Protection of Regulatory T Cells from Fragility and Inactivation in the Tumor Microenvironment.

Fragility of regulatory T (Treg) cells manifested by the loss of neuropilin-1 (NRP1) and expression of IFNγ undermines the immune suppressive functions of Treg cells and contributes to the success of immune therapies against cancers. Intratumoral Treg cells somehow avoid fragility; however, the mechanisms by which Treg cells are protected from fragility in the tumor microenvironment are not well understood. Here, we demonstrate that the IFNAR1 chain of the type I IFN (IFN1) receptor was downregulated on intratumoral Treg cells. Downregulation of IFNAR1 mediated by p38α kinase protected Treg cells from fragility and maintained NRP1 levels, which were decreased in response to IFN1. Genetic or pharmacologic inactivation of p38α and stabilization of IFNAR1 in Treg cells induced fragility and inhibited their immune suppressive and protumorigenic activities. The inhibitor of sumoylation TAK981 (Subasumstat) upregulated IFNAR1, eliciting Treg fragility and inhibiting tumor growth in an IFNAR1-dependent manner. These findings describe a mechanism by which intratumoral Treg cells retain immunosuppressive activities and suggest therapeutic approaches for inducing Treg fragility and increasing the efficacy of immunotherapies.

Targeting PARP11 to avert immunosuppression and improve CAR T therapy in solid tumors.

Evasion of antitumor immunity and resistance to therapies in solid tumors are aided by an immunosuppressive tumor microenvironment (TME). We found that TME factors, such as regulatory T cells and adenosine, downregulated type I interferon receptor IFNAR1 on CD8+ cytotoxic T lymphocytes (CTLs). These events relied upon poly-ADP ribose polymerase-11 (PARP11), which was induced in intratumoral CTLs and acted as a key regulator of the immunosuppressive TME. Ablation of PARP11 prevented loss of IFNAR1, increased CTL tumoricidal activity and inhibited tumor growth in an IFNAR1-dependent manner. Accordingly, genetic or pharmacologic inactivation of PARP11 augmented the therapeutic benefits of chimeric antigen receptor T cells. Chimeric antigen receptor CTLs engineered to inactivate PARP11 demonstrated a superior efficacy against solid tumors. These findings highlight the role of PARP11 in the immunosuppressive TME and provide a proof of principle for targeting this pathway to optimize immune therapies.

KAT6A and ENL Form an Epigenetic Transcriptional Control Module to Drive Critical Leukemogenic Gene-Expression Programs.

Epigenetic programs are dysregulated in acute myeloid leukemia (AML) and help enforce an oncogenic state of differentiation arrest. To identify key epigenetic regulators of AML cell fate, we performed a differentiation-focused CRISPR screen in AML cells. This screen identified the histone acetyltransferase KAT6A as a novel regulator of myeloid differentiation that drives critical leukemogenic gene-expression programs. We show that KAT6A is the initiator of a newly described transcriptional control module in which KAT6A-catalyzed promoter H3K9ac is bound by the acetyl-lysine reader ENL, which in turn cooperates with a network of chromatin factors to induce transcriptional elongation. Inhibition of KAT6A has strong anti-AML phenotypes in vitro and in vivo, suggesting that KAT6A small-molecule inhibitors could be of high therapeutic interest for mono-therapy or combinatorial differentiation-based treatment of AML. SIGNIFICANCE: AML is a poor-prognosis disease characterized by differentiation blockade. Through a cell-fate CRISPR screen, we identified KAT6A as a novel regulator of AML cell differentiation. Mechanistically, KAT6A cooperates with ENL in a "writer-reader" epigenetic transcriptional control module. These results uncover a new epigenetic dependency and therapeutic opportunity in AML. This article is highlighted in the In This Issue feature, p. 587.

Epigenetic and Transcriptional Control of the Epidermal Growth Factor Receptor Regulates the Tumor Immune Microenvironment in Pancreatic Cancer.

