Mcl-1 Inhibitors Research Articles

Regorafenib inhibits growth, survival and angiogenesis in nasopharyngeal carcinoma and is synergistic with Mcl-1 inhibitor.

Regorafenib is an oral multi-kinase inhibitor approved for various metastatic/advanced cancers, and has been investigated in clinical trials in many other tumour entities. The purpose of this study was to evaluate the therapeutic potential of regorafenib for nasopharyngeal carcinoma (NPC). Cellular proliferation, survival, apoptosis and colony formation assays were performed and combination index was determined. NPC xenograft tumour models were established. In vitro and In vivo angiogenesis assays were performed. Regorafenib is effective against a panel of NPC cell lines regardless of cellular origin and genetic profiling while sparing normal nasal epithelial cells. The predominant inhibitory effects of regorafenib in NPC are anchorage-dependent and anchorage-independent growth rather than survival. Apart from tumour cells, regorafenib potently inhibits angiogenesis. Mechanistically, regorafenib inhibits multiple oncogenic pathways including Raf/Erk/Mek and PI3K/Akt/mTOR. Regorafenib decreases Bcl-2 but not Mcl-1 level in NPC cells. The in vitro observations are evident in in vivo NPC xenograft mouse model. The combination of Mcl-1 inhibitor with regorafenib is synergistic in inhibiting NPC growth without causing systemic toxicity in mice. Our findings also support further clinical investigation of regorafenib and Mcl-1 inhibitor for NPC treatment.

Venetoclax, alone and in combination with the BH3 mimetic S63845, depletes HIV-1 latently infected cells and delays rebound in humanized mice

HIV-1 persists indefinitely in people living with HIV (PLWH) on antiretroviral therapy (ART). If ART is stopped, the virus rapidly rebounds from long-lived latently infected cells. Using a humanized mouse model of HIV-1 infection and CD4+ Tcells from PLWH on ART, we investigate whether antagonizing host pro-survival proteins can prime latent cells to die and facilitate HIV-1 clearance. Venetoclax, a pro-apoptotic inhibitor of Bcl-2, depletes total and intact HIV-1 DNA in CD4+ Tcells from PLWH exvivo. This venetoclax-sensitive population is enriched for cells with transcriptionally higher levels of pro-apoptotic BH3-only proteins. Furthermore, venetoclax delays viral rebound in a mouse model of persistent HIV-1 infection, and the combination of venetoclax with the Mcl-1 inhibitor S63845 achieves a longer delay in rebound compared with either intervention alone. Thus, selective inhibition of pro-survival proteins can induce death of HIV-1-infected cells that persist on ART, extending time to viral rebound.

HSP90 Inhibitor PU-H71 in Combination with BH3-Mimetics in the Treatment of Acute Myeloid Leukemia.

Targeting the molecular chaperone HSP90 and the anti-apoptotic proteins MCL1 and BCL2 may be a promising novel approach in the treatment of acute myeloid leukemia (AML). The HSP90 inhibitor PU-H71, MCL1 inhibitor S63845, and BCL2 inhibitor venetoclax were assessed as single agents and in combination for their ability to induce apoptosis and cell death in leukemic cells. AML cells represented all major morphologic and molecular subtypes including FLT3-ITD and TP53 mutant AML cell lines and a variety of patient-derived AML cells. Results: PU-H71 and combination treatments with MCL1 inhibitor S63845 or BCL2 inhibitor venetoclax induced cell cycle arrest and apoptosis in susceptible AML cell lines and primary AML. The majority of the primary AML samples were responsive to PU-H71 in combination with BH3 mimetics. Elevated susceptibility to PU-H71 and S63845 was associated with FLT3 mutated AML with CD34 < 20%. Elevated susceptibility to PU-H71 and venetoclax was associated with primary AML with CD117 > 80% and CD11b < 45%. The combination of HSP90 inhibitor PU-H71 and MCL1 inhibitor S63845 may be a candidate treatment for FLT3-mutated AML with moderate CD34 positivity while the combination of HSP90 inhibitor PU-H71 and BCL2 inhibitor venetoclax may be more effective in the treatment of primitive AML with high CD117 and low CD11b positivity.

