Polyphenon E Treatment Alters Gene Expression in Prostate Cancer Cells

Abstract

Prostate cancer (PCa) is the most common cancer among men in the United States. Polyphenon E (Poly E) is a standardized caffeine‐free green tea extract. Poly E has been reported to have chemopreventative activity against prostate cancer. The molecular mechanisms of this chemopreventative activity are not fully understood, however, the major constituent of Poly E, epigallocatechin‐3‐gallate (EGCG), has been reported to alter expression of some genes. It was hypothesized that Poly E treatment induces gene expression changes, which could provide insights to its molecular mechanism. The purpose of this experimental work was to identify genes that experienced substantially altered gene expression levels (changes 3‐fold or more) after Poly E treatment. Three human prostate cancer cell lines (DU 145, PC‐3, and LNCaP) were treated at three concentrations of Poly E (low, mid, high). Total cellular RNA was isolated from Poly E‐treated (and untreated) cells using RNA Easy Plus Miniprep Kit (Qiagen). RNA samples were subjected to qualitative and quantitative analysis using a BioAnalyzer RNA Chip (Agilent) at the Molecular Genomics Core facility at Moffitt Cancer Center. Analyzed RNAs at the mid‐concentration treatment were used for DNA microarray analyses with an Affymetrix Human U133 Plus 2.0 arrays. From the analysis of the microarray data acquired, numerous genes were identified with altered gene expression levels (increased or decreased >3‐fold) after Poly E treatment. From this list, selected genes were analyzed by TaqMan qRT‐PCR using the ΔΔCt method in a StepOne™ RT‐PCR System. Reported RQ values were reported as fold changes. Analyzed RNAs were converted into cDNA with DyNAmo cDNA Synthesis Kit (Fischer Scientific) and were used as templates for qRT‐PCR. Untreated samples served as the reference control and ACTB served as the endogenous control. At the highest Poly E concentrations tested, TaqMan qRT‐PCR results for SEC62 were consistent with less than three‐fold gene expression changes for all cell lines studied (2.0‐fold increase in DU 145, 1.8‐fold increase in PC‐3, 2.1‐fold decrease in LNCaP). WNT9a gene expression levels experienced minor changes at the highest treatment level in PC‐3 and LNCaP cell lines (decreased 1.4‐fold, increased 1.4‐fold, respectively), however, increased 7.8‐fold in DU 145 cells. RB1 expression levels decreased by 3.1‐fold in PC‐3 cells, but remained essentially unchanged in DU 145 and LNCaP (both increased 1.2‐fold). MXD1 gene expression levels increased by 5.6‐fold in PC‐3 cells and 13.0‐fold in DU 145, but remained essentially unchanged (increased 1.1‐fold) in LNCaP. RGS4 gene expression levels experienced a 102‐fold increase in DU 145 and a 5.1‐fold increase in PC‐3 cells, while LNCaP cells exhibited a 5.6‐fold decrease. DU 145 cells experienced increased gene expression levels for all genes analyzed, while LNCaP cells had either a marginal increase of less than 2‐fold (WNT9a, RB1, MXD1) or a substantial decrease in the gene expression level (SEC62 decreased 2.1‐fold; RGS4 decreased 5.6‐fold). Collectively, these data could provide insights into new avenues for the development of biomarker or therapeutic interventions for prostate cancer.Support or Funding InformationThis work was financially supported by a University of Tampa David Delo Research Professor Grant (PI: L. M. Carastro), a University of Tampa Dana Research Professor Grant (PI: L. M. Carastro). This work was also supported by funding from the Department of Chemistry, Biochemistry & Physics at the University of Tampa to support student research. We acknowledge the Moffitt Cancer Center Genomic Core Facility for excellent technical support in performing DNA microarray analyses.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.