Janeyuth Chaisakul

Effects of Animal Venoms and Toxins on Hallmarks of Cancer.

Animal venoms are a cocktail of proteins and peptides, targeting vital physiological processes. Venoms have evolved to assist in the capture and digestion of prey. Key venom components often include neurotoxins, myotoxins, cardiotoxins, hematoxins and catalytic enzymes. The pharmacological activities of venom components have been investigated as a source of potential therapeutic agents. Interestingly, a number of animal toxins display profound anticancer effects. These include toxins purified from snake, bee and scorpion venoms effecting cancer cell proliferation, migration, invasion, apoptotic activity and neovascularization. Indeed, the mechanism behind the anticancer effect of certain toxins is similar to that of agents currently used in chemotherapy. For example, Lebein is a snake venom disintegrin which generates anti-angiogenic effects by inhibiting vascular endothelial growth factors (VEGF). In this review article, we highlight the biological activities of animal toxins on the multiple steps of tumour formation or hallmarks of cancer. We also discuss recent progress in the discovery of lead compounds for anticancer drug development from venom components.

Hypotensive and vascular relaxant effects of phospholipase A2 toxins from Papuan taipan (Oxyuranus scutellatus) venom.

Phospholipase A2 (PLA2) toxins are common and abundant components of Australasian elapid venoms. These toxins are associated with a range of activities including neurotoxicity, myotoxicity and coagulation disturbances. We have recently reported that sudden cardiovascular collapse induced by Papuan taipan (Oxyuranus scutellatus) venom involves a combination of the release of dilator autacoids and a direct effect on the smooth muscle. In this study, we aimed to isolate PLA2 components from Papuan taipan venom and investigate their contribution to the hypotensive action of this venom. O. scutellatus venom was fractionated using size-exclusion high performance liquid chromatography (HPLC), and fractions screened for activity in anaesthetized rats. Fraction three from O. scutellatus venom (i.e. OSC3, 14.2±1.0% of whole venom) produced a 64% decrease in mean arterial pressure. Reverse-phase HPLC indicated that OSC3 consisted of two major components (i.e. OSC3a and OSC3b). OSC3a and OSC3b produced a significant hypotensive response in anaesthetized rats which were attenuated by prior administration of indomethacin or the combination of mepyramine and heparin. N-terminal analysis indicated that OSC3a and b displayed sequence homology to PLA2 toxins isolated from coastal taipan (O. scutellatus scutellatus) venom. These findings indicate that PLA2 components may play an important role in the development of hypotension and vascular relaxation which may contribute to the effects observed after envenoming by these Australasian elapids.

Differential Myotoxic and Cytotoxic Activities of Pre-synaptic Neurotoxins from Papuan Taipan (Oxyuranus scutellatus) and Irian Jayan Death Adder (Acanthophis rugosus) Venoms

Pre‐synaptic PLA2 neurotoxins are important components of many Australasian elapid snake venoms. These toxins disrupt neurotransmitter release. Taipoxin, a pre‐synaptic neurotoxin isolated from the venom of the coastal taipan (Oxyuranus scutellatus), causes necrosis and muscle degeneration. The present study examined the myotoxic and cytotoxic activities of venoms from the Papuan taipan (O. scutellatus) and Irian Jayan death adder (Acanthophis rugosus), and also tested their pre‐synaptic neurotoxins: cannitoxin and P‐EPTX‐Ar1a. Based on size‐exclusion chromatography analysis, cannitoxin represents 16% of O. scutellatus venom, while P‐EPTX‐Ar1a represents 6% of A. rugosus venom. In the chick biventer cervicis nerve‐muscle preparation, A. rugosus venom displayed significantly higher myotoxic activity than O. scutellatus venom as indicated by inhibition of direct twitches, and an increase in baseline tension. Both cannitoxin and P‐EPTX‐Ar1a displayed marked myotoxic activity. A. rugosus venom (50–300 μg/ml) produced concentration‐dependent inhibition of cell proliferation in a rat skeletal muscle cell line (L6), while 300 μg/ml of O. scutellatus venom was required to inhibit cell proliferation, following 24‐hr incubation. P‐EPTX‐Ar1a had greater cytotoxicity than cannitoxin, inhibiting cell proliferation after 24‐hr incubation in L6 cells. Lactate dehydrogenase levels were increased after 1‐hr incubation with A. rugosus venom (100–250 μg/ml), O. scutellatus venom (200–250 μg/ml) and P‐EPTX‐Ar1a (1–2 μM), but not cannitoxin (1–2 μM), suggesting venoms/toxin generated cell necrosis. Thus, A. rugosus and O. scutellatus venoms possess different myotoxic and cytotoxic activities. The proportion of pre‐synaptic neurotoxin in the venoms and PLA2 activity of the whole venoms are unlikely to be responsible for these activities.

