Evaluating the physicochemical properties and efficacy of recently expired and aged antivenom products from Thailand and Taiwan

doi:10.1016/j.toxicon.2019.02.010

Toxicon

Volume 160, 15 March 2019, Pages 55-58

https://doi.org/10.1016/j.toxicon.2019.02.010 Get rights and content

Highlights

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Recent-expired (within 2 years) and aged (18 years) antivenoms were evaluated.
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Gel filtration chromatography and SDS-PAGE revealed minimal protein degradation.
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Both retained immunoreactivity toward the homologous venom proteins.
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Hemotoxicity, neurotoxicity and lethal effects of venoms were neutralized.
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With proper storage, they remained stable beyond the indicated shelf life.

Abstract

Gel filtration chromatography and gel electrophoresis revealed minimal protein degradation in lyophilized antivenoms which were 2-year expired (Hemato Polyvalent, Neuro Polyvalent; Thailand) and 18-year expired (Hemato Bivalent, Neuro Bivalent; Taiwan). All expired antivenoms retained immunological binding activity, and were able to neutralize the hemotoxic or neurotoxic as well as lethal effects of the homologous snake venoms. The findings show that antivenoms under proper storage conditions may remain relatively stable beyond the indicated shelf life.

Graphical abstract

Section snippets

Acknowledgement

The authors are grateful to the University of Malaya for the provision of research facilities and funding (Grants no. RG352-15AFR and BKS003-2017).

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Cross reactivity and lethality neutralization of venoms of Indonesian Trimeresurus complex species by Thai Green Pit Viper Antivenom
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Cited by (10)

