Mcl-1 Inhibitors Research Articles

Osteosarcoma cells depend on MCL-1 for survival, and osteosarcoma metastases respond to MCL-1 antagonism plus regorafenib in vivo

Osteosarcoma is the most common form of primary bone cancer, which primarily afflicts children and adolescents. Chemotherapy, consisting of doxorubicin, cisplatin and methotrexate (MAP) increased the 5-year osteosarcoma survival rate from 20% to approximately 60% by the 1980s. However, osteosarcoma survival rates have remained stagnant for several decades. Patients whose disease fails to respond to MAP receive second-line treatments such as etoposide and, in more recent years, the kinase inhibitor regorafenib. BCL-2 and its close relatives enforce cellular survival and have been implicated in the development and progression of various cancer types. BH3-mimetics antagonize pro-survival members of the BCL-2 family to directly stimulate apoptosis. These drugs have been proven to be efficacious in other cancer types, but their use in osteosarcoma has been relatively unexplored to date. We investigated the potential efficacy of BH3-mimetics against osteosarcoma cells in vitro and examined their cooperation with regorafenib in vivo. We demonstrated that osteosarcoma cell lines could be killed through inhibition of MCL-1 combined with BCL-2 or BCL-xL antagonism. Inhibition of MCL-1 also sensitized osteosarcoma cells to killing by second-line osteosarcoma treatments, particularly regorafenib. Importantly, we found that inhibition of MCL-1 with the BH3-mimetic S63845 combined with regorafenib significantly prolonged the survival of mice bearing pulmonary osteosarcoma metastases. Together, our results highlight the importance of MCL-1 in osteosarcoma cell survival and present a potential therapeutic avenue that may improve metastatic osteosarcoma patient outcomes.

MCL1 inhibitor S63845 delivered by follicle-stimulating hormone modified liposome potentiates carboplatin efficacy in ovarian cancer

Platinum resistance cause therapeutic failure and poor prognosis in ovarian cancer, and evasion of apoptosis is a critical factor in chemoresistance. A limited number of FDA-approved anticancer drugs directly target apoptotic pathways. Here, we discovered that MCL1, a critical anti-apoptotic protein, is amplified and associated with platinum resistance and survival in ovarian cancer, assisting in personalized treatment. We further identified S63845 through drug-based screening, the most potent MCL1 inhibitor, which efficiently enhanced carboplatin (CBP) sensitivity in various ovarian cancer models, including primary ovarian cancer cells, orthotopic ovarian cancer, peritoneal metastasis, and human patient-derived xenograft (PDX) models. Mechanistically, S63845 competitively binds to MCL1, disrupts the binding of apoptosis effector (BAK and BAX) or pro-apoptotic BH3 protein (BIM) to MCL1 respectively, and eventually enhances CBP-induced apoptosis.To promote the clinical transformation of S63845, we developed follicle-stimulating hormone-modified liposome nanoparticles (S63845@Lipo-FSH) to enhance stability, membrane penetration, and tumor-targeting capabilities. S63845@Lipo-FSH exhibits a superior therapeutic efficacy and tumor targeting compared to free S63845, even when the dose of S63845 is reduced to one-fifth. Overall, targeting MCL1 by S63845@Lipo-FSH enhances CBP efficiency in ovarian cancer, with safety and efficacy, suggesting that this strategy is effective and promising for clinical application.

The dual BCL-2 and BCL-XL inhibitor AZD4320 acts on-target and synergizes with MCL-1 inhibition in B-cell precursor ALL

The dual BCL-2 and BCL-XL inhibitor AZD4320 acts on-target and synergizes with MCL-1 inhibition in B-cell precursor ALL

Unlocking Apoptotic Pathways: Overcoming Tumor Resistance in CAR-T-Cell Therapy.

