Journal articles on the topic 'Inhibitor Activity'
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Sumanadasa, Subathdrage D. M., Christopher D. Goodman, Andrew J. Lucke, Tina Skinner-Adams, Ishani Sahama, Ashraful Haque, Tram Anh Do, Geoffrey I. McFadden, David P. Fairlie, and Katherine T. Andrews. "Antimalarial Activity of the Anticancer Histone Deacetylase Inhibitor SB939." Antimicrobial Agents and Chemotherapy 56, no. 7 (April 16, 2012): 3849–56. http://dx.doi.org/10.1128/aac.00030-12.
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ABSTRACTHistone deacetylase (HDAC) enzymes posttranslationally modify lysines on histone and nonhistone proteins and play crucial roles in epigenetic regulation and other important cellular processes. HDAC inhibitors (e.g., suberoylanilide hydroxamic acid [SAHA; also known as vorinostat]) are used clinically to treat some cancers and are under investigation for use against many other diseases. Development of new HDAC inhibitors for noncancer indications has the potential to be accelerated by piggybacking onto cancer studies, as several HDAC inhibitors have undergone or are undergoing clinical trials. One such compound, SB939, is a new orally active hydroxamate-based HDAC inhibitor with an improved pharmacokinetic profile compared to that of SAHA. In this study, thein vitroandin vivoantiplasmodial activities of SB939 were investigated. SB939 was found to be a potent inhibitor of the growth ofPlasmodium falciparumasexual-stage parasitesin vitro(50% inhibitory concentration [IC50], 100 to 200 nM), causing hyperacetylation of parasite histone and nonhistone proteins. In combination with the aspartic protease inhibitor lopinavir, SB939 displayed additive activity. SB939 also potently inhibited thein vitrogrowth of exoerythrocytic-stagePlasmodiumparasites in liver cells (IC50, ∼150 nM), suggesting that inhibitor targeting to multiple malaria parasite life cycle stages may be possible. In an experimentalin vivomurine model of cerebral malaria, orally administered SB939 significantly inhibitedP. bergheiANKA parasite growth, preventing development of cerebral malaria-like symptoms. These results identify SB939 as a potent new antimalarial HDAC inhibitor and underscore the potential of investigating next-generation anticancer HDAC inhibitors as prospective new drug leads for treatment of malaria.
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Böhm, Martina, Manuela Krause, Charis Von Auer, Wolfgang Miesbach, and Inge Scharrer. "The Frequency and the Significance of ADAMTS-13 Neutralising Inhibitors in 62 Patients with Non-Familial Thrombotic Thrombocytopenic Purpura." Blood 104, no. 11 (November 16, 2004): 3945. http://dx.doi.org/10.1182/blood.v104.11.3945.3945.
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Abstract Neutralising inhibitors against ADAMTS-13 are detected in 51–67% of patients with Thrombotic Thrombocytopenic Purpura (TTP). These ADAMTS-13 inhibitors have not been very well characterised and the diagnostic or the prognostic value of these inhibitors is not established. In the present study, we measured ADAMTS-13 activity and the corresponding inhibitor titer in 96 samples from 62 patients with TTP at various stages of their disease. All patients presented with non-familial TTP. For patients with severe ADAMTS-13 activity without detectable inhibitor heritable ADAMTS-13 deficiency was excluded by either normalisation of ADAMTS-13 activity in remission or by detecting normal ADAMTS-13 activity in first degree family members. ADAMTS-13 activity was quantified by measuring the residual ristocetin cofactor activity of the substrate. The inhibitor against ADAMTS-13 was detected by mixing patient plasma, either neat or diluted, with normal plasma. The inhibitor concentration neutralising 50% of ADAMTS-13 activity in a 1:1 dilution of patient plasma with normal plasma was defined as 1 U/ml. Inhibitors were considered non-detectable (<0.4 U/ml), if residual ADAMTS-13 activity in the mixture was higher than 75%. Samples with ADAMTS-13 activity >6.25% were heat-inactivated (30 min at 56°C) before testing for inhibitory activity. We found severe ADAMTS-13 deficiency in 89% (24/27) of the samples from patients with acute untreated TTP. 87% (21/24) of these samples were positive for inhibitory activity. The inhibitor titer ranged from 0.4 to 62 U/ml with a median of 1 U/ml. One patient with acute TTP demonstrated ADAMTS-13 activity of 34% despite an inhibitor titer of 0.6 U/ml. The sensitivity of a positive inhibitor test for the diagnosis of TTP was thus 82%. The inhibitor titer before initiation of therapy could not be correlated with the platelet count, the CRP-level or the response to PE-therapy, if patients with an index episode and patients with a relapse were analysed separately. Severe ADAMTS-13 activity was detected in 15/31 samples collected from patients during plasma exchange therapy. 93% (14/15) of these samples were positive for an inhibitor with a titer ranging between 0.6 and 47 U/ml (median: 3 U/ml). 12 of 37 patients tested in remission presented with severe ADAMTS-13 deficiency, 6 of them were positive for inhibitory activity (range: 1–52 U/ml; median: 5 U/ml). The inhibitor titer for patients, which were analysed during acute untreated TTP as well as in remission (n=5), was notable not related to the stage of disease. Five patients with positive inhibitory activity at admission demonstrated mild ADAMTS-13 deficiency in remission without detectable inhibitor. In contrast, we detected inhibitory activity of 0,6–0,8 U/ml in 3 samples from two patients with measurable ADAMTS-13 activity. These low titer inhibitors were only detectable, if samples were heat inactivated before performing the inhibitor assay. Our data demonstrate, that inhibitors against ADAMTS-13 are very heterogeneous. It is highly suspected, that some of these inhibitors can either completely or partly neutralise ADAMTS-13 function in vivo without being detectable in vitro. Inhibitors against ADAMTS-13 might, on the other side, not always completely inhibit ADAMTS-13 function, since they can occur in patients with high residual ADAMTS-13 activity. Inhibitor titers show a wide variation and the clinical significance of the inhibitor titer before, during and after therapy needs to be further investigated.
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Aghaali, Negar, Mohammad Ghadamyari, Vahid Hosseininaveh, and Nasir Saberi Riseh. "PROTEASE INHIBITOR FROM THE CRUDE EXTRACT OF PLANT SEEDS AFFECTS THE DIGESTIVE PROTEASES IN HYPHANTRIA CUNEA (LEP.: ARCTIIDAE)." Journal of Plant Protection Research 53, no. 4 (October 1, 2013): 338–46. http://dx.doi.org/10.2478/jppr-2013-0051.
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Abstract Proteases are one of the most important digestive enzymes in the midgut of Hyphantria cunea Drury. Proteases are responsible for protein digestion. In the present study, we evaluated the efficiency of some plant inhibitors on proteases in the gut of the H. cunea. Last instar larvae were collected from mulberry trees. The digestive system of the larvae was used as an enzyme source. The total proteolytic and trypsin activity were assessed by the hemoglobin and BApNA, respectively, as the substrate. The evaluation of the total proteolytic and trypsin activities in various pHs showed the highest relative activity at a pH of 11. Also, the inhibitory effect of inhibitors extracted from Alhagi maurorum Medik., Lathyrus sativus L., Vicia faba L., Prosopis farcta (Banks & Sol.) Eig., and Panicum miliaceum L. on the digestive protease of the fall webworm was measured. Protease inhibitors extracted from A. maurorum, P. farcta and P. miliaceum showed negligible inhibition but L. sativus was able to inhibit 34.72% and 100% of the total activity of proteolytic and trypsin, respectively. Also, the total proteolytic and trypsin activities were inhibited by the inhibitor from V. faba, at 22.27% and 100%, respectively. The zymogram pattern of trypsin with nitro-cellulose membranes showed 2 isoforms in the gut of H. cunea. The inhibitor from L. sativus completely inhibited both isoforms. Gel electrophoresis of proteolitytic activity revealed at least 6 isoforms the inhibitor extracted from L. sativus; completely inhibiting some of them. The inhibitor from L. sativus was purified by ammonium sulfate precipitation and gel-filtration. The molecular mass of the inhibitor was determined as 45 kDa. The highest inhibition of trypsin activity by the inhibitor from L. sativus occurred at a pH of 10. The stability of the inhibitor from L. sativus was evaluated at different pHs and temperatures. The results showed that the inhibitor from L. sativus was stable at a pH of 11.0, and showed 45% inhibition on trypsin activity at a pH of 11. Also, this inhibitor revealed stability up to 50°C.
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Jang, Seong, Bill Strickland, Lynda Finis, Jeffrey J. Koojiman, Janneke J. Melis, Guido J. Zaman, and Jan V. Tornout. "Abstract 4014: Comparative biochemical kinase activity analysis identifies rivoceranib as the most selective VEGFR-2 inhibitor compared with other TKIs with known activity against VEGFR-2." Cancer Research 83, no. 7_Supplement (April 4, 2023): 4014. http://dx.doi.org/10.1158/1538-7445.am2023-4014.
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Abstract Introduction: Vascular endothelial growth factor receptor 2 (VEGFR-2) is a key regulator of tumor angiogenesis that is highly expressed in several tumor types and is a known target for anti-cancer therapy. Yet the clinical use of VEGFR-2 inhibitors has been challenged by limited efficacy and various side effects, potentially due to the low selectivity of these TKIs for VEGFR-2. Thus, potent VEGFR-2 inhibitors with improved selectivity are needed. Rivoceranib is an oral tyrosine kinase inhibitor (TKI) that potently and selectively inhibits VEGFR-2. A comparison of the potency and selectivity of VEGFR-2 inhibitors can provide a rationale for selecting a specific TKI for anticancer therapy in the clinic. Methods: Binding of rivoceranib to VEGFR-2 was determined on a Biacore T200. The affinity constant (KD) was derived from the association and dissociation rate constants. Inhibitory potency of rivoceranib and 10 FDA-approved reference inhibitors on kinase enzyme activity was determined using mobility shift assays (MSA) or immobilized metal ion affinity particle (IMAP) assays. The half-maximum inhibitory activity (IC50) of the 11 inhibitors on VEGFR-2 was determined in 10-point dose-response curves. The selectivity of the inhibitors was determined on 270 wild-type kinases at a fixed concentration of each inhibitor. Rivoceranib was tested at 10- and 100-times IC50 (160 nmol/L and 1.6 µmol/L). Reference inhibitors were tested at 1 µmol/L (35 to 1056-times IC50). Results: Rivoceranib had a KD of 3 nmol/L on VEGFR-2. In enzyme activity assays, rivoceranib had intermediate potency compared with the 10 reference inhibitors, with a VEGFR-2 kinase inhibition IC50 value of 16 nmol/L. Analysis of the residual activity of the panel of 270 kinases in the presence of rivoceranib or the reference inhibitors showed wide variation in selectivity for VEGFR-2, with rivoceranib identified as the most selective inhibitor (activity of 16 additional kinases inhibited by >50% at 1.6 µmol/L). Tivozanib, the most potent VEGFR-2 inhibitor, displayed greater than 50% inhibitory activity against more than 70 additional kinases. Sunitinib was identified as the least selective inhibitor included in this study, inhibiting 125 additional kinases by >50%. Conclusion: Variations in selectivity among TKIs with similar anti-VEGFR-2 potency can help explain differences in their clinical toxicity profiles, which may be partially due to variant inhibitory effects against TKIs other than VEGFR-2. This comparative biochemical analysis highlights the potential for rivoceranib to address clinical limitations associated with the poor selectivity of currently available VEGFR-2 inhibitors. Rivoceranib is under ongoing investigation as monotherapy and in combination with chemotherapy in various tumor types. Citation Format: Seong Jang, Bill Strickland, Lynda Finis, Jeffrey J. Koojiman, Janneke J. Melis, Guido J. Zaman, Jan V. Tornout. Comparative biochemical kinase activity analysis identifies rivoceranib as the most selective VEGFR-2 inhibitor compared with other TKIs with known activity against VEGFR-2. [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 4014.