Although immunotherapy has revolutionized cancer care, patients with pancreatic ductal adenocarcinoma (PDA) rarely respond to these treatments, a failure that is attributed to poor infiltration and activation of T cells in the tumor microenvironment (TME). We performed an in vivo CRISPR screen and identified lysine demethylase 3A (KDM3A) as a potent epigenetic regulator of immunotherapy response in PDA. Mechanistically, KDM3A acts through Krueppel-like factor 5 (KLF5) and SMAD family member 4 (SMAD4) to regulate the expression of the epidermal growth factor receptor (EGFR). Ablation of KDM3A, KLF5, SMAD4, or EGFR in tumor cells altered the immune TME and sensitized tumors to combination immunotherapy, whereas treatment of established tumors with an EGFR inhibitor, erlotinib, prompted a dose-dependent increase in intratumoral T cells. This study defines an epigenetic-transcriptional mechanism by which tumor cells modulate their immune microenvironment and highlights the potential of EGFR inhibitors as immunotherapy sensitizers in PDA. SIGNIFICANCE: PDA remains refractory to immunotherapies. Here, we performed an in vivo CRISPR screen and identified an epigenetic-transcriptional network that regulates antitumor immunity by converging on EGFR. Pharmacologic inhibition of EGFR is sufficient to rewire the immune microenvironment. These results offer a readily accessible immunotherapy-sensitizing strategy for PDA.This article is highlighted in the In This Issue feature, p. 521.

Tumor Cell-Intrinsic USP22 Suppresses Antitumor Immunity in Pancreatic Cancer.

Although immune checkpoint blockade (ICB) improves clinical outcome in several types of malignancies, pancreatic ductal adenocarcinoma (PDA) remains refractory to this therapy. Preclinical studies have demonstrated that the relative abundance of suppressive myeloid cells versus cytotoxic T cells determines the efficacy of combination immunotherapies, which include ICB. Here, we evaluated the role of the ubiquitin-specific protease 22 (USP22) as a regulator of the immune tumor microenvironment (TME) in PDA. We report that deletion of USP22 in pancreatic tumor cells reduced the infiltration of myeloid cells and promoted the infiltration of T cells and natural killer (NK) cells, leading to an improved response to combination immunotherapy. We also showed that ablation of tumor cell-intrinsic USP22 suppressed metastasis of pancreatic tumor cells in a T-cell-dependent manner. Finally, we provide evidence that USP22 exerted its effects on the immune TME by reshaping the cancer cell transcriptome through its association with the deubiquitylase module of the SAGA/STAGA transcriptional coactivator complex. These results indicated that USP22 regulates immune infiltration and immunotherapy sensitivity in preclinical models of pancreatic cancer.

Intersection of Epigenetic and Metabolic Regulation of Histone Modifications in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is one of the most lethal blood cancers, accounting for close to a quarter of a million annual deaths worldwide. Even though genetically heterogeneous, all AMLs are characterized by two interrelated features-blocked differentiation and high proliferative capacity. Despite significant progress in our understanding of the molecular and genetic basis of AML, the treatment of AMLs with chemotherapeutic regimens has remained largely unchanged in the past 30 years. In this review, we will consider the role of two cellular processes, metabolism and epigenetics, in the development and progression of AML and highlight the studies that suggest an interconnection of therapeutic importance between the two. Large-scale whole-exome sequencing of AML patients has revealed the presence of mutations, translocations or duplications in several epigenetic effectors such as DNMT3, MLL, ASXL1, and TET2, often times co-occuring with mutations in metabolic enzymes such as IDH1 and IDH2. These mutations often result in impaired enzymatic activity which leads to an altered epigenetic landscape through dysregulation of chromatin modifications such as DNA methylation, histone acetylation and methylation. We will discuss the role of enzymes that are responsible for establishing these modifications, namely histone acetyl transferases (HAT), histone methyl transferases (HMT), demethylases (KDMs), and deacetylases (HDAC), and also highlight the merits and demerits of using inhibitors that target these enzymes. Furthermore, we will tie in the metabolic regulation of co-factors such as acetyl-CoA, SAM, and α-ketoglutarate that are utilized by these enzymes and examine the role of metabolic inhibitors as a treatment option for AML. In doing so, we hope to stimulate interest in this topic and help generate a rationale for the consideration of the combinatorial use of metabolic and epigenetic inhibitors for the treatment of AML.