Chimeric antigen receptor (CAR)-T-cell therapy has transformed cancer treatment, leading to remarkable clinical outcomes. However, resistance continues to be a major obstacle, significantly limiting its efficacy in numerous patients. This review critically examines the challenges associated with CAR-T-cell therapy, with a particular focus on the role of apoptotic pathways in overcoming resistance. We explore various strategies to sensitize tumor cells to CAR-T-cell-mediated apoptosis, including the use of combination therapies with BH3 mimetics, Mcl-1 inhibitors, IAP inhibitors, and HDAC inhibitors. These agents inhibit anti-apoptotic proteins and activate intrinsic mitochondrial pathways, enhancing the susceptibility of tumor cells to apoptosis. Moreover, targeting the extrinsic pathway can increase the expression of death receptors on tumor cells, further promoting their apoptosis. The review also discusses the development of novel CAR constructs that enhance anti-apoptotic protein expression, such as Bcl-2, which may counteract CAR-T cell exhaustion and improve antitumor efficacy. We assess the impact of the tumor microenvironment (TME) on CAR-T cell function and propose dual-targeting CAR-T cells to simultaneously address both myeloid-derived suppressor cells (MDSCs) and tumor cells. Furthermore, we explore the potential of combining agents like PPAR inhibitors to activate the cGAS-STING pathway, thereby improving CAR-T cell infiltration into the tumor. This review highlights that enhancing tumor cell sensitivity to apoptosis and increasing CAR-T cell cytotoxicity through apoptotic pathways could significantly improve therapeutic outcomes. Targeting apoptotic proteins, particularly those involved in the intrinsic mitochondrial pathway, constitutes a novel approach to overcoming resistance. The insights presented herein lay a robust foundation for future research and clinical applications aimed at optimizing CAR-T cell therapies.

TMCO1 is upregulated in breast cancer and regulates the response to pro-apoptotic agents in breast cancer cells

AbstractThe release of Ca2+ ions from endoplasmic reticulum calcium stores is a key event in a variety of cellular processes, including gene transcription, migration and proliferation. This release of Ca2+ often occurs through inositol 1,4,5-triphosphate receptors and the activity of these channels and the levels of stored Ca2+ in the endoplasmic reticulum are important regulators of cell death in cancer cells. A recently identified Ca2+ channel of the endoplasmic reticulum is transmembrane and coiled-coil domains 1 (TMCO1). In this study, we link the overexpression of TMCO1 with prognosis in node-positive basal breast cancer patients. We also identify interacting proteins of TMCO1, which include endoplasmic reticulum-resident proteins involved in Ca2+ regulation and proteins directly involved in nucleocytoplasmic transport. Interacting proteins included nuclear transport proteins and TMCO1 was shown to have both nuclear and endoplasmic reticulum localisation in MDA-MB-231 basal breast cancer cells. These studies also define a role for TMCO1 in the regulation of breast cancer cells in their sensitivity to BCL-2/MCL-1 inhibitors, analogous to the role of inositol 1,4,5-triphosphate receptors in the regulation of cell death pathways activated by these agents.

KS18, a Mcl-1 inhibitor, improves the effectiveness of bortezomib and overcomes resistance in refractory multiple myeloma by triggering intrinsic apoptosis.

Despite a record number of clinical studies investigating various anti-myeloma treatments, the 5-year survival rate for multiple myeloma (MM) patients in the US is only 55%, and almost all patients relapse. Poor patient outcomes demonstrate that myeloma cells are "born to survive" which means they can adapt and evolve following treatment. Thus, new therapeutic approaches to combat survival mechanisms and target treatment resistance are required. Importantly, Mcl-1, anti-apoptotic protein, is required for the development of MM and treatment resistance. This study looks at the possibility of KS18, a selective Mcl-1 inhibitor, to treat MM and overcome resistance. Our investigation demonstrates that KS18 effectively induces cell death in MM by dual regulatory mechanisms targeting the Mcl-1 protein at both transcriptional and post-translational levels. Specifically, KS18 suppresses Mcl-1 activation via STAT-3 pathway and promotes Mcl-1 phosphorylation/ubiquitination/proteasome-dependent protein degradation (UPS). Significantly, KS18 triggered caspase-dependent apoptosis in MM patient samples and bortezomib-resistant cells, synergizing with venetoclax to boost apoptosis. KS18 promises to overcome bortezomib and venetoclax resistance and re-sensitize myeloma cells to chemotherapy. Furthermore, the study shows the tremendous impact of KS18 in inhibiting colony formation in bortezomib-resistant cells and demonstrates significant tumor shrinkage in KS18-treated NSG mice without notable toxicity signs after 4weeks of therapy with a single acceptable dose each week, indicating its powerful anti-neoplastic and anti-resistance characteristics. This study strongly implies that KS18 may treat MM and provide new hope to patients who are experiencing recurrence or resistance.

