Reprogramming of the stromal microenvironment in melanoma progression and therapeutic escape

尽管新型靶向治疗药物的批准，如BRAF抑制剂(BRAF)，MEK抑制剂(Meki)， 和免疫检查点抑制剂，已经彻底改变了黑色素瘤的治疗，长期疗效仍然 由于耐药性的发展，令许多患者感到失望。一个主要的促成因素 肿瘤的复原力和复发率是指“塑料”微环境的存在，这种微环境由以下因素组成 异质基质细胞群嵌入致密而坚硬的细胞外基质(ECM)中。尤其是， ECM不仅作为药物渗透和分布的屏障，而且还提供结构和 适应性信号，可以在黑色素瘤细胞中诱导治疗逃逸路径。基因稳定的癌症- 相关成纤维细胞(CAF)在肿瘤间质中是一种臭名昭著的ECM重塑机器。我们 已发现黑色素瘤间质中含有核β-连环蛋白的CAF数量显著增加 在患者接受BRAFi/Meki治疗后。我们已经证实在CAF中核β-连环蛋白的增加 是由BRAFi诱导的，而不是Meki。然而，BRAFi如何刺激CAF重新编程他们的生物 通过高激活的核β-连环蛋白活性实现的功能尚不清楚。我们已经取得了令人信服的成果 数据显示，CAF中β-连环蛋白的靶向缺失会削弱它们重塑肿瘤的能力 微环境，下调黑色素瘤细胞异常的BRAF/MAPK/ERK信号转导，并抑制 黑色素瘤细胞耐药的体内外研究。RNASeq数据表明，β-连环蛋白是CAF 通过创造CAF转录组来重塑ECM。我们已经鉴定了β-连环蛋白/TCF4靶基因Periostin (POSTN)作为一种重要的基质细胞蛋白，由CAF分泌，促进BRAFi抗性。中环 假说是，解码和靶向ECM重塑的CAF有可能产生药物敏感性 使黑色素瘤细胞对治疗剂敏感并提高其反应率的微环境。在AIM 1，我们将阐明BRAFi刺激CAF介导黑色素瘤药物的促激活途径(S) 耐药表型。我们将评估高激活的核β-连环蛋白在CAF功能中的作用。 黑色素瘤。在目标2中，我们将确定β-连环蛋白-tcf4转录复合体是否是信号中枢。 这控制着CAF驱动的ECM重塑和BRAFi/Meki抵抗。我们将评估是否扰乱了 β-连环蛋白-TCF4在CAF中的相互作用将在体内增强突变的BRAF黑色素瘤细胞对BRAFI/MeKI的敏感性。在AIM 3，我们将确定CAF来源的POSTN在黑色素瘤细胞生长和对BRAFi/Meki耐药中的作用。 我们将研究POSTN在黑色素瘤细胞中激活的信号通路，以促进其 增殖和对BRAFI的抗性。预期结果将是一种深入的机械性表征 CAF、BRAFi和ECM微环境之间的复杂相互作用促进了生长和 突变的BRAF黑色素瘤细胞的耐药性。所获得的知识将开启发展的可能性。 一种可以摧毁肿瘤利基以提高靶向治疗和优化患者结果的“神奇子弹”。

Although the approval of novel targeted therapy drugs, such as BRAF inhibitors (BRAFi), MEK inhibitors (MEKi), and immune checkpoint inhibitors, has revolutionized melanoma treatment, long-term outcomes are still disappointing for many patients because of the development of drug resistance. A major contributing factor to tumor resilience and relapse is the presence of a “plastic” microenvironment, which is comprised of heterogeneous stromal cell populations embedded in a dense and stiff extracellular matrix (ECM). Particularly, the ECM not only functions as a barrier to drug penetration and distribution and also provides structural and adaptive signals, which can induce therapeutic escape pathways in melanoma cells. Genetically stable cancer- associated fibroblasts (CAFs) are known to be a notorious ECM-remodeling machine in the tumor stroma. We have discovered that the number of CAFs with nuclear β-catenin in the melanoma stroma increases significantly after the patients are treated with BRAFi/MEKi. We have established that increased nuclear β-catenin in CAFs is induced by BRAFi but not MEKi. Nevertheless, how BRAFi stimulates CAFs to reprogram their biological functions via hyperactivated nuclear β-catenin activity remains to be understood. We have obtained compelling data demonstrating that targeted depletion of β-catenin in CAFs ablates their ability to remodel the tumor microenvironment, downregulates abnormal BRAF/MAPK/ERK signaling in melanoma cells, and suppresses melanoma cell drug resistance in vitro and in vivo. RNA-Seq data show that β-catenin is essential for CAF to remodel the ECM by coining the CAF transcriptome. We have identified the β-catenin/TCF4 target gene periostin (POSTN) as an important matricellular protein secreted by CAFs to promote BRAFi resistance. The central hypothesis is that decoding and targeting the ECM-remodeling CAFs has the potential to create a drug-sensitive microenvironment that sensitizes melanoma cells to therapeutic agents and increase their response rate. In Aim 1, we will elucidate the pro-activation pathway(s) by which BRAFi stimulates CAFs to mediate melanoma drug resistance phenotypes. We will assess the role of hyperactivated nuclear β-catenin in the function of CAFs in melanoma. In Aim 2, we will determine whether the β-catenin-TCF4 transcriptional complex is the signaling hub that controls CAF-driven ECM remodeling and BRAFi/MEKi resistance. We will evaluate whether disrupting the β-catenin-TCF4 interaction in CAFs will sensitize BRAF-mutant melanoma cells to BRAFi/MEKi in vivo. In Aim 3, we will determine the roles of CAF-derived POSTN in melanoma cell growth and resistance to BRAFi/MEKi. We will investigate signaling pathways that are activated by POSTN in melanoma cells to promote their proliferation and resistance to BRAFi. The expected outcomes are to be an in-depth mechanistic characterization of the complex interactions among CAFs, BRAFi, and the ECM microenvironment that promote the growth and drug resistance in BRAF-mutant melanoma cells. The knowledge obtained will open the possibility of developing a “magic bullet” that could destroy the tumor niche to improve targeted therapy and optimize patient outcomes.