Selective Small Molecule Inhibitor Research Articles

Practical Management of the Venetoclax-Treated Patient in Chronic Lymphocytic Leukemia and Acute Myeloid Leukemia

Venetoclax is a potent oral, highly selective small-molecule inhibitor of the antiapoptotic B-cell lymphoma 2 protein approved for chronic lymphocytic leukemia (CLL) or small lymphocytic lymphoma in treatment-naive patients (in combination with obinutuzumab) or for patients with relapsed/refractory CLL (in combination with rituximab). Venetoclax, in combination with azacitidine, decitabine, or low-dose cytarabine, is also approved in the United States for the treatment of newly diagnosed acute myeloid leukemia (AML) in adults who are ≥ 75 years or have comorbidities that preclude use of intensive induction chemotherapy. Clinical studies of patients with CLL or AML report both hematologic (e.g., neutropenia) and nonhematologic (e.g., gastrointestinal disorders and tumor lysis syndrome) adverse events associated with administration of venetoclax. It is therefore essential to provide information on the appropriate management of venetoclax-associated side effects. This article discusses the efficacy and safety of venetoclax administration and presents strategies specifically for the management of neutropenia and certain nonhematologic adverse events in patients receiving venetoclax for the treatment of AML and CLL.

A novel β-catenin/BCL9 complex inhibitor blocks oncogenic Wnt signaling and disrupts cholesterol homeostasis in colorectal cancer.

Dysregulated Wnt/β-catenin signaling is implicated in the pathogenesis of many human cancers, including colorectal cancer (CRC), making it an attractive clinical target. With the aim of inhibiting oncogenic Wnt activity, we developed a high-throughput screening AlphaScreen assay to identify selective small-molecule inhibitors of the interaction between β-catenin and its coactivator BCL9. We identified a compound that consistently bound to β-catenin and specifically inhibited in vivo native β-catenin/BCL9 complex formation in CRC cell lines. This compound inhibited Wnt activity, down-regulated expression of the Wnt/β-catenin signature in gene expression studies, disrupted cholesterol homeostasis, and significantly reduced the proliferation of CRC cell lines and tumor growth in a xenograft mouse model of CRC. This study has therefore identified a specific small-molecule inhibitor of oncogenic Wnt signaling, which may have value as a probe for functional studies and has important implications for the development of novel therapies in patients with CRC.

Protein proximity networks and functional evaluation of the casein kinase 1 gamma family reveal unique roles for CK1γ3 in WNT signaling

Aberrant activation or suppression of WNT/β-catenin signaling contributes to cancer initiation and progression, neurodegeneration, and bone disease. However, despite great need and more than 40 years of research, targeted therapies for the WNT pathway have yet to be fully realized. Kinases are considered exceptionally druggable and occupy key nodes within the WNT signaling network, but several pathway-relevant kinases remain understudied and “dark.” Here, we studied the function of the casein kinase 1γ (CSNK1γ) subfamily of human kinases and their roles in WNT signaling. miniTurbo-based proximity biotinylation and mass spectrometry analysis of CSNK1γ1, CSNK1γ2, and CSNK1γ3 revealed numerous components of the β-catenin–dependent and β-catenin–independent WNT pathways. In gain-of-function experiments, we found that CSNK1γ3 but not CSNK1γ1 or CSNK1γ2 activated β-catenin–dependent WNT signaling, with minimal effect on other signaling pathways. We also show that within the family, CSNK1γ3 expression uniquely induced low-density lipoprotein receptor–related protein 6 phosphorylation, which mediates downstream WNT signaling transduction. Conversely, siRNA-mediated silencing of CSNK1γ3 alone had no impact on WNT signaling, though cosilencing of all three family members decreased WNT pathway activity. Finally, we characterized two moderately selective and potent small-molecule inhibitors of the CSNK1γ family. We show that these inhibitors and a CSNK1γ3 kinase–dead mutant suppressed but did not eliminate WNT-driven low-density lipoprotein receptor–related protein 6 phosphorylation and β-catenin stabilization. Our data suggest that while CSNK1γ3 expression uniquely drives pathway activity, potential functional redundancy within the family necessitates loss of all three family members to suppress the WNT signaling pathway.