Initial AXL and MCL-1 inhibition contributes to abolishing lazertinib tolerance in EGFR-mutant lung cancer cells.

Lazertinib, a novel third-generation epidermal growth factor receptor tyrosine kinase inhibitor (EGFR-TKI), demonstrates marked efficacy in EGFR-mutant lung cancer. However, resistance commonly develops, prompting consideration of therapeutic strategies to overcome initial drug resistance mechanisms. This study aimed to elucidate the adaptive resistance to lazertinib and advocate novel combination treatments that demonstrate efficacy in preventing resistance as a first-line treatment for EGFR mutation-positive NSCLC. We found that AXL knockdown significantly inhibited lung cancer cell viability in the presence of lazertinib, indicating that AXL activation contributes to lazertinib resistance. However, long-term culture with a combination of lazertinib and AXL inhibitors led to residual cell proliferation and increased the MCL-1 expression level, which was mediated by the nuclear translocation of the transcription factor YAP. Triple therapy with an MCL-1 or YAP inhibitor in combination with lazertinib and an AXL inhibitor significantly reduced cell viability and increased the apoptosis rate. These results demonstrate that AXL and YAP/MCL-1 signals contribute to adaptive lazertinib resistance in EGFR-mutant lung cancer cells, suggesting that the initial dual inhibition of AXL and YAP/MCL-1 might be a highly effective strategy in eliminating lazertinib-resistant cells.

Increasing prostate cancer radiosensitivity by miR-7-5p knockdown of anti-apoptotic genes

Despite the success of radiotherapy for prostate cancer treatment, the recent discovery of radiation resistance prevents it from reaching its full potential. This study aims to use hsa-miR-7-5p for the expression of anti-apoptotic genes. The search for anti-apoptotic genes was carried out through databases. The selected genes included XIAP, MCL1, REL, and BIRC3. Our selection was based on the best miRNA because it has a greater impact on genes. The second step involved transfecting the miRNA into a prostate cancer cell line. Subsequently, radiosensitivity was tested using real-time PCR, clonogenic assay, and annexin V flow cytometry. The highest apoptosis rate in the transfected cells was at 0 Gy in hsa-miR-7-5p (28.88 ± 0.80), plenti III (18.81 ± 0.59), and the control group (4.10 ± 1.52) (P<0.001). Also, its rate was at 4 Gy in hsa-miR-7-5p (36.11 ± 1.93), plenti III (26.42 ± 0.42), and the control group (8.79 ± 2.29) (P<0.001). This study showed a decreasing trend in survival with increasing doses. Suppression of anti-apoptotic genes, including XIAP, MCL1, Birc3, and REL, enhanced radiosensitivity by increasing the expression of hsa-miR-7-5p in the PC3 and LNCaP cell lines. Hsa-miR-7-5p is a miRNA that can suppress the expression of anti-apoptotic genes and thus plays an essential role in the process of cell apoptosis. Targeting genes that are associated with apoptosis could potentially enhance the efficacy of treatments for patients with prostate cancer.

Contribution of A1 to macrophage survival in cooperation with MCL-1 and BCL-XL in a murine cell model of myeloid differentiation

Myeloid cells are the first line of defence against pathogens. Mitochondrial apoptosis signalling is a crucial regulator of myeloid cell lifespan and modulates the function of myeloid cells. The anti-apoptotic protein BCL-2-family protein BCL2A1/A1/BFL-1 is strongly upregulated in inflammation in macrophages. We analysed the contribution of A1 to apoptosis regulation in a conditional system of in vitro differentiation of murine macrophages from immortalised progenitors. We disabled the expression of A1 by targeting all murine A1 isoforms in the genome. Specific inhibitors were used to inactivate other anti-apoptotic proteins. Macrophage progenitor survival mainly depended on the anti-apoptotic proteins MCL-1, BCL-XL and A1 but not BCL-2. Deletion of A1 on its own had little effect on progenitor cell survival but was sensitised to cell death induction when BCL-XL or MCL-1 was neutralised. In progenitors, A1 was required for survival in the presence of the inflammatory stimulus LPS. Differentiated macrophages were resistant to inhibition of single anti-apoptotic proteins, but A1 was required to protect macrophages against inhibition of either BCL-XL or MCL-1; BCL-2 only had a minor role in these cells. Cell death by neutralisation of anti-apoptotic proteins completely depended on BAX with a small contribution of BAK only in progenitors in the presence of LPS. A1 and NOXA appeared to stabilise each other at the posttranscriptional level suggesting direct binding. Co-immunoprecipitation experiments showed the binding of A1 to NOXA and BIM. Interaction between A1 and Noxa may indirectly prevent neutralisation and destabilization of MCL-1. Our findings suggest a unique role for A1 as a modulator of survival in the macrophage lineage in concert with MCL-1 and BCL-XL, especially in a pro-inflammatory environment.

