Epigenetic reprogramming in breast cancer: From new targets to new therapies

Figures & data

Figure 1. Proposed model of epigenetic regulation of gene expression in breast cancer. DNA methylation is generally associated with transcription repression, while histone modifications can be either active or repressive. DNA is methylated by DNMTs and demethylated by a multistep process by the TETs. There are multiple post-translational modifications that occur on histone tails. A multitude of enzymes function to methylate (HMT: Set8, Set9, SUV39H1, EZH2), demethylate (KDM: JARID1B, LSD1/2, JMJD2C, JMJD3), acetylate (HAT), or deacetylate (HDAC) histones. DNA hypermethylation at a gene promoter in association with abnormal histone modifications in breast cancer cells may lead to the anomalous loss of genes that are important in curbing tumor growth. Established or investigational agents that can block many of the enzymes are shown in green boxes.

Figure 2. Epigenetic predisposition to breast cancer. Epigenetic regulation is instrumental in mammary gland development from the time the mammary streak is created in utero through rounds of pregnancy and lactation, and it is clearly involved in breast cancer development. Early in fetal development the genome is globally demethylated and remethylated in the appropriate patterns, but this process can be disrupted by toxin exposure and poor diet. The effects exerted by poor diet and toxin exposure can be dampened by maternal intake of folic acid or choline. During pregnancy the mammary gland DNA becomes globally hypomethylated to allow active transcription of genes needed to reorganize the tissue structure to develop alveolar buds and prepare for lactation. Once lactation commences and alveoli are mature, H3 and H4 in the chromatin of the gland become additionally hyperacetylated, allowing for activation of milk production genes. During breast cancer development, this process is abrogated and DNA is typically hypermethylated while H3 and H4 are generally hypoacetylated.

Table I. Select epigenetic agents with potential clinical applications in breast cancer. A list of epigenetic agents, their epigenetic targets, and characteristics and clinical implications is presented.

Epigenetic category	Epigenetic agent	Potential clinical applications
Methyl donors	Choline	Dietary constituent with potential for breast cancer prevention (65,67)
	Folate
DNA methyltransferase inhibitors	Azacitidine (5-azacytidine)	DNMTi with a very short half-life, FDA approved for treatment of MDS (96)
	Decitabine (5-aza-2’-deoxycytidine)	DNMTi with a very short half-life, FDA approved for treatment of MDS (96)
	SGI-110	Investigational DNMTi with a prolonged half-life due to resistance to cytosine deaminase (80,81,83)
	NPEOD-DAC
	Zebularine
Histone deacetylase inhibitors	Sulphoraphane	Dietary constituent in brassica vegetables with potential for breast cancer prevention (71–73,98)
	Phenethyl isothiocyanate (PEITC)
	Trichostatin A (TSA)	Laboratory tool without clinical application
	Vorinostat Romidepsin	FDA-approved agents for cutaneous T-cell lymphoma
Histone lysine demethylase inhibitors	Tranylcypromine (TCP)	Monoamine oxidase inhibitors showing inhibitory activity against LSD1 and/or LSD2 (26,31,116)
	Pargyline
	Polyamine analogues	Inhibiting LSD1 activity (25)
	JIB-04	Jumonji KDMi directly targets α-ketoglutarate (89)
Histone methyltransferase inhibitors	EPZ005687	SAM-competitive EZH2 inhibitor (92–94)
	EI1
	GSK126
Histone deacetylase and DNA methyltransferase inhibitor	Genistein	Dietary constituent in soy products; acts synergistically with tamoxifen to induce cell death in cancer cells lines (97)

Table II. Trials of epigenetic therapies in breast cancer. Selected clinical trials using epigenetic agents for the treatment of breast cancer, trial details, and available results are presented.