Metal-Catalyst-Free One-Pot Aqueous Synthesis of trans-1,2-Diols from Electron-Deficient α,β-Unsaturated Amides via Epoxidation Using Oxone as a Dual Role Reagent.

In organic synthesis, incorporating two functional groups into the carbon-carbon double bond of α,β-unsaturated amides is challenging due to the electron-deficient nature of the olefin moiety. Although a few examples of dihydroxylation of α,β-unsaturated amides have been demonstrated, producing cis-1,2-diols using either highly toxic OsO4 or other specialized metal reagents in organic solvents, they are limited to several specific amides. We describe herein a general and one-pot direct synthesis of trans-1,2-diols from electron-deficient α,β-unsaturated amides through dihydroxylation using oxone as a dual-role reagent in water. This reaction does not require any metal catalyst and produces non-hazardous and nontoxic K2SO4 as the sole byproduct. Moreover, epoxidation products could also be selectively formed by adjusting the reaction conditions. By the strategy, the intermediates of Mcl-1 inhibitor and antiallergic bioactive molecule can be synthesized in one pot. The gram-scale synthesis of trans-1,2-diol which is isolated and purified by recrystallization further shows the potential applications of this new reaction in organic synthesis.

Combined Mcl-1 and YAP1/TAZ inhibition for treatment of metastatic uveal melanoma.

Uveal melanoma is the most common intraocular tumor in adults, representing approximately 5% of all melanoma cases. Up to 50% of uveal melanoma patients develop metastases that are resistant to most of the commonly used antineoplastic treatments. Virtually all uveal melanoma tumors harbor activating mutations in GNAQ or GNA11 , encoding Gαq and Gα11, respectively. Constant activity of these proteins causes deregulation of multiple downstream signaling pathways including PKC, MAPK and YAP1/TAZ. While the importance of YAP1 signaling for the proliferation of uveal melanoma has recently been demonstrated, much less is known about the paralog of YAP1 transcriptional coactivator, named TAZ; however, similar to YAP1, TAZ is expected to be a therapeutic target in uveal melanoma. We performed a small-scale drug screen to discover a compound synergistically inhibiting uveal melanoma proliferation/survival in combination with YAP1/TAZ inhibition. We found that the combination of genetic depletion of YAP1/TAZ together with Mcl-1 inhibition demonstrates a synergistic inhibitory effect on the viability of uveal melanoma cell lines. Similarly, indirect attenuation of the YAP1/TAZ signaling pathway with an inhibitor of the mevalonate pathway, that is, the geranyl-geranyl transferase inhibitor GGTI-298, synergizes with Mcl-1 inhibition. This combination could be potentially used as a treatment for metastatic uveal melanoma.

Mcl-1 deficiency in murine livers leads to nuclear polyploidisation and mitotic errors: Implications for hepatocellular carcinoma

Background & AimsMcl-1, an antiapoptotic protein overexpressed in many tumours, including hepatocellular carcinoma (HCC), represents a promising target for cancer treatment. Although Mcl-1 non-apoptotic roles might critically influence the therapeutic potential of Mcl-1 inhibitors, these functions remain poorly understood. We aimed to investigate the effects of hepatic Mcl-1 deficiency (Mcl-1Δhep) on hepatocyte ploidy and cell cycle in murine liver in vivo and the possible implications on HCC. MethodsLivers of young Mcl-1Δhep and wild-type (WT) mice were analysed for ploidy profile, mitotic figures, in situ chromosome segregation, gene set enrichment analysis and were subjected to two-thirds partial hepatectomy to assess Mcl-1 deficiency effect on cell cycle progression in vivo. Mcl-1Δhep tumours in older mice were analysed for ploidy profile, chromosomal instability, and mutational signatures via whole exome sequencing. ResultsIn young mice, Mcl-1 deficiency leads to nuclear polyploidy and to high rates of mitotic errors with abnormal spindle figures and chromosome mis-segregation along with a prolonged spindle assembly checkpoint activation signature. Chromosomal instability and altered ploidy profile are observed in Mcl-1Δhep tumours of old mice as well as a characteristic mutational signature of currently unknown aetiology. ConclusionsOur study suggests novel non-apoptotic effects of Mcl-1 deficiency on nuclear ploidy, mitotic regulation, and chromosomal segregation in hepatocytes in vivo. In addition, the Mcl-1 deficiency characteristic mutational signature might reflect mitotic issues. These results are of importance to consider when developing anti-Mcl-1 therapies to treat cancer. Impact and implicationsAlthough Mcl-1 inhibitors represent promising hepatocellular carcinoma treatment, the still poorly understood non-apoptotic roles of Mcl-1 might compromise their successful clinical application. Our study shows that Mcl-1 deficiency leads to nuclear polyploidy, mitotic errors, and aberrant chromosomal segregation in hepatocytes in vivo, whereas hepatocellular tumours spontaneously induced by Mcl-1 deficiency exhibit chromosomal instability and a mutational signature potentially reflecting mitotic issues. These results have potential implications for the development of anti-Mcl-1 therapies to treat hepatocellular carcinoma, especially as hyperproliferative liver is a clinically relevant situation.

