Journal articles on the topic 'Inhibitor'
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1
Sever, Natasa, Metka Filipic, Joze Brzin, and Tamara T. Lah. "Effect of Cysteine Proteinase Inhibitors on Murine B16 Melanoma Cell Invasion in vitro." Biological Chemistry 383, no. 5 (May 15, 2002): 839–42. http://dx.doi.org/10.1515/bc.2002.088.
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Abstract Various types of proteinases are implicated in the malignant progression of human and animal tumors. Proteinase inhibitors may therefore be useful as therapeutic agents in antiinvasive and antimetastatic treatment. The aims of this study were (1) to estimate the relative importance of proteinases in B16 cell invasion in vitro using synthetic, classspecific proteinase inhibitors and (2) to assess the inhibitory effect of some naturally occurring cysteine proteinase inhibitors. Serine proteinase inhibitor reduced invasiveness by up to 24%, whereas inhibition of aspartic proteinases reduced invasion by 11%. Synthetic inhibitors of cysteine proteinases markedly impaired invasion: cathepsin B inhibitors, particularly Ca 074Me, inhibited invasion from 20 40%, whereas cathepsin L inhibitor Clik 148 reduced invasion by 11%. The potato cysteine proteinase inhibitor PCPI 8.7 inhibited invasion by 21%, whereas another potato inhibitor, PCPI 6.6, and the mushroom cysteine proteinase inhibitor clitocypin had no effects. As the inhibitors that inhibited cathepsin B were in general more efficient at impairing the invasiveness, we conclude that of the two cysteine proteinases, cathepsin B plays a more important role than cathepsin L in murine melanoma cell invasion.
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Jasim, Haider Hadi, Read Abd Al-Hussain, and Ahmed Shawqi Sadeq. "Evaluation the Efficiency of Various Types of Corrosion Inhibitors Used for Basrah Water Storage Tanks." Al-Nahrain Journal for Engineering Sciences 23, no. 3 (November 21, 2020): 267–76. http://dx.doi.org/10.29194/njes.23030267.
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In this paper, the efficiency of six different types of corrosion inhibitors used in Basrah drinking water tanks was assessed using a potentiostatic test method. The mechanism of adsorption of silicate and phosphate inhibitors in AISI 316 stainless steel surfaces and the effects of different water components in inhibitors are discussed in detail. The values of corrosion rate obtained from the Potentiostatic test showed that the protection against corrosion in the presence of inhibitors is better compared to the case of absence of inhibitors. The results of the six types of corrosion inhibitors tested showed that the inhibitory efficacy is higher below the temperatures 45oC, but when raise the temperature above 45oC the inhibitory efficiency becomes to decrease. Also, the test results indicated that the corrosion inhibitor involves silicate products provided more inhibited efficiency compared to the phosphate inhibitor alone or used the combined silicate/phosphate corrosion inhibitor. The inspection of the surface of the tested samples using optical methods shows that the pitting corrosion is demonstrated on the specimen surfaces after testing with or without inhibitors.
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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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Krulikas, Linas J., Ian M. McDonald, Benjamin Lee, Denis O. Okumu, Michael P. East, Thomas S. K. Gilbert, Laura E. Herring, et al. "Application of Integrated Drug Screening/Kinome Analysis to Identify Inhibitors of Gemcitabine-Resistant Pancreatic Cancer Cell Growth." SLAS DISCOVERY: Advancing the Science of Drug Discovery 23, no. 8 (May 9, 2018): 850–61. http://dx.doi.org/10.1177/2472555218773045.
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Continuous exposure of a pancreatic cancer cell line MIA PaCa-2 (MiaS) to gemcitabine resulted in the formation of a gemcitabine-resistant subline (MiaR). In an effort to discover kinase inhibitors that inhibited MiaR growth, MiaR cells were exposed to kinase inhibitors (PKIS-1 library) in a 384-well screening format. Three compounds (UNC10112721A, UNC10112652A, and UNC10112793A) were identified that inhibited the growth of MiaR cells by more than 50% (at 50 nM). Two compounds (UNC10112721A and UNC10112652A) were classified as cyclin-dependent kinase (CDK) inhibitors, whereas UNC10112793A was reported to be a PLK inhibitor. Dose–response experiments supported the efficacy of these compounds to inhibit growth and increase apoptosis in 2D cultures of these cells. However, only UNC10112721A significantly inhibited the growth of 3D spheroids composed of MiaR cells and GFP-tagged cancer-associated fibroblasts. Multiplexed inhibitor bead (MIB)–mass spectrometry (MS) kinome competition experiments identified CDK9, CLK1-4, DYRK1A, and CSNK1 as major kinase targets for UNC10112721A in MiaR cells. Another CDK9 inhibitor (CDK-IN-2) replicated the growth inhibitory effects of UNC10112721A, whereas inhibitors against the CLK, DYRK, or CSNK1 kinases had no effect. In summary, these studies describe a coordinated approach to discover novel kinase inhibitors, evaluate their efficacy in 3D models, and define their specificity against the kinome.
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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.
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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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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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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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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.
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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.
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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.
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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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Moon, Hyun-Jung, Su-Hoon Lee, Hak-Bong Kim, Kyoung-A. Lee, Chi-Dug Kang, and Sun-Hee Kim. "SIRT1 Inhibitor Enhances Hsp90 Inhibitor-mediated Abrogation of Hsp90 Chaperone Function and Potentiates the Cytotoxicity of Hsp90 Inhibitor in Chemo-resistant Human Cancer Cells." Journal of Life Science 26, no. 7 (July 30, 2016): 826–34. http://dx.doi.org/10.5352/jls.2016.26.7.826.
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Okamoto, Mika, Takashi Okamoto, and Masanori Baba. "Inhibition of Human Immunodeficiency Virus Type 1 Replication by Combination of Transcription Inhibitor K-12 and Other Antiretroviral Agents in Acutely and Chronically Infected Cells." Antimicrobial Agents and Chemotherapy 43, no. 3 (March 1, 1999): 492–97. http://dx.doi.org/10.1128/aac.43.3.492.
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ABSTRACT 8 - Difluoromethoxy - 1 - ethyl - 6 - fluoro - 1,4 - dihydro - 7 - [4 - (2 - methoxyphenyl) - 1 - piperazinyl] - 4 - oxoquinoline - 3 - carboxylic acid (K-12) has recently been identified as a potent and selective inhibitor of human immunodeficiency virus type 1 (HIV-1) transcription. In this study, we examined several combinations of K-12 and other antiretroviral agents for their inhibitory effects on HIV-1 replication in acutely and chronically infected cell cultures. Combinations of K-12 and a reverse transcriptase (RT) inhibitor, either zidovudine, lamivudine, or nevirapine, synergistically inhibited HIV-1 replication in acutely infected MT-4 cells. The combination of K-12 and the protease inhibitor nelfinavir (NFV) also synergistically inhibited HIV-1, whereas the synergism of this combination was weaker than that of the combinations with the RT inhibitors. K-12 did not enhance the cytotoxicities of RT and protease inhibitors. Synergism of the combinations was also observed in acutely infected peripheral blood mononuclear cells. The combination of K-12 and cepharanthine, a nuclear factor κB inhibitor, synergistically inhibited HIV-1 production in tumor necrosis factor alpha-stimulated U1 cells, a promonocytic cell line chronically infected with the virus. In contrast, additive inhibition was observed for the combination of K-12 and NFV. These results indicate that the combinations of K-12 and clinically available antiretroviral agents may have potential as chemotherapeutic modalities for the treatment of HIV-1 infection.