Lockdown, a selective small-molecule inhibitor of the integrin phosphatase PPM1F, blocks cancer cell invasion

Phosphatase PPM1F is a regulator of cell adhesion by fine-tuning integrin activity and actin cytoskeleton structures. Elevated expression of this enzyme in human tumors is associated with high invasiveness, enhanced metastasis, and poor prognosis. Thus, PPM1F is a target for pharmacological intervention, yet inhibitors of this enzyme are lacking. Here, we use high-throughput screening to identify Lockdown, a reversible and non-competitive PPM1F inhibitor. Lockdown is selective for PPM1F, because this compound does not inhibit other protein phosphatases invitro and does not induce additional phenotypes in PPM1F knockout cells. Importantly, Lockdown-treated glioblastoma cells fully re-capitulate the phenotype of PPM1F-deficient cells as assessed by increased phosphorylation of PPM1F substrates and corruption of integrin-dependent cellular processes. Ester modification yields LockdownPro with increased membrane permeability and prodrug-like properties. LockdownPro suppresses tissue invasion by PPM1F-overexpressing human cancer cells, validating PPM1F as a therapeutic target and providing an access point to control tumor cell dissemination.

Assessment of cytochrome P450 3A4-mediated drug\u2013drug interactions for ipatasertib using a fit-for-purpose physiologically based pharmacokinetic model

PurposeIpatasertib, a potent and highly selective small-molecule inhibitor of AKT, is currently under investigation for treatment of cancer. Ipatasertib is a substrate and a time-dependent inhibitor of CYP3A4. It exhibits non-linear pharmacokinetics at subclinical doses in the clinical dose escalation study. To assess the DDI risk of ipatasertib at the intended clinical dose of 400 mg with CYP3A4 inhibitors, inducers, and substrates, a fit-for-purpose physiologically based pharmacokinetic (PBPK) model of ipatasertib was developed.MethodsThe PBPK model was constructed in Simcyp using in silico, in vitro, and clinical data and was optimized and verified using clinical data.ResultsThe PBPK model described non-linear pharmacokinetics of ipatasertib and captured the magnitude of the observed clinical DDIs. Following repeated doses of 400 mg ipatasertib once daily (QD), the PBPK model predicted a 3.3-fold increase of ipatasertib exposure with itraconazole; a 2–2.5-fold increase with moderate CYP3A4 inhibitors, erythromycin and diltiazem; and no change with a weak CYP3A4 inhibitor, fluvoxamine. Additionally, in the presence of strong or moderate CYP3A4 inducers, rifampicin and efavirenz, ipatasertib exposures were predicted to decrease by 86% and 74%, respectively. As a perpetrator, the model predicted that ipatasertib (400 mg) caused a 1.7-fold increase in midazolam exposure.ConclusionThis study demonstrates the value of using a fit-for-purpose PBPK model to assess the clinical DDIs for ipatasertib and to provide dosing strategies for the concurrent use of other CYP3A4 perpetrators or victims.

Discovery of a selective and covalent small-molecule inhibitor of BFL-1 protein that induces robust apoptosis in cancer cells

Induction of apoptosis by the FDA-approved drug Venetoclax in cancer cells mainly derives from blocking the interactions between BCL-2 and BH3-only proteins. Anti-apoptotic BFL-1, a homolog of BCL-2, also competitively binds to the BH3-only proteins and is responsible for Venetoclax-induced drug resistance. Compared to BCL-2, small-molecule inhibitors of BFL-1 are relatively underexplored. In order to tackle this issue, in-house compound library was screened and a hit compound was identified and optimized to obtain 12 (ZH97) functioning as a covalent and selective inhibitor of BFL-1. 12 modifies BFL-1 at the C55 residue, blocks BFL-1/BID interaction in vitro, promotes cellular cytochrome c release from mitochondria, and induced apoptosis in BFL-1 overexpressing cancer cells. Mechanistic studies show that 12 inhibited BFL-1/PUMA interaction in cell lysate and is effective in cancer cells that harboring high expression level of BFL-1. In summary, blockade of BFL-1/BH3-only proteins interactions with a covalent small-molecule inhibitor induced apoptosis and elicited antitumor activity. Thus, our study demonstrates an appealing strategy for selective modulation of cellular BFL-1 for cancer therapy.

