Assessing TGF Beta1-Mediated ECM Remodeling in Metastatic Oral Squamous Cell Carcinoma

评估转移性口腔鳞状细胞癌中 TGF Beta1 介导的 ECM 重塑

基本信息

批准号：
10606396
负责人：
John Aleman
金额：
$ 4.31万
依托单位：
UNIVERSITY OF COLORADO DENVER
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-12-01 至 2025-11-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10606396
关键词：
Affect Atomic Force Microscopy Binding CD8-Positive T-Lymphocytes CD8B1 gene Cell Line Cells Characteristics Clustered Regularly Interspaced Short Palindromic Repeats Collagen Collagen Gene Collagen Type I Consensus Data Dependence Deposition Disease Distant Distant Metastasis Drops Esophagus Exclusion Extracellular Matrix Flow Cytometry Genes Genetic Goals Human Immune Immune checkpoint inhibitor Immune system Immunocompetent Immunomodulators In Vitro Infiltration Integrin Binding Integrins Investigational Therapies KRASG12D Knock-out Laboratories Laminin Lung Lymphocyte Lymphocyte Count Malignant Epithelial Cell Malignant Neoplasms Measures Mediating Metastatic Neoplasm to the Lung Metastatic Squamous Cell Carcinoma Modeling Mus Mutation Neoplasm Metastasis Non-Small-Cell Lung Carcinoma Nonmetastatic Outcome Patient Care Patients Primary Health Care Primary Neoplasm Production Prognostic Factor Prognostic Marker Proteins Regulation Risk Sampling Signal Transduction Squamous cell carcinoma Survival Rate TGFB1 gene Testing Transforming Growth Factor beta Transplantation Tumor-Infiltrating Lymphocytes anti-tumor immune response cancer cell cancer risk cancer stem cell cancer type cell motility checkpoint therapy crosslink high risk improved in vivo inhibitor insight lymphoid neoplasm malignant breast neoplasm malignant mouth neoplasm migration mouse model mouth squamous cell carcinoma response standard of care stem cell expansion stem cells success targeted treatment therapeutic target trafficking trait transcriptome sequencing tumor tumor microenvironment

项目摘要

Abstract Oral squamous cell carcinoma (OSCC) patients have a dismal survival rate due to distant metastases that escape primary care. Transforming growth factor beta (TGFβ) is a well-known driver of metastasis, modulator of immune cell activity, and regulator of extracellular matrix (ECM) genes. In breast cancer, a stiff ECM is generally attributed to excess type I collagen deposition and crosslinking that promotes cancer cell metastasis and cancer stem cell (CSC) expansion. In OSCC, a consensus has not been reached on whether a stiff or soft ECM increases metastatic potential and CSC expansion. The impact of OSCC ECM composition on immune cell trafficking is also unknown. Our laboratory has produced murine OSCC cell lines derived from Keratin15+ stem cells with Smad4 loss and KrasG12D mutation. Despite having the same genetic background, these cell lines have different metastatic ability, suggesting microenvironmental factors or cell intrinsic differences may mediate metastasis. RNAseq analysis revealed that metastatic OSCC cells have increased levels of laminins and laminin- binding integrins but downregulated type I collagen genes. Additionally, while OSCC cells cultured on stiff ECM demonstrate increased invasion, those cultured on a soft ECM display increased CSC characteristics. Treating metastatic OSCC cells with a TGFβ inhibitor reduced migration and invasion. Comprehensive immune profiling using flow cytometry revealed that metastatic tumors have decreased numbers of CD8+ T infiltrating lymphocytes (TILs) compared to non-metastatic OSCC tumors. These CD8+ TILs are instrumental for a robust anti-tumor immune response and the success of immune checkpoint inhibitor (ICI) therapy. Identifying responders to ICI and TGFβ inhibitors and sensitizing non-responders to these therapies persist as a major obstacle. Defining how OSCC cells interact with the ECM to promote their dissemination and enhancing current prognostic markers are both crucial to improve patient care and prolong survival. The goal of this proposal is to use unique murine models and human patient samples to define how ECM components and rigidity influence OSCC CSCs metastasis to improve the efficacy of emerging, targeted therapeutics to inhibit OSCC metastatic outgrowth. The hypothesis of this proposal is that elevated TGFβ signaling in OSCC induces increased integrin expression and a laminin/collagen imbalance in the ECM, altering ECM stiffness and modulating OSCC metastasis, OSCC CSCs, CD8+ TIL motility and ICI responsiveness. I will 1) evaluate TGFβ-dependency and function of laminins and associated integrins in OSCC metastasis; 2) determine how laminins contribute to ECM rigidity and subsequent impact on OSCC CSC characteristics and motility; and 3) assess if inhibiting laminin deposition enhances ICI. This study will provide insight into how enhanced laminin-binding integrin expression and laminin deposition facilitates OSCC metastasis, CSC characteristics, and CD8+ TIL exclusion. It will also identify predictive prognostic biomarkers for ICI and therapeutic targets for treating metastatic OSCC.

