Integrating Epigenetic Modulation into DNA Damage Repair

癌症中遗传性DNA修复突变的筛查随着同源重组修复而加速 (HRR)缺陷型肿瘤对DNA损伤剂和聚（ADP-核糖）聚合酶抑制剂反应良好 （PARPi）.最常见的HRD突变包括BRCA 1、BRCA 2、ATM和CHEK 2。这种突变传达了 在所有种族和民族背景中，40-80%的终生乳腺癌风险，并对以下疾病构成高风险： 卵巢癌前列腺癌和胰腺癌虽然毒性较小，但对PARP抑制剂的反应通常仍然很短， 尽管最初的答复率很高。PARP抑制剂在ATM和CHEK 2中的功效是机械预期的， 但尚未建立。重叠的毒性显著挑战了将联合收割机PARPi与 化疗和免疫疗法组合在大多数乳腺癌亚型中仍然是有限的益处。 为了提高PARPi的功效，我们研究了增加DNA捕获和DNA抑制的策略。 修复. DNA甲基转移酶（DNMT）直接调节DNA修复途径，并与DNA修复酶复合作用。 PARP修复单链DNA断裂。因此，我们假设DNMT抑制将显著地 改善PARP抑制在HRD癌症中的治疗益处。我们发现了增强的PARP捕获， 非常低剂量的DNMT抑制剂、地西他滨和PARPi的有希望的协同功效， 在体外和体内研究中，在基因工程HRR突变的癌细胞中显著增强 线和PDX模型。对联合治疗的反应因肿瘤（亚）组织背景和选择的HRD而异 基因突变我们的工作的初步数据已经导致了一项剂量探索I期试验的批准， 由联盟网络（A092003）提供。在本申请中，我们将探索敏感性的生物标志物和机制 以及对PARP和DNMT联合治疗的耐药性，以提供更深入的机制见解和指导 临时批准的大型随机ComboMatch试验（EAY 191 A4）中的患者选择有三个目的。 目的1：确定联合PARP和DNMT的协同作用机制和比较效果 在异种移植物中，在体外，在携带4种不同HRR途径基因的靶向突变的同基因细胞系中的抑制， 并与具有相似HRR突变但不同遗传背景的已建立PDX进行比较。 目的2：使用治疗前和治疗后的肿瘤活检和I期患者的系列血液样本 Alliance Network（A092003）试验，以详细了解患者的HRR突变， 基因组景观和HRR途径的功能状态。建立患者来源的异种移植物（PDX）， 目标3中关于耐药机制的工作的活检标本。评估循环肿瘤DNA（ctDNA） 用于预测治疗反应的生物标志物。 目的3：从临床转换回实验室，测试PDX中治疗反应的准确性， 类器官模型与源患者反应相比，识别PARPi+DNMTi治疗抗性 机制，测试对潜在后续治疗的敏感性。

Screening for hereditary DNA repair mutations in cancer has accelerated as Homologous Recombination Repair (HRR) deficient tumors respond well to DNA damaging agents and poly (ADP-ribose) polymerase inhibitors (PARPi). The most common HRD mutations include BRCA1, BRCA2, ATM, and CHEK2. Such mutations convey a 40-80% lifetime breast cancer risk across all racial and ethnic backgrounds, and pose elevated risks for ovarian, prostate and pancreatic cancer. While less toxic, responses to PARP inhibitors are still often short, despite a high initial response rate. Efficacy of PARP inhibitors in ATM and CHEK2 is mechanistically expected, but not yet established. Overlapping toxicities have significantly challenged the ability to combine PARPi with chemotherapy, and immunotherapy combinations remain of limited benefit in most breast cancer subtypes. In a quest to enhance PARPi efficacy, we studied strategies to increase DNA trapping and inhibition of DNA repair. DNA methyltransferases (DNMTs) directly modulate the DNA repair pathway and work in complex with PARP to repair single strand DNA breaks. As such, we hypothesize that DNMT inhibition would significantly improve the therapeutic benefit of PARP inhibition in HRD cancer. We found enhanced PARP trapping and promising synergistic efficacy with very low doses of the DNMT inhibitor, decitabine, and PARPi in preliminary in vitro and in vivo studies which was significantly enhanced in genetically engineered HRR mutated cancer cell lines and PDX models. Responsiveness to the combination varied by tumor (sub) tissue context and select HRD gene mutation. Preliminary data form our work has led to the approval of a dose finding phase I trial sponsored by the Alliance Network (A092003). In this application, we will explore biomarkers and mechanisms of sensitivity and resistance to combination PARP and DNMT treatment to provide deeper mechanistic insights and guide patient selection in the provisionally-approved large randomized ComboMatch trial (EAY191 A4) in three aims. Aim 1: Determine the mechanism of synergy and comparative effects of combined PARP and DNMT inhibition in isogenic cell lines bearing targeted mutation of 4 different HRR pathway genes, in vitro, in xenografts, and in comparison to established PDX with similar HRR mutations but different genetic backgrounds. Aim 2: Use pre- and posttreatment tumor biopsy and serial blood samples from patients in the Phase I Alliance Network (A092003) trial to generate a detailed understanding of patient’s HRR mutation, accompanying genomic landscape, and functional status of the HRR pathway. Establish Patient Derived Xenografts (PDX) from biopsy specimens for work in Aim 3 regarding resistance mechanisms. Assess circulating tumor DNA (ctDNA) for predictive biomarkers of therapeutic response. Aim 3: Translating from the clinic back to the bench, test the veracity of the therapeutic responses in PDX and organoid models compared to the source patient responses, identify PARPi+DNMTi therapeutic resistance mechanisms, test sensitivity to potential followup therapeutics.