PKCδ Inhibition Impairs Mammary Cancer Proliferative Capacity But Selects Cancer Stem Cells, Involving Autophagy

Protein kinase C (PKC) is a family of serine/threonine kinases that regulate diverse cellular functions including cell death, proliferation, and survival. Recent studies have reported that PKC delta, are involved in apoptosis or autophagy induction. In the present study we focused on how PKC delta regulates proliferation and cancer stem cell (CSC) properties of the hormone-independent mammary cancer cell line LM38-LP, using pharmacological and genetic approaches. We found that pharmacological inhibition of PKC delta, by Rottlerin treatment, impairs in vitro LM38-LP proliferation through cell cycle arrest, inducing the formation of cytoplasmic-vacuoles. Using immunofluorescence we confirmed that Rottlerin treatment induced the apparition of LC3 dots in cell cytoplasm, and increased autophagy flux. On the other side, the same treatment increased CSC growth rate and self-renewal. Furthermore, Rottlerin pre-treatment induced in CSC the development of a grape-like morphology when they are growing in 3D cultures (Matrigel), usually associated with a malignant phenotype, as well as an increase in the number of experimental lung metastasis when these cells were inoculated in vivo. The PKC delta knockdown, by RNA interference, induced autophagy and increased CSC number, indicating that these effects are indeed exerted through a PKC delta dependent pathway. Finally, the increase in the number of mammospheres could be reversed by a 3MA treatment, suggesting that autophagy mechanism is necessary for the increased of CSC self-renewal induced by PKC delta inhibition. Here we demonstrated that PKC delta activity exerts a dual role through the autophagy mechanism, decreasing proliferative capacity of mammary tumor cells but also regulating tumor stem cell self-renewal. J. Cell. Biochem. 117: 730-740, 2016. (C) 2015 Wiley Periodicals, Inc.