The monoacylglycerol acyltransferase pathway contributes to triacylglycerol synthesis in HepG2 cells

The monoacylglycerol acyltransferase (MGAT) pathway has a well-established role in the small intestine where it facilitates the absorption of dietary fat. In enterocytes, MGAT participates in the resynthesis of triacylglycerol using substrates (monoacylglycerol and fatty acids) generated in the gut lumen from the breakdown of triacylglycerol consumed in the diet. MGAT activity is also present in the liver, but its role in triacylglycerol metabolism in this tissue remains unclear. The predominant MGAT isoforms present in human liver appear to be MGAT2 and MGAT3. The objective of this study was to use selective small molecule inhibitors of MGAT2 and MGAT3 to determine the contributions of these enzymes to triacylglycerol production in liver cells. We found that pharmacological inhibition of either enzyme had no effect on TG mass in HepG2 cells but did alter lipid droplet size and number. Inhibition of MGAT2 did result in decreased DG and TG synthesis and TG secretion. Interestingly, MGAT2 preferentially utilized 2-monoacylglycerol derived from free glycerol and not from exogenously added 2-monoacylglycerol. In contrast, inhibition of MGAT3 had very little effect on TG metabolism in HepG2 cells. Additionally, we demonstrated that the MGAT activity of DGAT1 only makes a minor contribution to TG synthesis in intact HepG2 cells. Our data demonstrated that the MGAT pathway has a role in hepatic lipid metabolism with MGAT2, more so than MGAT3, contributing to TG synthesis and secretion.

A selective PPM1A inhibitor activates autophagy to restrict the survival of Mycobacterium tuberculosis

Metal-dependent protein phosphatases (PPMs) have essential roles in a variety of cellular processes, including inflammation, proliferation, differentiation, and stress responses, which are intensively investigated in cancer and metabolic diseases. Targeting PPMs to modulate host immunity in response to pathogens is an ambitious proposition. The feasibility of such a strategy is unproven because development of inhibitors against PPMs is challenging and suffers from poor selectivity. Combining a biomimetic modularization strategy with function-oriented synthesis, we design, synthesize and screen more than 500 pseudo-natural products, resulting in the discovery of a potent, selective, and non-cytotoxic small molecule inhibitor for PPM1A, SMIP-30. Inhibition of PPM1A with SMIP-30 or its genetic ablation (ΔPPM1A) activated autophagy through a mechanism dependent on phosphorylation of p62-SQSTM1, which restricted the intracellular survival of Mycobacterium tuberculosis in macrophages and in the lungs of infected mice. SMIP-30 provides proof of concept that PPMs are druggable and promising targets for the development of host-directed therapies against tuberculosis.

SS-4 is a highly selective small molecule inhibitor of STAT3 tyrosine phosphorylation that potently inhibits GBM tumorigenesis in vitro and in vivo

Glioblastoma (GBM) is a highly aggressive cancer with a dismal prognosis. Constitutively active STAT3 has a causal role in GBM progression and is associated with poor patient survival. We rationally designed a novel small molecule, SS-4, by computational modeling to specifically interact with STAT3. SS-4 strongly and selectively inhibited STAT3 tyrosine (Y)-705 phosphorylation in MT330 and LN229 GBM cells and inhibited their proliferation and induced apoptosis with an IC50 of ∼100 nM. The antiproliferative and apoptotic actions of SS-4 were Y-705 phosphorylation dependent, as evidenced by its lack of effects on STAT3 knockout (STAT3KO) cells or STAT3KO cells that overexpressed a phospho-Y705 deficient (STAT3Y705F) mutant, and the recovery of effects when wild-type STAT3 or a phospho-serine (S)727 deficient mutant was expressed in STAT3KO cells. SS-4 increased the expression of STAT3 repressed genes, while decreasing the expression of STAT3 promoted genes. Importantly, SS-4 markedly reduced the growth of GBM intracranial tumor xenografts. These data together identify SS-4 as a potent STAT3 inhibitor that selectively blocks Y705-phosphorylation, induces apoptosis, and inhibits growth of human GBM models in vitro and in vivo.

