Role Of MYC Research Articles

Overcoming retinoic acid resistance in HER2-enriched breast cancers: role of MYC.

HER2-enriched (HER2+) breast cancers express high levels of the growth-promoting HER2 protein. Although these cancers are treated with the HER2-targeted drug, trastuzumab, resistance to treatment is common. Retinoic acid (RA) is an anti-cancer agent that has been successfully used for the treatment of leukemia and holds promise for the treatment of solid cancers, including breast cancer. The HER2 gene is frequently co-amplified with RARA, a key determinant of RA sensitivity in breast cancers. It seems surprising, therefore, that HER2+ breast cancers are refractory to RA treatment. Here, we show that MYC mediates RA resistance by suppressing the expression of cellular retinoic acid binding protein 2 (CRABP2), resulting in RARα inactivation. CRABP2 is an intracellular RA transporter that delivers RA to the nuclear receptor RARα for its activation. Our results indicate that response to RA is enhanced by MYC depletion in HER2+ breast cancer cells and that RA treatment enhances trastuzumab responsiveness. Our findings support the use of RA and trastuzumab for the treatment of subsets of patients with breast cancers that are HER2-RARα co-amplified and have low levels of MYC.

Drosophila: a Tale of regeneration with MYC.

Regeneration is vital for many organisms, enabling them to repair injuries and adapt to environmental changes. The mechanisms underlying regeneration are complex and involve coordinated events at the cellular and molecular levels. Moreover, while some species exhibit remarkable regenerative capabilities, others, like mammals, have limited regenerative potential. Central to this process is the regulation of gene expression, and among the numerous genes involved, MYC emerges as a regulator of relevant processes during regeneration with roles conserved in several species, including Drosophila. This mini-review aims to provide valuable insights into the regeneration process in flies, focusing on significant organs where the role of MYC has been identified: from the imaginal discs, where MYC regulates cell growth, structure, and proliferation, to the gut, where it maintains the balance between renewal and differentiation of stem cells, and the central nervous system, where it influences the activities of neural stem cells and the interaction between glia and neuronal cells. By emphasizing the molecular mechanisms regulated by MYC, its significance in controlling regeneration mechanisms, and its conserved role in flies, we aim to offer valuable insights into the utility of Drosophila as a model for studying regeneration. Moreover, unraveling MYC's function in Drosophila during regeneration may help translate findings into the mechanisms underlying human tissue repair.

MYC and NCAPG2 as molecular targets of colorectal cancer and gastric cancer in nursing.

Colorectal cancer is a common malignant tumor in intestinal tract, the early symptoms are not obvious. Gastric cancer is a malignant tumor originating from the gastric mucosal epithelium. However, the role of MYC and non-SMC condensin II complex subunit G2 (NCAPG2) in colorectal cancer and gastric cancer remains unclear. The colorectal cancer datasets GSE49355 and gastric cancer datasets GSE19826 were downloaded from gene expression omnibus database. Differentially expressed genes (DEGs) were screened and weighted gene co-expression network analysis (WGCNA) was performed. Functional enrichment analysis, gene set enrichment analysis (GSEA) and immune infiltration analysis was performed. Construction and analysis of protein-protein interactions (PPI) network. Survival analysis and comparative toxicogenomics database (CTD) were performed. A heat map of gene expression was drawn. A total of 751 DEGs were obtained. According to the gene ontology (GO) analysis, in Biological process (BP) analysis, they are mainly enriched in cell differentiation, cartilage development, and skeletal development. In cellular component (CC) analysis, they are mainly enriched in the cytoskeleton of muscle cells and actin filaments. In molecular function (MF) analysis, they are mainly concentrated in Rho GTPase binding, DNA binding, and fibronectin binding. In Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis, they are mainly enriched in the MAPK signaling pathway, apoptosis, and cancer pathways. The soft threshold power for WGCNA analysis was set to 9, resulting in the generation of 40 modules. Ultimately, 2 core genes (MYC and NCAPG2) were identified. The heatmap of core gene expression showed high expression of MYC and NCAPG2 in colorectal cancer tissue samples and low expression in normal tissue samples, while they were core molecules in gastric cancer. Survival analysis indicated that MYC and NCAPG2 were risk factors, showing an upregulation trend with increasing risk scores. CTD analysis revealed associations of MYC and NCAPG2 with colorectal cancer, gastric cancer, inflammation, and immune system diseases. MYC and NCAPG2 are highly expressed in colorectal cancer. The higher the expression of MYC and NCAPG2, the worse the prognosis. MYC and NCAPG2 are core molecules in gastric cancer.

