Tumour suppressor EP300, a modulator of paclitaxel resistance and stemness, is downregulated in metaplastic breast cancerReportar como inadecuado




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Breast Cancer Research and Treatment

, Volume 163, Issue 3, pp 461–474

First Online: 24 March 2017Received: 06 February 2017Accepted: 13 March 2017DOI: 10.1007-s10549-017-4202-z

Cite this article as: Asaduzzaman, M., Constantinou, S., Min, H. et al. Breast Cancer Res Treat 2017 163: 461. doi:10.1007-s10549-017-4202-z

Abstract

PurposeWe have previously described a novel pathway controlling drug resistance, epithelial-to-mesenchymal transition EMT and stemness in breast cancer cells. Upstream in the pathway, three miRs miR-106b, miR-93 and miR-25 target EP300, a transcriptional activator of E-cadherin. Upregulation of these miRs leads to the downregulation of EP300 and E-cadherin with initiation of an EMT. However, miRs regulate the expression of many genes, and the contribution to EMT by miR targets other than EP300 cannot be ruled out.

MethodsWe used lentiviruses expressing EP300-targeting shRNA to downregulate its expression in MCF-7 cells as well as an EP300-knocked-out colon carcinoma cell line. An EP300-expression plasmid was used to upregulate its expression in basal-like CAL51 and MDA-MB-231 breast cancer cells. Drug resistance was determined by short-term proliferation and long-term colony formation assays. Stemness was determined by tumour sphere formation in both soft agar and liquid cultures as well as by the expression of CD44-CD24-ALDH markers. Gene expression microarray analysis was performed in MCF-7 cells lacking EP300. EP300 expression was analysed by immunohistochemistry in 17 samples of metaplastic breast cancer.

ResultsCells lacking EP300 became more resistant to paclitaxel whereas EP300 overexpression increased their sensitivity to the drug. Expression of cancer stem cell markers, as well as tumour sphere formation, was also increased in EP300-depleted cells, and was diminished in EP300-overexpressing cells. The EP300-regulated gene signature highlighted genes associated with adhesion CEACAM5, cytoskeletal remodelling CAPN9, stemness ABCG2, apoptosis BCL2 and metastasis TGFB2. Some genes in this signature were also validated in a previously generated EP300-depleted model of breast cancer using minimally transformed mammary epithelial cells. Importantly, two key genes in apoptosis and stemness, BCL2 and ABCG2, were also upregulated in EP300-knockout colon carcinoma cells and their paclitaxel-resistant derivatives. Immunohistochemical analysis demonstrated that EP300 expression was low in metaplastic breast cancer, a rare, but aggressive form of the disease with poor prognosis that is characterized by morphological and physiological features of EMT.

ConclusionsEP300 plays a major role in the reprogramming events, leading to a more malignant phenotype with the acquisition of drug resistance and cell plasticity, a characteristic of metaplastic breast cancer.

KeywordsMetastasis Cancer stem cells Drug resistance BCL2 ABCG2 EP300 signature AbbreviationsALDHAldehyde dehydrogenase

CSCCancer stem cell

DMEMDulbecco’s modified Eagle medium

EMTEpithelial-to-mesenchymal transition

IC50Drug concentration producing 50% cell death

miRMicro RNA

MTMECMinimally transformed mammary epithelial cell

QPCRQuantitative PCR

Electronic supplementary materialThe online version of this article doi:10.1007-s10549-017-4202-z contains supplementary material, which is available to authorized users.





Autor: Muhammad Asaduzzaman - Stephanie Constantinou - Haoxiang Min - John Gallon - Meng-Lay Lin - Poonam Singh - Selina Raguz - S

Fuente: https://link.springer.com/







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