Targeting FOXP3 mRNA splicing for breast cancer immunotherapy

项目摘要 包括免疫检查点抑制剂和过继细胞转移在内的肿瘤免疫治疗的进展 (ACT)为癌症治疗带来了革命性的变化。然而，成功仅限于相对较小的 患者和癌症类型的子集。调节性T细胞(Treg)在维持免疫系统中发挥核心作用 动态平衡和负性调节免疫介导性炎症，如自身免疫性疾病、哮喘 还有过敏症。然而，肿瘤浸润性Tregs也抑制了有效的抗肿瘤免疫。Foxp3是大师 Treg调节器的发展和功能。人FOXP3基因通过信使核糖核酸编码两种主要亚型 选择性剪接--一个长的全长异构体(FOXP3L)和一个缺少外显子2的较短的异构体(FOXP3S)。 为了研究FOXP3S在Treg中的功能，我们建立了一个仅表达Foxp3S的Foxp3S小鼠系 异构体。我们对小鼠原位乳腺肿瘤模型的初步研究表明，Foxp3S小鼠 完全抵抗肿瘤的发展，而表达Foxp3L的小鼠表现出 显著的肿瘤生长。乳腺肿瘤内注射促Foxp3S的吗啡诺药物(MO) 野生型小鼠肿瘤显著增加肿瘤中干扰素-γ+T细胞和CD8T细胞 肿瘤生长。有趣的是，来自癌症基因组图谱的转录数据的生物信息学分析 (TCGA)研究表明FOXP3S在乳腺癌中的表达与临床呈正相关。 结果。鉴于超过60%的肿瘤浸润性CD4T细胞是Tregs，并且来自 肿瘤内的Tregs是肿瘤反应性的，并识别特定的肿瘤新抗原，我们假设 促进FOXP3S亚型表达将重新编程肿瘤反应树并转换这些免疫 抑制Tregs对肿瘤特异性T辅助细胞的作用，从而促进抗肿瘤免疫。我们建议使用Unique 小鼠品系、促进FOXP3S的吗啉和患者衍生的有机化合物来验证我们的假设。我们会 确定FOXP3S介导的抗肿瘤免疫的机制，并进一步确定FOXP3S- 促进吗啡在乳腺癌治疗中的应用。成功完成这项研究将确定一部小说 乳腺癌免疫治疗的治疗策略。

Project Abstract The development of cancer immunotherapy including immune checkpoint inhibitors and adoptive cell transfer (ACT)-based therapies revolutionized cancer treatment. The success, however, is limited to a relatively small subset of patients and cancer types. Regulatory T cells (Tregs) play a central role in maintaining immune system homeostasis and negatively regulate immune-mediated inflammation such as autoimmune diseases, asthma and allergy. However, tumor infiltrating Tregs also suppress effective anti-tumor immunity. FOXP3 is a master regulator of Treg development and function. Human FOXP3 gene encodes two major isoforms through mRNA alternative splicing – a long full-length isoform (FOXP3L) and a shorter isoform lacking exon 2 region (FOXP3S). To study the function of FOXP3S in Tregs, we generated a Foxp3S mouse line expressing only the Foxp3S isoform. Our preliminary studies with orthotopic mouse breast tumor models demonstrate that the Foxp3S mice are completely resistant to tumor development, while the littermate wildtype mice expressing Foxp3L exhibit significant tumor growth. Intratumoral injection of Foxp3S-promoting morpholino drug (MO) into the breast tumors in wildtype mice markedly increases IFN-γ+ CD4 and CD8 T cells in the tumors and thus suppresses tumor growth. Interestingly, bioinformatics analysis of transcriptomic data from The Cancer Genome Atlas (TCGA) reveals that FOXP3S mRNA expression in human breast cancer is positively correlated with clinical outcomes. Given that over 60% of tumor-infiltrating CD4 T cells are Tregs and the most dominant TCRs from intratumoral Tregs were tumor-reactive and recognize specific tumor neoantigens, we hypothesize that promoting FOXP3S isoform expression will reprogram tumor-reactive Tregs and convert these immune suppressive Tregs to tumor-specific T helpers thus promoting antitumor immunity. We propose to use unique mouse lines, FOXP3S-promoting morpholinos and patient-derived organoids to test our hypothesis. We will determine the mechanisms for FOXP3S-mediated antitumor immunity and further define the efficacy of FOXP3S- promoting morpholinos in breast cancer treatment. Successful completion of this study will identify a novel therapeutic strategy for breast cancer immunotherapy.