Therapeutic category	Individual therapies	Study population	Trial design	Result (reference)
HDACi	Vorinostat	Advanced breast cancer	Phase II	No CR or partial response; 4 patients with SD of 4, 8, 9, and 14 months (102)
	Vorinostat vs. untreated controls	Newly diagnosed invasive breast cancer	Non-randomized; preoperative window study	Significant decreases in the gene expression of Ki67, STK15, and cyclin B1 following vorinostat, but not in other genes by the oncotype DX assay. No significant change in expression of expression of Ki67 or cleaved caspase-3 by IHC. No changes in methylation (103)
HDACi + endocrine therapy	Vorinostat + tamoxifen	43 ER+ metastatic breast cancer progressed on endocrine therapy	Phase II	ORR 19%; CBR 40%; significantly increased histone H4 acetylation and baseline HDAC2 levels in PBMCs in patients with a response or SD vs. non-responders (105)
	Entinostat + aromatase inhibitor	27 patients with ER+ metastatic breast cancer; progressed on AI	Phase II	One partial response; one SD > 6 months; 41% received study treatment for > 4 months (106)
	Exemestane + entinostat vs. exemestane + placebo	130 patients with advanced ER+ breast cancer progressed on AI	Randomized, double-blind, phase II	Improved PFS with combination 4.3 vs. 2.3 months; median OS 28.1 months with combination vs. 19.8 months (107)
	Panobinostat + letrozole	Triple-negative metastatic breast cancer	Phase I/II	No reported results for phase II (108)
	Vorinostat after aromatase inhibitors	ER+ metastatic breast cancer progressed on AI	Phase II	Not yet reported (109)
DNMTi + HDACi + endocrine therapy	5-AZA + entinostat + optional continuation phase with endocrine therapy at progression	40 patients with advanced breast cancer progressed on chemotherapy and endocrine therapy; one triple-negative and one HR+ cohort	Phase II	No response in triple-negative cohort; one partial response in the first stage of HR+ cohort, allowing for second stage enrollment; final results pending (104)
HDACi + DNMTi	Hydralazine + magnesium valproate + ac	16 patients with locally advanced breast cancer	Non-randomized; neoadjuvant	ORR 81%; 31% CR; 50% partial response; 6.6% pCR; significant decrease in global 5mC content and HDAC activity (110)
HDACi + chemotherapy	Vorinostat + paclitaxel + bevacizumab	54 patients with metastatic breast cancer	Phase I/II	55% ORR in phase II; paired tumor biopsies before and after the third vorinostat dose showed increased acetylation of Hsp90 and α-tubulin, and functional inhibition of Hsp90 (111)
	Vorinostat + capecitabine	14 evaluable patients with metastatic breast cancer	Phase I	No objective responses in 14 evaluable patients; 3 patients with SD > 6 months; 1 patient completed 42 cycles of therapy (113)
HDACi + chemotherapy	Carboplatin + nab-paclitaxel with vorinostat vs. placebo	62 patients with unresected stage II-III HER2-negative breast cancer	Neoadjuvant phase II double-blind	Initial results show 27.6% pCR for chemotherapy + vorinostat vs. 26.7% in placebo arm (114)
HDACi + anti-HER2 therapy	Vorinostat + trastuzumab	16 patients with resistant HER2 + metastatic breast cancer	Phase I/II	No objective response in centrally confirmed HER2 + patients; one objective response in patient with HER2-negative disease (115)

SD = stable disease.

Stefanska B, Karlic H, Varga F, Fabianowska-Majewska K, Haslberger A. Epigenetic mechanisms in anti-cancer actions of bioactive food components—the implications in cancer prevention. Br J Pharmacol. 2012;167:279–97.

 PubMed Web of Science ®Google Scholar

Gong Z, Ambrosone CB, McCann SE, Zirpoli G, Chandran U, Hong CC, et al. Associations of dietary folate, vitamins B6 and B12 and methionine intake with risk of breast cancer among African American and European American women. Int J Cancer. 2014;134: 1422–35.

 PubMed Web of Science ®Google Scholar

Vijayaraghavalu S, Dermawan JK, Cheriyath V, Labhasetwar V. Highly synergistic effect of sequential treatment with epigenetic and anticancer drugs to overcome drug resistance in breast cancer cells is mediated via activation of p21 gene expression leading to G2/M cycle arrest. Mol Pharm. 2013;10:337–52.

 PubMed Web of Science ®Google Scholar

Yoo CB, Jeong S, Egger G, Liang G, Phiasivongsa P, Tang C, et al. Delivery of 5-aza-2’-deoxycytidine to cells using oligodeoxynucleotides. Cancer Res. 2007;67:6400–8.

 PubMed Web of Science ®Google Scholar

Byun HM, Choi SH, Laird PW, Trinh B, Siddiqui MA, Marquez VE, et al. 2’-Deoxy-N4-[2-(4-nitrophenyl)ethoxycarbonyl]-5-azacytidine: a novel inhibitor of DNA methyltransferase that requires activation by human carboxylesterase 1. Cancer Lett. 2008; 266:238–48.

 Google Scholar

Billam M, Sobolewski MD, Davidson NE. Effects of a novel DNA methyltransferase inhibitor zebularine on human breast cancer cells. Breast Cancer Res Treat. 2010;120:581–92.

 PubMed Web of Science ®Google Scholar

Liu K, Cang S, Ma Y, Chiao JW. Synergistic effect of paclitaxel and epigenetic agent phenethyl isothiocyanate on growth inhibition, cell cycle arrest and apoptosis in breast cancer cells. Cancer Cell Int. 2013;13:10.

 PubMed Web of Science ®Google Scholar

Huang Y, Vasilatos SN, Boric L, Shaw PG, Davidson NE. Inhibitors of histone demethylation and histone deacetylation cooperate in regulating gene expression and inhibiting growth in human breast cancer cells. Breast Cancer Res Treat. 2012;131:777–89.

 PubMed Web of Science ®Google Scholar

Lim S, Janzer A, Becker A, Zimmer A, Schule R, Buettner R, et al. Lysine-specific demethylase 1 (LSD1) is highly expressed in ER-negative breast cancers and a biomarker predicting aggressive biology. Carcinogenesis. 2010;31:512–20.

 PubMed Web of Science ®Google Scholar

Karytinos A, Forneris F, Profumo A, Ciossani G, Battaglioli E, Binda C, et al. A novel mammalian flavin-dependent histone demethylase. J Biol Chem. 2009;284:17775–82.

 PubMed Web of Science ®Google Scholar

Zhu Q, Huang Y, Marton LJ, Woster PM, Davidson NE, Casero RA Jr. Polyamine analogs modulate gene expression by inhibiting lysine-specific demethylase 1 (LSD1) and altering chromatin structure in human breast cancer cells. Amino Acids. 2012;42:887–98.

 PubMed Web of Science ®Google Scholar

Wang L, Chang J, Varghese D, Dellinger M, Kumar S, Best AM, et al. A small molecule modulates Jumonji histone demethylase activity and selectively inhibits cancer growth. Nat Commun. 2013;4:2035.

 PubMed Web of Science ®Google Scholar

Li Y, Meeran SM, Patel SN, Chen H, Hardy TM, Tollefsbol TO. Epigenetic reactivation of estrogen receptor-alpha (ERalpha) by genistein enhances hormonal therapy sensitivity in ERalpha-negative breast cancer. Mol Cancer. 2013;12:9.

 PubMed Web of Science ®Google Scholar

Luu TH, Morgan RJ, Leong L, Lim D, McNamara M, Portnow J, et al. A phase II trial of vorinostat (suberoylanilide hydroxamic acid) in metastatic breast cancer: a California Cancer Consortium study. Clin Cancer Res. 2008;14:7138–42.

 PubMed Web of Science ®Google Scholar

Stearns V, Jacobs LK, Fackler M, Tsangaris TN, Rudek MA, Higgins M, et al. Biomarker modulation following short-term vorinostat in women with newly diagnosed primary breast cancer. Clin Cancer Res. 2013;19:4008–16.

 PubMed Web of Science ®Google Scholar

Munster PN, Thurn KT, Thomas S, Raha P, Lacevic M, Miller A, et al. A phase II study of the histone deacetylase inhibitor vorinostat combined with tamoxifen for the treatment of patients with hormone therapy-resistant breast cancer. Br J Cancer. 2011;104:1828–35.