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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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Shi, Qizhen, Erin L. Kuether, Jocelyn A. Schroeder, Crystal L. Perry, Scot A. Fahs, and Robert R. Montgomery. "Factor VIII Inhibitors: Von Willebrand Factor Makes A Difference In Vitro and In Vivo." Blood 116, no. 21 (November 19, 2010): 709. http://dx.doi.org/10.1182/blood.v116.21.709.709.
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Abstract Abstract 709 The important association between von Willebrand factor (VWF) and factor VIII (FVIII) has been investigated for decades, but the effect of VWF on FVIII inhibitors is still controversial. Studies have demonstrated that some anti-FVIII inhibitory antibodies inhibit VWF-FVIII interaction, while others rely on the presence of VWF to inhibit FVIII activities. The influence of VWF on the Bethesda assay, which is routinely used in the clinic to determine the titer of FVIII-neutralizing inhibitors, is still uncertain because the plasma from hemophilia A patients with inhibitors contains normal levels of VWF. To explore the effect of VWF on the reactivity of FVIII inhibitors, we immunized VWF and FVIII double knockout (VWFnullFVIIInull) mice with recombinant human B-domain deleted FVIII (rhFVIII) to induce anti-FVIII inhibitory antibody development. Inhibitory plasma was collected and the titer of inhibitors was determined by Bethesda assay. Murine plasma-derived VWF (from FVIIInull mice) or recombinant human VWF (rhVWF) was used to study the influence of VWF on inhibitor inactivation of FVIII activity (FVIII:C). The remaining FVIII:C after inactivation was determined by chromogenic assay. When inhibitory plasma was incubated with rhFVIII in the presence of 1 U/ml VWF, the residual FVIII activity recovered was higher than in the absence of VWF, resulting in 6.82 ± 1.12 (n = 27) fold lower apparent inhibitor titers. This protective effect is VWF dose dependent. The source of VWF (plasma-derived murine VWF vs. rhVWF) did not affect its protection of FVIII from inhibitor inactivation and VWF does not affect FVIII:C measured in the chromogenic assay in the absence of inhibitors. Interestingly, we found that inhibitor inactivation of FVIII:C in the absence of VWF occurred much faster than in its presence. When the usual 2 hr. incubation at 37°C was omitted from the Bethesda assay, adding rhVWF to rFVIII before mixing with inhibitory plasma resulted in 67.29 ± 20.18 (n = 5) fold lower apparent inhibitor titers than without added VWF. In contrast, if VWF was added to inhibitory plasma first and then mixed with rhFVIII, the inhibitor titers were only 11.04 ± 3.56 (n = 5) fold lower than without added VWF. These results indicate that rhFVIII present in a preformed VWF-FVIII complex is protected from inhibitory antibody inactivation. Conversely, when VWF and inhibitory plasma are added to rhFVIII at the same time, the VWF and inhibitors appear to compete to bind to rhFVIII. Inhibitor titers were lower than in the absence of VWF, but the protective effect is not as efficient as when VWF and rhFVIII were already associated with one another before encountering inhibitors. To confirm the protective effect of VWF on FVIII from inhibitor inactivation, we infused FVIIInull or VWFnullFVIIInull mice with inhibitory plasma and rhFVIII followed by a tail clip survival test. When rhFVIII was infused into FVIIInull mice to 2% followed by inhibitory plasma infusion, all mice with inhibitor titer of 2.5 BU/ml (n = 4) survived tail clipping, and 2 of 4 survived with either 25 BU/ml or 250 BU/ml. If inhibitory plasma was infused first followed by rhFVIII infusion, then only 2 of 6 mice with inhibitor titers of 2.5 BU/ml survived tail clip challenge and none survived with 25 BU/ml and 250 BU/ml. In the first set of mice the infused FVIII was able to form a protective complex with endogenous VWF before encountering inhibitors, while in the second set FVIII is exposed to VWF and pre-infused inhibitory antibodies at the same time, a competitive binding that appears to reduce VWF's protective effect. In contrast, when rhFVIII was infused into VWFnullFVIIInull mice followed by inhibitory plasma infusion, no animals (n = 4 for each group) survived tail clipping with inhibitor titers of 2.5 BU/ml or higher. In summary, our studies demonstrate that VWF exerts a protective effect, reducing inhibitor inactivation of FVIII, both in vitro and in vivo. While the role of VWF in stabilizing plasma FVIII in a milieu rich in proteases has been appreciated for decades, our results indicate that treatment utilizing products containing a complex of FVIII with VWF may be especially beneficial in hemophilia A patients with inhibitors. Disclosures: No relevant conflicts of interest to declare.
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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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Cunningham, Rochelle, Rajatsubhra Biswas, Marc Brazie, Deborah Steplock, Shirish Shenolikar, and Edward J. Weinman. "Signaling pathways utilized by PTH and dopamine to inhibit phosphate transport in mouse renal proximal tubule cells." American Journal of Physiology-Renal Physiology 296, no. 2 (February 2009): F355—F361. http://dx.doi.org/10.1152/ajprenal.90426.2008.
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The present experiments were designed to detail factors regulating phosphate transport in cultured mouse proximal tubule cells by determining the response to parathyroid hormone (PTH), dopamine, and second messenger agonists and inhibitors. Both PTH and dopamine inhibited phosphate transport by over 30%. The inhibitory effect of PTH was completely abolished in the presence of chelerythrine, a PKC inhibitor, but not by Rp-cAMP, a PKA inhibitor. By contrast, both chelerythrine and Rp-cAMP blocked the inhibitory effect of dopamine. Chelerythrine inhibited PTH-mediated cAMP accumulation but also blocked the inhibitory effect of 8-bromo-cAMP on phosphate transport. On the other hand, Rp-cAMP had no effect on the ability of DOG, a PKC activator, to inhibit phosphate transport. PD98059, an inhibitor of MAPK, had no effect on PTH- or dopamine-mediated inhibition of sodium-phosphate cotransport. Finally, compared with 8-bromo-cAMP, 8-pCPT-2′- O-Me-cAMP, an activator of EPAC, had no effect on phosphate transport. These results outline significant differences in the signaling pathways utilized by PTH and dopamine to inhibit renal phosphate transport. Our results also suggest that activation of MAPK is not critically involved in PTH- or dopamine-mediated inhibition of phosphate transport in mouse renal proximal tubule cells in culture.
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Honma, Masaru, Mark Stubbs, Ian Collins, Paul Workman, Wynne Aherne, and Fiona M. Watt. "Identification of Novel Keratinocyte Differentiation Modulating Compounds by High-Throughput Screening." Journal of Biomolecular Screening 11, no. 8 (December 2006): 977–84. http://dx.doi.org/10.1177/1087057106292556.