HydraMap v.2: Prediction of Hydration Sites and Desolvation Energy with Refined Statistical Potentials.

The complex network of water molecules within the binding pocket of a target protein undergoes alterations upon ligand binding, presenting a significant challenge for conventional molecular modeling methods to accurately characterize and compute the associated energy changes. We have previously developed an empirical method, HydraMap (J. Chem. Inf. Model. 2020, 60, 4359-4375), which employs statistical potentials to predict hydration sites and compute desolvation energy, achieving a reasonable balance between accuracy and speed. In this work, we present its improved version, namely, HydraMap v.2. We updated the statistical potentials for protein-water interactions through an analysis of 17 042 crystal protein structures. We also introduced a new feature to evaluate ligand-water interactions by incorporating statistical potentials derived from the solvated structures of 9878 small organic molecules produced by molecular dynamics simulations. By combining these potentials, HydraMap v.2 can predict and compare the hydration sites in a binding pocket before and after ligand binding, identifying key water molecules involved in the binding process, such as those forming bridging hydrogen bonds and unstable ones that can be replaced. We demonstrated the application of HydraMap v.2 in explaining the structure-activity relationship of a panel of MCL-1 inhibitors. The desolvation energies calculated by summing the energy change of each hydration site before and after ligand binding showed good correlation with known ligand binding affinities on six target proteins. In conclusion, HydraMap v.2 offers a cost-effective solution for estimating the desolvation energy during protein-ligand binding and also is practical in guiding lead optimization in structure-based drug discovery.

A Structure-Guided Designed Small Molecule Is an Anticancer Agent and Inhibits the Apoptosis-Related MCL-1 Protein

Cancer resistance to chemotherapy and radiation therapies presents significant challenges, necessitating the exploration of alternative approaches. Targeting specific proteins at the molecular level, particularly their active sites, holds promise in addressing this issue. We investigated the potential of 4′-methoxy-2-nitrochalcone (MNC) as an MCL-1 inhibitor, examining its chemical and structural characteristics to elucidate its biological activity and guide the selection of potential candidates. We conducted a docking study, followed by synthesis, structural characterization, theoretical calculations, and in vitro experiments to comprehensively evaluate MNC. The docking results revealed MNC’s excellent binding within the active site of MCL-1. At 50 µM, MNC demonstrated 99% inhibition of HCT116 cell proliferation, with an IC50 value of 15.18 µM after 24 h. Treatment with MNC at 30.36 and 15.18 µM resulted in reduced cell density. Notably, MNC exhibited marked cytotoxicity at concentrations of 15.58 µM and 7.79 µM, inducing high frequencies of plasma membrane rupture and apoptosis, respectively. Our findings highlight the significant biological potential of MNC as an MCL-1 inhibitor. Furthermore, we propose exploring chalcones with hydrogen bond acceptor substituents as promising candidates for studying inhibitors targeting this protein. In conclusion, our study addresses the challenge of cancer resistance by investigating MNC as an MCL-1 inhibitor. Through detailed characterization and experimental validation, we establish the efficacof MNC in inhibiting cell proliferation and inducing cytotoxic effects. These results underscore the potential of MNC as a valuable therapeutic agent and suggest the use of chalcones with hydrogen bond acceptor substituents as a basis for developing novel MCL-1 inhibitors.

