Authored by Connor O'Leary
Snakebite envenoming is a Neglected Tropical Disease (NTD) which kills over 100,000 people each year, as well as leaving hundreds of thousands with life-changing long-term morbidity1, and therefore may be considered as one of the world’s most severe NTDs. Snakes are abundant in warmer climates and so it is the rural populations of tropical countries of the developing world that suffer the greatest burden of the disease, with recent Disability-Adjusted Life-Years (DALY) reports highlighting the scale of this burden. Despite this, snakebite receives little attention from public health authorities, the pharmaceutical industry or governments when compared with diseases of similar impact, perpetuating the cycle of poverty.
Snakebite and the cycle of poverty
The United Nations Development Programme (UNDP) defines those who are ‘multidimensionally poor’, in accordance with the Multidimensional Poverty Index (MPI), as those suffering deprivations in 33% or more of the weighted indicators2. This definition of poverty provides a comprehensive and robust assessment of deprivation in the context of education, health and living standards. The relationship between snakebite envenoming and poverty is clearly demonstrated by the comparison of maps depicting the definition of poverty outlined by the MPI (Figure 1) and the snakebite mortality map (Figure 2), which displays a remarkable correlation. Furthermore, contact between snakes and the impoverished rural populations of tropical and sub-tropical countries is relatively common as a result of the non-mechanised, low-cost farming techniques upon which these populations rely, especially in the rainy season when human agricultural activity coincides with the snakes’ breeding season3.
Although antivenom remains an effective life-saving treatment for snakebite cases, the notoriously complex and specific protein mixtures that comprise snake venom makes antivenom production expensive, whilst severely impeding economies of scale6. When one considers these limitations to the commercial incentives for antivenom development, it comes as little surprise that over the past few decades several major antivenom manufacturers such as Syntex and Behringwerke have discontinued African antivenom production. This has had devastating implications on the burden of snakebite in African nations such as Nigeria, in which non-specific and ineffective antivenoms, fake antivenoms and dangerous traditional treatments are exacerbating the mortality and morbidity risks posed by the NTD. This is a particular concern in regions such as north-eastern Nigeria in which up to 70% of hospital beds are occupied by victims of snakebite in the sowing and harvesting seasons7.
Assessing the burden of snakebite envenoming
Moreover, a further burden snakebite envenoming imposes on victims is the therapeutic limitations of antivenom, which although life-saving, has limited efficacy against effects of snakebite such as local tissue damage3. Pharmaceutical advances in the efficacy of snakebite treatments will require greater recognition of the burden of the NTD and investment from public health authorities and ‘large pharma’. Additionally, this inevitably results in an alarming DALY assessment of the NTD, as outlined in a DALY report of the disease’s burden in 16 west African countries by Habib et al. (summarised in Figure 3), ranking snakebite envenoming as the fourth highest burden out of the twenty NTDs7. DALY assessments are based on the sum of years of life lost and years of life lived with disability, generating a quantitative analysis of the burden incurred by the disease (Figure 4). This study found that snakebite envenoming accounted for between 320,000-1,200,000 DALYs in these 16 countries, with the lower estimate greater than the worldwide estimated burden of leprosy and the African burden of leishmaniasis. These results produce a damning assessment of the mismatch between the NTD’s burden and its resources. Furthermore, research by Shiffman into the allocation of funding for disease control, which evaluates funding from 42 major donors and pharmaceutical companies, found no evidence of any contributions made to snakebite envenoming during the period of the survey, compounding the scale of this tragedy8,9.
Mode of action
Venomous snakes produce venoms with complex compositions which contain phospholipase A2 toxins (PLA2), which cause a range of pathological alterations. PLA2 ‘myotoxins’ act mainly on skeletal muscles, causing extensive damage due to their high selectivity for plasma membrane targets in muscle fibres. Irreversible muscle cell damage occurs as a result of the mode of action of myotoxins, causing calcium influx into the cytosol which causes mitochondrial disfunction and other degenerative effects. Following necrosis (localised death of living tissue), myotoxic PLA2s cause an extensive inflammatory reaction caused by their action on inflammatory cells, leading to intense pain, haemorrhages and blistering. This tissue damage further enhances the risk of infection, which can exacerbate tissue loss and require amputation of the affected limb. Conversely, ‘neurotoxins’ such as taipoxin and textilotoxin, in snake venom compositions act specifically on the peripheral nervous system, paralysing prey rapidly and are therefore responsible for the death of the prey11.
Myotoxins and neurotoxins in snake venom further catalyse the production of lysophospholipids (LysoPLs) and fatty acids, which causes membrane damage. The presence of LysoPLs and fatty acids in the plasma membrane inhibits endocytosis and increases membrane permeability to ions, characterised by a prominent influx of Ca2+ which promotes a complex series of cellular derangements. Fatty acids in particular are associated with the inhibition of the biochemically essential organic chemical adenosine triphosphate (ATP) and therefore mitochondrial respiration. However, PLA2 toxins are unable to cross the blood-brain barrier and therefore do not reach the central nervous system3.
