Ben Newman and Dr. Chin-Yo Lin, Microbiology and Molecular Biology
Introduction
Breast cancer is the most prevalent type of cancer found in women and the majority of such cancers are estrogen receptor (ER) positive. This means the cancer is dependent on estrogen for growth. In such cancers, ER acts as a ligand-dependent transcription factor, changing gene expression when bound to estrogen. The altered expression of ER target genes in breast cancer is a likely cause of the uncontrolled cell growth.
Estrogen receptor also plays important physiological roles in the body such as bone density maintenance and cardioprotection. Direct inhibition of ER in breast cancer treatment would therefore have detrimental consequences. To inhibit ER function in breast tissue while allowing it in other areas of the body, drugs called selective estrogen receptor modulators (SERMs) were developed. The anti-ER function of these drugs is tissue specific.
Most previous studies on ER target genes focused on those genes that are over-expressed following exposure to estrogen. For example, the action of SERMs was characterized based on up-regulated genes such as PS2. Previous research in our lab verified microarray data showing the under-expression of several ER target genes. To better understand the effect SERMs have on these newly discovered down-regulated genes, we measured gene expression in MCF-7 breast cancer cells following drug treatment. Tamoxifen, Raloxifene, and ICI-182 were the SERMs used to treat cells and expression levels of the genes HES1, MME, NFIA, PSCA, SLC35a1, ZNRF3, COBL, and CYP1A1 were measured.
Results
We used PS2, a well known up-regulated ER target gene, as a positive control. Following estrogen (E2) treatment to activate ER, cells showed a 3.5 fold increase in PS2 expression. This indicates that the ER response was working. The PS2 over-expression was abolished when accompanied by SERM treatment (Figure 1). This established that the drugs were successfully inhibiting ER function. Treating the cells with the drugs in the absence of E2 showed less gene expression than when untreated, likely due to inhibition of basal PS2 levels.
We measured gene expression of eight genes that we previously had found to be down-regulated following E2 treatment. These genes were chosen based on previous associations with cancer or other diseases and physiological functions in the body. ZNRF3 and COBL did not show significant down-regulation when treated with E2 and no significant effect on expression was observed when SERM treatment was included. HES1, MME, PSCA, SLC35a1, and CYP1A1 all showed down-regulation following E2 treatment. In these genes, ICI and Raloxifene prevented the down-regulation more consistently than Tamoxifen. NFIA actually showed a slight up-regulation following E2 treatment, while drug treatments strongly lessened its expression levels.
Discussion
Unlike the up-regulated control gene which showed expected behavior following SERM treatment, these down-regulated genes show a variety of responses to SERM treatment. Not all genes respond in a similar way to treatment with a particular SERM. Treatment of different SERMs can have different results on expression levels of a particular gene. The wide array of responses to SERM treatments indicate that different SERMs will have different specificity and efficacy in targeting ER. Understanding these differences could have direct clinical relevance in breast cancer treatment. Adverse side-effects and off-target effects may be better explained and avoided as more is understood about how these drugs work. Such knowledge may also aid in identifying more effective SERMs in the future.
References
- American Cancer Society. Cancer Facts & Figures 2006. Atlanta: American Cancer Society; 2006.
- Gruber, et al. Production and Actions of Estrogens. N Engl J Med. Vol. 346, No. 5. 2002.
- Lin, C-Y, et al. Discovery of estrogen receptor a target genes and response elements in breast tumor cells. Genome Biology. Vol. 5, Issue 9, Article R66. 2004.