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1
Meyer, Stefanie, Sofija Vlasevska, Laura Garcia Ibanez, Claudio Scuoppo, Riccardo Dalla-Favera, and Laura Pasqualucci. "Targeting Histone Acetyltransferase Gene Inactivation in Diffuse Large B Cell Lymphoma." Blood 132, Supplement 1 (November 29, 2018): 671. http://dx.doi.org/10.1182/blood-2018-99-117542.
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Abstract Diffuse Large B-cell Lymphoma (DLBCL) is the most common form of non-Hodgkin lymphoma, accounting for ~30% of de-novo diagnoses and also arising as a frequent clinical evolution of indolent lymphomas. Although curable in a substantial fraction of cases, one third of patients do not achieve durable remissions, highlighting the need for novel, targeted therapies. Over the past decade, we and others have identified the CREBBP acetyltransferase and, less frequently, its paralogue EP300 as highly recurrent targets of inactivating somatic mutations/deletions in DLBCL and follicular lymphoma (FL) (30% and 60% of patients, respectively), indicating a prominent role in the pathogenesis of these tumors (Pasqualucci et al., Nature 2011). In most cases, mutations are heterozygous and the residual wildtype (WT) allele is expressed, suggesting a haploinsufficient tumor suppressor role. Indeed, germinal center (GC)-specific loss of Crebbp perturbs the expression of genes that are relevant to the normal biology of this structure, i.e. the lymphoma cell of origin, and cooperates with BCL2 deregulation to increase the incidence of tumors recapitulating the features of the human disease (Zhang et al., Cancer Discovery 2017). Intringuingly, while CREBBP binds to virtually all GC-specific superenhancers, no detrimental effects were observed upon its deletion in mice, suggesting the existence of compensatory mechanisms. Consistent with this hypothesis, inactivation of CREBBP and EP300 rarely coexist in human DLBCL and FL, suggesting that cells require a certain amount of acetyltransferase activity. To investigate whether EP300 compensates for CREBBP loss in the GC, we analyzed the GC responses in compound mouse models engineered to specifically delete these two genes (alone and in combination) upon SRBC immunization and induction of a Cγ1-driven Cre-recombinase. While CrebbpKOmice showed a mild increase in GC formation, as reported, loss of Ep300 led to ~40% reduction in the percentage of GC cells (mean: 1.8% vs 3.1% in WT littermates; p<0.05), documenting that these two enzymes play non-entirely overlapping roles in this population. Importantly, GC formation was completely abrogated in CrebbpKOEp300KO mice and dramatically impaired in CrebbpHETEp300KO mice, as compared to both WT and single EP300KO mice. These data suggest that GC B cells require a minimum amount of acetyltransferase activity, and reveal a potential therapeutically exploitable dependency of Crebbp-mutated GC B cells on Ep300. In order to probe if a similar dependency exists in neoplastic GC B cells, we used an inducible CRISPR/Cas9 system to delete EP300 (or a control non-genic region) in 4 DLBCL cell lines representative of the various CREBBP genotypes found in DLBCL, and monitored cell proliferation and survival in competition assays over 12 days. Compared to CREBBPWT, CREBBP heterozygous and homozygous mutant cells were significantly counter-selected from the total population following doxycycline induced EP300 deletion (~30% at day 7). Moreover, no EP300-edited clones were recovered from the CREBBP mutant lines in single cell plating assays, compared to CREBBP WT (p<0.01). Thus, DLBCL cells remain addicted to the residual EP300 aceyltransferase activity, supporting the existence of a therapeutic window for EP300 inhibitors. To explore this concept further, we generated isogenic DLBCL clones carrying WT or defective CREBBP alleles (n=4 each), and performed drug-sensitivity assays with 2 novel small molecule inhibitors that specifically target the CREBBP/EP300 HAT or BRD domain. While, at higher doses, both inhibitors interfered with cell growth in all clones, CREBBPKO cells were significantly more sensitive than their isogenic WT pairsat low nanomolar ranges (IC50: 60nM vs 300nM). Importantly, we were able to design an in vitro protocol that was toxic to CREBBPKO cells but tolerated by CREBBPWT cells, providing a proof of concept for therapeutically targeting these molecules. In conclusion, we show that CREBBP and EP300 have differential roles in normal GC B cell development and that CREBBP mutated cells are addicted to the residual EP300 activity. This dependency is maintained in DLBCL cells, providing the basis for the potential application of acetyl transferase inhibition into the clinical settings. Disclosures No relevant conflicts of interest to declare.
2
Ying, Hsia-Yuan, Yanwen Jiang, Ana Ortega-Molina, Huimin Geng, Dylan McNally, Ling Wang, Ashley Doane, et al. "Crebbp Mutations Disrupt Dynamic Enhancer Acetylation in B-Cells, Enabling HDAC3 to Drive Lymphomagenesis." Blood 128, no. 22 (December 2, 2016): 735. http://dx.doi.org/10.1182/blood.v128.22.735.735.
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Abstract Somatic mutations in CREBBP occur frequently in germinal center derived lymphomas such as DLBCL and FL. However whether or how these mutations might contribute to lymphomagenesis is still largely unknown. Most CREBBP mutations are predicted to result in loss of function since they target the histone acetyltransferase (HAT) domain or give rise to premature stop codon prior to the HAT domain. Here, we show that Crebbp shRNA knockdown (KD) accelerated lymphomagenesis in VavP-Bcl2 transgenic mice, a model that recapitulates human GC-derived lymphomas. The median time to lymphoma onset in VavP-Bcl2/CrebbpKD mice was 114 days, significantly shorter than control VavP-Bcl2/GFP mice (193 days, p=0.04). Histopathology revealed that VavP-Bcl2/CrebbpKD lymphomas were more aggressive and widely disseminated than VavP-Bcl2/GFP lymphomas. CREBBP can regulate gene enhancer function through H3K27 acetylation. ChIP-seq in VavP-Bcl2/CrebbpKD lymphoma cells revealed significant reduction of H3K27ac peaks compared to control lymphoma cells (N = 1717, Kolmogorov-Smirnov test, p<2.2E-16). Loss of H3K27ac was markedly skewed towards enhancers. We observed similar loss of enhancer H3K27ac in human DLBCL cells after CREBBP shRNA KD. Enhancer H3K27ac loss was significantly associated with repression of nearby genes in both murine (FDR q=0.044) and human lymphoma cells (FDR q=0). RNA-seq performed in three independent FL or DLBCL patient cohorts revealed a characteristic CREBBP mutant gene expression signature featuring prominent transcriptional repression (p=1.32E-14, p=0.001, and p=0.0002 respectively). Notably, the human patient CREBBP mutant signature was highly enriched in murine and human cell line CREBBP shRNA profiles (FDR=0, GSEA), indicating that CREBBP KD signature was highly similar to CREBBP mutant signature in humans. Functional analysis of the CREBBP mutant/KD signature showed significant enrichment of GC exit pathways including genes induced by CD40, IRF4 and plasma cell differentiation; as well as immune response processes including antigen processing and presentation, such as MHC class II genes (BH-adjusted p<0.05). To better understand mechanism we performed an integrative analysis of CREBBP signatures against databases of B-cell transcription factor and epigenome profiles. This analysis yielded significant enrichment (BH-adjusted p<0.05) for i) enhancers bound by the BCL6 transcriptional repressor and its SMRT/HDAC3 corepressor complex, ii) enhancers that are normally deacetylated in GC B-cells, and iii) genes induced by BCL6 siRNA. This is notable because in normal GCs BCL6 represses enhancers by recruiting SMRT/HDAC3 complexes to deacetylate H3K27. Hence our data suggest that CREBBP is a counteracting HAT to BCL6/SMRT/HDAC3. Indeed, conditional knockout of Hdac3 in GC B-cells in mice resulted in impaired GC formation and a transcriptional signature featuring upregulation of the same genes that are repressed by CREBBP KD (GSEA FDR=0). Moreover, CREBBP KD in DLBCL cells resulted in H3K27ac loss at BCL6/SMRT/HDAC3 regulated enhancers, including those nearby CDKN1A, NFATC1, FOXP1, and MHC II genes, such as HLA-DQA1 and HLA-DRB5. CREBBP KD also resulted in silencing of these genes. Since we show HDAC3 is the opposing HDAC to CREBBP then we reasoned that CREBBP mutant DLBCLs might be especially dependent on HDAC3. Indeed we observed that HDAC3 shRNA resulted in profound suppression of CREBBP mutant DLBCL cells in vitro and in vivo (DLBCL xenografts in mice, p=0.005), whereas CREBBP WT cell lines were barely affected by HDAC3 KD. The opposing effects of BCL6/SMRT/HDAC3 and CREBBP on MHC class II could have implications for immune surveillance. Accordingly CREBBP KD induced significant loss of cell surface HLA-DR molecules (p<0.05), and these cells exhibited up to 90% less capability to stimulate T-cell response in allogeneic mixed lymphocyte reaction experiments. The loss of MHC class II molecules and T-cell response was rescued when CREBBP loss of function cells were exposed to a specific HDAC3 inhibitor. In summary, CREBBP mutations drive lymphomagenesis by enabling unopposed suppression of enhancers by BCL6/SMRT/HDAC3 complexes, resulting in a repressive transcriptional programming that disrupts GC exit and evades immune surveillance. HDAC3 targeted therapy may rescue these effects and serve as a precision approach for CREBBP mutant lymphomas. Disclosures Scott: Celgene: Consultancy; Roche: Honoraria; Janssen: Consultancy; BC Cancer Agency: Patents & Royalties: Inventor on a patent licensed to NanoString Technologies. Tam:Millennium Pharmaceuticals, Inc.: Consultancy. Melnick:Janssen: Research Funding.
3
Lamble, Adam J., Robert B. Gerbing, Jenny L. Smith, Rhonda E. Ries, Edward A. Kolb, Todd A. Alonzo, and Soheil Meshinchi. "Crebbp Alterations Are Associated with a Poor Prognosis in De Novo AML." Blood 138, Supplement 1 (November 5, 2021): 3451. http://dx.doi.org/10.1182/blood-2021-154052.
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Abstract Introduction: The translocation, t(8;16)(p11;p13), results in the fusion between KAT6A and CREBBP and has been associated with a poor prognosis in both pediatric and adult acute myeloid leukemia (AML). This lesion has therefore been re-classified as high risk on the active Phase 3 Children's Oncology Group (COG) trial for de novo AML, AAML1831 (NCT04293562). Less is known about the prognostic significance of CREBBP sequence variants. Methods: CREBBP variant status was determined in patients with AML enrolled on 4 successive COG trials for de novo pediatric AML (NCT00003790, NCT00070174, NCT01407757, NCT01371981). Fusions involving CREBBP were prospectively obtained via conventional cytogenetics and retrospectively confirmed via RNAseq. Insertions and deletions (indels) leading to frameshift mutations and single nucleotide variants (SNVs) were retrospectively interrogated via next generation sequencing. Results: Of 2216 patients (age: 0-29.8 years), 55 (2.5%) patients had an alteration involving CREBBP. Sixteen (29%) of these were a fusion involving CREBBP (CREBBP/fus), with KAT6A being the most common translocation partner (n=15) and the remaining translocation involving ANK1. The remaining 39 patients (71%) had a CREBBP mutation (CREBBP/mut), including 19 with an indel (CREBBP/indel) leading to a frameshift mutation and 20 with a SNV (CREBBP/SNV). We compared clinical and biologic characteristics between the three cohorts. CREBBP/fus patients were significantly younger than CREBBP/indel and CREBBP/SNV patients (median ages of 2.6 vs. 7.8 vs. 11.9 years; p=0.027). There was a higher prevalence of t(8;21)/RUNX1-RUNX1T1 in CREBBP/indel patients compared to CREBBP/SNV patients (42.1% vs. 5%; p=0.008). In contrast, CREBBP/SNV patients were more likely to be associated with a normal karyotype (40% vs. 5.3%; p=0.02). There was a similar prevalence of co-occurring high-risk lesions in CREBBP/indel (n=5; CBFA2T3-GLIS2, KMT2A-AFF1, KMT2A-MLLT4, MLLT10-PICALM, NUP98-HOXA9) and CREBBP/SNV (n=7; DEK-NUP214, ETV6-FOXO1, FUS-ERG, NUP98-NSD1, ETV6-MNX1, FLT3-ITDx2) patients. There was otherwise no difference between presenting WBC count, FLT3-ITD, NPM1, CEBPA, remission rates or MRD status after Induction 1 therapy. Patients with any CREBBP alteration had a significantly worse 5-year event free survival (EFS) compared to patients without (25.9% vs. 45.2%; p=0.002) and this inferior EFS overlaps with contemporarily defined high-risk patients (Figure 1a). Evaluation of outcomes based on type of alteration demonstrated a similar 5-year EFS of 33.3% and 23.1% between CREBBP/fus and CREBBP/mut patients, respectively (Figure 1b; p=0.832). This poor EFS was maintained in the CREBBP/indel patients with a co-occurring t(8;21) (n=8, 5-year EFS 12.5%). When patients with co-occurring high-risk lesions were excluded from analysis, the remaining CREBBP/mut (n=27) patients maintained their poor EFS (29.6%). Despite their poor EFS, CREBBP/mut patients had an analogous overall survival (OS) to non-CREBBP patients (57.4% vs. 62.3%; p=0.499, Figure 1c), demonstrating that these patients could be successfully salvaged following relapse. In contrast, all patients with CREBBP/fus that relapsed subsequently died from their disease (OS 33.3%). Conclusions: In a large study of CREBBP alterations in pediatric patients with de novo AML, we show that these patients have a dismal EFS, regardless of alteration type. Further, despite enrichment of t(8;21), the favorable prognosis typically conferred by this alteration was abrogated by the co-occurrence of CREBBP/indel. Similarly, by excluding patients with co-occurring high-risk lesions from analysis, we show that these poor outcomes persist in a cohort of patients that would otherwise be considered low risk. Translocations between CREBBP and KAT6A in patients over 90 days of age are considered high risk on the active COG phase 3 trial. Given the inferior EFS and high salvage rates associated with other CREBBP alterations, intensification of upfront treatment, including hematopoietic stem cell transplant, should be considered in this population. The authors would like to acknowledge Astellas Pharma Global Development, Inc. Figure 1 Figure 1. Disclosures No relevant conflicts of interest to declare.
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Zhu, Yu, Zi Wang, Yanan Li, Hongling Peng, Jing Liu, Ji Zhang, and Xiaojuan Xiao. "The Role of CREBBP/EP300 and Its Therapeutic Implications in Hematological Malignancies." Cancers 15, no. 4 (February 14, 2023): 1219. http://dx.doi.org/10.3390/cancers15041219.
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Disordered histone acetylation has emerged as a key mechanism in promoting hematological malignancies. CREB-binding protein (CREBBP) and E1A-binding protein P300 (EP300) are two key acetyltransferases and transcriptional cofactors that regulate gene expression by regulating the acetylation levels of histone proteins and non-histone proteins. CREBBP/EP300 dysregulation and CREBBP/EP300-containing complexes are critical for the initiation, progression, and chemoresistance of hematological malignancies. CREBBP/EP300 also participate in tumor immune responses by regulating the differentiation and function of multiple immune cells. Currently, CREBBP/EP300 are attractive targets for drug development and are increasingly used as favorable tools in preclinical studies of hematological malignancies. In this review, we summarize the role of CREBBP/EP300 in normal hematopoiesis and highlight the pathogenic mechanisms of CREBBP/EP300 in hematological malignancies. Moreover, the research basis and potential future therapeutic implications of related inhibitors were also discussed from several aspects. This review represents an in-depth insight into the physiological and pathological significance of CREBBP/EP300 in hematology.
