Citation: Araki, T.; Hamada, K.; Myat, A.B.; Ogino, H.; Hayashi, K.; Maeda, M.; Tong, Y.; Murakami,Y.; Nakao, K.; Masutani, M. Enhanced Cytotoxicity on Cancer Cells by Combinational Treatment of PARP Inhibitor and 5-Azadeoxycytidine Accompanying Distinct Transcriptional Proﬁles. Cancers 2022, 14, 4171. https://doi.org/10.3390/ cancers14174171 Academic Editor: Angela Celetti Received: 5 June 2022 Accepted: 24 August 2022 Published: 28 August 2022 Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional afﬁl- iations. Copyright: © 2022 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/). cancers Article Enhanced Cytotoxicity on Cancer Cells by Combinational Treatment of PARP Inhibitor and 5-Azadeoxycytidine Accompanying Distinct Transcriptional Proﬁles Tomonori Araki 1,2,† , Kensuke Hamada 3,4,5,†,‡ , Aung Bhone Myat 1 , Hideki Ogino 3,‡ , Kohei Hayashi 1,2 , Miho Maeda 3,4,‡ , Ying Tong 1 , Yasufumi Murakami 4 , Kazuhiko Nakao 2 and Mitsuko Masutani 1,3, * ,‡ 1 Department of Molecular and Genomic Biomedicine, Center for Bioinformatics and Molecular Medicine, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523, Japan 2 Department of Gastroenterology and Hepatology, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8501, Japan 3 Biochemistry Division, National Cancer Center Research Institute, Tokyo 104-0045, Japan 4 Departmentof Biological Science and Technology, Faculty of Industrial Science and Technology, Tokyo University of Science, Tokyo 162-8601, Japan 5 Medical Research Institute, Tokyo Medical and Dental University, Chiyoda-ku, Tokyo 113-8510, Japan * Correspondence: mmasutan@nagasaki-u.ac.jp; Tel.: +81-95-819-8502 † These authors contributed equally to this work. ‡ Previous afﬁliation. Simple Summary: We investigated the effect of combinational use of PARP inhibitors on cytotoxicity of 5-aza-dC in human cancer cell lines. The combinational treatment of 5-aza-dC and PARP inhibitor PJ-34 exhibited a stronger cytotoxicity compared with their treatment alone in blood cancer HL-60, U937, and colon cancer HCT116 and RKO cells. In microarray analysis, combinational treatment with PJ-34 and 5-aza-dC caused different broad changes in gene expression proﬁles compared with their single treatments in both HCT116 and RKO cells. The proﬁles of reactivation of silenced genes were also different in combination of PJ-34 and 5-aza-dC and their single treatments. The results suggest that a combination of 5-aza-dC and PARP inhibitor may be useful by inducing distinct transcriptional proﬁle changes. Abstract: Poly(ADP-ribose) polymerase (PARP) is involved in DNA repair and chromatin regulation. 5-Aza-2 ′ -deoxycytidine (5-aza-dC) inhibits DNA methyltransferases, induces hypomethylation, blocks DNA replication, and causes DNA single strand breaks (SSBs). As the PARP inhibitor is expected to affect both DNA repair and transcriptional regulations, we investigated the effect of combinational use of PARP inhibitors on cytotoxicity of 5-aza-dC in human cancer cell lines. The combinational treatment of 5-aza-dC and PARP inhibitor PJ-34 exhibited a stronger cytotoxicity compared with their treatment alone in blood cancer HL-60, U937, and colon cancer HCT116 and RKO cells. Treatment with 5-aza-dC but not PJ-34 caused SSBs in HCT116 cell lines. Global genome DNA demethylation was observed after treatment with 5-aza-dC but not with PJ-34. Notably, in microarray analysis, combinational treatment with PJ-34 and 5-aza-dC caused dissimilar broad changes in gene expression proﬁles compared with their single treatments in both HCT116 and RKO cells. The proﬁles of reactivation of silenced genes were also different in combination of PJ-34 and 5-aza-dC and their single treatments. The results suggest that the combinational use of 5-aza-dC and PARP inhibitor may be useful by causing distinct transcriptional proﬁle changes. Keywords: PARP inhibitor; 5-aza-2 ′ -deoxycytidine; microarray; transcription; DNA methylation 1. Introduction DNA damage, genomic, and epigenomic instability induce the activation of oncogenes and the inactivation of tumor suppressor genes [1]. This results in malignant transformation Cancers 2022, 14, 4171. https://doi.org/10.3390/cancers14174171 https://www.mdpi.com/journal/cancers