Hold the MYCrophone: MYC Invades Enhancers to Control Cancer-Type Gene Programs.

MYC is an oncogenic transcription factor that binds gene promoters to facilitate oncogenic gene expression. When overexpressed, as is the case in most human cancers, MYC also invades active enhancers-cis-regulatory elements that are critical for regulating gene expression. In previous studies, the regulatory significance of MYC enhancer invasion in cancer cells has been debated. In their study published in Nature Genetics, Jakobsen and colleagues establish a new role for MYC in enhancer regions: regulating cancer type-specific gene programs. Their work reveals a mechanism in which MYC cooperates with other oncogenic transcription factors to recruit epigenetic regulators to enhancers, resulting in an epigenetic "switch" that promotes enhancer activation through BRD4 and RNA polymerase II. This activity was highly cancer-type specific, highlighting gene expression programs that predicted clinical outcome in a subtype-specific manner in patients with breast cancer.

Abstract 5976: R9: A novel NuRD complex inhibitor for targeted therapy in triple negative breast cancer (TNBC)

Abstract Triple Negative Breast Cancer (TNBC) is a highly aggressive breast cancer subtype, disproportionately affecting young and African American populations. TNBC constitutes up to 15% of breast cancers that lack effective targeted therapy. The Nucleosome Remodeling and Deacetylase (NuRD) complex, a pivotal chromatin remodeling entity, plays a crucial role in cancer development and progression. Specifically, the NuRD complex associates with SALL4 in cancer cells, contributing to the silencing of tumor-suppressor genes like PTEN. In this study, we discover R9, a novel small-molecule nucleotide analog, a first-in-class NuRD inhibitor. Utilizing computational and cellular methods, R9 demonstrated selective engagement with RBBP4, a key NuRD complex subunit. Cellular target engagement technology has been utilized to verify direct interaction of R9 with NuRD complex as well as to guide medicinal chemistry efforts. Mass spectrometry thermal proteome profiling (TPP) affirmed RBBP4 as the primary target of R9 with minimal off-target effects. In vitro studies reveal R9 displaying robust toxicity in TNBC cells, while sparing normal tissue derived cells. Encouragingly, R9 effectively impeded TNBC tumors growth in mice without inducing toxicity. Mechanistically, we demonstrate R9 induces DNA damage in sensitive TNBC cells, and the sensitivity correlates to DNA damage-vulnerable cancer cells, indicating DNA damage is a major effector of NuRD complex inhibition in TNBCs. We further demonstrate that R9 induces DNA damage through a c-MYC dependent manner. Ongoing studies aim to elucidate key transcription factors involved in NuRD complex inhibition and the role of MYC and DNA damage in response to NuRD complex inhibition. This research positions R9 as a promising candidate for targeted therapy in TNBC, offering potential insights into the intricate molecular pathways involved in TNBC tumor progression. Citation Format: Kun Zhang, Sheeba Jacob, Mikhail Dozmorov, Ivan Babic, Elmar Nurmemmedov, Anthony C. Faber. R9: A novel NuRD complex inhibitor for targeted therapy in triple negative breast cancer (TNBC) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5976.

Pharmacological inhibition of MYC to mitigate chemoresistance in preclinical models of squamous cell carcinoma.

Rationale: Cisplatin-based chemotherapy is the first-line treatment for late-stage head and neck squamous cell carcinoma (HNSCC). However, resistance to cisplatin has become a major obstacle for effective therapy. Cancer stem cells (CSCs) are critical for tumor initiation, growth, metastasis, and chemoresistance. How to effectively eliminate CSCs and overcome chemoresistance remains a key challenge. Herein, we confirmed that MYC plays critical roles in chemoresistance, and explored targeting MYC to overcome cisplatin resistance in preclinical models. Methods: The roles of MYC in HNSCC cisplatin resistance and cancer stemness were tested in vitro and in vivo. The combined therapeutic efficiency of MYC targeting using the small molecule MYC inhibitor MYCi975 and cisplatin was assessed in a 4‑nitroquinoline 1-oxide-induced model and in a patient-derived xenograft model. Results: MYC was highly-expressed in cisplatin-resistant HNSCC. Targeting MYC using MYCi975 eliminated CSCs, prevented metastasis, and overcame cisplatin resistance. MYCi975 also induced tumor cell-intrinsic immune responses, and promoted CD8+ T cell infiltration. Mechanistically, MYCi975 induced the DNA damage response and activated the cGAS-STING-IRF3 signaling pathway to increase CD8+ T cell-recruiting chemokines. Conclusions: Our findings suggested that targeting MYC might eliminate CSCs, prevent metastasis, and activate antitumor immunity to overcome cisplatin resistance in HNSCC.

