Characterizing oncogenic function of ITCH in melanoma

表征 ITCH 在黑色素瘤中的致癌功能

• 批准号: 10454357
• 负责人: Lixin Wan
• 金额: $ 36.92万
• 依托单位: H. LEE MOFFITT CANCER CTR & RES INST
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-01 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10454357
• 关键词: Adopted; Autoimmune; BRAF gene; Binding; Biochemical; Bioinformatics; Biology; Cell Culture Techniques; Cell Proliferation; Cell physiology; Cells; Communication; Data; Development; Ectopic Expression; Exhibits; Fostering; Future; Genetically Engineered Mouse; Goals; Human; Hyperactivity; Immune; Immune Evasion; Immune checkpoint inhibitor; Immune response; Immune system; Immunology; Immunosuppression; Immunotherapy; In Vitro; Investigation; JNK-activating protein kinase; Knockout Mice; Knowledge; Leukocytes; Link; MAP Kinase Gene; MEK inhibition; MEKs; Malignant Neoplasms; Melanoma Cell; Modeling; Molecular Conformation; Mus; N-terminal; Nature; Oncogenic; Pathologic; Pathway interactions; Patients; Phenotype; Phosphoric Monoester Hydrolases; Phosphorylation; Physiological; Play; Polyubiquitin; Process; Protein Phosphatase 2A Regulatory Subunit PR53; Proteins; Proteomics; Pruritus; Publications; Reader; Reporting; Research; Resistance; Role; Signal Transduction; Testing; Therapeutic; Transforming Growth Factor beta; Tumor-infiltrating immune cells; Ubiquitination; Xenograft procedure; Yin; base; cell growth; cytokine; fitness; in vivo; inhibitor; insight; knock-down; melanocyte; melanoma; melanomagenesis; migration; mouse model; mutant; neoplastic cell; novel strategies; tumor; tumor-immune system interactions; tumorigenesis; tumorigenic; ubiquitin-protein ligase; upstream kinase

Abstract The ITCH ubiquitin E3 ligase has been well characterized as a key molecule in immune cells. The roles of ITCH in tumorigenesis are rather controversial due to its versatile functions in both tumor and immune cells. Therefore, an in-depth characterization of ITCH in tumorigenesis considering both tumor and immune cells is warranted to fill the gap in our current knowledge of ITCH’s function in cancer. We recently reported an oncogenic function of ITCH through promoting BRAF activation in BRAF wild-type melanoma cells. Our newly obtained preliminary results unveil a multifaceted role of ITCH in melanoma cells by modulating CRAF and other oncogenic signals. Our results also suggest an immunosuppressive function of ITCH in the melanoma tumor immune microenvironment. Based on these data, we hypothesize that in BRAF wild-type melanoma cells, ITCH is a therapeutic vulnerability in both melanoma cells and the tumor immune microenvironment. In melanoma cells, ITCH sustains oncogenic pathways including RAF-MEK-ERK and others to maintain their fitness; activation of ITCH in immune cells, on the other hand, fosters an immunosuppressive tumor microenvironment. Hence targeting ITCH could be a novel strategy to alter the cancer-immune set point. Three aims are proposed to test our hypothesis. In Aim 1, we will define how K27-linked ubiquitination of RAF proteins by ITCH modulates MEK-ERK signaling. We will characterize if ITCH facilitates the atypical ubiquitination and subsequent activation of CRAF in a similar way to BRAF, especially in the cellular contexts where CRAF plays a central role in activating MEK-ERK signals. Moreover, we will determine if the B55-PP2A phosphatase functions as the reader protein for the poly-ubiquitin chains assembled on RAF proteins to remove the inhibitory N-terminal phosphorylations. In Aim 2, we will characterize RAF-independent oncogenic functions of ITCH. We will examine if ITCH facilitates the activation of other oncogenic pathways that fulfill its pro-survival and pro-metastatic roles. In Aim 3, we will focus on investigating both melanoma cell-autonomous and non-autonomous functions of ITCH in vivo. We will dissect these roles by using several mouse melanoma models. To define if ITCH creates an immunosuppressive microenvironment, mouse syngeneic melanoma models will be generated using Itch+/+ and Itch-/- mice, and immuno-profiling will be carried out to search for the underlying mechanism(s). Further, Itch+/+ and Itch-/- mice will be crossed with well-characterized murine melanoma models to assess its roles in both the tumor cell and immune microenvironment.

摘要 ITCH泛素E3连接酶已被充分表征为免疫细胞中的关键分子。的作用 由于ITCH在肿瘤和免疫细胞中的多功能性，其在肿瘤发生中的作用是相当有争议的。 因此，深入表征ITCH在肿瘤发生中的作用，同时考虑肿瘤和免疫细胞， 填补了我们目前对ITCH在癌症中作用的认识的差距。我们最近报道了一个 ITCH通过促进BRAF野生型黑色素瘤细胞中BRAF活化的致癌功能。我们新 获得的初步结果揭示了ITCH在黑色素瘤细胞中的多方面作用， 其他致癌信号。我们的研究结果还表明ITCH在黑色素瘤中具有免疫抑制功能。 肿瘤免疫微环境基于这些数据，我们假设在BRAF野生型黑素瘤中， 在黑色素瘤细胞中，ITCH是黑色素瘤细胞和肿瘤免疫微环境中的治疗脆弱性。在 在黑色素瘤细胞中，ITCH维持致癌途径，包括RAF-MEK-ERK和其他途径，以维持它们的功能。 另一方面，免疫细胞中ITCH的激活会促进免疫抑制性肿瘤的形成。 微环境因此，靶向ITCH可能是一种改变癌症免疫设定点的新策略。三 目的是为了检验我们的假设。在目标1中，我们将定义K27连接的RAF泛素化是如何发生的。 ITCH蛋白调节MEK-ERK信号传导。我们将描述ITCH是否促进非典型 CRAF的泛素化和随后的激活以类似于BRAF的方式，特别是在细胞环境中 其中CRAF在激活MEK-ERK信号中起核心作用。此外，我们将确定B55-PP 2A是否 磷酸酶作为RAF蛋白上组装的多聚泛素链的阅读蛋白， 去除抑制性N-末端磷酸化。在目标2中，我们将描述RAF非依赖性致癌基因 ITCH的功能。我们将研究ITCH是否促进了其他致癌通路的激活，这些通路实现了ITCH的功能。 促进存活和促进转移的作用。在目标3中，我们将专注于研究黑色素瘤细胞自主 和ITCH在体内的非自主功能。我们将通过使用几种小鼠黑色素瘤来剖析这些角色 模型为了确定ITCH是否产生免疫抑制微环境，小鼠同基因黑色素瘤 将使用瘙痒+/+和瘙痒-/-小鼠产生模型，并进行免疫分析以寻找 （一）基本机制。此外，将瘙痒+/+和瘙痒-/-小鼠与充分表征的鼠杂交。 黑色素瘤模型，以评估其在肿瘤细胞和免疫微环境中的作用。