A selective inhibitor of the NLRP3 inflammasome as a potential therapeutic approach for neuroprotection in a transgenic mouse model of Huntington\u2019s disease

BackgroundHuntington’s disease (HD) is a neurodegenerative disorder caused by the expansion of the CAG repeat in the huntingtin (HTT) gene. When the number of CAG repeats exceeds 36, the translated expanded polyglutamine-containing HTT protein (mutant HTT [mHTT]) interferes with the normal functions of many cellular proteins and subsequently jeopardizes important cellular machineries in major types of brain cells, including neurons, astrocytes, and microglia. The NACHT, LRR, and PYD domain-containing protein 3 (NLRP3) inflammasome, which comprises NLRP3, ASC, and caspase-1, is involved in the activation of IL-1β and IL-18 and has been implicated in various biological functions. Although the existence of the NLRP3 inflammasome in the brain has been documented, the roles of the NLRP3 inflammasome in HD remain largely uncharacterized. MCC950 is a highly selective and potent small-molecule inhibitor of NLRP3 that has been used for the treatment of several diseases such as Alzheimer’s disease. However, whether MCC950 is also beneficial in HD remains unknown. Therefore, we hypothesized that MCC950 exerts beneficial effects in a transgenic mouse model of HD.MethodsTo evaluate the effects of MCC950 in HD, we used the R6/2 (B6CBA-Tg[HDexon1]62Gpb/1J) transgenic mouse model of HD, which expresses exon 1 of the human HTT gene carrying 120 ± 5 CAG repeats. Male transgenic R6/2 mice were treated daily with MCC950 (10 mg/kg of body weight; oral administration) or water for 5 weeks from the age of 7 weeks. We examined neuronal density, neuroinflammation, and mHTT aggregation in the striatum of R6/2 mice vs. their wild-type littermates. We also evaluated the motor function, body weight, and lifespan of R6/2 mice.ResultsSystematic administration of MCC950 to R6/2 mice suppressed the NLRP3 inflammasome, decreased IL-1β and reactive oxygen species production, and reduced neuronal toxicity, as assessed based on increased neuronal density and upregulation of the NeuN and PSD-95 proteins. Most importantly, oral administration of MCC950 increased neuronal survival, reduced neuroinflammation, extended lifespan, and improved motor dysfunction in R6/2 mice.ConclusionsCollectively, our findings indicate that MCC950 exerts beneficial effects in a transgenic mouse model of HD and has therapeutic potential for treatment of this devastating neurodegenerative disease.

TWIST1-EP300 Expedites Gastric Cancer Cell Resistance to Apatinib by Activating the Expression of COL1A2.

The association between collagen type I alpha (COL1A) and chemoresistance has been verified in cancers. However, the specific role of COL1A2 in gastric cancer (GC) cell resistance to apatinib, a highly selective small-molecule inhibitor of vascular endothelial growth factor receptor 2, has not been investigated before. The purpose of this study was to explore the potential factors associated with COL1A2 regulation on GC cell apatinib resistance in vitro. With the aid of the Oncomine database and integrated bioinformatics methods, we identified COL1A2 overexpression in GC and its prognostic value. Mechanistically, the COL1A2 promoter has a distinct H3K27ac modification site and that E1A binding protein p300 (EP300) and twist family bHLH transcription factor 1 (TWIST1) can bind to the COL1A2 promoter, which in turn transcriptionally activated COL1A2 expression. In addition, overexpression of COL1A2 significantly promoted resistance to apatinib in GC cells, but knockdown of EP300 or TWIST1 remarkably inhibited COL1A2 expression and promoted sensitivity of GC cells to apatinib. Our findings demonstrated that the combination of EP300 and TWIST1 has a synergistically regulatory effect on COL1A2 expression, thus contributing to apatinib resistance in GC cells.

Inhibition of the NLRP3 Inflammasome Activation by Manoalide Ameliorates Experimental Autoimmune Encephalomyelitis Pathogenesis.

