4-Hydroxytamoxifen enhances sensitivity of estrogen receptor α-positive breast cancer to docetaxel in an estrogen and ZNF423 SNP-dependent fashion
- 157 Downloads
In early stage, ERα-positive breast cancer, concurrent use of endocrine therapy and chemotherapy has not been shown to be superior to sequential use. We hypothesized that genetic biomarkers can aid in selecting patients who would benefit from chemo-endocrine therapy. Our previous studies revealed that ZNF423 is a transcription factor for BRCA1 and an intronic single nucleotide polymorphism (SNP) in ZNF423, rs9940645, determines tamoxifen response. Here, we identified mitosis-related genes that are regulated by ZNF423 which led us to investigate taxane response in a rs9940645 SNP- and tamoxifen-dependent fashion.
The Cancer Genome Atlas (TCGA) breast cancer dataset was used to identify genes correlated with ZNF423. Quantitative reverse transcription PCR, chromatin immunoprecipitation, and luciferase reporter assays were used to validate the gene regulation. We used CRISPR/Cas9 to engineer paired ZR-75-1 cells which differ only in ZNF423 rs9940645 SNP genotype to test SNP-dependent phenotypes including cell cycle and cell viability. We validated our findings in an additional two breast cancer cell lines, Hs578T-ERα and HCC1500.
Mitosis-related genes VRK1 and PBK, which encode histone H3 kinases, were experimentally validated to be regulated by ZNF423. ZNF423 knockdown decreased VRK1 and PBK expression and activity. Additionally, ZNF423 knockdown enhanced docetaxel-induced G2/M arrest and cytotoxicity through VRK1 or PBK regulation. Lastly, cells carrying the rs9940645 variant genotype had increased G2/M arrest and decreased cell viability when treated with docetaxel in combination with estradiol and 4-OH-TAM.
We identified ZNF423 regulated genes involved in the G2/M phase of the cell cycle. 4-OH-TAM sensitized ERα-positive breast cancer cells to docetaxel in a ZNF423 SNP-dependent manner. Our findings suggest that patients with rs9940645 variant genotype may benefit from concurrent tamoxifen and docetaxel. This would impact a substantial proportion of patients because this SNP has a minor allele frequency of 0.47.
KeywordsBreast cancer ZNF423 rs9940645 Chemo-endocrine therapy Single nucleotide polymorphism Precision medicine
Calmodulin-like protein 3
Clustered, regularly interspaced short palindromic repeats
Estrogen response element
Genome-wide association study
Lymphoblastoid cell line
National surgical adjuvant breast and bowel project
PDZ binding kinas
Quantitative reverse transcription polymerase chain reaction
Selective estrogen receptor modulator
Single nucleotide polymorphism
The Cancer Genome Atlas
Vaccinia-related kinase 1
Zinc finger protein 423
We would like to acknowledge Thomas Spelsberg, Ph.D. for providing the Hs578T-ERα cell line.
GW, SQ, and JZ participated in data acquisition, data analysis, and manuscript writing. GW designed the study and drafted the manuscript. SQ generated the ZR75-1 CRISPR Cas9 genome edited cell line. ML carried out the ZNF423 and 4-OH-TAM screens. JNI provided invaluable clinical expertise and assisted in manuscript preparation. RMW conceived the study, participated in the study design, and provided guidance in data interpretation. KS provided clinical advice and helped apply for the ChuYing Charity Foundation support. LW conceived the study, participated in the study design, coordinated the study, and is responsible for all data as described. All authors approved the final manuscript.
This work was supported by The Breast Cancer Research Foundation (BCRF-18-076) and Eisenberg Foundation. GW was supported by ChuYing Charity Foundation. JZ was supported by the Mayo Clinic Medical Scientist Training Program (T32 GM065841) and Initiative for Maximizing Student Development (R25 GM055252).
Compliance with ethical standards
Conflict of interest
ML is currently affiliated with AbbVie and, however, was not at the time of involvement in this study. LW and RMW are co-founders and stockholders in OneOme, LLC, a pharmacogenomic decision support company.
All experiments performed in this publication comply with US laws. There were no human participants or animals used in this study.