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Hashwah, Hind, Corina A. Schmid, Sabrina Kasser, Katrin Bertram, Anna Stelling, Markus G. Manz, and Anne Müller. "Inactivation of CREBBP expands the germinal center B cell compartment, down-regulates MHCII expression and promotes DLBCL growth." Proceedings of the National Academy of Sciences 114, no. 36 (August 22, 2017): 9701–6. http://dx.doi.org/10.1073/pnas.1619555114.
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The genes encoding the histone acetyl-transferases (HATs) CREB binding protein (CREBBP) and EP300 are recurrently mutated in the activated B cell-like and germinal center (GC) B cell-like subtypes of diffuse large B cell lymphoma (DLBCL). Here, we introduced a patient mutation into a human DLBCL cell line using CRISPR and deleted Crebbp and Ep300 in the GC B cell compartment of mice. CREBBP-mutant DLBCL clones exhibited reduced histone H3 acetylation, expressed significantly less MHCII, and grew faster than wild-type clones in s.c. and orthotopic xenograft models. Mice lacking Crebbp in GC B cells exhibited hyperproliferation of their GC compartment upon immunization, had reduced MHCII surface expression on GC cells, and developed accelerated MYC-driven lymphomas. Ep300 inactivation reproduced some, but not all, consequences of Crebbp inactivation. MHCII deficiency phenocopied the effects of CREBBP loss in spontaneous and serial transplantation models of MYC-driven lymphomagenesis, supporting the idea that the mutational inactivation of CREBBP promotes immune evasion. Indeed, the depletion of CD4+ T cells greatly facilitated the engraftment of lymphoma cells in serial transplantation models. In summary, we provide evidence that both HATs are bona fide tumor suppressors that control MHCII expression and promote tumor immune control; mutational inactivation of CREBBP, but not of EP300, has additional cell-intrinsic engraftment and growth-promoting effects.
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Zimmer, Stephanie N., Qing Zhou, Ting Zhou, Ziming Cheng, Sherry L. Abboud-Werner, Diane Horn, Mike Lecocke, et al. "Crebbp haploinsufficiency in mice alters the bone marrow microenvironment, leading to loss of stem cells and excessive myelopoiesis." Blood 118, no. 1 (July 7, 2011): 69–79. http://dx.doi.org/10.1182/blood-2010-09-307942.
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Abstract CREB-binding protein (CREBBP) is important for the cell-autonomous regulation of hematopoiesis, including the stem cell compartment. In the present study, we show that CREBBP plays an equally pivotal role in microenvironment-mediated regulation of hematopoiesis. We found that the BM microenvironment of Crebbp+/− mice was unable to properly maintain the immature stem cell and progenitor cell pools. Instead, it stimulates myeloid differentiation, which progresses into a myeloproliferation phenotype. Alterations in the BM microenvironment resulting from haploinsufficiency of Crebbp included a marked decrease in trabecular bone that was predominantly caused by increased osteoclastogenesis. Although CFU-fibroblast (CFU-F) and total osteoblast numbers were decreased, the bone formation rate was similar to that found in wild-type mice. At the molecular level, we found that the known hematopoietic modulators matrix metallopeptidase-9 (MMP9) and kit ligand (KITL) were decreased with heterozygous levels of Crebbp. Lastly, potentially important regulatory proteins, endothelial cell adhesion molecule 1 (ESAM1) and cadherin 5 (CDH5), were increased on Crebbp+/− endothelial cells. Our findings reveal that a full dose of Crebbp is essential in the BM microenvironment to maintain proper hematopoiesis and to prevent excessive myeloproliferation.
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Huntly, Brian J. P., Sarah Jayne Horton, George Giotopoulos, Haiyang Yun, Shabana Vohra, Olivia Sheppard, Rachael Bashford-Rogers, et al. "Early Loss of CREBBP Confers Malignant Stem Cell Properties on Lymphoid Progenitors." Blood 128, no. 22 (December 2, 2016): 460. http://dx.doi.org/10.1182/blood.v128.22.460.460.
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Abstract Loss-of-function mutations of the cyclic-AMP response element binding protein, binding protein (CREBBP) gene have recently been described at high frequencies across a spectrum of lymphoid malignancies, particularly follicular lymphoma (FL) and diffuse large B-cell lymphoma (DLBCL). The multiple effects of this epigenetic regulator on developmental and homeostatic processes have been extensively studied, however, exactly how CREBBP functions as a tumor suppressor and the reasons for its particular predilection for suppression of lymphoid tumors remains unclear. In addition, for many mature lymphoid malignancies, the existence of cancer stem cells is unproven and their provenance and the initial target cell for transformation a source of ongoing debate. Here we use multiple mouse strains to model loss of Crebbp in different lymphoid compartments to address these questions. We demonstrate that early loss of Crebbp in hematopoietic stem and progenitor cells (HSPC), through disruption by the Mx1-Cre recombinase, leads to increased development of hematological malignancies, particularly of the B-lymphoid lineage that mimic features of human lymphomas. Theses B-cell malignancies are of long latency and are preceded by significant alterations in the proliferation, self-renewal and differentiation of lymphoid progenitors, allowing hyperproliferative lymphoid progenitors whose differentiation is blocked to accumulate. Using an aberrant surface phenotype that resembles the eventual tumor, we identify a pre-malignant population in the peripheral blood of animals that is often evident many months before any disease characteristics. We demonstrate pre-malignant stem cell characteristics for this population in functional experiments, where it generates high level reconstitution of peripheral blood in transplant recipients, but only gives rise to disease in these animals after a long latency. We also utilize this unique cellular population in longitudinal genome scale analyses (clonality, RNA-Seq, ChIP-Seq and exome sequencing) to document the mechanisms of malignant evolution. Linking the increased rate of mutation we describe to Crebbp loss, we also demonstrate increased DNA damage and an altered DNA-damage response in premalignant lymphoid progenitors. Importantly, using a Cd19-Cre recombinase that excises only within committed lymphoid cells, we are able to demonstrate that when Crebbp is lost at a later stage of lymphoid development, the marked cellular abnormalities described above are completely lost and the development of tumors is no different from normal (Figure, below). Taken together, these findings define the developmental stage-specific tumor suppressor functions of Crebbp and shed light on the cellular origins and subsequent evolution of lymphoid malignancies. In addition, the altered response to DNA damage that we demonstrate upon loss of Crebbp, allied to the increased exposure to physiological DNA-damage during lymphoid ontogeny offers an explanation for the high incidence of CREBBP mutations in mature lymphoid malignancies. Figure Left panel, Kaplan Meier graph for Mx1-Cre Crebbp mice with loss of Crebbp in the HSPC compartment demonstrates significantly shorter survival vs WT littermates with intact expression of Crebbp. In contrast, when Crebbp is excised in a later lymphoid compartment through Cd19-Cre mediated recombination, right panel, no difference in survival is noted from WT littermate controls. Figure. Left panel, Kaplan Meier graph for Mx1-Cre Crebbp mice with loss of Crebbp in the HSPC compartment demonstrates significantly shorter survival vs WT littermates with intact expression of Crebbp. In contrast, when Crebbp is excised in a later lymphoid compartment through Cd19-Cre mediated recombination, right panel, no difference in survival is noted from WT littermate controls. Disclosures Huntly: Novartis: Speakers Bureau; BMS: Speakers Bureau; Ariad: Speakers Bureau; Pfizer: Speakers Bureau.
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Idoia, García-Ramírez, Shashank Shrishrimal, Ines Gonzalez-Herrero, Alberto Martín-Lorenzo, Guillermo rodriguez-Hernandez, Romain Duval, Dalia Moore, et al. "CREBBP Loss Cooperates with BCL2 Over-Expression to Promote Lymphoma in Mice." Blood 128, no. 22 (December 2, 2016): 458. http://dx.doi.org/10.1182/blood.v128.22.458.458.
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Abstract Follicular lymphoma (FL) is genetically characterized by translocations of the BCL2 oncogene that are found in ~90% of patients, and mutations of chromatin modifying genes that are found in up to 96% of patients. The latter include inactivating mutations of KMT2D and CREBBP, and activating mutations of EZH2, among others. However, CREBBP has yet to be investigated using this approach. We recently defined the evolutionary hierarchy of somatic mutations in FL and found that CREBBP mutations were most frequently acquired as early events during disease evolution and were maintained throughout disease progression and transformation. Recent studies, using transgenic mouse models, have shown that inactivation of KMT2D and introduction of the activating EZH2 mutation results in perturbed B-cell development and lymphomagenesis. Here, we extended upon these observations by performing targeted next generation sequencing of an additional cohort of tumors allowing the identification of the spectrum of CREBBP mutations across 200 FLs. This identified CREBBP mutations in 55% of tumors, and found that 31% of these mutations reside within the lysine acetyltransferase domain. Furthermore, 30% of mutations altered a single amino acid, arginine 1408, to either a cysteine or histidine residue. We performed a sensitive in vitro acetyltransferase assay for these point mutants and show that they result in >90% loss of catalytic activity. As our results show that CREBBP mutations result in a loss of function, we modeled these events in mice by floxing one or both alleles of Crebbp and crossing with the Mb1-cre strain. This yielded mice that deleted Crebbp specifically in B-cells. We additionally crossed these mice with the EµBcl2 strain that over-expresses Bcl2 in B-cells. Inactivation of Crebbp in B-cells was associated with deficits in B-cell development, with significantly reduced numbers of total B-cells that were contributed to by reductions in multiple B-cell subsets. These deficits were partially rescued by the EµBcl2 transgene. After 14-21 months, some mice became ill and necropsy revealed lymphadenopathy and splenomegaly as a result of B-cell lymphoma. We noted increased penetrance and decreased latency of lymphoma with one vs two alleles of Crebbp deleted, and with absence vs presence of the EµBcl2 transgene (Figure 1). We investigated the molecular etiology of these tumors by isolating splenic B-cells from these mice and performing transcriptome profiling and epigenetic profiling for the histone H3 lysine 18 acetylation (H3K18Ac) mark that is catalyzed by Crebbp. Transcriptional profiling identified a signature of 335 genes with increased expression and 370 genes with decreased expression, including an incremental increase in Myc expression when one or both alleles of Crebbp were deleted, respectively. Surprisingly, changes in transcript abundance were not associated with changes in H3K18Ac in the proximal regulatory regions of those genes. Regions of significantly altered H3K18Ac were instead localized primarily to intragenic regions. Analysis of the DNA sequences in these regions identified a significant enrichment of motifs that contained Myc consensus sequences, and these were present in >60% of regions with altered H3K18Ac. In addition, ChIP-seq data from the ENCODE database showed a strong level of Myc binding to the center of these regions with altered H3K18Ac. Together, our results demonstrate that inactivating mutations of Crebbp may have a role in altering B-cell development. The significant induction of Myc expression that was associated with Crebbp deletion, and epigenetic changes in regions that are bound by Myc, suggest that Crebbp inactivation may have a role in the induction of Myc expression and activity. This may be important with respect to transformation of FL, which may proceed via induction of MYC. However, our results also demonstrate some important discrepancies between the role of CREBBP mutations in human FL, and the role of Crebbp deletion in murine models. Disclosures Lunning: Celgene: Consultancy; Spectrum: Consultancy; TG Therapeutics: Consultancy; Gilead: Consultancy; Genentech: Consultancy; Juno: Consultancy; Bristol-Myer-Squibb: Consultancy; AbbVie: Consultancy; Pharmacyclics: Consultancy.
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Dixon, Zach, Julie A. E. Irving, and Lindsay Nicholson. "Crebbp K nockdown Does Not Impact on Glucocorticoid Induced Apoptosis in Childhood Acute Lymphoblastic Leukemia." Blood 126, no. 23 (December 3, 2015): 1429. http://dx.doi.org/10.1182/blood.v126.23.1429.1429.
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Abstract Childhood acute lymphoblastic leukemia (ALL) is the most common childhood cancer and, despite a cure rate approaching 90%, relapse is a significant cause of death in young people. Recently it has been shown that inactivating mutations in the histone acetyltransferase, CREB binding protein (CREBBP or CBP) are frequently seen at relapse in childhood ALL, with enrichment in high hyperdiploid and hypodiploid cases. Mutations are usually heterozygous, suggesting haploinsufficiency, and are often acquired at relapse, implying a role in drug resistance. Since glucocorticoid (GC) response genes are known targets of CREBBP and, given the pivotal role of GCs in ALL therapy, it has been postulated that CREBBP mutations confer GC chemoresistance. CREBBP is a multifunctional protein, playing a role in cAMP dependent signalling, acetylation mediated activation of p53 and inactivation of BCL6 and a range of DNA damage repair pathways including base excision repair (BER) and direct DNA damage repair. To assess the role of CREBBP haploinsufficiency in ALL, RNAi techniques were used to create isogenic CREBBP knockdown models of ALL. CREBBP knockdown was carried out using small hairpin RNA (shRNA) transduction (termed shCBP cells) or small interfering RNA (siRNA) transfection (termed siCBP cells) in the PreB 697 B-cell precursor cell line (t(1;19)) and the hypodiploid MHH-CALL-2 cell line, as well as high hyperdiploid primagraft ALL cells. Knockdown of at least 50% of control was confirmed at both mRNA and protein level. The functional impact of CREBBP knockdown in cells was determined by analysis of known CREBBP target residues; acetyl H3K18 and H3K27, and transcription of cAMP dependent genes (CXCR4, MKNK2, DUSP5, DUSP10 and RGS16). To assess the impact of CREBBP knockdown on response to GCs, cells were treated with dexamethasone and expression of the classic glucocorticoid receptor (GR) targets; GILZ and FKBP51, was assessed by quantitative reverse transcriptase PCR (QRT-PCR). Alamar blue cell viability assays were used to determine the sensitivity of each CREBBPknockdown model to dexamethasone compared to isogenic controls. Three out of four cell models displayed a reduction in H3K18 or H3K27 acetylation compared to isogenic control, indicating a relevant functional impact of CREBBP knockdown. Cell lines showed a trend towards reduced induction of some of the selected cAMP dependent targets but statistical significance was not achieved (p values >0.2). Gene expression profiling and Ingenuity Pathway Analysis of PreB 697 shCBP cells compared to isogenic control predicted that upstream transcription of NR3C1, the gene encoding the GR, would be affected in CREBBP knockdown cells. However, while induction of GILZ and FKBP51 in PreB 697 shCBP cells in response to GC was significantly impaired in knockdown compared to control cells (GILZ p=0.009, FKBP51 p=0.03), they were no more resistant to dexamethasone (p=0.9). This was mirrored in siCBP cell lines and primagraft cells, where a significant impairment in basal expression of GILZ and/or FKBP51 was seen in some lines (GILZ reduction; p=0.03 PreB 697 shCBP, p=0.02 PreB 697 siCBP, FKBP51 reduction; p=0.01 primagraft siCBP cells) but no significant impairment in the transcriptional induction of these genes in response to GC compared to isogenic control was observed (p values >0.5). Importantly, no decreased sensitivity to dexamethasone was seen in any model after CREBBP knockdown (p values >0.1). CREBBP knockdown in ALL cells had no significant effect on the induction of cAMP dependent genes, had a variable effect on GR target expression, but consistently showed no impact on GC sensitivity, regardless of cytogenetic context. These data show that the acquisition of CREBBP mutations at relapse in childhood ALL is not mediated through GC resistance and suggest that other CREBBP associated mechanisms, such as DNA damage repair, may influence drug response. Understanding the role of CREBBP in carcinogenesis and drug resistance is crucial as it is implicated as a tumour suppressor in a growing number of cancers, making it a potential multi-tumour target for novel therapies. Disclosures No relevant conflicts of interest to declare.