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The authors have designed high-throughput screens to identify compounds that promote or inhibit terminal differentiation of primary human epidermal keratinocytes. Eleven known inhibitors of signaling pathways and approximately 4000 compounds of diverse structure were screened using an In-Cell Western system based on immunofluorescent staining of the terminal differentiation marker, involucrin. Staurosporine, a nonspecific protein kinase C inhibitor, and H89, a protein kinase A inhibitor, promoted expression of involucrin. Conversely, U0126, a MEK inhibitor, and SAHA or SBHA, 2 histone deacetylase inhibitors, reduced the expression of involucrin during calcium-induced stratification. In addition, the authors found 1 novel compound that induced keratinocyte differentiation and 2 novel compounds that were inhibitory to calcium-induced differentiation. The differentiation-inducing compound also inhibited growth of a human squamous cell carcinoma line by stimulating both differentiation and apoptosis. Because the compound affected the tumor cells at a lower concentration than primary keratinocytes, it may have potential as an antitumor therapy.
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Potempa, J., J. J. Enghild, and J. Travis. "The primary elastase inhibitor (elastasin) and trypsin inhibitor (contrapsin) in the goat are serpins related to human α1-anti-chymotrypsin." Biochemical Journal 306, no. 1 (February 15, 1995): 191–97. http://dx.doi.org/10.1042/bj3060191.
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Two primary serine proteinase inhibitors in goat plasma have been isolated and characterized. The N-terminal sequence analysis of the purified proteins revealed that they are closely related to each other and are highly homologous to human alpha 1-anti-chymotrypsin rather than alpha 1-proteinase inhibitor. However, despite structural similarities the inhibitory specificity of the goat inhibitors differed from each other and from that of anti-chymotrypsin. In contrast with human anti-chymotrypsin, one of the goat inhibitors was shown to be a strong and specific inhibitor of trypsin (k(ass.) = 1.9 x 10(6) M-1.s-1), whereas the other was an efficient inhibitor of neutrophil elastase (k(ass.) = 1.5 x 10(6) M-1.S-1). Differences in the inhibitory specificity of each protein could readily be attributed to the amino acid sequence within the reactive site region. The trypsin inhibitor with an assumed arginine residue at the P1 position of the reactive-site peptide bond is referred to as ‘contrapsin’, and indicates that the occurrence of contrapsins is not restricted to rodents. In contrast, the inhibitory specificity, resistance to oxidative and proteolytic inactivation and the presence of a P1 leucine residue in the elastase inhibitor is unique among inhibitory serpins that have been characterized to date. Because this serpin is apparently the major elastase inhibitor in goat plasma, it is likely to be involved in the control of goat neutrophil elastase. Therefore, we suggest the name ‘elastasin’, and extend it to any other anti-chymotrypsin related serpins possessing neutrophil-elastase- inhibitory activity.
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Tang, Xiaopeng, Mengrou Chen, Zilei Duan, James Mwangi, Pengpeng Li, and Ren Lai. "Isolation and Characterization of Poecistasin, an Anti-Thrombotic Antistasin-Type Serine Protease Inhibitor from Leech Poecilobdella manillensis." Toxins 10, no. 11 (October 26, 2018): 429. http://dx.doi.org/10.3390/toxins10110429.
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Antistasin, first identified as a potent inhibitor of the blood coagulation factor Xa, is a novel family of serine protease inhibitors. In this study, we purified a novel antistasin-type inhibitor from leech Poecilobdella manillensis called poecistasin. Amino acid sequencing of this 48-amino-acid protein revealed that poecistasin was an antistasin-type inhibitor known to consist of only one domain. Poecistasin inhibited factor XIIa, kallikrein, trypsin, and elastase, but had no inhibitory effect on factor Xa and thrombin. Poecistasin showed anticoagulant activities. It prolonged the activated partial thromboplastin time and inhibited FeCl3-induced carotid artery thrombus formation, implying its potent function in helping Poecilobdella manillensis to take a blood meal from the host by inhibiting coagulation. Poecistasin also suppressed ischemic stroke symptoms in transient middle cerebral artery occlusion mice model. Our results suggest that poecistasin from the leech Poecilobdella manillensis plays a crucial role in blood-sucking and may be an excellent candidate for the development of clinical anti-thrombosis and anti-ischemic stroke medicines.
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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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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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Kim, Min-Jeong, Sarita Pandit, and Jun-Goo Jee. "Discovery of Kinase and Carbonic Anhydrase Dual Inhibitors by Machine Learning Classification and Experiments." Pharmaceuticals 15, no. 2 (February 16, 2022): 236. http://dx.doi.org/10.3390/ph15020236.
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A multi-target small molecule modulator is advantageous for treating complicated diseases such as cancers. However, the strategy and application for discovering a multi-target modulator have been less reported. This study presents the dual inhibitors for kinase and carbonic anhydrase (CA) predicted by machine learning (ML) classifiers, and validated by biochemical and biophysical experiments. ML trained by CA I and CA II inhibitor molecular fingerprints predicted candidates from the protein-specific bioactive molecules approved or under clinical trials. For experimental tests, three sulfonamide-containing kinase inhibitors, 5932, 5946, and 6046, were chosen. The enzyme assays with CA I, CA II, CA IX, and CA XII have allowed the quantitative comparison in the molecules’ inhibitory activities. While 6046 inhibited weakly, 5932 and 5946 exhibited potent inhibitions with 100 nM to 1 μM inhibitory constants. The ML screening was extended for finding CAs inhibitors of all known kinase inhibitors. It found XMU-MP-1 as another potent CA inhibitor with an approximate 30 nM inhibitory constant for CA I, CA II, and CA IX. Differential scanning fluorimetry confirmed the direct interaction between CAs and small molecules. Cheminformatics studies, including docking simulation, suggest that each molecule possesses two separate functional moieties: one for interaction with kinases and the other with CAs.
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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.
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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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Lange, Falko, Benjamin Franz, Claudia Maletzki, Michael Linnebacher, Maja Hühns, and Robert Jaster. "Biological and Molecular Effects of Small Molecule Kinase Inhibitors on Low-Passage Human Colorectal Cancer Cell Lines." BioMed Research International 2014 (2014): 1–13. http://dx.doi.org/10.1155/2014/568693.
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Low-passage cancer cell lines are versatile tools to study tumor cell biology. Here, we have employed four such cell lines, established from primary tumors of colorectal cancer (CRC) patients, to evaluate effects of the small molecule kinase inhibitors (SMI) vemurafenib, trametinib, perifosine, and regorafenib in anin vitrosetting. The mutantBRAF(V600E/V600K) inhibitor vemurafenib, but also the MEK1/2 inhibitor trametinib efficiently inhibited DNA synthesis, signaling through ERK1/2 and expression of genes downstream of ERK1/2 inBRAFmutant cells only. In case of the AKT inhibitor perifosine, three cell lines showed a high or intermediate responsiveness to the drug while one cell line was resistant. The multikinase inhibitor regorafenib inhibited proliferation of all CRC lines with similar efficiency and independent of the presence or absence ofKRAS, BRAF, PIK3CA, andTP53mutations. Regorafenib action was associated with broad-range inhibitory effects at the level of gene expression but not with a general inhibition of AKT or MEK/ERK signaling. In vemurafenib-sensitive cells, the antiproliferative effect of vemurafenib was enhanced by the other SMI. Together, our results provide insights into the determinants of SMI efficiencies in CRC cells and encourage the further use of low-passage CRC cell lines as preclinical models.