Recent advancements in snake antivenom production
2023, International Journal of Biological Macromolecules
Snakebite envenoming (SBE), a neglected tropical disease, claims lives of about 138,000 people globally, and antivenom is the only approved treatment worldwide. However, this century-old therapy has serious limitations, including limited efficacy and some side effects. Although alternative and adjunct therapies are being developed, their commercialization will take time. Hence, improving existing antivenom therapy is crucial for immediate reduction in the global SBE burden. The neutralization potential and immunogenicity of antivenoms depend primarily on the venom pool used for animal immunization and the production host, along with antivenom purification procedure and quality control. Enhancing antivenom quality and production capacity are also critical actions of the World Health Organization (WHO) roadmap 2021 against SBE. The present review details the latest developments in antivenom production, such as immunogen preparation, production host, antibody purification methods, antivenom testing-including alternative animal models, in vitro assays, and proteomics and in silico methodologies, and storage, reported from 2018 to 2022. Based on these reports, we propose that production of broadly specific, affordable, safe, and effective (BASE) antivenoms is fundamental to realizing the WHO roadmap and reducing the global SBE burden. This concept can also be applied during the designing of alternative antivenoms.
Clinical aspects of snakebite envenoming and its treatment in low-resource settings
2023, The Lancet
There is increasing recognition of the public health importance of snakebite envenoming. Worldwide annual incidence is likely to be 5 million bites, with mortality exceeding 150 000 deaths, and the resulting physical and psychological morbidity leads to substantial social and economic repercussions. Prevention through community education by trained health workers is the most effective and economically viable strategy for reducing risk of bites and envenoming. Clinical challenges to effective treatment are most substantial in rural areas of low-resource settings, where snakebites are most common. Classic skills of history taking, physical examination, and use of affordable point-of-care tests should be followed by monitoring of evolving local and systemic envenoming. Despite the profusion of new ideas for interventions, hyperimmune equine or ovine plasma-derived antivenoms remain the only specific treatment for snakebite envenoming. The enormous interspecies and intraspecies complexity and diversity of snake venoms, revealed by modern venomics, demands a radical redesign of many current antivenoms.
Sharp-nosed Pit Viper (Deinagkistrodon acutus) from Taiwan and China: A comparative study on venom toxicity and neutralization by two specific antivenoms across the Strait
2022, Acta Tropica
Citation Excerpt :
DaMAV-Taiwan was also more effective than DaMAV-China in neutralizing the procoagulant and lethal effects of its homologous Da-Taiwan venom, indicating that the venom lethality is systemically related to the procoagulant activity of the venom (which causes coagulopathy as described above). DaMAV-China, in comparison to DaMAV-Taiwan, neutralized the procoagulant and lethal effects of its homologous Da-China venom less efficaciously, indicating that the proportions of neutralizing antibodies in the antivenoms varied presumably due to differences in the production techniques, e.g., the immunization protocol and antibody purification method adopted (León et al., 2018; WHO, 2016b), and probably DaMAV-China as a liquid formulation might have a lower product stability than the lyophilized DaMAV-Taiwan (Tan et al., 2019; WHO, 2016b). Besides the amount of neutralizing antibodies, the production techniques might also affect the pharmacokinetics and binding affinity of the antibody toward the target proteins to form stable immunocomplexes in vivo for effective neutralization (Gutiérrez et al., 2003).
In East Asia, the Sharp-nosed Pit Viper (Deinagkistrodon acutus) is a medically important venomous snake in Taiwan and China, two geographical areas long separated by the Taiwan Strait. Yet, snake venom variation is little known between specimens found across the Strait. This study thus investigated the intra-species variation of D. acutus venoms from Taiwan (Da-Taiwan) and China (Da-China) in their profiles of gel electrophoresis, toxicity, immunoreactivity and neutralization effect by antivenom. Da-China venom exhibited higher procoagulant, hemorrhagic and lethal activities than Da-Taiwan venom, presumably attributed to the higher abundance of moderate-to-high molecular weight toxins (procoagulants and hemorrhagins) in the venom. The mono-specific antivenoms produced in Taiwan (DaMAV-Taiwan) and China (DaMAV-China) were immunoreactive toward both venoms, and were able to neutralize the venom toxicity to different extents. DaMAV-Taiwan was more efficacious in neutralizing the venom procoagulant and lethal effects, while DaMAV-China was more potent against hemorrhagic effect. The discrepancy in efficacy between the two antivenoms could be due to varying proportions of neutralizing antibodies in the respective products, influenced by techniques of antibody raising and purification. Further study is warranted to elucidate variation in the proteome and antigenicity of D. acutus venom between snakes from Taiwan and China.
Potential para-specific and geographical utility of Thai Green Pit Viper (Trimeresurus albolabris) Monovalent Antivenom: Neutralization of procoagulant and hemorrhagic activities of diverse Trimeresurus pit viper venoms
2021, Toxicon
Citation Excerpt :
Furthermore, the formulation and protein concentrations of the antivenoms were evaluated. SDS-PAGE profiling revealed that all three antivenoms were composed of proteins of ∼120 kDa, which were reduced to fragments corresponding to the heavy chains (∼28 kDa) and the light chains (∼22 kDa) of an Fab unit (Tan et al., 2019b, 2020). It is thus deduced that the antivenoms contained F(ab’)2 fragments derived from the plasma IgG of horses upon pepsin digestion to remove the Fc fragment, which is a common strategy adopted by many manufacturers to reduce the risk of anaphylactic reaction and anti-immunoglobulin response (León et al., 2001).
