The effects of soy supplementation on gene expression in breast cancer: a randomized placebo-controlled study

Abstract

Background: There are conflicting reports on the impact of soy on breast carcinogenesis. This study examines the effects of soy supplementation on breast cancer-related genes and pathways.

Methods: Women (n = 140) with early-stage breast cancer were randomly assigned to soy protein supplementation (n = 70) or placebo (n = 70) for 7 to 30 days, from diagnosis until surgery. Adherence was determined by plasma isoflavones: genistein and daidzein. Gene expression changes were evaluated by NanoString in pre- and posttreatment tumor tissue. Genome-wide expression analysis was performed on posttreatment tissue. Proliferation (Ki67) and apoptosis (Cas3) were assessed by immunohistochemistry.

Results: Plasma isoflavones rose in the soy group (two-sided Wilcoxon rank-sum test, P < .001) and did not change in the placebo group. In paired analysis of pre- and posttreatment samples, 21 genes (out of 202) showed altered expression (two-sided Student's t-test, P < .05). Several genes including FANCC and UGT2A1 revealed different magnitude and direction of expression changes between the two groups (two-sided Student's t-test, P < .05). A high-genistein signature consisting of 126 differentially expressed genes was identified from microarray analysis of tumors. This signature was characterized by overexpression (>2-fold) of cell cycle transcripts, including those that promote cell proliferation, such as FGFR2, E2F5, BUB1, CCNB2, MYBL2, CDK1, and CDC20 (P < .01). Soy intake did not result in statistically significant changes in Ki67 or Cas3.

Conclusions: Gene expression associated with soy intake and high plasma genistein defines a signature characterized by overexpression of FGFR2 and genes that drive cell cycle and proliferation pathways. These findings raise the concerns that in a subset of women soy could adversely affect gene expression in breast cancer.

Trial registration: ClinicalTrials.gov NCT00597532.

Figure 1.

CONSORT diagram for study design and availability of samples for analysis.

Figure 2.

Plasma genistein and daidzein following soy intake. Post-treatment isoflavone levels increased compared with pretreatment levels following intake of soy (P < .001) but not placebo. The changes (post-pre) in plasma isoflavones were statistically significantly greater for women receiving soy compared with those receiving placebo (P < .001). * Indicates within-group statistical significance by the Wilcoxon matched-pairs signed rank test, P < .001. † Indicates statistical significance for the comparison of the fold-change between treatment groups by the Wilcoxon rank-sum test (P < .001).

Figure 3.

Expression of FANCC and UGT2A1. Gene expression was measured in paired samples (pre/post) using NanoString. Expression levels of FANCC (A), UGT2A1 (C), and SERPINE1 (E) were increased following consumption of soy (P < .05), but not placebo. The expression fold changes (FC) were statistically significantly greater in tumors exposed to soy compared with placebo for FANCC (B) and UGT2A1 (D) (P < .05), but not SERPINE1 (P = .26) (F). A heat map and hierarchical clustering of the post/pre FC of the 21 genes DE between paired samples (P < .05) (G). Positive FC are colored red, negative FC are colored blue, and treatment group is indicated below the sample dendogram. *Indicates within-group statistical significance by paired t test P < .05, †Indicates statistical significance for the comparison of fold change between treatment groups by unpaired t test (P < .05).

Figure 4.

Hierarchical clustering of DE genes in high versus low- genistein subsets. Clustering of samples and 126 DE genes between high- and low-genistein samples (fold change > 2 and t-test P < .1) as identified by microarrays. Clustering was performed using Euclidian distance. Gene expression values are log2 transformed and standardized. ER status, menopausal status, and genistein plasma concentrations for each sample are indicated. Twelve tumors of the soy group with high (>16ng/mL) plasma genistein defined the high-genistein subset, and 22 tumors of the placebo group with low (<6.8ng/mL) genistein defined the low-genistein subset. FGFR2 was overexpressed in three of 12 tumors of the high-genistein group (arrow, P < .01).

Figure 5.

Differential expression of FGFR2. FGFR2 expression was increased 2.4 fold in the high-genistein subset of the soy group by microarray (P = .006) (A). FGFR2 overexpression by qPCR in soy vs placebo (P = .03) (B). Nanostring analysis of pre/post samples. Although there were no statistically significant differences for the group as a whole, in two samples from the soy group there was a marked increase in FGFR2 expression (C). *Indicates statistically significant overexpression (P < .05).