MCL1 Inhibition Overcomes the Aggressiveness Features of Triple-Negative Breast Cancer MDA-MB-231 Cells.

Triple-Negative Breast Cancer (TNBC) is a particularly aggressive subtype among breast cancers (BCs), characterized by anoikis resistance, high invasiveness, and metastatic potential as well as Epithelial-Mesenchymal Transition (EMT) and stemness features. In the last few years, our research focused on the function of MCL1, an antiapoptotic protein frequently deregulated in TNBC. Here, we demonstrate that MCL1 inhibition by A-1210477, a specific BH3-mimetic, promotes anoikis/apoptosis in the MDA-MB-231 cell line, as shown via an increase in proapoptotic markers and caspase activation. Our evidence also shows A-1210477 effects on Focal Adhesions (FAs) impairing the integrin trim and survival signaling pathways, such as FAK, AKT, ERK, NF-κB, and GSK3β-inducing anoikis, thus suggesting a putative role of MCL1 in regulation of FA dynamics. Interestingly, in accordance with these results, we observed a reduction in migratory and invasiveness capabilities as confirmed by a decrease in metalloproteinases (MMPs) levels following A-1210477 treatment. Moreover, MCL1 inhibition promotes a reduction in EMT characteristics as demonstrated by the downregulation of Vimentin, MUC1, DNMT1, and a surprising re-expression of E-Cadherin, suggesting a possible mesenchymal-like phenotype reversion. In addition, we also observed the downregulation of stemness makers such as OCT3/4, SOX2, NANOG, as well as CD133, EpCAM, and CD49f. Our findings support the idea that MCL1 inhibition in MDA-MB-231 could be crucial to reduce anoikis resistance, aggressiveness, and metastatic potential and to minimize EMT and stemness features that distinguish TNBC.

Molecular docking analysis of MCL-1 inhibitors for breast cancer management.

Myeloid leukemia 1 (MCL-1), a BCL-2 protein family member, acts as an anti-apoptotic protein by interacting with pro-apoptotic BCL-2 proteins. Its overexpression is frequently observed in numerous cancer types including breast cancer, and is closely linked to the initiation and progression of tumors as well as poor prognosis and resistance to therapeutic interventions. Here, a database of 3402 chemicals with established therapeutic activity against various diseases was chosen and systematically screened against the MCL-1 protein. Visual inspection and binding energy analysis revealed that the compounds OSU-03012, Raltitrexed, Ostarine (MK-2866), Dovitinib (TKI-258), and Varespladib (LY315920) had strong binding affinity for the MCL-1 protein. Notably, their binding affinity was higher than that of the control compounds. These compounds exhibited strong interactions with critical amino acid residues of the MCL-1 protein. Furthermore, these compounds shared several common amino acid residue interactions with the control compounds. These findings suggest that these compounds may be useful as MCL-1 inhibitors in the treatment of breast cancer. However, additional experimental validation is required to confirm these findings.

A PKPD Case Study: Achieving Clinically Relevant Exposures of AZD5991 in Oncology Mouse Models.

Capturing human equivalent drug exposures preclinically is a key challenge in the translational process. Motivated by the need to recapitulate the pharmacokinetic (PK) profile of the clinical stage Mcl-1 inhibitor AZD5991 in mice, we describe the methodology used to develop a refined mathematical model relating clinically relevant concentration profiles to efficacy. Administration routes were explored to achieve target exposures matching the clinical exposure of AZD5991. Intravenous infusion using vascular access button (VAB) technology was found to best reproduce clinical target exposures of AZD5991 in mice. Exposure-efficacy relationships were evaluated, demonstrating that dissimilar PK profiles result in differences in target engagement and efficacy outcomes. Thus, these data underscore the importance of accurately ascribing key PK metrics in the translational process to enable clinically meaningful predictions of efficacy.