Abstract C104: Targeted epigenomic inhibition of MYC enhances responses to immune checkpoint and EGFR inhibitors in preclinical models of NSCLC

Abstract Aberrant expression of the transcription factor MYC is found in most cancers including non-small cell lung cancer (NSCLC) and has critical roles in a wide range of oncogenic processes including immune evasion and EGFR inhibitor resistance. Despite MYC’s therapeutic attractiveness, potent inhibitors of MYC activity have been elusive, due to its intrinsically disordered protein structure and tightly autoregulated expression. We have developed NSCLC-specific programmable epigenomic mRNA therapy, termed a MYC Omega epigenomic controller (NSCLC MYC-OEC) that is designed to target the insulated genomic domain of MYC. We have previously shown that NSCLC MYC-OEC effectively downregulates MYC expression pre-transcriptionally and have demonstrated that NSCLC MYC-OEC inhibits NSCLC patient-derived organoid viability in vitro and abrogates xenograft tumor growth in vivo. To identify combination treatments with NSCLC MYC-OEC, we retrospectively evaluated baseline MYC expression and real-world progression-free survival (PFS) in a cohort of 283 NSCLC patients treated with pembrolizumab, an anti-PD1 monoclonal antibody. This analysis revealed that NSCLC patients with MYC-overexpressing tumors exhibited shorter PFS when treated with pembrolizumab. Moreover, analysis of PD1 or PD-L1 transcript levels in tumors from 13,230 lung adenocarcinoma patients revealed a significant positive correlation between PD1 or PD-L1 and MYC mRNA levels, consistent with a role for MYC in regulation of the immune response. In vivo evaluation of NSCLC MYC-OEC activity and immune checkpoint immunotherapy (ICI) treatment of preclinical Lewis lung carcinoma syngeneic tumor models demonstrated that the combination of NSCLC MYC-OEC with anti-PD-L1 antibody enhanced inhibition of tumor growth and prolonged survival over administration when compared to either NSCLC MYC-OEC or anti-PD-L1 antibody monotherapy. Retrospective analysis of a cohort of 301 NSCLC patients also showed that patients with MYC-overexpressing tumors exhibited significantly shorter PFS when treated with osimertinib, a third generation EGFR inhibitor. To further understand this, we investigated the effects of NSCLC MYC-OEC and osimertinib treatment alone or in combination in EGFR mutant cells, H1975 and PC9. While NSCLC MYC-OEC and osimertinib monotherapy treatments resulted in downregulation of MYC protein expression in both cell lines, combination treatment led to a significantly greater decrease in MYC expression. Furthermore, the combination synergistically reduced H1975 and PC9 cell viability in vitro and led to a significantly greater inhibition of H1975 xenograft growth relative to each single agent in vivo. Together, these data provide preclinical proof-of-concept in NSCLC for development of OEC-mediated pre-transcriptional epigenomic inhibition of MYC expression in combination with immune checkpoint and EGFR inhibitors. Furthermore, these findings suggest that NSCLC MYC-OEC combination with ICI or EGFR-targeted therapy may improve patient outcomes. Citation Format: Defne Yarar, Eugine Lee, Padraich Flahardy, Cameron Vergato, Graeme Hodgson, Thomas McCauley. Targeted epigenomic inhibition of MYC enhances responses to immune checkpoint and EGFR inhibitors in preclinical models of NSCLC [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2023 Oct 11-15; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2023;22(12 Suppl):Abstract nr C104.

Mechanism of co-operation of mutant IL-7Rα and mutant NRAS in acute lymphoblastic leukemia: role of MYC.

Acute lymphoblastic leukemia (ALL) is an aggressive leukemia which can be derived from either T-cell or B-cell precursors. With current treatments, the survival rate is high, but the treatments are highly toxic with severe side effects. Individual mutations in IL7Ra and RAS pathways have been previously shown to be prevalent in ALL, and especially in relapsed patients. The relationship of IL-7Ra and RAS was investigated by transducing immature mouse thymocytes with the combination of these mutants. The resultant ALL cells were analyzed to identify the regulators and the oncoproteins that are up-regulated or down-regulated by the combination of IL7Ra with NRAS. Leukemia cells showed a significant increase in IL7Ra-mediated BCL2 expression, and an increase in MYC protein levels was mainly induced by NRAS signaling. MYC was both necessary and sufficient to replace mutant NRAS, and drugs targeting the MYC pathway showed a therapeutic benefit in IL-7Ra/NRAS T-ALL. We suggest that MYC protein stability can be regulated by PLK-1 kinase, which was increased mainly by the NRAS signal. These studies identify novel pathways of oncogenesis and new targets for intervention that could lead to better therapeutic development.