The activation of NLRP3 inflammasome leads to cell pyroptosis and inflammatory cytokines secretion and gets involved in the development of many diseases, such as neuroinflammation and metabolic syndrome, but the drugs targeting NLRP3 are not clinically available for now. Through screening the small molecule library, we found that manoalide is a highly selective small molecule inhibitor of NLRP3. Mechanismly, manoalide inhibited the NLRP3 inflammasome activation by acting downstream of potassium efflux, chloride efflux and mitochondrial dysfunction. Moreover, manoalide blocked the interaction between NEK7 and NLRP3 by covalently binding to Lys 377 of the NLRP3 protein. Treatment of manoalide relieved the pathogenesis of experimental autoimmune encephalomyelitis (EAE) in mice. Thus, our results identify manoalide as a selective and covalent NLRP3 inhibitor and suggest it has the potential for the treatment of NLRP3-associated diseases.

Abstract P5-08-14: Maternal embryonic leucine zipper kinase is associated with metastasis in triple-negative breast cancer

Abstract Background: Triple-negative breast cancer (TNBC) has high rates of relapse and metastasis. It has a high proportion of cancer stem-like cells (CSCs), which possess self-renewal and tumor initiation capacity. MELK (maternal embryonic leucine zipper kinase), a protein kinase of the Snf1/AMPK kinase family, plays a critical role in promoting CSC maintenance, cell proliferation, and malignant transformation. Here, we assessed the role of MELK in TNBC aggressiveness. Methods: The clinical relevance of MELK in TNBC was analyzed using the World IBC Consortium dataset (n = 314). The effects of MELK knockdown (siRNA), knockout (CRISPR-Cas9), and MELK inhibition with a novel selective small molecule inhibitor MELK-In-7 on migration and invasion were determined using the migration and invasion assays, respectively. MELK’s role in regulating CSC phenotypes was assessed using the mammosphere assay and flow cytometry. The impact of MELK knockdown on epithelial-to-mesenchymal transition (EMT) was determined using Western blotting. MELK’s protumorigenic role was determined in vitro using soft agar assay and in vivo using TNBC xenograft mouse models. Results: MELK mRNA expression level was higher in TNBC tumors than in tumors expressing hormone receptor, HER2, or both (P < 0.0001). In univariate analysis, breast cancer patients with high-MELK-expressing tumors had worse overall survival (OS, P < 0.001) and distant metastasis-free survival (DMFS, P < 0.01) than patients with low-MELK-expressing tumors, suggesting a prognostic impact of MELK in breast cancer. MELK knockdown using siRNA or MELK inhibition using MELK-In-7 significantly reduced migration, invasion, mammosphere formation, CD24-/CD44+ subpopulations, and ALDH activity, and reversed EMT in TNBC cells. This result suggests a role for MELK in regulating CSCs and EMT, which play crucial roles in tumor invasion and metastasis. Nude mice injected with MELK-knockout MDA-MB-231 cells exhibited suppression of lung metastasis and significantly improved OS compared with the mice injected with parental MDA-MB-231 cells (P < 0.05). Further, MELK-selective inhibitor (MELK-In-7) suppressed the growth of 4T1 mouse TNBC tumors in a dose-dependent manner in syngeneic BALB/c mice (P < 0.001). These results indicate an essential role for MELK in promoting TNBC tumor growth and metastasis. To uncover the underlying molecular mechanism of MELK’s regulation of TNBC metastasis, we performed microarray analysis and identified SERPINE1, a serine protease inhibitor known to regulate metastasis, as a downstream target of MELK. We validated that mRNA levels of SERPINE1 were reduced in MELK-knockout MDA-MB-231 cells and MELK-In-7-treated MDA-MB-231 cells. Moreover, SERPINE1 knockdown using siRNA reduced migration and invasion of TNBC cells. These results suggest that MELK regulates TNBC cell motility through SERPINE1. Conclusion: This work confirms MELK as a driver of aggressiveness and metastasis in TNBC. The mechanisms underlying MELK’s regulation of TNBC metastasis via SERPINE1 require further studies. Citation Format: Xuemei Xie, Gaurav B. Chauhan, Ramakrishna Edupuganti, Takahiro Kogawa, Jihyun Park, Moises T Tacam, Fnu Vidhu, Diane D. Liu, Juliana M. Taliaferro, Mary Kathryn Pitner, Yu Shen, Naoto T. Ueno, Savitri Krishnamurthy, Gabriel N. Hortobagyi, Debu Tripathy, Steven J. Van Laere, Geoffrey Bartholomeusz, Kevin Dalby, Chandra Bartholomeusz. Maternal embryonic leucine zipper kinase is associated with metastasis in triple-negative breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr P5-08-14.