Platinum-Based Mcl-1 Inhibitor Targeting Mitochondria Achieves Enhanced Antitumor Activity as a Single Agent or in Combination with ABT-199.

Discovery of small molecule inhibitors targeting Mcl-1 (Myeloid cell leukemia 1) confronts many challenges. Based on the fact that Mcl-1 is mainly localized in mitochondria, we propose a new strategy of targeting mitochondria to improve the binding efficiency of Mcl-1 inhibitors. We report the discovery of complex 9, the first mitochondrial targeting platinum-based inhibitor of Mcl-1, which selectively binds to Mcl-1 with high binding affinity. Complex 9 was mainly concentrated in the mitochondria of tumor cells which led to an enhanced antitumor efficacy. Complex 9 induced Bax/Bak-dependent apoptosis in LP-1 cells and synergized with ABT-199 to kill ABT-199 resistant cells in multiple cancer models. Complex 9 was effective and tolerable as a single agent or in combination with ABT-199 in mouse models. This research work demonstrated that developing mitochondria-targeting Mcl-1 inhibitors is a new potentially efficient strategy for tumor therapy.

Simultaneous Inhibition of Mcl-1 and Bcl-2 Induces Synergistic Cell Death in Hepatocellular Carcinoma.

Despite the recent approval of new therapies, the prognosis for patients with hepatocellular carcinoma (HCC) remains poor. There is a clinical need for new highly effective therapeutic options. Here, we present a combined application of BH3-mimetics as a potential new treatment option for HCC. BH3-mimetics inhibit anti-apoptotic proteins of the BCL-2 family and, thus, trigger the intrinsic apoptosis pathway. Anti-apoptotic BCL-2 proteins such as Bcl-2 and Mcl-1 are frequently overexpressed in HCC. Therefore, we analyzed the efficacy of the two BH3-mimetics ABT-199 (Bcl-2 inhibitor) and MIK665 (Mcl-1 inhibitor) in HCC cell lines with differential expression levels of endogenous Bcl-2 and Mcl-1. While administration of one BH3-mimetic alone did not substantially trigger cell death, the combination of two inhibitors enhanced induction of the intrinsic apoptosis pathway. Both drugs acted synergistically, highlighting the effectivity of this specific BH3-mimetic combination, particularly in HCC cell lines. These results indicate the potential of combining inhibitors of the BCL-2 family as new therapeutic options in HCC.

Abstract B068: Treatment combinations targeting MCL-1 and BCL-xL exert synergistic anti-tumor effects in AR-V7 expressing CRPC cell lines

Abstract Therapeutic agents that overcome acquired resistance to current androgen receptor (AR) pathway targeted therapies in patients with castration-resistant prostate cancer (CRPC) remain an unmet clinical need. A multitude of pathways are implicated in acquired treatment resistance, including the PI3K/mTOR/AKT, WNT/β-catenin, and DNA damage repair pathways. Notably, the androgen receptor splice variant 7 (AR-V7) activates the AR pathway in the absence of androgens, playing a crucial role in CRPC progression. An unbiased high throughput drug screen evaluating an array of inhibitors targeting diverse signaling pathways revealed highly synergistic activity of BH3 mimetics in AR-V7 expressing CRPC cell lines (e.g. LNCaP95, 22Rv1, VCaP-CR). Combinations targeting MCL-1 and BCL-xL (S63845 with A-1331852 or Navitoclax) displayed increased antitumor activity compared to those targeting MCL-1 and BCL-2 (S63845 with Venetoclax). Synergy among BCL-xL and MCL-1 targeted combinations was demonstrated using 2D and 3D CellTiter-Glo viability assays. Combination treatments in 3D spheroid cultures of 22RV1 and LNCaP95 cells confirmed the synergy observed in 2D assays. Furthermore, combined BCL-xL and MCL-1 inhibition altered AR-V7 expression and activated c-PARP cleavage. Drug combinations synergistically targeting BCL-xL and MCL-1 should be further explored for the treatment of therapy-resistant CRPC. Citation Format: Giulia C. Napoli, Andres F. Leon, Erica L. Beatson, Keith T. Schmidt, Emily N. Risdon, Cindy H. Chau, Douglas K. Price, William D. Figg. Treatment combinations targeting MCL-1 and BCL-xL exert synergistic anti-tumor effects in AR-V7 expressing CRPC cell lines [abstract]. In: Proceedings of the AACR Special Conference: Advances in Prostate Cancer Research; 2023 Mar 15-18; Denver, Colorado. Philadelphia (PA): AACR; Cancer Res 2023;83(11 Suppl):Abstract nr B068.

