Troglitazone-Induced PRODH/POX-Dependent Apoptosis Occurs in the Absence of Estradiol or ERβ in ER-Negative Breast Cancer Cells
by
, , ,
Sylwia Lewoniewska
1,†, 
Ilona Oscilowska
2,†,
Thi Yen Ly Huynh
1,
Izabela Prokop
1,
Weronika Baszanowska
1,
Katarzyna Bielawska
1 and
Jerzy Palka
1,* 1
Department of Medicinal Chemistry, Medical University of Bialystok, Kilinskiego 1, 15-089 Bialystok, Poland
2
Department of Analysis and Bioanalysis of Medicines, Medical University of Bialystok, Kilinskiego 1, 15-089 Bialystok, Poland
*
Author to whom correspondence should be addressed.
†
These authors contributed equally to this work.
J. Clin. Med. 2021, 10(20), 4641; https://doi.org/10.3390/jcm10204641
Submission received: 23 August 2021
/
Revised: 28 September 2021
/
Accepted: 8 October 2021
/
Published: 10 October 2021
(This article belongs to the Section Oncology)
Simple Summary
PRODH/POX (proline dehydrogenase/proline oxidase) is a mitochondrial enzyme that catalyzes proline degradation generating reactive oxygen species (ROS). Estrogens limit proline availability for PRODH/POX by stimulating collagen biosynthesis. It has been considered that estrogens determine efficiency of troglitazone (TGZ)-induced PRODH/POX-dependent apoptosis in breast cancer cells. The studies were performed in wild-type and PRODH/POX-silenced estrogen-dependent MCF-7 cells and estrogen-independent MDA-MB-231 cells. DNA and collagen biosynthesis were determined by radiometric method, ROS production was measured by fluorescence assay, protein expression was determined by Western blot and proline concentration by LC/MS analysis. We found that: i/TGZ-induced apoptosis in MDA-MB-231 occurs only in the absence of estradiol or ERβ, ii/the process is mediated by PRODH/POX, iii/and is facilitated by proline availability for PRODH/POX by TGZ-dependent inhibition of collagen biosynthesis (proline utilizing process). The data suggest that combined TGZ and anti-estrogen treatment could be considered in experimental therapy of ER negative breast cancers.
Abstract
The impact of estradiol on troglitazone (TGZ)-induced proline dehydrogenase/proline oxidase (PRODH/POX)-dependent apoptosis was studied in wild-type and PRODH/POX-silenced estrogen receptor (ER) dependent MCF-7 cells and ER-independent MDA-MB-231 cells. DNA and collagen biosynthesis were determined by radiometric method, prolidase activity evaluated by colorimetric method, ROS production was measured by fluorescence assay. Protein expression was determined by Western blot and proline concentration by LC/MS analysis. PRODH/POX degrades proline yielding reactive oxygen species (ROS). Estrogens stimulate collagen biosynthesis utilizing free proline and limiting its availability for PRODH/POX-dependent apoptosis. TGZ cytotoxicity was highly pronounced in wild-type MDA-MB-231 cells cultured in medium without estradiol or in the cells cultured in medium with estradiol but deprived of ERβ (by ICI-dependent degradation), while in PRODH/POX-silenced cells the process was not affected. The TGZ cytotoxicity was accompanied by increase in PRODH/POX expression, ROS production, expression of cleaved caspase-3, caspase-9 and PARP, inhibition of collagen biosynthesis, prolidase activity and decrease in intracellular proline concentration. The phenomena were not observed in PRODH/POX-silenced cells. The data suggest that TGZ-induced apoptosis in MDA-MB-231 cells cultured in medium without estradiol or deprived of ERβ is mediated by PRODH/POX and the process is facilitated by proline availability for PRODH/POX by TGZ-dependent inhibition of collagen biosynthesis. It suggests that combined TGZ and antiestrogen treatment could be considered in experimental therapy of estrogen receptor negative breast cancers.
Keywords:
apoptosis; PRODH/POX; collagen; estrogen; breast cancer cells
