Design of miRNA sponges for MDV-1 as a therapeutic strategy against lymphomas

摘要

Lymphomas are solid-type tumors containing lymphoid cells. Some of latent herpesvirus infections established in B and/or T-lymphocytes could result in the formation of lymphomas. Marek's disease virus serotype 1 (MDV-1) is an avian herpes virus causing to lymphoproliferative tumors in birds, known as Marek’s disease (MD). MD has often been used as an ideal biological model for studying the pathogenesis of lymphoma diseases caused by viruses. Therefore, we used it as a research subject to study the effect of miRNA sponges on its tumorigenicity, and to develop the theoretical basis for a new anti-tumor small molecule. The miRNA sponges designed in this study specifically bind to and degrade the miRNAs of meq gene cluster of MDV-1, including miR-M2-3p, miR-M3-5p, miR-M5-3p, miR-M9-5p and miR-M12-3p.qPCR results showed that the knockdown efficiency was 85.03%, 74.97%, 47.06%, 75.33% and 62.55%, respectively. EDU staining and CCK-8 results showed that miRNA sponges inhibited the proliferation of MDV-1 transformed MSB-1 cells in vitro, and the proliferation rate of miRNA sponges-treated cells was about 50% of the control group. DAPI staining and Annxin V-FITC/PI double staining showed that miRNA sponges induced apoptosis in MSB-1 cells, and the apoptotic rate was increased by about 27.87% compared with the control group. The results of transwell showed that miRNA sponges could inhibit the invasion of MSB-1 cells in vitro, and the inhibitory rate was about 64.52%. The soft agar assay showed that miRNA sponges could inhibit the tumorigenic ability of MSB-1 cells in vitro, and the inhibitory rate was about 66.44%.The 60-days animal study showed that miRNA sponges could alleviate the growth inhibition of MSB-1 cells (about 14.78%) and reduce the mortality (about 16.00%). In addition, the tumor formation rate was 0 (8–12% in the control group).This study suggests that miRNA sponges can serve as an effective anti-tumor small molecule for the tumors caused by herpesvirus, with potential clinical implications.