AKT inhibition sensitizes acute leukemia cells to S63845-induced apoptosis

ABSTRACTThe MCL1 inhibitors are undergoing clinical testing for multiple leukemia. However, because that MCL1 inhibition has on-target hematopoietic, hepatic and cardiac toxicities, there is substantial interest in finding agents can sensitize leukemia cells to the MCL1 inhibitors. Here we describe that the AKT inhibitors MK-2206 and Gsk690693 sensitize multiple leukemia cells to the MCL1 inhibitor S63845. Further experiments demonstrate that MK-2206 and Gsk690693 sensitize S63845 through the mitochondrial apoptosis pathway. Moreover, MK-2206 downregulates the anti-apoptotic protein BCLXL and induces the BH3-only pro-apoptotic protein BAD dephosphorylation and mitochondrial translocation. Knockdown of BAD significantly inhibits MK-2206-induced sensitization to S63845. Thus, our results suggest that MK-2206 sensitizes multiple leukemia cells to S63845-induced apoptosis, with the mechanisms involving BAD dephosphorylation and BCLXL downregulation.

Update on Small Molecule Targeted Therapies for Acute Myeloid Leukemia.

The search for effective therapies for the highly heterogenous disease acute myeloid leukemia (AML) has remained elusive. While cytotoxic therapies can induce complete remission and even, at times, long-term survival, this approach is associated with significant toxic effects to visceral organs and worsening of immune dysfunction and marrow suppression leading to death. Sophisticated molecular studies have revealed defects within the AML cell that can be exploited by utilizing small molecule agents to target these defects, often dubbed "target therapy." Several medications have already established new standards of care for many patients with AML, including FDA-approved agents that inhibitor IDH1, IDH2, FLT3, and BCL-2. Emerging small molecules hold additional to add to the armamentarium of AML treatment options including MCL-1 inhibitors, TP53 inhibitors, menin inhibitors, and E-selectin antagonists. Moreover, the increasing options also mean that future combinations of these agents need to be explored, including with cytotoxic drugs and other newer emerging strategies such as immunotherapies for AML. Recent investigations continue to show that overcoming many of the challenges of treating AML finally is on the horizon.

Design of rigid protein–protein interaction inhibitors enables targeting of undruggable Mcl-1

The structure-based design of small-molecule inhibitors targeting protein-protein interactions (PPIs) remains a huge challenge as the drug must bind typically wide and shallow protein sites. A PPI target of high interest for hematological cancer therapy is myeloid cell leukemia 1 (Mcl-1), a prosurvival guardian protein from the Bcl-2 family. Despite being previously considered undruggable, seven small-molecule Mcl-1 inhibitors have recently entered clinical trials.Here, we report the crystal structure of the clinical-stage inhibitor AMG-176 bound to Mcl-1 and analyze its interaction along with clinical inhibitors AZD5991 and S64315. Our X-ray data reveal high plasticity of Mcl-1 and aremarkable ligand-induced pocket deepening. Nuclear Magnetic Resonance (NMR)-based free ligand conformer analysis demonstrates that such unprecedented induced fit is uniquely achieved by designing highly rigid inhibitors, preorganized in their bioactive conformation. By elucidating key chemistry design principles, this work provides a roadmap for targeting the largely untapped PPI class more successfully.

