Pretreatment with Huperzine A-Loaded Poly(lactide-co-glycolide) Nanoparticles Protects against Lethal Effects of Soman-Induced in Mice

Abstract:

Huperzine A (HupA), an alkaloid isolated from the Chinese club moss, is a reversible inhibitor of cholinesterases which cross the blood-brain barrier and show high specificity for acetylcholinesterase (AChE). However, HupA induces unwanted side effects in an effective dose against nerve agent poisoning. In the present study, HupA–loaded poly (lactide-co-glycolide) nanoparticles (HupA-PLGA-NP) were prepared using the O/W emulsion solvent evaporation method. The results of SEM demonstrated that HupA-PLGA-NP had an spherical shape and a smooth surface without pores. It’s mean diameter and PDI were 208.5±3.6nm and 0.09±0.01 respectively. The Zeta potential was-35.3±1.8mV and the drug loading was 2.86±0.6%. In vitro drug release studies showed that HupA-PLGA-NP had a sustained-release behavior in phosphate buffer solution, The accumulated amount of HupA was about 72.1% at 48h with a low burst release within 30min. The LD₅₀ values of HupA and HupA-PLGA-NP were 1.40 and 4.85mg/kg respectively, showing that the toxicity of HupA was reduced by 3.5 times. We evaluated the protective efficacy for different doses of HupA or HupA-PLGA-NP against 1.0×LD₉₅(143.0μg/kg) soman toxicity. The results confirmed that HupA (0.3~0.5mg/kg) or HupA-PLGA-NP (0.5~1.5mg/kg) could ensure animals survive. However, about 10% of the animals injected with HupA (0.8mg/kg) died, while no animals died when injected with HupA-PLGA-NP (1.5mg/kg). Aim to 100% survival rate, the effective protective time (12h) of HupA-PLGA-NP (0.5mg/kg,iv) against 1.0×LD₉₅ soman toxicity in mice was significantly prolonged compared with that of HupA (4h). The study of AChE activity showed that whole-blood and supernatant of brain diluted by 80-fold and 10-fold respectively were optimum in this study. AChE inhibition after administration of HupA and HupA-PLGA-NP (0.5mg/kg,iv) was recorded and analyzed, The peak values of AChE inhibition in whole-blood and brain by HupA-PLGA-NP (17.6% and 21.8%) were lower than those by HupA (33.7% and 31.9%) and AChE inhibition time by HupA-PLGA-NP was longer than that by HupA. These data confirmed that HupA-PLGA-NP had less toxic and more longer time than HupA against 1.0×LD₉₅ soman poisoning and warrant further development as a potent medical countermeasure against chemical warfare nerve agents (CWNAs) poisoning.

• Periodical: Key Engineering Materials (Volumes 645-646)
• Pages: 1374-1382
• DOI: https://doi.org/10.4028/www.scientific.net/KEM.645-646.1374
• Online since: May 2015
• Authors: Rui Hua Zhang, Li Qin Li*, Chen Wang, Xiao Jing Lu, Tong Shi, Jian Fu Xu, Liang Cai Song, Hui Fang Wang
• Keywords: Acetylcholinesterase, Huperzine A, Nanoparticle, Nerve Agent, Soman

References

[1] M. Nagao, T. Takatori, Y. Matsuda, M. Nakajima, H. Iwase, K. Iwadate, Definitive evidence for the acute sarin poisoning diagnosis in the Tokyo subway, Toxicol. Appl. Pharmacol. 144 (1997) 198-203.

DOI: 10.1006/taap.1997.8110

[2] N.M. Herkert, S. Schulz, T. Wille, H. Thiermann, R.A. Hatz, F. Worek, Pre- and post-treatment effect of physostigmine on soman-inhibited human erythrocyte and muscle acetylcholinesterase in vitro, Toxicol. Appl. Pharmacol. 253 (2011) 7–13.

DOI: 10.1016/j.taap.2011.03.005

[3] K. Cannard, The acute treatment of nerve agent exposure, J. Neurol. Sci. 249 (2006): 86–94.

[4] J. Bajgar, Organophosphates/nerve agent poisoning: mechanism of action, diagnosis, prophylaxis, and treatment, Adv. Clin. Chem. 38 (2004): 151–216.

DOI: 10.1016/s0065-2423(04)38006-6

[5] H.P.M. van Helden, M.J.A. Joosen, I.H.C. Philippens, Non-enzymatic pretreatment of nerve agent(soman)poisoning: A brief state-of-the-art review, Toxicol. Lett. 206 (2011)35-40.

DOI: 10.1016/j.toxlet.2011.04.021

[6] J.R. Haigh, S.R. Johnston, A. Peppernay, P.J. Mattern, G.E. Garcia, B.P. Doctor, R.K. Gordon, P.S. Aisen, Protection of red blood cell acetylcholinerterase by oral huperzineA against ex vivo soman exposure: Next generation prophylaxis and sequestering of acetylcholinerterase over butyrylcholinesterase, Chem. Bio. Interact. 175 (2008).

