Modulation of the tumor microenvironment with probiotic therapies

癌症免疫治疗的最新进展为三阴性患者提供了有希望的治疗选择 乳腺癌(TNBC)。尽管这些恶性肿瘤的治疗总体上取得了成功，但免疫治疗方法 面临一些独特的挑战：(1)非靶标副作用造成的剂量限制，(2)相加毒性 联合治疗；(3)乳腺癌免疫原性较低。为了克服这些限制， 这项提议寻求改造益生菌菌株，选择性地定植于乳腺癌和局部 释放免疫疗法。最终目标是诱导更强大和多样化的抗肿瘤T细胞免疫 并促进原发癌和转移性乳腺癌结缔组织的清除和系统性生长 乳腺癌衍生的病灶。随之而来的项目将首先专注于定义 用抗体介导的耗竭方法研究益生菌E.coliNissle 1917(ECN)的瘤内趋向性 和定向基因敲除，以确定调节肿瘤特异性的宿主免疫途径 成长。使用合成生物学方法，ECN将被设计成稳定表达和释放检查点 肿瘤内局部靶向CD47、PD-L1和CTLA-4的抑制物纳米体。促炎细胞因子 将额外表达以促进抗原递呈和增强细胞毒性T细胞反应。 这项建议的主要创新之处在于结合两种方法发展了更好的理解 益生菌对肿瘤的定植，以及工程益生菌作为免疫治疗递送载体。 具体地说，这种方法比目前的治疗策略有几个优点，包括：(1)识别 新的ECN宿主株及其肿瘤定植机制的研究 在工程疗法中，(2)肿瘤特异性免疫疗法的产生，(3)导致 新型免疫疗法和脂多糖佐剂的有效释放，以及(4)局部递送 具有全身毒性的新型免疫治疗组合。这项工作旨在克服当前的 免疫疗法的局限性，通过提供一个有针对性的载体来在当地提供免疫疗法，以刺激 抗肿瘤免疫，同时预防全身毒性，减轻免疫相关不良反应。

Recent advances in cancer immunotherapy have provided promising treatment options for patients with triple-negative breast cancer (TNBC). Despite overall success in treating these malignancies, immunotherapeutic approaches face a number of unique challenges: (1) dose limitation due to off-target side effects, (2) additive toxicity of combination therapies, (3) and relatively low immunogenicity of breast cancer. To overcome these limitations, this proposal seeks to engineer probiotic strains of bacteria that selectively colonize breast cancer and locally release immunotherapeutics. The ultimate goal is to elicit more robust and diversified antitumor T cell immunity and promote the clearance of colonized primary and metastatic breast cancer lesions and systemically growing breast cancer-derived foci. The accompanying project will first focus on deciphering mechanisms that define the intratumoral tropism of the probiotic strain E. coli Nissle 1917 (EcN) by using antibody-mediated depletion approaches and targeted genetic knockouts to pinpoint host immunological pathways that regulate tumor-specific growth. Using synthetic biology approaches, EcN will then be engineered to stably express and release checkpoint inhibitor nanobodies targeting CD47, PD-L1, and CTLA-4 locally inside of tumors. Pro-inflammatory cytokines will additionally be expressed to promote antigen presentation and enhance cytotoxic T cell responses. The primary innovations of this proposal are in the combined approach of both developing a better understanding of probiotic colonization of tumors, along with engineering probiotics as an immunotherapeutic delivery vector. Specifically, this approach has several advantages over current therapeutic strategies, including: (1) identification of novel EcN host strains and mechanistic understanding of their tumor colonization for further improvements in engineered therapies, (2) tumor-specific production of immunotherapeutics, (3) bacteria lysis that leads to effective release of novel immunotherapeutics and lipopolysaccharides (LPS) adjuvant, and (4) local delivery of novel immunotherapeutic combinations that are toxic to deliver systemically. This work seeks to overcome current limitations of immunotherapies, by providing a targeted vehicle to locally deliver immunotherapies that stimulate antitumor immunity while preventing systemic toxicity and mitigating immune-related adverse effects.