Pharmacological Inhibition of PKCα and Retinoic Acid synergize to inhibit tumour progression and impairs selfrenewal of cancer stem cells through RARγ2 modulation in a murine triple negative mammary cancer model.

BERARDI; DÍAZ BESSONE; URTREGER; TODARO

Congreso; Brazilian Society for Biochemistry and Molecular Biology; 2016

INTRODUCTIONRetinoids exert different effects on malignant phenotype reversion through the crosstalk with PKC pathway.OBJECTIVESWe proposed to: A) Evaluate the combined effect of AllTransRetinoic Acid (ATRA) and PKCα inhibitor (Gö6976) on: 1) cellproliferation, migration and cancer stem cells (CSC) selfrenewaland differentiation 2) Retinoic Acid Receptors (RARs) levelsB) Study the effect of ATRA/PKCα inhibitor treatment on tumour progression in vivo.MATERIALS AND METHODSWe employed a murine mammary triplenegativecell model (LM38LP),composed by luminal (LEP), myoepithelial (MEP) andCSC. We perform RTPCRto measure RARs. Drug interaction was analyzed by ChouTalalay?smethod. Migration potentialwas studied by woundhealingassay. We employed mammoesphere assays to evaluated CSC. 3D culture in Matrigel toevaluate differentiation. LM38LPcells were inoculated orthotopically to get tumors in BALB/c mice?s, for in vivo studies.DISCUSSION AND RESULTSWe found an inhibitory effect on cell proliferation exerted by ATRA/PKCα; inhibitor that was synergistic by ChouTalalay´smethod. Furthermore, ATRA/PKCα; inhibitor treatment reduced LM38LPmigration more efficiently than each treatment alone,showing a synergic effect. In a 3D matrigel culture assay, ATRA pretreatedCSC formed polarized colonies with presence oflumen. PKCα; inhibition impairs lumen formation induced by ATRA but led to smaller colonies. ATRA/PKCα; inhibitortreatment synergized to reduce mammospheres growth. Surprisingly, ATRA treatment induced CSC selfrenewalbut theblockage of PKCα; activity impairs this effect. Interestingly, we found that ATRA treatment increased RARβ2 and RARγ2 inCSC. In addition, we observed that only the combined treatment induced a decrease of RARγ2 in the migratory MEP cells andimpairs RARγ2 induction by ATRA in CSC. Finally, we observed that the combined treatment reduced LM38LPtumor growthand metastatic spread in a synergic manner.CONCLUSIONOur findings suggest that ATRA and