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Cheng, Liwei, Limin Wang, Zhi Li, Bei Liu, and Guangjin Chen. "Inhibition Effect of Kinetic Hydrate Inhibitors on the Growth of Methane Hydrate in Gas–Liquid Phase Separation State." Energies 12, no. 23 (November 25, 2019): 4482. http://dx.doi.org/10.3390/en12234482.
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The effect of kinetic hydrate inhibitors (KHIs) on the growth of methane hydrate in the gas–liquid phase separation state is studied at the molecular level. The simulation results show that the kinetic inhibitors, named PVP and PVP-A, show good inhibitory effects on the growth of methane hydrate under the gas–liquid phase separation state, and the initial position of the kinetic hydrate inhibitors has a major effect on the growth of methane hydrates. In addition, inhibitors at different locations exhibit different inhibition performances. When the inhibitor molecules are located at the gas–liquid phase interface, increasing the contact area between the groups of the inhibitor molecules and methane is beneficial to enhance the inhibitory performance of the inhibitors. When inhibitor molecules are located at the solid–liquid phase interface, the inhibitor molecules adsorbed on the surface of the hydrate nucleus and decreased the direct contact of hydrate nucleus with the surrounding water and methane molecules, which would delay the growth of hydrate nucleus. Moreover, the increase of hydrate surface curvature and the Gibbs–Thomson effect caused by inhibitors can also reduce the growth rate of methane hydrate.
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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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Yoo Im, Sonia, Camila Ramalho Bonturi, Adriana Miti Nakahata, Clóvis Ryuichi Nakaie, Arnildo Pott, Vali Joana Pott, and Maria Luiza Vilela Oliva. "Differences in the Inhibitory Specificity Distinguish the Efficacy of Plant Protease Inhibitors on Mouse Fibrosarcoma." Plants 10, no. 3 (March 23, 2021): 602. http://dx.doi.org/10.3390/plants10030602.
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Metastasis, the primary cause of death from malignant tumors, is facilitated by multiple protease-mediated processes. Thus, effort has been invested in the development of protease inhibitors to prevent metastasis. Here, we investigated the effects of protease inhibitors including the recombinant inhibitors rBbKI (serine protease inhibitor) and rBbCI (serine and cysteine inhibitor) derived from native inhibitors identified in Bauhinia bauhinioides seeds, and EcTI (serine and metalloprotease inhibitor) isolated from the seeds of Enterolobium contortisiliquum on the mouse fibrosarcoma model (lineage L929). rBbKI inhibited 80% of cell viability of L929 cells after 48 h, while EcTI showed similar efficacy after 72 h. Both inhibitors acted in a dose and time-dependent manner. Conversely, rBbCI did not significantly affect the viability of L929 cells. Confocal microscopy revealed the binding of rBbKI and EcTI to the L929 cell surface. rBbKI inhibited approximately 63% of L929 adhesion to fibronectin, in contrast with EcTI and rBbCI, which did not significantly interfere with adhesion. None of the inhibitors interfered with the L929 cell cycle phases. The synthetic peptide RPGLPVRFESPL-NH2, based on the BbKI reactive site, inhibited 45% of the cellular viability of L929, becoming a promising protease inhibitor due to its ease of synthesis.
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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.
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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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Luan, Hengjie, Mingkang Liu, Qinglin Shan, Yujing Jiang, Peng Yan, and Xiaoyu Du. "Experimental Study on the Effect of Mixed Thermodynamic Inhibitors with Different Concentrations on Natural Gas Hydrate Synthesis." Energies 17, no. 9 (April 26, 2024): 2078. http://dx.doi.org/10.3390/en17092078.
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Natural gas hydrate (NGH) is a potential future energy resource. More than 90% of NGH resources exist in the pore medium of seafloor sediments. During the development of deep-sea oil and gas fields, wellbore pipelines are often clogged due to the synthesis of gas hydrates, and the addition of thermodynamic inhibitors is a common solution to prevent hydrate synthesis. In this paper, the effects of two single inhibitors, sodium chloride and ethylene glycol, as well as hybrid inhibitors combining these two inhibitors on the synthesis of methane hydrates were investigated using the self-developed one-dimensional gas hydrate exploitation simulation test apparatus. The effects of single and hybrid inhibitors were investigated in terms of the hydrate synthesis volume and gas–water two-phase conversion rate. The results show that the hybrid inhibitor has a better inhibitory effect on hydrate synthesis with the same initial synthesis driving force. When the concentration of inhibitors is low, salt inhibitors can have a better inhibitory effect than alcohol inhibitors. However, in the mixed inhibitor experiment, increasing the proportion of ethylene glycol in the mixed inhibitor can more effectively inhibit the synthesis of hydrates than increasing the proportion of sodium chloride in the mixed inhibitor.
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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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Chen, Chang, Jin-Lai Gao, Ming-Yu Liu, Shan-Liang Li, Xiu-Chen Xuan, Xin-Zi Zhang, Xi-Yue Zhang, et al. "Mitochondrial Fission Inhibitors Suppress Endothelin-1-Induced Artery Constriction." Cellular Physiology and Biochemistry 42, no. 5 (2017): 1802–11. http://dx.doi.org/10.1159/000479536.
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Background/Aims: Endothelin-1 is implicated in the pathogenesis of hypertension, but the underlying mechanisms remained elusive. Our previous study found that inhibition of mitochondrial fission of smooth muscle cells suppressed phenylephrine- and high K+-induced artery constriction. Here, we studied the effects of mitochondrial fission inhibitors on endothelin-1-induced vasoconstriction. Methods: The tension of rat mesenteric arteries and thoracic aorta was measured by using a multi-wire myograph system. Mitochondrial morphology of aortic smooth muscle cells was observed by using transmission electron microscopy. Results: Dynamin-related protein-1 selective inhibitor mdivi-1 relaxed endothelin-1-induced constriction, and mdivi-1 pre-treatment prevented endothelin-1-induced constriction of rat mesenteric arteries with intact and denuded endothelium. Mdivi-1 had a similar inhibitory effect on rat thoracic aorta. Another mitochondrial fission inhibitor dynasore showed similar effects as mdivi-1 in rat mesenteric arteries. Mdivi-1 inhibited endothelin-1-induced increase of mitochondrial fission in smooth muscle cells of rat aorta. Rho-associated protein kinase inhibitor Y-27632 which relaxed endothelin-1-induced vasoconstriction inhibited endothelin-1-induced mitochondrial fission in smooth muscle cells of rat aorta. Conclusion: Endothelin-1 increases mitochondrial fission in vascular smooth muscle cells, and mitochondrial fission inhibitors suppress endothelin-1-induced vasoconstriction.
