DOI: 10.1002/cbic.201200081 Cardiotoxin-I: An Unexpectedly Potent Insulinotropic Agent Thi Tuyet Nhung Nguyen, [a, b, d] Benjamin Folch, [a] Myriam LØtourneau, [a, d] David Vaudry, [c, d] Nam Hai Truong, [b] Nicolas Doucet, [a] David Chatenet,* [a, d] and Alain Fournier* [a, d] Introduction Diabetes is a condition caused by a deficiency or a diminished effectiveness of endogenous insulin secretion. As such, this en- docrine disorder, characterized by hyperglycemia and dysfunc- tional metabolism, has potential long term complications that can affect several organs, including kidneys, eyes, heart, blood vessels, and nerves. [1] Various hormonal and neural stimuli can trigger insulin secretion through the activation of G protein- coupled receptors, for example, the glucagon-like peptide- 1 (GLP-1) receptors, [2] but also through modulation of ion channel activity. [3] Insulinotropic toxins are potent, naturally occurring com- pounds of animal or plant origin that stimulate insulin secre- tion from pancreatic b-cells, a subset of endocrine islet cells that act as blood glucose sensors. [4] Several compounds with therapeutic potential have been isolated from nature, particu- larly in animal venoms that represent a rich source of bioactive molecules such as peptides, proteins, and enzymes. [5] As pri- mary molecular targets of venom include receptors, ion chan- nels, and enzymes, and because survival of the animal depends on the potency and efficacy of its venom, high-affinity bio- active molecules could be isolated from this source. [6] Thus, toxins or molecules isolated from venoms represent valuable tools for the design of highly selective drugs that might be useful in the treatment of specific pathologies. Venomous organisms synthesize a great diversity of toxic molecules that can act on G protein-coupled receptors or affect membrane excitability. Several compounds isolated from natural sources have been investigated for their potential as novel pharmacological/therapeutic agents in the treatment of type 2 diabetes due to their insulin-releasing and b-cell growth-promoting properties. For example, Mastoparan, a tet- radecapeptide from wasp venom, was shown to stimulate in- sulin secretion from b-TC3 and INS-1 cells via direct activation of Rho protein. [7] TsTx-V, a sodium channel agonist isolated from scorpion venom, potentiated glucose-induced insulin release from isolated islets of Langerhans by enhancing b-cell membrane depolarization and increasing the relative duration of electrical activity during the active phase. [8] Guangxitoxin, a toxin isolated from spider venom, is a potent inhibitor of Kv2.1/Kv2.2 channels and was shown to inhibit most of the delayed-rectifier K + currents that contribute to action potential repolarization, thereby modulating insulin secretion. [9] Finally, Exendin-4, isolated from Gila monster saliva, is a 39-amino acid peptide acting through GLP-1 receptors that is currently sold as a therapeutic drug, Exenatide, for the treatment of type 2 diabetes. [2] In this work, we report the isolation and characterization of a new insulinotropic peptide, identified as cardiotoxin-I (CTX-I), from the vietnamese snake Naja kaouthia. This compound, Insulin secretion from pancreatic b-cells is a complex process, involving the integration and interaction of multiple external and internal stimuli, in which glucose plays a major role. Un- derstanding the physiology leading to insulin release is a crucial step toward the identification of new targets. In this study, we evaluated the presence of insulinotropic metabolites in Naja kaouthia snake venom. Only one fraction, identified as cardio- toxin-I (CTX-I) was able to induce insulin secretion from INS-1E cells without affecting cell viability and integrity, as assessed by MTT and LDH assays. Interestingly, CTX-I was also able to stimulate insulin secretion from INS-1E cells even in the ab- sence of glucose. Although cardiotoxins have been character- ized as potent hemolytic agents and vasoconstrictors, CTX-I was unable to induce direct hemolysis of human erythrocytes or to induce potent vasoconstriction. As such, this newly iden- tified insulin-releasing toxin will surely enrich the pool of exist- ing tools to study b-cell physiology or even open a new thera- peutic avenue. [a] T. T. N. Nguyen, Dr. B. Folch, M. LØtourneau, Prof. N. Doucet, Dr. D. Chatenet, Prof. A. Fournier INRS–Institut Armand-Frappier, UniversitØ du QuØbec 531 Boulevard des Prairies, Ville de Laval, QuØbec, H7V 1B7 (Canada) E-mail : david.chatenet@iaf.inrs.ca alain.fournier@iaf.inrs.ca [b] T. T.N. Nguyen, Prof. N. H. Truong Institute of Biotechnology 18 Hoang Quoc Viet, Caugiay, Hanoi (Vietnam) [c] Dr. D. Vaudry Neuronales & Neuroendocrines, IFRMP 23, UniversitØ de Rouen 76821 Mont-Saint-Aignan (France) [d] T. T.N. Nguyen, M. LØtourneau, Dr. D. Vaudry, Dr. D. Chatenet, Prof. A. Fournier Laboratoire International AssociØ Samuel de Champlain (INSERM—INRS—UniversitØ de Rouen) Supporting information for this article is available on the WWW under http ://dx.doi.org/10.1002/cbic.201200081. ChemBioChem 0000, 00, 1 – 9  2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim &1& These are not the final page numbers! ÞÞ