Abstract OT2-01-02: First in human phase 1 dose escalation and expansion study of the safety and pharmacokinetics of the oral CDK7 inhibitor XL102 as a single-agent and in combination therapy in patients with inoperable locally advanced or metastatic solid tumors, including breast cancer

Abstract Background: XL102 is an orally bioavailable, selective, and covalent small-molecule inhibitor of cyclin-dependent kinase 7 (CDK7). CDK7 is a serine/threonine kinase which controls cell cycle progression via the phosphorylation of CDKs (1, 2, 4 & 6) and regulates transcription through phosphorylation of RNA polymerase II. CDK7 overexpression has been reported in multiple tumor types including hormone receptor positive (HR+) breast cancer (BC) and triple negative breast cancer (TNBC). XL102 induced cell death in a number of cancer cell lines and caused tumor regression in murine xenograft models of multiple tumor types including HR+ BC and TNBC. Here, we present the study design of an ongoing phase 1 trial in solid tumors which includes cohorts with advanced HR+ BC, TNBC, epithelial ovarian cancer (EOC), and metastatic castration-resistant prostate cancer (mCRPC). Trial Design and Statistical Methods: This phase 1, open-label trial will assess the safety, tolerability, pharmacokinetics, anti-tumor activity and effect on biomarkers of XL102 administered orally alone and in combination regimens in selected tumor types (NCT04726332). The study consists of dose-escalation stages and disease-specific cohort expansion stages. In the dose-escalation stages (modified interval 3+3 design), a maximum tolerated (MTD) and/or recommended XL102 dose (RD) for use alone and in combination therapy with standard dose fulvestrant (HR+BC) or abiraterone/prednisone (mCRPC) will be established in independent strata. Upon reaching the MTD/RD, the expansion stage will enroll according to Simon’s Two-Stage Minimax design, assuming a power of 80% and one-sided α of 15%, for single agent XL102 (HR+BC, TNBC, EOC, and mCRPC) and XL102 in combination therapy (HR+BC and mCRPC). Eligibility criteria: All patients must have adequate organ function and be able to take oral medications. Patients with CNS disease are eligible if treated and stable, provided they meet the other inclusion and exclusion criteria. Patients enrolled in expansion cohorts must have measurable disease. Patients in HR+ BC expansion cohorts must have received ≥ 1 prior endocrine anticancer therapy and 1 prior CDK4/6 inhibitor therapy either sequentially or concurrently and may have received ≤ 2 additional prior lines of systemic anticancer therapy for locally advanced or metastatic disease. Patients in TNBC and EOC expansion cohorts must have received ≥ 1 line but < 3 lines of prior systemic anticancer therapy for locally advanced or metastatic disease or platinum-resistant ovarian cancer (eg, cytotoxic therapy, targeted therapy, immunotherapy), respectively. Patients in EOC expansion cohort may include primary peritoneal cancer and fallopian tube cancer who have platinum-resistant disease following treatment with a platinum-containing chemotherapy. Patients with ovarian borderline epithelial tumors and platinum-refractory disease are ineligible. Study objectives: The primary objective of the expansion stage is the objective response rate of XL102 alone and in combination therapy as assessed by the investigator per RECIST 1.1. The secondary objectives are to evaluate safety, tolerability, and pharmacokinetics. Exploratory objectives include duration of response, progression-free survival, overall survival, and correlation of tumor and blood biomarkers with response. Accrual: The study began enrolling patients in February 2021 and is ongoing. Total enrollment estimated to be up to 298 subjects. Citation Format: Amita Patnaik, Minal Barve, Manali Bhave, Vivek Subbiah, Sumandeep Atwal, Keerti Sharma, Christian Scheffold, Cynthia Wetmore, Geoffrey Shapiro. First in human phase 1 dose escalation and expansion study of the safety and pharmacokinetics of the oral CDK7 inhibitor XL102 as a single-agent and in combination therapy in patients with inoperable locally advanced or metastatic solid tumors, including breast cancer [abstract]. In: Proceedings of the 2021 San Antonio Breast Cancer Symposium; 2021 Dec 7-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2022;82(4 Suppl):Abstract nr OT2-01-02.

