors (TKIs), interferon alpha (IFN-α) was the treatment of choice in CML and prolonged survival of responding patients. Recent studies suggest that combination of IFN-α with TKI improves therapy outcome. Significantly, a proportion of IFN-α treated patients in prolonged complete cytogenetic remission (CCyR) have been able to discontinue treatment without disease relapse (Mahon et al. JCO 2002). The mechanism of action of IFN-α therapy is incompletely understood; the drug exerts both direct cytotoxic and immunomodulatory effects on leukemic cells. The aim of this project was to study the immunomodulatory effects of IFN-α in CML patients in prolonged remission and isolate biological markers predicting response. Patients and methods. The study population consisted of CML patients treated with IFN-α monotherapy (n = 10, median therapy time 146 months, range 63–231 months) and CML patients who had discontinued IFN-α therapy but remained in remission for >2 years (n = 9, median therapy time 120 months, range 80–184; median time without therapy 53 months, range 24–96). None of the patients were previously treated with TKI therapy. In addition, non-CML patients (3 patients with essential trombocythemia and one patient with polycythemia vera) treated with IFN-α and healthy volunteers (n = 43) were included as controls. Lymphocyte populations in all four groups were characterized with comprehensive immunophenotyping panels. The clonality of T-cells was analyzed by a TCR γ/δ rearrangement assay by PCR. Lymphocytes were further sorted into CD3+ TCR αβ+ and CD3+ TCR γδ+ populations (n = 8). Additionally, plasma levels of 25 cytokines were measured with a multiplex bead-based cytokine assay (Luminex®). Results. The proportion of NK-cells from lymphocytes was significantly increased in IFN-α discontinued patients (median 26%, range 18–51%) compared to healthy volunteers (11%, 5–21%) or patients on IFN-α therapy (12%, 6–31%)(P=0.0005). Similarly the proportion of CD8+ cells from T-cells was significantly increased in both CML IFN-α groups (55% in IFN-α discontinued patients, 44% in IFN-α treated patients vs. 31% in healthy volunteers; P<0.05 for both groups). Also a larger proportion of T-cells expressed the long-term memory antigen CD45RO in IFN-α patients (74%, 58% vs. 44% in healthy controls, P<0.01). The proportion of regulatory T-cells (CD4+CD25+FoxP3+) was increased in IFN-α groups (6.1%, 5.2% vs. 3.8% in healthy volunteers, P=0.01). Similar changes in immunoprofile were not observed in IFN-treated non-CML patients. Clonal TCR γ/δ rearrangements were observed in 18 of 19 (95%) IFN-treated CML patients as compared to 3 of 22 (14%) in healthy volunteers (P<0.01). In both IFN-α CML patient groups a unique rearrangement pattern was observed: 14/19 (79%) of patients had the Vγ9 gene clonally rearranged. This clonal rearrangement resided in CD3+ γδ+ T-cell population, as assessed by cell sorting. Two of four non-CML patients treated with IFN-α had the same clonal rearrangement, as well as one healthy control (1/22; 5%). Similar clonality patterns have not been observed in dasatinib or imatinib treated CML patients (Kreutzman et al. Blood 2010). IFN-α treatment was associated with a distinct plasma cytokine profile in CML patients. IP-10, IL-6, IL-12, eotaxin, MCP-1, and IFN-γ levels were significantly increased in IFN-α treated CML patients. In particular, eotaxin and MCP-1 levels differed significantly between healthy controls and IFN-α patients who had successfully discontinued IFN-α therapy (428 vs. 1173 pg/ml, P<0.0001 and 107 vs. 459 pg/ml, P=0.0003, respectively). In IFN-α treated non-CML patients, eotaxin or MCP-1 levels were not increased. Conclusions. Our results show that IFN-α treatment induces distinct changes in the immunoprofile of CML patients. Patients who had successfully discontinued IFN-α therapy differed markedly from healthy controls. IFN-α therapy was associated with increased numbers of NK-cells and clonal γδ+ T-cells. These cells possess potent anti-leukemic activity and may contribute to the prolonged therapy responses in this group of patients. Furthermore, plasma cytokine profile could be a helpful biomarker when considering which patients can discontinue the IFN-α treatment without imminent disease relapse. Disclosures: Faber: BMS, Novartis: Consultancy, Honoraria. Porkka:BMS, Novartis: Consultancy, Honoraria, Research Funding. Mustjoki:BMS, Novartis: Honoraria.