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Xu, Abai, and Tingting Chen. "Abstract 5102: Correlation analysis of CREBBP mutation with tumor mutation burden and effect of immune checkpoint therapy in bladder cancer." Cancer Research 82, no. 12_Supplement (June 15, 2022): 5102. http://dx.doi.org/10.1158/1538-7445.am2022-5102.
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Abstract Background: CREB-binding protein (CREBBP, hereafter CBP) encoded the protein having intrinsic histone acetyltransferase activity, which stabilized the additional protein interactions with the transcription complex. The mutation of CREBBP has been reported that caused Rubinstein-Taybi syndrome (RTS) and associated with acute myeloid leukemia, but not been studied in bladder cancer (BLCA). Immune checkpoint therapy (ICI) has significantly became one of the primary treatment of BLCA, and Tumor mutation burden (TMB) has been regarded as the most prevalent biomarker to predict immunotherapy. The CREBBP mutation status correlation with TMB and immune response remain unknown. Methods: Whole exome sequencing (WES) data and clinical data of 406 BLCA patients was obtained from the Cancer Genome Altas (TCGA). The mutation data of 206 FFPE tumor samples from Chinese BLCA patients were sequenced by targeted next-generation sequencing (NGS, 3DMed panel). NGS data and clinical data of 192 advanced BLCA patients treated with ICI were obtained from the MSK-IMPACT Clinical Sequencing cohort (MSKCC). The association between CREBBP mutation and TMB level in BLCA were explored. Survival analysis was determined by Kaplan-Meier (KM) analysis. Results: In total, The CREBBP mutation frequency was 16.25% (66/406) of BLCA patients in TCGA, 14.56% (30/206) in Chinese cohort and 13.54 (26/192). In Chinese cohort, the alteration frequency of NOTCH signal pathways affected was 48.54% (100/206), in which the highest frequency genes was CREBBP. The TMB level in CREBBP mutant group was higher than wild-type group both in Chinese group (Wilcoxon test, p = 0.001) and MSKCC cohort (Wilcoxon test, p < 0.001). The survival analysis were performed on patients from TCGA cohorts without treatment information and MSKCC BLCA patients treated with ICIs, separately. In TCGA cohort, there are not significantly difference between CREBBP mutation group (n=65) and wild-type group (n=340). While in MSKCC ICIs treatment cohort, the overall survival (OS) of CREBBP mutation group (n=26) were significantly longer than wild-type group (n=166) (median OS, mutation vs wild-type = NE vs 15 months; HR 0.468 [95% CI 0.26-0.84]; P = 0.0431). Conclusion: The CREBBP gene mutation was associated with higher TMB level. Clinical cohort analysis results suggested that CREBBP might be a predictive biomarker of immune checkpoint therapy but not a prognostic factor in BLCA. Citation Format: Abai Xu, Tingting Chen. Correlation analysis of CREBBP mutation with tumor mutation burden and effect of immune checkpoint therapy in bladder cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5102.
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Feng, Dapeng, Zhengwei Li, Liang Yang, Haidong Liang, Hongtao He, Lin Liu, and Wei Zhang. "BMSC-EV-derived lncRNA NORAD Facilitates Migration, Invasion, and Angiogenesis in Osteosarcoma Cells by Regulating CREBBP via Delivery of miR-877-3p." Oxidative Medicine and Cellular Longevity 2022 (March 1, 2022): 1–19. http://dx.doi.org/10.1155/2022/8825784.
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Bone marrow mesenchymal stem cells (BMSCs) can boost osteosarcoma (OS) cell proliferation and invasion, yet the function of extracellular vesicles (EVs) derived from BMSCs on OS is scarcely known. This study is aimed at examining the role of BMSC-EVs in OS cells. BMSCs and BMSC-EVs were isolated and identified. The effect of EVs and EVs-si-NORAD on OS cell proliferation, invasion, migration, and angiogenesis was determined. Expressions of NORAD, miR-877-3p, and CREBBP were detected. The binding relationship among NORAD, miR-877-3p, and CREBBP was verified. The miR-877-3p inhibitor or pc-CREBBP was delivered into OS cells treated with EVs-si-NORAD for in vitro analysis. The nude mouse model of the subcutaneous tumor xenograft was established for in vivo analysis. BMSC-EVs promoted OS cell proliferation, invasion, migration, and angiogenesis. BMSC-EVs carried NORAD into OS cells and upregulated CREBBP by sponging miR-877-3p. miR-877-3p downregulation or CREBBP overexpression partly inverted the inhibitory effect of EVs by silencing NORAD on OS cell proliferation, invasion, migration, and angiogenesis. In vivo experiments validated that BMSC-EV-derived NORAD facilitated tumor growth by upregulating CREBBP via miR-877-3p. To conclude, BMSC-EV-derived NORAD facilitated OS cell proliferation, invasion, migration, and angiogenesis by modulating CREBBP via miR-877-3p, which may offer new insights into OS treatment.
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García-Ramírez, Idoia, Saber Tadros, Inés González-Herrero, Alberto Martín-Lorenzo, Guillermo Rodríguez-Hernández, Dalia Moore, Lucía Ruiz-Roca, et al. "Crebbp loss cooperates with Bcl2 overexpression to promote lymphoma in mice." Blood 129, no. 19 (May 11, 2017): 2645–56. http://dx.doi.org/10.1182/blood-2016-08-733469.
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Key Points Crebbp inactivation perturbs B-cell development, but cooperates with Bcl2 overexpression to promote lymphoma. Transcriptional and epigenetic signatures of Crebbp loss implicate Myc in disease etiology.
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Tokunaga, Kenji, Shunichiro Yamaguchi, Eisaku Iwanaga, Tomoko Nanri, Taizo Shimomura, Hitoshi Suzushima, Mitsuya Hiroaki, and Norio Asou. "Crebbp HAT Domain Mutations Are Frequently Detected in Adult Acute Lymphoblastic Leukemia." Blood 120, no. 21 (November 16, 2012): 1419. http://dx.doi.org/10.1182/blood.v120.21.1419.1419.
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Abstract Abstract 1419 Aims: Molecular pathogenesis of acute lymphoblastic leukemia (ALL) has largely been verified in pediatric patients and the identification of genetic alterations have contributed to stratifying therapeutic applications. In adult patients with ALL, cytogenetic and genetic abnormalities have not sufficiently been elucidated and therapeutic improvement has been hindered. CREB binding protein (CREBBP) is a transcriptional coactivator that interacts with a diverse range of transcription factors and regulates transcription by histone acetylation in hematopoiesis. Mutations of the CREBBP gene are recently found in approximately 2–4% of pediatric patients with ALL. Especially in relapsed cases, the mutations prevail (18–63%) and are possible markers for prediction of relapse in pediatric ALL. In adult patients with ALL, the clinical significance of CREBBP mutations remains to be determined. Here we examined adult ALL patients in an attempt to determine the incidence, clinical characteristics and prognostic impact of the CREBBP mutations. Methods: We investigated 71 adult patients with newly diagnosed ALL treated with JALSG protocols between 1986 and 2010. Age ranged from 15 to 86 years, with a median of 54 years. CREBBP mutations are dominantly identified in histone acetyltransferase (HAT) domain. HAT domain in the CREBBP gene was amplified with RT-PCR using RNA isolated from the peripheral blood or bone marrow mononuclear cells at diagnosis and was subjected to direct sequencing. We compared clinical profiles between patients with and without CREBBPHAT domain mutations. This study was approved by the Institutional Review Boards and informed consent was obtained from each patient according to guidelines based on the revised Declaration of Helsinki. Results: CREBBP HAT domain mutations were detected in 8 of 71 (11.3%) patients: one nonsense mutation, five insertion mutations with frameshifts, and five missense mutations. Two patients harbored biallelic mutations. The mutations at diagnosis in adult patients were seen more frequently than those in pediatric patients ever reported. Such mutations were not completely identical to those detected in pediatric ALL, but were seen in the region within the HAT domain, indicating that such mutations are loss-of-function mutations. The mutations were found in both B-cell (6/53: 11.3%) and T-cell (1/9: 11.1%) ALL, and distributed in patients harboring IKZF1 alterations (3/31: 9.7%) or the BCR-ABL fusion gene (2/19: 10.5%). There were no statistical difference in age, sex, leukocyte, platelet counts and complete remission rate between patients with and without the CREBBP HAT domain mutations. Patients with the mutations had a trend with worse cumulative incidence of relapse (P=0.4637), relapse-free survival (P=0.4195) and OS (P=0.2349) compared to patients lacking the mutations, but statistical significance was not detected in this small cohort. Conclusions: CREBBP HAT domain mutations at diagnosis in adult ALL are found more frequently than in pediatric ALL. This may be one of the mechanisms that adult ALL has been associated with poor OS compared with pediatric ALL. In this study, CREBBP HAT domain mutations were observed in various subtypes of ALL: both B-cell and T-cell ALL, and both Philadelphia chromosome positive and negative ALL. In pediatric ALL, CREBBP mutations were frequently seen in relapsed patients but not in previously untreated patients. These observations suggest that CREBBP mutations play an important role in an additional late event(s) leading to the development and progression of ALL. Our study implies the possibility that mutations of the CREBBP gene are associated with the pathogenesis and prognostic marker of adult ALL and represent specific epigenetic modifiers in adult ALL, serving as potential therapeutic targets. Disclosures: No relevant conflicts of interest to declare.
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Camos, Mireia, Jordi Esteve, Dolors Colomer, Pedro Jares, Maria Rozman, Neus Villamor, Dolors Costa, et al. "Gene Expression Signature of Acute Myeloid Leukemia (AML) with T(8;16)(P11;P13) and MYST3-CREBBP Rearrangement: A Microarray Study Validated by Multiple Real-Time PCR." Blood 106, no. 11 (November 16, 2005): 3009. http://dx.doi.org/10.1182/blood.v106.11.3009.3009.
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Abstract AML with t(8;16)(p11;p13) is an infrequent leukemia subtype with characteristic clinical-biological features. The t(8;16)(p11;p13) translocation leads to the fusion of MYST3 and CREBBP genes, probably resulting in a disturbed transcriptional program of a myelo-monocytic precursor. In this study, the genetic signature of MYST3-CREBBP AML was compared with other well-defined AML subtypes. Genotypic analyses using oligonucleotide U133A arrays (Affymetrix) were performed on RNA of 23 AML patients, including three MYST3-CREBBP cases, PML-RARa (n=3), RUNX1-CBF2T1 (n=3), CBFβ-MYH11 (n=3), t(9;11)/AF9-MLL (n=1), monocytic AML (FAB M4/M5), n=8, and two cases of AML with multilineage dysplasia. Forty-six genes differentially expressed in MYST3-CREBBP cases were analyzed by multiple real-time RT-PCR using low-density arrays in an additional series of 40 patients, which included 7 MYST3-CREBBP cases, 18 AML samples with well characterized rearrangements (PML-RARa, n=3; RUNX1-CBF2T1, n=3; CBFβ-MYH11, n=3, MLL-rearranged AML, n=9), and 15 patients with normal karyotype AML. After unsupervised analysis, MYST3-CREBBP cases clustered together, displaying a distinctive expression signature. The analysis by RT-PCR confirmed the gene expression pattern found in the high-density array study. Thus, overexpressed genes included oncogene RET, several homeobox (HOXA9, HOXA10), genes involved in apoptosis (DAP) and prolactin gene. In contrast, cyclinD2, STAT5A, STAT5B and WT1 were underexpressed. Interestingly, MYST3-CREBBP cases showed up-regulation of a subgroup of genes (HOXA9, MEIS1, AKR7A2, CHD3, FLT3 and APBA2) that were also found overexpressed in MLL-rearranged leukemias. In summary, this study showed the distinctive genetic signature of MYST3-CREBBP AML, which harbours some similarities with MLL-rearranged AML. In addition, the low-density array methodology validated the results of the microarray analysis and allowed to study a larger series of patients, including samples not suitable for microarray analysis.
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Chafai Elalaoui, Siham, Wiam Smaili, Julien Van-Gils, Patricia Fergelot, Ilham Ratbi, Mariam Tajir, Benoit Arveiler, Didier Lacombe, and Abdelaziz Sefiani. "Clinical description and mutational profile of a Moroccan series of patients with Rubinstein Taybi syndrome." African Health Sciences 21, no. 2 (August 2, 2021): 960–67. http://dx.doi.org/10.4314/ahs.v21i2.58.
Full textAbstract:
Background: Rubinstein-Taybi syndrome (RSTS; OMIM 180849) is a rare autosomal dominant developmental disorder with an estimated prevalence of one case per 125,000 live births. RSTS is characterized by typical face, broad thumbs and halluces, short stature, and intellectual disability. Facial dysmorphy is characteristic with microcephaly, low frontal hairline, arched eyebrows, long eyelashes, convex profile of nose, narrow palate, and micrognathia. RSTS is mainly due to mutations or microdeletions of the CREBBP gene (about 60%) and more rarely of the EP300 gene (8%).
Objective: Clinical description and identification of mutations of patients with Rubinstein Taybi syndrome.
Methods: PCR and direct sequencing of CREBBP gene.
Results: We report here, the clinical and molecular data of a series of six Moroccan patients with a phenotype of RSTS. The molecular study of the major gene CREBBP (by Sanger Sequencing followed by CGH array, if sequence normal) revealed point mutations in five patients. For the sixth patient, CGH array revealed a microdeletion carrying the CREBBP gene. Through this work, we emphasize the importance of clinical expertise in the diagnosis, management and genetic counseling in Rubinstein Taybi syndrome.
Keywords: Rubinstein Taybi syndrome; CREBBP gene; mutation; Moroccan.
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Al-Qattan, Mohammad M., Zuhair A. Rahbeeni, Zuhair N. Al-Hassnan, Abdulaziz Jarman, Atif Rafique, Nehal Mahabbat, and Faris A. S. Alsufayan. "Chromosome 16p13.3 Contiguous Gene Deletion Syndrome including the SLX4, DNASE1, TRAP1, and CREBBP Genes Presenting as a Relatively Mild Rubinstein–Taybi Syndrome Phenotype: A Case Report of a Saudi Boy." Case Reports in Genetics 2020 (January 9, 2020): 1–5. http://dx.doi.org/10.1155/2020/6143050.
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The classic Rubinstein–Taybi syndrome Type 1 (RSTS1, OMIM 180849) is caused by heterozygous mutations or deletions of the CREBBP gene. Herein, we describe the case of a Saudi boy with chromosome 16p13.3 contiguous gene deletion syndrome (OMIM 610543) including the SLX4, DNASE1, TRAP1, and CREBBP genes, but presenting with a relatively mild RSTS1 syndrome phenotype. Compared with previously reported cases with severe phenotypes associated with 16p13.3 contiguous gene deletions, our patient had partial deletion of the CREBBP gene (with a preserved 5′ region), which might explain his relatively mild phenotype.
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Bommi-Reddy, Archana, Sungmi Park-Chouinard, David N. Mayhew, Esteban Terzo, Aparna Hingway, Michael J. Steinbaugh, Jonathan E. Wilson, Robert J. Sims, and Andrew R. Conery. "CREBBP/EP300 acetyltransferase inhibition disrupts FOXA1-bound enhancers to inhibit the proliferation of ER+ breast cancer cells." PLOS ONE 17, no. 3 (March 30, 2022): e0262378. http://dx.doi.org/10.1371/journal.pone.0262378.
