Differential response to retinoid treatment in mouse and human mammary tumor cell lines with alterations in protein kinase C (PKC) expression

MARIA I. DIAZ BESSONE; DAMIAN E. BERARDI; STÉFANO M. CIRIGLIANO; CAROLINA FLUMIAN; ELISA BAL DE KIER JOFFÉ; LAURA B. TODARO; ALEJANDRO J. URTREGER

Filadelfia

Congreso; 106th Annual Meeting of the American Associaton for Cancer Research; 2015

American Associaton for Cancer Research (AACR)

PKC is a serine-threonine kinase family that controls malignant transformation and metastatic dissemination. ATRA is the main active metabolite of vitamin A. Some evidences indicate that PKCd may regulate the expression of some retinoid acid (RA) dependent genes, and others indicate that retinoids could alter PKCa intracellular localization; both processes would lead to cell differentiation. In this work we have developed human (MDA-MB 231) and murine (LM3) cell models overexpressing PKCa or PKCd, in order to determine whether PKC expression alters the sensitivity to retinoids treatment (ATRA). The effect of ATRA was studied in vitro, analyzing biological responses related to tumor growth. In LM3 cells, only PKCa overexpression was able to reduce in vitro population doubling time (PDT) as compared to control (PDT: 14,8±2,3 h vs 21,2±3,1 h for LM3-PKCa y LM3-Vector respectively). Moreover, these cells also responded to retinoid treatment with a significant delay in cell proliferation (PDT: 24,3±4,2 h vs. 14,8±2,3 h for LM3-PKCa ATRA treated or not respectively. In MDA-MB 231 derived cell lines, PKC overexpression as well as ATRA treatment have no effect on proliferative potential. No differences were observed on migratory and invasive capabilities either. Interestingly, in LM3, PKCd overexpression induced an important increase in proteolytic enzymes secretion, which correlates with its major invasiveness, but this increase had no impact on in vivo metastatic dissemination. Only PKCa overexpression increased this parameter (lung nodes, median (range): 55 (20-75) vs 0 (0-10) for LM3-PKCa y LM3-Vector respectively). Contrary to LM3-PKCd in MDA-PKCd cells we could detect a decrease on proteolytic enzymes production and invasive capability. Moreover, colonies growing in Matrigel as 3D cultures showed a small and branched structures. Finally we studied whether the overexpression of a and d PKC isoforms is able to alter the activity of retinoic acid responsive elements (RARE) through a reporter gene assay. On LM3 model, the constitutive expression of PKCd highly increased RARE dependent activity. Surprisingly, in MDA-MB231 derived sublines, we could detect a significant increase on RARE activity when cells were treated with ATRA. Altogether, these results suggest that PKCa overexpression confers a more aggressive phenotype in LM3 model, but also makes these cells sensitive to ATRA effects. We could hypothesize, that the absence of response to ATRA treatment displayed by MDA-MB 231 sublines can be explained by their lack of RARb, which is implied in AP-1 transrepression in response to ATRA. Among others phenomena, AP-1 is involved in cell proliferation and proteolytic enzymes production. Regarding the differences of response to PKCd overexpression of both models, it has been reported that this PKC isoform has a differential role, pro or anti tumorigenic, depending on cellular context.