Tumor cell-intrinsic EPHA2 suppresses anti-tumor immunity by regulating PTGS2 (COX-2).

Resistance to immunotherapy is one of the biggest problems of current oncotherapeutics. WhileT cell abundance is essential for tumor responsiveness to immunotherapy, factors that define the T cell inflamed tumor microenvironment are not fully understood. We conducted an unbiased approach to identify tumor-intrinsic mechanisms shaping the immune tumor microenvironment(TME), focusing on pancreatic adenocarcinoma because it is refractory to immunotherapy and excludes T cells from the TME. From human tumors, we identified EPHA2 as a candidate tumor intrinsic driver of immunosuppression. Epha2 deletion reversed T cell exclusion and sensitized tumors to immunotherapy. We found that PTGS2, the gene encoding cyclooxygenase-2, lies downstream of EPHA2 signaling through TGFß and is associated with poor patient survival. Ptgs2 deletion reversed T cell exclusion and sensitized tumors to immunotherapy; pharmacological inhibition of PTGS2 was similarly effective. Thus, EPHA2-PTGS2 signaling in tumor cells regulates tumor immune phenotypes; blockade may represent a novel therapeutic avenue for immunotherapy-refractory cancers. Our findings warrant clinical trials testing the effectiveness of therapies combining EPHA2-TGFß-PTGS2 pathway inhibitors with anti-tumor immunotherapy, and may change the treatment of notoriously therapy-resistant pancreatic adenocarcinoma.

Tumor Cell-Intrinsic Factors Underlie Heterogeneity of Immune Cell Infiltration and Response to Immunotherapy.

The biological and functional heterogeneity between tumors-both across and within cancer types-poses a challenge for immunotherapy. To understand the factors underlying tumor immune heterogeneity and immunotherapy sensitivity, we established a library of congenic tumor cell clones from an autochthonous mouse model of pancreatic adenocarcinoma. These clones generated tumors that recapitulated T cell-inflamed and non-T-cell-inflamed tumor microenvironments upon implantation in immunocompetent mice, with distinct patterns of infiltration by immune cell subsets. Co-injecting tumor cell clones revealed the non-T-cell-inflamed phenotype is dominant and that both quantitative and qualitative features of intratumoral CD8+ T cells determine response to therapy. Transcriptomic and epigenetic analyses revealed tumor-cell-intrinsic production of the chemokine CXCL1 as a determinant of the non-T-cell-inflamed microenvironment, and ablation of CXCL1 promoted T cell infiltration and sensitivity to a combination immunotherapy regimen. Thus, tumor cell-intrinsic factors shape the tumor immune microenvironment and influence the outcome of immunotherapy.

MyD88 NEDDylation negatively regulates MyD88-dependent NF-κB signaling through antagonizing its ubiquitination.

Myeloid differentiation factor 88 (MyD88) plays a central role in innate immunity response, however, how its activity is tightly regulated remains largely unknown. In this study, we identify MyD88 as a novel substrate of NEDD8, and demonstrate that MyD88 NEDDylation antagonizes its ubiquitination. Interestingly, in response to the stimulation of IL-1ß, MyD88 NEDDylation is downregulated while its ubiquitination is upregulated. We also show that deNEDDylase NEDP1 serves as a regulator of this process. Furthermore, we demonstrate that NEDD8 negatively regulates the dimerization of MyD88 and suppresses MyD88-dependent NF-κB signaling. Taken together, this study reveals that NEDDylation of MyD88 regulates NF-κB activity through antagonizing its ubiquitination, suggesting a novel mechanism of modulating NF-κB signaling pathway.