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Seo, Songyi, Mi-Kyung Kim, Ryul-I. Kim, Yeongju Yeo, Koung Li Kim, and Wonhee Suh. "Evogliptin, a dipeptidyl peptidase-4 inhibitor, attenuates pathological retinal angiogenesis by suppressing vascular endothelial growth factor-induced Arf6 activation." Experimental & Molecular Medicine 52, no. 10 (October 2020): 1744–53. http://dx.doi.org/10.1038/s12276-020-00512-8.
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Abstract Dipeptidyl peptidase-4 (DPP-4) inhibitors are used for the treatment of type 2 diabetes mellitus (DM). Recent studies have shown that beyond their effect in lowing glucose, DPP-4 inhibitors mitigate DM-related microvascular complications, such as diabetic retinopathy. However, the mechanism by which pathological retinal neovascularization, a major clinical manifestation of diabetic retinopathy, is inhibited is unclear. This study sought to examine the effects of evogliptin, a potent DPP-4 inhibitor, on pathological retinal neovascularization in mice and elucidate the mechanism by which evogliptin inhibits angiogenesis mediated by vascular endothelial growth factor (VEGF), a key factor in the vascular pathogenesis of proliferative diabetic retinopathy (PDR). In a murine model of PDR, an intravitreal injection of evogliptin significantly suppressed aberrant retinal neovascularization. In human endothelial cells, evogliptin reduced VEGF-induced angiogenesis. Western blot analysis showed that evogliptin inhibited the phosphorylation of signaling molecules associated with VEGF-induced cell adhesion and migration. Moreover, evogliptin substantially inhibited the VEGF-induced activation of adenosine 5′-diphosphate ribosylation factor 6 (Arf6), a small guanosine 5′-triphosphatase (GTPase) that regulates VEGF receptor 2 signal transduction. Direct activation of Arf6 using a chemical inhibitor of Arf-directed GTPase-activating protein completely abrogated the inhibitory effect of evogliptin on VEGF-induced activation of the angiogenic signaling pathway, which suggests that evogliptin suppresses VEGF-induced angiogenesis by blocking Arf6 activation. Our results provide insights into the molecular mechanism of the direct inhibitory effect of the DPP-4 inhibitor evogliptin on pathological retinal neovascularization. In addition to its glucose-lowering effect, the antiangiogenic effect of evogliptin could also render it beneficial for individuals with PDR.
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Savitri, Erna Noor, Putut Marwoto, and Sunyoto Eko Nugroho. "The Effectiveness of a Combination of Lime (Citrus aurantifolia.), Lerak (Sapindus rarak) and Jasmine Flower (Jasminum nudiflorum) Extracts as and Environmentally Friendly Corrosion Inhibitor." Jurnal Penelitian Pendidikan IPA 10, no. 3 (March 30, 2024): 1019–24. http://dx.doi.org/10.29303/jppipa.v10i3.6364.
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Corrosion can also be defined as the forced destruction of metal by the surrounding medium which is usually a liquid (corrosive agent). Corrosion prevention processes can be carried out, including by coating the metal surface, cathodic protection, adding corrosion inhibitors and so on. Several types of inhibitors that have been widely used in industrial applications are synthetic chemical inhibitors. However, the compounds of inhibitors are environmentally unfriendly, toxic and expensive. To overcome this problem, it is necessary to develop an environmentally friendly alternative corrosion inhibitor or better known as a green inhibitor. This research aims to test the effectiveness of using a combination of lime (Citrus aurantifolia.), Lerak (Sapindus rarak) and jasmine flower (Jasminum nudiflorum) extracts as an environmentally friendly corrosion inhibitor. The results obtained were inhibitors in the form of a combination which were added to the corrosive medium HCl which could reduce the rate of aluminum corrosion. This research also shows that time and concentration influence the corrosion rate. A higher concentration (200 ppm) has a greater inhibitory power than a concentration of 100 ppm. The best inhibitory power is found in 200 ppm inhibitor with a soaking time of 20 minutes
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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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Kim, Dong Wook, Young Suk Jo, Hye Sook Jung, Hyo Kyun Chung, Jung Hun Song, Ki Cheol Park, Su Hyeon Park, et al. "An Orally Administered Multitarget Tyrosine Kinase Inhibitor, SU11248, Is a Novel Potent Inhibitor of Thyroid Oncogenic RET/Papillary Thyroid Cancer Kinases." Journal of Clinical Endocrinology & Metabolism 91, no. 10 (October 1, 2006): 4070–76. http://dx.doi.org/10.1210/jc.2005-2845.
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Abstract Context: The oncogenic RET/PTC tyrosine kinase causes papillary thyroid cancer (PTC). The use of inhibitors specific for RET/PTC may be useful for targeted therapy of PTC. Objective: The objective of the study was to evaluate the efficacies of the recently developed kinase inhibitors SU11248, SU5416, and SU6668 in inhibition of RET/PTC. Design: SU11248, SU5416, and SU6668 were synthesized, and their inhibitory potencies were evaluated using an in vitro RET/PTC kinase assay. The inhibitory effects of the compounds on RET/PTC were evaluated by quantifying the autophosphorylation of RET/PTC, signal transducer and activator of transcription (STAT)-3 activation, and the morphological reversal of RET/PTC-transformed cells. Results: An in vitro kinase assay revealed that SU5416, SU6668, and SU11248 inhibited phosphorylation of the synthetic tyrosine kinase substrate peptide E4Y by RET/PTC3 in a dose-dependent manner with IC50 of approximately 944 nm for SU5416, 562 nm for SU6668, and 224 nm for SU11248. Thus, SU11248 effectively inhibits the kinase activity of RET/PTC3. RET/PTC-mediated Y705 phosphorylation of STAT3 was inhibited by addition of SU11248, and the inhibitory effects of SU11248 on the tyrosine phosphorylation and transcriptional activation of STAT3 were very closely correlated with decreased autophosphorylation of RET/PTC. SU11248 caused a complete morphological reversion of transformed NIH-RET/PTC3 cells and inhibited the growth of TPC-1 cells that have an endogenous RET/PTC1. Conclusion: SU11248 is a highly effective tyrosine kinase inhibitor of the RET/PTC oncogenic kinase.
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Lopez, Tyler, Aaron Ramirez, Chris Benitez, Zahid Mustafa, Henry Pham, Ramon Sanchez, and Xin Ge. "Selectivity Conversion of Protease Inhibitory Antibodies." Antibody Therapeutics 1, no. 2 (September 2018): 75–83. http://dx.doi.org/10.1093/abt/tby008.
