Synthesis and evaluation of 3-salicyloylpyridine derivatives as cytotoxic mitochondrial apoptosis inducers Alisha Sood a , Vishal Sharma a , Ashun Chaudhry b , Rakesh Kumar b , Saroj Arora b , Rajnikant c , Vivek Gupta c , Mohan Paul S. Ishar a,⇑ a Bio-Organic and Photochemistry Laboratory, Department of Pharmaceutical Sciences, Guru Nanak Dev University, Amritsar 143005, Punjab, India b Department of Botanical and Environmental Sciences, Guru Nanak Dev University, Amritsar 143005, Punjab, India c Post-Graduate Department of Physics, University of Jammu, Jammu Tawi, 180 006, India article info Article history: Received 16 April 2014 Revised 17 July 2014 Accepted 5 August 2014 Available online 12 August 2014 Keywords: 3-Formylchromone 3-Salicyloylpyridines Cytotoxic activity Apoptosis abstract A series of novel 3-salicyloylpyridines (4a–h) were synthesized with good yield by modified Knoevena- gel–Stobbel method; o-allylation with allyl bromide lead to formation of compounds (5a–h). The synthe- sized compounds were characterized by spectroscopic techniques and evaluated for cytotoxic activity against human cancer cell lines. Compounds bearing hydroxyl group displayed high cytotoxicity (4a–h) as compared to o-allylated molecules (5a–h). The most active compound 4b was selected for further investigation to look for mechanism of cell death in prostate cancer (PC-3) cells. The apoptotic bodies induced by 4b in PC-3 cells were scanned by confocal microscopy and confirmed by scanning electron microscopy (SEM). Further results obtained from spectrofluorimetric determination of mitochondrial membrane potential (DWm) and intracellular reactive oxygen species (ROS) in treated PC-3 cells revealed that mitochondria dependent apoptosis was involved in the cell death. Ó 2014 Elsevier Ltd. All rights reserved. Cancer is a mortiferous disease characterized by unregulated proliferation of cells leading to tremendous health costs with asso- ciated high level of mortality and morbidity. 1 Despite the availabil- ity of many effective anticancer agents, still chemotherapy is placed in the third line for the treatment of cancer due to severe side effects. Therefore, search for novel anticancer drugs without any side effects is the major goal for the drug designing program of medicinal chemists. Pyridine is a six member nitrogen contain- ing heterocyclic compound and its derivatives exhibit remarkable biological activities such as antiviral, 2 anticancer, 3 antimicrobial, 4 antidiabetic, 5 antidote, 6 antileishmanial, 7 antioxidant, 8 anticoagu- lant, 9 antitubercular 10 etc. In the biological system, pyridine is present as basic nucleus in important vitamins like niacin (vitamin B 3 ) and pyridoxine (vitamin B 6 ), and it also exist as pharmacophore of over 7000 existing drugs with a variety of biological activities. 11 Camptothecin analogues having pyridine nucleus such as topotec- an and irinotecan are used clinically as anticancer drugs, which act as an effective poison of DNA topoisomerase I (TopI) by forming a ternary complex with TopI-DNA. 12 Recently, we have reported cytotoxic pyridine based isoxazolidines as highly active against ovarian (IGROV-1), breast (MCF-7) and human glioblastoma (SF-295) cancer cells. 13 According to literature reports, pyridine based compounds have been found to possess anticancer activity through varied mechanisms such as blocking the activity of enzyme CHK1 protein kinase, 14 selective inhibition of telomerase, 15 inhibition of topoisomerase II, 16 inhibition of tyrosine kinases, 17 and inhibition of microtubule assembly. 18 Taking cognizance of promising anticancer activity of pyridine based molecules, it was decided to synthesize some novel pyridine analogues and evaluates their cytotoxic potential against various human cancer cell lines. A variety of approaches have been employed for the con- struction of pyridine nucleus. 19–23 In the present study, modified Knoevenagel–Stobbel method was utilized for the construction of substituted pyridines. 24 Substituted pyrano[4,3-b][1]benzopyrans (3a–h) were synthe- sized by the hetero Diels–Alder reaction [4+2] of substituted 3- formylchromones (1a–h) with excess of ethyl vinyl ether (2) in dichloromethane at room temperature, 25,26 which on reaction with ammonium acetate in ethanol, under refluxing conditions for an hour, afforded 3-salicyloylpyridines (4a–h, Scheme 1, Table 1). Lit- erature reports suggested that o-allylated/prenylated compounds have shown promising bioactivities, 27 therefore, taking into con- sideration the substituted 3-salicyloylpyridines (4a–h) were ally- lated with allyl bromide in the presence of fused potassium carbonate to obtain 3-(2 0 -allyloxybenzoyl)pyridines (5a–h, Table 1). After completion of reactions (Tlc), the residues obtained http://dx.doi.org/10.1016/j.bmcl.2014.08.010 0960-894X/Ó 2014 Elsevier Ltd. All rights reserved. ⇑ Corresponding author. Tel.: +91 1832258802 09x3321; fax: +91 1832258820. E-mail address: mpsishar@yahoo.com (M.P.S. Ishar). Bioorganic & Medicinal Chemistry Letters 24 (2014) 4724–4728 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl