BRMS1-mediated suppression of metastases in p53 mutant lung adenocarcinoma

在美国，转移性肺癌每年导致18万人死亡，而只有约10%的肺癌患者 腺癌(LUAD)具有靶向性的驱动基因改变。仍然有一个未得到满足的需求 创新疗法。BRMS1是一种转移抑制基因，在LUAD中降低，并与 肿瘤移行、侵袭增加，预后不良。P53基因DNA结合域(DBD)的突变 在LUAD中很常见，并与转移增加有关。我们做了一个新奇的观察 BRMS1在p53mut中作为选定的转移相关基因的转录因子而不是 P53WT LUAD。具体地说，我们发现BRMS1结合了含有活性P53反应的启动子区域 P53mut中转录调控转移相关基因Maspin和SERPINE1的元件，而不是p53WT 路德。此外，我们还证明BRMS1的表达与生物学上不同的组织亚型有关。 使用CK2抑制剂CX4945恢复LUAD和BRMS1导致显著减少细胞 P53mut与p53WT LUAD的迁移和侵袭比较。我们的首要目标是确定 使用临床相关的人类LUAD在p53突变的背景下维持BRMS1的重要性 样本、患者衍生的有机化合物(PDO)和条件GEMM模型。两个具体的目标将考验我们的 假设：目的1)确定BRMS1介导的转录的下游靶点和相关成分 在p53mut LUAD中。我们将使用芯片序列来鉴定p53mut中全基因组特异性的brms1-dna结合区。 LUAD细胞。然后我们将确定BRMS1-DNA结合在转录水平上的功能意义 在具有不同P53状态的BRMS1WT和BRMS1KO LUAD等基因细胞中执行RNA-SEQ的调节。 接下来，使用shRNA筛选和报告基因分析相结合的方法来鉴定基因特异性成分， 我们验证了BRMS1通过与独特转录相互作用来差异调节转录的假设 P53mut LUAD中的辅助因素。目的2)确定BRMS1在调节人体转移中的需要量 P53mut LUAD。我们将利用我们广泛的、经过临床注释的&gt；2000人类LUAD生物信息库 具有不同的p53突变谱、病理分期和组织亚型的标本来表征 LUAD肿瘤谱。然后，我们研究了CK2抑制剂CX4945在预防BRMS1方面的有效性 我们新开发的应用于特定组织学亚型的LUAD的降解和减少转移 LUAD患者衍生器官(PDO)转移模型。我们使用这个经过修改的PDO模型 可诱导表达系统评估maspin和SERPINE1在BRMS1介导下的重要性 转移抑制。最后，使用我们新创建的Brms1-/-/KrasG12D GEMM和气管内交付 P53mut CRISPR我们建立了转移性LUAD小鼠模型来评估BRMS1对CX4945的需求- 介导性抑制转移。影响：我们的工作将提供机械和翻译证据 支持以p53mut LUAD中BRMS1为靶点的未来临床试验，这些试验具有生物学上不同的组织学亚型。

Metastatic lung cancer kills >180,000 people in the United States annually and only ~10% of patients with lung adenocarcinoma (LUAD) have targetable driver genomic alterations. There remains an unmet need for innovative therapies. BRMS1 is a metastasis suppressor gene that is decreased in LUAD and is associated with increased cancer migration, invasion, and poor prognosis. Mutations in DNA binding domain (DBD) of p53 are common in LUAD and are associated with increased metastases. We have made a novel observation that BRMS1 functions as transcription factor for selected metastasis-related genes in p53mut but not p53WT LUAD. Specifically, we show that BRMS1 binds promoter regions containing active p53 responsive elements to transcriptionally regulate metastasis-related genes, Maspin and Serpine1, in p53mut but not p53WT LUAD. Moreover, we demonstrate that BRMS1 expression is related to biologically distinct histologic subtypes of LUAD and that restoration of BRMS1 using the CK2 inhibitor CX4945 results in significantly less cell migration and invasion in p53mut compared with p53WT LUAD. Our overarching goal is to determine the importance of maintaining BRMS1 in the context of p53 mutations using clinically relevant, human LUAD samples, patient-derived organoids (PDO), and conditional GEMM models. Two Specific Aims will test our hypotheses: Aim 1) Identify downstream targets and associated components of BRMS1-mediated transcription in p53mut LUAD. We will use ChIP-seq to identify genome-wide specific BRMS1-DNA binding regions in p53mut LUAD cells. We will then determine the functional significance of BRMS1-DNA binding on transcriptional regulation by performing RNA-seq in BRMS1WT and BRMS1KO LUAD isogenic cells with different p53 status. Next, using shRNA screens combined with reporter gene assays to characterize gene-specific components, we test the hypothesis that BRMS1 differentially regulates transcription via interaction with unique transcription co-factors in p53mut LUAD. Aim 2) Determine the requirement for BRMS1 in regulating metastases in human p53mut LUAD. We will leverage our extensive, clinically-annotated biorepository of >2000 human LUAD specimens with varying p53 mutational profiles, pathologic stages, and histologic subtypes to characterize the spectrum of LUAD tumors. We then examine the efficacy of the CK2 inhibitor, CX4945, in preventing BRMS1 degradation and decreasing metastases in specific histological subtypes of LUAD using our newly developed LUAD patient-derived organoid (PDO) model of metastases. We use this PDO model modified with an inducible expression system to assess the importance of maspin, and serpine1 on BRMS1-mediated metastasis suppression. Finally, using our newly created Brms1-/-/KrasG12D GEMM with intratracheal delivery of p53mut CRISPR we create a metastatic LUAD mouse model to assess the requirement of BRMS1 for CX4945- mediated suppression of metastases. Impact: Our work will provide mechanistic and translational evidence to support future clinical trials that target BRMS1 in p53mut LUAD with biologically distinct histologic subtypes.