Head-to-Tail Cyclization of α-Conotoxin TxID Leads to Enhanced Stability in Serum

Year 2024, , 593 - 604, 01.06.2024

Nurcan Biçen Karakoç , Muharrem Akcan

https://doi.org/10.35378/gujs.1263833

Abstract

Peptide biomolecules have important bioactivities and hence their use in drug design and development studies has increased in recent years. Conotoxins are natural peptides that obtained from cone snail venoms and have the potential to be used for chronic pain treatment, Parkinson's disease, schizophrenia, obesity and cancer due to their effects on the nervous system. However, their use as medicines has been limited as they can be easily broken down by many proteolytic enzymes in the body. Several modification methods are used to overcome these disadvantages. Cyclization of the peptide backbone is one such method and has been used to stabilize various linear peptides. In this study, the N- and C-termini of α-conotoxin TxID with two disulfide bridges were joined using a six amino acid long GGAAGG linker peptide chain to cyclize the peptide backbone and the serum stability of the cyclized peptide was examined. The cyclic TxID peptide remained intact about 50% in human serum after 24 hours.

Keywords

Conotoxins, Peptide synthesis, Cyclization, Cyclic peptides, Serum stability

Supporting Institution

TUBITAK

Thanks

This study was supported by TUBITAK 2209-A University Students Research Projects Support Program 2018/2.