Exploring the role of lactylation-related genes in osteosarcoma: A deep dive into prognostic significance and therapeutic potential.

Osteosarcoma (OS), notorious for its complex pathogenesis and formidable prognosis, represents a significant medical quandary. This research embarked on a quest to unravel the implications of lactylation-related genes (LRGs) in OS, offering a novel lens through which to interpret its intricacies. A meticulous evaluation of 329 LRGs within the TARGET dataset spotlighted 27 paramount genes, intricately intertwined with survival. These genes highlighted metabolic processes-particularly amino acid metabolism-as key players, as evidenced in both GO and KEGG analyses. Utilizing consensus clustering and principal component analysis, the 93 OS samples were segmented into two distinct groups, differing notably in overall and event-free survival. Cluster 2 demonstrated a heightened immune response, contrasting the other cluster. Machine learning techniques, like generalized boosted model, CoxBoost, and RSF, spotlighted MYC and GOT2 as critical genes. Using multivariate Cox regression, a risk model was developed, categorizing patients into high and low-risk groups, each displaying varied survival patterns. Additionally, a contrast was observed between MYC and GOT2's associations with HLA molecules, emphasizing their distinct roles in antigen presentation. Potential therapeutic avenues were identified for each risk group, with special attention to mutations in MYC, particularly amplifications, hinting at its role in tumor progression. Finally, delving deeper into the role of MYC, Western blot analyses exhibited amplified myc protein levels in OS cells U-2 and MG-63 when juxtaposed against human osteoblastic cells Hfob1.19. A focused knockdown of myc in OS cells subsequently confirmed its influence on cell proliferation and migration, with reduced myc expression resulting in inhibited cell activities. Furthermore, immunofluorescence assays corroborated myc's heightened expression in OS cells relative to normal osteoblastic cells. In summary, this study accentuates the vital role of LRGs and specifically MYC in OS, ushering in a horizon of tailored therapeutic strategies.

Tumor Suppressive Mir-7 Targeting PSME3 Improves the Efficacy of Histone Deacetylases Inhibitors in Multiple Myeloma

Background: Histone deacetylases (HDACs) play a tumor-promoting role in multiple myeloma (MM), contributing to disease progression and treatment resistance. Panobinostat, an HDAC inhibitor, is clinicallyrelapsed or refractory MM. However, HDAC inhibitors have several limitations, such as drug resistance, poor tolerance, and limited use of combination drugs. Our previous comprehensive analysis of microRNAs modulated by HDAC inhibitors revealed new molecular pathogenesis and potential therapeutic targets in malignant lymphomas. However, the relationship between HDAC inhibitors and microRNAs in MM remains largely unknown. Aim and methods: We treated the MM cell lines (MM.1S and KMS-11) with HDAC inhibitors (10 nM panobinostat and 1 µM vorinostat), collected RNA 48 h later, and conducted a comprehensive microarray analysisFurthermore, we explored the novel clinical impact of HDAC inhibitors via functional analysis of microRNAs. Results: Microarray analysis showed that no microRNAs were commonly upregulated; however, only hsa-miR-7-5p (miR-7) was commonly downregulated in MM.1S and KMS-11 cells treated with both HDAC inhibitors. After exposing MM cell lines (MM.1S, KMS-11, RPMI-8226, H929, and U266) and patient samples to 10 nM panobinostat and 1 µM vorinostat for 48 h, we performed quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and observed significant downregulation in the expression of miR-7. Next, we examined the function of miR-7 in MM by introducing miR-7 into MM cell lines. Introducing miR-7 significantly suppressed the proliferation of MM.1S and KMS-11 cells. We hypothesized that the introduction of miR-7 enhances the effects of HDAC inhibitors. We introduced miR-7 into MM.1S and cultured them for 48 h with panobinostat, followed by an apoptosis assay. The introduction of miR-7 significantly enhanced the apoptosis-inducing effect of panobinostat, suggesting a tumor-suppressive role of miR-7 in MM. Furthermore, we examined the regulatory mechanisms of miR-7 in MM cells. Because it has been reported that MYC promotes miR-7 transcription, we focused on the role of MYC in miR-7 expression in MM. After exposing MM.1S and KMS-11 cells to HDAC inhibitors for 24 h, western blot analysis revealed that MYC was downregulated. After exposing MM.1S and KMS-11 cells to the BRD4 inhibitor JQ1, which suppresses MYC, at concentrations of 50 and 100 nM for 48 h, qRT-PCR showed that the expression of miR-7 was significantly downregulated in these cell lines. Similarly, introducing siRNA for MYC into MM.1S cells downregulated the expression of miR-7. These results suggest that HDAC inhibitors downregulate miR-7 expression by suppressing MYC expression. In addition, we examined the effect of bortezomib, a proteasome inhibitor, in combination with MM on the expression of miR-7. After exposing MM.1S, KMS-11, and H929 cells to 5 nM and 10 nM bortezomib for 48 h, we performed qRT-PCR and observed a significant upregulation in the expression of miR-7. Therefore, anti-myeloma effects of panobinostat by restoring miR-7 expression. Finally, we investigated miR-7 targets that mediate the anti-myeloma effects. We introduced miR-7 into MM.1S cells via electroporation, collected RNA 48 h later, and performed microarray analysis. As a result, 26 genes were identified, and their expression was downregulated by less than one-third. Among these, four genes ( SLC6A9, LRRC59, EXOSC2, and PSME3) possess conserved binding sites for miR-7. PSME3 (proteasome activator subunit 3：PA28ɤ) has been reported as an oncogene in MM, and we focused on PSME3. Introducing miR-7 into MM.1S cells via electroporation significantly suppressed PSME3 expression. Furthermore, when MM.1S cells were exposed to HDAC inhibitors, PSME3 expression was significantly upregulated. It has been reported that the inhibition of PSME3 increases sensitivity to proteasome inhibitors in MM. The favorable combination effect of bortezomib with HDAC inhibitors could be due to the modulation of the miR-7- PSME3 axis. Conclusion: We revealed that miR-7 exerts anti-myeloma effects and that PSME3 is one of the targets of miR-7. miR-7 is an interesting microRNA because it is positively regulated by MYC, even though it is tumor-suppressive. Elucidating the role of miR-7 may lead to the re-discovery of HDAC inhibitors in MM therapeutic strategies.

