METHODS: In this study, we evaluated the feasibility of intraperitoneal administration of non-virally engineered MSCs to co-deliver CDUPRT/5-Flucytosine prodrug system and IFNb to potentially enhance the cGAS-STING signalling axis. Here, MSCs were engineered to express CDUPRT or CDUPRT-IFNb. Expression of CDUPRT and IFNb was confirmed by flow cytometry and ELISA, respectively. The anti-cancer efficacy of the engineered MSCs was evaluated in both in vitro and in vivo model. ES2, HT-29 and Colo-205 were cocultured with engineered MSCs at various ratio. The cell viability with or without 5-flucytosine was measured with MTS assay. To further compare the anti-cancer efficacy of the engineered MSCs, peritoneal carcinomatosis mouse model was established by intraperitoneal injection of luciferase expressing ES2 stable cells. The tumour burden was measured through bioluminescence tracking.
RESULTS: Firstly, there was no changes in phenotypes of MSCs despite high expression of the transgene encoding CDUPRT and IFNb (CDUPRT-IFNb). Transwell migration assays and in-vivo tracking suggested the co-expression of multiple transgenes did not impact migratory capability of the MSCs. The superiority of CDUPRT-IFNb over CDUPRT expressing MSCs was demonstrated in ES2, HT-29 and Colo-205 in-vitro. Similar observations were observed in an intraperitoneal ES2 ovarian cancer xenograft model. The growth of tumor mass was inhibited by ~ 90% and 46% in the mice treated with MSCs expressing CDUPRT-IFNb or CDUPRT, respectively.
CONCLUSIONS: Taken together, these results established the effectiveness of MSCs co-expressing CDUPRT and IFNb in controlling and targeting PC growth. This study lay the foundation for the development of clinical trial using multigene-armed MSCs for PC.
方法:在本研究中,我们评估了腹膜内给予非病毒工程化MSCs共同递送CDUPRT/5-氟胞嘧啶前药系统和IFNb以潜在增强cGAS-STING信号轴的可行性.这里,将MSC工程化以表达CDUPRT或CDUPRT-IFNb。流式细胞术和ELISA证实CDUPRT和IFNb的表达,分别。在体外和体内模型中评估工程化MSC的抗癌功效。将ES2、HT-29和Colo-205与工程化MSC以各种比例共培养。用MTS测定法测量有或没有5-氟胞嘧啶的细胞活力。为了进一步比较工程MSCs的抗癌功效,通过腹腔注射表达荧光素酶的ES2稳定细胞建立腹膜癌变小鼠模型。通过生物发光跟踪测量肿瘤负荷。
结果:首先,尽管编码CDUPRT和IFNb(CDUPRT-IFNb)的转基因高表达,但MSCs的表型没有变化。Transwell迁移测定和体内跟踪表明,多种转基因的共表达不会影响MSC的迁移能力。在体外ES2、HT-29和Colo-205中证明了CDUPRT-IFNb优于表达CDUPRT的MSC。在腹膜内ES2卵巢癌异种移植模型中观察到类似的观察结果。在用表达CDUPRT-IFNb或CDUPRT的MSCs治疗的小鼠中,肿瘤块的生长被抑制约90%和46%,分别。
结论:综合来看,这些结果确立了共表达CDUPRT和IFNb的MSC在控制和靶向PC生长中的有效性。本研究为开发使用多基因武装的MSCs进行PC的临床试验奠定了基础。