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ABSTRACT Background: Proteases are one of the largest pharmaceutical targets for drug developments. Their dysregulations result in a wide variety of diseases. Because proteolytic networks usually consist of protease family members that share high structural and catalytic homology, distinguishing them using small molecule inhibitors is often challenging. To achieve specific inhibition, this study described a novel approach for the generation of protease inhibitory antibodies. As a proof of concept, we aimed to convert a matrix metalloproteinase (MMP)-14 specific inhibitor to MMP-9 specific inhibitory antibodies with high selectivity. Methods: An error-prone single-chain Fv (scFv) library of an MMP-14 inhibitor 3A2 was generated for yeast surface display. A dual-color competitive FACS was developed for selection on MMP-9 catalytic domain (cdMMP-9) and counter-selection on cdMMP-14 simultaneously, which were fused/conjugated with different fluorophores. Isolated MMP-9 inhibitory scFvs were biochemically characterized by inhibition assays on MMP-2/-9/-12/-14, proteolytic stability tests, inhibition mode determination, competitive ELISA with TIMP-2 (a native inhibitor of MMPs), and paratope mutagenesis assays. Results: We converted an MMP-14 specific inhibitor 3A2 into a panel of MMP-9 specific inhibitory antibodies with dramatic selectivity shifts of 690–4,500 folds. Isolated scFvs inhibited cdMMP-9 at nM potency with high selectivity over MMP-2/-12/-14 and exhibited decent proteolytic stability. Biochemical characterizations revealed that these scFvs were competitive inhibitors binding to cdMMP-9 near its reaction cleft via their CDR-H3s. Conclusions: This study developed a novel approach able to convert the selectivity of inhibitory antibodies among closely related protease family members. This methodology can be directly applied for mAbs inhibiting many proteases of biomedical importance. Statement of Significance To achieve high selectivity required for therapies, we developed a novel approach for the generation of protease inhibitory antibodies with nM potency and decent proteolytic stability. The methodology demonstrated here can be readily applied to many proteases of biomedical importance.
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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.
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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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Yoshida, Hideo, Yosuke Okamura, Naohide Watanabe, Yasuo Ikeda, and Makoto Handa. "Shear-dependent suppression of platelet thrombus formation by phosphodiesterase 3 inhibition requires low levels of concomitant Gs-coupled receptor stimulation." Thrombosis and Haemostasis 105, no. 03 (2011): 487–95. http://dx.doi.org/10.1160/th10-07-0439.
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SummaryPhosphodiesterase (PDE)3 inhibitors exert potent antiplatelet effects through maintaining elevated intracellular cyclic adenosine monophosphate levels, but do not prolong bleeding time. To resolve this discrepancy, we hypothesised that PDE3 inhibitors effectively suppress shear-induced platelet thrombus formation initiated by the interaction of the platelet receptor GPIb/V/IX with its ligand, von Willebrand factor (VWF), since arterial thrombosis is more dependent on shear stress as compared with haemostatic plug formation. To test the hypothesis, we compared the in vitro effects of K-134 (a PDE3 inhibitor), tirofiban (a GPIIb/IIIa inhibitor) and acetylsalicylic acid (ASA) on ristocetin-induced platelet aggregation and platelet thrombus formation on VWF or collagen surfaces under flow conditions. K-134 inhibited GPIIb/IIIa-dependent platelet aggregation to the same extent as tirofiban and more potently than ASA. Likewise, K-134 and tirofiban effectively inhibited stable platelet thrombus formation (platelet firm adhesion and subsequent aggregation) on the VWF or collagen surface under high shear, but ASA only inhibited aggregation. Notably, inhibition by K-134 became evident only when a low concentration of PGE1 was present. These inhibitors did not block shear-induced initial platelet contact with VWF via GPIb/V/IX. In contrast, under low shear, the inhibitory effects of K-134 on platelet aggregation on the collagen surface were lower than tirofiban or ASA. The observed shear-dependent suppression of platelet thrombus formation by PDE3 inhibitor in the presence of low levels of adenylate cyclase stimulator may contribute to high therapeutic benefit with low risk of bleeding.
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Gledhill, J. R., and J. E. Walker. "Inhibitors of the catalytic domain of mitochondrial ATP synthase." Biochemical Society Transactions 34, no. 5 (October 1, 2006): 989–92. http://dx.doi.org/10.1042/bst0340989.
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An understanding of the mechanism of ATP synthase requires an explanation of how inhibitors act. The catalytic F1-ATPase domain of the enzyme has been studied extensively by X-ray crystallography in a variety of inhibited states. Four independent inhibitory sites have been identified by high-resolution structural studies. They are the catalytic site, and the binding sites for the antibiotics aurovertin and efrapeptin and for the natural inhibitor protein, IF1.
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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.
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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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Syrbu, Sergei I., Waltraut H. Waterman, Thaddeus F. P. Molski, Deepa Nagarkatti, Jean-Jacques Hajjar, and Ramadan I. Sha’afi. "Phosphorylation of Cytosolic Phospholipase A2 and the Release of Arachidonic Acid in Human Neutrophils." Journal of Immunology 162, no. 4 (February 15, 1999): 2334–40. http://dx.doi.org/10.4049/jimmunol.162.4.2334.
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Abstract Kinases mediating phosphorylation and activation of cytosolic phospholipase A2 (cPLA2) in intact cells remain to be fully characterized. Platelet-activating factor stimulation of human neutrophils increases cPLA2 phosphorylation. This increase is inhibited by PD 98059, a mitogen-activated protein (MAP)/extracellular signal-regulating kinase (erk) 1 inhibitor, but not by SB 203580, a p38 MAP kinase inhibitor, indicating that this action is mediated through activation of the p42 MAP kinase (erk2). However, platelet-activating factor-induced arachidonic acid release is inhibited by both PD 98059 and SB 203580. Stimulation by TNF-α increases cPLA2 phosphorylation, which is inhibited by SB 203580, but not PD 98059, suggesting a role for p38 MAP kinase. LPS increases cPLA2 phosphorylation and arachidonic acid release. However, neither of these actions is inhibited by either PD 98059 or SB 203580. PMA increases cPLA2 phosphorylation. This action is inhibited by PD 98059 but not SB 203580. Finally, FMLP increases cPLA2 phosphorylation and arachidonic acid release. Interestingly, while the FMLP-induced phosphorylation of cPLA2 is not affected by the inhibitors of the p38 MAP kinase or erk cascades, both inhibitors significantly decrease arachidonic acid release stimulated by FMLP. SB 203580 or PD 98059 has no inhibitory effects on the activity of coenzyme A-independent transacylase.
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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.
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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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Takuma, T. "Evidence against direct involvement of cyclic AMP-dependent protein phosphorylation in the exocytosis of amylase." Biochemical Journal 256, no. 3 (December 15, 1988): 867–71. http://dx.doi.org/10.1042/bj2560867.
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To examine whether or not the activation of cyclic AMP-dependent protein kinase is coupled to the exocytosis of amylase from rat parotid cells, the effect of protein kinase inhibitors on amylase release and protein phosphorylation was studied. A membrane-permeable inhibitor of cyclic AMP-dependent protein kinase, N-[2-(methylamino)ethyl]-5-isoquinolinesulphonamide (H-8), and peptide fragments of the heat-stable protein kinase inhibitor [PKI-(5-24)-peptide and PKI-(14-24)-amide] strongly inhibited cyclic AMP-dependent protein kinase activity in the cell homogenate. However, H-8 had no inhibitory effect on amylase release from either intact or saponin-permeabilized parotid cells stimulated by isoproterenol or cyclic AMP. Moreover, PKI-(5-24)-peptide and PKI-(14-24)-amide did not inhibit cyclic AMP-evoked amylase release from saponin-permeabilized cells, whereas cyclic AMP-dependent phosphorylations of 21 and 26 kDa proteins in intact or permeabilized cells were markedly inhibited by these inhibitors. These results suggest that cyclic AMP-dependent protein phosphorylation is not directly involved in the exocytosis of amylase regulated by cyclic AMP.
