Synthesis and ribonuclease A inhibition activity of resorcinol and phloroglucinol derivatives of catechin and epicatechin: Importance of hydroxyl groups Sansa Dutta, Amit Basak * , Swagata Dasgupta * Department of Chemistry, Indian Institute of Technology, Kharagpur 721 302, India article info Article history: Received 17 May 2010 Revised 18 June 2010 Accepted 19 June 2010 Available online 25 June 2010 Keywords: Catechin Epicatechin Inhibition RNase A Angiogenin CAM assay abstract The reported ribonuclease A inhibitory activity of the green tea extracts prompted us to synthesize novel catechin/epicatechin based conjugates with resorcinol and phloroglucinol with the aim to increase the number of phenolic OH groups. These are found to be more effective inhibitors of ribonuclease A as com- pared to catechin and epicatechin thus indicating the importance of number of phenolic OH groups for the inhibition of ribonucleolytic activity. Fluorescence studies have been carried out to evaluate the bind- ing parameters. The protein–ligand docking studies are also performed to gain insight into the protein– polyphenols interactions. The epicatechin based polyphenols 1 and 2 also showed inhibition of angioge- nin-induced angiogenesis, as determined by chorioallantoic membrane (CAM) assay. Ó 2010 Elsevier Ltd. All rights reserved. 1. Introduction Replication, transcription and translation form the basis of all life processes. The genetic flow of information is controlled by DNA polymerases involved in DNA replication, RNA polymerases for RNA synthesis and RNA depolymerases (usually called ‘ribonu- cleases’) for RNA degradations. Mammalian Ribonucleases (RNas- es) are endonucleases that catalyze the degradation of RNA via a two-step transphosphorylation-hydrolytic reaction. 1,2 RNases can be cytotoxic because undesired cleavage of RNA renders its en- coded information undecipherable. They inhibit the translation processes that subsequently cause cell death by adsorbing specifi- cally to certain cells, entering the cytosol and degrading the RNA. Hence, the past few years has seen a surge in the search for low molecular weight inhibitors of ribonucleases. This has also been stimulated largely by research demonstrating that several pancre- atic RNase A homologues, including angiogenin, 3 eosinophil-de- rived neurotoxin (EDN), 4 and bovine seminal RNase A, 5 utilize their enzymatic activities for potent physiological effects. 6 The ribonucleolytic centre of RNase A consists of multiple subsites (e.g., P 1 ,B 1 and B 2 ) that bind to the phosphate, nucleobase and ri- bose components of RNA molecule, respectively. However, the cleavage of phosphodiester bond occurs at the P 1 site comprising of His 12, Lys 41 and His 119. Apart from the amino acid residues directly involved in the catalytic process, residues present at vari- ous subsites are known to play an indirect role in the catalytic mechanism. So a strategy can be adopted such that the inhibitors target these subsites other than the active site resulting in the loss of enzymatic activity. Ribonuclease inhibitors, either synthetic or natural, have been intensively sought after for therapeutic pur- poses. This also finds application in angiogenesis research. The pri- mary reason is that angiogenesis is frequently promoted by angiogenin, a pancreatic ribonuclease homologue, and inhibition of angiogenin is expected to result in suppression of growth and metastasis of solid tumours. The ribonucleolytic activity of angio- genin is essential for its angiogenic activity. It is speculated that these inhibitors may also be used in a broader aspect to inhibit en- zymes, like angiogenin, that belong to the ribonuclease superfamily. In this regard, polyphenols, once known as Vitamin P, have drawn keen interest because of possible health benefits of all types of polyphenols. Recent studies indicate that polyphenols may have antioxidant characteristics with potential health benefits 7 and that they may reduce the risk of cardiovascular disease and cancer. 8,9 The reports suggest that the green tea extracts comprising of epi- catechin (EC), epigallocatechin (EGC), epicatechin gallate (ECG) and epigallocatechin gallate (EGCG) showed inhibition of ribonuc- leolytic activity of ribonuclease A and the order of inhibition is EGCG > ECG > EGC > EC. 10,11 The earlier studies have prompted us to investigate whether increasing the number of phenolic hydroxyl groups in the form of phloroglucinol and resorcinol at C-4 in the 0968-0896/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmc.2010.06.077 * Corresponding authors. Tel.: +91 3222 283300; fax: +91 3222 282252. E-mail address: absk@chem.iitkgp.ernet.in (A. Basak). Bioorganic & Medicinal Chemistry 18 (2010) 6538–6546 Contents lists available at ScienceDirect Bioorganic & Medicinal Chemistry journal homepage: www.elsevier.com/locate/bmc