Engineered self-destructing Salmonella as a colorectal cancer cure

描述（由申请人提供）：我们构建了减毒的高侵袭性肠炎沙门氏菌血清型鼠伤寒沙门氏菌（以下简称鼠伤寒沙门氏菌），它是减毒的，但能够合成选定的蛋白质，并含有编码选定蛋白质的改进的DNA疫苗载体。在这些鼠伤寒沙门氏菌菌株中设计的程序自毁特征允许细菌在宿主组织中积累后通过细胞裂解释放细胞内容物。如果选定的蛋白质能够触发肿瘤细胞凋亡，那么经过改造的自毁沙门氏菌可以作为程序化的“生物定时炸弹”，通过在肿瘤组织中定植后释放肿瘤杀伤物质来破坏肿瘤组织，这一想法促进了这一提议。结直肠癌是美国癌症相关死亡的第二大原因（仅次于肺癌）。治疗癌症的一个主要挑战是很难将治疗带到灌注不良的实体肿瘤区域，这些区域通常对化疗和放疗最有抵抗力。运动兼性鼠伤寒沙门氏菌菌株被静止癌细胞产生的化合物特别吸引，可以克服传统的治疗屏障。众所周知，减毒鼠伤寒沙门氏菌更倾向于瞄准并渗透到肿瘤组织中，使其在肿瘤中的积累量比在其他器官中多2000倍，在其他器官中，它们可以克服扩散限制，攻击不受标准化疗和放疗影响的静止癌细胞。因此，我们希望探索使用工程自毁沙门氏菌作为结直肠癌治疗的潜力。我们的目标是设计、构建和评估一种新型廉价、可快速修饰的重组减毒鼠伤寒沙门氏菌（RAS）递送系统，该系统将作为一颗“定时炸弹”，与多种功能生物治疗剂一起作为一种最佳的结直肠癌治疗方法，以克服化疗和放疗的耐药性。具有新开发特征的（RAS）传递系统将(i)具有高侵袭性，（ii）选择性地合成细菌丝氨酸和天冬氨酸化学受体，以促进肿瘤组织的最大定植，（iii）显示受调节的鼠伤寒沙门氏菌T3SS效应物SopE2的延迟合成，刺激先天免疫反应，（iv）显示受调节的延迟裂解，以有效递送细菌毒力因子TlpA。一种改良的DNA疫苗载体，编码肿瘤特异性合成的死亡配体Fas，可触发肿瘤细胞死亡，并吸引免疫细胞攻击肿瘤细胞。这种RAS递送系统将具有使其对人类施用工程沙门氏菌完全安全的特点，并且无法在免疫个体中持续存在或如果排出则无法存活。此外，我们相信设计、构建和评估一种基于沙门氏菌的工程癌症治疗方法将是一种非常有效的方法，可以减少癌症治疗的全身毒性，并可能导致从宿主中根除肿瘤。

DESCRIPTION (provided by applicant): We have constructed the attenuated hyper-invasive Salmonella enterica serovar Typhimurium (hereinafter S. Typhimurium), that are attenuated, yet capable of synthesizing selected protein and harboring an improved DNA vaccine vector encoding a selected protein. The programmed self-destructing features designed into these S. Typhimurium strains allow release of the cell contents by cell lysis after bacteria accumulated in host tissues. This proposal was promoted by the idea that if the selected proteins are able to trigger tumor cell apoptosis, then the engineered self-destructing Salmonella could serve as a programmed "bio-time-bomb" to destroy tumor tissues by release of tumor killing materials after colonization in tumor tissues. Colorectal cancer is the second leading cause of cancer-related deaths in the United States (after lung cancer). A major challenge in treating cancer is the difficulty of bringing therapy to poorly perfused areas of solid tumors, which are often most resistant to chemo- and radiotherapy. Motile facultative S. Typhimurium strains, which are specifically attracted to compounds produced by quiescent cancer cells and could overcome the traditional therapeutic barrier. It is known that attenuated S. Typhimurium preferably target and penetrate into tumor tissue allowing to accumulate about 2,000-fold more in tumors than in other organs, where they could overcome diffusion limitations and attack quiescent cancer cells that are impervious to standard chemo- and radiotherapy. Therefore we wish to explore the potential to use the engineered self-destructing Salmonella as a colorectal cancer cure. Our objective is to design, construct and evaluate a novel inexpensive rapidly modifiable recombinant attenuated S. Typhimurium (RAS) delivery system that will be a "time bomb" with multiple functional biotherapy agents as an optimal colorectal cancer cure to overcome chemo- and radiotherapeutic resistance. The (RAS) delivery system with newly developed features will (i) be hyper- invasive, (ii) selectively synthesize bacterial serine and aspartate chemoreceptors to facilitate maximal colonization of tumor tissues, (iii) displays regulated delayed synthesis of S. Typhimurium T3SS effector SopE2 that stimulates innate immune responses, and (iv) exhibits regulated delayed lysis to efficiently deliver the bacterial virulence factor TlpA, that directed by Salmonella promoter preferentially activated inside tumors, to induce apoptosis in tumor cells, and an improved DNA vaccine vector encoding a tumor-specifically synthesized death ligand Fas to trigger tumor cell death and also to attract immune cells to attack tumor cells. This RAS delivery system will have features to render it completely safe for humans administered the engineered Salmonella and to be unable to persist in immunized individuals or survive if excreted. Furthermore, we believe that the designing, constructing and evaluating an engineered Salmonella-based cancer cure would represent a highly effective means to reduce systemic toxicity of cancer treatment and perhaps lead to eradication of tumors from the host. ) PUBLIC HEALTH RELEVANCE: The engineered self-destructing Salmonella could serve as a programmed "bio-time-bomb" to destroy tumor tissues by release of tumor killing materials after colonization in tumor tissues. The designing, constructing and evaluating an engineered Salmonella-based colorectal cancer cure would represent a highly effective means to overcome chemo- and radiotherapeutic resistance, to reduce systemic toxicity of cancer treatment and perhaps lead to eradication of tumors from the host.