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Therapeutic targeting of the estrogen receptor (ER) is a clinically validated approach for estrogen receptor positive breast cancer (ER+ BC), but sustained response is limited by acquired resistance. Targeting the transcriptional coactivators required for estrogen receptor activity represents an alternative approach that is not subject to the same limitations as targeting estrogen receptor itself. In this report we demonstrate that the acetyltransferase activity of coactivator paralogs CREBBP/EP300 represents a promising therapeutic target in ER+ BC. Using the potent and selective inhibitor CPI-1612, we show that CREBBP/EP300 acetyltransferase inhibition potently suppresses in vitro and in vivo growth of breast cancer cell line models and acts in a manner orthogonal to directly targeting ER. CREBBP/EP300 acetyltransferase inhibition suppresses ER-dependent transcription by targeting lineage-specific enhancers defined by the pioneer transcription factor FOXA1. These results validate CREBBP/EP300 acetyltransferase activity as a viable target for clinical development in ER+ breast cancer.
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Mullighan, Charles, Jinghui Zhang, Lawryn H. Kasper, Stephanie Lerach, Debbie Payne-Turner, Letha A. Phillips, J. Racquel Collins-Underwood, et al. "CREBBP Mutations In Relapsed Acute Lymphoblastic Leukemia." Blood 116, no. 21 (November 19, 2010): 413. http://dx.doi.org/10.1182/blood.v116.21.413.413.
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Abstract Abstract 413 Relapsed acute lymphoblastic leukemia (ALL) is a leading cause of death due to disease in young people, but the biologic determinants of treatment failure remain poorly understood. To identify novel sequence mutations contributing to relapsed in ALL, we resequenced 300 genes in matched diagnosis and relapse samples from 23 patients with ALL. The cohort included B-progenitor ALL with high hyperdiploidy (N=3), TCF3-PBX1 (N=1), ETV6-RUNX1 (N=3), rearrangement of MLL (N=3), BCR-ABL1 (N=3), and low hyperdiploid, pseudodiploid, or miscellaneous karyotypes (N=10). This identified 52 somatic non-synonymous mutations in 32 genes, many of which were novel, including mutations in the transcriptional coactivators CREBBP and NCOR1, the transcription factors ERG, SPI1, TCF4 and TCF7L2, components of the Ras signalling pathway, histone genes, genes involved in histone modification (CREBBP and CTCF), and genes previously shown to be targets of recurring DNA copy number alteration in ALL. Analysis of an extended cohort of 63 diagnosis-relapse cases and 200 acute leukaemia cases that did not relapse found that 19% of relapse cases had sequence or deletion mutations of CREBBP, which encodes the transcriptional coactivator and histone acetyltransferase (HAT) CREB-binding protein (CBP). The mutations were either present at diagnosis, acquired at relapse, or duplicated to homozygosity at the time of relapse. Moreover, several mutations acquired at relapse were detected in subclones at diagnosis, suggesting that the mutations confer a selective advantage and promote resistance to therapy. The mutations either resulted in truncated alleles or deleterious substitutions in highly conserved residues of the HAT domain. To examine the functional consequences of the mutations, we introduced wild type or mutant Crebbp alleles into Cbp/Ep300flox/flox murine embryonic fibroblasts, (dKO MEFs), and examined histone acetylation, expression of CREBBP target genes, and cellular proliferation. The HAT domain mutations resulted in impaired acetylation of the key Crebbp substrate, H3K18, and resulted in impaired transcriptional regulation of multiple CREBBP targets and pathways, including cAMP, dsRNA and dexamethasone responsive genes. The latter observation suggests that CREBBP mutations may directly result in resistance to corticosteroid therapy, which is a hallmark of high risk ALL. Together, these data these results extend the landscape of genetic alterations in leukemia, and identify mutations targeting transcriptional and epigenetic regulation as a mechanism of resistance in ALL. Disclosures: Pui: EUSA Pharma: Honoraria; Enzon: Honoraria; Sanofi-Aventis: Honoraria.
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Schroers-Martin, Joseph G., Joanne Soo, Gabriel Brisou, Florian Scherer, David M. Kurtz, Brian Sworder, Michael S. Khodadoust, et al. "Recurrent Crebbp Mutations in Follicular Lymphoma Appear Localized to the Committed B-Cell Lineage." Blood 136, Supplement 1 (November 5, 2020): 30–31. http://dx.doi.org/10.1182/blood-2020-142761.
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Background: Follicular lymphoma (FL) is genetically characterized by translocations involving the BCL2 locus on chromosome 18q21. However, up to 70% of healthy individuals also carry detectable t(14;18)-positive cells, suggesting BCL2 translocation is critical but not sufficient for FL development. Chromatin modifying genes (CMGs) including KMT2D, CREBBP, EZH2, and EP300 are almost ubiquitously mutated in FL. We previously reported the direct characterization by ultra-deep sequencing of pre-diagnostic blood and tissue specimens from 19 subjects who ultimately developed FL. CREBBP lysine acetyltransferase (KAT) domain mutations were the most commonly observed precursor lesions, detected in blood a median of 7.5 years before diagnosis in patients developing FL (8/19, 42%) but not in healthy adults with or without detected BCL2 translocations (0/13, p=0.01 and 0/20, p<0.001, respectively) (Schroers-Martin et al, ASH Annual Meeting 2017). While the BCL2 translocation is thought to occur in pre-B-cell precursors in the bone marrow, the high prevalence of CMG mutations raises the possibility that, analogous to myeloid age-related clonal hematopoiesis, such early lesions could occur in hematopoietic stem cells (HSCs) rather than in the committed B-cell lineage. Methods: To address this question of mutational hierarchy, we studied blood, bone marrow aspirate, and lymph node samples from 6 patients with CREBBP or KMT2D mutations in their FL tumor. We analyzed flow-sorted purified hematopoietic cell populations by deep sequencing, including isotype-specific mature B-cell, mature T-cell, and precursor populations encompassing HSCs and common lymphoid progenitors (CLPs). Patients additionally underwent routine clinical sequencing of tumor biopsy and marrow specimens. Results: Bulk sequencing of a diagnostic bone marrow aspirate from patient FL002 revealed a CREBBP mutation concordant with FL tumor biopsy. To ascertain the population bearing this mutation, we sorted to high purity viable marrow aspirate cells (Fig. A). The CREBBP mutation was confirmed in the mature B-cell compartment at an AF of 40.3% but was not detected in other cell populations. To validate this finding, a similar sorting strategy was employed on viable bone marrow aspirate or peripheral blood samples from another 4 FL patients bearing CREBBP mutations (Fig B). In each case, CREBBP was absent from the CD34+ precursor population. In patient CIML004 a characteristic KMT2D stop mutation was likewise absent in precursors. An atypical case sheds additional light on the localization of early FL mutations. Patient LYM267 was diagnosed with Grade 1-2 FL bearing CREBBP and NRAS mutations. Eight years into a prolonged remission after chemoimmunotherapy, he developed cutaneous and gingival myeloid sarcoma without radiographic or histopathological evidence of FL recurrence. While the CREBBP mutation was not detected in myeloid sarcoma or bone marrow biopsies, concordant NRAS mutations and clonal VDJ rearrangements were seen in all 3 compartments (Fig. C). This unusual clonal lineage favors the occurrence of the CREBBP mutation later than the branch point between morphologically distinct lymphoid and myeloid tumors, likely in the committed B-cell lineage after pre-BCR rearrangements (Fig D). Conclusions: HSCs are believed to be the cell of origin in several lymphoid leukemias, and mouse models have demonstrated lymphoma development with induced CREBBP lesions in HSCs (Horton et al Nat Cell Bio 2017). However, in sorted hematopoietic cell populations from marrow and peripheral blood, we observed CREBBP mutations in B-cell lineages but never in CD34+/CD20- precursor populations or paired lymphoid/myeloid disease. Our data therefore are not in support of HSCs as a precursor reservoir in FL. Given that cells harboring the t(14;18) translocation in healthy individuals appear derived from the germinal center, recurrent mutations in CREBBP are likely to occur after the pre-B-cell stage. Figure Disclosures Kurtz: Genentech: Consultancy; Foresight Diagnostics: Other: Ownership; Roche: Consultancy. Khodadoust:Kyowa Kirin: Consultancy; Seattle Genetics: Consultancy. Nadel:Innate Pharma: Research Funding; Institut Roche: Research Funding. Diehn:RefleXion: Consultancy; Varian Medical Systems: Research Funding; Illumina: Research Funding; Roche: Consultancy; BioNTech: Consultancy; AstraZeneca: Consultancy. Roulland:Celgene/BMS: Research Funding; Roche: Honoraria. Alizadeh:Pharmacyclics: Consultancy; Genentech: Consultancy; Janssen: Consultancy; Celgene: Consultancy; Chugai: Consultancy; Gilead: Consultancy; Pfizer: Research Funding; Roche: Consultancy.
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Sima, Aurora, Roxana Elena Smădeanu, Anca Angela Simionescu, Florina Nedelea, Andreea-Maria Vlad, and Cristina Becheanu. "Menke–Hennekam Syndrome: A Literature Review and a New Case Report." Children 9, no. 5 (May 22, 2022): 759. http://dx.doi.org/10.3390/children9050759.
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Background: Menke–Hennekam syndrome (MHS) is a rare and recently described syndrome consecutive to the variants in exon 30 or 31 in CREBBP (CREB-binding protein gene). The CREB-binding protein (CREBBP) and EP300 genes are two commonly expressed genes whose products possess acetyltransferase activity for histones and various other proteins. Mutations that affect these two genes are known to cause Rubinstein–Taybi syndrome (RTS); however, with the application of whole exome sequencing (WES) there were reports of variants that affect specific regions of exon 30 or 31 of these two genes but without the specific phenotype of RTS. Material and Methods: A review of the available literature was conducted, aimed at underscoring the difficulties in diagnosing MHS based on phenotype particularities. Results: Five applicable studies were identified by searching PubMed, Web of Science, and Scopus databases for publications up to November 2021 using the key terms “Menke–Hennekam syndrome” and “CREBBP”. Conclusions: In this paper, we present a new case and highlight the importance of exome sequencing to identify different mutations of exons 30 and 31 of the CREBBP gene involved in MHS, and we make formal recommendations based on our literature review.
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Melanie, Schoof, Gefion Dorothea Epplen, Carolin Walter, Annika Ballast, Dörthe Holdhof, Carolin Göbel, Sina Neyazi, Thomas Albert, Kornelius Kerl, and Ulrich Schüller. "MODL-11. The tumor suppressor CREBBP and the oncogene MYCN cooperate to induce malignant brain tumors in mice." Neuro-Oncology 24, Supplement_1 (June 1, 2022): i170—i171. http://dx.doi.org/10.1093/neuonc/noac079.634.
Full textAbstract:
Abstract CREBBP (cAMP response element-binding protein binding protein) and MYCN (v-myc avian myelocytomatosis viral oncogene neuroblastoma derived homolog), two essential proteins in central nervous system development that are described to be genetically altered in the same brain tumor entities, such as high grade glioma or medulloblastoma. Therefore, we hypothesized that alterations in both genes cooperate to induce brain tumor formation. In order to investigate molecular mechanisms and potential tumor formation, we established a novel mouse model combining a deletion of Crebbp with an overexpression of MYCN in neural stem cells (hGFAP-cre::CrebbpFl/Fl::lsl-MYCN). The animals developed highly aggressive forebrain tumors with 75 % penetrance already detectable in seven-day old mice, leading to tumor-related death within eight months. Although tumors mostly developed in the olfactory bulb region, we hypothesized that these tumors develop from subventricular stem cells that migrate into the olfactory bulb, where they settle and establish large solid tumors. In order to test this hypothesis, we analyzed neural stem cells (NSCs) derived from our mouse model. Compared to control cells, NSCs from transgenic mice have a significantly increased cell viability and proliferation in vitro and did not undergo full glial differentiation. In order to gain further insights into the cellular origin and biology of the tumors, we finally employed single cell RNA sequencing. Apart from a high intratumoral heterogeneity within the tumor cell population, data comparison with non-neoplastic CNS cell populations revealed a high similarity with transit-amplifying NSCs of the subventricular zone. Together, we were able to show the oncogenic potential of a combined loss of Crebbp and aberrant expression of MYCN in neural stem cells. This novel mouse model will be an important tool for understanding tumor-driving mechanisms and a valuable model for preclinical testing and will thereby help to improve patient care.
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Camos, Mireia, Jordi Esteve, Pedro Jares, Maria Rozman, Dolors Colomer, Neus Villamor, Dolors Costa, et al. "Gene Expression Profile of Acute Myeloid Leukemia (AML) with t(8;16)(p11;p13) and MYST3/CREBBP Rearrangement." Blood 104, no. 11 (November 16, 2004): 2054. http://dx.doi.org/10.1182/blood.v104.11.2054.2054.
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Abstract Translocation t(8;16)(p11;p13) is an infrequent chromosomal abnormality in de novo and secondary AML cases, leading to the fusion of MYST3 (MOZ) and CREBBP (CBP) genes, both of them harboring histone lysine acetyl-transferase activity. This AML variety displays specific clinical and biological features, although its gene expression profile is currently unknown. In this study, the genetic signature of AML cases with MYST3/CREBBP rearrangement was compared with the genetic profile of other well-defined AML subtypes. Genotypic analyses using oligonucleotide U133A arrays (Affymetrix) were performed on RNA of 19 AML samples, including t(8;16)-AML (n=3), t(15;17) (n=3), t(8;21) (n=2), inv(16)/t(16;16) (n=3), t(9;11) with AF9/MLL rearrangement (n=2), 3 cases with normal karyotype and flt-3 internal tandem duplication (flt-3 ITD), the three remaining samples corresponding to monocytic cases (M4/M5) without MLL rearrangement nor flt-3 ITD. After unsupervised analysis, cases of AML with t(8;16) clustered together, displaying a differential expression profile. Supervised analysis allowed the identification of the top 53 up-regulated and 28 down-regulated genes. Among the set of genes overexpressed, genes involved in chromatin remodelling and transcription (HOXA9, HOXA10, MEIS1, CHD3, SATB1) and protooncogenes (RET, flt-3, LMO2) were identified. In contrast, CREBBP gene and several members of the JAK-STAT pathway (STAT3, STAT5B, JAK2) were underexpressed. Interestingly, overexpression of multiple homeobox genes was detected in flt-3 ITD cases, some of them as a distinctive finding (HOXA2, HOXA3, HOXB6), and others (HOXA9, HOXA10, MEIS1) were found to be highly expressed in MYST3/CREBBP and MLL-rearranged samples. In conclusion, AML with t(8;16) and MYST3/CREBBP rearrangement shows a distinctive gene expression profile, with some similarities with MLL rearranged leukemias and flt-3 ITD AML cases, thus suggesting a partially common leukemogenic pathway.
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Bruno Marcelo Rocha Freire. "Identificação da fusão gênica MYST3-CREBBP em crianças com LMA e hemofagocitose." Revista Científica Hospital Santa Izabel 1, no. 4 (May 18, 2020): 42–44. http://dx.doi.org/10.35753/rchsi.v1i4.167.
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LMA apresenta uma translocação (8;16) (p11; p13) que gera a fusão gênica MYST3-CREBBP, que é mais comumente vista em adultos. Essa translocação (8;16) (p11; p13) está associada à idade precoce, diferenciação monocítica e hemofagocitose e sendo indicativo de pior prognóstico. Método: série de 266 casos de LMA em pacientes pediátricos no período de 2003 a 2012. Foram incluídas crianças com menos de 2 anos de idade, com LMA e presença de hemofagocitose. Foram incluídos 48 casos de LMA sem hemofagocitose. O método de hibridização fluorescente in situ (FISH) para os rearranjos MLL foi realizado ao diagnóstico, assim como o FISH para rearranjo MYST3-CREBBP. Resultados: dos casos analisados, 5 estavam associados à fusão MYST3-CREBBP e tinham associação com: hepatoesplenomegalia, lesões de pele ou cloroma localizados. Discussão: a alta frequência de LMA MYST3-CREBBP em bebês (idade < 24 meses) e casos de LMA congênita favorecem a hipótese de que a leucemia ocorre durante a vida intrauterina e pode ser utilizada para explorar melhor o entendimento da leucemogênese da LMA.