DOI: 10.1016/j.cbi.2008.04.033

[7] Y. Wang, Y.L. Wei, S. Oguntayo, N. Jensen, B.P. Doctor ,M.P. Nambiar, [+]-Huperzine A Protects Against Soman Toxicity in Guinea Pigs, Neurochem. Res. 36 (2011)2381–2390.

DOI: 10.1007/s11064-011-0564-5

[8] P. Masson, Evolution of and perspectives on therapeutic approaches to nerve agent poisoning. Toxicol. Lett. 206 (2011)5-13.

[9] Y.Q. Liang, X.C. Tang, Comparative effects of huperzine A, donepezil and rivastigmine on cortical acetylcholine level and acetylcholinesterase activity in rats, Neurosci. Lett. 361 (2004) 56–59.

DOI: 10.1016/j.neulet.2003.12.071

[10] Y. Ashani, J.O. Peggins, B.P. Doctor, Mechanism of inhibition of cholinesterases by HUP A, Biochem. Biophys. Res. Commun. 30 ( 1992)719-726.

DOI: 10.1016/0006-291x(92)90649-6

[11] E.X. Albuquerque, E.F.R. Pereira, Y. Aracava, W.P.F. Fawcett, M. Oliveira, W.R. Randall, Effective countermeasure against poisoning by organophosphorus insecticedes and nerve agents, Proc. Natl. Acad. Sci. 103 (2006)13220-13225.

DOI: 10.1073/pnas.0605370103

[12] G. Lallement, V. Baille, D. Baubichon, P. Carpentier, J.M. Collombet, P. Filliat, A. Foquin, Review of the value of Huperzine as pretreatment of organophosphate poisoning, Neurotoxicology. 23 (2002) 1–5.

DOI: 10.1016/s0161-813x(02)00015-3

[13] L.S. Tonduli, G. Testylier, C. Masqueliez, G. Lallement, P. Monmaur, Effects of Huperzine used as pre-treatment against soman induced seizures. Neurotoxicology. 22 (2001)29–37.

DOI: 10.1016/s0161-813x(00)00015-2

[14] B.R. Coleman, R.H. Ratcliffe, S.A. Oguntayo, X. Shi, B.P. Doctor, R.K. Gordon, M.P. Nambiar, [+]-Huperzine A treatment protects against N-methyl-D-aspartate-induced seizure/status epilepticus in rats, Chem. Bio. Interact. 175 (2008)387–395.

DOI: 10.1016/j.cbi.2008.05.023

[15] J. Grunwald, L. Raveh, B.P. Doctor, Y. Ashani, Huperzine A as a pretreatment candidate drug against nerve agent toxicity, Life. Sci, 54 (1994) 991–997.

DOI: 10.1016/0024-3205(94)00501-x

[16] R.K. Gordon a, J.R. Haigha, G.E. Garcia, S.R. Feaster, M.A. Riel, D.E. Lenz, P.S. Aisen, B.P. Doctor, Oral administration of pyridostigmine bromide and huperzine A protects human whole blood cholinesterases from ex vivo exposure to soman, Chem. Bio. Interact. 157–158 (2005).

DOI: 10.1016/j.cbi.2005.10.031

[17] G. Lallement, J. Veyret, C. Masqueliez, S. Aubriot, M.F. Burckhart, D. Baubichon, Efficacy of huperzine in preventing soman-induced seizures, neuropathological changes and lethality. Fund. Clin. Pharmacol. 11 (1997) 387–394.

DOI: 10.1111/j.1472-8206.1997.tb00200.x

[18] T.M. Fahmy, S.L. Demento, M.J. Caplan, I. Mellman, W.M. Saltzman, Design opportunities for actively targeted nanoparticle vaccines, Nanomed. 3 (2008)343–355.

DOI: 10.2217/17435889.3.3.343

[19] Kohei Tahara, Yuta Miyazaki, Yoshiaki Kawashima , Hiromitsu Yamamoto, Brain targeting with surface-modified poly(D, L-lactic-co-glycolic acid) nanoparticles delivered via carotid artery administration, Eur. J. Pharm. Biopharm. 77 (2011) 84–88.

DOI: 10.1016/j.ejpb.2010.11.002

[20] F.Y. Sun, C. Sui, L.S. Teng, X.M. Liu, L.R. Teng, Q.F. Meng, Y.X. Li, Studies on the preparation, characterization and pharmacological evaluation of tolterodine PLGA microspheres Int. J. Pharmaceu. 397 (2010) 44–49.

DOI: 10.1016/j.ijpharm.2010.06.042

[21] R.H. Zhang, L.Q. Li, T. Shi, C. Wang, J.F. Xu, A blood cholinesterase assay method for organophosphorus compounds in vitro, Mil. Med. Sci. 36 (2012)424–426.

[22] G. Mittal, D.K. Sahana, V. Bhardwaj, M.N.V. Ravi Kumar, Estradiol loaded PLGA nanoparticles for oral administration: effect of polymer molecular weight and copolymer composition on release behavior in vitro and in vivo, J. Control. Rel. 119 (2007).

DOI: 10.1016/j.jconrel.2007.01.016