Sorafenib and SIAIS361034, a novel PROTAC degrader of BCL-xL, display synergistic antitumor effects on hepatocellular carcinoma with minimal hepatotoxicity

The overexpression of BCL-xL is closely associated with poor prognosis in hepatocellular carcinoma (HCC). While the strategy of combination of BCL-xL and MCL-1 for treating solid tumors has been reported, it presents significant hepatotoxicity. SIAIS361034, a novel proteolysis targeting chimera (PROTAC) agent, selectively induces the ubiquitination and subsequent proteasomal degradation of BCL-xL through the CRBN-E3 ubiquitin ligase. When combined with sorafenib, SIAIS361034 showed a potent synergistic effect in inhibiting hepatocellular carcinoma development both in vitro and in vivo. Since SIAIS361034 exhibits a high degree of selectivity for degrading BCL-xL in hepatocellular carcinoma, the hepatotoxicity typically associated with the combined inhibition of BCL-xL and MCL-1 is significantly reduced, thereby greatly enhancing safety. Mechanistically, BCL-xL and MCL-1 sequester the BH3-only protein BIM on mitochondria at baseline. Treatment with SIAIS361034 and sorafenib destabilizes BIM/BCL-xL and BIM/MCL1 association, resulting in the liberation of more BIM proteins to trigger apoptosis. Additionally, we discovered a novel compensatory regulation mechanism in hepatocellular carcinoma cells. BIM can rapidly respond to changes in the balance between BCL-xL and MCL-1 through their co-transcription factor MEF2C to maintain apoptosis resistance. In summary, the combination therapy of SIAIS361034 and sorafenib represents an effective and safe approach for inhibiting hepatocellular carcinoma progression. The novel balancing mechanism may also provide insights for combination and precision therapies in the treatment of hepatocellular carcinoma.

MCL1 inhibitor BRD-810 kills cancer cells while minimizing risk of cardiotoxicity.

MCL1 inhibitor BRD-810 kills cancer cells while minimizing risk of cardiotoxicity.

Synthesis, Biological Evaluation, and Molecular Docking Studies of 7-Azaindole-Acylsulfonamide Derivatives as Mcl-1 Inhibitors

Synthesis, Biological Evaluation, and Molecular Docking Studies of 7-Azaindole-Acylsulfonamide Derivatives as Mcl-1 Inhibitors

A Phase 1 First-in-Human Study of the MCL-1 Inhibitor AZD5991 in Patients with Relapsed/Refractory Hematologic Malignancies.

AZD5991, a human MCL-1 inhibitor, was assessed for safety, tolerability, pharmacokinetics, and antitumor activity as monotherapy and in combination with venetoclax in patients with relapsed or refractory hematologic malignancies. In the monotherapy cohort (n = 61), patients with hematologic malignancies received AZD5991 intravenously in escalating doses either once or twice weekly, following intrapatient dose escalation, during a 3-week cycle. In the combination cohort (n = 17), patients with acute myeloid leukemia and myelodysplastic syndrome received escalating doses of AZD5991 and venetoclax during either a 3- or 4-week cycle. Primary objectives were safety and maximum tolerated dose; secondary objectives included plasma pharmacokinetics and antitumor activity. The most common (≥30%) adverse events were diarrhea (59.0%), nausea (55.1%), and vomiting (47.4%). Four deaths occurred because of adverse events: cardiac arrest, sepsis, tumor lysis syndrome, and acute respiratory failure; only tumor lysis syndrome was related to AZD5991. Dose-limiting toxicities occurred in five patients. Three patients with myelodysplastic syndrome achieved an objective response: one marrow complete remission without hematologic improvement, one partial remission with AZD5991 monotherapy, and one marrow complete remission with AZD5991 + venetoclax. Asymptomatic elevations of troponin I or T were observed in eight (10.3%) patients. Post hoc retrospective analysis revealed elevated troponin T in 14/31 patients before any AZD5991 dose and in 54/65 patients after any AZD5991 dose at or after Cycle 1. No associations were found between elevated troponin and cardiovascular risk factors. Treatment with AZD5991 was associated with high incidence of laboratory troponin elevation and a low overall response rate.

