The Battle for HOXC10 in the War Against Breast Cancer
Tamoxifen and aromatase inhibitors (AIs) are both treatment options for patients with breast tumors that express estrogen receptor (ER) α and that require estrogen for growth. Tamoxifen competes with estrogen in binding to the ER, but does not activate the receptor. In comparison, AIs lower estrogen levels in the body by blocking the conversion of testosterone to estrogen. However, this is only offered to postmenopausal women because the ovaries in premenopausal woman would still continue to produce estrogen, negating the effects of treatment. Treatment with AI for postmenopausal ER+ breast cancer has proven to be much more effective than tamoxifen. Unfortunately, resistance to these endocrine therapies is currently a major clinical problem and while possible mechanisms of resistance to tamoxifen have been identified, mechanisms of resistance to AIs are largely unknown. One model, recently published by Pathiraja, T.N, et. al., suggests that the apoptotic and growth-inhibitory gene, HOXC10, may be repressed by estrogen and thus activated by AIs- but long-term AI treatment results in epigenetic reprogramming and permanent repression of HOXC10, which leads to acquired resistance.
The researchers used sub-lines C4-12 (ER–) and LTED (ER+), derived from cell line MCF-7 (ER+), that are resistant to estrogen deprivation to test the proposed model. The cell lines were first subjected to a genome-wide methylation binding domain pull-down experiment focusing on a list of the commonly hypermethylated genes between the cell lines, given the relationship between hypermethylation and gene silencing. HOXC10 was chosen from this list for subsequent studies due to its hypermethylated status in both C4-12 and LTED, and the importance of HOX genes in tumorigenesis. Quantification of HOXC10 mRNA in C4-12, LTED, and MCF-7 before and after treatment with a DNA methylation inhibitor and histone deacetylase inhibitor suggests that both DNA methylation of the CpG island shore in the HOXC10 proximal promoter and histone modifications are involved in transcriptional silencing of HOXC10. A larger study of the effect of methylation on additional breast cancer cell lines indicated that methylation at the CpG island shore is inversely correlated with expression. Given that most of the ER+ cell lines expressed HOXC10 above the average level, Pathiraja T.N., et al. thought that HOXC10 expression might be regulated by estrogen. Quantification of HOXC10 mRNA expression and ChIP studies of cell lines treated with estradiol and antiestrogen suggested not only that estrogen causes repression of HOXC10 mRNA expression in non-resistant cells, but that short-term deprivation of estrogen is associated with increased EZH2 and H3K27me3. The researchers then asked whether resistance to estrogen deprivation could be attributed to the loss of HOXC10 expression and subsequent proliferation of the cancer. They designed short hairpin RNAs to decrease HOXC10 expression in MCF-7 cells and found that decreased expression allowed for increased cell growth and motility, and decreased apoptosis and sensitivity to estrogen deprivation therapies. In vivo experiments using an endocrine resistant xenograft model also supported the hypothesis that repression of HOXC10 promotes resistance to estrogen deprivation. Additionally, HOXC10 levels were measured by immunohistochemistry (IHC) or qPCR in matched primary-recurrent tumor pairs from patients treated with tamoxifen and were found to be lower in recurrent tumor samples.
The model of epigenetic reprogramming in endocrine resistance proposed by Prathiraja T.N., et al. was supported not only by experiments in cell lines, but in clinical samples as well. This paper focused on only one of dozens of hypermethylated genes that surfaced from the genome-wide MBD-PD and there may be many more genes that play a role in endocrine resistance. Nevertheless, the findings in this paper suggest that one approach to the treatment of breast cancer with AI may have to include mechanisms to overcome methylation of targeted regions.
Pathiraja TN, Nayak SR, Xi Y, Jiang S, Garee JP, Edwards DP, Lee AV, Chen J, Shea MJ, Santen RJ, Gannon F, Kangaspeska S, Jelinek J, Issa JP, Richer JK, Elias A, McIlroy M, Young LS, Davidson NE, Schiff R, Li W, & Oesterreich S (2014). Epigenetic reprogramming of HOXC10 in endocrine-resistant breast cancer. Science translational medicine, 6 (229) PMID: 24670685