References

• [1] Craik, D.J., Fairlie, D.P., Liras, S., and Price, D., “The Future of Peptide-based Drugs”, Chemical Biology & Drug Design, 81(1): 136–147, (2013).
• [2] Reiss, S., Sieber, M., Oberle, V., Wentzel, A., Spangenberg, P., Claus, R., Kolmar, H., Lösche, W., “Inhibition of platelet aggregation by grafting RGD and KGD sequences on the structural scaffold of small disulfide-rich proteins ” ,Platelets, 17(3): 153–157, (2006).
• [3] Kadin, H., “Captopril.” In: Florey KBT-AP of DS, editor. Academic Press, 79–137, (1982).
• [4] Terlau, H., and Olivera, B.M., “Conus venoms: a rich source of novel ion channel-targeted peptides”, Physiological Reviews, 84(1): 41–68, (2004).
• [5] Shen, G.S., Layer, R.T., and McCabe, R.T., “Conopeptides: From deadly venoms to novel therapeutics”, Drug Discovery Today, 5(3): 98–106, (2000).
• [6] Alonso, D., Khalil, Z., Satkunanthan, N., and Livett, B., “Drugs From the Sea: Conotoxins as Drug Leads for Neuropathic Pain and Other Neurological Conditions”, Mini-Reviews in Medicinal Chemistry, 3(7): 785–787, (2005).
• [7] Miljanich, G.P., “Ziconotide: Neuronal Calcium Channel Blocker for Treating Severe Chronic Pain”, Current Medicinal Chemistry, 11(23): 3029–3040, (2012).
• [8] Conibear, A.C., Chaousis, S., Durek, T., Johan Rosengren, K., Craik, D.J., and Schroeder, C.I., “Approaches to the stabilization of bioactive epitopes by grafting and peptide cyclization”, Biopolymers, 106(1): 89–100, (2016).
• [9] Gopal, R., Kim, Y.J., Seo, C.H., Hahm, K.S. and Park, Y., “Reversed sequence enhances antimicrobial activity of a synthetic peptide”, Journal of Peptide Science, 17(5): 329–334, (2011).
• [10] De Araujo, A.D., Callaghan, B., Nevin, S.T., Daly, N.L., Craik, D.J., Moretta, M., Hopping, G., Christie, J.M., Adams, J.D., Alewood, F.P., “Total synthesis of the analgesic conotoxin MrVIB through selenocysteine-assisted folding”, Angewandte Chemie - International Edition, 50(29), 6527–6529, (2011).
• [11] Chhabra, S., Belgi, A., Bartels, P., Van Lierop, B.J., Robinson, S.D., Kompella, S.N., Hung, A., Callaghan, P.B., Adams, J.D., Robinson, J.A., Norton, S.R., “Dicarba analogues of α-conotoxin RgIA. Structure, stability, and activity at potential pain targets”, Journal of Medicinal Chemistry, 57(23): 9933–9944, (2014).
• [12] Gori, A., Wang, C.I.A., Harvey, P.J., Rosengren, K.J., Bhola, R.F., Gelmi, M.L. Longhi, R., Christie, J.M., Lewis, J.R., Alewood, F.P., Brust, A., “Stabilization of the cysteine-rich conotoxin MrIA by using a 1,2,3-triazole as a disulfide bond mimetic”, Angewandte Chemie - International Edition, 54(4): 1361–1364, (2015).
• [13] Gerwig, G.J., Hocking, H.G., Stöcklin, R., Kamerling, J.P., and Boelens, R., “Glycosylation of conotoxins”, Marine Drugs, 11(3): 623–642, (2013).
• [14] Blanchfield, J.T., Dutton, J.L., Hogg, R.C., Gallagher, O.P., Craik, D.J., Jones, A., Adams, J.D., Lewis, J.R., Alewood, F.P., Toth, I., “Synthesis, structure elucidation, in vitro biological activity, toxicity, and Caco-2 cell permeability of lipophilic analogues of α-conotoxin MII”, Journal of Medicinal Chemistry, 46(7): 1266–1272, (2003).
• [15] Clark, R.J., Akcan, M., Kaas, Q., Daly, N.L., and Craik, D.J., “Cyclization of conotoxins to improve their biopharmaceutical properties”, Toxicon, 59(4): 446–455, (2012).
• [16] Rubin, S., and Qvit, N., “Cyclic peptides for protein–protein interaction targets: Applications to human disease”, Critical Reviews in Eukaryotic Gene Expression, 26(3): 199–221, (2016).
• [17] Borel, J.F., Feurer, C., Gubler, H.U., and Stähelin, H., “Biological effects of cyclosporin A: A new antilymphocytic agent”, Agents and Actions, 6(4): 468–475, (1976).
• [18] Cebrián, R., Rodríguez-Cabezas, M.E., Martín-Escolano, R., Rubiño, S., Garrido-Barros, M., Montalbán-López, M., Rosales, J.M., Sánchez-Moreno, M., Valdivia, E., Martínez-Bueno, M., Marín, C., Gálvez, J., Maqueda, M., “Preclinical studies of toxicity and safety of the AS-48 bacteriocin.” Journal of Advanced Research, 20: 20129–20139, (2019).
• [19] Colgrave, M.L., Korsinczky, M.J.L., Clark, R.J., Foley, F., and Craik, D.J., “Sunflower trypsin inhibitor-1, proteolytic studies on a trypsin inhibitor peptide and its analogs”, Biopolymers, 94(5): 665–672, (2010).
• [20] Craik, D.J., Čemažar, M., and Daly, N.L., “The chemistry and biology of cyclotides”, Current Opinion in Drug Discovery and Development, 10(2): 176–184, (2007).
• [21] Wang, C.K., and Craik, D.J., “Designing macrocyclic disulfide-rich peptides for biotechnological applications perspective”, Nature Chemical Biology, 14(5): 417–427, (2018).
• [22] Wu, X., Huang, Y.H., Kaas, Q., and Craik, D.J., “Cyclisation of Disulfide-Rich Conotoxins in Drug Design Applications”, European Journal of Organic Chemistry, 2016(21): 3462–3472, (2016).
• [23] Clark, R.J., Fischer, H., Dempster, L., Daly, N.L., Rosengren, K.J., Nevin, S.T., Meunier, A. F., Adams, J. D., Craik, J. D., “Engineering stable peptide toxins by means of backbone cyclization: Stabilization of the α-conotoxin MII”, Proceedings of the National Academy of Sciences of the United States of America, 102(39): 13767–13772, (2005).