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Li, Qi, Hongyu Yang, Jun Mo, Yao Chen, Yue Wu, Chen Kang, Yuan Sun, and Haopeng Sun. "Identification by shape-based virtual screening and evaluation of new tyrosinase inhibitors." PeerJ 6 (January 26, 2018): e4206. http://dx.doi.org/10.7717/peerj.4206.
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Targeting tyrosinase is considered to be an effective way to control the production of melanin. Tyrosinase inhibitor is anticipated to provide new therapy to prevent skin pigmentation, melanoma and neurodegenerative diseases. Herein, we report our results in identifying new tyrosinase inhibitors. The shape-based virtual screening was performed to discover new tyrosinase inhibitors. Thirteen potential hits derived from virtual screening were tested by biological determinations. Compound 5186-0429 exhibited the most potent inhibitory activity. It dose-dependently inhibited the activity of tyrosinase, with the IC50 values 6.2 ± 2.0 µM and 10.3 ± 5.4 µM on tyrosine and L-Dopa formation, respectively. The kinetic study of 5186-0429 demonstrated that this compound acted as a competitive inhibitor. We believe the discoveries here could serve as a good starting point for further design of potent tyrosinase inhibitor.
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Sato, Shun, Kana Yamamoto, Moeno Ito, Katsutoshi Nishino, Takanao Otsuka, Kazuhiro Irie, and Masaya Nagao. "Enhancement of Inhibitory Activity by Combining Allosteric Inhibitors Putatively Binding to Different Allosteric Sites on Cathepsin K." Molecules 28, no. 10 (May 19, 2023): 4197. http://dx.doi.org/10.3390/molecules28104197.
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Background: Cathepsin K, which is involved in bone resorption, is a good target for treating osteoporosis, but no clinically approved medicine has been developed. Recently, allosteric inhibitors with high specificity and few side effects have been attracting attention for use in new medicines. Methods: Cathepsin K inhibitors were isolated from the methanol extract of Chamaecrista nomame (Leguminosae) using cathepsin K inhibition activity-assisted multi-step chromatography. Standard kinetic analysis was employed to examine the mechanism of cathepsin K inhibition when an isolated inhibitor and its derivative were used. The allosteric binding of these cathepsin K inhibitors was supported by a docking study using AutoDock vina. Combinations of allosteric cathepsin K inhibitors expected to bind to different allosteric sites were examined by means of cathepsin K inhibition assay. Results: Two types of cathepsin K inhibitors were identified in the methanol extract of Chamaecrista nomame. One type consisted of cassiaoccidentalin B and torachrysone 8-β-gentiobioside, and inhibited both cathepsin K and B with similar inhibitory potential, while the other type of inhibitor consisted of pheophytin a, and inhibited cathepsin K but not cathepsin B, suggesting that pheophytin a binds to an allosteric site of cathepsin K. Kinetic analysis of inhibitory activity suggested that pheophytin a and its derivative, pheophorbide b, bind allosterically to cathepsin K. This possibility was supported by a docking study on cathepsin K. The cathepsin K inhibitory activity of pheophytin a and pheophorbide b was enhanced by combining them with the allosteric inhibitors NSC 13345 and NSC94914, which bind to other allosteric sites on cathepsin K. Conclusions: Different allosteric inhibitors that bind to different sites in combination, as shown in this study, may be useful for designing new allosteric inhibitory drugs with high specificity and few side effects.
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Pintus, Francesca, Delia Spanò, Angela Corona, and Rosaria Medda. "Antityrosinase activity ofEuphorbia characiasextracts." PeerJ 3 (October 13, 2015): e1305. http://dx.doi.org/10.7717/peerj.1305.
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Tyrosinase is a well-known key enzyme in melanin biosynthesis and its inhibitors have become increasingly important because of their potential use as hypopigmenting agents. In the present study, the anti-melanogenic effect of aqueous and ethanolic extracts fromEuphorbia characiasleaves, stems, and flowers in cell-free and cellular systems was examined. All the extracts showed inhibitory effects against mushroom tyrosinase with leaf extracts exhibiting the lowest IC50values of 24 and 97 µg/mL for aqueous and ethanolic extracts respectively. Enzyme kinetic analysis indicated that leaf aqueous extract acts as a mixed type inhibitor, while ethanolic extract shows a competitive inhibition effect on mushroom tyrosinase using L-DOPA as substrate. In addition, the inhibitory effect of leaf extracts on tyrosinase activity and melanin production was examined in murine melanoma B16F10 cells. Cellular tyrosinase activity as well as levels of melanin synthesis are reduced in a dose-dependent manner by extracts in cells treated withα-melanocyte stimulating hormone (α-MSH). The effects are comparable, and sometimes even better, than that of kojic acid, a well known tyrosinase inhibitor used for reference. All these results suggest thatE. characiascould be a great source of the natural inhibitors from tyrosinase and has the potential to be used as a whitening agent in therapeutic fields.
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Hs, Ranjini, Padmanabha Udupa Eg, Shobha U. Kamath, Manjunath Setty, Basavaraj Hadapad, and Asha Kamath. "AN IN VITRO STUDY OF CINNAMOMUM ZEYLANICUM AS NATURAL INHIBITOR OF ANGIOTENSIN-CONVERTING ENZYME (ACE) ON SHEEP (OVIS ARIES) TISSUES." Asian Journal of Pharmaceutical and Clinical Research 9, no. 5 (September 1, 2016): 249. http://dx.doi.org/10.22159/ajpcr.2016.v9i5.13424.
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ABSTRACTObjective: The present study was aimed to find the angiotensin-converting enzyme (ACE) inhibitory activity using the methanolic extract ofCinnamomum zeylanicum (as a natural inhibitor) on sheep tissues as the enzyme source.Methods: Hippuryl-histidyl-leucine (HHL) as a substrate, tissue ACE activity was measured spectrophotometrically at 228 nm. For an incubationperiod of 30 minutes at 37°C, the linearity of ACE activity of kidney, lung, and testis enzyme was established. A known medicinal plant C. zeylanicumwas used as natural inhibitor of ACE. In this enzyme assay, inhibitory effect of methanolic extract of C. zeylanicum on kidney, lung and testicular ACEwas determined. ACE activity was confirmed by captopril, a standard inhibitor of ACE.Results: In the presence of a methanolic extract of C. zeylanicum (10:1), ACE activity was determined and this has inhibited ACE activity verysignificantly. C. zeylanicum leaves extract has reduced sheep kidney, lung, and testis ACE activity by 70.06%, 12.63%, and 20.23%, respectively.Conclusion: Significant inhibition was observed in the kidney ACE than in lung and testis ACE activity. This can propose that there may be a possiblerole in controlling blood pressure or reduction in cardiovascular diseases. Some plants with the great medicinal property may be considered aspromising sources of natural inhibitors of ACE for medicine and commercial uses. This comprehensive study may show numerous beneficial effects asa potential therapeutic agent for lowering blood pressure.Keywords: Angiotensin-converting enzyme, Natural angiotensin-converting enzyme inhibitor, Kinetic assay, Hippuryl-histidyl-leucine, Cinnamomumzeylanicum, Cardiovascular diseases.
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CHEN, Ching-Jiunn, Hui-Sheng HUANG, Yu-Tsong LEE, Chia-Yi YANG, and Wen-Chang CHANG. "Characterization and purification of a lipoxygenase inhibitor in human epidermoid carcinoma A431 cells." Biochemical Journal 327, no. 1 (October 1, 1997): 193–98. http://dx.doi.org/10.1042/bj3270193.
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A lipoxygenase inhibitor in the cytosolic fraction of human epidermoid carcinoma A431 cells was characterized and purified. The cytosolic inhibitor lost the inhibitory activity upon heating at 75 °C for 15 min or pretreating with 1 mg/ml trypsin at 37 °C for 60 min. Cytosol, after dialysis, lost the inhibitory activity but its inhibitory activity recovered when 1 mM GSH was added to the dialysate. The inhibitory activity of cytosol was also abolished by treatment either with 1 mM iodoacetate at 4 °C for 1 h or with 0.5 mM H2O2. The pI of the inhibitor was approx. 7.0. In addition to 12-lipoxygenase, the inhibitor inhibited the activities of 5-lipoxygenase and fatty acid cyclo-oxygenase in a cell-free system. The inhibitor was purified by a series of column chromatographies using CM Sephadex C-50, Sephadex G-100 SF and Mono P columns. A major 22 kDa protein was obtained that was distinct from selenium-dependent glutathione peroxidase.
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Miao, Yuxi, Guanzhu Chen, Xinping Xi, Chengbang Ma, Lei Wang, James F. Burrows, Jinao Duan, Mei Zhou, and Tianbao Chen. "Discovery and Rational Design of a Novel Bowman-Birk Related Protease Inhibitor." Biomolecules 9, no. 7 (July 14, 2019): 280. http://dx.doi.org/10.3390/biom9070280.
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Anuran amphibian skin secretions are a rich source of peptides, many of which represent novel protease inhibitors and can potentially act as a source for protease inhibitor drug discovery. In this study, a novel bioactive Bowman-Birk type inhibitory hexadecapeptide of the Ranacyclin family from the defensive skin secretion of the Fukien gold-striped pond frog, Pelophlax plancyi fukienesis, was successfully isolated and identified, named PPF-BBI. The primary structure of the biosynthetic precursor was deduced from a cDNA sequence cloned from a skin-derived cDNA library, which contains a consensus motif representative of the Bowman-Birk type inhibitor. The peptide was chemically synthesized and displayed a potent inhibitory activity against trypsin (Ki of 0.17 µM), as well as an inhibitory activity against tryptase (Ki of 30.73 µM). A number of analogues of this peptide were produced by rational design. An analogue, which substituted the lysine (K) at the predicted P1 position with phenylalanine (F), exhibited a potent chymotrypsin inhibitory activity (Ki of 0.851 µM). Alternatively, a more potent protease inhibitory activity, as well as antimicrobial activity, was observed when P16 was replaced by lysine, forming K16-PPF-BBI. The addition of the cell-penetrating peptide Tat with a trypsin inhibitory loop resulted in a peptide with a selective inhibitory activity toward trypsin, as well as a strong antifungal activity. This peptide also inhibited the growth of two lung cancer cells, H460 and H157, demonstrating that the targeted modifications of this peptide could effectively and efficiently alter its bioactivity.
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Domoney, C., and T. Welham. "Trypsin inhibitors in Pisum: variation in amount and pattern of accumulation in developing seed." Seed Science Research 2, no. 3 (September 1992): 147–54. http://dx.doi.org/10.1017/s0960258500001276.
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AbstractA survey of Pisum genotypes for seed trypsin inhibitors revealed a tenfold range in the extent of inhibition. Approximately 90% of trypsin inhibitory activity was associated with two albumin fractions in selected variant lines. The differences among extreme variants were consistent in three environments, between two sources of trypsin tested and whether expressed on a unit protein or dry weight basis.A study of the appearance of trypsin inhibitors during seed development in selected highand low-inhibitor lines showed differences in the accumulation pattern of active inhibitors. An endogenous protease was identified in Pisum seed protein preparations, whose in vitro trypsin-like activity was predominant in protein from early stages of seed development, when little or no trypsin inhibitor was present. However, there was no correlation between the amount of this protease and the extent of trypsin inhibitory activity in lines that varied for inhibitor content.
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KUMAR, Priyadarsini, and Donal A. WALSH. "A dual-specificity isoform of the protein kinase inhibitor PKI produced by alternate gene splicing." Biochemical Journal 362, no. 3 (March 8, 2002): 533–37. http://dx.doi.org/10.1042/bj3620533.
