{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,10,12]],"date-time":"2025-10-12T02:20:59Z","timestamp":1760235659180,"version":"build-2065373602"},"reference-count":25,"publisher":"MDPI AG","issue":"18","license":[{"start":{"date-parts":[[2021,9,17]],"date-time":"2021-09-17T00:00:00Z","timestamp":1631836800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Melanoma is the most lethal form of skin cancer, which is intrinsically resistant to conventional chemotherapy. Combination therapy has been developed to overcome this challenge and show synergistic anticancer effects on melanoma. Notably, the histone deacetylase inhibitor, valproic acid (VPA), has been indicated as a potential sensitizer of chemotherapy drugs on various metastatic cancers, including advanced melanoma. In this study, we explored whether VPA could serve as an effective sensitizer of chemotherapy drug etoposide (ETO) on B16-F10 and SK-MEL-2-Luc melanoma cell lines in response to drug-induced DNA damages. Our results demonstrated that the VPA-ETO simultaneous combined treatment and ETO pretreated sequential combined treatment generated higher inhibitory effectivities than the individual treatment of each drug. We found the VPA-ETO simultaneous combined treatment contributed to the synergistic inhibitory effect by the augmented DNA double-strand breaks, accompanied by a compromised homologous recombination activity. In comparison, the ETO pretreated sequential combined treatment led to synergistic inhibitory effect via enhanced apoptosis. Surprisingly, the enhanced homologous recombination activity and G2\/M phase arrest resulted in the antagonistic effect in both cells under VPA pretreated sequential combined treatment. In summary, our findings suggested that sequential order and effective dose of drug administration in VPA-ETO combination therapy could induce different cellular responses in melanoma cells. Such understanding might help potentiate the effectiveness of melanoma treatment and highlight the importance of sequential order and effective dose in combination therapy.<\/jats:p>","DOI":"10.3390\/ijms221810029","type":"journal-article","created":{"date-parts":[[2021,9,17]],"date-time":"2021-09-17T21:23:29Z","timestamp":1631913809000},"page":"10029","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Synergistic Effect of Simultaneous versus Sequential Combined Treatment of Histone Deacetylase Inhibitor Valproic Acid with Etoposide on Melanoma Cells"],"prefix":"10.3390","volume":"22","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3338-2020","authenticated-orcid":false,"given":"Yueh-Ming","family":"Shyu","sequence":"first","affiliation":[{"name":"Institute of Bioinformatics and Structural Biology, National Tsing Hua University, Hsinchu 30013, Taiwan"},{"name":"Department of Medical Science, National Tsing Hua University, Hsinchu 30013, Taiwan"},{"name":"Department of Internal Medicine, Division of Cardiology, Hsinchu MacKay Memorial Hospital, Hsinchu 30071, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2101-9574","authenticated-orcid":false,"given":"Lawrence Yu-Min","family":"Liu","sequence":"additional","affiliation":[{"name":"Department of Internal Medicine, Division of Cardiology, Hsinchu MacKay Memorial Hospital, Hsinchu 30071, Taiwan"},{"name":"Department of Medicine, MacKay Medical College, New Taipei City 25245, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-6022-0107","authenticated-orcid":false,"given":"Yung-Jen","family":"Chuang","sequence":"additional","affiliation":[{"name":"Institute of Bioinformatics and Structural Biology, National Tsing Hua University, Hsinchu 30013, Taiwan"},{"name":"Department of Medical Science, National Tsing Hua University, Hsinchu 30013, Taiwan"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2021,9,17]]},"reference":[{"key":"ref_1","first-page":"1071","article-title":"Cutaneous melanoma: From pathogenesis to therapy","volume":"52","author":"Leonardi","year":"2018","journal-title":"Int. J. Oncol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"525","DOI":"10.1016\/j.det.2012.04.011","article-title":"Radiation therapy for cutaneous melanoma","volume":"30","author":"Barker","year":"2012","journal-title":"Derm. Clin."},{"key":"ref_3","doi-asserted-by":"crossref","first-page":"553","DOI":"10.2174\/156652411800615153","article-title":"Resistance to chemotherapy and molecularly targeted therapies: Rationale for combination therapy in malignant melanoma","volume":"11","author":"Wu","year":"2011","journal-title":"Curr. Mol. Med."},{"key":"ref_4","doi-asserted-by":"crossref","first-page":"291","DOI":"10.1038\/499291a","article-title":"Cancer: Calculated treatment","volume":"499","author":"Komarova","year":"2013","journal-title":"Nature"},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"38022","DOI":"10.18632\/oncotarget.16723","article-title":"Combination therapy in combating cancer","volume":"8","author":"Mokhtari","year":"2017","journal-title":"Oncotarget"},{"key":"ref_6","unstructured":"Terrie, Y. (2021, May 26). Monitoring Combinationdrug Therapy. Available online: https:\/\/www.pharmacytimes.com\/publications\/issue\/2010\/january2010\/rxfocuscombination-0110."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"185","DOI":"10.4161\/cbt.6.2.3578","article-title":"Valproic acid induces apoptosis, p16INK4A upregulation and sensitization to chemotherapy in human melanoma cells","volume":"6","author":"Valentini","year":"2007","journal-title":"Cancer Biol."