Remotely controlled listerial bactodrones for cancer immunotherapy

项目总结 通过静脉注射传播的弱毒株单核细胞增多性李斯特菌(Lm)， 在原发肿瘤和转移瘤中积累和繁殖，同时从健康组织中迅速清除。 我们打算将这些菌株用作远程控制的肿瘤特异性抗癌有效载荷载体， 无人巴氏杆菌。在这个项目中，我们将工程LM合成和分泌环状二核苷酸(c-di-NMP)作为 肿瘤微环境内强大的先天免疫系统刺激物。C-di-NMPs的现场积累 会通过刺痛的先天免疫途径诱导型干扰素的产生。这将改善 LM诱导免疫原性肿瘤细胞死亡并导致肿瘤相关抗原释放的能力 这将促进肿瘤特异性CD8 T细胞的募集。肿瘤定位的持续性c-di-NMP 生产将保持T细胞和其他抗癌免疫细胞的激活。评估以下项目的可行性和有效性 通过基因工程的LM将c-di-NMP转移到肿瘤内，我们将追求两个目标。在目标1中，我们将 工程LM在免疫和肿瘤细胞中分泌合成c-di-NMP的酶。在《目标2》中，设计的 Lm将通过一种被称为杆菌感染的过程，传递编码c-di-NMP合成酶的质粒。两者都有 这些方法有望将肿瘤微环境中的受感染细胞转变为c-diNMP的产生。 工厂，并确保持久的刺痛激活。重要的是，LM介导的c-di-NMP传递系统将是 可用良性化学诱导剂诱导，这将允许暂时控制刺痛的激活和 限制与全身暴露c-di-NMP相关的毒性。在对LM型杆状无人机进行优化之后 在体外，在乳腺癌细胞系中，我们将测试定期注射杆状病毒对转移的小鼠的疗效 乳腺癌模型。我们预计，无人驾驶飞机将成为定位肿瘤的有效工具， 用于各种抗肿瘤活动的时间控制和廉价的基因有效载荷的输送。

PROJECT SUMMARY Attenuated and avirulent strains of Listeria monocytogenes (Lm), that are delivered via intravenous injections, accumulate and propagate in primary tumors and metastases while being quickly cleared from healthy tissues. We intend to use these strains as remotely controlled, tumor-specific anticancer payload delivery vehicles, bactodrones. In this project we will engineer Lm to synthesize and secrete cyclic dinucleotides (c-di-NMPs) as potent innate immune system stimulators inside tumor microenvironments. On-site accumulation of c-di-NMPs will induce production of type I interferon via the STING innate immunity pathway. This will improve the capacity of Lm to induce immunogenic tumor cell death and lead to the release of tumor-associated antigens, which will facilitate recruitment of tumor-specific CD8 T cells. The sustained tumor-localized c-di-NMP production will keep T cells and other anticancer immune cells activated. To assess feasibility and efficacy of delivering intratumoral c-di-NMP via genetically engineered Lm, we will pursue two aims. In aim 1, we will engineer Lm to secrete enzymes for c-di-NMP synthesis in immune and tumor cells. In aim 2, the engineered Lm will deliver plasmids encoding a c-di-NMP synthases, via a process known as bactofection. Both approaches are expected to turn infected cells in the tumor microenvironment into c-di-NMP producing factories and ensure durable STING activation. Importantly, Lm-mediated c-di-NMP delivery systems will be made inducible with a benign chemical inducer, which will enable temporal control of STING activation and limit toxicity associated with systemic c-di-NMP exposure. Following optimization of the Lm bactodrones in vitro, and in breast cancer cell line, we will test efficacy of periodic bactodrone injections in a mouse metastatic breast cancer model. We anticipate that Lm bactodrones will become efficient vehicles for tumor-localized, temporally controlled and inexpensive delivery of genetic payloads for various antitumor activities.