Publicado na Rev. Bras. Hematol hemoter. 2016;38(4):291–297, com mesmo título. Autores: Francianne Gomes Andrade, Elda Pereira Noronha, Rosania Maria Baseggio, Teresa Cristina Cardoso Fonseca, Bruno Marcelo Rocha Freire, Isis M. Quezado Magalhães, Ilana R. Zalcberg, Maria S. Pombo-de-Oliveira.
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Green, Michael R., Shingo Kihira, Chih Long Liu, Ramesh V. Nair, Raheleh Salari, Andrew J. Gentles, Jonathan Irish, et al. "Mutations in early follicular lymphoma progenitors are associated with suppressed antigen presentation." Proceedings of the National Academy of Sciences 112, no. 10 (February 23, 2015): E1116—E1125. http://dx.doi.org/10.1073/pnas.1501199112.
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Follicular lymphoma (FL) is incurable with conventional therapies and has a clinical course typified by multiple relapses after therapy. These tumors are genetically characterized by B-cell leukemia/lymphoma 2 (BCL2) translocation and mutation of genes involved in chromatin modification. By analyzing purified tumor cells, we identified additional novel recurrently mutated genes and confirmed mutations of one or more chromatin modifier genes within 96% of FL tumors and two or more in 76% of tumors. We defined the hierarchy of somatic mutations arising during tumor evolution by analyzing the phylogenetic relationship of somatic mutations across the coding genomes of 59 sequentially acquired biopsies from 22 patients. Among all somatically mutated genes, CREBBP mutations were most significantly enriched within the earliest inferable progenitor. These mutations were associated with a signature of decreased antigen presentation characterized by reduced transcript and protein abundance of MHC class II on tumor B cells, in line with the role of CREBBP in promoting class II transactivator (CIITA)-dependent transcriptional activation of these genes. CREBBP mutant B cells stimulated less proliferation of T cells in vitro compared with wild-type B cells from the same tumor. Transcriptional signatures of tumor-infiltrating T cells were indicative of reduced proliferation, and this corresponded to decreased frequencies of tumor-infiltrating CD4 helper T cells and CD8 memory cytotoxic T cells. These observations therefore implicate CREBBP mutation as an early event in FL evolution that contributes to immune evasion via decreased antigen presentation.
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Zhou, Ting, Stephanie N. Zimmer, Ziming Cheng, and Vivienne I. Rebel. "Myelodysplastic/Myeloproliferative Neoplasm in Crebbp+/− Mice Is a Transplantable Disease in Which Multiple Hematopoietic Cell Populations Are Involved, As Well As the Microenvironment." Blood 120, no. 21 (November 16, 2012): 1705. http://dx.doi.org/10.1182/blood.v120.21.1705.1705.
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Abstract Abstract 1705 Myelodysplastic/myeloproliferative neoplasms (MDS/MPNs) are myeloid malignancies that display features of both MDS and MPN, but cannot be properly assigned to either MDS or MPN. It is currently not known whether it originates from the hematopoietic stem cell (HSC) compartment (like MDS), from a more committed myeloid progenitor population, or a combination thereof. Fifteen to 40% of MDS/MPN patients develop acute myeloid leukemia (AML); whether the transformation occurs in a particular cell population is also unknown. We previously demonstrated that mice heterozygous for the CREB binding protein gene (Crebbp) develop MDS/MPN at 9–12 months of age and ∼40% of them progress to develop a hematologic malignancy. Thus, Crebbp+/− mice are an excellent model to address the before mentioned questions, which is important for the development of better strategies to treat MDS/MPN. For this purpose, we harvested and combined bone marrow from 1.5-year old Crebbp+/− mice (10 donors per experiment, thereby ensuring that the marrow of ∼4 donors harbored malignant hematopoietic cells) and transplanted it into lethally irradiated, wild-type recipients. Groups of mice either received unfractionated whole bone marrow (WBM) or populations purified by fluorescence-activated cell sorting. Naive Crebbp+/− mice had demonstrated functional and/or quantitative abnormalities in long-term and short-term HSCs, common myeloid progenitors (CMPs) and granulocyte/macrophage progenitors (GMPs) and we therefore focused on these populations. All transplant recipients also received unfractionated wild-type “helper cells” to increase survival. Mice were closely monitored and those suspected of having developed a hematopoietic disease were sacrificed and their hematopoietic system analyzed. Four independent experiments were performed and data were combined for analysis. Among the 18 recipients who received Crebbp+/− WBM, 8 recipients (44%) developed an early-onset AML with myelofibrosis, 2–7 months after the transplant, which was not preceded by MDS. The other 10 recipients (56%) developed MDS/MPN, 12–18 months after the transplant. These mice displayed ineffective hematopoiesis, evidenced by a normocellular bone marrow, significant leukopenia, and trilineage dysplasia. One of these 10 Crebbp+/− WBM recipients that developed MDS/MPN subsequently progressed to AML. In contrast, none of the 15 recipients of Crebbp+/− HSCs (defined as Lin−Sca-1+c-Kit++ (LSK) cells, including long-term and short-term progenitors, as well as lymphoid-restricted progenitors) developed early-onset AML. Instead, 1 developed MDS/MPN while the remainder developed MDS by 11–17 months after the transplant, with one of them progressing to a disease resembling human mature T-cell leukemia. Transplantation of Crebbp+/− CMPs and GMPs also failed to cause early-onset AML and, as expected, gave rise to extremely low long-term reconstitution. Thus, these mice were mostly reconstituted by the co-transplanted wild-type “helper cells”. However, unexpectedly, 9 out of 24 (38%) showed <10% dysplastic cells in 1 or more lineages, while 4 (17%) developed overt MDS, i.e. >10% dysplastic cells and 2 (8%) developed MPD or AML with myelofibrosis. Control mice, i.e., recipients of wild-type BM cells remained healthy for the duration of the experiments. The results of these transplantation experiments show that in this mouse model, MDS/MPN is transplantable. However, it requires transplantation of WBM, since the transplantation of LSK cells resulted in MDS, suggesting that the microenvironment may play a crucial role in the etiology of MDS/MPN. This notion is in concordance with our previous study, demonstrating that Crebbp+/− mice transplanted with wild-type cells developed MPD that originated from the transplanted wild-type cells. This notion is further supported by the outcome of the CMP and GMP transplantation experiments, suggesting that abnormal myeloid progenitors are also important factors for MDS/MPN disease development. Moreover, malignant transformation seems to occur in a non-LSK cell that is more differentiated than CMPs and GMPs. Alternatively, malignant transformation requires all hematopoietic and non-hematopoietic cells to be present, again suggesting that MDS/MPN is a complex disease where both the hematopoietic compartment and its bone marrow microenvironment are affected. Disclosures: No relevant conflicts of interest to declare.
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Sokol, Ethan, Smruthy Sivakumar, Brennan Decker, Jeffrey Ross, and Priti Hegde. "Abstract P5-13-02: Serially biopsied BRCA1/2 mutant breast tumors frequently acquire alterations in BRCA1, BRCA2, and CREBBP." Cancer Research 82, no. 4_Supplement (February 15, 2022): P5–13–02—P5–13–02. http://dx.doi.org/10.1158/1538-7445.sabcs21-p5-13-02.
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Abstract Introduction Tumors with alterations in BRCA1 or BRCA2 (BRCAm) are sensitive to poly ADP ribose polymerase inhibitors (PARPi), with significant benefit in ovary, breast, prostate, and pancreatic cancers. Despite strong early responses on PARPi, many patients eventually exhibit relapse. Previous literature has identified BRCA1/2 reversion as a common class of acquired alteration in the resistance setting; however, additional resistance pathways have not been well characterized. Here, we examined 100 BRCAm patients who were profiled with serial biopsy during clinical care to identify possible therapy resistance alterations. Methods Comprehensive genomic profiling (CGP) was carried out in a Clinical Laboratory Improvement Amendments (CLIA)-certified, CAP (College of American Pathologists)-accredited laboratory (Foundation Medicine Inc., Cambridge, MA, USA). Tissue biopsy CGP was performed on FFPE blocks examining at least 324 genes for all classes of alterations (FoundationOne® and FoundationOne® CDx). Liquid biopsy CGP was performed examining at least 62 genes (FoundationACT®, FoundationOne®Liquid).One hundred breast cancer patients with baseline BRCAm were serially profiled with CGP testing during routine clinical care (n=50 tissue then liquid; n=50 tissue for both tests). As a comparator, we examined 1,294 patients without a baseline BRCAm (BRCAwt) profiled with serial biopsy (n = 585 tissue then liquid; n=709 tissue for both tests). Co-occurrence analyses (Fisher’s exact) were run on the full research dataset, including 12,198 breast-biopsied (local) and 16,586 metastatic-biopsied samples. All classes of genomic alterations were included in the analysis. Results Of the serially biopsied samples, 7.2% (100/1,394) had baseline BRCAm. In patients with baseline BRCAm, acquired alterations were frequently observed in BRCA1/2 (17%), ESR1 (16%), TP53 (15%), MYC (12%), CREBBP (10%), RB1 (9%), PIK3CA (9%), and NF1 (5%). To understand if any of these are specific to the BRCAm population, we compared the milieu of acquired alterations to a serially biopsied BRCAwt cohort (n=1,294). While many of the alterations were common and shared across both cohorts, likely due to endocrine therapy use, acquired alterations in BRCA1, BRCA2, and CREBBP were specifically enriched in the BRCAm population (all p<0.005). Acquired BRCA1/2 mutations were predicted reversion mutations and spanned a range of mechanisms, including large deletion/rearrangement events that remove the exon of the truncal mutation, point mutations that change a nonsense BRCA1/2 to a missense event, upstream indels that revert the frame of the truncal BRCA1/2 event, and splice site mutations that lead to a skipping of the truncal BRCA1/2. Of note, a majority of cases with a reversion involved a rearrangement event (9/17; 53%). For patients with a baseline BRCA1/2 short variant mutation, there was a trend towards a higher rate of reversion in BRCA2 v BRCA1 (23.1% v 18.5%, p>0.05). Acquired BRCAm were never observed in cases with baseline deletion of BRCA1/2. The high rate of acquired CREBBP alterations was specific to the BRCAm population (10% BRCAm v 1% BRCAwt; p = 0.0013), suggesting a possible role for CREBBP in platinum or PARPi resistance. Consistent with this, BRCAm and CREBBP significantly co-occur in the metastatic setting (Odds ratio, OR = 1.6; p = 0.016) but not in the local setting (OR = 1.1; p = 0.68). Conclusions Analysis of serially biopsied BRCAm breast cases revealed frequent acquisition of BRCA1/2 reversion mutations and CREBBP alterations that are not frequently observed in BRCAwt samples. Additional studies are warranted to investigate the possible role of CREBBP in PARPi therapy resistance. Citation Format: Ethan Sokol, Smruthy Sivakumar, Brennan Decker, Jeffrey Ross, Priti Hegde. Serially biopsied BRCA1/2 mutant breast tumors frequently acquire alterations in BRCA1, BRCA2, and CREBBP [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-13-02.
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Zhang, Hui, Maoxiang Qian, Shirley, Kow Yin Kham, Shuguang Liu, Chuang Jiang, Xujie Zhao, Yi Lu, et al. "Whole Transcriptome Sequencing Identified a Distinct Subtype of Acute Lymphoblastic Leukemia with Abnormalities of CREBBP and EP300." Blood 128, no. 22 (December 2, 2016): 3912. http://dx.doi.org/10.1182/blood.v128.22.3912.3912.
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Abstract While acute lymphoblastic leukemia (ALL) is a prototype of cancer that can be cured by chemotherapy alone, current ALL treatment regimens rely primarily on conventional cytotoxic agents with significant acute and long-term side effects. Better understanding of genomic landscape of ALL is critical for developing molecularly targeted therapy and implementing genomics-based precision medicine in this cancer. In particularly, sentinel chromosomal translocations are common in ALL and often involve key transcription factors important for hematopoiesis. Epigenetic regulator genes are also frequently targeted by somatic genomic alterations such as sequence mutations (e.g., CREBBP) and gene fusions (e.g., MLL, EP300). To comprehensively define transcriptomic abnormalities in childhood ALL, we performed RNA-seq of an unselected cohort of 231 children enrolled on the MaSpore frontline ALL protocols in Singapore or Malaysia. In total, we identified 58 putatively functional and predominant fusion genes in 125 patients (54.1%), the majority of which have not been reported previously. In particular, we described a distinct ALL subtype with a characteristic gene expression signature driven by chromosomal rearrangements of the ZNF384 gene with different partners (i.e., histone acetyl-transferases EP300 and CREBBP, TAF15, and TCF3). In 9 of 11 ALL cases with ZNF384 rearrangements, the breakpoint in this gene was invariably between exon 2 and exon 3, resulting in deletion of the 5'-UTR and then in-frame fusion of the entire ZNF384 coding sequence with the partner genes. The top two most significantly up-regulated genes in the ZNF384-rearranged group were CLCF1 and BTLA, whose expression levels were 15.5- and 15.0-fold higher than in ALL cases with wildtype ZNF384, respectively. In fact, ZNF384 binding was identified within the CLCF1 and BTLA loci (particularly the promoter regions) by chromatin immunoprecipitation sequencing in B lymphoblasoid cells. Using luciferase transcription driven by CLCF1 promoter in HEK293T cells as a model system, we observed significantly greater transcription activity with EP300-ZNF384 fusion compared to cells expressing wildtype ZNF384, suggesting that this chimeric gene resulted in gain of ZNF384 function. Similar results were obtained with luciferase transcription assay driven by the BTLA promoter. In human ALL cells, CLCF1 and BTLA promoter activities were consistently and significantly higher in ZNF384-rearranged ALL than in ALL cell line with wildtype ZNF384. To examine the effects of ZNF384 fusion on hematopoietic stem and progenitor cell (HSPCs) function, we also evaluated colony forming potential of HSPC in vitro upon ectopic expression of ZNF384 fusions. While there was marked suppression of colonies from myeloid and erythoid lineages, expression of EP300-ZNF384 or CREBBP-ZNF384 significantly stimulated preB cell colony formation. However, neither EP300- nor CREBBP-ZNF384 fusion was able to transform mouse hematopoietic precursor cell Ba/f3 in vitro, but instead increased the transforming potential of other oncogenic mutations (NRASG12D). EP300-ZNF384 and CREBBP-ZNF384 fusion proteins lacked the histone acetyltransferase (HAT) domain, and showed only 25% and 10% of HAT activity of full-length EP300 and CREBBP, respectively, with dominant-negative effects. Also, expression of EP300-ZNF384 led to significant decrease in global H3 acetylation in Ba/f3 cells in vitro. Finally, in NRASG12D-transformed Ba/f3 cells, co-expression of EP300-ZNF384 or CREBBP-ZNF384 substantially potentiated cytotoxic effects of histone deacetylase inhibitor vorinostat. Similarly, in a panel of human ALL cell lines, ZNF384-rearrangement was also associated with increased sensitivity to vorinostat, suggesting that some ZNF384-rearranged ALL may benefit from therapeutic agents targeting histone acetylation regulation. In conclusion, our results indicate that gene fusion is the major class of genomic abnormalities in childhood ALL and chromosomal rearrangements involving EP300 and CREBBP may cause global epigenetic deregulation in ALL with potentials for therapeutic targeting. Disclosures No relevant conflicts of interest to declare.
