z Biological Chemistry & Chemical Biology Design, Synthesis and Characterisation of Novel Phenothiazine-Based Triazolopyridine Derivatives: Evaluation of Anti-Breast Cancer Activity on Human Breast Carcinoma Tanisha Sachdeva, [a] May Lee Low, [b] Chun-Wai Mai, [b, c] Siew Lee Cheong, [b] Yun Khoon Liew, [d] and Marilyn Daisy Milton* [a] A series of novel phenothiazine based [1,2,4]triazolo[4, 3-a] pyridine scaffolds were designed and synthesized in good yields by the oxidative cyclisation of phenothiazine pyridylhy- drazones. Biological responses of all compounds toward a panel of human breast cancer cells (MDA-MB-231, MDA-MB- 468, MCF7, SKBR3 and T47D) and human non-tumorigenic epithelial breast cells (MCF10 A) were evaluated. Structure- activity relationship revealed that compound with pendant phenyl ring on phenothiazine exhibited significant cytotoxic activity and apoptotic induction effects against breast cancer cell line with IC 50 value 10.2 to 17.6 μM. Notably, the cytotoxic effect was 3.5 fold higher on cancer than non-cancer cells, indicating potential control of breast cancer with lower side effects. Molecular docking studies confirmed the presence of hydrophobic contacts between appended phenyl ring, triazolo- pyridine and phenothiazine moieties with adjacent residues within the binding pocket of tubulin. One of the nitrogen in the triazolo ring also showed hydrogen bonding with tubulin. These tubulin interactions were also found with the taxane ring of paclitaxel. Cell cycle analysis confirmed the G2/M arrest induced by this compound on human breast cancer cells. Therefore, the potential anti-cancer, pro-apoptotic, and cell cycle arrest warrant further development of this molecule as a new class of anticancer agent. 1. Introduction Breast cancer is the most commonly diagnosed disease and second most leading cause of deaths among more than 200 types of tissue specific cancers. [1–2] In 2017, the American Cancer Society estimated 250,000 new cases of invasive breast cancer in women, an approximate 63,000 additional cases of in situ breast cancer were also reported. [3] Because of the numerous undesirable side effects of available treatment, development of drug resistance, recurrences and relapses of cancer, it is crucial to develop new chemotherapeutic agents which display less toxicity, potent activity, tissue selectivity and novel modes of action. [4–7] Now-a-days, combination of two or more pharmacophores into a single molecule is considered as a more promising approach to design and synthesize new chemotherapeutic agents. [8–10] Several phenothiazine molecules are known to possess chemotherapeutic properties (Figure 1), for instance, pheno- thiazine-sulphonamide derivatives I showed promising in-vitro anticancer activity against human breast cancer cell line (T47D) with aromatase inhibitory effect which consequently induces apoptosis. [11] A molecule based on phenothiazine and triazole II has also been reported to induce apoptosis against MCF-7 cells [a] T. Sachdeva, Prof. M. D. Milton Department of Chemistry, University of Delhi, Delhi-110007, India E-mail: mdmilton@chemistry.du.ac.in [b] Dr. M. L. Low, Dr. C.-W. Mai, Dr. S. L. Cheong Department of Pharmaceutical Chemistry, School of Pharmacy, Interna- tional Medical University, No. 126, Jalan Jalil Perkasa 19, Bukit Jalil, 57000 Kuala Lumpur, Malaysia [c] Dr. C.-W. Mai Center for Cancer and Stem Cell Research, Institute for Research, Development and Innovation, International Medical University, No. 126, Jalan Jalil Perkasa 19, Bukit Jalil 57000 Kuala Lumpur, Malaysia [d] Dr. Y. K. Liew Department of Life Sciences, School of Pharmacy, International Medical University, No. 126, Jalan Jalil Perkasa 19, Bukit Jalil, 57000 Kuala Lumpur, Malaysia Supporting information for this article is available on the WWW under https://doi.org/10.1002/slct.201903203 Figure 1. Examples of phenothiazine scaffolds showing anticancer activity against human cancer cell lines. Full Papers DOI: 10.1002/slct.201903203 12701 ChemistrySelect 2019, 4, 12701–12707 © 2019 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim