Polycomb repressor complex 2 suppresses interferon-responsive MHC-II expression in melanoma cells and is associated with anti-PD-1 resistance.

BACKGROUND: Despite the remarkable success of immunotherapy in treating melanoma, understanding of the underlying mechanisms of resistance remains limited. Emerging evidence suggests that upregulation of tumor-specific major histocompatibility complex-II (tsMHC-II) serves as a predictive marker for the response to anti-programmed death-1 (PD-1)/programmed death ligand 1 (PD-L1) therapy in various cancer types. The genetic and epigenetic pathways modulating tsMHC-II expression remain incompletely characterized. Here, we provide evidence that polycomb repressive complex 2 (PRC2)/EZH2 signaling and resulting H3K27 hypermethylation suppresses tsMHC-II. METHODS: RNA sequencing data from tumor biopsies from patients with cutaneous melanoma treated with or without anti-PD-1, targeted inhibition assays, and assays for transposase-accessible chromatin with sequencing were used to observe the relationship between EZH2 inhibition and interferon (IFN)-γ inducibility within the MHC-II pathway. RESULTS: We find that increased EZH2 pathway messenger RNA (mRNA) expression correlates with reduced mRNA expression of both presentation and T-cell genes. Notably, targeted inhibition assays revealed that inhibition of EZH2 influences the expression dynamics and inducibility of the MHC-II pathway following IFN-γ stimulation. Additionally, our analysis of patients with metastatic melanoma revealed a significant inverse association between PRC2-related gene expression and response to anti-PD-1 therapy. CONCLUSIONS: Collectively, our findings demonstrate that EZH2 inhibition leads to enhanced MHC-II expression potentially resulting from improved chromatin accessibility at CIITA, the master regulator of MHC-II. These insights shed light on the molecular mechanisms involved in tsMHC-II suppression and highlight the potential of targeting EZH2 as a therapeutic strategy to improve immunotherapy efficacy.

Multi-omics analysis identifies therapeutic vulnerabilities in triple-negative breast cancer subtypes.

Triple-negative breast cancer (TNBC) is a collection of biologically diverse cancers characterized by distinct transcriptional patterns, biology, and immune composition. TNBCs subtypes include two basal-like (BL1, BL2), a mesenchymal (M) and a luminal androgen receptor (LAR) subtype. Through a comprehensive analysis of mutation, copy number, transcriptomic, epigenetic, proteomic, and phospho-proteomic patterns we describe the genomic landscape of TNBC subtypes. Mesenchymal subtype tumors display high mutation loads, genomic instability, absence of immune cells, low PD-L1 expression, decreased global DNA methylation, and transcriptional repression of antigen presentation genes. We demonstrate that major histocompatibility complex I (MHC-I) is transcriptionally suppressed by H3K27me3 modifications by the polycomb repressor complex 2 (PRC2). Pharmacological inhibition of PRC2 subunits EZH2 or EED restores MHC-I expression and enhances chemotherapy efficacy in murine tumor models, providing a rationale for using PRC2 inhibitors in PD-L1 negative mesenchymal tumors. Subtype-specific differences in immune cell composition and differential genetic/pharmacological vulnerabilities suggest additional treatment strategies for TNBC.

MEK inhibition activates STAT signaling to increase breast cancer immunogenicity via MHC-I expression.

AIM: Immunotherapy and immune checkpoint inhibitors (ICI) have changed cancer care for many patients; however, breast cancers have exhibited minimal response to single agent ICI therapy. There is a significant need to identify novel targets capable of increasing cancer cell immunogenicity and response to ICIs in breast cancer. Mitogen activated protein kinase (MAPK) signaling is essential for many cellular processes but the relationship between MAPK signaling and cancer cell immunogenicity is less well understood. Recent reports suggest that MEK inhibition (MEKi) affects the tumor-immune microenvironment by altering the expression of interferon responsive PD-L1 and MHC-I through unknown mechanisms. METHODS: Using western blotting and flow cytometry, we sought to determine whether MEKi affects JAK-STAT signaling upstream of PD-L1 and MHC-I expression in a panel of mouse mammary cancer and triple negative breast cancer cell lines. RESULTS: The cell lines tested exhibited increased STAT activation in response to MEKi treatment. Furthermore, MEKi-induced MHC-I and PD-L1 expression are dependent upon STAT1 in MMTV-Neu cells. Interestingly, MEKi-induced STAT activation and interferon-responsive protein expression are abrogated with ErbB-family inhibitor co-treatment in MMTV-Neu cells, suggesting ErbB receptor signaling dependence, but not in basal-like cell lines. Importantly, analysis of basal-like breast cancer patient samples exhibited an inverse relationship between STAT1 and Ras/MAPK activation signatures. CONCLUSION: These findings suggest that MAPK signaling and STAT activation are inversely related in both mouse and human mammary tumors. This work also supports further study of MEKi to increase STAT signaling and potentially, immunotherapy responses through increased MHC-I and PD-L1 expression.