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Pasqualucci, Laura, David Dominguez-Sola, Annalisa Chiarenza, Giulia Fabbri, Adina Grunn, Vladimir Trifonov, Lawryn H. Kasper, et al. "Genome-Wide Analysis Reveals Frequent Inactivating Mutations of Acetyltransferase Genes In B-Cell Lymphoma." Blood 116, no. 21 (November 19, 2010): 474. http://dx.doi.org/10.1182/blood.v116.21.474.474.
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Abstract Abstract 474 B cell non-Hodgkin lymphoma (B-NHL) comprises a variety of biologically and clinically distinct diseases whose pathogenesis is associated with largely unique genetic lesions affecting oncogenes and tumor suppressor genes. The identification of the complete set of genes and cellular pathways that are altered in the lymphoma cell is critical for a full understanding of the genesis and maintenance of these malignancies. Using genome-wide copy number analysis and high throughput whole exome sequencing, we have recently found that the two most common types of B-NHL, follicular lymphoma (FL) and diffuse large B cell lymphoma (DLBCL), are associated with frequent structural alterations inactivating the genes that encode CREBBP and, in a smaller fraction of cases, EP300, two highly related histone and non-histone acetyltranferases (HAT) that act as transcriptional coactivators in multiple signaling pathways. Overall, ~29% (n=39/134) of DLBCL and 32% (n=15/47) of FL samples display genomic deletions and/or somatic point mutations that remove or inactivate the HAT domain of CREBBP. In 19 additional cases (15 DLBCL and 4 FL), mutations and deletions were found in the EP300 gene, also predicting its functional loss. With few exceptions, structural alterations of CREBBP and EP300 were mutually exclusive, suggesting that inactivation of these loci is at least in part functionally equivalent. Thus, in total ~36% of all DLBCL (n=49/134) and at least 40% of FL cases (n=19/47) display genetic aberrations targeting acetyltransferase genes. Interestingly, while these lesions were observed in both DLBCL phenotypic subtypes, their frequency appears to be significantly higher in germinal center B cell type DLBCL, where they account for ~46% of the cases (n=30/65) as compared to 27% in activated B cell type DLBCL (n=11/69; p<0.01). CREBBP and EP300 structural alterations commonly affect a single allele, suggesting that reduction in HAT dosage is important for lymphomagenesis. While the functional consequences of these lesions are likely to be broad on gene transcription, we demonstrate specific defects in the acetylation-mediated inactivation of the BCL6 onco-protein and activation of the p53 tumor suppressor, both of which contribute to transformation. These results identify CREBBP/EP300 alterations as a major pathogenetic mechanism shared by common forms of B-NHL, and have direct implications for the development of therapeutic regimens targeted to acetylation/deacetylation mechanisms. Disclosures: No relevant conflicts of interest to declare.
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Cortopassi, Wilian A., Kiran Kumar, and Robert S. Paton. "Cation–π interactions in CREBBP bromodomain inhibition: an electrostatic model for small-molecule binding affinity and selectivity." Organic & Biomolecular Chemistry 14, no. 46 (2016): 10926–38. http://dx.doi.org/10.1039/c6ob02234k.
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Tokunaga, Kenji, Shunichro Yamaguchi, Taizo Shimomura, Hitoshi Suzushima, Yutaka Okuno, Hiroaki Mitsuya, and Norio Asou. "Accumulation Of Gene Alterations Of TP53, Crebbp and IKZF1 Is a Prognostic Factor In Adult Acute Lymphoblastic Leukemia." Blood 122, no. 21 (November 15, 2013): 1386. http://dx.doi.org/10.1182/blood.v122.21.1386.1386.
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Abstract Aims Mutations of the genes associating with cell differentiation or proliferation are recognized as factors of tumorigenesis or prognosis in hematological malignancies. In pediatric acute lymphoblastic leukemia (ALL), alterations of IKZF1 (a factor of lymphocyte differentiation), TP53 (a cell cycle regulator) and CREBBP (a histone modifier) are found as possible prognostic markers for stratification of treatments. On the other hand, in adult ALL, clinical significance of such alterations remains to be determined. In the present work, we examined whether the mutations in those genes affected the incidence and prognosis in adult ALL patients. Methods We investigated 87 adult patients with newly diagnosed ALL treated with JALSG protocols between 1986 and 2011. Age ranged from 15 to 86 years, with a median of 51 years. We obtained cDNA and genomic DNA from the peripheral blood or bone marrow mononuclear cells at diagnosis. CREBBP mutations are dominantly identified in the histone acetyltransferase (HAT) domain. HAT domain in the CREBBP gene was amplified by PCR using cDNA and was subjected to direct sequencing. Additionally other histone modifiers, EZH2, EED, and UTX, were sequenced as the same as in CREBBP. TP53 exons 5 – 8 and 10, in which mutations were commonly reported, were sequenced using genomic DNA. We amplified IKZF1 using RT-PCR for detecting aberrant dominant negative isoforms: Ik6 and Ik10. Genomic deletions of IKZF1 were assessed with RQ-PCR or genomic DNA PCR. We compared clinical profiles between patients with and without such gene mutations. The present study was approved by the Institutional Review Boards and informed consent was obtained from each patient according to guidelines based on the revised Declaration of Helsinki. Results In 87 adult patients with ALL, alterations of CREBBP, EED, TP53 and IKZF1 were detected in 7 (9.5%), 3 (4.8%), 6 (6.9%) and 42 (50%), respectively. None of EZH2 and UTX mutation was found. The alterations of CREBBP and IKZF1 at diagnosis in adult patients were more frequent than those in pediatric patients ever reported. Some gene mutations were not found frequently. Each gene mutation per se did not significantly affect prognosis. We tried to predict the prognosis by scoring gene mutations and chromosomal abnormalities. Philadelphia chromosome (Ph) has great impact to prognosis of patients with ALL. We scored the number of mutated genes and Ph for each patient. As the score was higher, adult patients with ALL had poorer relapse-free survival (P=0.0439) and OS (P=0.4819), but statistical significance was not detected in this small cohort. Conclusions and Discussion Single gene mutations, such as IKZF1, can predict the prognosis in pediatric ALL. In adult ALL, however, only few gene mutations are reported to be promising prognostic factors which have impacts to treatment outcomes. Scoring system may be a useful method for predicting prognosis and stratifying treatment in adult ALL. Our study implies the possibility that a variety and heterogeneity of genetic alterations in adult ALL are associated with the pathogenesis for treatment resistance and prognostic marker of adult ALL. Disclosures: No relevant conflicts of interest to declare.
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Mosquera Orgueira, Adrián, Roi Ferreiro Ferro, José Ángel Díaz Arias, Carlos Aliste Santos, Beatriz Antelo Rodríguez, Laura Bao Pérez, Natalia Alonso Vence, et al. "Detection of new drivers of frequent B-cell lymphoid neoplasms using an integrated analysis of whole genomes." PLOS ONE 16, no. 5 (May 4, 2021): e0248886. http://dx.doi.org/10.1371/journal.pone.0248886.
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B-cell lymphoproliferative disorders exhibit a diverse spectrum of diagnostic entities with heterogeneous behaviour. Multiple efforts have focused on the determination of the genomic drivers of B-cell lymphoma subtypes. In the meantime, the aggregation of diverse tumors in pan-cancer genomic studies has become a useful tool to detect new driver genes, while enabling the comparison of mutational patterns across tumors. Here we present an integrated analysis of 354 B-cell lymphoid disorders. 112 recurrently mutated genes were discovered, of which KMT2D, CREBBP, IGLL5 and BCL2 were the most frequent, and 31 genes were putative new drivers. Mutations in CREBBP, TNFRSF14 and KMT2D predominated in follicular lymphoma, whereas those in BTG2, HTA-A and PIM1 were more frequent in diffuse large B-cell lymphoma. Additionally, we discovered 31 significantly mutated protein networks, reinforcing the role of genes such as CREBBP, EEF1A1, STAT6, GNA13 and TP53, but also pointing towards a myriad of infrequent players in lymphomagenesis. Finally, we report aberrant expression of oncogenes and tumor suppressors associated with novel noncoding mutations (DTX1 and S1PR2), and new recurrent copy number aberrations affecting immune check-point regulators (CD83, PVR) and B-cell specific genes (TNFRSF13C). Our analysis expands the number of mutational drivers of B-cell lymphoid neoplasms, and identifies several differential somatic events between disease subtypes.
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Los-de Vries, G. Tjitske, Wendy B. C. Stevens, Erik van Dijk, Carole Langois-Jacques, Andrew J. Clear, Phylicia Stathi, Margaretha G. M. Roemer, et al. "Genomic and microenvironmental landscape of stage I follicular lymphoma, compared with stage III/IV." Blood Advances 6, no. 18 (September 26, 2022): 5482–93. http://dx.doi.org/10.1182/bloodadvances.2022008355.
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Abstract Although the genomic and immune microenvironmental landscape of follicular lymphoma (FL) has been extensively investigated, little is known about the potential biological differences between stage I and stage III/IV disease. Using next-generation sequencing and immunohistochemistry, 82 FL nodal stage I cases were analyzed and compared with 139 FL stage III/IV nodal cases. Many similarities in mutations, chromosomal copy number aberrations, and microenvironmental cell populations were detected. However, there were also significant differences in microenvironmental and genomic features. CD8+ T cells (P = .02) and STAT6 mutations (false discovery rate [FDR] <0.001) were more frequent in stage I FL. In contrast, programmed cell death protein 1–positive T cells, CD68+/CD163+ macrophages (P < .001), BCL2 translocation (BCL2trl+) (P < .0001), and KMT2D (FDR = 0.003) and CREBBP (FDR = 0.04) mutations were found more frequently in stage III/IV FL. Using clustering, we identified 3 clusters within stage I, and 2 clusters within stage III/IV. The BLC2trl+ stage I cluster was comparable to the BCL2trl+ cluster in stage III/IV. The two BCL2trl– stage I clusters were unique for stage I. One was enriched for CREBBP (95%) and STAT6 (64%) mutations, without BLC6 translocation (BCL6trl), whereas the BCL2trl– stage III/IV cluster contained BCL6trl (64%) with fewer CREBBP (45%) and STAT6 (9%) mutations. The other BCL2trl– stage I cluster was relatively heterogeneous with more copy number aberrations and linker histone mutations. This exploratory study shows that stage I FL is genetically heterogeneous with different underlying oncogenic pathways. Stage I FL BCL2trl– is likely STAT6 driven, whereas BCL2trl– stage III/IV appears to be more BCL6trl driven.
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Thomas, Anish. "CREBBP-mutated cancers HAT-tricked." Science Translational Medicine 7, no. 317 (December 9, 2015): 317ec212. http://dx.doi.org/10.1126/scitranslmed.aad8032.
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van Belzen, Martine, Oliver Bartsch, Didier Lacombe, Dorien J. M. Peters, and Raoul C. M. Hennekam. "Rubinstein–Taybi syndrome (CREBBP, EP300)." European Journal of Human Genetics 19, no. 1 (July 28, 2010): 3. http://dx.doi.org/10.1038/ejhg.2010.124.
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О.Р., Исмагилова,, Адян, Т.А., Бескоровайная, Т.С., and Поляков, А.В. "Molecular-genetic analysis of Rubinstein-Taybi syndrome in Russia." Nauchno-prakticheskii zhurnal «Medicinskaia genetika, no. 9 (September 30, 2022): 48–51. http://dx.doi.org/10.25557/2073-7998.2022.09.48-51.
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Синдром Рубинштейна-Тейби (СРТ) - редкая наследственная патология, характеризующаяся умственной отсталостью и задержкой физического развития в сочетании с определённым комплексом внешних проявлений и аномалиями различных органов и систем. Диагностический алгоритм нацелен на поиск мутаций в двух основных генах: CREBBP и EP300, обнаруживаемых примерно в 60% клинически определённых случаев СРТ. В настоящей работе представлены результаты молекулярно-генетического анализа выборки российских пациентов с СРТ. Несмотря на то, что около 40% пациентов не получают подтверждения клинического диагноза, существуют перспективы для более глубокого понимания патогенеза заболевания и усовершенствования принципов диагностики. Rubinstein-Taybi syndrome (RTS) - rare hereditary disorder characterized by intellectual disability and growth retardation in conjunction with specific craniofacial and skeletal features and a wide range of multiple congenital anomalies. To date, mutations in two genes: CREBBP and EP300 can be discovered in about 60% of clinically identified patients with RTS. We herein report the result of molecular analysis in a cohort of Russian RTS patients. Despite the fact that we cannot confirm the diagnosis of RTS in 40% of causes, there are possibility for a better understanding of causative molecular mechanisms and improving the diagnostic process.
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Chueire, Ana Flávia Wendpap, Marcelle Lupi Gasparini, and Clarissa Torresan. "Síndrome de Rubinstein-Taybi, genética e clínica: uma revisão integrativa." Brazilian Journal of Health Review 5, no. 5 (October 31, 2022): 21727–42. http://dx.doi.org/10.34119/bjhrv5n5-309.
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Introdução: A Síndrome de Rubinstein-Taybi é uma doença genética rara, com amplo espectro fenotípico. É conhecida classicamente por sua alteração nos genes CREBBP e EP300 e clinicamente, pelo atraso intelectual, microcefalia, fácies típica, hálux alargado e alterações cognitivas. Entretanto, há uma variedade de quadros clínicos e genéticos muito ampla e pouco difundidas, o que dificulta o diagnóstico. Nesse sentido, esta revisão tem o objetivo de apresentar os diversos achados clínicos e facilitar o diagnóstico. Além disso, expõe os aspectos genéticos e a mutação mais frequente encontrada. Metodologia: foi realizada uma revisão integrativa, com o método PRISMA, com o intuito de demonstrar os principais aspectos genéticos e clínicos da síndrome de Rubinstein-Taybi, com a seleção final de 25 artigos. Resultados: foi observada uma clínica muito variada, com destaque para alterações não citadas no quadro clínico clássico e complicações em diversos sistemas. Na parte genética, foi encontrada maior mutação no gene CREBBP em relação ao EP300. Discussão: não foi possível relacionar o quadro genético com os fenótipos de forma muito específica.
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Spriano, Filippo, Eugenio Gaudio, Chiara Tarantelli, Gaetanina Golino, Luciano Cascione, Emanuele Zucca, Anastasios Stathis, Francis J. Giles, and Francesco Bertoni. "Targeting Both BET and Crebbp/EP300 Proteins with the Novel Dual Inhibitor NEO2734 Leads to More Preclinical Anti-Tumor Activity in Diffuse Large B Cell Lymphomathan with Single BET or Crebbp/EP300 Inhibitors." Blood 132, Supplement 1 (November 29, 2018): 4174. http://dx.doi.org/10.1182/blood-2018-99-114900.