The Trimeresurus complex consists of diverse medically important venomous pit vipers that cause snakebite envenomation. Antivenoms, however, are in limited supply, and are specific to only two out of the many species across Asia. This study thus investigated the immunoreactivities of regional pit viper antivenoms toward selected Trimeresurus pit viper venoms, and examined the neutralization of their hemotoxic activities. Trimeresurus albolabris Monovalent Antivenom (TaMAV, Thailand) exhibited a higher immunoreactivity than Hemato Bivalent Antivenom (HBAV, raised against Trimeresurus stejnegeri and Protobothrops mucrosquamatus, Taiwan) and Gloydius brevicaudus Monovalent Antivenom (GbMAV, China), attributed to its monovalent nature and conserved antigens in the Trimeresurus pit viper venoms. The venoms showed moderate-to-strong in vitro procoagulant and in vivo hemorrhagic effects consistent with hemotoxic envenomation, except for the Sri Lankan Trimeresurus trigonocephalus venom which lacked hemorrhagic activity. TaMAV was able to differentially neutralize both in vitro and in vivo hemotoxic effects of the venoms, with the lowest efficacy shown against the procoagulant effect of T. trigonocephalus venom. The findings suggest that TaMAV is a potentially useful treatment for envenomation caused by hetero-specific Trimeresurus pit vipers, in particular those in Southeast Asia and East Asia. Clinical study is warranted to establish its spectrum of para-specific effectiveness, and dosages need be tailored to the different species in respective regions.
Cross-reactivity of some Micrurus venoms against experimental and therapeutic anti-Micrurus antivenoms
2021, Toxicon
Citation Excerpt :
In most of the experiments the antivenoms were used before their expiration date. In some experiments, however, they were used after their expiration date, an approach ensuring batch consistency and valid based on the available bibliography on the efficacy of antivenoms after their expiration date, if properly conserved (O'Leary et al., 2009; Sánchez et al., 2019; Tan et al., 2019). In addition all antivenoms used were confirmed to have the physicochemical and neutralizing potencies declared and required for their use at the moment of experimentation (ThNAmMAv 90 i.v. LD50/ml of M. nigrocinctus venom and ThSAmMAv 1 mg/ml of M. pyrrhocryptus venom).
We developed experimental equine polyvalent and monovalent antivenoms against the venoms of Micrurus (M.) fulvius, M. nigrocinctus and M. surinamensis and studied their immunochemical reactivity on the venoms used as immunogens and on M. pyrrhocryptus, M altirostris and M. balyocoriphus venoms. Assessment of the neutralizing capacity of the polyvalent experimental antivenom was based on inhibition of lethality (preincubation and rescue assay experiments in mice) and indirect hemolytic and phospholipase activities. The immunochemical reactivity and neutralizing capacity were compared with those of two therapeutic antivenoms used for the treatment of coral snake envenomation in North America and in Argentina. In general, the experimental antivenom conferred a comparable level of neutralization against the venoms used as immunogens when compared to the therapeutic antivenoms and a certain level of cross-neutralization against the other venoms. The results suggest the need for additional venoms in the immunogenic mixture used, in order to obtain a broad spectrum anti-Micrurus antivenom with a good neutralizing potency. Paraspecific neutralization of South American coral snake venoms, although present at a higher level than the neutralization conferred by available nonspecific Micrurus therapeutic antivenoms, was rather low in relation to the specific neutralizing capacity.
The neutralization efficacy of expired polyvalent antivenoms: An alternative option
2019, Toxicon
Citation Excerpt :
In doing so, this will put forth the urgency of the pathology of snake envenoming attracting more attention by governmental agencies and other powers of wills, which in turn, we hope, will provide more resources to allow for the improvements of antivenom availability, accessibility, affordability, and efficacy. Presently, there are antivenoms that are marked with expiration dates that are potentially functional (Tan et al., 2019) and could be an alternative for snakebite therapy when companies cease to produce them due to the vicious cycle of antivenom market decline, which is created by the high prices of antivenoms that drives down their affordability; thereby, reducing the sales of antivenoms. In addition, these expired antivenoms could also be used in the event of emergencies by non-indigenous venomous snakebites in which the specific antivenoms of the country of origin cannot be obtained on time.
The expense of production and distribution of snakebite antivenom, as well as its relatively infrequent use, has caused antivenom to be increasingly difficult to obtain and ultimately producing an alarming global shortage. Unused, expired antivenom may represent a significant, untapped resource to ameliorate this crisis. This study examines the efficacy of expired antivenom over time using in vitro, whole blood clotting, and platelet function statistics. Representatives from three years for four different global brands of polyvalent antivenom were chosen and tested against their corresponding venoms as well as other venoms that could display cross-reactivity. These antivenoms include Wyeth Polyvalent (U.S.; exp. 1997, 2001, 2003), Antivipmyn^® (Mexico; exp. 2005, 2013, 2017), Biotecfars Polyvalent (Venezuela; exp. 2010, 2014, 2016), and SAIMR (South Africa; exp. 1997, 2005, 2017). Venoms of species tested were Crotalus atrox against Wyeth; C. atrox and Crotalus vegrandis against Antivipmyn^®; C. atrox, C. vegrandis and Bothrops colombiensis against Biotecfar; and Bitis gabonica and Echis carinatus against South African Institute for Medical Research (SAIMR). Parameters recorded were activated clotting time (ACT), clotting rate (CR), and platelet function (PF). Preliminary results are encouraging as the antivenoms maintained significant efficacy even 20 y after their expiration date. We anticipate these results will motivate further studies and provide hope in the cases of snakebite emergencies when preferable treatments are unavailable.

