Journal Article

HIF-1 activation induces doxorubicin resistance in MCF7 3-D spheroids via P-glycoprotein expression: a potential model of the chemo-resistance of invasive micropapillary carcinoma of the breast

Doublier, Sophie, Belisario, Dimas C, Annaratone, Laura, Sapino, Anna, Allia, Elena, Ghigo, Dario, Polimeni, Manuela, Bosia, Amalia, Riganti, Chiara

2012

Overview

Background Invasive micropapillary carcinoma (IMPC) of the breast is a distinct and aggressive variant of luminal type B breast cancer that does not respond to neoadjuvant chemotherapy. It is characterized by small pseudopapillary clusters of cancer cells with inverted cell polarity. To investigate whether hypoxia-inducible factor-1 (HIF-1) activation may be related to the drug resistance described in this tumor, we used MCF7 cancer cells cultured as 3-D spheroids, which morphologically simulate IMPC cell clusters. Methods HIF-1 activation was measured by EMSA and ELISA in MCF7 3-D spheroids and MCF7 monolayers. Binding of HIF-1α to MDR-1 gene promoter and modulation of P-glycoprotein (Pgp) expression was evaluated by ChIP assay and FACS analysis, respectively. Intracellular doxorubicin retention was measured by spectrofluorimetric assay and drug cytotoxicity by annexin V-FITC measurement and caspase activity assay. Results In MCF7 3-D spheroids HIF-1 was activated and recruited to participate to the transcriptional activity of MDR-1 gene, coding for Pgp. In addition, Pgp expression on the surface of cells obtained from 3-D spheroids was increased. MCF7 3-D spheroids accumulate less doxorubicin and are less sensitive to its cytotoxic effects than MCF7 cells cultured as monolayer. Finally, HIF-1α inhibition either by incubating cells with 3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole (a widely used HIF-1α inhibitor) or by transfecting cells with specific siRNA for HIF-1α significantly decreased the expression of Pgp on the surface of cells and increased the intracellular doxorubicin accumulation in MCF7 3-D spheroids. Conclusions MCF7 breast cancer cells cultured as 3-D spheroids are resistant to doxorubicin and this resistance is associated with an increased Pgp expression in the plasma membrane via activation of HIF-1. The same mechanism may be suggested for IMPC drug resistance.

• Publisher: BioMed Central,Springer Nature B.V,BMC
• Subject: 3-D spheroids, Elastase / ABC transporters / Aggressiveness / Annexins - analysis / Antineoplastic Agents - pharmacology / ATP-Binding Cassette, Sub-Family B, Member 1 - metabolism / Biomedical and Life Sciences / Biomedicine / Breast cancer / Breast Neoplasms - drug therapy / Breast Neoplasms - metabolism / Cancer Research / Cancer therapies / Carcinoma, Papillary - drug therapy / Carcinoma, Papillary - metabolism / Caspases - analysis / Deoxyribonucleic acid / DNA / DNA repair / Doxorubicin - pharmacology / Doxorubicin resistance / Drug resistance / Drug Resistance, Neoplasm - physiology / Experimental therapeutics and drug development / Experiments / Female / Fluorides / Health Promotion and Disease Prevention / HIF-1α / Humans / Hypoxia / Hypoxia-Inducible Factor 1 - metabolism / Invasive micropapillary breast carcinoma / Kinases / Medicine/Public Health / MUC-1 / Nitric oxide / Oncology / P-glycoprotein / Research Article / Spheroids, Cellular / Studies / Surgical Oncology / Tumor Cells, Cultured