Epigenetic Regulators Open the Door to SCLC Plasticity.

Small-cell lung cancer (SCLC) is a neuroendocrine tumor type with limited treatment options and poor prognosis. SCLC comprises multiple molecular subtypes that are defined by the expression of the lineage-related transcription factors ASCL1, NEUROD1, POU2F3, and more controversially, YAP1. SCLC exhibits remarkable plasticity with the capacity to transition between molecular states; because these states are associated with unique therapeutic susceptibilities, SCLC has been likened to a moving therapeutic target. While MYC's role in driving the ASCL1-to-NEUROD1 (A-to-N) transition is established, additional mechanisms governing SCLC plasticity remain largely obscure. A recent study by Duplaquet and colleagues, published in Nature Cell Biology, employs an innovative genetically engineered mouse model of SCLC harboring loss of KDM6A-a histone lysine demethylase mutated in approximately 2% of SCLC cases. KDM6A loss in SCLC alters chromatin accessibility and increases the potential for A-to-N plasticity in vivo. Through characterization of the epigenetic landscape, Duplaquet and colleagues identified histone methylation as a key regulator of SCLC plasticity. These findings provide not only a new model system for studying SCLC plasticity, but also identify new epigenetic mechanisms involved, which will ultimately be critical for designing more effective therapies.

Unraveling MYC's Role in Orchestrating Tumor Intrinsic and Tumor Microenvironment Interactions Driving Tumorigenesis and Drug Resistance.

The transcription factor MYC plays a pivotal role in regulating various cellular processes and has been implicated in tumorigenesis across multiple cancer types. MYC has emerged as a master regulator governing tumor intrinsic and tumor microenvironment interactions, supporting tumor progression and driving drug resistance. This review paper aims to provide an overview and discussion of the intricate mechanisms through which MYC influences tumorigenesis and therapeutic resistance in cancer. We delve into the signaling pathways and molecular networks orchestrated by MYC in the context of tumor intrinsic characteristics, such as proliferation, replication stress and DNA repair. Furthermore, we explore the impact of MYC on the tumor microenvironment, including immune evasion, angiogenesis and cancer-associated fibroblast remodeling. Understanding MYC's multifaceted role in driving drug resistance and tumor progression is crucial for developing targeted therapies and combination treatments that may effectively combat this devastating disease. Through an analysis of the current literature, this review's goal is to shed light on the complexities of MYC-driven oncogenesis and its potential as a promising therapeutic target.

Myc beyond Cancer: Regulation of Mammalian Tissue Regeneration.