Biomarker predictors for immunotherapy benefit in breast: beyond PD-L1.

PURPOSE OF REVIEW: Immune checkpoint blockade (ICB) has changed the clinical course of multiple cancer types and durable responses have now been observed in breast cancer (BC) patients. Most data suggest that, compared to other subtypes, triple-negative BC (TNBC) patients are more responsive to ICB, and anti-PD-L1 therapy is now approved in PD-L1+ metastatic TNBC, in combination with chemotherapy. RECENT FINDINGS: Nearly 40% of PD-L1+ TNBC patients did not respond to this combination. Thus, additional biomarkers appear to be necessary to more precisely identify potential responders. A comprehensive analysis of the breast tumor microenvironment (TME) and peripheral blood may identify potential biomarkers for a more accurate selection of patients likely to respond to ICB. SUMMARY: Herein, we summarize key features of the breast TME, and beyond, that may hold predictive power in determining immunotherapy benefit. Incorporation of these features in controlled clinical trials may help further guide personalized care for BC immunotherapy.

Treatment-Induced Tumor Cell Apoptosis and Secondary Necrosis Drive Tumor Progression in the Residual Tumor Microenvironment through MerTK and IDO1.

Efferocytosis is the process by which apoptotic cells are cleared from tissue by phagocytic cells. The removal of apoptotic cells prevents them from undergoing secondary necrosis and releasing their inflammation-inducing intracellular contents. Efferocytosis also limits tissue damage by increasing immunosuppressive cytokines and leukocytes and maintains tissue homeostasis by promoting tolerance to antigens derived from apoptotic cells. Thus, tumor cell efferocytosis following cytotoxic cancer treatment could impart tolerance to tumor cells evading treatment-induced apoptosis with deleterious consequences in tumor residual disease. We report here that efferocytosis cleared apoptotic tumor cells in residual disease of lapatinib-treated HER2+ mammary tumors in MMTV-Neu mice, increased immunosuppressive cytokines, myeloid-derived suppressor cells (MDSC), and regulatory T cells (Treg). Blockade of efferocytosis induced secondary necrosis of apoptotic cells, but failed to prevent increased tumor MDSCs, Treg, and immunosuppressive cytokines. We found that efferocytosis stimulated expression of IFN-γ, which stimulated the expression of indoleamine-2,3-dioxegenase (IDO) 1, an immune regulator known for driving maternal-fetal antigen tolerance. Combined inhibition of efferocytosis and IDO1 in tumor residual disease decreased apoptotic cell- and necrotic cell-induced immunosuppressive phenotypes, blocked tumor metastasis, and caused tumor regression in 60% of MMTV-Neu mice. This suggests that apoptotic and necrotic tumor cells, via efferocytosis and IDO1, respectively, promote tumor 'homeostasis' and progression. SIGNIFICANCE: These findings show in a model of HER2+ breast cancer that necrosis secondary to impaired efferocytosis activates IDO1 to drive immunosuppression and tumor progression.

Selenoprotein Expression in Macrophages Is Critical for Optimal Clearance of Parasitic Helminth Nippostrongylus brasiliensis.

The plasticity of macrophages is evident in helminthic parasite infections, providing protection from inflammation. Previously we demonstrated that the micronutrient selenium induces a phenotypic switch in macrophage activation from a classically activated (pro-inflammatory; M1/CAM) toward an alternatively activated (anti-inflammatory; M2/AAM) phenotype, where cyclooxygenase (COX)-dependent cyclopentenone prostaglandin J2 (15d-PGJ2) plays a key role. Here, we hypothesize that dietary selenium modulates macrophage polarization toward an AAM phenotype to assist in the increasing clearance of adult Nippostrongylus brasiliensis, a gastrointestinal nematode parasite. Mice on a selenium-adequate (0.08 ppm) diet significantly augmented intestinal AAM presence while decreasing adult worms and fecal egg production when compared with infection of mice on selenium-deficient (<0.01 ppm) diet. Further increase in dietary selenium to supraphysiological levels (0.4 ppm) had very little or no impact on worm expulsion. Normal adult worm clearance and enhanced AAM marker expression were observed in the selenium-supplemented Trsp(fl/fl)Cre(WT) mice that express selenoproteins driven by tRNA(Sec) (Trsp), whereas N. brasiliensis-infected Trsp(fl/fl)Cre(LysM) selenium-supplemented mice showed a decreased clearance, with lowered intestinal expression of several AAM markers. Inhibition of the COX pathway with indomethacin resulted in delayed worm expulsion in selenium-adequate mice. This was rescued with 15d-PGJ2, which partially recapitulated the effect of selenium supplementation on fecal egg output in addition to increasing markers of AAMs in the small intestine. Antagonism of PPARγ blocked the effect of selenium. These results suggest that optimal expression of selenoproteins and selenium-dependent production of COX-derived endogenous prostanoids, such as Δ(12)-PGJ2 and 15d-PGJ2, may regulate AAM activation to enhance anti-helminthic parasite responses.