 PubMed Web of Science ®Google Scholar

Wardley AM, Stein R, McCaffrey J, Crown J, Malik Z, Rea D, Barrett-Lee PJ, et al. Phase II data for entinostat, a class 1 selective histone deacetylase inhibitor, in patients whose breast cancer is progressing on aromatase inhibitor therapy. 2010 ASCO Annual Meeting Abstract. J Clin Oncol. 2010;28(15 suppl):1052.

 Google Scholar

Yardley DA, Ismail-Khan RR, Melichar B, Lichinitser M, Munster PN, Klein PM, et al. Randomized phase II, double-blind, placebo-controlled study of exemestane with or without entinostat in postmenopausal women with locally recurrent or metastatic estrogen receptor-positive breast cancer progressing on treatment with a nonsteroidal aromatase inhibitor. J Clin Oncol. 2013;31:2128–35.

 PubMed Web of Science ®Google Scholar

Tan W, Allred JB, Moreno-Aspitia A, Northfelt DW, Ingle JN, Perez EA. Phase I study of panobinostat (LBH589) and letrozole in post-menopausal women with metastatic breast cancer. J Clin Oncol. 2012;30(suppl); abstr e13501.

 Google Scholar

Linden HM, Kurland BF, Link J, Gadi VK, Specht JM, Gralow J, et al. Vorinostat to restore sensitivity to aromatase inhibitor therapy in metastatic breast cancer: a phase II clinical trial with ER imaging correlates. J Clin Oncol. 2012;30(suppl): abstr TPS3109.

 Google Scholar

Connolly RM, Jankowitz RC, Zahnow CA, Zhang Z, Rudek MA, Jeter SC, et al. A phase 2 study investigating the safety, efficacy and surrogate biomarkers of response of 5-azacitidine (5-AZA) andentinostat (MS-275) in patients with triple-negative advanced breast cancer. Cancer Res. 2013;73(8 Suppl 1).

 Google Scholar

Arce C, Perez-Plasencia C, Gonzalez-Fierro A, de la Cruz-Hernandez E, Revilla-Vazquez A, Chavez-Blanco A, et al. A proof-of-principle study of epigenetic therapy added to neoadjuvant doxorubicin cyclophosphamide for locally advanced breast cancer. PLoS One. 2006;1:e98.

 PubMed Web of Science ®Google Scholar

Ramaswamy B, Fiskus W, Cohen B, Pellegrino C, Hershman DL, Chuang E, et al. Phase I-II study of vorinostat plus paclitaxel and bevacizumab in metastatic breast cancer: evidence for vorinostat- induced tubulin acetylation and Hsp90 inhibition in vivo. Breast Cancer Res Treat. 2012;132:1063–72.

 PubMed Web of Science ®Google Scholar

James ES, Chung GG, DiGiovanna M, Sanft TB, Hofstatter EW, Sowers N, et al. Phase I study of the HDAC inhibitor vorinostat in combination with capecitabine in a biweekly schedule in advanced breast cancer. J Clin Oncol 2013;31(suppl); abstr 2587.

 Google Scholar

Connolly RM, Jeter S, Zorzi J, Zhang Z, Armstrong DK, Fetting JH, et al. A multi-institutional double-blind phase II study evaluating response and surrogate biomarkers to carboplatin and nab-paclitaxel (CP) with or without vorinostat as preoperative systemic therapy (PST) in HER2- negative primary operable breast cancer (TBCRC008). 2010 ASCO Annual Meeting Abstracts. J Clin Oncol. 2010;28(15 suppl):TPS111.

 Google Scholar

Swaby R, Sparano J, Bhalla K, Meropol N, Falkson C, Pellegrino C, et al. A phase II study of the histone deacetylase inhibitor, vorinostat, in combination with trastuzumab in patients with advanced metastatic and/or local chest wall recurrent HER-2 amplified breast cancer resistant to transtuzumab-containing therapy: (E1104) a trial of the Eastern Cooperative Oncology Group. Cancer Res. 2009;69(24 Suppl):5084.

 Google Scholar