- 1.Siegel RL, Miller KD, Jemal A (2018) Cancer statistics, 2018. CA: a cancer journal for clinicians 68 (1):7–30. https://doi.org/10.3322/caac.21442
- 3.Albain KS, Barlow WE, Ravdin PM, Farrar WB, Burton GV, Ketchel SJ, Cobau CD, Levine EG, Ingle JN, Pritchard KI, Lichter AS, Schneider DJ, Abeloff MD, Henderson IC, Muss HB, Green SJ, Lew D, Livingston RB, Martino S, Osborne CK (2009) Adjuvant chemotherapy and timing of tamoxifen in postmenopausal patients with endocrine-responsive, node-positive breast cancer: a phase 3, open-label, randomised controlled trial. Lancet 374(9707):2055–2063. https://doi.org/10.1016/s0140-6736(09)61523-3 CrossRefGoogle Scholar
- 4.Bedognetti D, Sertoli MR, Pronzato P, Del Mastro L, Venturini M, Taveggia P, Zanardi E, Siffredi G, Pastorino S, Queirolo P, Gardin G, Wang E, Monzeglio C, Boccardo F, Bruzzi P (2011) Concurrent vs sequential adjuvant chemotherapy and hormone therapy in breast cancer: a multicenter randomized phase III trial. Journal of the National Cancer Institute 103(20):1529–1539. https://doi.org/10.1093/jnci/djr351 CrossRefGoogle Scholar
- 6.Ingle JN, Liu M, Wickerham DL, Schaid DJ, Wang L, Mushiroda T, Kubo M, Costantino JP, Vogel VG, Paik S, Goetz MP, Ames MM, Jenkins GD, Batzler A, Carlson EE, Flockhart DA, Wolmark N, Nakamura Y, Weinshilboum RM (2013) Selective estrogen receptor modulators and pharmacogenomic variation in ZNF423 regulation of BRCA1 expression: individualized breast cancer prevention. Cancer Discov 3(7):812–825. https://doi.org/10.1158/2159-8290.CD-13-0038 CrossRefGoogle Scholar
- 7.Qin S, Ingle JN, Liu M, Yu J, Wickerham DL, Kubo M, Weinshilboum RM, Wang L (2017) Calmodulin-like protein 3 is an estrogen receptor alpha coregulator for gene expression and drug response in a SNP, estrogen, and SERM-dependent fashion. Breast Cancer Res 19(1):95. https://doi.org/10.1186/s13058-017-0890-x CrossRefGoogle Scholar
- 8.Zerbino DR, Achuthan P, Akanni W, Amode MR, Barrell D, Bhai J, Billis K, Cummins C, Gall A, Girón CG, Gil L, Gordon L, Haggerty L, Haskell E, Hourlier T, Izuogu OG, Janacek SH, Juettemann T, To JK, Laird MR, Lavidas I, Liu Z, Loveland JE, Maurel T, McLaren W, Moore B, Mudge J, Murphy DN, Newman V, Nuhn M, Ogeh D, Ong CK, Parker A, Patricio M, Riat HS, Schuilenburg H, Sheppard D, Sparrow H, Taylor K, Thormann A, Vullo A, Walts B, Zadissa A, Frankish A, Hunt SE, Kostadima M, Langridge N, Martin FJ, Muffato M, Perry E, Ruffier M, Staines DM, Trevanion SJ, Aken BL, Cunningham F, Yates A, Flicek P (2018) Ensembl 2018. Nucl Acids Res 46(D1):D754–D761. https://doi.org/10.1093/nar/gkx1098 CrossRefGoogle Scholar
- 9.Liu D, Ho MF, Schaid DJ, Scherer SE, Kalari K, Liu M, Biernacka J, Yee V, Evans J, Carlson E, Goetz MP, Kubo M, Wickerham DL, Wang L, Ingle JN, Weinshilboum RM (2017) Breast cancer chemoprevention pharmacogenomics: deep sequencing and functional genomics of the ZNF423 and CTSO genes. NPJ Breast Cancer 3:30. https://doi.org/10.1038/s41523-017-0036-4 CrossRefGoogle Scholar
- 10.Niu N, Qin Y, Fridley BL, Hou J, Kalari KR, Zhu M, Wu TY, Jenkins GD, Batzler A, Wang L (2010) Radiation pharmacogenomics: a genome-wide association approach to identify radiation response biomarkers using human lymphoblastoid cell lines. Genome Res 20(11):1482–1492. https://doi.org/10.1101/gr.107672.110 CrossRefGoogle Scholar
- 12.del Puerto-Nevado L, Marin-Arango JP, Fernandez-Aceñero MJ, Arroyo-Manzano D, Martinez-Useros J, Borrero-Palacios A, Rodriguez-Remirez M, Cebrian A, Gomez del Pulgar T, Cruz-Ramos M, Carames C, Lopez-Botet B, Garcia-Foncillas J (2016) Predictive value of vrk 1 and 2 for rectal adenocarcinoma response to neoadjuvant chemoradiation therapy: a retrospective observational cohort study. BMC Cancer 16:519. https://doi.org/10.1186/s12885-016-2574-9 CrossRefGoogle Scholar
