Abstract 879: Multidrug resistant tumor cells-derived microparticles induce a multifactorial resistance phenotype through regulation of oncogenic-related pathways in sensitive cancer cells.
Authors: Silva de Souza Paloma , de Souza Cruz André L. , de Biaso Viola Joao Paulo , Maia Raquel Ciuvalschi 
Published: August 2014

Background: Multidrug resistance (MDR) is associated with unsuccessful clinical cancer treatment. Different proteins are enhanced in MDR including the efflux pump P-glycoprotein (Pgp/MDR1) and inhibitor of apoptosis proteins (IAP). Newly, it has been reported that Pgp and microRNAs (miR) associated with it can be transferred through of membrane microparticles (MPs) from MDR cells to sensitive cells. Based on that, we analyzed intercellular transfer of Pgp, IAPs (c-IAP1, survivin and XIAP) and miRs related to Pgp and tumorigenesis, from MDR cells to sensitive cells through MPs. Besides that, we also analyzed NFκB and YB-1 oncogenic pathway in sensitive cells after co-culture with MDR cells-derived MPs - both transcriptional Pgp and/or IAPs regulators.

Methods: MDR cells-derived MPs were isolated by ultracentrifugation, identified by flow citometry and analyzed for proteins (Pgp/MDR1 and IAPs) by Western blot, for mRNA (Pgp/MDR1 and IAPs) and for miRs (miR-27a, -451 and -21) by qRT-PCR. Sensitive cancer cells were co-culture with MDR cells for 24 separated by Transwell inserts, with isolated MPs, and culture with conditioned medium from MDR cells; and then analyzed for proteins (Pgp/MDR1, IAPs, IBα and YB-1), immunofluorescence (Pgp, NFκB and YB-1), mRNA (Pgp, IAPs and YB-1), miRs (miR-27a, -451 and -21), and drug resistance profile by annexin-V/PI staining.

Results: It was observed that MDR cells-derived MPs carry miR-27a, -451 and -21; Pgp and IAPs mRNA and proteins except for XIAP mRNA. After co-culture, sensitive cells acquired Pgp mRNA and protein and showed clusters of Pgp suggesting the presence of MPs. It was also observed an enhancement of IAPs (mRNA and protein) except for cIAP1; and a reduction of drug-inducible apoptosis index. No changes were observed in Pgp or IAPs in cells cultured with conditioned medium, which show that co-culture inducing proteins enhancement is not related with shedding soluble molecules by MDR cells. In parallel, we observed equivalent Pgp and IAPs enhancement when sensitive cells were co-culture with MDR cells-derived MPs, which show that co-culture inducing Pgp and IAPs changes are direct related with MPs. Also, sensitive cells co-cultured with MDR cells showed NFκB translocated to the nucleus and peri or nuclear YB-1 localization suggesting theirs participation on Pgp and IAPs endogenous expression. In addition, we observed an enhancement on miR-21, - 27a and -451, which also suggest a positive endogenous regulation of Pgp and IAPs.

Conclusion: Our data show that sensitive cells acquired multifactorial resistance through intercellular transfer of resistant cells-derived MPs, and suggest that oncogenic pathways could be involved with this phenomenon. These findings contribute to our knowledge for the emergence of MDR in cancer cells and could be helpful for new treatment approaches.

Citation Format: Paloma Silva de Souza, André L. de Souza Cruz, Joao Paulo de Biaso Viola, Raquel Ciuvalschi Maia. Multidrug resistant tumor cells-derived microparticles induce a multifactorial resistance phenotype through regulation of oncogenic-related pathways in sensitive cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 879. doi:10.1158/1538-7445.AM2013-879