A prospective, multicenter, dose-escalation, and dose-expansion study evaluating apatinib plus chemotherapy in second-line therapy of patients with postoperatively gastric cancer (ACHIEVE).

TPS362 Background: Apatinib, a highly selective small-molecule tyrosinase inhibitor of VEGFR2, is the third-line standard therapy for patients (pts) with advanced gastric cancer. Many previous studies have preliminarily demonstrated the efficacy and safety of apatinib combined with chemotherapy in the second-line treatment of gastric cancer. Comparing with pts who have not undergone surgery, previous pharmacological study has found that pts who have undergone gastric cancer surgery have lower bioavailability and can tolerate higher doses of apatinib (Clin Pharmacokinet. 2017 Jan; 56(1):65-76). However, there is no prospective evidence of apatinib plus chemotherapy for second line treatment in pts with postoperatively gastric cancer. Methods: To explore the efficacy, safety and the maximum tolerated dose (MTD) of apatinib from apatinib+chemotherapy for second line treatment in pts with postoperatively gastric cancer. We designed ACHIEVE as a multicenter, dose-escalation, and dose-expansion trial in pts with gastric or gastroesophageal junction cancer who had prior undergone gastrectomy. In part 1 (dose escalation), pts who had received previously first-line treatment will be assigned to 1 of 3 three dose groups: (1) apatinib (250mg qd) + chemotherapy drug; (2) apatinib (500mg qd) + chemotherapy drug; (3) apatinib (750mg qd) + chemotherapy drug. The therapeutic drugs for second-line chemotherapy are determined by the investigator, it’s recommended to use docetaxel or albumin-bound paclitaxel, and chemotherapeutics are used for up to 6 cycles. In part 2 (dose expansion), pts will be treated with apatinib (MTD determined by dose escalation, qd) + chemotherapy drug. Whether PD-1 or PD-L1 inhibitors have been used in the first-line treatment will be analyzed as a subgroup analysis factor. Primary endpoints are progression-free survival (PFS) and MTD. Secondary endpoints include confirmed objective response rate (ORR), disease control rate (DCR), duration of response (DOR), overall survival (OS), and safety. AEs will be monitored throughout the study and graded per NCI CTCAE v5.0. Tumor response will be assessed by RECIST v1.1 every 6 weeks in the first 18 weeks, and every 9 weeks thereafter. Clinical trial information: ChiCTR2100048288.

Targeting pancreatic cancer by TAK-981: a SUMOylation inhibitor that activates the immune system and blocks cancer cell cycle progression in a preclinical model

ObjectivePancreatic ductal adenocarcinoma (PDAC) has the characteristics of high-density desmoplastic stroma, a distinctive immunosuppressive microenvironment and is profoundly resistant to all forms of chemotherapy and immunotherapy, leading to a 5-year...

P019 GS-1069518 is a potent and selective small molecule α4β7 inhibitor

Abstract Background The interaction between the adhesion molecule MAdCAM-1 and the α4β7 integrin heterodimer enables the trafficking of gut-antigen specific lymphocytes to the small and large intestines and to gut-associated lymphoid tissues. The role of the α4β7 integrin in the pathophysiology of IBD has been clinically demonstrated, and targeted therapeutics have been shown to mitigate inflammation and mucosal damage mediated by α4β7+ lymphocytes. The purpose of this study was to determine the potency and selectivity of the small molecule α4β7 inhibitor GS-1069518. Methods The selectivity of GS-1069518 and relevant reference molecules was determined using in vitro cell capture assays with natural integrin ligands and by measuring receptor occupancy in human and cynomolgus monkey whole blood. The number of RPMI-8866 cells binding to MAdCAM-1-coated surfaces (a natural α4β7 ligand) and Jurkat cells binding to VCAM-1-coated surfaces (a natural α4β1 ligand) were quantified with high content imaging. Receptor occupancy on CD3+CD4+CD45RO+β7+ (human) or CD3+CD4+CD45RA-β7+ (cynomolgus monkey) and CD3+CD4+CD45RO+α4+β1+β7- (human) or CD3+CD4+CD45RA-α4+β1+β7- (cynomolgus monkey) memory T cells was measured by flow cytometry with fluorescent probes that detect occupancy in the ligand-binding sites. Results In a MAdCAM-1 cell capture assay, the EC50 of GS-1069518 was 0.034 nM, with 71-fold and >10,000-fold selectivity over α4β1 and αLβ2, respectively. In α4β7 receptor occupancy assays the EC50 of GS-1069518 was 0.36 nM in human whole blood and 0.4 nM in cynomolgus monkey whole blood. GS-1069518 was 482-fold selective for α4β7 over α4β1 in human whole blood and 119-fold selective for α4β7 in cynomolgus monkey blood. Conclusion GS-1069518 is a highly potent and selective small molecule inhibitor of the α4β7-MAdCAM-1 ligand binding interaction.