Myc is one of the most well-known oncogenes driving tumorigenesis in a wide variety of tissues. From the brain to blood, its deregulation derails physiological pathways that grant the correct functioning of the cell. Its action is carried out at the gene expression level, where Myc governs basically every aspect of transcription. Indeed, in addition to its role as a canonical, chromatin-bound transcription factor, Myc rules RNA polymerase II (RNAPII) transcriptional pause-release, elongation and termination and mRNA capping. For this reason, it is evident that minimal perturbations of Myc function mirror malignant cell behavior and, consistently, a large body of literature mainly focuses on Myc malfunctioning. In healthy cells, Myc controls molecular mechanisms involved in pivotal functions, such as cell cycle (and proliferation thereof), apoptosis, metabolism and cell size, angiogenesis, differentiation and stem cell self-renewal. In this latter regard, Myc has been found to also regulate tissue regeneration, a hot topic in the research fields of aging and regenerative medicine. Indeed, Myc appears to have a role in wound healing, in peripheral nerves and in liver, pancreas and even heart recovery. Herein, we discuss the state of the art of Myc's role in tissue regeneration, giving an overview of its potent action beyond cancer.

A MYC-controlled redox switch protects B lymphoma cells from EGR1-dependent apoptosis

Refractory and relapsed B cell lymphomas are often driven by the difficult-to-target oncogene MYC. Here, we report that high MYC expression stimulates proliferation and protects B lymphoma cells from apoptosis under normal oxidative stress levels and that compounds including N-acetylcysteine (NAC) and vitamin C (VitC) induce apoptosis by reducing oxidative stress. NAC and VitC injections effectively reduce tumor growth in lymphoma cells with high MYC expression but not in those with low MYC expression. MYC knockdown confers tumor resistance to NAC and VitC, while MYC activation renders B cells sensitive to these compounds. Mechanistically, NAC and VitC stimulate MYC binding to EGR1 through Cys117 of MYC, shifting its transcriptional output from cell cycle to apoptosis gene expression. These results identify a redox-controlled mechanism for MYC's role in maintaining proliferation and preventing apoptosis, offering a potential therapeutic rationale for evaluating NAC or VitC in patients with MYC-driven B cell lymphoma.

Abstract 6256: Inhibition of class IIa HDACs potentiates MYC inhibitor-driven cytotoxicity by inducing MYC depletion and oxidative stress in non-small cell lung cancer

Abstract Lung cancer is the most common cause of cancer mortality worldwide and non-small cell lung cancer (NSCLC) accounts for approximately 85% of all lung cancers. Dysregulation and/or overexpression of MYC are widely implicated in NSCLC, which suggests that MYC may be a promising therapeutic target for the treatment of NSCLC. Recently, two direct MYC inhibiting agents with improved in vivo efficacy and tolerability, MYCi975 and Omomyc, have been developed. To define new drug strategies that could potentiate the therapeutic effects of MYC inhibitors, transcriptome datasets from MYCi975 and Omomyc studies were reanalyzed. These data revealed the induction of two class IIa histone deacetylases (HDACs), HDAC5 and HDAC9, upon MYC inhibition. Importantly, class IIa HDACs are involved in cancer proliferation, aggressiveness, and prognosis. In addition, novel agents that selectively inhibit class IIa HDACs have demonstrated promising anti-tumor efficacy; thus, suggesting potential therapeutic benefits from dual targeting of MYC and class IIa HDACs. We assessed the treatment efficacy of MYCi plus class IIa HDACi across 18 NSCLC cell lines, 10 of which demonstrated substantial reduction of cell viability upon combination treatment. Querying of driver mutation spectra associated with differential drug response in our cell line panel defined that EGFR mutant cell lines exhibited resistance, while STK11 or Ras mutant cell lines were sensitive to our drug paradigm. Gene set enrichment analysis comparing the transcriptomes of sensitive versus resistant cell lines revealed that MYC Targets, Oxidative Phosphorylation, and Reactive Oxygen Species (ROS) pathways were activated in combination treatment responsive cell lines. RNA sequencing on treated cell lines identified large-scale transcriptional shifts facilitated by combination treatment, including suppression of MYC, cell cycle, and mitochondrial pathways. Furthermore, G1/S arrest and elevated mitochondrial ROS levels in combination drug-treated cells were confirmed using flow cytometry. Concordant with the role of MYC as a key cell cycle regulator, MYC protein levels were decreased in the combination treatment group as compared to the vehicle or mono-treatment groups. Importantly, both MYC overexpression and ROS scavenger treatment partially rescued the reduction of cell viability by combination treatment, suggesting both MYC depletion and ROS elevation as contributors to therapeutic efficacy driven by combination treatment. Expanding these studies to in vivo models, we discern that combination MYC and class IIa HDAC inhibition significantly reduced tumor burden in a patient-derived xenograft model of human NSCLC. In summary, we define here a novel drug paradigm combining MYC and class IIa HDAC inhibitors, which potentiates anti-tumor efficacy in NSCLC via MYC depletion and oxidative stress. Citation Format: Jina Park, Ying-Yu Chen, Jennie Cao, Michelle Vaz, Ray-Whay C. Yen, Stephen B. Baylin, Michael J. Topper. Inhibition of class IIa HDACs potentiates MYC inhibitor-driven cytotoxicity by inducing MYC depletion and oxidative stress in non-small cell lung cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6256.

