HTERT Downregulation Attenuates Resistance to DOX, Impairs FAK-Mediated Adhesion, and Leads to Autophagy Induction in Breast Cancer Cells.

Aleksandra Romaniuk-Drapała,Wojciech Barczak,Przemysław Kopczyński,Ewa Totoń,Dagmar Kowal,Mariusz Kaczmarek,Marta Machnik,Natalia Konieczna,Błażej Rubiś

doi:10.3390/cells10040867

Aleksandra Romaniuk-Drapała, Wojciech Barczak + Show 7 more

Open Access

https://doi.org/10.3390/cells10040867

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Abstract

Telomerase is known to contribute to telomere maintenance and to provide cancer cell immortality. However, numerous reports are showing that the function of the enzyme goes far beyond chromosome ends. The study aimed to explore how telomerase downregulation in MCF7 and MDA-MB-231 breast cancer cells affects their ability to survive. Consequently, sensitivity to drug resistance, proliferation, and adhesion were assessed. The lentiviral-mediated human telomerase reverse transcriptase (hTERT) downregulation efficiency was performed at gene expression and protein level using qPCR and Western blot, respectively. Telomerase activity was evaluated using the Telomeric Repeat Amplification Protocol (TRAP) assay. The study revealed that hTERT downregulation led to an increased sensitivity of breast cancer cells to doxorubicin which was demonstrated in MTT and clonogenic assays. During a long-term doubling time assessment, a decreased population doubling level was observed. Interestingly, it did not dramatically affect cell cycle distribution. hTERT downregulation was accompanied by an alteration in β1-integrin- and by focal adhesion kinase (FAK)-driven pathways together with the reduction of target proteins phosphorylation, i.e., paxillin and c-Src. Additionally, autophagy activation was observed in MDA-MB-231 cells manifested by alternations in Atg5, Beclin 1, LC3II/I ratio, and p62. These results provide new evidence supporting the possible therapeutic potential of telomerase downregulation leading to induction of autophagy and cancer cells elimination.

Highlights

One of the most important directions in the development of targeted cancer therapy is to obtain specificity of action with limited side effects
Our result showed no activation of the p53 pathway, but a significant accumulation of p21 was observed in both cell lines after human telomerase reverse transcriptase (hTERT) downregulation
Our study revealed that hTERT downregulation correlated with a significant decrease in breast cancer cell migration and adhesion

Summary

Introduction

One of the most important directions in the development of targeted cancer therapy is to obtain specificity of action with limited side effects. Among cancer cells’ critical features, we can list the migratory and invasion potential, resistance to therapy, immortality, or ability to escape from cell death. The enzyme’s main function is the maintenance of telomere length that prevents DNA degradation, chromosomal fusion (potentially leading to polyploidization and cell death), and DNA repair activities [2]. Telomere shortening induces replicative senescence or may eventually lead to cell death [4,5,6,7]. Some noncanonical functions are associated with modifications of drug resistance, proliferation, and adhesion abilities of cancer cells that may affect their survival [9,10,11,12]

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