抽象的口腔鳞状细胞癌（OSCC）患者由于远处转移而生存率很低逃避初级保健。转化生长因子β（TGFβ）是众所周知的转移驱动因素和调节剂。免疫细胞活性和细胞外基质 (ECM) 基因的调节因子在乳腺癌中，ECM 通常是僵硬的。归因于过量的 I 型胶原蛋白沉积和交联，促进癌细胞转移和癌症在 OSCC 中，对于硬 ECM 还是软 ECM 尚未达成共识。增加转移潜力和 CSC 扩张 OSCC ECM 成分对免疫细胞的影响。我们的实验室已经生产了源自 Keratin15+ 干细胞的小鼠 OSCC 细胞系。具有 Smad4 缺失和 KrasG12D 突变的细胞尽管具有相同的遗传背景，但这些细胞系具有相同的遗传背景。不同的转移能力，表明微环境因素或细胞内在差异可能介导 RNAseq 分析显示，转移性 OSCC 细胞的层粘连蛋白和层粘连蛋白水平增加。此外，当 OSCC 细胞在坚硬的 ECM 上培养时，它会结合整合素，但下调 I 型胶原蛋白基因。表明侵袭增加，在软 ECM 上培养的细胞显示出增加的 CSC 特征。使用 TGFβ 抑制剂可减少转移性 OSCC 细胞的迁移和侵袭。使用流式细胞术显示转移性肿瘤的 CD8+ T 浸润数量减少淋巴细胞 (TIL) 与非转移性 OSCC 肿瘤相比，这些 CD8+ TIL 有助于保持稳健。抗肿瘤免疫反应和免疫检查点抑制剂（ICI）治疗的成功。对 ICI 和 TGFβ 抑制剂有反应的人以及对这些疗法无反应的人持续成为主要的治疗方法定义 OSCC 细胞如何与 ECM 相互作用以促进其传播并增强电流。预后标志物对于改善患者护理和延长生存至关重要。使用独特的小鼠模型和人类患者样本来定义 ECM 成分和刚性如何影响 OSCC CSC 转移，以提高抑制 OSCC 转移的新兴靶向治疗的疗效该提议的假设是 OSCC 中 TGFβ 信号传导的升高会导致细胞生长增加。 ECM 中的整合素表达和层粘连蛋白/胶原蛋白失衡，改变 ECM 硬度并调节 OSCC 转移、OSCC CSC、CD8+ TIL 运动性和 ICI 反应性我将 1) 评估 TGFβ 依赖性和层粘连蛋白和相关整合素在 OSCC 转移中的功能；2) 确定层粘连蛋白如何促进 ECM 刚性及其对 OSCC CSC 特征和运动性的影响；以及 3) 评估是否抑制层粘连蛋白沉积增强 ICI 本研究将深入了解如何增强层粘连蛋白结合整合素表达。层粘连蛋白沉积也会促进 OSCC 转移、CSC 特征和 CD8+ TIL 排除。确定 ICI 的预测预后生物标志物和治疗转移性 OSCC 的治疗靶点。