Acyl-CoA synthetase-4 is implicated in drug resistance in breast cancer cell lines involving the regulation of energy-dependent transporter expression

ORLANDO ULISES; SOLANO, ANGELA ROSARIA; MEDRANO, MAYRA AGUSTINA RÍOS; PODESTA ERNESTO JORGE; CASTILLO ANA FERNANADA; MALOBERTI, PAULA MARIANA

San Pablo

Congreso; Second AACR International Conference Translational Cancer Medicine Cancer Discoveries for Clinical Application; 2018

American Association for Cancer Research. Latin American Cooperative, Oncology Group

Chemotherapeutic treatment of disseminated breast cancer is usually faced with obstacles associated totherapy resistance. The most common reason for the acquisition of drug resistance is the expression ofATP-binding cassette (ABC) transporters which detect and eject anticancer drugs from cells. In addition,we have previously shown that acyl-CoA synthetase 4 (ACSL4) is implicated in hormone therapyresistance. In this context, the aim of the present work was to study the role of ACSL4 in cell resistance tocisplatin, doxorubicin and paclitaxel treatment and the involvement of ABC transporters in the underlyingmechanisms.To this end, we used MCF-7 Tet-Off/ACSL4 and MDA-MB-231 mock cells, which overexpress ACSL4, andcontrol line MCF7 Tet-Off empty vector, MDA-MB-231 shRNA ACSL4 and MDA-MB-231 wild type cells.Assays were conducted on cell viability (MTT), cell proliferation (BrdU), drug efflux (flow cytometry),ACSL4-responsive drug resistance ABC transporters (RNA-Seq), transporter mRNA expression, proteinlevels and signaling pathway participation (real-time PCR and Western blot).MTT assays rendered higher survival rates upon chemotherapeutic treatment in MCF7 Tet-Off/ACSL4 andMDA-MB-231 mock cells as compared to MCF-7 Tet-Off empty vector and MDA-MB-231 shRNA ACSL4,respectively. Doxycycline-induced ACSL4 inhibition in MCF7 Tet-Off/ACSL4 cells counteracted cell survival,rendering results comparable to MCF7 Tet-Off empty vector. BrdU incorporation experiments showed asynergic effect on the inhibition of MDA-MB-231 wild type cell proliferation upon treatment with ACSL4inhibitor triacsin C and chemotherapeutic drugs. RNA-Seq studies identified ACSL4-responsive drugresistance genes which were significantly and differentially expressed in MCF-7 Tet-Off/ACSL4 ascompared to MCF-7 Tet-Off empty vector. Real time PCR and Western blot results validated the changesobserved in the expression of transporter genes ABCG2, ABCC4, and ABCC8. MCF-7 Tet-Off/ACSL4 cellsshowed an increase in protein expression of ABC transporters, while MDA-MB-231 shRNA ACSL4 cellsshowed inhibition of both protein expression and chemotherapeutic drug efflux. Also, ABCG2 and ABCC4exhibited higher activity in cells overexpressing ACSL4. Cell survival was further analyzed in the presenceof Ko 143 and Ceefourin 1, inhibitors of ABCG2 and ABCC4, respectively, upon chemotherapeutictreatment, with results showing greater participation of ABCG2 in drug anticancer drug resistance in cellsoverexpressing ACSL4. In addition, ACSL4 inhibition and chemotherapeutic treatment combined withrapamycin-induced mTOR inhibition synergically inhibited proliferation and reduced ABCG2 expression inIn sum, ACSL4 may be regarded as a novel therapeutic target regulating the expression of transportersinvolved in anticancer drug resistance through the mTOR pathway to restore drug sensitivity in tumorswith poor prognosis for disease-free and overall survival.