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Baader, E., G. Tschank, K. H. Baringhaus, H. Burghard, and V. Günzler. "Inhibition of prolyl 4-hydroxylase by oxalyl amino acid derivatives in vitro, in isolated microsomes and in embryonic chicken tissues." Biochemical Journal 300, no. 2 (June 1, 1994): 525–30. http://dx.doi.org/10.1042/bj3000525.
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The potency of oxalyl amino acid derivatives as inhibitors of prolyl 4-hydroxylase was studied in vitro, in isolated microsomes and in chicken embryonic-tissue culture. These compounds represent structural analogues of 2-oxoglutarate in which the -CH2- moiety at C-3 is replaced by -NH-, with or without further structural modifications. The most efficient inhibitor of purified prolyl 4-hydroxylase was oxalylglycine. Its mode of inhibition was competitive with respect to 2-oxoglutarate. The Ki value varied between 1.9 and 7.8 microM, depending on the variable substrate used. Oxalylalanine inhibited purified enzyme with a Ki of 40 microM. Other oxalyl amino acid derivatives showed little inhibitory activity. In microsomes isolated from embryonic chicken bone, oxalylglycine and oxalylalanine inhibited prolyl hydroxylation with IC50 values of 23 and 120 microM respectively. Dimethyloxalylglycine was not an inhibitor of purified prolyl 4-hydroxylase and only weakly active in the microsomal system, but efficiently suppressed hydroxyproline synthesis in embryonic chicken calvaria and lung. The data suggest that dimethyloxalyl amino acids are converted into active inhibitors in intact cells, most likely in the cytoplasmic compartment.
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Umezawa, Koji, and Isao Kii. "Druggable Transient Pockets in Protein Kinases." Molecules 26, no. 3 (January 27, 2021): 651. http://dx.doi.org/10.3390/molecules26030651.
Full textAbstract:
Drug discovery using small molecule inhibitors is reaching a stalemate due to low selectivity, adverse off-target effects and inevitable failures in clinical trials. Conventional chemical screening methods may miss potent small molecules because of their use of simple but outdated kits composed of recombinant enzyme proteins. Non-canonical inhibitors targeting a hidden pocket in a protein have received considerable research attention. Kii and colleagues identified an inhibitor targeting a transient pocket in the kinase DYRK1A during its folding process and termed it FINDY. FINDY exhibits a unique inhibitory profile; that is, FINDY does not inhibit the fully folded form of DYRK1A, indicating that the FINDY-binding pocket is hidden in the folded form. This intriguing pocket opens during the folding process and then closes upon completion of folding. In this review, we discuss previously established kinase inhibitors and their inhibitory mechanisms in comparison with FINDY. We also compare the inhibitory mechanisms with the growing concept of “cryptic inhibitor-binding sites.” These sites are buried on the inhibitor-unbound surface but become apparent when the inhibitor is bound. In addition, an alternative method based on cell-free protein synthesis of protein kinases may allow the discovery of small molecules that occupy these mysterious binding sites. Transitional folding intermediates would become alternative targets in drug discovery, enabling the efficient development of potent kinase inhibitors.
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Na, Bon Hyang, Thi Xoan Hoang, and Jae Young Kim. "Hsp90 Inhibition Reduces TLR5 Surface Expression and NF-κB Activation in Human Myeloid Leukemia THP-1 Cells." BioMed Research International 2018 (2018): 1–8. http://dx.doi.org/10.1155/2018/4319369.
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Tumors highly express active heat shock protein 90 (Hsp90), which is involved in tumor survival and progression. Enhanced Toll-like receptor (TLR) 5 expression and signaling were reported to be associated with acute myeloid leukemia. In the present study, we investigated the possible modulatory effects of Hsp90 inhibitors on TLR5 expression and signaling in the human myeloid leukemia cell line THP-1. Cells were pretreated with various concentrations of the Hsp90 inhibitor geldanamycin (GA) or the Hsp70 inhibitor VER155008, followed by stimulation with bacterial flagellin. Flagellin-induced nuclear factor-κB (NF-κB) activation was significantly reduced by treatment with GA or VER155008. To elucidate the underlying mechanism of this effect, mRNA and cell surface expression of TLR5 was examined. TLR5 mRNA expression was enhanced by both GA and VER155008, whereas cell surface expression of TLR5 was reduced by three different Hsp90 inhibitors, including GA, 17-(allylamino)-17-demethoxygeldanamycin, and radicicol, and an Hsp70 inhibitor. The inhibitory effect of Hsp90 inhibitors was much higher than that of Hsp70 inhibitor. Our results suggest that Hsp90 inhibitors suppress TLR5 surface expression and activation of NF-κB in THP-1 cells in response to TLR5 ligand, and these inhibitory effects may be associated with the possible mechanisms by which Hsp90 inhibitors suppress myeloid leukemia.
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Tran, Nhu Nguyen Quynh, and Kwang-Hoon Chun. "ROCK2-Specific Inhibitor KD025 Suppresses Adipocyte Differentiation by Inhibiting Casein Kinase 2." Molecules 26, no. 16 (August 5, 2021): 4747. http://dx.doi.org/10.3390/molecules26164747.
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KD025, a ROCK2 isoform-specific inhibitor, has an anti-adipogenic activity which is not mediated by ROCK2 inhibition. To identify the target, we searched binding targets of KD025 by using the KINOMEscanTM screening platform, and we identified casein kinase 2 (CK2) as a novel target. KD025 showed comparable binding affinity to CK2α (Kd = 128 nM). By contrast, CK2 inhibitor CX-4945 and ROCK inhibitor fasudil did not show such cross-reactivity. In addition, KD025 effectively inhibited CK2 at a nanomolar concentration (IC50 = 50 nM). We examined if the inhibitory effect of KD025 on adipocyte differentiation is through the inhibition of CK2. Both CX-4945 and KD025 suppressed the generation of lipid droplets and the expression of proadipogenic genes Pparg and Cebpa in 3T3-L1 cells during adipocyte differentiation. Fasudil exerted no significant effect on the quantity of lipid droplets, but another ROCK inhibitor Y-27632 increased the expression of Pparg and Cebpa. Both CX-4945 and KD025 acted specifically in the middle stage (days 1–3) but were ineffective when treated at days 0–1 or the late stages, indicating that CX-4945 and KD025 may regulate the same target, CK2. The mRNA and protein levels of CK2α and CK2β generally decreased in 3T3-L1 cells at day 2 but recovered thereafter. Other well-known CK2 inhibitors DMAT and quinalizarin inhibited effectively the differentiation of 3T3-L1 cells. Taken together, the results of this study confirmed that KD025 inhibits ROCK2 and CK2, and that the inhibitory effect on adipocyte differentiation is through the inhibition of CK2.
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Taylor, A. S., R. C. Howe, A. R. Morrison, H. Sprecher, and J. H. Russell. "Inhibition of cytotoxic T lymphocyte-mediated lysis by ETYA: effect independent of arachidonic acid metabolism." Journal of Immunology 134, no. 2 (February 1, 1985): 1130–35. http://dx.doi.org/10.4049/jimmunol.134.2.1130.