Discovery of a Myeloid Cell Leukemia 1 (Mcl-1) Inhibitor That Demonstrates Potent In Vivo Activities in Mouse Models of Hematological and Solid Tumors.

Myeloid cell leukemia 1 (Mcl-1) is a key regulator of the intrinsic apoptosis pathway. Overexpression of Mcl-1 is correlated with high tumor grade, poor survival, and both intrinsic and acquired resistance to cancer therapies. Herein, we disclose the structure-guided design of a small molecule Mcl-1 inhibitor, compound 26, that binds to Mcl-1 with subnanomolar affinity, inhibits growth in cell culture assays, and possesses low clearance in mouse and dog pharmacokinetic (PK) experiments. Evaluation of 26 as a single agent in Mcl-1 sensitive hematological and solid tumor xenograft models resulted in regressions. Co-treatment of Mcl-1-sensitive and Mcl-1 insensitive lung cancer derived xenografts with 26 and docetaxel or topotecan, respectively, resulted in an enhanced tumor response. These findings support the premise that pro-apoptotic priming of tumor cells by other therapies in combination with Mcl-1 inhibition may significantly expand the subset of cancers in which Mcl-1 inhibitors may prove beneficial.

Targeting MCL1 with Sanggenon C overcomes MCL1-driven adaptive chemoresistance via dysregulation of autophagy and endoplasmic reticulum stress in cervical cancer

BackgroundCervical cancer ranks as one of the most prevalent malignancies among women worldwide and poses a significant threat to health and quality of life. MCL1 is an antiapoptotic protein closely linked to tumorigenesis, drug-resistance and poor prognosis in various cancers. Sanggenon C, a natural flavonoid derived from Morus albal., exhibits multiple activities, including anti-oxidant, anti-inflammatory, antivirus, and antitumor properties. However, the molecular mechanisms by which Sanggenon C exerts antitumor effects on in cervical cancer remain unclear. PurposeTo investigate the oncogenic role of MCL1 and elucidate the antitumor activity of Sanggenon C, along with its molecular mechanisms, in cervical cancer. MethodsIn vitro, the effects of Sanggenon C on proliferation, the cell cycle, apoptosis, and autophagy were explored. Transcriptome sequencing was employed to analyze critical genes and pathways. The expression of genes or proteins was evaluated via immunofluorescence, qRT-PCR, immunohistochemistry, and Western blotting. To identify targets of Sanggenon C, various techniques such as clinical database analysis, molecular docking, cellular thermal shift assays, co-immunoprecipitation, and ubiquitination assays were utilized. Additionally, Xenograft mouse models were established to further investigate Sanggenon C as a novel MCL1 inhibitor and its anti-tumor activity in vivo. ResultsOur investigation reveals that Sanggenon C effectively inhibits cervical cancer cell proliferation both in vitro and in vivo. Furthermore, Sanggenon C induces endoplasmic reticulum stress and triggers protective autophagy via activation of the ATF4-DDIT3-TRIB3-AKT-MTOR signaling axis. Furthermore, Sanggenon C specifically targets MCL1 to exert its antitumor effects by modulating MCL1 protein stability through SYVN1-mediated ubiquitination. Notably, MCL1 overexpression attenuates the Sanggenon C-induced decrease in cell viability and apoptosis. Our study further characterizes the role of MCL1 in cisplatin resistance and identifies MCL1 as a promising target for Sanggenon C, which effectively inhibits proliferation and induces apoptosis in cisplatin-resistant cervical cancer cells. Importantly, combining Sanggenon C with an autophagy inhibitor represents a promising strategy to enhance therapeutic outcomes in cisplatin-resistant cervical cancer cells. ConclusionOur findings demonstrates that Sanggenon C induces endoplasmic reticulum stress and highlights the potential of targeting MCL1 to exploit vulnerabilities in drug-resistant cervical cancer cells. Sanggenon C emerges as a promising therapeutic agent against MCL1-driven adaptive chemoresistance through disruption of autophagy and endoplasmic reticulum stress in cervical cancer.

