Role of TIEG1 in Foxp3+Treg development and tumor progression

描述（由申请人提供）：转化科学：癌症作为一种全身性疾病（15-CA-115）：尽管肿瘤疫苗可以诱导针对肿瘤抗原的CD4辅助细胞和CD8细胞毒性反应，但在临床上，它们在引起肿瘤消退方面基本上无效。这是因为肿瘤细胞获得了许多机制来逃避宿主的免疫监视程序。Foxp3+CD4+CD25+ treg介导的免疫抑制已成为肿瘤重要的免疫逃避机制之一，也是肿瘤免疫治疗成功的主要障碍。包括前列腺癌细胞在内的大多数恶性细胞分泌大量TGF-¿，并通过诱导Foxp3表达将效应T细胞转化为肿瘤抗原特异性Tregs。这种肿瘤诱导的Tregs不仅抑制了抗肿瘤效应细胞的启动和效应功能，而且形成了一个广泛的自扩增免疫抑制网络。因此，克服肿瘤诱导的扩增和Tregs的新生对于设计有效的免疫治疗策略以成功治疗癌症至关重要。我们已经证明TGF-诱导的早期基因1 （TIEG1）在TGF-处理的CD4T细胞中对Foxp3的转录调节中起关键作用。E3连接酶瘙痒介导的单泛素化对核易位和TIEG1的转录激活至关重要。然而，在瞬时过表达系统中，Itch靶向TIEG1进行单泛素化和多泛素化。我们的初步研究表明抑制TGF-¿诱导的Foxp3表达的IL-6可能通过多泛素化作用诱导TIEG1的蛋白酶体降解。tyk2介导的TIEG1磷酸化似乎是TIEG1多泛素化的识别信号。因此，我们假设当CD4T细胞受到TGF-¿或IL-6刺激时，Itch对TIEG1的单泛素化和多泛素化作用存在差异，并调节其激活和降解。尽管关于Foxp3在Treg发育和功能中的作用的数据越来越多，但Foxp3的转录是如何被调节的尚不清楚。我们已经确定了Foxp3启动子上一致的NFAT和TIEG1结合位点相邻。由于大多数转录因子与其他紧密结合的因子协同工作，我们假设NFAT和TIEG1在Foxp3启动子上相互作用，调节染色质重塑和Foxp3表达。清楚地了解CD4T细胞中印记Foxp3转录的分子组合和交叉对话将有助于设计策略来破坏肿瘤微环境中Tregs的抑制网络。我们将利用分泌大量TGF-¿的前列腺癌trump - c2细胞，分析TIEG1缺乏对Treg发生和肿瘤进展的影响。由于TIEG1不影响胸腺中nTreg的发育，因此靶向TIEG1是阻断treg从头诱导的一种有吸引力的策略。这种策略有望消除大多数抑制抗肿瘤免疫反应的肿瘤特异性treg，而不会引发自身免疫的风险。

DESCRIPTION (provided by applicant): Translational Science: Cancer as a systemic disease (15-CA-115): Although tumor vaccines can induce CD4 helper and CD8 cytotoxic response against tumor antigens, they have been largely ineffective in causing tumor regression in the clinic. This is because the tumor cells acquire many mechanisms to evade the immune surveillance program of the host. Foxp3+CD4+CD25+Treg-mediated immune suppression has emerged as one of the crucial tumor immune evasion mechanisms and main obstacle of successful tumor immunotherapy. Most malignant cells including prostate cancer cells secret large amounts of TGF-¿ and has been shown to convert the effector T cells into tumor antigen specific Tregs by inducing Foxp3 expression. Such tumor induced Tregs not only suppress the priming and effector function of anti-tumor effector cells but also form a broad network of self-amplifying immunosuppressive network. Therefore, overcoming tumor induced expansion and de novo generation of Tregs is critically important for the design of effective immunotherapeutic strategies for successful cancer treatment. We have demonstrated a critical role of TGF-¿ inducible early gene-1 (TIEG1) in the transcriptional regulation of Foxp3 in CD4T cells treated with TGF-¿. E3 ligase Itch-mediated monoubiquitination is essential for nuclear translocation, and transcriptional activation of TIEG1. However, in transient overexpression systems Itch targets TIEG1 for both mono and polyubiquitination. Our preliminary studies suggest that IL-6 which inhibits TGF-¿ induced Foxp3 expression induces proteasomal degradation of TIEG1 possibly through polyubiquitination. Tyk2-mediated phosphorylation of TIEG1 seems to act as a recognition signal for polyubiquitination of TIEG1. Therefore, we hypothesize that Itch targets TIEG1 differentially for mono and polyubiquitination when the CD4T cells are stimulated with TGF-¿ or IL-6 and regulates its activation and degradation. Despite the growing body of data on the role of Foxp3 in Treg development and function, how Foxp3 transcription is regulated is not clear. We have identified consensus NFAT and TIEG1 binding sites adjacent to each other on Foxp3 promoter. Since, most transcription factors work cooperatively with other factors binding in close proximity we hypothesize that NFAT and TIEG1 interact on Foxp3 promoter and regulate chromatin remodeling and Foxp3 expression. A clear understanding of molecular combinations and cross-talks that imprint Foxp3 transcription in CD4T cells will aid in designing strategies to disrupt the inhibitory network of Tregs in tumor microenvironment. Using prostate cancer TRAMP-C2 cells which secrete large amount of TGF-¿, we will analyze the effect of TIEG1 deficiency on Treg development and tumor progression. Since TIEG1 does not effect nTreg development in the thymus, targeting TIEG1 is an appealing strategy to block the de novo induction of Tregs. Such a strategy is expected to eliminate most potent tumor specific Tregs that inhibit anti-tumor immune response without the risk of triggering autoimmunity. PUBLIC HEALTH RELEVANCE: The application is in response to the Recovery Act Challenge Grant; Research Area: Translational Science Topic; (15-CA-115), Cancer as a systemic disease. This innovative application proposes to study the role of TGF-¿ inducible early gene-1(TIEG1) in the induction of Foxp3+Tregs by TGF-¿ secreted from tumor cells. The results obtained from these studies are expected gain significant new insights to maximize current immunotherapeutic approaches. In addition it complies with the spirit of the Challenge Grant as it will necessitate the hiring of new employees, especially in Detroit, Michigan which suffers from the highest unemployment rate in the country.