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We have previously shown that the protein kinase inhibitor β (PKIβ) form of the cAMP-dependent protein kinase inhibitor exists in multiple isoforms, some of which are specific inhibitors of the cAMP-dependent protein kinase, whereas others also inhibit the cGMP-dependent enzyme [Kumar, Van Patten and Walsh (1997), J. Biol. Chem. 272, 20011–20020]. We have now demonstrated that the switch from a cAMP-dependent protein kinase (PKA)-specific inhibitor to one with dual specificity arises as a consequence of alternate gene splicing. We have confirmed using bacterially produced pure protein that a single inhibitor species has dual specificity for both PKA and cGMP-dependent protein kinase (PKG), inhibiting each with very high and closely similar inhibitory potencies. The gene splicing converted a protein with 70 amino acids into one of 109 amino acids, and did not change the inhibitory potency to PKA, but changed it from a protein that had no detectable PKG inhibitory activity to one that now inhibited PKG in the nanomolar range.
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Tsujimura, Akane, Yukimasa Shiotsu, Hitoshi Kiyoi, Yuichi Ishikawa, Hiroshi Ishida, Tsutomu Toki, Etsuro Ito, and Tomoki Naoe. "Anti–leukemia Activity of a Novel KIT Selective Inhibitor KI–328." Blood 114, no. 22 (November 20, 2009): 4146. http://dx.doi.org/10.1182/blood.v114.22.4146.4146.
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Abstract Abstract 4146 Introduction KIT is a type III receptor tyrosine kinase together with FLT3, PDGFR and FMS. The interaction of KIT and its ligand stem cell factor (SCF) plays an important role in the cell survival, proliferation and differentiation. Activating mutations of KIT have been demonstrated in several kinds of human malignancies, such as mastocytoma, gastrointestinal stromal tumor, and acute myeloid leukemia (AML). Since KIT mutation seems a poor prognostic factor in CBF leukemia and the KIT expression is observed in most AML cells, KIT serves a molecular target for the treatment of AML. To date, several small molecules have been demonstrated to have a potency against KIT kinase, while KIT selective inhibitors are not yet developed for the clinical use. We recently developed a novel KIT selective inhibitor KI-328, and evaluated here its inhibitory effect on wild-type (Wt) and mutant KIT kinases. Methods We identified 5 types of KIT mutations (D816V, M541L, V540L, T417F/del418-419 and N822K) in AML cells, and established these mutant-KIT, as well as Wt-KIT, expressing IL-3-dependent mouse myeloid precursor 32D cells. Using these Wt- and mutant KIT expressing 32D cells, we examined the anti-leukemia activity of KI-328 in comparison with another potent KIT inhibitors. Results In Wt- and M541L-KIT expressing cells, KITs were phosphorylated by the SCF stimulation. In contrast, mutant KITs were constitutively phosphorylated in D816V-, V540L-, T417F-, and N822K-KIT expressing cells. However, the autonomous proliferation was observed only in D816V-KIT expressing cells, and the other mutant KIT expressing cells required SCF for their proliferations like Wt-KIT expressing cells. These results were confirmed by the colony formation ability in the semi-liquid media, where only D816V-KIT expressing cells could form the colony without any growth factors. The growth inhibitory effect of KI-328 was, therefore, examined in the existence of 50 ng/ml of SCF. KI-328 inhibited the growth of Wt-, M541L-, V540L-, T417F- and N822K-KIT expressing cells with the GI50 value 127 nM, 229 nM, 575 nM, 445 nM and 997 nM, respectively. The cell cycle analysis showed the KI-328 increased sub-G1 populations in these cells at each GI50 value. In consistent with the growth inhibitory effects, KI-328 potently inhibited the phosphorylations of Wt- and mutant KITs except D816V as well as their downstream molecules STAT3, AKT, MAPK at the concentration of over the GI50 value, indicating the proof of concept that KI-328 inhibits the growth of these cells by the KIT kinase inhibition. However, the significant growth inhibition was not observed in D816V-KIT expressing cells up to the 5 μM, and more than 2 μM of KI-328 were required for the de-phosphorylation of D816V-KIT. We further examined whether another potent KIT inhibitors showed the different sensitivities between D816V-KIT and Wt-KIT. Multi-kinase inhibitors such as dasatinib and sunitinib showed the same growth inhibitory effects on D816V- and Wt-KIT expressing cells: each GI50 value against D816V- and Wt-KIT was 43 nM and 72 nM, and 116 nM and 206 nM, respectively. In contrast, imatinib, which is relatively selective against KIT kinase, did not inhibit the growth of the D816V-KIT expressing cells like KI-328. Conclusions We demonstrated that KI-328 is a potent and selective KIT inhibitor. Although KI-328 did not show the significant growth inhibitory effect on the D816V-KIT expressing 32D cells up to the 5 μM, G-CSF mediating neutrophil maturation was observed when those were treated with less than 1 μM of KI-328, indicating that KI-328 has a weak potency against the D816V-KIT kinase. Therefore, the combination therapy with another potent KIT inhibitors, such as HSP90 inhibitor, might conquer the resistance against the D816V-KIT kinase. Since the kinase inhibitory profile seemed to be associated with the resistance against the D816V-KIT kinase, the structural analysis of the D816V-KIT is required for developing more potent inhibitors against all mutant KIT kinases. Disclosures: Shiotsu: Kyowa Hakko Kirin Co., Ltd.: Employment. Kiyoi:Kyowa Hakko Kirin Co. Ltd.: Consultancy; Novartis Pharma Co. Ltd.: Research Funding. Ishida:Kyowa Hakko Kirin Co., Ltd.: Employment. Naoe:Kyowa Hakko Kirin Co., Ltd. : Research Funding; Chugai Pharmaceutical Co.,Ltd.: Research Funding; Wyeth K.K.: Research Funding.
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Sellitepe, Hasan Erdinç, Jong Min Oh, İnci Selin Doğan, Sercan Yildirim, Ahmet Buğra Aksel, Geum Seok Jeong, Ahmed Khames, et al. "Synthesis of N′-(4-/3-/2-/Non-substituted benzylidene)-4-[(4-methylphenyl)sulfonyloxy] Benzohydrazides and Evaluation of Their Inhibitory Activities against Monoamine Oxidases and β-Secretase." Applied Sciences 11, no. 13 (June 23, 2021): 5830. http://dx.doi.org/10.3390/app11135830.
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Nineteen tosylated acyl hydrazone derivatives were synthesized, and their inhibitory activities against monoamine oxidases (MAOs), acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and β-secretase (BACE-1) were evaluated. Compound 3o was the most potent inhibitor of MAO-A, with an IC50 value of 1.54 µM, followed by 3a (IC50 = 3.35 µM). A structural comparison with 3a indicated that the 3-F group in 3o increased its inhibitory activity against MAO-A. Compound 3s was the most potent inhibitor of MAO-B, with an IC50 value of 3.64 µM, followed by 3t (IC50 = 5.69 µM). The MAO-B inhibitory activity increased in the order of 3- > 4- > 2-NO2 groups in 3s, 3t, and 3r, respectively. All the compounds weakly inhibited AChE and BChE, which retained >50% residual activity at 10 µM, except for 3a, which inhibited BChE with an IC50 value of 16.1 µM. Interestingly, 3e, 3f, and 3n inhibited BACE-1 with IC50 values of 8.63, 9.92, and 8.47 µM, respectively, which were lower than the IC50 of the quercetin reference. Compounds 3o and 3s were found to be reversible competitive inhibitors of MAO-A and MAO-B, respectively, with Ki values of 0.35 ± 0.074 and 1.97 ± 0.65 µM, respectively. Moreover, compounds 3e, 3f, and 3n were effective BACE-1 inhibitors. The lead molecules were further investigated by molecular docking studies to elucidate the binding interactions with the target enzymes.
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Wang, Wei-Ya, Chien-Kei Wei, Che-Ming Teng, and Chin-Chung Wu. "Combined blockade of thrombin anion binding exosite-1 and PAR4 produces synergistic antiplatelet effect in human platelets." Thrombosis and Haemostasis 105, no. 01 (2011): 88–95. http://dx.doi.org/10.1160/th10-05-0305.
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SummaryThrombin exosite-1 mediates the specific binding of thrombin with fibrinogen and protease-activated receptor (PAR) 1. Exosite-1 inhibitors have been shown to effectively decrease the clotting activity of thrombin, while their antiplatelet effects are relatively weak. In the present study, the inhibitory effects of two exosite-1 inhibitors, hirugen and HD1, but not the exosite-2 inhibitor HD22, on thrombin-induced platelet aggregation and P-selectin expression were dramatically enhanced by a PAR4 antagonist, YD-3. In contrast, the PAR1 antagonist SCH-79797 did not affect the antiplatelet effects of exosite-1 inhibitors. The exosite-1 inhibitors and YD-3 prevented the Ca2+ spike and the prolonged Ca2+ response in thrombin-stimulated platelets, respectively; and combination of these two classes of agents led to abolishment of Ca2+ signal. Unlike exosite-1 inhibitors, the antiplatelet effects of the active site inhibitor PPACK and the bivalent inhibitor bivalirudin were not significantly enhanced by YD-3. In addition, the platelet-stimulating activity of γ-thrombin, an autolytic product of α-thrombin which lacks exosite-1, was inhibited by YD-3. These results suggest that the synergistic antiplatelet effects of exosite-1 inhibitor and PAR4 antagonist are resulted from combined blockade of PAR1 and PAR4 in platelets. In fibrinogen or plasma clotting assay, YD-3 neither prolonged the clotting time on its own nor enhanced the anticoagulant activity of exosite-1 inhibitors. Therefore, the combined blockade of exosite-1 and PAR4 may offer a potential strategy for improving the balance of benefits and risks of antithrombotic therapy.
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Volk, Kenneth A., Russell F. Husted, Peter M. Snyder, and John B. Stokes. "Kinase regulation of hENaC mediated through a region in the COOH-terminal portion of the α-subunit." American Journal of Physiology-Cell Physiology 278, no. 5 (May 1, 2000): C1047—C1054. http://dx.doi.org/10.1152/ajpcell.2000.278.5.c1047.
Full textAbstract:
In an effort to gain insight into how kinases might regulate epithelial Na+ channel (ENaC) activity, we expressed human ENaC (hENaC) in Xenopus oocytes and examined the effect of agents that modulate the activity of some kinases. Activation of protein kinase C (PKC) by phorbol ester increased the activity of ENaC, but only in oocytes with a baseline current of <2,000 nA. Inhibitors of protein kinases produced varying effects. Chelerythrine, an inhibitor of PKC, produced a significant inhibition of ENaC current, but calphostin C, another PKC inhibitor, had no effect. The PKA/protein kinase G inhibitor H-8 had no effect, whereas the p38 mitogen-activated protein kinase inhibitor, SB-203580 had a significant inhibitory effect. Staurosporine, a nonspecific kinase inhibitor, was the most potent tested. It inhibited ENaC currents in both oocytes and in M-1 cells, a model for the collecting duct. Site-directed mutagenesis revealed that the staurosporine effect did not require an intact COOH terminus of either the β- or γ-hENaC subunit. However, an intact COOH terminus of the α-subunit was required for this effect. These results suggest that an integrated kinase network regulates ENaC activity through an action that requires a portion of the α-subunit.
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Coutinho, M., K. S. Aulak, and A. E. Davis. "Functional analysis of the serpin domain of C1 inhibitor." Journal of Immunology 153, no. 8 (October 15, 1994): 3648–54. http://dx.doi.org/10.4049/jimmunol.153.8.3648.
Full textAbstract:
Abstract To analyze the role of the heavily glycosylated amino-terminal domain of C1 inhibitor in protease inhibitory activity, two truncated C1 inhibitor molecules were constructed. The abilities of the recombinant truncated inhibitors to complex with target proteases were compared with that of the wild-type recombinant protein. One recombinant truncated molecule consisted of amino acid residues 76 to 478 (C-serp(76)) and the other of residues 98 to 478 (C-serp(98)). The recombinant proteins were each expressed in similar quantities. The thermal denaturation profiles of the two truncated proteins were similar to that of the wild-type protein. Identical binding of C1s, C1r, kallikrein, and beta factor XIIa was observed with the three molecules. Furthermore, the truncated molecules also effectively inhibited C1 activity in hemolytic assays. These studies therefore clearly demonstrate that the amino-terminal domain of C1 inhibitor does not influence complex formation with target proteases.
