Elucidating mechanisms of therapy response in BRCA2 mutant prostate cancers

摘要 前列腺癌是男性中最常见的非皮肤恶性肿瘤，预计在2010年将导致34，500人死亡。 2022年，仅在美国。晚期致死性去势抵抗性前列腺癌的基因测序研究 （CRPC）已经确定了致病性BRCA 2突变的高发生率（约13%）。这些发现使 临床试验和随后的食品和药物管理局（FDA）批准的聚（ADP-核糖）聚合酶 （PARP）抑制剂（PARPis）奥拉帕尼和rucaparib治疗携带致病性BRCA 2的晚期CRPC患者 突变。尽管最初有反应，但对PARPis的治疗耐药性很常见。然而，分子 BRCA 2突变CRPC中发生的对PARPi应答的适应性尚不清楚，这是由于缺乏 生物学和临床相关模型。我们提出的研究利用了两种新的患者衍生模型 致病性BRCA 2突变CRPC系统将阐明PARPi中涉及的生物学机制 治疗反应，并帮助解决关键的临床未满足的需求，以防止或克服对PARPis的耐药性。 在本提案中，我们将使用CRPC中致病性BRCA 2突变的两种新模型，包括40511 细胞系和匹配的PARPi敏感性和抗性LTL-610 PDX。基因集富集分析（GSEA）和 利用这些新模型的RNA测序数据的过度代表性分析（ORA）指出， 参与细胞外基质（ECM）调节的基因上调， PARPi疗法特别地，ECM相关基因SERPINE 1编码蛋白质纤溶酶原 激活物抑制剂1（派-1）是奥拉帕尼治疗72小时后最显著相关的基因， GSEA前沿分析。由于派-1规范地防止ECM降解，我们然后使用Masson的 三色染色以评估PARPi抗性LTL-610 PDX，发现I型PDX显著增加， 与PARPi敏感亲本系相比的胶原蛋白沉积。因为基质改变会影响 癌细胞存活，我们假设BRCA 2中PARPis诱导的ECM基因如SERPINE 1 突变的CRPC导致增强的肿瘤基质，并使治疗抗性成为可能。两个具体目标是 在本研究中，我们提出了研究这一假设：在目标1中，我们将阐明SERPINE 1信号转导在 体外、离体和体内BRCA 2突变CRPC中的ECM沉积。在目标2中，我们将研究机制 BRCA 2突变CRPC中SERPINE 1的转录激活响应PARPi。结果从这些 研究将使系统的方法来调节ECM的变化，以响应BRCA 2突变体中的PARPi CRPC。

ABSTRACT Prostate cancer is the most common non-skin malignancy in men and is projected to cause 34,500 deaths in 2022 in the United States alone. Sequencing studies of advanced lethal castrate resistant prostate cancer (CRPC) have identified a high incidence (~13%) of pathogenic BRCA2 mutations. These findings have enabled clinical trials and subsequent Food and Drug Administration (FDA) approval of the poly (ADP-ribose) polymerase (PARP) inhibitors (PARPis) olaparib and rucaparib in advanced CRPC patients harboring a pathogenic BRCA2 mutations. Despite initial responses, therapy resistance to PARPis is common. However, the molecular adaptations that occur in BRCA2 mutant CRPC in response to PARPi are poorly understood, due to a lack of biologically and clinically relevant models. Our proposed studies leveraging two new patient-derived model systems of pathogenic BRCA2 mutant CRPC will elucidate the biological mechanisms implicated in PARPi therapy response and help address a critical clinical unmet need to prevent or overcome resistance to PARPis. In this proposal, we will use two new models of pathogenic BRCA2 mutations in CRPC, including the 40511 cell line and matched PARPi-sensitive and resistant LTL-610 PDXs. Gene Set Enrichment Analysis (GSEA) and Over-Representation Analysis (ORA) of RNA-sequencing data utilizing these novel models point to significant upregulation in genes involved in Extracellular Matrix (ECM) modulation in response to both short and long term PARPi therapy. In particular, the ECM associated gene SERPINE1, which encodes for the protein Plasminogen Activator Inhibitor 1, (PAI-1) is the most significantly implicated gene after 72 hours of olaparib treatment via GSEA leading edge analysis. Since PAI-1 canonically prevents ECM degradation, we then used Masson’s Trichrome staining to evaluate the PARPi resistant LTL-610 PDX and found dramatically increased Type I Collagen deposition compared to its PARPi sensitive parental line. Since stromal alterations are known to affect cancer cell survival, we hypothesize that the induction of ECM genes like SERPINE1 by PARPis in BRCA2 mutant CRPC results in enhanced tumor stroma, and enables therapy resistance. Two specific aims are proposed in this grant to study this hypothesis: in Aim 1, we will elucidate the role of SERPINE1 signaling in ECM deposition in BRCA2 mutant CRPC in vitro, ex vivo, and in vivo. In Aim 2, we will investigate the mechanism of transcriptional activation of SERPINE1 in BRCA2 mutant CRPC in response to PARPi. The results from these studies will enable systematic approaches to modulate ECM alterations in response to PARPi in BRCA2 mutant CRPCs.