- 14.Lee N, Kwon JH, Kim YB, Kim SH, Park SJ, Xu W, Jung HY, Kim KT, Wang HJ, Choi KY (2015) Vaccinia-related kinase 1 promotes hepatocellular carcinoma by controlling the levels of cell cycle regulators associated with G1/S transition. Oncotarget 6(30):30130–30148. https://doi.org/10.18632/oncotarget.4967 CrossRefGoogle Scholar
- 15.Salzano M, Vazquez-Cedeira M, Sanz-Garcia M, Valbuena A, Blanco S, Fernandez IF, Lazo PA (2014) Vaccinia-related kinase 1 (VRK1) confers resistance to DNA-damaging agents in human breast cancer by affecting DNA damage response. Oncotarget 5(7):1770–1778. https://doi.org/10.18632/oncotarget.1678 CrossRefGoogle Scholar
- 19.Ohashi T, Komatsu S, Ichikawa D, Miyamae M, Okajima W, Imamura T, Kiuchi J, Kosuga T, Konishi H, Shiozaki A, Fujiwara H, Okamoto K, Tsuda H, Otsuji E (2017) Overexpression of PBK/TOPK relates to tumour malignant potential and poor outcome of gastric carcinoma. Br J Cancer 116(2):218–226. https://doi.org/10.1038/bjc.2016.394 CrossRefGoogle Scholar
- 20.Ohashi T, Komatsu S, Ichikawa D, Miyamae M, Okajima W, Imamura T, Kiuchi J, Nishibeppu K, Kosuga T, Konishi H, Shiozaki A, Fujiwara H, Okamoto K, Tsuda H, Otsuji E (2016) Overexpression of PBK/TOPK contributes to tumor development and poor outcome of esophageal squamous cell carcinoma. Anticancer Res 36(12):6457–6466. https://doi.org/10.21873/anticanres.11244 CrossRefGoogle Scholar
- 21.Pirovano G, Ashton TM, Herbert KJ, Bryant RJ, Verrill CL, Cerundolo L, Buffa FM, Prevo R, Harrap I, Ryan AJ, Macaulay V, McKenna WG, Higgins GS (2017) TOPK modulates tumour-specific radiosensitivity and correlates with recurrence after prostate radiotherapy. Br J Cancer 117(4):503–512. https://doi.org/10.1038/bjc.2017.197 CrossRefGoogle Scholar
- 24.Nguyen PL, Taghian AG, Katz MS, Niemierko A, Abi Raad RF, Boon WL, Bellon JR, Wong JS, Smith BL, Harris JR (2008) Breast cancer subtype approximated by estrogen receptor, progesterone receptor, and HER-2 is associated with local and distant recurrence after breast-conserving therapy. J Clin Oncol 26(14):2373–2378. https://doi.org/10.1200/JCO.2007.14.4287 CrossRefGoogle Scholar
- 26.Ades F, Zardavas D, Bozovic-Spasojevic I, Pugliano L, Fumagalli D, de Azambuja E, Viale G, Sotiriou C, Piccart M (2014) Luminal B breast cancer: molecular characterization, clinical management, and future perspectives. J Clin Oncol 32(25):2794–2803. https://doi.org/10.1200/JCO.2013.54.1870 CrossRefGoogle Scholar
- 27.Casoni F, Croci L, Bosone C, D’Ambrosio R, Badaloni A, Gaudesi D, Barili V, Sarna JR, Tessarollo L, Cremona O, Hawkes R, Warming S, Consalez GG (2017) Zfp423/ZNF423 regulates cell cycle progression, the mode of cell division and the DNA-damage response in Purkinje neuron progenitors. Development 144(20):3686–3697. https://doi.org/10.1242/dev.155077 CrossRefGoogle Scholar
- 29.Payton M, Bush TL, Chung G, Ziegler B, Eden P, McElroy P, Ross S, Cee VJ, Deak HL, Hodous BL, Nguyen HN, Olivieri PR, Romero K, Schenkel LB, Bak A, Stanton M, Dussault I, Patel VF, Geuns-Meyer S, Radinsky R, Kendall RL (2010) Preclinical evaluation of AMG 900, a novel potent and highly selective pan-aurora kinase inhibitor with activity in taxane-resistant tumor cell lines. Cancer Res 70(23):9846–9854. https://doi.org/10.1158/0008-5472.CAN-10-3001 CrossRefGoogle Scholar
- 31.Ingle JN, Everson LK, Wieand HS, Martin JK, Votava HJ, Wold LE, Krook JE, Cullinan SA, Paulsen JK, Twito DI et al (1988) Randomized trial of observation versus adjuvant therapy with cyclophosphamide, fluorouracil, prednisone with or without tamoxifen following mastectomy in postmenopausal women with node-positive breast cancer. J Clin Oncol 6(9):1388–1396. https://doi.org/10.1200/jco.1918.104.22.1688 CrossRefGoogle Scholar
- 32.Ingle JN, Everson LK, Wieand HS, Cullinan SA, Wold LE, Hagen JB, Martin JK, Krook JE, Fitzgibbons RG, Foley JF et al (1989) Randomized trial to evaluate the addition of tamoxifen to cyclophosphamide, 5-fluorouracil, prednisone adjuvant therapy in premenopausal women with node-positive breast cancer. Cancer 63(7):1257–1264CrossRefGoogle Scholar