Rapamycin Induces Apoptosis When Autophagy is Inhibited in T-47D Mammary Cells and Both Processes are Regulated by Phlda1

Abstract

Autophagy is an evolutionarily conserved lysosomal degradation pathway and plays a critical role in the homeostatic process of recycling proteins and organelles. Functional relationships have been described between apoptosis and autophagy. Perturbations in the apoptotic machinery have been reported to induce autophagic cell deaths. Inhibition of autophagy in cancer cells has resulted in cell deaths that manifested hallmarks of apoptosis. However, the molecular relationships and the circumstances of which molecular pathways dictate the choice between apoptosis and autophagy are currently unknown. This study aims to identify specific gene expression of rapamycin-induced autophagy and the effects of rapamycin when the autophagy process is inhibited. In this study, we have demonstrated that rapamycin is capable of inducing autophagy in T-47D breast carcinoma cells. However, when the autophagy process was inhibited by 3-MA, the effects of rapamycin became apoptotic. The Phlda1 gene was found to be up-regulated in both autophagy and apoptosis and silencing this gene was found to reduce both activities, strongly suggests that Phlda1 mediates and positively regulates both autophagy and apoptosis pathways.

Pleckstrin homology-like domain, family A, member 1 (PHLDA1) and cancer

“…Upregulation of PHLDA1 has been identified in autophagy and apoptosis induced by rapamycin in T47D breast cancer cells (26). In addition, stress-induced heat-shock proteins (Hsp; including Hsp70, Hsp110 and Hsp40) bind directly to the pleckstrin domain of PHLDA1 and suppress its proapoptotic function in mouse embryonic fibroblast (MEF) cells (27).…”

Untangling knots between autophagic targets and candidate drugs, in cancer therapy

“…Other candidates such as RAD001, dexamethasone, imipramine, PM02734, bufalin, oroxylin A and calix[6]arene have been shown to have the ability to induce pro-death autophagy through the PI3KCI/Akt/mTORC1 pathway (46,47,51,74,84,102,104,105,129). In addition, some candidate drugs such as rapamycin and resveratrol also have a double-faced role, by inducing pro-death or pro-survival autophagy in different cancer cells (40,41,54,142). Exact mechanisms of the different effects that different candidate drugs cause by regulating the PI3KCI/Akt/mTORC1 pathway remains unclear and calls for considerable further investigation.…”

Estudo funcional do gene PHLDA1 Pleckstrin Homology-like Domain, Family A, Member 1 em células epiteliais de mama, MCF10A

“…A relação de PHLDA1 com câncer já foi descrita em diferentes tipos tumorais, incluindo câncer de mama (Nagai et al 2007;Marchiori et al 2008;Oberst et al 2008;Johnson et al 2011;Li et al 2014;Kastrati et al 2014;Moad et al 2013;Nagai 2016), melanoma (Neef et al 2002), câncer de pulmão (Joo et al 2007), câncer colorretal (Chiu 2005), câncer renal (Tavares et al 2008) e carcinoma oral (Coutinho-Camillo et al 2013). …”

“…Estes efeitos foram diminuídos após silenciamento de PHLDA1 indicando regulação do processo autofágico e apoptótico por PHLDA1 nestas células (Moad et al 2013 onde correlacionou-se a expressão de PHLDA1 com propriedades do tipo célula tronco.…”

“…O docetaxel pertence à família dos taxanos e corresponde a um quimioterápico que interfere, entre outros processos, na divisão mitótica provocando a estabilização dos microtúbulos e, consequentemente, podendo desencadear a morte celular (Morse et al 2005). A apoptose é o mecanismo de morte predominante que ocorre em resposta ao tratamento com esta droga (Wang et al 1998) e seu uso é aprovado para o tratamento de câncer de mama, câncer de cabeça e pescoço, câncer gástrico, câncer de próstata e câncer de pulmão de não pequenas células (Pazdur 2014 Conforme mencionado anteriormente, estudos pioneiros do nosso grupo mostraram PHLDA1 como um forte marcador de prognóstico no câncer de mama a partir de um estudo em uma coorte de cerca de setecentos pacientes (Nagai et al 2007) e estudos subsequentes indicaram seu envolvimento na regulação de apoptose, autofagia e efeito inibitório do crescimento em diferentes linhagens celulares: células HME16C transformadas com Ras; células do subtipo basal-like, MDA-MB-231; células Her2+, SKBR3 e ER+, MCF7 (Oberst et al 2008;Johnson et al 2011;Moad et al 2013;Li et al 2014). Todavia, PHLDA1 vem também sendo reportado como um marcador folicular de célula tronco em uma série de estudos (Ohyama et al 2006;Sellheyer & Nelson 2011;Yeh et al 2012;Sellheyer & Krahl 2011;Sellheyer et al 2012;Battistella et al 2014) e foi adicionalmente descrito possuindo funções oncogências em células MCF7 (Kastrati et al 2014).…”