9MO - C-Myc regulates the cancer stem-cell phenotype differentially induced by the individual silencing of AKT isoforms in pancreatic adenocarcinoma

Background

Cancer stem cells (CSCs) represent a small proportion of tumor cells characterized by their high self-renewal and proliferative potential, and they have been proposed as one of the main explanations for the transience of the effect of antineoplastic agents. However, whether they arise from pre-existing populations after selection or are the result of metabolic adaptation of differentiated cells is still under debate. By individually silencing AKT isoforms in pancreatic adenocarcinoma we have observed the acquisition of stem-cell characteristics, with sharp differences between each isoform.

Methods

We have individually silenced AKT isoforms using short hairpin RNAs (shRNAs) delivered by lentivirus. When cells adapted to the modification, high-throughput quantitative proteomics analyses were performed to evaluate the differentially altered molecular routes. We have evaluated accepted CSCs features as mitochondrial mass and metabolism, CD44 and EpCAM expression, ALDH activity, NANOG expression, spheroid growth and autophagy. We have generated constitutively active and dominant negative c-MYC pancreatic adenocarcinoma cell lines.

Results

The adaptation induced deep proteomic changes that affected cells differently depending on the AKT isoform silenced. c-MYC was the main transcription factor associated to potential regulation of the proteomes induced by the silencing of AKT isoforms. c-MYC expression was downregulated in shAKT1 expressing cell line. Mitochondrial mass was increased in the 3 isoforms, although only shAKT1 had greater mitochondrial respiration. Higher basal autophagy was observed in shAKT1 and shAKT3 compared with shAKT2 and control, as well as spheroid formation, regarding both number and size. An increase in NANOG was apparent in shAKT1. CD44 and EpCAM co-expression was increased only in shAKT1, while it was abrogated in c-MYC T58A expressing cell line.

Conclusions

The silencing of each AKT isoform induce distinct degrees of CSC dedifferentiation in pancreatic adenocarcinoma. c-MYC is a main regulator of this process.

Clinical trial identification

Editorial acknowledgement

Legal entity responsible for the study

The authors.

Funding

AECC, Instituto de Salud Carlos III.

Disclosure

All authors have declared no conflicts of interest.