Mechanisms of liver metastasis and associated resistance to immunotherapy

项目总结 黑色素瘤的发病率在过去的30年里翻了一番，预计在2005年将超过10万例 2020年的美国。2019年，美国有近7000人死于黑色素瘤。黑色素瘤有很强的倾向 通过淋巴和血液途径传播。最常见的转移部位包括淋巴。 结节、肺、肝和脑。尽管新的治疗方法，如免疫疗法，提高了患者的存活率 对于患有转移性疾病的患者，转移模式会影响对此类治疗的反应。特别是， 肝转移的患者对免疫检查点的反应和存活率较低 与转移到其他器官的患者相比，抑制剂的作用更大。肝转移的分子基础 对药物的发展及其对药物反应和耐药性的影响知之甚少。这在一定程度上是由于 类似于患者转移模式的临床前模型，以及稀疏的基因组 人类肝脏转移性病变的特征。这项提案的目标是确定黑色素瘤的驱动因素 肝转移(MLM)与免疫治疗相关耐药机制。为此，我们 描述并使用了一种新的同基因小鼠模型，该模型忠实地概括了基因组、转移和 在病人身上看到的反应模式。我们进行了大规模的活体CRISPR-Cas9筛查，并发现 促进MLM发展，但不促进原发肿瘤生长或转移到其他器官的扰动 网站。在这个提议的第一个目标中，我们阐明了促进传销的细胞内在信号机制。下一首, 利用集成的高复合成像，免疫图谱和单细胞测序，我们确定了免疫 MLM的浸润性和并发的肺转移以及免疫隔间如何影响对 临床上使用的免疫疗法。最后，我们收集了一大批肝转移癌患者的活检标本。 以及来自其他器官部位以及外周血单个核细胞(PBMC)的并发转移 基因组和免疫图谱。总之，这些研究将提供一幅基因组决定因素的图景， 传销的信号机制、免疫环境及其对免疫治疗应答的影响 并有可能确定合理的药物开发的新途径。

PROJECT SUMMARY The incidence of melanoma has doubled in the last three decades and is projected to exceed 100,000 cases in the US in 2020. Almost 7,000 people died of melanoma in the US in 2019. Melanoma has a strong propensity to spread both via lymphatic and hematogenous routes. The most common sites of metastasis include lymph nodes, lung, liver and brain. Although novel therapies, such as immunotherapies have improved survival of patients with metastatic disease, metastatic patterns can influence responses to such therapies. In particular, patients with metastases to the liver have a lower response and survival rates in response to immune checkpoint inhibitors compared to patients with metastases to other organs. The molecular underpinnings of liver metastasis development and the impact on drug response and resistance are poorly understood. This is in part due to limited pre-clinical models that resembles metastatic patterns seen in patients, and a sparsity of genomic characterization of human liver metastatic lesions. The goal of this proposal is to determine drivers of melanoma liver metastasis (MLM) and mechanisms of associated resistance to immunotherapies. For this purpose, we characterized and used a novel syngeneic mouse model that faithfully recapitulates genomic, metastatic and response patterns seen in patients. We performed a large-scale in vivo CRISPR-Cas9 screen and identified perturbations that promote development of MLM, but not primary tumor growth or metastasis to other organ sites. In the first aim of this proposal, we elucidate the cell intrinsic signaling mechanisms promoting MLM. Next, using integrated high-plex imaging, immune-profiling and single-cell sequencing, we determine the immune infiltrates of MLM and concurrent lung metastases and how the immune compartment impacts the response to clinically used immunotherapies. Finally, we assembled a large cohort of patient biopsies from liver metastases and concurrent metastases from other organ sites as well as peripheral blood mononuclear cells (PBMCs) for genomic and immune profiling. Together, these studies will provide a landscape of genomic determinants, signaling mechanisms and immune environments of MLM and their impact on responses to immunotherapies and have the potential to identify novel avenues for rationale drug development.