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Highlights

Abstract

Graphical abstract

Section snippets

Acknowledgement

References (26)

Proteomics, functional characterization and antivenom neutralization of the venom of Pakistani Russell's viper (Daboia russelii) from the wild

J. Proteom.

Effect of preservatives on IgG aggregation, complement-activating effect and hypotensive activity of horse polyvalent antivenom used in snakebite envenomation

Biologicals

Cross neutralization of common Southeast Asian viperid venoms by a Thai polyvalent snake antivenom (Hemato Polyvalent Snake Antivenom)

Acta Trop.

An examination of the activity of expired and mistreated commercial Australian antivenoms

Trans. R. Soc. Trop. Med. Hyg.

Venom proteome of Bungarus sindanus (Sind krait) from Pakistan and in vivo cross-neutralization of toxicity using an Indian polyvalent antivenom

J. Proteom.

Cross neutralization of Hypnale hypnale (hump-nosed pit viper) venom by polyvalent and monovalent Malayan pit viper antivenoms in vitro and in a rodent model

Acta Trop.

Cross reactivity and lethality neutralization of venoms of Indonesian Trimeresurus complex species by Thai Green Pit Viper Antivenom

Toxicon

Venom proteome of the yellow-lipped sea krait, Laticauda colubrina from Bali: insights into subvenomic diversity, venom antigenicity and cross-neutralization by antivenom

J. Proteom.

Venomics, lethality and neutralization of Naja kaouthia (monocled cobra) venoms from three different geographical regions of Southeast Asia

J Proteomics

Venomics of Naja sputatrix, the Javan spitting cobra: a short neurotoxin-driven venom needing improved antivenom neutralization

J Proteomics

Snake venomics and antivenomics of Protobothrops mucrosquamatus and Viridovipera stejnegeri from Taiwan: keys to understand the variable immune response in horses

J. Proteom.

The snakebite fight

Nature

Antivenin (Micrurus fulvius)(Equine Origin). North American coral snake antivenin

Cited by (10)

Recent advancements in snake antivenom production

Clinical aspects of snakebite envenoming and its treatment in low-resource settings

Sharp-nosed Pit Viper (Deinagkistrodon acutus) from Taiwan and China: A comparative study on venom toxicity and neutralization by two specific antivenoms across the Strait

Potential para-specific and geographical utility of Thai Green Pit Viper (Trimeresurus albolabris) Monovalent Antivenom: Neutralization of procoagulant and hemorrhagic activities of diverse Trimeresurus pit viper venoms

Cross-reactivity of some Micrurus venoms against experimental and therapeutic anti-Micrurus antivenoms

The neutralization efficacy of expired polyvalent antivenoms: An alternative option

Short communicationEvaluating the physicochemical properties and efficacy of recently expired and aged antivenom products from Thailand and Taiwan

Highlights

Abstract

Graphical abstract

Section snippets

Acknowledgement

J. Proteom.

Biologicals

Acta Trop.

Trans. R. Soc. Trop. Med. Hyg.

J. Proteom.

Acta Trop.

Toxicon

J. Proteom.

J Proteomics

J Proteomics

J. Proteom.

The snakebite fight

Nature

Antivenin (Micrurus fulvius)(Equine Origin). North American coral snake antivenin

Short communication
Evaluating the physicochemical properties and efficacy of recently expired and aged antivenom products from Thailand and Taiwan