Developing a MicroRNA-Based Strategy for Targeting Uterine Leiomyosarcoma

MicroRNAs (miRNAs) are a novel class of small, non-coding RNA transcripts that broadly regulate patterns of gene expression. Recently, we used Next Generation Sequencing to dentify 49 individual miRNAs differentially expressed in uterine leiomyosarcoma (ULMS). We also identified potential targets for these miRNAs by using bioinformatic algorithms to screen gene products differentially expressed when genome-wide transcriptional profiling was used to examine our specimens. Based on these results, we hypothesize that miR-143 plays a central role in the pathogenesis of ULMS. To test this hypothesis, we intend to examine the function of miR-143 in uterine smooth muscle and smooth muscle tumors both n vitro and in vivo. Mimics and inhibitors for miR-143 will be transfected into an established LMS cell line (SK-LMS), primary cell strains derived from metastastic ULMS (285, 505 and 987) and telomerase-immortalized lines derived from human myometrium (HM) and eiomyoma (DD). Standard MTT and caspase assays will be used to compare rates of Droliferation and apoptosis. Flow cytometry will be used to measure the impact of altered miR-143 levels on the cell cycle. Scratch assays will be used to measure cell migration. Once biologic functions for miR-143 have been established, target genes will be validated using quantitative PCR and Western blotting. We also intend to test whether miR-143 sensitizes SK-LMS and primary ULMS cell strains to gemcitabine and docetexel. Lastly, we plan to examine whether miR-143 regulates the growth and metastasis of ULMS in vivo, using clones of SK-LMS stably transfected with miR-143 mimic, miR-143 inhibitor or scrambled control to create xenografts in nude mice.

We believe that the insight generated by the proposed work will establish a functional role for miR-143 in ULMS and set the stage for the clinical application of this insight. Future work would focus on how best to deliver miR-143 for therapeutic purposes.