Immuno-Genetic Changes Underlie Response to Immune Checkpoint Blockade Therapy in Richter's Syndrome Mouse Models

Richter's syndrome (RS) represents one of the foremost clinical challenges in CLL, due to the disappointing responses to both standard chemotherapy and novel agents. Checkpoint blockade (CPB) with anti-PD-1 antibody treatment has shown promise in RS (but minimal efficacy in CLL), suggesting that distinct immuno-genetic features underlie differential response to immunotherapy of these two entities. We previously generated CRISPR-based multiplexed gene-edited mouse models of RS in immune competent backgrounds with genetic, epigenetic, transcriptional and functional similarity with human disease, thus enabling interrogation of changes in tumor microenvironmental (TME) features underlying CLL transformation. To this end, we analyzed the T-cell and monocyte/macrophage composition of 5 CLL, 3 CLL/RS (i.e. cases with equal co-existence of CLL and RS), and 9 murine RS cases by multi-parametric flow cytometry of splenocyte preparations, and compared them with 5 age-matched wild-type controls (15-18 month old). Whereas the overall proportional abundance of CD3+ T cells was not significantly changed across groups, we detected decreased CD4+ and increased CD8+ T cell populations in RS cases (P≤0.04, ANOVA). Both CD4+ and CD8+ T cells showed a prominent skewing toward the effector memory subset (CD44+ CD62L-), when compared to naïve (CD44- CD62L+) and central memory (CD44+ CD62L+). CD4+ and CD8+ T cell populations from RS cases also showed significant upregulation of exhaustion markers including PD-1, TIGIT, TIM3 and LAG3 (P≤0.05). The monocytic compartment was similarly skewed, with RS-associated enrichment in non-classical monocytes (CD43+ LyC6med/low, known to have lower antigen presentation capacity), when compared to classical inflammatory CD43+ LyC6high cells (P=0.04). Importantly, we measured increased abundance of CD11b+ Ly6C- tumor associated macrophages in RS cases (P=0.02), which expressed significantly reduced MHC class II (P=0.003), and increased PD-1 surface levels (P=0.02). Further, in 11 cases (3 CLL/RS, 8 RS) where at least 5-10% residual CLL cells co-existed with RS within the same animal, and the two populations were discernable by size, we measured increased expression levels of both PD-1 and PD-L1 ligands on RS compared to CLL by flow cytometry (P≤0.001, paired t test), consistent with surface PD-1 observed in human clonally related RS. Interestingly, surface PD-L1 levels were particularly enriched in RS cases carrying clonal deletion in Mga, a negative regulator of MYC (P=0.01, Mann Whitney), in line with the known role of MYC in upregulating PD-L1 gene expression. In light of these changes, we asked whether our animal models could be utilized as a reliable platform to study differential response to CPB. To this end, we leveraged transplantable models in immune competent mice. In our initial study, 40 million RS cells from a Trp53-deleted primary tumor were transplanted into 20 CD45.1 recipients (2 million/recipient), which were randomized either 1 [median disease burden: 2%(1-13.5)] or 2 weeks [37%(7-54)] post-transplant through peripheral blood (PB) flow cytometric assessment of circulating disease. Treatment with 200 micrograms rat IgG2a anti-PD-1 or isotype control (n=5 mice/group) was performed for 3 weeks, every 3 days, and animals euthanized 3 days after the last dose. Whereas animals with highest disease burden at treatment start (2 weeks post-transplant) showed minimal, if any, efficacy of CPB treatment, animals with lower disease burden at time of randomization (1 week post-transplant) showed significant reduction in RS infiltration of PB, spleen and bone marrow at euthanasia (P≤0.04, Mann Whitney). This suggests that disease burden may impact differential response to CPB. In conclusion, our analyses demonstrate marked exhaustion of the T cell and monocytic compartment and increased expression of PD-1 and PD-L1 on tumor cells and TME components upon RS transformation. Initial analyses showed variable benefit from single agent anti-PD-1 therapy in Trp53-mutant disease. We are currently exploring combinatorial strategies with targeted therapy (including PI3K and BTK inhibitors) to improve in vivo response rates in the context of tumor lines with distinct genetics and characterizing the immune microenvironmental changes induced by these treatments.