In Vitro Toxic Effects of Puff Adder (Bitis arietans) Venom, and Their Neutralization by Antivenom

This study investigated the in vitro toxic effects of Bitis arietans venom and the ability of antivenom produced by the South African Institute of Medical Research (SAIMR) to neutralize these effects. The venom (50 µg/mL) reduced nerve-mediated twitches of the chick biventer muscle to 19% ± 2% of initial magnitude (n = 4) within 2 h. This inhibitory effect of the venom was significantly attenuated by prior incubation of tissues with SAIMR antivenom (0.864 µg/µL; 67% ± 4%; P < 0.05; n = 3–5, unpaired t-test). Addition of antivenom at t50 failed to prevent further inhibition or reverse the inhibition of twitches and responses to agonists. The myotoxic action of the venom (50 µg/mL) was evidenced by a decrease in direct twitches (30% ± 6% of the initial twitch magnitude) and increase in baseline tension (by 0.7 ± 0.3 g within 3 h) of the chick biventer. Antivenom failed to block these effects. Antivenom however prevented the venom induced cytotoxic effects on L6 skeletal muscle cells. Venom induced a marginal but significant reduction in plasma clotting times at concentrations above 7.8 µg/100 µL of plasma, indicating poor procoagulant effects. In addition, the results of western immunoblotting indicate strong immunoreactivity with venom proteins, thus warranting further detailed studies on the neutralization of the effects of individual venom toxins by antivenom.

A pharmacological examination of the cardiovascular effects of Malayan krait (Bungarus candidus) venoms

Cardiovascular effects (e.g., tachycardia, hypo- and/or hypertension) are often clinical outcomes of snake envenoming. Malayan krait (Bungarus candidus) envenoming has been reported to cause cardiovascular effects that may be related to abnormalities in parasympathetic activity. However, the exact mechanism for this effect has yet to be determined. In the present study, we investigated the in vivo and in vitro cardiovascular effects of B. candidus venoms from Southern (BC-S) and Northeastern (BC-NE) Thailand. SDS-PAGE analysis of venoms showed some differences in the protein profile of the venoms. B. candidus venoms (50 µg/kg–100 µg/kg, i.v.) caused dose-dependent hypotension in anaesthetised rats. The highest dose caused sudden hypotension (phase I) followed by a return of mean arterial pressure to baseline levels and a decrease in heart rate with transient hypertension (phase II) prior to a small decrease in blood pressure (phase III). Prior administration of monovalent antivenom significantly attenuated the hypotension induced by venoms (100 µg/kg, i.v.). The sudden hypotensive effect of BC-NE venom was abolished by prior administration of hexamethonium (10 mg/kg, i.v.) or atropine (5 mg/kg, i.v.). BC-S and BC-NE venoms (0.1 µg/kg–100 µg/mL) induced concentration-dependent relaxation (EC50 = 8 ± 1 and 13 ± 3 µg/mL, respectively) in endothelium-intact aorta. The concentration–response curves were markedly shifted to the right by pre-incubation with L-NAME (0.2 mM), or removal of the endothelium, suggesting that endothelium-derived nitric oxide (NO) is likely to be responsible for venom-induced aortic relaxation. Our data indicate that the cardiovascular effects caused by B. candidus venoms may be due to a combination of vascular mediators (i.e., NO) and autonomic adaptation via nicotinic and muscarinic acetylcholine receptors.

Perinatal Taurine Supplementation Prevents Metabolic and Cardiovascular Effects of Maternal Diabetes in Adult Rat Offspring

This study tests the hypothesis that perinatal taurine supplementation prevents diabetes mellitus and hypertension in adult offspring of maternal diabetic rats. Female Wistar rats were fed normal rat chow and tap water with (Diabetes group) or without diabetic induction by intraperitoneal streptozotocin injection (Control group) before pregnancy. Then, they were supplemented with 3% taurine in water (Control+T and Diabetes+T groups) or water alone from conception to weaning. After weaning, both male and female offspring were fed normal rat chow and tap water throughout the study. Blood chemistry and cardiovascular parameters were studied in 16-week old rats. Body, heart, and kidney weights were not significantly different among the eight groups. Further, lipid profiles except triglyceride were not significantly different among male and female groups, while male Diabetes displayed increased fasting blood glucose, decreased plasma insulin, and increased plasma triglyceride compared to other groups. Compared to Control, mean arterial pressures significantly increased and baroreflex control of heart rate decreased in both male and female Diabetes, while heart rates significantly decreased in male but increased in female Diabetes group. Although perinatal taurine supplementation did not affect any measured parameters in Control groups, it abolished the adverse effects of maternal diabetes on fasting blood glucose, plasma insulin, lipid profiles, mean arterial pressure, heart rate, and baroreflex sensitivity in adult male and female offspring. The present study indicates that maternal diabetes mellitus induces metabolic and cardiovascular defects more in male than female adult offspring, and these adverse effects can be prevented by perinatal taurine supplementation.