Bladder cancer PD-L1 control of homologous recombination: Basic mechanisms applied to novel treatments

膀胱癌 PD-L1 对同源重组的控制：应用于新疗法的基本机制

• 批准号: 10688261
• 负责人: Tyler J. Curiel
• 金额: $ 62.07万
• 依托单位: DARTMOUTH-HITCHCOCK CLINIC
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2027-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10688261
• 关键词: Adoptive Cell Transfers; BARD1 gene; BRCA1 gene; Basic Science; Biochemical; Biological Assay; Biology; Breast; CRISPR library; Cancer Biology; Cells; Clinical; Clinical Trials; Collaborations; Complex; Computer software; Critiques; Cytotoxic agent; DNA; DNA Damage; DNA Repair; DNA Repair Pathway; Data; Engineering; Excision; FDA approved; Follow-Up Studies; Funding; Genetic; Histology; Human; Image Analysis; Immune; Immunity; Immunotherapy; In Vitro; In complete remission; Malignant Neoplasms; Malignant neoplasm of urinary bladder; Mediating; Modeling; Mus; Organoids; Ovary; Pathogenesis; Pathologic; Pathway interactions; Pharmaceutical Preparations; Phenocopy; Phosphoproteins; Poly(ADP-ribose) Polymerase Inhibitor; Pre-Clinical Model; Prediction of Response to Therapy; Productivity; Proteins; Proteomics; Recording of previous events; Recurrence; Reporter; Research; Role; Signal Transduction; Stimulator of Interferon Genes; Testing; Translating; Transplantation; Treatment Efficacy; Treatment outcome; Tumor Biology; Tumor Immunity; Western Blotting; Work; Writing; Xenograft procedure; cancer cell; cancer immunotherapy; cancer therapy; clinical translation; clinically actionable; clinically relevant; confocal imaging; digital imaging; domain mapping; drug repurposing; experience; genetic manipulation; homologous recombination; human model; humanized mouse; immune checkpoint blockade; immunogenicity; improved; in vitro Assay; in vitro testing; in vivo; in vivo evaluation; insight; loss of function; melanoma; mouse model; mutant; novel; p53-binding protein 1; pharmacologic; programmed cell death ligand 1; response; response biomarker; small molecule; success; transcriptome sequencing; translational potential; treatment response; treatment strategy; tumor

This revised MPI proposal assembles experts in bladder cancer, tumor biology, DNA damage response (DDR) and pre-clinical models with a long-standing history of productive collaborations to study effects of intracellular bladder cancer PD-L1 signals on BRCA1-mediated DDR, and how such signals alter sensitivity to PARP inhibitors and immune checkpoint blockade immunotherapy in orthotopic/metastatic mouse and human models. We test transplantable mouse bladder cancer lines MB49 and MBT-2, and the human bladder cancer lines RT4, UM-UC3 and UM-UC14, representing basal and luminal histologies plus new PDX, xenograft and organoid models and our novel GEMM all per reviewer request . Intracellular PD-L1 signals are interrogated using control versus genetically manipulated tumors and other approaches including new non-biochemical approaches per reviewer request . We will use these clinically relevant models together to test our overarching hypothesis that bladder cancer PD-L1 signals promote the homologous recombination DDR pathway that suppresses PARP inhibitor and immune checkpoint blockade immunotherapy efficacy. Aim 1: Define mechanisms for PD-L1-mediated control of HR. We will use well-validated genetic, biochemical and pharmacologic approaches plus definitive cell reporter assays for homologous recombination, RNA-seq, proteomics, RPPA, immunoblots, CRISPR library screens, organoids and digital imaging with image analysis software to understand how PD-L1 regulates BRCA1 functionality and to test mechanisms, including for tumor immunogenicity. We identified FDA-approved pharmacologic agents that we repurposed to deplete tumor PD-L1 and inhibit HR, for which we define mechanisms. Aim 2: Define treatment consequences of PD-L1 controlled homologous recombination/BRCA1 effects. Control versus genetically PD-L1 depleted cells with genetic perturbations of BRCA1 and related molecules will be tested for in vivo effects by challenging engineered cells into mice and treating with single agents or combinations, based on findings from Aim 1 to assess treatment efficacy and specific mechanisms. We use transplantable, syngeneic mouse models, and human xenograft and humanized mouse models in which we have extensive experience. Notably, we will assess tumor microenvironmental and immune contributions to treatments using genetically immune altered mice, adoptive cell transfers and immune blocking/neutralizing molecules. In vivo data are mechanistically refined in in vitro assays in which we are expert. PDX, organoids

这 修改 MPI建议组装专家膀胱癌，肿瘤生物学，DNA损伤反应（DDR） 和临床前模型，具有长期存在的生产性合作史，以研究细胞内的影响 BRCA1介导的DDR上的膀胱癌PD-L1信号，以及此类信号如何改变对PARP的敏感性 原位/转移小鼠和人类模型中的抑制剂和免疫检查点阻断免疫疗法。 我们测试可移植的小鼠膀胱癌系MB49和MBT-2，以及人膀胱癌系RT4， UM-UC3和UM-UC14，代表基础和腔液组织学以及新的PDX，异种移植物和类器官 模型和我们的小说Gemm都是每个审稿人的要求 。使用控制对细胞内PD-L1信号进行询问 与遗传操纵的肿瘤和其他方法 包括新的非生物化学方法 审阅者请求 。我们将共同使用这些临床相关模型来测试我们的总体假设 膀胱癌PD-L1信号促进了抑制的同源重组DDR途径 PARP抑制剂和免疫检查点封锁免疫疗法功效。 AIM 1：定义PD-L1介导的HR控制的机制。我们将使用验证良好的遗传， 生化和药理学方法以及确定的细胞记者分析，用于同源重组， RNA-seq，蛋白质组学，RPPA，免疫印迹， CRISPR图书馆屏幕，器官 和图像的数字成像 分析软件以了解PD-L1如何调节BRCA1功能和测试机制，包括 用于肿瘤免疫原性。我们确定了由FDA批准的药理学剂，我们重新使用以耗尽 肿瘤PD-L1和抑制HR，为此定义机制。 AIM 2：定义PD-L1控制同源重组/BRCA1效应的治疗后果。 对照与遗传性PD-L1耗尽的细胞具有BRCA1和相关分子的遗传扰动 通过将工程细胞挑战小鼠并用单一药物或单一剂或 基于AIM 1的发现，以评估治疗功效和特定机制的组合。我们使用 可移植，合成小鼠模型和人异种移植物和人源化小鼠模型 我们有丰富的经验。值得注意的是，我们将评估肿瘤微环境和免疫 使用遗传免疫改变的小鼠，产卵细胞转移和免疫对治疗的贡献 阻断/中和分子。体内数据在我们是专家的体外测定中是机械化的。 PDX，类器官