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Rymerson, Robert T., and Robert P. Bodnaryk. "GUT PROTEINASE ACTIVITY IN INSECT PESTS OF CANOLA." Canadian Entomologist 127, no. 1 (February 1995): 41–48. http://dx.doi.org/10.4039/ent12741-1.
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AbstractThe digestive proteinases of three important pests of canola, Brassica napus L. and B. rapa L., in western Canada were characterized by assessing the proteolytic activity of homogenates of their midguts against azocasein or azoalbumin at various pH levels and in the presence of diagnostic proteinase inhibitors. The midgut of larvae of the bertha armyworm, Mamestra configurata Wlk., had maximum proteolytic activity at pH 10.5 which was inhibited 45–60% by serine proteinase inhibitors such as the soybean trypsin inhibitor. The midgut of larvae of the diamondback moth, Plutella xylostella L., had maximum proteolytic activity at pH 10 which was inhibited 56–75% by serine proteinase inhibitors. The two lepidopterans thus use a serine-like proteinase in digestion. The midgut of adults of the flea beetle, Phyllotreta cruciferae Goeze, exhibited maximum proteolytic activity at pH 5 which was inhibited 33–61% by specific cysteine proteinase inhibitors such as cystatin and trans-epoxysuccinyl-L-leucylamido (4-guanidino)-butane (E-64) and was activated strongly by L-cysteine. Aspartic proteinase inhibitors such as pepstatin A also decreased proteolytic activity by 21–50%. Serine proteinase inhibitors were without effect. Therefore, P. cruciferae appears to use both cysteine- and aspartic-like proteinases in digestion. Cotyledons and first true leaves of canola, B. napus cv. Westar, contained inhibitory activity against serine, cysteine, and aspartic proteinases when tested against bovine trypsin, papain, or porcine pepsin, but the level of antiproteinase activity is insufficient to provide significant resistance against any of these pests.
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Milner, Malgorzata, Jadwiga Chroboczek, and Wlodzimierz Zagorski-Ostoja. "Engineered resistance against proteinases." Acta Biochimica Polonica 54, no. 3 (September 6, 2007): 523–36. http://dx.doi.org/10.18388/abp.2007_3226.
Full textAbstract:
Exogenous proteinase inhibitors are valuable and economically interesting protective biotechnological tools. We examined whether small proteinase inhibitors when fused to a selected target protein can protect the target from proteolytic degradation without simultaneously affecting the function and activity of the target domain. Two proteinase inhibitors were studied: a Kazal-type silk proteinase inhibitor (SPI2) from Galleria mellonella, and the Cucurbita maxima trypsin inhibitor I (CMTI I). Both inhibitors target serine proteinases, are small proteins with a compact structure stabilized by a network of disulfide bridges, and are expressed as free polypeptides in their natural surroundings. Four constructs were prepared: the gene for either of the inhibitors was ligated to the 5' end of the DNA encoding one or the other of two selected target proteins, the coat protein (CP) of Potato potyvirus Y or the Escherichia coli beta-glucuronidase (GUS). CMTI I fused to the target proteins strongly hampered their functions. Moreover, the inhibitory activity of CMTI I was retained only when it was fused to the CP. In contrast, when fused to SPI2, specific features and functions of both target proteins were retained and the inhibitory activity of SPI2 was fully preserved. Measuring proteolysis in the presence or absence of either inhibitor, we demonstrated that proteinase inhibitors can protect target proteins used either free or as a fusion domain. Interestingly, their inhibitory efficiency was superior to that of a commercial inhibitor of serine proteinases, AEBSF.
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ALIAS, ZAZALI, and NORA ASYIKIN RAMLI. "PURIFICATION AND PARTIAL CHARACTERISATION OF A PROTEASE INHIBITOR FROM Mimosa diplotricha." Malaysian Applied Biology 51, no. 4 (October 31, 2022): 169–75. http://dx.doi.org/10.55230/mabjournal.v51i4.26.
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Search for inhibitors to insect proteases is one of many strategies to control pests. Previous work has demonstrated successful purification of effective inhibitors from plant origin. Thus, the current study attempted to purify protease inhibitors from locally available medicinal plants. The study demonstrated that the ethanolic extracts of Mimosa diplotricha leaves caused a significant 80% reduction in bovine trypsin activity. The inhibitory property of the proteinaceous nature of the extract was reconfirmed through qualitative analysis using the detection of trypsin inhibitors on the SDS-PAGE technique. The ammonium precipitated trypsin inhibitor was purified using Hi-Trap G25 and resolved into a single band with a molecular weight of approximately 20.8 kDa. By using the Dixon plot the competitive inhibitor has a Ki value of 2.16 × 10-4 mM. The purified protein inhibited the protease extract of Chrysomya megacephala at IC50 of 28 μg/mL. The results highlighted the presence of trypsin inhibitor in Mimosa diplotricha and its potential as a pest control agent.
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Curry, V. A., I. M. Clark, H. Bigg, and T. E. Cawston. "Large inhibitor of metalloproteinases (LIMP) contains tissue inhibitor of metalloproteinases (TIMP)-2 bound to 72,000-Mr progelatinase." Biochemical Journal 285, no. 1 (July 1, 1992): 143–47. http://dx.doi.org/10.1042/bj2850143.
Full textAbstract:
Connective-tissue cells in culture produce a family of metalloproteinases which, once activated, can degrade all the components of the extracellular matrix. These potent enzymes are all inhibited by the tissue inhibitor of metalloproteinases (TIMP), and it was thought that this inhibitor was solely responsible for the inhibition of these enzymes within connective tissue. However, other inhibitors have recently been described, including large inhibitor of metalloproteinases (LIMP) present in the culture medium of human foetal lung fibroblasts. Here we show that a large proportion of the inhibitory activity of LIMP consists of 72,000-M(r)-progelatinase bound to TIMP-2, a recently discovered low-M(r) metalloproteinase inhibitor closely related to TIMP. The physiological implications of the secretion of a complex of 72,000-M(r) progelatinase and TIMP-2 are discussed, and the separation of the complex in 6 M-urea is described.
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Pujirahayu, Niken, Debu Kumar Bhattacharjya, Toshisada Suzuki, and Takeshi Katayama. "α-Glucosidase Inhibitory Activity of Cycloartane-Type Triterpenes Isolated from Indonesian Stingless Bee Propolis and Their Structure–Activity Relationship." Pharmaceuticals 12, no. 3 (July 1, 2019): 102. http://dx.doi.org/10.3390/ph12030102.
Full textAbstract:
This study reports on the antioxidant activity and α-glucosidase inhibitory activity of five cycloartane-type triterpenes isolated from Indonesian stingless bee (Tetragonula sapiens Cockerell) propolis and their structure–activity relationships. The structure of the triterpenes was determined to include mangiferolic acid (1), Cycloartenol (2), ambonic acid (3), mangiferonic acid (4), and ambolic acid (5). The inhibitory test results of all isolated triterpenes against α-glucosidase showed a high potential for inhibitory activity with an IC50 range between 2.46 and 10.72 µM. Among the compounds tested, mangiferonic acid (4) was the strongest α-glucosidase inhibitor with IC50 2.46 µM compared to the standard (–)-epicatechin (1991.1 µM), and also had antioxidant activities with IC50 values of 37.74 ± 6.55 µM. The study on the structure–activity relationships among the compounds showed that the ketone group at C-3 and the double bonds at C-24 and C-25 are needed to increase the α-glucosidase inhibitory activity. The carboxylic group at C-26 is also more important for increasing the inhibitory activity compared with the methyl group. This study provides an approach to help consider the structural requirements of cycloartane-type triterpenes from propolis as α-glucosidase inhibitors. An understanding of these requirements is deemed necessary to find a new type of α-glucosidase inhibitor from the cycloartane-type triterpenes or to improve those inhibitors that are known to help in the treatment of diabetes.
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Wu, Wei, Hai-Ying Sun, Xiu-Ling Deng, and Gui-Rong Li. "EGFR tyrosine kinase regulates human small-conductance Ca2+-activated K+ (hSKCa1) channels expressed in HEK-293 cells." Biochemical Journal 452, no. 1 (April 25, 2013): 121–29. http://dx.doi.org/10.1042/bj20121324.
Full textAbstract:
SKCa (small-conductance Ca2+-activated K+) channels are widely distributed in different tissues, including the brain, pancreatic islets and myocardium and play an important role in controlling electrical activity and cellular functions. However, intracellular signal modulation of SKCa channels is not fully understood. The present study was designed to investigate the potential regulation of hSKCa1 (human SKCa1) channels by PTKs (protein tyrosine kinases) in HEK (human embryonic kidney)-293 cells expressing the hSKCa1 (KCNN1) gene using approaches of whole-cell patch voltage-clamp, immunoprecipitation, Western blotting and mutagenesis. We found that the hSKCa1 current was inhibited by the broad-spectrum PTK inhibitor genistein, the selective EGFR (epidermal growth factor receptor) kinase inhibitors T25 (tyrphostin 25) and AG556 (tyrphostin AG 556), but not by the Src-family kinases inhibitor PP2. The inhibitory effect of these PTK inhibitors was significantly antagonized by the PTP (protein tyrosine phosphatase) inhibitor orthovanadate. The tyrosine phosphorylation level of hSKCa1 channels was reduced by genistein, T25 or AG556. The reduced tyrosine phosphorylation was countered by orthovanadate. Interestingly, the Y109F mutant hSKCa1 channel lost the inhibitory response to T25 or AG556, and showed a dramatic reduction in tyrosine phosphorylation levels and a reduced current density. These results demonstrate the novel information that hSKCa1 channels are inhibited by genistein, T25 and AG556 via EGFR tyrosine kinase inhibition, which is related to the phosphorylation of Tyr109 in the N-terminus. This effect may affect electrical activity and cellular functions in brain, pancreatic islets and myocardium.
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Korn, J. H., K. M. Brown, E. Downie, Z. H. Liao, and D. L. Rosenstreich. "Augmentation of IL 1-induced fibroblast PGE2 production by a urine-derived IL 1 inhibitor." Journal of Immunology 138, no. 10 (May 15, 1987): 3290–94. http://dx.doi.org/10.4049/jimmunol.138.10.3290.
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Abstract IL 1, a monocyte-derived cytokine, has potent biologic effects in a variety of target tissues. The existence of naturally occurring inhibitors of IL 1 activity has been recently described; these inhibitors blocked one IL 1 effect: stimulation of thymocyte responses to mitogens. We examined the effect of one well-characterized inhibitor of IL 1, isolated from the urine of febrile patients, on a second IL 1 effect, stimulation of fibroblast PGE synthesis. In this system, purified preparations of the urinary inhibitor that completely blocked murine thymocyte proliferative responses to mitogen failed to block PGE synthetic responses to IL 1. Rather, inhibitor preparations markedly enhanced fibroblast PGE synthetic responses to IL 1. When partially purified inhibitor preparations were fractionated by ion exchange chromatography, inhibitory activity for the IL 1 effect on thymocytes and PGE stimulatory activity co-eluted. Augmentation of the IL 1-induced PGE response was seen with both low (1:1 unit) and high (400:1) ratios of inhibitor to IL 1. Inhibitor preparations alone did not stimulate fibroblast PGE synthesis. The augmentation of fibroblast PGE synthesis by inhibitor preparations was not due to contaminating endotoxin. Active inhibitor preparations contained less than 15 pg of endotoxin/U activity, and the PGE stimulatory effect was not blocked by the addition of polymyxin B, whereas polymyxin B reversed the effects of exogenous endotoxin. It appears that the inhibition of IL 1 effects by naturally occurring inhibitors may have target cell and/or functional specificity.