Advances in Understanding of Metabolism of B-Cell Lymphoma: Implications for Therapy.

There have been significant recent advances in the understanding of the role of metabolism in normal and malignant B-cell biology. Previous research has focused on the role of MYC and mammalian target of rapamycin (mTOR) and how these interact with B-cell receptor signaling and hypoxia to regulate glycolysis, glutaminolysis, oxidative phosphorylation (OXPHOS) and related metabolic pathways in germinal centers. Many of the commonest forms of lymphoma arise from germinal center B-cells, reflecting the physiological attenuation of normal DNA damage checkpoints to facilitate somatic hypermutation of the immunoglobulin genes. As a result, these lymphomas can inherit the metabolic state of their cell-of-origin. There is increasing interest in the potential of targeting metabolic pathways for anti-cancer therapy. Some metabolic inhibitors such as methotrexate have been used to treat lymphoma for decades, with several new agents being recently licensed such as inhibitors of phosphoinositide-3-kinase. Several other inhibitors are in development including those blocking mTOR, glutaminase, OXPHOS and monocarboxylate transporters. In addition, recent work has highlighted the importance of the interaction between diet and cancer, with particular focus on dietary modifications that restrict carbohydrates and specific amino acids. This article will review the current state of this field and discuss future developments.

Abstract 1466: MYC localizes to the nuclear pore to promote DNA repair during replication stress

Abstract Genomic instability is a hallmark of cancer which promotes oncogenic mutations in pancreatic ductal adenocarcinoma (PDAC), leading to more aggressive tumors with greater potential for drug resistance. Deregulation of the master transcription factor, MYC, is found in virtually all PDAC tumors and promotes genomic alterations by increasing replication stress, augmenting DNA repair pathways, and promoting cell survival. To resolve replication stress, stalled forks are trafficked to the nuclear pores where the necessary machinery accumulates. Activated MYC is spatially reorganized to the nuclear pores as well, a mechanism that is exacerbated in cancer. Whether MYC activity at the nuclear pores contributes to resolving replication stress in PDAC remains to be known. To investigate this mechanism, we are creating genomic and proteomic tools to determine MYC’s function at the nuclear pores during replication stress induced by olaparib treatment in PDAC cells. We found that olaparib significantly increases MYC accumulation at the nuclear pore in low-passage cell lines we established from PDAC patient tumors. Furthermore, we showed that MYC interacts with the nuclear pore resident SUMO protease, SENP1, which plays a key role in resolving stalled forks. Our preliminary data suggest a role for MYC resolving replication stress at nuclear pores, and could indicate MYC activity as a therapeutic target in olaparib-resistant PDAC. Citation Format: Gabriel Cohn, Colin J. Daniel, Daniel F. Liefwalker, Rosalie C. Sears. MYC localizes to the nuclear pore to promote DNA repair during replication stress [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1466.

Myc Supports Self-Renewal of Basal Cells in the Esophageal Epithelium.

It is widely believed that cellular senescence plays a critical role in both aging and cancer, and that senescence is a fundamental, permanent growth arrest that somatic cells cannot avoid. Here we show that Myc plays an important role in self-renewal of esophageal epithelial cells, contributing to their resistance to cellular senescence. Myc is homogeneously expressed in basal cells of the esophageal epithelium and Myc positively regulates their self-renewal by maintaining their undifferentiated state. Indeed, Myc knockout induced a loss of the undifferentiated state of esophageal epithelial cells resulting in cellular senescence while forced MYC expression promoted oncogenic cell proliferation. A superoxide scavenger counteracted Myc knockout-induced senescence, therefore suggesting that a mitochondrial superoxide takes part in inducing senescence. Taken together, these analyses reveal extremely low levels of cellular senescence and senescence-associated phenotypes in the esophageal epithelium, as well as a critical role for Myc in self-renewal of basal cells in this organ. This provides new avenues for studying and understanding the links between stemness and resistance to cellular senescence.

MYC Inhibition Halts Metastatic Breast Cancer Progression by Blocking Growth, Invasion, and Seeding.

While MYC role in metastasis has been long controversial, this manuscript demonstrates that MYC inhibition by either transgenic expression or pharmacologic use of the recombinantly produced Omomyc miniprotein exerts antitumor and antimetastatic activity in breast cancer models in vitro and in vivo, suggesting its clinical applicability.