The use of epigenetic therapy to activate an immune response in ovarian cancer

描述（由申请人提供）：大约67名美国妇女中有一名在她的一生中会患上上皮性卵巢癌（EOC）。晚期疾病占病例的70%，五年生存率不到25%。在目前的护理标准下，手术后化疗，EOC女性的生存率在过去三十年中变化不大。显然需要EOC的新疗法。表观遗传药物正在被探索作为EOC和其他实体瘤的新疗法。肺癌患者的临床成功表明，DNA甲基转移酶抑制剂5-氮杂胞苷（AZA）和组蛋白去乙酰化酶抑制剂恩替诺特的作用可能会刺激免疫应答并靶向免疫细胞以根除肿瘤。此外，AZA已被FDA批准用于治疗急性髓性白血病。癌症的特征在于在肿瘤抑制基因的启动子处抑制染色质，包括DNA甲基化和组蛋白的脱乙酰化。AZA抑制DNA甲基化，恩替司他抑制组蛋白脱乙酰酶，打开染色质并引起受抑制基因的转录。在上皮性卵巢癌中，免疫基因的高表达和免疫细胞浸润到肿瘤中预示着更好的预后。我们从23个EOC细胞系用AZA处理的初步数据显示参与免疫应答的基因上调。这些AZA诱导的免疫基因（AIM基因）代表先天性和适应性免疫的途径，聚集来自TCGA的EOC肿瘤，并指示一组具有低AIM表达的肿瘤。在这项授权中，我建议使用AZA来刺激EOC细胞的免疫反应。这种疗法可以单独对一部分患者起作用，但应该使更多的患者对随后的免疫疗法敏感。我假设AZA诱导的AIM去抑制将导致癌细胞死亡和宿主免疫细胞向肿瘤的募集，并且可以使卵巢癌细胞对随后的免疫治疗敏感。AZA激活上皮细胞免疫反应的机制尚不清楚。AZA的去甲基化可引起免疫主调节因子的重新表达。我观察到转录正常沉默的RNA，如人类内源性逆转录病毒（ERVs）和AZA治疗后的重复元件，这可能会激活免疫（病毒防御）反应。另外，AZA可以诱导肿瘤上免疫检查点配体PD-L1的表达。因此，我提出将表观遗传免疫调节与PD-1/PD-L1通路的抑制相结合可能会导致对EOC肿瘤的协同抗肿瘤作用。目的1：确定AZA免疫应答的表型效应和机制。目的2：确定内源性逆转录病毒和转座因子RNA是否参与AZA免疫应答。目标3：我将在卵巢癌的免疫活性小鼠模型中用AZA刺激免疫应答，然后用抗PD-1抗体阻断免疫逃避，并评估宿主免疫细胞活化和肿瘤细胞杀伤。因此，我将评估表观遗传学与治疗相结合的疗效，以打破人类最致命的恶性肿瘤之一的免疫耐受。

DESCRIPTION (provided by applicant): Approximately one out of 67 U.S. women will get epithelial ovarian cancer (EOC) in her lifetime. Late-stage disease accounts for 70% of cases, with a dismal five year survival rate of less than 25%. Under the current standard of care, surgery followed by chemotherapy, survival of women with EOC has changed very little over the past three decades. Novel therapies for EOC are clearly needed. Epigenetic agents are being explored as new therapies for EOC and other solid tumors. Clinical success in patients with lung cancer suggests that the actions of the DNA methyltransferase inhibitor 5-azacytidine (AZA) and the histone deacetylase inhibitor entinostat may stimulate the immune response and target immune cells to eradicate tumors. Furthermore AZA has FDA approval for treatment of acute myeloid leukemia. Cancers are characterized by repressive chromatin, including DNA methylation and deacetylation of histones, at promoters of tumor suppressor genes. AZA inhibits DNA methylation and entinostat inhibits histone deacetylases, opening up chromatin and causing transcription of repressed genes. In EOC, higher expression of immune genes and immune cell infiltration into tumors predict better prognosis. Our preliminary data from 23 EOC cell lines treated with AZA show an upregulation of genes involved in the immune response. These AZA-Induced iMmune genes (AIM genes), representing pathways of both innate and adaptive immunity, cluster EOC tumors from TCGA and indicate a group of tumors with low AIM expression. In this grant I propose to use AZA to stimulate an immune response in EOC cells. This therapy could work alone for a subset of patients but should sensitize a larger group of patients to subsequent immune therapy. I hypothesize that an AZA-induced de-repression of AIMs will lead to cancer cell death and recruitment of host immune cells to the tumor and can sensitize ovarian cancer cells to subsequent immunotherapy. The mechanism by which AZA activates an epithelial cell immune response is unknown. Demethylation by AZA could cause re-expression of immune master regulators. I observe transcription of normally silenced RNAs such as human endogenous retroviruses (ERVs) and repetitive elements upon AZA treatment, which may activate the immune (viral defense) response. Separately, AZA can induce expression of the immune checkpoint ligand PD-L1 on tumors. Thus, I propose that combining epigenetic immune modulation with inhibition of the PD-1/PD-L1 pathway may result in synergistic anti-tumor effects against EOC tumors. Aim 1: I will determine the phenotypic effects and mechanism of the AZA immune response. Aim 2: I will determine whether endogenous retrovirus and transposable element RNAs are responsible for the AZA immune response. Aim 3: I will treat with AZA to stimulate the immune response, then an anti-PD-1 antibody to block immune evasion, in immunocompetent mouse models of ovarian cancer and assess host immune cell activation and tumor cell killing. I will thus assess the efficacy of combining epigenetic with therapy to break immune tolerance in one of the deadliest human malignancies.