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Thøgersen, I. B., G. Salvesen, F. H. Brucato, S. V. Pizzo, and J. J. Enghild. "Purification and characterization of an α-macroglobulin proteinase inhibitor from the mollusc Octopus vulgaris." Biochemical Journal 285, no. 2 (July 15, 1992): 521–27. http://dx.doi.org/10.1042/bj2850521.
Full textAbstract:
The cell-free haemolymph of the mollusc Octopus vulgaris inhibited the proteolytic activity of the thermolysin against the high-molecular-mass substrate hide powder azure. The purified inhibitor was a glycoprotein composed of two identical 180 kDa disulphide-linked subunits. In addition to the inhibition of the metalloproteinase thermolysin, the protein inhibited the serine proteinases human neutrophil elastase, pig pancreatic elastase, bovine chymotrypsin, bovine trypsin and the cysteine proteinase papain. A fraction of the proteinase-inhibitor complex resisted dissociation after denaturation indicating that some of the proteinase molecules became covalently bound. The nucleophile beta-aminopropionitrile decreased the covalent binding of proteinases to the Octopus vulgaris protein, suggesting that this interaction is mediated by an internal thiol ester; the reactivity and the amino acid sequence flanking the reactive residues of the putative thiol ester were consistent with this hypothesis. Bound trypsin remained active against the low-molecular-mass chromatogenic substrate H-D-Pro-Phe-Arg p-nitroanilide and was protected from inhibition by active-site-directed protein inhibitors of trypsin; however, the bound trypsin was readily inhibited by small synthetic inhibitors. This indicates that the inhibition of proteinases is accomplished by steric hindrance. The proteinase-inhibitory activity of this protein is characteristic of inhibition by mammalian alpha-macroglobulins and the presence of a putative thiol ester suggests that the Octopus vulgaris proteinase inhibitor is a homologue of human alpha 2-macroglobulin.
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Chandrashekharaiah, K. S. "Studies on the Amylase Inhibitors from the Seeds of Adenanthera Pavonina." Biosciences, Biotechnology Research Asia 14, no. 3 (September 25, 2017): 1009–15. http://dx.doi.org/10.13005/bbra/2535.
Full textAbstract:
ABSTRACT: An α-Amylase inhibitor was isolated and purified employing ammonium sulphate fractionation, molecular sieve chromatography on sephadex G-10 and G-50 and HPLC from the seeds of Adenanthera pavonina. The molecular weight was found to be 10 - 12 kDa as determined by gel-permeation chromatography on Sephadex G-100. The specific inhibitor activity, fold purity and the yield obtained for Adenanthera pavonina amylase inhibitor was 32.12, 51 and 13.07, respectively. The purified inhibitor was heat stable and retained more than 52% activity at 65°C. The optimum pH obtained for purified inhibitor was 6.3 and 100% Zone of inhibition was observed when it was added on the plated organisms. The Adenanthera pavonina amylase inhibitor inhibited the activity of human salivary α-amylase and inhibitory activity of α-amylase inhibitor against mammalian amylases could suggest its potential in treatment of diabetes and related nutritional problems results in obesity.
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Ellis, Dorette Z., James A. Nathanson, and Kathleen J. Sweadner. "Carbachol inhibits Na+-K+-ATPase activity in choroid plexus via stimulation of the NO/cGMP pathway." American Journal of Physiology-Cell Physiology 279, no. 6 (December 1, 2000): C1685—C1693. http://dx.doi.org/10.1152/ajpcell.2000.279.6.c1685.
Full textAbstract:
Secretion of cerebrospinal fluid by the choroid plexus can be inhibited by its cholinergic innervation. We demonstrated that carbachol inhibits the Na+-K+-ATPase in bovine choroid tissue slices and investigated the mechanism. Many of the actions of cholinergic agents are mediated by nitric oxide (NO), which plays important roles in fluid homeostasis. The inhibition of Na+-K+-ATPase was blocked by the NO synthase inhibitor [ N ω-nitro-L-arginine methyl ester] and was quantitatively mimicked by the NO agonists sodium nitroprusside (SNP) and diethylenetriamine NO. Inhibition by SNP correlated with an increase in tissue cGMP and was abolished by 1 H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one, an inhibitor of soluble guanylate cyclase. Inhibition was mimicked by the protein kinase G activator 8-bromo-cGMP and by okadaic acid, an inhibitor of protein phosphatases 1 and 2A. cGMP-dependent protein kinase inhibitors Rp-8-pCPT-cGMP (0.5–5 μM) and KT-5823 (2.0 μM) did not block the effects of SNP, but higher concentrations of the more selective inhibitor (Rp-8-pCPT-cGMP) had a pharmacological inhibitory effect on Na+-K+-ATPase. The data suggest that cholinergic regulation of the Na+-K+-ATPase is mediated by NO and involves activation of guanylate cyclase and elevation of cGMP.
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Buzzelli, Mark D., Murali Nagarajan, John F. Radtka, Margaret L. Shumate, Maithili Navaratnarajah, Charles H. Lang, and Robert N. Cooney. "Nuclear Factor-κB Mediates the Inhibitory Effects of Tumor Necrosis Factor-α on Growth Hormone-Inducible Gene Expression in Liver." Endocrinology 149, no. 12 (August 21, 2008): 6378–88. http://dx.doi.org/10.1210/en.2007-1574.
Full textAbstract:
TNF inhibits serine protease inhibitor 2.1 (Spi 2.1) and IGF-I gene expression by GH in CWSV-1 hepatocytes. The current study describes construction of a GH-inducible IGF-I promoter construct and investigates mechanisms by which TNF and nuclear factor-κB (NFκB) inhibit GH-inducible gene expression. CWSV-1 cells were transfected with GH-inducible Spi 2.1 or IGF-I promoter luciferase constructs, incubated with TNF signaling inhibitors (fumonisin B1 for sphingomyelinase and SP600125 for c-Jun N-terminal kinase), treated with or without TNF, and then stimulated with recombinant human GH. The 5- to 6-fold induction of Spi 2.1 and IGF-I promoter activity by GH was inhibited by TNF. Neither fumonisin B1 nor SP600125 prevented the inhibitory effects of TNF on GH-inducible promoter activity. Dominant-negative inhibitor-κBα (IκBα) expression vectors (IκBαS/A or IκBαTrunc), p65 and p50 expression vectors, and p65 deletion constructs were used to investigate the NFκB pathway. IκBαS/A and IκBαTrunc ameliorated the inhibitory effects of TNF on GH-inducible Spi 2.1 and IGF-I promoter activity. Cotransfection of CWSV-1 cells with expression vectors for p65 alone or p50 and p65 together inhibited GH-inducible Spi 2.1 and IGF-I promoter activity. Cotransfection with a C-terminal p65 deletion (1–450) enhanced GH-inducible promoter activity, whereas the N-terminal deletion (31–551) was inhibitory for IGF-I but not Spi 2.1. Cycloheximide did not antagonize the inhibitory effects of TNF on GH-inducible IGF-I expression. We conclude the inhibitory effects of TNF on GH-inducible promoter activity are mediated by NFκB, especially p65, by a mechanism that does not require protein synthesis.
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Scaffa, P. M. C., C. M. P. Vidal, N. Barros, T. F. Gesteira, A. K. Carmona, L. Breschi, D. H. Pashley, et al. "Chlorhexidine Inhibits the Activity of Dental Cysteine Cathepsins." Journal of Dental Research 91, no. 4 (January 19, 2012): 420–25. http://dx.doi.org/10.1177/0022034511435329.
Full textAbstract:
The co-expression of MMPs and cysteine cathepsins in the human dentin-pulp complex indicates that both classes of enzymes can contribute to the endogenous proteolytic activity of dentin. Chlorhexidine (CHX) is an efficient inhibitor of MMP activity. This study investigated whether CHX could also inhibit cysteine cathepsins present in dentin. The inhibitory profile of CHX on the activity of dentin-extracted and recombinant cysteine cathepsins (B, K, and L) was monitored in fluorogenic substrates. The rate of substrate hydrolysis was spectrofluorimetrically measured, and inhibitory constants were calculated. Molecular docking was performed to predict the binding affinity between CHX and cysteine cathepsins. The results showed that CHX inhibited the proteolytic activity of dentin-extracted cysteine cathepsins in a dose-dependent manner. The proteolytic activity of human recombinant cathepsins was also inhibited by CHX. Molecular docking analysis suggested that CHX strongly interacts with the subsites S2 to S2′ of cysteine cathepsins B, K, and L in a very similar manner. Taken together, these results clearly showed that CHX is a potent inhibitor of the cysteine cathepsins-proteolytic enzymes present in the dentin-pulp complex.
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Han, Di, Huiqun Wang, Wei Cui, Beibei Zhang, and Bo-Zhen Chen. "Computational insight into the mechanisms of action and selectivity of Afraxis PAK inhibitors." Future Medicinal Chemistry 12, no. 5 (March 2020): 367–85. http://dx.doi.org/10.4155/fmc-2019-0273.
Full textAbstract:
Aim: The p21-activated kinases (PAKs) are involved in many important biological activity regulations. FRAX019, FRAX414, FRAX597, FRAX1036 and G-5555 were identified as PAKs inhibitors. Their detailed inhibitory mechanisms deserve further investigation. Results: Molecular dynamics simulations and further calculations for the PAK1/inhibitor and PAK4/inhibitor complexes indicate that their binding free energies are basically consistent with the trend of experimental activity data. Conclusion: The anchoring of residues Leu347PAK1 and Leu398PAK4 is the structural basis for designing Afraxis PAK inhibitors. This study discloses the inhibitory mechanisms of FRAX019, FRAX414, FRAX597, FRAX1036 and G-5555 toward PAK1 and PAK4 and some clues to enhance kinase activities and selectivities, which will provide valuable information to the development of more potent and selective PAK inhibitors.
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Jacquemin, Marc, Abdellah Benhida, Kathelijne Peerlinck, Benoı̂t Desqueper, Luc Vander Elst, Renaud Lavend'homme, Roseline d'Oiron, et al. "A human antibody directed to the factor VIII C1 domain inhibits factor VIII cofactor activity and binding to von Willebrand factor." Blood 95, no. 1 (January 1, 2000): 156–63. http://dx.doi.org/10.1182/blood.v95.1.156.
Full textAbstract:
Abstract The occurrence of factor VIII (fVIII) inhibitory antibodies is a rare complication of fVIII substitution therapy in mild/moderate hemophilia A patients. fVIII mutations in certain regions such as the C1 domain are, however, more frequently associated with inhibitor, for reasons which remain unclear. To determine whether inhibitors could map to the mutation site, we analyzed at the clonal level the immune response of such a patient with an inhibitor to wild-type but not self-fVIII and an Arg2150His substitution in the C1 domain. Immortalization of the patient B lymphocytes provided a cell line producing an anti-fVIII IgG4κ antibody, LE2E9, that inhibited fVIII cofactor activity, following type 2 kinetics and prevented fVIII binding to von Willebrand factor. Epitope mapping with recombinant fVIII fragments indicated that LE2E9 recognized the fVIII C1 domain, but not the Arg2150His-substituted C1 domain. Accordingly, LE2E9 did not inhibit Arg2150His fVIII activity. These observations identify C1 as a novel target for fVIII inhibitors and demonstrate that Arg2150His substitution alters a B-cell epitope in the C1 domain, which may contribute to the higher inhibitor incidence in patients carrying such substitution. (Blood. 2000; 95:156-163)
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Jacquemin, Marc, Abdellah Benhida, Kathelijne Peerlinck, Benoı̂t Desqueper, Luc Vander Elst, Renaud Lavend'homme, Roseline d'Oiron, et al. "A human antibody directed to the factor VIII C1 domain inhibits factor VIII cofactor activity and binding to von Willebrand factor." Blood 95, no. 1 (January 1, 2000): 156–63. http://dx.doi.org/10.1182/blood.v95.1.156.001k50_156_163.