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Abstract Background: Lymphoma cells have frequent deregulation of their epigenome. The Bromodomain (BRD) and Extra-Terminal domain (BET) proteins are key regulators of the transcription process (Stathis & Bertoni, Cancer Discovery 2018). The acetyltransferases cyclic AMP response element binding protein (CREB)-binding protein (CBP) and the E1A interacting protein of 300 kDa (EP300 or P300) are highly homologous BRD-containing transcriptional co-activators and their genes are often mutated in diffuse large B cell lymphoma (DLBCL) (Pasqualucci et al, Nature 2011; Morin et al, Nature 2011). Targeting the individual classes of proteins is a new therapeutic approach, as shown especially by BET inhibitors with both preclinical and early clinical anti-lymphoma activity (Stathis & Bertoni, Cancer Discovery 2018). NEO2734 (Epigene Therapeutics Inc) is a novel oral dual inhibitor of BET and CREBBP/EP300 proteins with pre-clinical activity in a spectrum of solid tumors (Giles et al, ESMO 2018). Here, we present the first data exploring its anti-tumor activity in DLBCL models. Methods: Lymphoma cell lines were exposed to increasing doses of compounds for 72h. Cell proliferation was measured by using the MTT assay. Results: Twenty-seven DLBCL cell lines were exposed to NEO2734. The compound showed anti-tumor activity with a median IC50 of 157 nM (95% C.I., 135-214). Cell lines derived from activated B-cell-like DLBCL (ABC DLBCL) (n.=7) were more sensitive than the ones derived from germinal center B-cell (GCB) DLBCL (n.=20) (P = 0.04). No difference were observed based on MYC gene status (translocation: yes, n=8; no, n.=13), BCL2 gene status (translocation: yes, n=12; no, n.=6), TP53 gene status (inactive: yes, n=14; no, n.=6), double hit MYC/BCL2 (yes, n.=6; no, n.=14), CREBBP gene status (mutated, n.=10; wild type, n.=16), or EP300 gene status (mutated, n.=5; wild type, n.=20). As comparison, all the cell lines were also exposed to a BET inhibitor (birabresib, OTX015) (Boi et al, Clinical Cancer Res 2015) and to a CREBBP/EP300 inhibitor (CBP30) (Hammitzsch et al, PNAS 2015). The median IC50 values of the two molecules were 237 nM (95% C.I., 171-344) and 5.5 μM (95% C.I., 4.2-8.3 μM), respectively. The data obtained for birabresib were in agreement with what we had previously reported (Boi et al, Clinical Cancer Res 2015). The three compounds presented a similar pattern of anti-proliferative activity across all the cell lines (NEO2734 and birabresib: R2 =0.84, P < 0.001; NEO2734 and CBP30, R2 = 0.73, P < 0.001; birabresib and CBP30, R2 = 0.73, P < 0.001) but with different degrees of IC50. Both NEO2734 and birabresib were more potent than CBP30 (P <0.0001). The novel dual inhibitor was more potent than birabresib (P=0.0182) and the difference was even bigger considering the compounds' IC90 (P = 0.0025): median values were 1.1 μM (95% C.I., 735 nM - 2.7 μM) and 20 μM (95% C.I., 2 - 30 μM) for the dual inhibitor and for the BET inhibitor, respectively. Conclusions: The novel dual BET and CREBBP/EP300 inhibitor NEO2734 showed strong in vitro anti-tumor activity across a large panel of DLBCL cell lines and it appeared more potent than single BET or CREBBP/EP300 inhibitors. Disclosures Zucca: Celltrion: Consultancy; AstraZeneca: Consultancy. Stathis:Oncology Therapeutic Development: Research Funding. Giles:Actuate Therapeutics Inc: Employment, Equity Ownership.
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Schroers-Martin, Joseph G., Joanne Soo, Gabriel Brisou, Florian Scherer, David M. Kurtz, Brian Sworder, Michael S. Khodadoust, et al. "Tumor-Confirmed Follicular Lymphoma Mutations Are Detectable in Peripheral Blood Years Prior to Clinical Diagnosis." Blood 138, Supplement 1 (November 5, 2021): 709. http://dx.doi.org/10.1182/blood-2021-151058.
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Abstract Background: Mutations in chromatin modifying genes (CMGs) including KMT2D, CREBBP, EZH2, and EP300 have been inferred as early events in follicular lymphoma (FL) by truncal status in mature tumors and persistence between diagnosis and relapse. We previously reported frequent detection of CREBBP lysine acetyltransferase (KAT) domain mutations in pre-diagnostic blood and tissue specimens from individuals later developing FL (Schroers-Martin et al, ASH Annual Meeting 2017). However, the limited availability of paired tumor biopsies has precluded confirmation of concordance between precursor lesions and subsequent clinical malignancy. Methods: The American Cancer Society (ACS) Cancer Prevention Study-II (CPS-II) LifeLink cohort collected screening blood or saliva samples from over 100,000 cancer-free American participants between 1998 and 2002. To evaluate detection of tumor-confirmed variants in pre-diagnostic specimens, we identified 29 FL patients with available FFPE tumor biopsy and screening sample (Fig A). The median age at screening was 71 years (range 56-83) with a median time to FL diagnosis of 56 months (TTD, range 6-139). Tumor biopsies were sequenced utilizing hybrid capture sequencing for commonly mutated lymphoma genes. DNA extracted from pre-diagnostic blood or saliva cell pellet specimens was sequenced utilizing error-corrected CAPP-Seq (Newman et al Nat Biotech 2016) to a median depth of 5204x. We sequenced to similar depths peripheral blood DNA from control cohorts of individuals with detectable t(14;18) but no subsequent lymphoma diagnosis (n=14) and healthy individuals without detectable t(14;18) by PCR (n=20). Results: Coding mutations were identified from all tumors with a mutational distribution similar to prior FL sequencing studies. Tumor-derived variants were detected in 7 of 29 paired pre-diagnostic specimens (24%) at a median TTD of 44 months (range 11-112 months, Fig B). The statistical significance of detection was assessed using a previously described approach based on Monte Carlo sampling (Newman et al Nat Biotech 2016) and the error distribution of affected loci in control cohorts. While an outlier case contained concordant TNFRSF14, FOXO1, and STAT6 mutations 90 months pre-diagnosis at an elevated allelic fraction (AF) of 1.8%, the mean AF of other detected precursor variants was 0.091%. Individuals with detected variants were not older (Fig C) nor significantly closer to clinical diagnosis (Fig D). The most frequently detected lesions were CREBBP (6/15 cases, 40%) and BCL2 (3/13, 23%) with one case demonstrating a fuller mutational profile including FOXO1 and ARID1A at 44 months before diagnosis. All detected precursor CREBBP variants localized to the KAT domain, reflecting prior observations in pre-diagnostic samples without confirmatory biopsy (Fig E). Of note, saliva cell pellets may contain 30% or more hematopoietic DNA (Kaur et al Chimerism 2012) and we detected tumor-confirmed variants in both saliva and blood screening specimens (Fig F) with no significant difference in AF (Fig G). In an illustrative independent case with available imaging, a patient undergoing radical prostatectomy was found to have involvement of a pelvic lymph node with in situ follicular neoplasia (ISFN). Staging PET/CT showed no evidence of FL (Fig H) and he was followed expectantly for 4 years without emergent disease. Eight years after surgery he presented with inguinal swelling and bilateral FDG-avid adenopathy on PET/CT. Excisional biopsy confirmed low grade FL and sequencing for M7-FLIPI revealed a CREBBP KAT domain variant. Retrospective sequencing of serial peripheral blood specimens from his initial surveillance showed detectable CREBBP R1446C at the earliest collected time point (AF range 0.019-0.046%) rising to AF 0.082% after clinical diagnosis. Conclusions: Precursor FL mutations are detectable in peripheral blood and saliva years prior to clinical diagnosis with a spectrum of variants enriched in CREBBP and BCL2 and concordant with subsequent FL tumors. Such lesions may assist in stratifying individuals at elevated risk of clinical malignancy, including after identification of pathologic precursors such as ISFN. Figure 1 Figure 1. Disclosures Kurtz: Foresight Diagnostics: Consultancy, Current holder of stock options in a privately-held company; Roche: Consultancy; Genentech: Consultancy. Khodadoust: Alexion, AstraZeneca Rare Disease: Other: Study investigator; CRISPR Therapeutics, Nutcracker Therapeutics: Research Funding; Myeloid Therapeutics: Membership on an entity's Board of Directors or advisory committees. Diehn: Roche: Consultancy; AstraZeneca: Consultancy; RefleXion: Consultancy; BioNTech: Consultancy; Varian Medical Systems: Research Funding; Illumina: Research Funding; CiberMed: Current holder of stock options in a privately-held company, Patents & Royalties; Foresight Diagnostics: Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company. Roulland: BMS: Research Funding. Alizadeh: Bristol Myers Squibb: Research Funding; Gilead: Consultancy; CAPP Medical: Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company; Celgene: Consultancy, Research Funding; Roche: Consultancy, Honoraria; Janssen Oncology: Honoraria; Foresight Diagnostics: Consultancy, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company; Cibermed: Consultancy, Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company; Forty Seven: Current holder of individual stocks in a privately-held company, Current holder of stock options in a privately-held company.
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Van Gils, Julien, Frederique Magdinier, Patricia Fergelot, and Didier Lacombe. "Rubinstein-Taybi Syndrome: A Model of Epigenetic Disorder." Genes 12, no. 7 (June 24, 2021): 968. http://dx.doi.org/10.3390/genes12070968.
Full textAbstract:
The Rubinstein-Taybi syndrome (RSTS) is a rare congenital developmental disorder characterized by a typical facial dysmorphism, distal limb abnormalities, intellectual disability, and many additional phenotypical features. It occurs at between 1/100,000 and 1/125,000 births. Two genes are currently known to cause RSTS, CREBBP and EP300, mutated in around 55% and 8% of clinically diagnosed cases, respectively. To date, 500 pathogenic variants have been reported for the CREBBP gene and 118 for EP300. These two genes encode paralogs acting as lysine acetyltransferase involved in transcriptional regulation and chromatin remodeling with a key role in neuronal plasticity and cognition. Because of the clinical heterogeneity of this syndrome ranging from the typical clinical diagnosis to features overlapping with other Mendelian disorders of the epigenetic machinery, phenotype/genotype correlations remain difficult to establish. In this context, the deciphering of the patho-physiological process underlying these diseases and the definition of a specific episignature will likely improve the diagnostic efficiency but also open novel therapeutic perspectives. This review summarizes the current clinical and molecular knowledge and highlights the epigenetic regulation of RSTS as a model of chromatinopathy.
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Mata-Rocha, Minerva, Angelica Rangel-López, Elva Jiménez-Hernández, Blanca Angélica Morales-Castillo, Carolina González-Torres, Javier Gaytan-Cervantes, Enrique Álvarez-Olmos, et al. "Identification and Characterization of Novel Fusion Genes with Potential Clinical Applications in Mexican Children with Acute Lymphoblastic Leukemia." International Journal of Molecular Sciences 20, no. 10 (May 15, 2019): 2394. http://dx.doi.org/10.3390/ijms20102394.
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Acute lymphoblastic leukemia is the most common type of childhood cancer worldwide. Mexico City has one of the highest incidences and mortality rates of this cancer. It has previously been recognized that chromosomal translocations are important in cancer etiology. Specific fusion genes have been considered as important treatment targets in childhood acute lymphoblastic leukemia (ALL). The present research aimed at the identification and characterization of novel fusion genes with potential clinical implications in Mexican children with acute lymphoblastic leukemia. The RNA-sequencing approach was used. Four fusion genes not previously reported were identified: CREBBP-SRGAP2B, DNAH14-IKZF1, ETV6-SNUPN, ETV6-NUFIP1. Although a fusion gene is not sufficient to cause leukemia, it could be involved in the pathogenesis of the disease. Notably, these new translocations were found in genes encoding for hematopoietic transcription factors which are known to play an important role in leukemogenesis and disease prognosis such as IKZF1, CREBBP, and ETV6. In addition, they may have an impact on the prognosis of Mexican pediatric patients with ALL, with the potential to be included in the current risk stratification schemes or used as therapeutic targets.
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Di Fede, Elisabetta, Valentina Massa, Bartolomeo Augello, Gabriella Squeo, Emanuela Scarano, Anna Maria Perri, Rita Fischetto, et al. "Expanding the phenotype associated to KMT2A variants: overlapping clinical signs between Wiedemann–Steiner and Rubinstein–Taybi syndromes." European Journal of Human Genetics 29, no. 1 (July 8, 2020): 88–98. http://dx.doi.org/10.1038/s41431-020-0679-8.
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AbstractLysine-specific methyltransferase 2A (KMT2A) is responsible for methylation of histone H3 (K4H3me) and contributes to chromatin remodeling, acting as “writer” of the epigenetic machinery. Mutations in KMT2A were first reported in Wiedemann–Steiner syndrome (WDSTS). More recently, KMT2A variants have been described in probands with a specific clinical diagnosis comprised in the so-called chromatinopathies. Such conditions, including WDSTS, are a group of overlapping disorders caused by mutations in genes coding for the epigenetic machinery. Among them, Rubinstein–Taybi syndrome (RSTS) is mainly caused by heterozygous pathogenic variants in CREBBP or EP300. In this work, we used next generation sequencing (either by custom-made panel or by whole exome) to identify alternative causative genes in individuals with a RSTS-like phenotype negative to CREBBP and EP300 mutational screening. In six patients we identified different novel unreported variants in KMT2A gene. The identified variants are de novo in at least four out of six tested individuals and all of them display some typical RSTS phenotypic features but also WDSTS specific signs. This study reinforces the concept that germline variants affecting the epigenetic machinery lead to a shared molecular effect (alteration of the chromatin state) determining superimposable clinical conditions.
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Liu, Xi-yu, and Jiang-feng Xu. "Reduced Histone H3 Acetylation in CD4+T Lymphocytes: Potential Mechanism of Latent Autoimmune Diabetes in Adults." Disease Markers 2015 (2015): 1–7. http://dx.doi.org/10.1155/2015/285125.
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Aims. Latent autoimmune diabetes in adults (LADA) is the result of gene-environment interactions. Histone acetylation regulates gene expression and maybe interpret how environmental factors modify LADA. Hence, we studied the histone acetylation patterns in CD4+T lymphocytes from LADA patients.Methods. Blood CD4+T lymphocytes from 28 patients with LADA and 28 healthy controls were obtained to detect histone H3 acetylation and H4 acetylation. The gene expression of histone acetyltransferases (P300 and CREBBP) and histone deacetylases (HDAC1, HDAC2, and HDAC7) was measured by real-time polymerase chain reaction (RT-PCR).Results. Compared to healthy controls, reduced global H3 acetylation was observed in LADA patients’ CD4+T lymphocytes (P<0.05). Global level of H4 acetylation was not statistically different. Among LADA, CD4+T lymphocytes H3 acetylation was associated with glycosylated hemoglobin (HbA1c) and GADA titer. Compared to healthy controls, the expression of histone acetyltransferases CREBBP in LADA patients was downregulated, and the expression of histone deacetylases HDAC1 and HDAC7 was upregulated.Conclusion. A concerted downregulation of histone H3 acetylation was found in CD4+T lymphocytes of LADA patients, and this might provide evidence of a novel epigenetic explanation for the pathogenesis of LADA and its complications.
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Perveze, I., and R. Hammer. "A Rare Case of Therapy Related Monoblastic Leukemia with t(8;16)(p11.2;p13.3)/KAT6A-CREBBP Fusion and Hemophagocytosis." American Journal of Clinical Pathology 158, Supplement_1 (November 1, 2022): S112—S113. http://dx.doi.org/10.1093/ajcp/aqac126.239.