Genetic events associated with venetoclax resistance in CLL identified by whole-exome sequencing of patient samples

AbstractAlthough BCL2 mutations are reported as later occurring events leading to venetoclax resistance, many other mechanisms of progression have been reported though remain poorly understood. Here, we analyze longitudinal tumor samples from 11 patients with disease progression while receiving venetoclax to characterize the clonal evolution of resistance. All patients tested showed increased in vitro resistance to venetoclax at the posttreatment time point. We found the previously described acquired BCL2-G101V mutation in only 4 of 11 patients, with 2 patients showing a very low variant allele fraction (0.03%-4.68%). Whole-exome sequencing revealed acquired loss(8p) in 4 of 11 patients, of which 2 patients also had gain (1q21.2-21.3) in the same cells affecting the MCL1 gene. In vitro experiments showed that CLL cells from the 4 patients with loss(8p) were more resistant to venetoclax than cells from those without it, with the cells from 2 patients also carrying gain (1q21.2-21.3) showing increased sensitivity to MCL1 inhibition. Progression samples with gain (1q21.2-21.3) were more susceptible to the combination of MCL1 inhibitor and venetoclax. Differential gene expression analysis comparing bulk RNA sequencing data from pretreatment and progression time points of all patients showed upregulation of proliferation, B-cell receptor (BCR), and NF-κB gene sets including MAPK genes. Cells from progression time points demonstrated upregulation of surface immunoglobulin M and higher pERK levels compared with those from the preprogression time point, suggesting an upregulation of BCR signaling that activates the MAPK pathway. Overall, our data suggest several mechanisms of acquired resistance to venetoclax in CLL that could pave the way for rationally designed combination treatments for patients with venetoclax-resistant CLL.

Discovery of an Oral, Beyond-Rule-of-Five Mcl-1 Protein-Protein Interaction Modulator with the Potential of Treating Hematological Malignancies.

Avoidance of apoptosis is critical for the development and sustained growth of tumors. The pro-survival protein myeloid cell leukemia 1 (Mcl-1) is an anti-apoptotic member of the Bcl-2 family of proteins which is overexpressed in many cancers. Upregulation of Mcl-1 in human cancers is associated with high tumor grade, poor survival, and resistance to chemotherapy. Therefore, pharmacological inhibition of Mcl-1 is regarded as an attractive approach to treating relapsed or refractory malignancies. Herein, we disclose the design, synthesis, optimization, and early preclinical evaluation of a potent and selective small-molecule inhibitor of Mcl-1. Our exploratory design tactics focused on structural modifications which improve the potency and physicochemical properties of the inhibitor while minimizing the risk of functional cardiotoxicity. Despite being in the "non-Lipinski" beyond-Rule-of-Five property space, the developed compound benefits from exquisite oral bioavailability in vivo and induces potent pharmacodynamic inhibition of Mcl-1 in a mouse xenograft model.

Combined inhibition of BCL-2 and MCL-1 overcomes BAX deficiency-mediated resistance of TP53-mutant acute myeloid leukemia to individual BH3 mimetics

TP53-mutant acute myeloid leukemia (AML) respond poorly to currently available treatments, including venetoclax-based drug combinations and pose a major therapeutic challenge. Analyses of RNA sequencing and reverse phase protein array datasets revealed significantly lower BAX RNA and protein levels in TP53-mutant compared to TP53–wild-type (WT) AML, a finding confirmed in isogenic CRISPR-generated TP53-knockout and -mutant AML. The response to either BCL-2 (venetoclax) or MCL-1 (AMG176) inhibition was BAX-dependent and much reduced in TP53-mutant compared to TP53-WT cells, while the combination of two BH3 mimetics effectively activated BAX, circumventing survival mechanisms in cells treated with either BH3 mimetic, and synergistically induced cell death in TP53-mutant AML and stem/progenitor cells. The BH3 mimetic–driven stress response and cell death patterns after dual inhibition were largely independent of TP53 status and affected by apoptosis induction. Co-targeting, but not individual targeting of BCL-2 and MCL-1 in mice xenografted with TP53-WT and TP53-R248W Molm13 cells suppressed both TP53-WT and TP53-mutant cell growth and significantly prolonged survival. Our results demonstrate that co-targeting BCL-2 and MCL-1 overcomes BAX deficiency-mediated resistance to individual BH3 mimetics in TP53-mutant cells, thus shifting cell fate from survival to death in TP53-deficient and -mutant AML. This concept warrants clinical evaluation.