• [24] Akcan, M., Stroud, M.R., Hansen, S.J., Clark, R.J., Daly, N.L., Craik, D.J., Olson M.J., “Chemical re-engineering of chlorotoxin improves bioconjugation properties for tumor imaging and targeted therapy”, Journal of Medicinal Chemistry, 54(3): 782–787, (2011).
• [25] Armishaw, C.J., Jensen, A.A., Balle, L.D., Scott, K.C.M., Sørensen, L., and Strømgaard, K., “Improving the stability of α-conotoxin AuIB through N-to-C cyclization: The effect of linker length on stability and activity at nicotinic acetylcholine receptors”, Antioxidants and Redox Signaling, 14(1): 65–76, (2011).
• [26] Clark, R.J., Jensen, J., Nevin, S.T., Callaghan, B.P., Adams, D.J., and Craik, D.J., “The engineering of an orally active conotoxin for the treatment of neuropathic pain”, Angewandte Chemie - International Edition, 37(122): 6695-6698. (2010).
• [27] Li, X., Wang, S., Zhu, X., Zhangsun, D., Wu, Y., and Luo, S., “Effects of Cyclization on Activity and Stability of α-Conotoxin TxIB”, Marine Drugs, 18(4): 180-190. (2020).
• [28] Hemu, X., Taichi, M., Qiu, Y., Liu, D.X., and Tam, J.P., “Biomimetic synthesis of cyclic peptides using novel thioester surrogates”, Biopolymers, 100(5): 492–501, (2013).
• [29] Hemu, X., and Tam, J.P., “Macrocyclic Antimicrobial Peptides Engineered from ω-Conotoxin”, Current Pharmaceutical Design, 23(14): 2131–2138, (2017).
• [30] Dutton, J., and Craik, D., “Alpha Conotoxins Nicotinic Acetylcholine Receptor Antagonists as Pharmacological Tools and Potential Drug Leads”, Current Medicinal Chemistry, 8(4): 327–344, (2012).
• [31] Luo, S., Zhangsun, D., Zhu, X., Wu, Y., Hu, Y., Christensen, S., Harvey, J.P., Akcan, M., Craik, J.D., McIntosh, J.M., “Characterization of a novel α-conotoxin TxID from Conus textile that potently blocks rat α3β4 nicotinic acetylcholine receptors”, Journal of Medicinal Chemistry, 56(23): 9655–9663, (2013).
• [32] Yu, J., Zhu, X., Harvey, P.J., Kaas, Q., Zhangsun, D., Craik, D.J., Luo, S., “Single Amino Acid Substitution in α-Conotoxin TxID Reveals a Specific α3β4 Nicotinic Acetylcholine Receptor Antagonist”, Journal of Medicinal Chemistry, 61(20): 9256–9265, (2018).
• [33] Wu, Y., Zhangsun, D., Zhu, X., Kaas, Q., Zhangsun, M., Harvey, P.J., Craik, J.D., McIntosh, J.M., Luo, S., “α-Conotoxin [S9A]TxID Potently Discriminates between α3β4 and α6/α3β4 Nicotinic Acetylcholine Receptors”, Journal of Medicinal Chemistry, 60(13): 5826–5833, (2017).
• [34] Zhao, W., Xiong, Y., Zhangsun, D., Luo, S., “DSPE-PEG modification of α-conotoxin TXID”, Marine Drugs, 17(6): 342-352, (2019).
• [35] Wang, S., Ren, J., Li, R., Li, X., Zhangsun, D., Wu, Y., Luo, S., “Synthesis and evaluation of disulfide-rich cyclic α-conotoxin [S9A]TxID analogues as novel α3β4 nAChR antagonists”, Bioorganic Chemistry, 112: 104875-104881, (2021).
• [36] Koradi, R., Billeter, M., and Wüthrich, K., “MOLMOL: A program for display and analysis of macromolecular structures”, Journal of Molecular Graphics, 14(1): 51–55, (1996).
• [37] Cheneval, O., Schroeder, C.I., Durek, T., Walsh, P., Huang, Y.H., Liras, S., Price, A.D., Craik J.D., “Fmoc-based synthesis of disulfide-rich cyclic peptides”, Journal of Organic Chemistry, 79(12): 5538–5544, (2014).
• [38] Malesevic, M., Strijowski, U., Bachle, D., and Sewald, N., “An improved method for the solution cyclization of peptides under pseudo-high dilution conditions”, Journal of Biotechnology, 112(1–2): 73–77, (2004).
• [39] Wu, Y., Wu, X., Yu, J., Zhu, X., Zhangsun, D., and Luo, S., “Influence of disulfide connectivity on structure and bioactivity of α-conotoxin TxIA”, Molecules, 19(1): 966–979, (2014).
• [40] Bulaj, G., and Olivera, B.M., “Folding of conotoxins: Formation of the native disulfide bridges during chemical synthesis and biosynthesis of Conus peptides”, Antioxidants and Redox Signaling, 10(1): 141–155, (2008).
• [41] Jin, A.H., Brandstaetter, H., Nevin, S.T., Tan, C., Clark, R.J., Adams, D.J., Alewood, P.F., Craik, J.D., Daly, L.N., “Structure of α-conotoxin BuIA: Influences of disulfide connectivity on structural dynamics”, BMC Structural Biology, 7: 1-13, (2007).
• [42] Nielsen, J.S., Buczek, P., and Bulaj, G., “Cosolvent-assisted oxidative folding of a bicyclic α-conotoxin ImI”, Journal of Peptide Science, 10(5): 249–256, (2004).
• [43] Halai, R., Callaghan, B., Daly, N.L., Clark, R.J., Adams, D.J., and Craik, D.J., “Effects of cyclization on stability, structure, and activity of α-conotoxin RgIA at the α9α10 nicotinic acetylcholine receptor and GABA B receptor”, Journal of Medicinal Chemistry, 54(19): 6984–6992, (2011).
• [44] Carstens, B.B., Swedberg, J., Berecki, G., Adams, D.J., Craik, D.J., and Clark, R.J., “Effects of linker sequence modifications on the structure, stability, and biological activity of a cyclic α-conotoxin”, Biopolymers, 106(6): 864–875, (2016).