ATR inhibitor-mediated reversal of PARP inhibitor resistance in high-grade serous ovarian cancer (HGSOS)

总结 高级别浆液性卵巢癌（HGSOC）经常在DNA修复途径中存在缺陷， 同源重组（HR）修复缺陷，以及停滞的DNA复制的稳定性受损 叉子这些缺陷通常涉及BRCA改变，并赋予对聚（ADP-核糖）抑制剂的敏感性。 聚合酶（PARP）抑制剂。这些特工现在已经进入了常规的HGSOC医疗设备， 高级和维护设置。该项目的首要目标是解决新出现的问题， 随着PARP抑制剂使用的增加，PARP抑制剂耐药性将变得更加重要。项目 重点是抑制共济失调毛细血管扩张和Rad 3相关（ATR）作为一种策略，旨在扭转这两个 获得性PARP抑制剂耐药的主要机制，包括HR修复的恢复和 DNA复制叉。提出了三个具体目标。在目标1中，我们将评估活动和机制 ATR抑制剂AZD 6738作为单一疗法和与PARP抑制剂奥拉帕尼组合在体外和体内的作用。 HGSOC的体内BRCA突变细胞系、类器官培养物和患者来源的异种移植物（PDX）模型， 获得性PARP抑制剂耐药性。这项工作将为Aim 2联合AZD 6738的临床试验铺平道路 和奥拉帕尼，在PARP受体耐药BRCA患者中使用最大化ATR抑制的方案， 突变的HGSOC一旦确定了最大耐受剂量，我们将确认耐受性并评估 在12名患者的扩展队列中的初步抗肿瘤活性。将进行成对活检， 机制验证药效学研究，其中使用免疫组织化学RAD 51评估HR 测定和复制叉稳定性用类器官培养物中的DNA纤维测定来评估。在目标3中，我们 在HGSOC中测定单独和与吉西他滨组合的ATR抑制活性， 但本质上对PARP抑制无反应。AZD 6738和吉西他滨将 在具有高水平CCNE 1或MYC扩增的细胞系、类器官和PDX模型中研究。我们将分析 这些模型中的复制应激及其通过ATR抑制和吉西他滨的加重， ATR途径活化的免疫组织化学、细胞学和基因表达特征生物标志物。最后， 我们将利用最近完成的吉西他滨与吉西他滨联合ATR抑制剂的研究 M6620在铂类耐药HGSOC患者中的疗效，其中联合用药在伴有以下症状的患者中具有上级优势： 无铂间隔时间少于3个月。我们将检验这组人群中肿瘤富集的假设 携带高度复制应激，由CCNE 1或MYC扩增以及生物标志物定义 在临床前模型中开发的。总之，这些目标将使我们能够建立ATR抑制的作用 在HGSOC医疗设施中，为预后不良的HGSOC人群确定新的治疗途径 并提供对PARP抑制剂抗性生物学的见解。

SUMMARY High-grade serous ovarian cancer (HGSOC) frequently harbors defects in DNA repair pathways that confer homologous recombination (HR) repair deficiency, as well as compromised stability of stalled DNA replication forks. Such defects frequently involve BRCA alterations and confer sensitivity to inhibitors of poly (ADP-ribose) polymerase (PARP) inhibitors. These agents have now entered the routine HGSOC armamentarium both in the advanced and maintenance settings. An overarching objective of this project is to address the emerging problem of PARP inhibitor resistance that will assume greater importance as PARP inhibitor use increases. The project focuses on inhibition of Ataxia telangiectasia and Rad3-related (ATR) as a strategy designed to reverse the two major mechanisms of acquired PARP inhibitor resistance, including restoration of HR repair and stabilization of DNA replication forks. Three Specific Aims are proposed. In Aim 1, we will assess the activity and mechanisms of the ATR inhibitor AZD6738 as monotherapy and in combination the PARP inhibitor olaparib in in vitro and in vivo BRCA-mutated cell line, organoid culture and patient-derived xenograft (PDX) models of HGSOC with acquired PARP inhibitor resistance. This work will prepare the way for a clinical trial in Aim 2 combining AZD6738 and olaparib with a schedule that maximizes ATR inhibition in patients with PARP inhibitor-resistant BRCA- mutated HGSOC. Once the maximum tolerated doses are established, we will confirm tolerability and assess preliminary antitumor activity in an expansion cohort of twelve patients. Paired biopsies will be procured for proof-of-mechanism pharmacodynamic studies in which HR is assessed with an immunohistochemical RAD51 assay and replication fork stability is assessed with a DNA fiber assay in organoid cultures. In Aim 3, we will determine the activity of ATR inhibition alone and in combination with gemcitabine in HGSOCs with a high degree of replication stress but are intrinsically unresponsive to PARP inhibition. AZD6738 and gemcitabine will be studied in cell line, organoid and PDX models harboring high-level CCNE1 or MYC amplification. We will analyze replication stress in these models and its exacerbation by ATR inhibition and gemcitabine using immunohistochemical, cytological and gene expression signature biomarkers of ATR pathway activation. Lastly, we will leverage a recently completed study of gemcitabine vs. gemcitabine combined with the ATR inhibitor M6620 in platinum-resistant HGSOC patients, in which the combination was superior among patients with a platinum-free interval of less than 3 months. We will test the hypothesis that this group was enriched with tumors carrying a high degree of replicative stress, defined by CCNE1 or MYC amplification, as well as by biomarkers developed in the preclinical models. Taken together, these aims will allow us to establish a role for ATR inhibition in the HGSOC armamentarium, identify new therapeutic avenues for HGSOC populations with poor prognosis and provide insights into the biology of PARP inhibitor resistance.