Full textAbstract:
The occurrence of factor VIII (fVIII) inhibitory antibodies is a rare complication of fVIII substitution therapy in mild/moderate hemophilia A patients. fVIII mutations in certain regions such as the C1 domain are, however, more frequently associated with inhibitor, for reasons which remain unclear. To determine whether inhibitors could map to the mutation site, we analyzed at the clonal level the immune response of such a patient with an inhibitor to wild-type but not self-fVIII and an Arg2150His substitution in the C1 domain. Immortalization of the patient B lymphocytes provided a cell line producing an anti-fVIII IgG4κ antibody, LE2E9, that inhibited fVIII cofactor activity, following type 2 kinetics and prevented fVIII binding to von Willebrand factor. Epitope mapping with recombinant fVIII fragments indicated that LE2E9 recognized the fVIII C1 domain, but not the Arg2150His-substituted C1 domain. Accordingly, LE2E9 did not inhibit Arg2150His fVIII activity. These observations identify C1 as a novel target for fVIII inhibitors and demonstrate that Arg2150His substitution alters a B-cell epitope in the C1 domain, which may contribute to the higher inhibitor incidence in patients carrying such substitution. (Blood. 2000; 95:156-163)
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Javaid, Bushra. "Anti-trypsin and antihypertensive activity of protein extracts from Nigella sativa seeds." Pakistan Journal of Agricultural Sciences 58, no. 04 (September 1, 2021): 1237–43. http://dx.doi.org/10.21162/pakjas/21.204.
Full textAbstract:
Protease inhibitors (PIs) are a ubiquitous, diverse group of molecules present in multiple forms in all organisms. These inhibitors inactivate proteases from predators/pathogens in addition to regulating intracellular proteolysis. In addition to intracellular localization, storage organs of plants are also a potential site of protease inhibitors. Proteins with trypsin inhibitory activity were isolated from Nigella sativa seed extracts by ammonium sulphate precipitation. Extraction conditions were optimized by choosing an optimum solvent, temperature and incubation period. The highest inhibitory activity of protein extracts was achieved by using 50 mM Tris buffer as solvent and overnight incubation of the suspension at 4°C. The crude seed extract fractionated at 60% ammonium sulphate concentration exhibited highest trypsin inhibitory activity, i.e., 60.15 ± 2.95 %, which was comparable to soybean trypsin inhibitor used as positive control. Ammonium sulphate precipitation of crude extract yielded 39.83-fold purification. Partially purified trypsin inhibitor exhibited 2.39±0.23 TIU mg-1. Additionally, Nigella sativa protein extracts were also investigated for their health-promoting effects against two important proteases, α- Dipeptidyl peptidase-IV (DPP-IV) and angiotensin-converting enzyme (ACE). Highest inhibitory activity against ACE was shown by the crude extract of N. sativa. Among AS fractions, 30% ammonium sulphate concentration exhibited highest inhibition activity against ACE and DPP-IV. Our results suggest that the widely believed role of N. sativa in control of hypertension may at least be partially shared by inhibition of ACE. This is the first study conducted to evaluate the biological activity of N. sativa protein extracts suggesting a potential use of N. sativa proteins in management of hypertension as well as an important source of trypsin inhibitor. Further identification, purification and characterization of different bioactive compounds from N. sativa are being carried out.
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Demiroglu, Asuman, E. Joanna Steer, Carol Heath, Kerry Taylor, Mark Bentley, Steven L. Allen, Prasad Koduru, et al. "The t(8;22) in chronic myeloid leukemia fuses BCR to FGFR1: transforming activity and specific inhibition of FGFR1 fusion proteins." Blood 98, no. 13 (December 15, 2001): 3778–83. http://dx.doi.org/10.1182/blood.v98.13.3778.
Full textAbstract:
Abstract This report describes 2 patients with a clinical and hematologic diagnosis of chronic myeloid leukemia (CML) in chronic phase who had an acquired t(8;22)(p11;q11). Analysis by fluorescence in situ hybridization (FISH) and reverse transcription-polymerase chain reaction (RT-PCR) indicated that both patients were negative for the BCR-ABL fusion, but suggested that the BCR gene was disrupted. Further FISH indicated a breakpoint within fibroblast growth factor receptor 1 (FGFR1), the receptor tyrosine kinase that is known to be disrupted in a distinctive myeloproliferative disorder, most commonly by fusion to ZNF198. RT-PCR confirmed the presence in both cases of an in-frame messenger RNA fusion between BCR exon 4 and FGFR1 exon 9. Expression of BCR-FGFR1 in the factor-dependent cell line Ba/F3 resulted in interleukin 3-independent clones that grew at a comparable rate to cells transformed with ZNF198-FGFR1. The growth of transformed cells was inhibited by the phosphatidylinositol 3-kinase inhibitor LY294002, the farnesyltransferase inhibitors L744832 and manumycin A, the p38 inhibitors SB202190 and SB203580 but not by the MEK inhibitor PD98059. The growth of BaF3/BCR-FGFR1 and BaF3/ZNF198-FGFR1 was not significantly inhibited by treatment with STI571, but was inhibited by SU5402, a compound with inhibitory activity against FGFR1. Inhibition with this compound was associated with decreased phosphorylation of ERK1/2 and BCR-FGFR1 or ZNF198-FGFR1, and was dose dependent with an inhibitory concentration of 50% of approximately 5 μM. As expected, growth of BaF3/BCR-ABL was inhibited by STI571 but not by SU5402. The study demonstrates that the BCR-FGFR1 fusion may occur in patients with apparently typical CML. Patients with constitutively active FGFR1 fusion genes may be amenable to treatment with specific FGFR1 inhibitors.
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Ganea, D., D. Cearlock, J. Minowada, and S. Dray. "A serine proteinase inhibitor produced by an HTLV I virus-transformed human T lymphocyte line." Journal of Immunology 138, no. 4 (February 15, 1987): 1208–14. http://dx.doi.org/10.4049/jimmunol.138.4.1208.
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Abstract A previous report from our laboratory indicated that a proteinase inhibitor is produced by rabbit T lymphocytes. We now report that a human T cell line, C91/PL, produces a proteinase inhibitor which inhibits the enzymatic activity of trypsin and kallikrein. This newly identified proteinase inhibitor (LPI 1) did not inhibit the enzymatic activity of four other serine proteinases (thrombin, plasmin, chymotrypsin, or pancreatic elastase), a thiol proteinase (papain), or a carboxyl proteinase (pepsin). Active synthesis of LPI 1 by the C91/PL cell line was shown by the appearance of similar levels of inhibitory activity in sequential cell supernatants, lack of appearance of inhibitor in supernatants of cells killed by heat or sodium azide or of viable cells in the presence of cyclohexamide, and incorporation of a radiolabeled amino acid into newly synthesized inhibitor. Although both the inhibitor of rabbit origin and of human origin are proteins produced by T cells and have similar inhibitory specificity, important differences were observed: LPI 1 is sensitive to boiling and the two inhibitors migrate differently upon electrophoresis in substrate-containing polyacrylamide gel. Furthermore, LPI 1 was produced by a cell line of the T4 phenotype which had been established by in vitro viral transformation of human cord blood lymphocytes with HTLV 1 whereas the inhibitor of rabbit origin was produced by normal splenic T cells. Three other human T cell lines of the T4 phenotype, MOLT-13, KE-37, and HPB-ALL, from patients with acute lymphoblastic leukemia did not produce a proteinase inhibitor. Thus, the production of proteinase inhibitors does not appear to be a general characteristic of human T cell lines nor of the T4 subset. Proteinase inhibitors produced by T cells may have an immunoregulatory role in proteinase-mediated physiological processes.
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Cao, Jiangying, Wei Zhao, Chunlong Zhao, Qian Liu, Shunda Li, Guozhen Zhang, C. James Chou, and Yingjie Zhang. "Development of a Bestatin-SAHA Hybrid with Dual Inhibitory Activity against APN and HDAC." Molecules 25, no. 21 (October 28, 2020): 4991. http://dx.doi.org/10.3390/molecules25214991.
Full textAbstract:
With five histone deacetylase (HDAC) inhibitors approved for cancer treatment, proteolysis-targeting chimeras (PROTACs) for degradation of HDAC are emerging as an alternative strategy for HDAC-targeted therapeutic intervention. Herein, three bestatin-based hydroxamic acids (P1, P2 and P3) were designed, synthesized and biologically evaluated to see if they could work as HDAC degrader by recruiting cellular inhibitor of apoptosis protein 1 (cIAP1) E3 ubiquitin ligase. Among the three compounds, the bestatin-SAHA hybrid P1 exhibited comparable even more potent inhibitory activity against HDAC1, HDAC6 and HDAC8 relative to the approved HDAC inhibitor SAHA. It is worth noting that although P1 could not lead to intracellular HDAC degradation after 6 h of treatment, it could dramatically decrease the intracellular levels of HDAC1, HDAC6 and HDAC8 after 24 h of treatment. Intriguingly, the similar phenomenon was also observed in the HDAC inhibitor SAHA. Cotreatment with proteasome inhibitor bortezomib could not reverse the HDAC decreasing effects of P1 and SAHA, confirming that their HDAC decreasing effects were not due to protein degradation. Moreover, all three bestatin-based hydroxamic acids P1, P2 and P3 exhibited more potent aminopeptidase N (APN, CD13) inhibitory activities than the approved APN inhibitor bestatin, which translated to their superior anti-angiogenic activities. Taken together, a novel bestatin-SAHA hybrid was developed, which worked as a potent APN and HDAC dual inhibitor instead of a PROTAC.
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Vest, Rebekah S., Kurtis D. Davies, Heather O'Leary, J. David Port, and K. Ulrich Bayer. "Dual Mechanism of a Natural CaMKII Inhibitor." Molecular Biology of the Cell 18, no. 12 (December 2007): 5024–33. http://dx.doi.org/10.1091/mbc.e07-02-0185.
Full textAbstract:
Ca2+/calmodulin (CaM)-dependent protein kinase II (CaMKII) is a major mediator of cellular Ca2+ signaling. Several inhibitors are commonly used to study CaMKII function, but these inhibitors all lack specificity. CaM-KIIN is a natural, specific CaMKII inhibitor protein. CN21 (derived from CaM-KIIN amino acids 43–63) showed full specificity and potency of CaMKII inhibition. CNs completely blocked Ca2+-stimulated and autonomous substrate phosphorylation by CaMKII and autophosphorylation at T305. However, T286 autophosphorylation (the autophosphorylation generating autonomous activity) was only mildly affected. Two mechanisms can explain this unusual differential inhibitor effect. First, CNs inhibited activity by interacting with the CaMKII T-site (and thereby also interfered with NMDA-type glutamate receptor binding to the T-site). Because of this, the CaMKII region surrounding T286 competed with CNs for T-site interaction, whereas other substrates did not. Second, the intersubunit T286 autophosphorylation requires CaM binding both to the “kinase” and the “substrate” subunit. CNs dramatically decreased CaM dissociation, thus facilitating the ability of CaM to make T286 accessible for phosphorylation. Tat-fusion made CN21 cell penetrating, as demonstrated by a strong inhibition of filopodia motility in neurons and insulin secrection from isolated Langerhans' islets. These results reveal the inhibitory mechanism of CaM-KIIN and establish a powerful new tool for dissecting CaMKII function.
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Bishop, P. B., J. Young, T. Peng, and J. F. Richards. "An inhibitor of ornithine decarboxylase in the thymus and spleen of dexamethasone-treated rats." Biochemical Journal 226, no. 1 (February 15, 1985): 105–12. http://dx.doi.org/10.1042/bj2260105.