Structure of snake PLA2 toxins
Secreted neurotoxic and myotoxic PLA2 toxins fold in a characteristic scaffold consisting of four main helices, held together by seven disulfide bonds, and two long anti-parallel disulfide-linked alpha helices which define a hydrophobic channel. The hydrophobic channel, which leads the phospholipid substrate to the catalytic site, is characterised by four amino acid residues: histidine-48, aspartic acid-49, tyrosine-52 and aspartic acid-99. In the mode of action, the histidine residue hydrogen-bonds the water molecule involved in hydrolysis, whilst aspartic acid-49 coordinates the Ca2+ ion which binds the phosphate and carbonyl groups of the phospholipid molecule during hydrolysis10.
Tragically, for decades the public health significance of snakebite envenoming has remained grossly underappreciated by international and national health authorities, which has precluded efforts to reduce the impact and burden of the disease. However, the decision of the World Health Organisation (WHO) to relist snakebite envenoming into Category A on the formal list of Neglected Tropical Diseases12 in 2017 represents a significant step forward in achieving the global recognition that will be essential in attaining resources commensurate with the disease’s pathological impact, as well as the interest of the pharmaceutical industry.
With an increasing international focus on snakebite envenoming, there have been numerous promising advances in the field of medicinal chemistry in recent years which take aim at tackling the efficacy and affordability challenges of snakebite treatments. One notable focus in research has been the development of treatments for venom-induced local tissue destruction, which represents the most significant limitation to the therapeutic efficacy of antivenom. Antivenom is largely ineffective in the prevention of local tissue damage, often necessitating amputation, because of the discrepancies between the pharmacokinetic profiles of the low molecular weight toxins – reaching tissue targets rapidly after injection – and antivenoms13. Indeed, it is reported that approximately 400,000 snakebite-related amputations are undertaken each year14. For example, PLA2 inhibitors, such as varespladib and methyl-varespladib (Figure 6), have shown potential in delaying the local effects of envenomation, whilst crucially being low-cost15. Other ongoing research focuses include peptide inhibitors of matrix metalloproteinases, VHH fragments of camelid immunoglobulin G (IgG) and polymer nanoparticles14,15.
 J. Slagboom, J. Kool, R.A. Harrison and N.R. Casewell, British Journal of Haematology, 2017, 177(6), 947-959.
 Multidimensional Poverty Index (MPI), United Nations Development Programme Human Development Reports, http://hdr.undp.org/en/content/multidimensional-poverty-index-mpi (accessed September 2018).
 J.M. Gutierrez, J.J. Calvete, A.G. Habib, R.A. Harrison, D.J. Williams and D.A. Warrell, Nature, 2017, DOI: 10.1038/nrdp.2017.63.
 Global Extreme Poverty, Our World in Data, https://ourworldindata.org/extreme-poverty (accessed September 2018).
 A. Kasturiratne, A.R. Wickremasinghe, N. de Silva, N.K. Gunawardena and A. Pathmeswaran, PLoS Med, 2008, 5, 1591-1604.
 R.A. Harrison, A. Hargreaves, S.C. Wagstaff, B. Faragher and D.G. Lalloo, PLOS Neglected Tropical Diseases, 2009, 3(12), 569.
 R.D.G. Theakston and D.A. Warrell, Lancet, 2000, 356, 2104.
 A.G. Habib, A. Kuznik, M. Hamza, M.I. Abdullahi, B.A. Chedi, J. Chippaux and D.A. Warrell, PLOS Neglected Tropical Diseases, 2015, DOI: doi.org/10.1371/journal.pntd.0004088.
 J. Shiffman, Health Policy and Planning, 2006, 21(6), 411-420.
 ‘The burden of disease and what it means in England’, Public Health England (PHE), https://publichealthmatters.blog.gov.uk/2015/09/15/the-burden-of-disease-and-what-it-means-in-england/ (accessed September 2018).
 C. Montecucco, J.M. Gutierrez and B. Lamonte, B. Cell. Mol. Life Sci., 2008, 65, 2897.
 ‘Snakebite envenomation turns again into a neglected tropical disease!’, World Health Organisation, http://www.who.int/snakebites/resources/s40409-017-0127-6/en/ (accessed September 2018).
 J.M. Gutierrez, G. Leon and B. Lomonte, Clinical Pharmacokinetics, 2003, 42(8), 721-741.
 R.A. Harrison, D.A. Cook, C. Renjifo, N.R. Casewell, R.B. Currier and S.C. Wagstaff, Journal of Proteomics, 2011, 74, 1768-1780.
 M. Lewin, S. Samuel, J. Merkel and P. Bickler, Toxins (Basel), 2016, 8, 248.