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Abstract Introduction/Objective Therapy related leukemias are often myeloid and occur as a late complication of cytotoxic therapy or radiotherapy. Most common etiologies are alkylating agents and topoisomerase II inhibitors leading to inaccurate base pairing and chromosomal breakages resulting in complex karyotypes. We describe an extremely rare case of therapy related monobalstic leukemia with t(8;16)(p11.2;p13.3)/KAT6A-CREBBP Fusion and hemophagocytosis. Methods/Case Report An 81-year-old female with history of mixed cellularity Hodgkin’s lymphoma status post chemotherapy with AVD (Adriamycin, Vinblastine and Dacarbazine) and in remission presented with a complaint of newly developed skin rash on her chest. Punch biopsy showed a diffuse infiltrate of blastoid mononuclear cells with a component of monocytic differentiation present throughout the dermis. Complete blood count revealed bicytopenia (Hemoglobin 8.4 g/dl, platelets 32 x 109 per liter) and peripheral blood smear examination showed 54% blasts. Bone marrow biopsy was remarkable for infiltration by 79.2% blasts with monocytic differentiation and prominent hemophagocytosis. Immunophenotypic analysis was notable for weak positivity for p53 and a Ki67 proliferation index of 20%. These results led to the diagnosis of monobasic leukemia with hemophagocytosis. Cytogenetic analysis was abnormal with 19 out of 20 metaphases showing t(8;16)(p11.2;p13.3). Interestingly, FISH revealed the rare KAT6A- CREBBP fusion gene. The patient was hospitalized for induction chemotherapy and started on Decitabine. Her hospital course was complicated with acute kidney injury and pulmonary edema with rapid deterioration along with persistent elevation of blasts in the blood. The patient opted for hospice due to lack of improvement and died thereafter. Results (if a Case Study enter NA) NA. Conclusion AML with t(8,16) is an extremely rare entity accounting for 0.2 to 0.4% of AML cases. AML with t(8,16)/KAT6-CREBBP is frequently associated with monocytic or myelomonocytic differentiation, leukemia cutis and hemophagocytosis. While spontaneous remission can occur in a subset of neonatal t(8;16)(p11.2;p13.3), the majority of cases are therapy-related and carry a poor prognosis as seen in our patient.
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Alari, Valentina, Paolo Scalmani, Paola Francesca Ajmone, Sara Perego, Sabrina Avignone, Ilaria Catusi, Paola Adele Lonati, et al. "Histone Deacetylase Inhibitors Ameliorate Morphological Defects and Hypoexcitability of iPSC-Neurons from Rubinstein-Taybi Patients." International Journal of Molecular Sciences 22, no. 11 (May 28, 2021): 5777. http://dx.doi.org/10.3390/ijms22115777.
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Rubinstein-Taybi syndrome (RSTS) is a rare neurodevelopmental disorder caused by mutations in CREBBP or EP300 genes encoding CBP/p300 lysine acetyltransferases. We investigated the efficacy of the histone deacetylase inhibitor (HDACi) Trichostatin A (TSA) in ameliorating morphological abnormalities of iPSC-derived young neurons from P149 and P34 CREBBP-mutated patients and hypoexcitability of mature neurons from P149. Neural progenitors from both patients’ iPSC lines were cultured one week with TSA 20 nM and, only P149, for 6 weeks with TSA 0.2 nM, in parallel to neural progenitors from controls. Immunofluorescence of MAP2/TUJ1 positive cells using the Skeletonize Image J plugin evidenced that TSA partially rescued reduced nuclear area, and decreased branch length and abnormal end points number of both 45 days patients’ neurons, but did not influence the diminished percentage of their neurons with respect to controls. Patch clamp recordings of TSA-treated post-mitotic P149 neurons showed complete/partial rescue of sodium/potassium currents and significant enhancement of neuron excitability compared to untreated replicas. Correction of abnormalities of P149 young neurons was also affected by valproic acid 1 mM for 72 h, with some variation, with respect to TSA, on the morphological parameter. These findings hold promise for development of an epigenetic therapy to attenuate RSTS patients cognitive impairment.
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Gotti, Giacomo, Maneka Puligandla, Kristen E. Stevenson, Brenton G. Mar, Barbara L. Asselin, Uma Athale, Luis A. Clavell, et al. "Frequency and Prognostic Significance of Recurrent Gene Mutations in Pediatric B-ALL: Report from the DFCI ALL Consortium." Blood 136, Supplement 1 (November 5, 2020): 15–16. http://dx.doi.org/10.1182/blood-2020-136190.
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The application of next-generation sequencing (NGS) approaches to leukemia markedly expanded our understanding of the molecular landscape of pediatric acute lymphoblastic leukemia (ALL), but NGS integration into clinical care and therapeutic decision making is still limited. The prognostic impact of discovered mutations in uniformly treated patients with newly diagnosed B-ALL is not well characterized, and there is no consensus regarding the clinical significance of many of these findings. We investigated the frequency of mutations affecting common molecular pathways in pediatric B-ALL that are potentially druggable with targeted therapies, to define the prognostic role of these mutations, as measured by end-induction minimal residual disease (MRD) and event-free survival (EFS). We analyzed 159 patients (median age 6 years, range 1-18) with newly diagnosed Ph-negative B-ALL treated between 2005-2015 according to the Dana-Farber Cancer Institute (DFCI) ALL Consortium Protocols 05-001 and 11-001. Diagnostic leukemia samples were sequenced using a validated clinical NGS panel. We focused on mutations affecting the following pathways: Ras (NRAS, KRAS, PTPN11, NF1 and BRAF), cell cycle regulation (CCND3, CDKN2A/B and RB1), PI3K signaling (PTEN, PIK3CA, PIK3R1, PIK3C2B, MTOR, TSC1 and TSC2), Polycomb repressive complex 2 (PRC2) (EED, EZH2 and SUZ12), and JAK/STAT signaling (CRLF2, JAK2, JAK3, MPL, SH2B3 and SOCS1). Additionally, we analyzed mutations affecting CREBBP, FLT3, PAX5, SETD2 and TP53. Mutations occurring in the ExAC database at a frequency greater than 0.01% were excluded. A Fisher exact test and Wilcoxon rank sum test were used for categorical and continuous variables. High end-induction MRD, defined as > 10-3, was assessed by an Ig-TCR PCR assay. EFS was estimated using Kaplan and Meier method and tested between groups using a log-rank test. Induction death/failure, relapse or death were considered as events. Overall, 108 of the 159 patients (68%) carried at least one mutation in the studied genes. Most common mutations were in the Ras pathway (47%), CREBBP (9%), JAK pathway (8%), FLT3 (8%), PI3K pathway (5%), PAX5 (4%), SETD2 (4%), TP53 (4%), cell cycle regulation (3%) and PRC2 complex (3%). We investigated the distribution of mutations among common cytogenetic groups: ETV6-RUNX1, high hyperdiploidy (HHD) (51-65 chromosomes), hypodiploidy (<45 chromosomes), KMT2A-rearranged, TCF3-PBX1, and intra-amplification chromosome 21 (iAMP21). There was a strong association between TP53 and hypodiploidy (43% vs 2%; p=0.001). Ras pathway and FLT3 mutations were enriched in HHD (67% vs 39%; p=0.004) and (17% vs 4%; p=0.016), respectively. FLT3 mutations were mutually exclusive with ETV6-RUNX1 (12% vs. 0%, p=0.038), and Ras pathway mutations were rare in this subgroup (p=0.018). PAX5 mutations were enriched in children >10 years of age (p=0.035). There were no associations with sex. Ras pathway mutations were associated with high end-induction MRD (68% vs 42%; p=0.045), and this association was stronger for clonal (variant allele frequency (VAF) >25%) mutations (n=33) (63% vs 19%; p=0.0002). Focusing on HHD, CREBBP mutations frequently co-occurred with clonal Ras pathway mutations (83% vs 17%; p=0.032). Among HHD patients with evaluable MRD (n=33), 7 patients had high MRD, and 3 of these had CREBBP mutations (p=0.052). The analyzed cohort was enriched for higher risk ALL disease (Table 1). The 5-year EFS was 80%±3% among these 159 patients. Overall, no EFS difference was observed based on Ras pathway (p=0.35). Among HHD ALL, the presence of CREBBP or clonal Ras pathway mutations was significantly associated with inferior 5-yr EFS (50%±20% vs 97%±3%; p=0.0006, Figure 1) and (70%±13% vs 100%; p=0.007, Figure 2), respectively. In conclusion, our findings provide insight into the prognostic significance of the most common mutations in pediatric HHD B-ALL in a uniformly treated cohort of patients as part of the DFCI ALL Consortium. The presence of CREBBP and clonal Ras pathway mutations may be associated with upfront chemotherapy resistance as demonstrated by high end-induction MRD. Further analysis of Ras pathway mutations segregated by VAF is warranted. Future trials may integrate these findings into risk stratification of HHD ALL. With prospective continued clinical use of NGS assays, we will further clarify the role of mutations and their contribution to disease outcomes in B-ALL. Disclosures Mar: Blueprint Medicines Corporation: Current Employment, Current equity holder in publicly-traded company. Stegmaier:Novartis: Research Funding; Auron Therapeutics: Consultancy. Silverman:Takeda: Other: advisory board; Servier: Other: advisory board; Syndax: Other: advisory board.
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Sadeghi, Hossein, Sahra Esmkhani, Reihaneh Pirjani, Mona Amin-Beidokhti, Milad Gholami, Ghasem Azizi Tabesh, Mohammad Reza Ghasemi, Latif Gachkar, and Reza Mirfakhraie. "CREB-binding protein (CREBBP) and preeclampsia: a new promising target gene." Molecular Biology Reports 48, no. 3 (February 24, 2021): 2117–22. http://dx.doi.org/10.1007/s11033-021-06215-1.
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Yumul, Rhea Camille R., and Mary Anne D. Chiong. "Rubinstein–Taybi Syndrome in a Filipino Infant with a Novel CREBBP Gene Pathogenic Variant." Case Reports in Genetics 2022 (May 21, 2022): 1–5. http://dx.doi.org/10.1155/2022/3388879.
Full textAbstract:
Rubinstein–Taybi syndrome (RSTS) is a rare genetic disorder characterized by dysmorphic facial features, broad thumbs and halluces, intellectual disability, and postnatal growth retardation. This report presents a male infant with microcephaly and characteristic facial features, namely, low anterior hairline, hirsutism, thin upper lip and micrognathia, broad thumbs and first toes, cryptorchidism, recurrent pneumonia, developmental delay, and growth retardation. Genetic testing showed a novel pathogenic variant in the CREBBP gene which is consistent with the clinical diagnosis of RSTS.
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Dutto, Ilaria, Claudia Scalera, Micol Tillhon, Giulio Ticli, Gianluca Passaniti, Ornella Cazzalini, Monica Savio, et al. "Mutations in CREBBP and EP300 genes affect DNA repair of oxidative damage in Rubinstein-Taybi syndrome cells." Carcinogenesis 41, no. 3 (August 29, 2019): 257–66. http://dx.doi.org/10.1093/carcin/bgz149.
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Abstract Rubinstein-Taybi syndrome (RSTS) is an autosomal-dominant disorder characterized by intellectual disability, skeletal abnormalities, growth deficiency and an increased risk of tumors. RSTS is predominantly caused by mutations in CREBBP or EP300 genes encoding for CBP and p300 proteins, two lysine acetyl-transferases (KAT) playing a key role in transcription, cell proliferation and DNA repair. However, the efficiency of these processes in RSTS cells is still largely unknown. Here, we have investigated whether pathways involved in the maintenance of genome stability are affected in lymphoblastoid cell lines (LCLs) obtained from RSTS patients with mutations in CREBBP or in EP300 genes. We report that RSTS LCLs with mutations affecting CBP or p300 protein levels or KAT activity, are more sensitive to oxidative DNA damage and exhibit defective base excision repair (BER). We have found reduced OGG1 DNA glycosylase activity in RSTS compared to control cell extracts, and concomitant lower OGG1 acetylation levels, thereby impairing the initiation of the BER process. In addition, we report reduced acetylation of other BER factors, such as DNA polymerase β and Proliferating Cell Nuclear Antigen (PCNA), together with acetylation of histone H3. We also show that complementation of CBP or p300 partially reversed RSTS cell sensitivity to DNA damage. These results disclose a mechanism of defective DNA repair as a source of genome instability in RSTS cells.
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Mensah, Afua Adjeiwaa, Sergio Valente, Milos Matkovic, Giulio Sartori, Chiara Falzarano, Chiara Tarantelli, Luciano Cascione, et al. "Abstract 3279: Dual inhibition of EZH2 and histone deacetylases for the treatment of lymphomas with epigenetic aberrations." Cancer Research 82, no. 12_Supplement (June 15, 2022): 3279. http://dx.doi.org/10.1158/1538-7445.am2022-3279.
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Abstract Background: Lymphomas are characterized by aberrations in epigenetic proteins that contribute to establishing and maintaining the malignant phenotype. Gain of function mutations in EZH2 histone methyltransferase and inactivating mutations in CREBBP acetyltransferase occur in up to 30% of diffuse large B cell lymphomas (DLBCLs). Half of DLBCLs with mutated EZH2 have also mutated CREBBP. In vitro, mutated EZH2 DLBCLs have a lower sensitivity to HDAC inhibitors, HDACi (Mensah et al, 2021). Here, we explored the dual pharmacological inhibition of EZH2 and HDAC for an improved anti-lymphoma activity. Methods: Biochemical evaluation and characterization of target engagement were done using fluorescence polarization, thermal shift, surface plasmon resonance, isothermal titration calorimetry and microscale thermophoresis. Computer modelling was performed using available EZH2 and HDACs crystal structures. Anti-proliferative activity was assessed after 7 days (d) using MTT and live imaging in DLBCL cells (n = 5). For cell cycle analysis by flow cytometry, cells were treated, fixed then stained with 7-AAD. Results: We designed and synthesized 2 dual EZH2/HDAC inhibitors, MC4343 and MC4355, starting from the structures of EZH2 inhibitor tazemetostat (taz) and HDACi vorinostat. In biochemical assays, MC4343 and MC4355 had equal potencies towards EZH2 (0.032 nM) but different specificities towards class I and class II HDACs: MC4355 showed 7.5-fold greater inhibition of HDAC3 compared to MC4343 (0.38 µM and 2.85 µM, respectively) and more potently inhibited HDACs 6 and 8 (0.016, 0.17 µM and no activity, respectively). Computational modelling showed that the coordinative functional group of MC4343, but not of MC4355, caused steric clashes with several HDACs in increasing order: HDAC1 = HDAC3 > HDAC8 >> HDAC4 = HDAC6. These results closely mirrored those obtained from the biochemical analysis. Both compounds inhibited proliferation in DLBCL cell lines irrespective of EZH2 or CREBBP mutational status but EZH2 and/or CREBBP mutants were more sensitive. MC4355 (IC50 range = 0.17 - 1.68 µM; median = 0.2 µM) was more potent than MC4343 (IC50 range = 0.17-2.72 µM; median = 1.78 µM). This was confirmed by live imaging analyses. SUDHL4, with EZH2 Y666N, showed poorer sensitivity to both inhibitors compared to DLBCLs with EZH2 Y646N/S. MC4343 and MC4355 induced cell death and G1 arrest in a dose-dependent manner. Pfeiffer, KARPAS422, WSUDLCL2, most sensitive to taz alone (IC50 = 4, 16, 77 nM), were most sensitive to MC4355 (IC50 = 200, 174, 171 nM). Notably, dual inhibitor treatment of Toledo and SUDHL4 with low sensitivity to taz (IC50 = 9, 14 µM), partially rescued sensitivity (IC50 = 1.7, 1.6 µM). Conclusions: We designed and synthesized 2 novel dual EZH2/HDAC inhibitors, MC4343 and MC4355, with robust anti-proliferative effects in DLBCL. Our data show the efficacy of this novel class of epigenetic agents in lymphomas. Citation Format: Afua Adjeiwaa Mensah, Sergio Valente, Milos Matkovic, Giulio Sartori, Chiara Falzarano, Chiara Tarantelli, Luciano Cascione, Stefano A. Pileri, Andrea Cavalli, Antonello Mai, Francesco Bertoni. Dual inhibition of EZH2 and histone deacetylases for the treatment of lymphomas with epigenetic aberrations [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 3279.
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Mullighan, Charles G., Jinghui Zhang, Lawryn H. Kasper, Stephanie Lerach, Debbie Payne-Turner, Letha A. Phillips, Sue L. Heatley, et al. "CREBBP mutations in relapsed acute lymphoblastic leukaemia." Nature 471, no. 7337 (March 2011): 235–39. http://dx.doi.org/10.1038/nature09727.
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