Discovery and development of thiazolidine-2,4-dione derivatives as Bcl-2/Mcl-1 dual inhibitors

Increasing the levels of antiapoptotic Bcl-2 proteins is an important way that cancer cells utilize to get out of apoptosis, underscoring their significance as promising targets for anticancer therapies. Lately, a primary compound 1 bearing thiazolidine-2,4-dione was discovered to exhibit comparable Mcl-1 inhibitory activity in comparison to WL-276. Herein, thirty-nine thiazolidine-2,4-dione analogs were yielded through incorporating different biphenyl moieties (R1), amino acid side chains (R2) and sulfonamides (R3) on 1. The findings indicated that certain compounds exhibited favorable inhibitory effects against Bcl-2/Mcl-1, while demonstrating limited or negligible binding affinity towards Bcl-xL. In particular, compounds 16 and 20 exhibited greater Bcl-2/Mcl-1 inhibition compared to AT-101, WL-276 and 1. Moreover, they demonstrated notable antiproliferative effects and significantly induced apoptosis in U937 cells. The western blot and co-immunoprecipitation assays confirmed that 20 could induce alterations in the expression of apoptosis-associated proteins to result in apoptosis through on-target Bcl-2 and Mcl-1 inhibition. In addition, 20 exhibited favorable stability profiles in both rat plasma and rat liver microsomes. In total, 20 could be used as a promising compound to discover Bcl-2/Mcl-1 dual inhibitors with favorable therapeutic properties.

Development of a scalable route to a complex cyclobutane linchpin via a diastereoselective CuI-catalyzed vinylation of a cyclobutanyl aldehyde

A robust and scalable CuI-catalyzed diastereoselective vinylation of an aliphatic aldehyde was developed to enable clinical supply of investigational Mcl-1 inhibitors in our oncology pipeline. Through process design and impurity control, a telescoped process consisting of a benzotriazole salt break and diastereoselective vinylation was developed. This process eliminated the need to perform distillation operations, improved reaction performance and efficiency, and was successfully scaled to >100 kg.

Discovery of Phenylpyrazole Derivatives as a New Class of Selective Inhibitors of MCL-1 with Antitumor Activity.

MCL-1, an antiapoptotic member of the BCL-2 family, is dysregulated and overexpressed in various tumors. In tumors with MCL-1 overexpression, selective inhibitors of MCL-1 are expected to overcome the drug resistance caused by BCL-2 inhibitors currently used in clinical treatment. Here, we employed docking-based virtual screening to identify an active hit, LC126, with binding affinity around 10 μM for MCL-1 and BCL-2. Under the guidance of structure-based design, we obtained a few selective inhibitors of MCL-1 after three rounds of structural optimization. The representative compound GQN-B37-E exhibited binding affinity for MCL-1 at the submicromolar range (K i = 0.6 μM) without apparent binding to BCL-2 or BCL-XL. 15N-heteronuclear single-quantum coherence NMR spectra suggested that this compound binds to the BH3-domain-binding pocket in the MCL-1 surface. Cellular assays revealed that GQN-B37-Me, the precursor of GQN-B37-E, is effective particularly on leukemia cells (such as H929 and MV-4-11) to induce caspase-dependent apoptosis. Its interaction with MCL-1 in cells was confirmed by coimmunoprecipitation. Administration of GQN-B37-Me to MV-4-11 xenograft mice at 50 mg/kg every 2 days for 20 days led to 43% tumor growth inhibition. GQN-B37-Me also exhibited reasonable in vitro stability in GSH and liver microsomes from several species. This new class of MCL-1 inhibitor may have potential to be further developed into a preclinical candidate for treating leukemia.