Full textAbstract:
A marked decrease in activity of ornithine decarboxylase in thymus and spleen occurs soon after treatment of rats with a glucocorticoid. In the present study, evidence was obtained that extracts of these tissues prepared 5 h after administration of dexamethasone, when the enzyme activity is very low, contain an inhibitor of ornithine decarboxylase. The inhibitor is also present at 12 h after treatment and, in lesser amount, at 2.5 h, but was not evident at 24 h. The inhibitory activity was destroyed by treatment with heat or with trypsin, and was not lost on dialysis of the extract. Preliminary experiments indicate that the Mr of the inhibitor is greater than 50 000, which differentiates it from antizyme, an inhibitor of ornithine decarboxylase found in several other cell types. The inhibitor seems to act by a non-catalytic and non-competitive mechanism. The inhibition is dependent on the amount of inhibitor and does not change with time. Since inhibition is not changed by dialysis of the inhibitory extract, its activity apparently does not require small-Mr substances. This differentiates it from inhibitors which inactivate ornithine decarboxylase by covalent modification, such as the polyamine-dependent protein kinase or transglutaminase. The formation of this inhibitor is an early event in lymphoid tissues in response to dexamethasone and may be important in causing the inhibition of cell division which precedes the destruction of lymphocytes.
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Dong, Wenxiu, Fei Luo, Yuguang Du, Xuefang Bai, and Xianzhen Li. "Production and properties of an inhibitor of the Pseudomonas autoinducer by Pseudomonas aeruginosa." Canadian Journal of Microbiology 51, no. 9 (September 1, 2005): 783–89. http://dx.doi.org/10.1139/w05-066.
Full textAbstract:
An inhibitor was found in the culture fluid of Pseudomonas aeruginosa PAO1, which could inhibit the activity of the Pseudomonas autoinducer (PAI). The maximal inhibitory activity occurred in stationary phase culture sup ernatant. The PAI inhibitor did not influence the cell growth and the PAI production by P. aeruginosa PAO1 when the PAI inhibitor was added into culture medium. The induced expression of lacZ in the reporter strain Agrobacterium tumefaciens NT1 was suppressed by this PAI inhibitor, whereas inhibition could be relieved by increasing the auto inducer concentration. The quorum sensing of P. aeruginosa was inhibited presumably by inhibiting the inducing activity of Pseudomonas autoinducer but not by inhibiting the production of Pseudomonas autoinducer. It was demonstrated that the structure of the PAI inhibitor was different from that of acyl-homoserine lactones.Key words: quorum sensing, autoinducer, PAI inhibitor, Pseudomonas aeruginosa, N-acylhomoserine lactone.
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Suzuki, Suzuki, Morio Arai, Kagehiro Amano, Kazuhiko Kagawa, and Katsuyuki Fukutake. "Factor VIII Inhibitor Antibodies with C2 Domain Specificity Are Less inhibitory to Factor VIII Complexed with von Willebrand Factor." Thrombosis and Haemostasis 76, no. 05 (1996): 749–54. http://dx.doi.org/10.1055/s-0038-1650655.
Full textAbstract:
SummaryIn order to clarify the potential role of von Willebrand factor (vWf) in attenuating the inactivation of factor VIII (fVIII) by those antibodies with C2 domain specificity, we investigated a panel of 14 human antibodies to fVIII. Immunoblotting analysis localized light chain (C2 domain) epitopes for four cases, heavy chain (A2 domain) epitopes in five cases, while the remaining five cases were both light and heavy chains. The inhibitor titer was considerably higher for Kogenate, a recombinant fVIII concentrate, than for Haemate P, a fVIII/vWf complex concentrate, in all inhibitor plasmas that had C2 domain specificity. In five inhibitor plasmas with A2 domain specificity and in five with both A2 and C2 domain specificities, Kogenate gave titers similar to or lower than those with Haemate P. The inhibitory effect of IgG of each inhibitor plasma was then compared with recombinant fVIII and its complex with vWf. When compared to the other 10 inhibitor IgGs, IgG concentration, which inhibited 50% of fVIII activity (IC50), was remarkably higher for the fVIII/vWf complex than for fVIII in all the inhibitor IgGs that had C2 domain reactivity. Competition of inhibitor IgG and vWf for fVIII binding was observed in an ELISA system. In 10 inhibitors that had C2 domain reactivity, the dose dependent inhibition of fVIII-vWf complex formation was observed, while, in the group of inhibitors with A2 domain specificity, there was no inhibition of the complex formation except one case. We conclude that a subset of fVIII inhibitors, those that bind to C2 domain determinants, are less inhibitory to fVIII when it is complexed with vWf that binds to overlapping region in the C2 domain.
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Murthy, Karnam S., Huiping Zhou, and Gabriel M. Makhlouf. "PKA-dependent activation of PDE3A and PDE4 and inhibition of adenylyl cyclase V/VI in smooth muscle." American Journal of Physiology-Cell Physiology 282, no. 3 (March 1, 2002): C508—C517. http://dx.doi.org/10.1152/ajpcell.00373.2001.
Full textAbstract:
Regulation of adenylyl cyclase type V/VI and cAMP-specific, cGMP-inhibited phosphodiesterase (PDE) 3 and cAMP-specific PDE4 by cAMP-dependent protein kinase (PKA) and cGMP-dependent protein kinase (PKG) was examined in gastric smooth muscle cells. Expression of PDE3A but not PDE3B was demonstrated by RT-PCR and Western blot. Basal PDE3 and PDE4 activities were present in a ratio of 2:1. Forskolin, isoproterenol, and the PKA activator 5,6-dichloro-1-β-d-ribofuranosyl benzimidazole 3′,5′-cyclic monophosphate, SP-isomer, stimulated PDE3A phosphorylation and both PDE3A and PDE4 activities. Phosphorylation of PDE3A and activation of PDE3A and PDE4 were blocked by the PKA inhibitors [protein kinase inhibitor (PKI) and H-89] but not by the PKG inhibitor (KT-5823). Sodium nitroprusside inhibited PDE3 activity and augmented forskolin- and isoproterenol-stimulated cAMP levels; PDE3 inhibition was reversed by blockade of cGMP synthesis. Forskolin stimulated adenylyl cyclase phosphorylation and activity; PKI blocked phosphorylation and enhanced activity. Stimulation of cAMP and inhibition of inositol 1,4,5-trisphosphate-induced Ca2+release and muscle contraction by isoproterenol were augmented additively by PDE3 and PDE4 inhibitors. The results indicate that PKA regulates cAMP levels in smooth muscle via stimulatory phosphorylation of PDE3A and PDE4 and inhibitory phosphorylation of adenylyl cyclase type V/VI. Concurrent generation of cGMP inhibits PDE3 activity and augments cAMP levels.
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Liang, Mingyu, and Franklyn G. Knox. "Nitric oxide activates PKCα and inhibits Na+-K+-ATPase in opossum kidney cells." American Journal of Physiology-Renal Physiology 277, no. 6 (December 1, 1999): F859—F865. http://dx.doi.org/10.1152/ajprenal.1999.277.6.f859.
Full textAbstract:
Nitric oxide (NO) reduces the molecular activity of Na+-K+-ATPase in opossum kidney (OK) cells, a proximal tubule cell line. In the present study, we investigated the cellular mechanisms for the inhibitory effect of NO on Na+-K+-ATPase. Sodium nitroprusside (SNP), a NO donor, inhibited Na+-K+-ATPase in OK cells, but not in LLC-PK1cells, another proximal tubule cell line. Similarly, phorbol 12-myristate 13-acetate, a protein kinase C (PKC) activator, inhibited Na+-K+-ATPase in OK, but not in LLC-PK1, cells. PKC inhibitors staurosporine or calphostin C, but not the protein kinase G inhibitor KT-5823, abolished the inhibitory effect of NO on Na+-K+-ATPase in OK cells. Immunoblotting demonstrated that treatment with NO donors caused significant translocation of PKCα from cytosolic to particulate fractions in OK, but not in LLC-PK1, cells. Furthermore, the translocation of PKCα in OK cells was attenuated by either the phospholipase C inhibitor U-73122 or the soluble guanylate cyclase inhibitor 1H-[1,2,4]oxadiazolo-[4,3-a]quinoxalin-1-one. U-73122 also blunted the inhibitory effect of SNP on Na+-K+-ATPase in OK cells. The phospholipase A2inhibitor AACOCF3 did not blunt the inhibitory effect of SNP on Na+-K+-ATPase in OK cells. AACOCF3 alone, however, also decreased Na+-K+-ATPase activity in OK cells. In conclusion, our results demonstrate that NO activates PKCα in OK, but not in LLC-PK1, cells. The activation of PKCα in OK cells by NO is associated with inhibition of Na+-K+-ATPase.
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Saleh, Neveen M., Yasmine S. Moemen, Sara H. Mohamed, Ghady Fathy, Abdullah A. S. Ahmed, Ahmed A. Al-Ghamdi, Sami Ullah, and Ibrahim El-Tantawy El Sayed. "Experimental and Molecular Docking Studies of Cyclic Diphenyl Phosphonates as DNA Gyrase Inhibitors for Fluoroquinolone-Resistant Pathogens." Antibiotics 11, no. 1 (January 1, 2022): 53. http://dx.doi.org/10.3390/antibiotics11010053.
Full textAbstract:
DNA gyrase and topoisomerase IV are proven to be validated targets in the design of novel antibacterial drugs. In this study, we report the antibacterial evaluation and molecular docking studies of previously synthesized two series of cyclic diphenylphosphonates (1a–e and 2a–e) as DNA gyrase inhibitors. The synthesized compounds were screened for their activity (antibacterial and DNA gyrase inhibition) against ciprofloxacin-resistant E.coli and Klebsiella pneumoniae clinical isolates having mutations (deletion and substitution) in QRDR region of DNA gyrase. The target compound (2a) that exhibited the most potent activity against ciprofloxacin Gram-negative clinical isolates was selected to screen its inhibitory activity against DNA gyrase displayed IC50 of 12.03 µM. In addition, a docking study was performed with inhibitor (2a), to illustrate its binding mode in the active site of DNA gyrase and the results were compatible with the observed inhibitory potency. Furthermore, the docking study revealed that the binding of inhibitor (2a) to DNA gyrase is mediated and modulated by divalent Mg2+ at good binding energy (–9.08 Kcal/mol). Moreover, structure-activity relationships (SARs) demonstrated that the combination of hydrazinyl moiety in conjunction with the cyclic diphenylphosphonate based scaffold resulted in an optimized molecule that inhibited the bacterial DNA gyrase by its detectable effect in vitro on gyrase-catalyzed DNA supercoiling activity.
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Rajotte, D., S. Tremblay, A. Pelletier, P. Salois, L. Bourgon, R. Coulombe, S. Mason, L. Lamorte, C. F. Sturino, and R. Bethell. "Identification and Characterization of a Novel HIV-1 Nucleotide-Competing Reverse Transcriptase Inhibitor Series." Antimicrobial Agents and Chemotherapy 57, no. 6 (April 1, 2013): 2712–18. http://dx.doi.org/10.1128/aac.00113-13.
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ABSTRACTSeveral groups have recently reported on the identification of nucleotide-competing reverse transcriptase inhibitors (NcRTIs), a new class of RT inhibitors. NcRTIs reversibly inhibit binding of the incoming nucleotide to the RT active site but do not act as chain terminators, unlike the nucleos(t)ide reverse transcriptase inhibitor (NRTI) class. We identified a novel benzo[4,5]furo[3,2,d]pyrimidin-2-one NcRTI chemical series. Structure-activity relationship evaluation of this series with both RT and viral replication assays led to the identification of compound A, a new NcRTI. Compound A inhibited HIV-1 RT in a primer extension assay (50% inhibitory concentration, 2.6 nM) but had no measurable activity against human DNA polymerase γ at 10 μM. It potently inhibited HIV-1 replicationin vitro(50% effective concentration, 1.5 nM). The antiviral potency of compound A was unaffected by the presence of nonnucleotide RT inhibitor (NNRTI) mutations tested (L100I, K103N/Y181C, V106A, or Y188L). Notably, viruses encoding K65R were hypersusceptible to inhibition by compound A. Compound A also retained full activity against viruses encoding M184V.In vitroselection for resistant virus to compound A led to the selection of a single substitution within RT: W153L. A recombinant virus encoding the RT W153L was highly resistant to compound A (fold change, 160). W153 is a highly conserved residue in HIV RT and has not been previously associated with drug resistance. In summary, a novel NcRTI series with optimized antiviral activity, minimal cross-resistance to existing RT inhibitor classes, and a distinct resistance profile has been discovered. These results further establish NcRTIs as an emerging class of antiretroviral agents.