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Abstract Cytotoxic T lymphocyte (CTL)-mediated lysis of target cells was inhibited by 5,8,11,14-eicosatetraynoic acid (ETYA) and other inhibitors of the lipoxygenase pathway at concentrations that inhibited arachidonic acid metabolism in mixed lymphocyte cultures. Inhibition was reversible and selective for the "lethal hit" stage in the CTL-target interaction. Studies to define CTL-specific arachidonic acid metabolites demonstrated that cloned CTL populations have little or no capacity to metabolize arachidonic acid. Therefore, inhibitor actions appear to be independent of the effects on CTL arachidonic acid metabolism. Alternative explanations for inhibitory effects are discussed.
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Wang, Jintao, Yan Chen, Junyan Chen, Ling Jiang, Xiaofen Lin, Chaojun Cai, Minhua Zhang, Ming Li, and Justin Gu. "Abstract 670: Preclinical candidate LAE120, a novel selective USP1 inhibitor shows effective anticancer and combination activity with PARP inhibitor." Cancer Research 84, no. 6_Supplement (March 22, 2024): 670. http://dx.doi.org/10.1158/1538-7445.am2024-670.
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Abstract The Fanconi anemia (FA) and translesion synthesis (TLS) are DNA damage tolerance and repair pathways that have been found to be regulated by reversible ubiquitination in which ubiquitin-specific protease 1 (USP1) plays a key role. Numerous studies have indicated that USP1 inhibitors have therapeutic effects on tumors with homologous recombination deficiency (HRD), including mutations in BRCA1/2. Due to the emergence of resistance to PARP inhibitors in clinical applications, new strategies are needed to address this issue. Here we present LAE120, a novel, allosteric and highly potent USP1 inhibitor, displayed monotherapy potency and combination activity with PARP inhibitor in HRD cancers. LAE120 has a unique tail group distinct from all the published USP1 inhibitors and is predicted to induce a different conformational change at the USP1 allosteric site. In the biochemical assay, LAE120 strongly inhibited USP1 enzymatic activity with IC50 of 5.6 nM. LAE120 potently inhibited proliferation of BRCA-mutated MDA-MB-436 breast cancer cell (IC50:19 nM) and exhibited anti-proliferative activity in NCI-H1792 (IC50: 69 nM) and K562 (IC50: 38 nM) cell lines. In the xenograft model of MDA-MB-436 cells with BRCA mutations, LAE120 demonstrated robust single-agent tumor inhibitory activity (TGI: 88%, 10 mg/kg BID) and a favorable dose-dependent response (TGI: 106%, 25 mg/kg BID). We also observed strong tumor-inhibition efficacy of LAE120 as a single agent in the K562 xenograft model with Bcr-Abl expression (TGI: 86%, 10 mg/kg BID). Current research indicates that USP1 inhibitors exhibit synergistic activity in combination with PARP inhibitors and re-sensitizes PARP refractory tumors. Similar results have been confirmed in the xenograft model of BRCA-mutated MDA-MB-436 cells. The combination of LAE120 and Olaparib (PARP inhibitor) yielded a more robust and durable anti-tumor efficacy at low dose (TGI: 110%, LAE120, 5 mg/kg BID + Olaparib, 50 mg/kg QD). LAE120 also displayed favorable ADME properties and PK profiles. Preclinical-tox studies indicated the molecule was well tolerated without significant toxicities and hematological observations. Both in vitro and in vivo data strongly support further investigation of LAE120, whether as a single agent or in combination therapy. Citation Format: Jintao Wang, Yan Chen, Junyan Chen, Ling Jiang, Xiaofen Lin, Chaojun Cai, Minhua Zhang, Ming Li, Justin Gu. Preclinical candidate LAE120, a novel selective USP1 inhibitor shows effective anticancer and combination activity with PARP inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 670.
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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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Michisaka, Saki, Hitoshi Sase, Yukako Tachibana, Masami Hasegawa, Toshihiko Fujii, Shino Kuramoto, Norihito Shibahara, et al. "Abstract 1664: Combination of LUNA18, a novel RAS inhibitor, with KRAS G12C inhibitors augments anti-tumor activity via inhibition of MAPK pathway reactivation." Cancer Research 84, no. 6_Supplement (March 22, 2024): 1664. http://dx.doi.org/10.1158/1538-7445.am2024-1664.
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Abstract RAS gene alterations, found in approximately 15% of all cancers and known oncogenes, are recognized as promising therapeutic targets. KRAS G12C inhibitors were the first FDA-approved RAS targeting agent, but their efficacy of monotherapy is limited. Reactivation of the MAPK pathway is reported to be the one of the causes for attenuating efficacy of KRAS G12C inhibitors. LUNA18 is the cyclic peptide that binds to RAS mutants and wildtype, including KRAS, NRAS and HRAS, thereby inhibiting protein-protein interaction (PPI) between the inactive form of RAS and GEFs (guanine nucleotide exchange factors).We observed the rebound of ERK activation 24-72 hours after the treatment with a KRAS G12C inhibitor in cells with the KRAS G12C mutation though suppression of KRAS activity was maintained. This rebound was not suppressed by the KRAS G12C inhibitor retreatment. On the other hand, LUNA18 persistently suppressed ERK activity as well as KRAS activity up to 72 hours. LUNA18 also suppressed ERK reactivation caused by long-term treatment with the KRAS G12C inhibitor. The combination of the KRAS G12C inhibitor with siRNA for RAS wildtype showed durable inhibition of ERK activity, suggesting that RAS wildtype plays a key role for the rebound of ERK activity and inhibitory activity to RAS wildtype by LUNA18 contribute to the persistent inhibition of RAS signaling by LUNA18. In the experiment model that with strong RAS wildtype activation by stimulation with GFs (growth factors), the KRAS G12C inhibitor also did not suppress ERK activation. Conversely, LUNA18 suppressed it. Cell proliferation induced by GF stimulation was more strongly inhibited by LUNA18 than by the KRAS G12C inhibitors. Our data indicates that MAPK pathway reactivation via WT-RAS activation leads to resistance to KRAS G12C inhibitors and that the combination of LUNA18 and KRAS G12C inhibitors has the potential to overcome this resistance mechanism. In the in vitro cell growth assay, combination of LUNA18 and the KRAS G12C inhibitor showed more potent cell growth inhibition than single agents. We orally administered LUNA18 and the KRAS G12C inhibitor to a xenograft model, which showed temporal response and then acquired resistance to the KRAS G12C inhibitor, and found that LUNA18 significantly suppressed emergence of resistance to the KRAS G12C inhibitor. These results suggest that LUNA18 could be a promising candidate for combination therapy with KRAS G12C inhibitors. Citation Format: Saki Michisaka, Hitoshi Sase, Yukako Tachibana, Masami Hasegawa, Toshihiko Fujii, Shino Kuramoto, Norihito Shibahara, Atsushi Ohta, Mikihisa Tanada, Takuya Shiraishi, Hitoshi Iikura, Takehisa Kitazawa, Hiroshi Tanaka. Combination of LUNA18, a novel RAS inhibitor, with KRAS G12C inhibitors augments anti-tumor activity via inhibition of MAPK pathway reactivation [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1664.