Abstract PR07: SETD1B mutations confer apoptosis resistance and BCL2 independence in B-cell lymphoma

Abstract The translocation t(14;18) activates BCL2 and is considered the initiating genetic lesion in over ninety percent of follicular lymphomas (FL). Surprisingly, FL patients respond poorly to clinically approved inhibitor, venetoclax. The mechanisms underlying BCL2 independence in FL are poorly understood, and it has been speculated that epigenetic rewiring of cell death programs may play a role. We performed separate genome-wide CRISPR/Cas9 screens to explore the mechanisms of resistance to Venetoclax and an experimental MCL-1 inhibitor (S63845) and unveiled the histone lysine methyltransferase SETD1B (KMT2G) as an unexpected mechanism of resistance to both inhibitors in lymphoma. We show that mutations and deletions affecting SETD1B occur in 7% of FLs and 16% of diffuse large B cell lymphomas (DLBCL). In vivo, SETD1B acts as a tumor suppressor gene and cooperates with the related histone-lysine N-methyltransferase 2D (KMT2D) gene leading to a more aggressive disease presentation. This oncogenic cooperation is reflected in the genomes of human lymphoma, where SETD1B mutations typically co-occur with KMT2D mutations. Mechanistically, SETD1B is required to express pro-apoptotic BCL2 family proteins like BIM and BIK. Inhibitors of the KDM5 histone H3K4 de-methylase restore the expression of BIM and BIK, venetoclax sensitivity and lead to synergistic drug effects in SETD1B deficient lymphoma xenografts. Our results highlight a role for SETD1B in the epigenetic regulation of cell death proteins in B cell lymphomas, connecting the loss of SETD1B to lymphoma development and reduced sensitivity to venetoclax. Citation Format: Ana Portelinha, Shenqiu Wang, Sara Parsa, Man Jiang, Sagarajit Mohanty, Soumya Sharma, Neeraj Arya, Omid Tavana, Jiayu Wen, Jude Fitzgibbon, Ahmet Dogan, Ana Younes, Ari M. Melnick, Hans-Guido Wendel. SETD1B mutations confer apoptosis resistance and BCL2 independence in B-cell lymphoma [abstract]. In: Proceedings of the Fourth AACR International Meeting on Advances in Malignant Lymphoma: Maximizing the Basic-Translational Interface for Clinical Application; 2024 Jun 19-22; Philadelphia, PA. Philadelphia (PA): AACR; Blood Cancer Discov 2024;5(3_Suppl):Abstract nr PR07.

DIPG-35. OVERCOMING THE BLOOD-BRAIN BARRIER CHALLENGE IN DIFFUSE MIDLINE GLIOMA

Abstract Diffuse midline gliomas (DMGs) are aggressive paediatric brainstem tumours with no known cure. The development of effective treatments is greatly impeded by the failure of most anti-tumour agents to penetrate the blood-brain barrier (BBB). Our research, along with that of others, has revealed that the BBB remains intact in DMG. We employed DMG orthotopic models to explore potential region-specific variations in response to the therapeutic agent temsirolimus as well as confocal microscopy and single-cell RNA sequencing (scRNAseq) to elucidate the impact of DMGs on brain endothelial morphology and pathways governing BBB integrity. We first assessed the effectiveness of the mTOR inhibitor temsirolimus when administered in cortex-injected DMG models compared to brainstem-injected DMG. We found significantly increased survival rates, elevated temsirolimus levels, and reduced tumour cell proliferation in the cortex versus the brainstem. Confocal imaging analysis indicated no structural differences between DMG and Matrigel-injected animals. However, scRNAseq unveiled distinct transcriptomic changes: brainstem-injected DMG exhibited downregulation of genes related to inflammatory and apoptotic pathways, whereas cortex-injected DMG showed downregulation of interferon pathways. To overcome the tightened barrier in the brainstem, we explored three MCL1 inhibitors and SNGR-TNFa (targeting CD13) as potential blood-brain barrier modulators. These inhibitors significantly reduced transendothelial electrical resistance, increased tracer dye leakage in an in vitro DMG BBB model, and decreased the protein expression of claudin-5. Moreover, in vivo, we found that administration of a single dose of either SNGR-TNFa or the MCL1 inhibitor S63845 improved penetration of Texas-Red (3KDa) in DMG-engrafted animals, indicating the effective opening of the BBB. In summary, our study revealed that DMG directly affects BBB pathways in endothelial cells, tightening the barrier and reducing treatment efficacy. MCL1 inhibitors and SNGR-TNFa show promise in modulating the BBB and have the potential to enhance therapeutic activity.