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Zhang, Yilin, Yong Yan, Lufan Liang, Jie Feng, Xuejun Wang, Li Li, and Kewu Yang. "Halogen-Substituted Triazolethioacetamides as a Potent Skeleton for the Development of Metallo-β-Lactamase Inhibitors." Molecules 24, no. 6 (March 25, 2019): 1174. http://dx.doi.org/10.3390/molecules24061174.
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Metallo-β-lactamases (MβLs) are the target enzymes of β-lactam antibiotic resistance, and there are no effective inhibitors against MβLs available for clinic so far. In this study, thirteen halogen-substituted triazolethioacetamides were designed and synthesized as a potent skeleton of MβLs inhibitors. All the compounds displayed inhibitory activity against ImiS with an IC50 value range of 0.032–15.64 μM except 7. The chlorine substituted compounds (1, 2 and 3) inhibited NDM-1 with an IC50 value of less than 0.96 μM, and the fluorine substituted 12 and 13 inhibited VIM-2 with IC50 values of 38.9 and 2.8 μM, respectively. However, none of the triazolethioacetamides exhibited activity against L1 at inhibitor concentrations of up to 1 mM. Enzyme inhibition kinetics revealed that 9 and 13 are mixed inhibitors for ImiS with Ki values of 0.074 and 0.27μM using imipenem as the substrate. Docking studies showed that 1 and 9, which have the highest inhibitory activity against ImiS, fit the binding site of CphA as a replacement of ImiS via stable interactions between the triazole group bridging ASP120 and hydroxyl group bridging ASN233.
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Kitano, Mitsutaka, Atsuko Yamamoto, Takeshi Noshi, Makoto Kawai, Ryu Yoshida, Akihiko Sato, Takao Shishido, and Akira Naito. "Synergistic Antiviral Activity of S-033188/S-033447, a Novel Inhibitor of Influenza Virus Cap-Dependent Endonuclease, in Combination with Neuraminidase Inhibitors In Vitro." Open Forum Infectious Diseases 4, suppl_1 (2017): S371. http://dx.doi.org/10.1093/ofid/ofx163.910.
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Abstract Background S-033447, an active form of orally available prodrug S-033188, is a novel small molecule inhibitor of cap-dependent endonuclease that is essential for influenza virus transcription and replication. In this study, we evaluated the inhibitory effect of S-033188 in combination with neuraminidase inhibitors on the replication of influenza A/H1N1 virus in cultured cells. Methods The inhibitory effects of S-033447 in combination with NA inhibitors on the cytopathic effect of A/PR/8/34 strain in Madin–Darby canine kidney cells cultured for 2 days were tested and EC50 were determined. The combination index (CI), which were obtained when S-033188 and NA inhibitor were added at the closest ratio of each EC50 value, were used for the evaluation of these combinational effects (Table 1). CI values were calculated by the Chou and Talalay method, in which combinational effect were determined according to the criteria as follows: synergistic if CI ≤ 0.8, additive if 0.8 < CI < 1.2, and antagonistic if CI ≥ 1.2. CI = (DA/A + B)/DA + (DB/A + B)/DB + (DA/A + B × DB/A + B)/(DA × DB) DA: the EC50 of S-033447 DB: the EC50 of NA inhibitor DA/A + B: the concentration of S-033447 giving 50% inhibition in combination with NA inhibitor at the closest ratio of each EC50 value DB/A + B: the concentration of NA inhibitor giving 50% inhibition in combination with S-033447 at the closest ratio of each EC50 value Results All CI values were lower than 0.8, under the condition that both S-033447 and NA inhibitor (oseltamivir acid, zanamivir hydrate, laninamivir, or peramivir trihydrate) were added at the closest ratio of each EC50 value (Table 1). Conclusion S-033447 in combination with oseltamivir acid, zanamivir hydrate, laninamivir, or peramivir trihydrate synergistically inhibited the replication of influenza A/H1N1 virus in MDCK cells. Disclosures All authors: No reported disclosures.
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Yan, Daojing, Jiakun Xu, Xiang Wang, Jiaxing Zhang, Gang Zhao, Yingwu Lin, and Xiangshi Tan. "Spiro-Oxindole Skeleton Compounds Are Efficient Inhibitors for Indoleamine 2,3-Dioxygenase 1: An Attractive Target for Tumor Immunotherapy." International Journal of Molecular Sciences 23, no. 9 (April 23, 2022): 4668. http://dx.doi.org/10.3390/ijms23094668.
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Indoleamine 2,3-dioxygenase 1 (IDO1) is an attractive heme enzyme for its significant function in cancer immunotherapy. Potent IDO1 inhibitors have been discovered for decades, whereas no clinical drugs are used for cancer treatment up to now. With the goal of developing medically valuable IDO inhibitors, we performed a systematic study of SAR405838 analogs with a spiro-oxindole skeleton in this study. Based on the expression and purification of human IDO1, the inhibitory activity of spiro-oxindole skeleton compounds to IDO1 was evaluated by IC50 and Ki values. The results demonstrated that inhibitor 3 exhibited the highest IDO1 inhibitory activity with IC50 at 7.9 μM among all inhibitors, which is ~six-fold of the positive control (4−PI). Moreover, inhibitor 3 was found to have the most effective inhibition of IDO1 in MCF-7 cancer cells without toxic effects. Molecular docking analysis revealed that the hydrophobic interaction stabilized the binding of inhibitor 3 to the IDO1 active site and made an explanation for the uncompetitive mode of inhibitors. Therefore, this study provides valuable insights into the screen of more potent IDO1 inhibitors for cancer immunotherapy.
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Zhu, Yongxia, Xinyi Chen, Qiangsheng Zhang, Lihong Shi, Luoting Yu, and Hongtao Xiao. "Anti-tumor activity of SKLB-0322, a novel EZH2 covalent inhibitor, in ovarian cancer." Journal of Clinical Oncology 39, no. 15_suppl (May 20, 2021): e15052-e15052. http://dx.doi.org/10.1200/jco.2021.39.15_suppl.e15052.
Full textAbstract:
e15052 Background: Enhancer of Zeste Homolog 2 (EZH2) is the catalytic subunit of the Polycomb Repressive Complex 2 (PRC2) that regulate downstream target genes expression, and then promotes tumor cell proliferation, metastasis and drug resistance. EZH2 also performs some functions in a PRC2-independent manner. Most of reported EZH2 inhibitors are S-adenosyle-methionine (SAM)-competitive inhibitor, and are less selective for EZH2 close homolog EZH1, which resulted in safety concerns and insufficient efficacy. To obtain irreversible EZH2 inhibitor, a novel covalent inhibitor was developed and characterized. Methods: SKLB-0322 and its derivatives were designed, synthesized and confirmed as EZH2 covalent inhibitor by us. The anti-tumor activities of SKLB-0322 were investigated by MTT assay, flow cytometry, and western blot assay. The reversible analog of SKLB-0322 (SKLB-0322’) was used as negative control. Results: SKLB-0322 inhibited EZH2 methyltransferase activity with nanomolar potency, while the inhibitory activities of SKLB-0322’ was reduced. The mass spectrometry (MS) analyses revealed that SKLB-0322 could efficiently forms a single modified covalent adduct. SKLB-0322 displayed noteworthy potency against ovarian cancer cell lines at low micromolar level and reduced the expression level of H3K27me3 in a concentration-dependent manner, which was about 5-fold more active than the reversible negative control SKLB-0322’. Besides, SKLB-0322 caused G2/M phase cell cycle arrest in A2780 and PA-1 cells. Furthermore, SKLB-0322 induced A2780 and PA-1 cell apoptosis in a time- and concentration- dependent manner. Conclusions: Our data clarified that SKLB-0322 is an EZH2 covalent inhibitor for ovarian cancer therapy which is worthy of further evaluation.
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Jo, Sooheum, Jin-Hee Kim, Jiyeon Lee, Youngjun Park, and Jaebong Jang. "Azumamides A-E: Isolation, Synthesis, Biological Activity, and Structure–Activity Relationship." Molecules 27, no. 23 (December 2, 2022): 8438. http://dx.doi.org/10.3390/molecules27238438.
Full textAbstract:
Cyclic peptides are one of the important chemical groups in the HDAC inhibitor family. Following the success of romidepsin in the clinic, naturally occurring cyclic peptides with a hydrophilic moiety have been intensively studied to test their function as HDAC inhibitors. Azumamides A-E, isolated from Mycale izuensis, are one of the powerful HDAC inhibitor classes. Structurally, azumamides A-E consist of three D-α-amino acids and unnatural β-amino acids such as 3-amino-2-methyl-5-nonenedioic acid-9-amide (Amnna) and 3-amino-2-methyl-5-nonenoic-1,9-diacid (Amnda). Moreover, azumamides have a retro-arrangement peptide backbone, unlike other naturally occurring cyclopeptide HDAC inhibitors, owing to the D-configuration of all residues. This review summarizes the currently available synthetic methods of azumamides A-E focusing on the synthesis of β-amino acids and macrocyclization. In addition, we overview the structure–activity relationship of azumamides A-E based on reported analogs. Collectively, this review highlights the potentiality of azumamides A-E as an HDAC inhibitor and provides further developmental insight into naturally occurring cyclic peptides in HDAC inhibition.
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Arita, Minetaro, Takaji Wakita, and Hiroyuki Shimizu. "Cellular kinase inhibitors that suppress enterovirus replication have a conserved target in viral protein 3A similar to that of enviroxime." Journal of General Virology 90, no. 8 (August 1, 2009): 1869–79. http://dx.doi.org/10.1099/vir.0.012096-0.
Full textAbstract:
Previously, we identified a cellular kinase inhibitor, GW5074, that inhibits poliovirus (PV) and enterovirus 71 replication strongly, although its target has remained unknown. To identify the target of GW5074, we searched for cellular kinase inhibitors that have anti-enterovirus activity similar or related to that of GW5074. With this aim, we performed screenings to identify cellular kinase inhibitors that could inhibit PV replication cooperatively with GW5074 or synthetically in the absence of GW5074. We identified MEK1/2 inhibitors (SL327 and U0126), an EGFR inhibitor (AG1478) and a phosphatidylinositol 3-kinase inhibitor (wortmannin) as compounds with a cooperative inhibitory effect with GW5074, and an Akt1/2 inhibitor (Akt inhibitor VIII) as a compound with a synthetic inhibitory effect with MEK1/2 inhibitors and AG1478. Individual treatment with the identified kinase inhibitors did not affect PV replication significantly, but combined treatment with MEK1/2 inhibitor, AG1478 and Akt1/2 inhibitor suppressed the replication synthetically. The effect of AG1478 in this synthetic inhibition was compensated by other receptor tyrosine kinase inhibitors (IGF-1R inhibitor II and Flt3 inhibitor II). We isolated mutants resistant to Flt3 inhibitor II and GW5074 and found that these mutants had cross-resistance to each treatment. These mutants had a common mutation in viral protein 3A that results in an amino acid change at position 70 (Ala to Thr), a mutation that was previously identified in mutants resistant to a potent anti-enterovirus compound, enviroxime. These results suggest that cellular kinase inhibitors and enviroxime have a conserved target in viral protein 3A to suppress enterovirus replication.