A Structure-based Design Approach for Generating High Affinity BRD4 D1-Selective Chemical Probes.

Chemical probes for epigenetic proteins are essential tools for dissecting the molecular mechanisms for gene regulation and therapeutic development. The bromodomain and extra-terminal (BET) proteins are master transcriptional regulators. Despite promising therapeutic targets, selective small molecule inhibitors for a single bromodomain remain an unmet goal due to their high sequence similarity. Here, we address this challenge via a structure-activity relationship study using 1,4,5-trisubstituted imidazoles against the BRD4 N-terminal bromodomain (D1). Leading compounds 26 and 30 have 15 and 18 nM affinity against BRD4 D1 and over 500-fold selectivity against BRD2 D1 and BRD4 D2 via ITC. Broader BET selectivity was confirmed by fluorescence anisotropy, thermal shift, and CETSA. Despite BRD4 engagement, BRD4 D1 inhibition was unable to reduce c-Myc expression at low concentration in multiple myeloma cells. Conversely, for inflammation, IL-8 and chemokine downregulation were observed. These results provide new design rules for selective inhibitors of an individual BET bromodomain.

Screening for Small-Molecule Inhibitors of Histone Methyltransferases.

Potent and highly selective small-molecule inhibitors are needed to unravel the biological complexities of histone methyltransferases and to reveal their therapeutic potential. A prerequisite to developing these inhibitors is the identification of validated chemical matter for initiating a medicinal chemistry campaign. For the most part, finding these initial starting points occurs through screening of large, unbiased compound libraries. The size and nature of these libraries, coupled with the complexities of the bisubstrate utilizing histone methyltransferases, necessitates that the primary screen and subsequent hit triage be carefully considered.In this chapter, using EZH2 as a representative example, we describe a screening and hit triage campaign that identified validated chemical matter allowing initiation of medicinal chemistry studies. Moreover, we discuss a cell-based assay to support lead identification and optimization. The approach described here entailing a mixture of biochemical, biophysical and cell-based assays should be applicable to identifying validated starting points for other histone methyltransferases.

Small molecule STING inhibition improves myocardial infarction remodeling

AimsMyocardial infarction (MI) is a major global cause of death. Massive cell death leads to inflammation, which is necessary for ensuing wound healing. Extensive inflammation, however, promotes infarct expansion and adverse remodeling. The DNA sensing receptor cyclic GMP-AMP synthase and its downstream signaling effector stimulator of interferon genes (cGAS-STING) is central in innate immune reactions in infections or autoimmunity. Cytosolic double-strand DNA activates the pathway and down-stream inflammatory responses. Recent papers demonstrated that this pathway is also active following MI and that its genetic targeting improves outcome. Thus, we investigated if pharmacologic pathway inhibition is protective after MI in order to test its translational potential. Main methodsWe investigated novel and selective small-molecule STING inhibitors that inhibit STING palmitoylation and multimerization and thereby downstream pathway activation in a preclinical murine MI model. We assessed structural and functional cardiac remodeling, infarct expansion and fibrosis, as well as cardiomyocyte hypertrophy and the expression of inflammatory genes. Key findingsPharmacologic STING inhibition did not reduce mortality due to myocardial rupture in non-reperfused MI. Infarct size at day one was comparable. However, three weeks of pharmacologic STING inhibition after reperfused MI decreased infarct expansion and scarring, increased left ventricular systolic function to levels approaching normal values, and reduced myocardial hypertrophy. SignificanceSelective small-molecule STING inhibition after myocardial infarction has the potential to improve wound healing responses and pathological remodeling and thereby attenuate the development of ischemic heart failure.