脐带间充质干细胞运载呼肠孤病毒对急性髓系白血病细胞的溶瘤效应及机制探讨

Acute myeloid Leukemia, (AML)is a severe disease caused by hematopoietic stem cells/ancestral cells, characterized by the abnormal proliferation of primary and juvenile myeloid cells in bone and peripheral blood. The onset can be at any age and chemotherapy is still the main treatment option. The chemotherapy has limited effect and the recurrence rate is high. The total remission rate of chemotherapy plus allogeneic hematopoietic stem cell transplantation, (alloHSCT)) is around 60%. Therefore, how to increase the survival time of AML on a long term base becomes the main clinical problem. In recent years, the rise of oncolytic viral therapy provides a new approach of AML treatment. Oncolytic viruses refer to those non-pathogenic viral granules who may infect tumor cells and lead to their death without negative influence on normal tissues. Reovirus has been used in clinical experiments in more than 30 tumors for its safety and good resistance. Therefore, treating AML with reovirus is a good direction. Like other oncolytic cells, however, reovirus produces neutralizing antibody and lessens the effect when used in human. Researches find that cell carrier is capable of carrying viruses while blocking the function of neutralizing antibody. Therefore, this study plans to use mesenchymalstem cells(MSCs) capable of chemotaxis to carry reovirus, in vivo and in vitro observation will be made on their combined killing effect, and discussions will be held on the treatment mechanism, to provide theoretical evidence on developing novel AML treatment technique and drugs.

急性髓系白血病(AML)是髓系造血干/祖细胞恶性疾病，以骨髓与外周血中原始和幼稚髓性 细胞异常增生为特征，各个年龄段均可发病，目前以化疗为主，难治和复发率高，如何提高AML的长期生存是临床面临的主要问题。近年，溶瘤病毒(oncolytic virus)治疗技术为AML的治疗提供了一条新思路。溶瘤病毒是可感染肿瘤细胞并致其死亡，而对正常组织没有影响的非致病性病毒颗粒。呼肠孤病毒(reovirus)因其安全、耐受性好，已在30多个肿瘤中开展临床治疗试验。但呼肠孤病毒在人体使用 时会产生中和抗体而减轻疗效，遂采用细胞运载工具，即可携带病毒又屏蔽了宿主的中和抗体作用。本研究拟采用对肿瘤有趋化作用的间充干细胞(MSCs)荷载呼肠孤病毒，通过体内外实验观察对AML的协同杀伤作用并深入探讨其治疗机制，为开发新的AML治疗技术和药物提供理论依据。

近年来，溶瘤病毒（oncolytic virus）为急性髓系白血病（acute myeloid leukemia，AML）的治疗提供了新思路，为解决溶瘤呼肠孤病毒静脉系统给药会由于血循环中和抗体而减轻治疗疗效这一技术难题，本项目以脐带间充质干细胞（umbilical cord mesenchymal stem cells，UC-MSCs）运载呼肠孤病毒（reovirus，REO）对AML的治疗为中心，提出MSCs作为reovirus的运载工具既可携带病毒又可屏蔽中和抗体的作用的假设，通过体内外实验观察对AML的杀伤作用并深入探讨其治疗机制。研究发现UC-MSCs运载reovirus的最佳条件是病毒滴度为1MOI，病毒与细胞孵育的时间为72h，UC-MSCs感染reovirus后维持了细胞基本生物学特征；UC-MSCs运载reovirus通过激活ROS途径诱导了THP-1凋亡和自噬，该运载系统诱导的自噬可能导致AML细胞株THP-1的死亡，抑制THP-1凋亡可增强THP-1自噬，而抑制自噬则可减少凋亡；UC-MSCs在体内外均可保护reovirus免遭中和抗体的破坏作用传递reovirus到白血病肿瘤细胞，发挥抗肿瘤作用；适度病毒滴度的reovirus感染可维持UC-MSCs的趋化迁移能力和正常生物学功能，并促进CXCL10等细胞因子的释放，更有利于对肿瘤细胞的趋化性；UC-MSCs通过囊泡释放的方式传递病毒到肿瘤细胞，并通过直接溶解和释放细胞细胞因子等机制杀伤白血病肿瘤细胞。本研究提出，UC-MSCs是一个较好的溶瘤病毒运载细胞工具，同时能增强和更好靶向溶瘤病毒对白血病细胞的杀伤作用。