Integrating Epigenetic Modulation into DNA Damage Repair

随着同源重组修复的加速，癌症中遗传性DNA修复突变的筛查已经加速 (HRR)缺陷肿瘤对DNA损伤剂和多聚(ADP-核糖)聚合酶抑制剂有良好的反应 (PARPI)。最常见的HRD突变包括BRCA1、BRCA2、ATM和CHEK2。这样的突变传达了 一生中患乳腺癌的风险在所有种族和民族背景中都有40%-80%，并对 卵巢癌、前列腺癌和胰腺癌。虽然毒性较小，但对PARP抑制剂的反应仍然很短， 尽管最初的应答率很高。PARP抑制剂在ATM和CHEK2中的疗效是机械预期的， 但还没有建立起来。重叠的毒性显著地挑战了将PARPI与 化疗和免疫联合治疗在大多数乳腺癌亚型中的益处仍然有限。 为了提高PARPI的有效性，我们研究了增加DNA捕获和DNA抑制的策略 修理。DNA甲基转移酶(DNMT)直接调节DNA修复途径，并与 PARP修复单链DNA断裂。因此，我们假设DNMT的抑制将显著 提高PARP抑制治疗HRD癌的疗效。我们发现了增强的PARP陷阱和 DNMT抑制剂、地西他滨和PARPI极低剂量的初步协同疗效 基因工程HRR突变癌细胞的体内外研究显着增强 系列和PDX型号。对组合的反应因肿瘤(亚)组织背景和选择的HRD而不同 基因突变。来自我们工作的初步数据导致了剂量发现第一阶段试验的批准 由联盟网络提供(A092003)。在这项应用中，我们将探索生物标志物和敏感性机制 以及对PARP和DNMT联合治疗的抵抗力，以提供更深层次的机制见解和指导 在临时批准的大型随机组合试验(EAY191 A4)的三个目标中的患者选择。 目的1：确定PARP与DNMT联合应用的协同作用机制及效果比较 对携带4种不同HRR途径基因靶向突变的同基因细胞系的抑制作用，体外，在异种移植瘤中， 与已建立的具有相似HRR突变但遗传背景不同的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 联盟网络(A092003)试验，以产生对患者HRR突变的详细了解，伴随 基因组景观，以及HRR途径的功能状态。建立患者来源的异种移植(PDX) 目标3中有关耐药机制工作的活组织标本。检测循环肿瘤DNA(CtDNA) 用于预测治疗反应的生物标志物。 目标3：从临床转回工作台，测试PDX治疗反应的准确性 将器官模型与来源患者的反应进行比较，确定PARPI+DNMTi治疗耐药 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.