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Herrmann, Oliver, Caroline CA Weßling, Michael Huber, Shaoguang Li, Tim H. Brümmendorf, Steffen Koschmieder, and Mirle Schemionek. "Genetic Depletion of Fc Gamma Receptor 2b Affects CML Stem Cell Biology." Blood 124, no. 21 (December 6, 2014): 4528. http://dx.doi.org/10.1182/blood.v124.21.4528.4528.
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Abstract Chronic myeloid leukemia (CML) is caused by the acquisition of the Philadelphia chromosome t(9;22) at the hematopoietic stem cell (HSC) level. The resulting Bcr-Abl protein loses the Abl autoinhibitory function and gains the Bcr oligomerization domain resulting in the constitutive activation of the Abl tyrosine kinase. The implementation of Bcr-Abl tyrosine kinase inhibitors (TKI) has greatly improved therapy outcome of CML patients. However, various mechanisms such as the acquisition of tyrosine kinase domain mutations or kinase-independent alterations have been described to result in therapy resistance. Using an inducible transgenic SCLtTAxBcr-Abl CML mouse model, we previously demonstrated that Bcr-Abl inhibition leads to eradication of mature leukemic cells but spares disease-driving leukemic stem cells (LSC) due to a lack of oncogene-addiction (Schemionek et al., BLOOD 2010; Hamilton, Helgason, Schemionek et al., BLOOD 2012). Aiming to identify new Bcr-Abl targets that could overcome therapy resistance of stem cells, we previously performed a microarray analysis using murine LSC. Expression of FcγRIIb (Fc gamma receptor 2b; CD32) was significantly increased 2.79-fold in Bcr-Abl positive vs. control Lin-Sca1+c-kit+(LSK) cells. The FcγRIIb contains an immuno receptor tyrosine-based inhibitory motif (ITIM) and inhibits intracellular signaling upon activation. Phosphorylation of the receptor is mainly mediated via Src kinase Lyn and enables the receptor to recruit Src homology 2 domain-containing phosphatases. Using qRT-PCR, we confirmed 2.83-fold upregulation of FcγRIIb within the LSK population of CML mice. Retroviral infection of 32D cells and primary BM cells with Bcr-Abl significantly increased FcγRIIb expression level (90.3-fold and 13.2-fold, resp., p<0.005 each vs. emtpy vector). TKI treatment of Bcr-Abl positive 32D cells did not affect FcγRIIb mRNA levels or surface expression. Bcr-Abl infection of lineage-depleted BM cells decreased growth factor independent colony forming capability (5.5-fold, p<0.05) and the proliferation potential of FcγRIIb-/- lineage negative bone marrow (cells) compared to wild type (wt) in the absence of cytokines. Transplantation of Bcr-Abl infected FcγRIIb-/- BM cells significantly reduced white blood cell count (WBC) of the recipients as compared to wt Bcr-Abl positive cells by 2.4-fold (p<0.05). Interestingly, WBC of Bcr-Abl positive FcγRIIb-/- mice was similar to FcγRIIb-/- empty vector control animals (p=0.875) as well as wt control mice (p=0.241). CML development was analyzed upon autopsy by FACS of BM and spleen cells and showed a 2.0-fold reduction (p<0.05) of leukemic c-kit positive cells as well as a reduction in spleen weight in recipients of Bcr-Abl transduced FcγRIIb-/- cells (293mg ± 30) vs. Bcr-Abl transduced wt controls (381mg ± 13; p<0.05). Moreover, total numbers of leukemic LSK cells were significantly decreased by 3.5-fold in BM and 3-fold in spleen of Bcr-Abl transduced FcγRIIb-/- BM recipients compared to recipients of Bcr-Abl transduced wt cells. Transplantation of empty vector transduced FcγRIIb-/- vs. empty vector transduced wt control cells did not result in a decrease of knock-out cells showing that the observed effects were specific for impairment of Bcr-Abl transforming activity by loss of FcγRIIb but not due to FcγRIIb depletion alone. In conclusion, we show that FcγRIIb is upregulated in the stem cell compartment of CML mice and that this is due to Bcr-Abl expression. The absence of FcγRIIb impairs leukemic cell growth in vitro and in vivo. Moreover, genetic depletion of the receptor decreases LSC numbers and affects CML development in vivo suggesting that FcγRIIb plays an important role in Bcr-Abl leukemogenesis, making it an attractive novel stem cell specific target for CML therapy. (*Authors contributed equally) Disclosures Brümmendorf: Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties, Research Funding. Koschmieder*:Novartis: Consultancy, Honoraria, Other, Research Funding.