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"1","DOI":"10.1097\/00001813-200201000-00001","article-title":"Histone deacetylase inhibitors in cancer treatment","volume":"13","author":"Vigushin","year":"2002","journal-title":"Anticancer Drugs"},{"key":"ref_9","doi-asserted-by":"crossref","first-page":"2479","DOI":"10.1158\/1078-0432.CCR-08-1931","article-title":"Potentiation of a topoisomerase I inhibitor, karenitecin, by the histone deacetylase inhibitor valproic acid in melanoma: Translational and phase I\/II clinical trial","volume":"15","author":"Daud","year":"2009","journal-title":"Clin. Cancer Res."},{"key":"ref_10","doi-asserted-by":"crossref","unstructured":"Kalal, B.S., Pai, V.R., Behera, S.K., and Somashekarappa, H.M. (2019). HDAC2 inhibitor valproic acid increases radiation sensitivity of drug-resistant melanoma cells. Med. Sci., 7.","DOI":"10.3390\/medsci7030051"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"704","DOI":"10.1080\/07357900802653480","article-title":"Cytotoxicity and mitochondrial apoptosis induced by etoposide in melanoma cells","volume":"27","author":"Rudolf","year":"2009","journal-title":"Cancer Investig."},{"key":"ref_12","doi-asserted-by":"crossref","unstructured":"Eckschlager, T., Plch, J., Stiborova, M., and Hrabeta, J. (2017). Histone deacetylase inhibitors as anticancer drugs. Int. J. Mol. Sci., 18.","DOI":"10.3390\/ijms18071414"},{"key":"ref_13","doi-asserted-by":"crossref","first-page":"159","DOI":"10.1007\/s11060-007-9402-7","article-title":"Valproic acid induces p21 and topoisomerase-II (alpha\/beta) expression and synergistically enhances etoposide cytotoxicity in human glioblastoma cell lines","volume":"85","author":"Das","year":"2007","journal-title":"J. Neuro-Oncol."},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"28","DOI":"10.1038\/sj.bjc.6604817","article-title":"A phase I-II study of the histone deacetylase inhibitor valproic acid plus chemoimmunotherapy in patients with advanced melanoma","volume":"100","author":"Rocca","year":"2009","journal-title":"Br. J. Cancer"},{"key":"ref_15","unstructured":"(2021, May 26). GraphPad Prism 8 Curve Fitting Guide\u2014Equation: Two Phase Association. Available online: https:\/\/www.graphpad.com\/guides\/prism\/8\/curve-fitting\/reg_exponential_association_2phase.htm."},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"27","DOI":"10.1016\/0065-2571(84)90007-4","article-title":"Quantitative analysis of dose-effect relationships: The combined effects of multiple drugs or enzyme inhibitors","volume":"22","author":"Chou","year":"1984","journal-title":"Adv. Enzym. Regul."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"621","DOI":"10.1124\/pr.58.3.10","article-title":"Theoretical basis, experimental design, and computerized simulation of synergism and antagonism in drug combination studies","volume":"58","author":"Chou","year":"2006","journal-title":"Pharm. Rev."},{"key":"ref_18","first-page":"95","article-title":"Molecular mechanisms of etoposide","volume":"14","author":"Montecucco","year":"2015","journal-title":"EXCLI J."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"1803","DOI":"10.1038\/sj.onc.1204252","article-title":"Regulation of the G2\/M transition by p53","volume":"20","author":"Taylor","year":"2001","journal-title":"Oncogene"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"209","DOI":"10.1016\/S0099-5428(05)32008-9","article-title":"Valproic Acid and sodium valproate: Comprehensive profile","volume":"32","author":"Alsarra","year":"2005","journal-title":"Profiles Drug Subst. Excip. Relat. Methodol."},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"9","DOI":"10.1186\/2041-9414-3-9","article-title":"Pathway choice in DNA double strand break repair: Observations of a balancing act","volume":"3","author":"Brandsma","year":"2012","journal-title":"Genome Integr."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"3067","DOI":"10.1158\/0008-5472.CAN-15-2680","article-title":"Enhanced histone deacetylase activity in malignant melanoma provokes RAD51 and FANCD2-triggered drug resistance","volume":"76","author":"Krumm","year":"2016","journal-title":"Cancer Res."},{"key":"ref_23","first-page":"607","article-title":"The same, only different\u2014DNA damage checkpoints and their reversal throughout the cell cycle","volume":"128","author":"Shaltiel","year":"2015","journal-title":"J. Cell Sci."},{"key":"ref_24","first-page":"61","article-title":"Flow cytometry measurement of DNA fragmentation in the course of cell death via apoptosis. New techniques for evaluation of DNA status for the pathologist","volume":"17","author":"Lizard","year":"1997","journal-title":"Ann. Pathol."},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"6661","DOI":"10.21873\/anticanres.13881","article-title":"Analysis of DNA Damage Responses After Boric Acid-mediated Boron Neutron Capture Therapy in Hepatocellular Carcinoma","volume":"39","author":"Chen","year":"2019","journal-title":"Anticancer Res."}],"container-title":["International Journal of Molecular Sciences"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/1422-0067\/22\/18\/10029\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,11]],"date-time":"2025-10-11T07:01:12Z","timestamp":1760166072000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/1422-0067\/22\/18\/10029"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2021,9,17]]},"references-count":25,"journal-issue":{"issue":"18","published-online":{"date-parts":[[2021,9]]}},"alternative-id":["ijms221810029"],"URL":"https:\/\/doi.org\/10.3390\/ijms221810029","relation":{},"ISSN":["1422-0067"],"issn-type":[{"type":"electronic","value":"1422-0067"}],"subject":[],"published":{"date-parts":[[2021,9,17]]}}}