Development of a new polyspecific antivenom for snakebite envenoming in Sri Lanka: Analysis of its preclinical efficacy as compared to a currently available antivenom.

A new whole IgG, freeze-dried, polyspecific antivenom was prepared from the plasma of horses immunized with the venoms of the snakes Daboia russelii, Echis carinatus, Hypnale hypnale, and Naja naja from Sri Lanka. The preclinical neutralizing ability of this antivenom against several toxic and enzymatic activities of these four venoms was analyzed, and compared with that of a batch of VINS antivenom (India) being currently used in Sri Lanka. The activities tested were: lethality, hemorrhagic, in vitro coagulant, proteinase and phospholipase A2. Both antivenoms neutralized, to a different extent, these activities of the venom of D. russelii, E. carinatus, and N. naja. In general, the polyspecific Sri Lankan antivenom was more effective than the Indian antivenom in the neutralization of the venoms of D. russelii and E. carinatus, whereas the Indian antivenom showed a higher efficacy against the venom of N. naja. Regarding H. hypnale, the new Sri Lankan antivenom was effective in the neutralization of all activities tested, whereas the Indian antivenom neutralized lethality but not hemorrhagic, coagulant, proteinase and PLA2 activities, in agreement with the fact that this venom is not included in the immunization mixture for this antivenom. Results suggest that the new polyspecific Sri Lankan antivenom has a satisfactory preclinical neutralizing profile and compares favorably with the Indian antivenom. This is ready to be tested in a clinical trial to evaluate its efficacy and safety in human victims of snakebite envenomings by D. russelii, E. carinatus and H. hypnale in Sri Lanka.

Combined active and passive immunization against nicotine: minimizing monoclonal antibody requirements using a target antibody concentration strategy.

Nicotine vaccines have shown preliminary evidence of efficacy for enhancing smoking cessation rates, but the serum nicotine-specific antibody (NicAb) concentrations produced are highly variable and many subjects do not develop effective levels. As an alternative to vaccination, passive immunization with nicotine-specific monoclonal antibodies could produce more uniform serum NicAb concentrations, but its use is limited by their high cost and shorter elimination half-life. This study investigated supplementing vaccination with monoclonal antibodies in a targeted fashion to increase vaccine efficacy while minimizing the required monoclonal antibody dose. Rats were vaccinated and then given individualized supplemental doses of the nicotine-specific monoclonal antibody Nic311 to achieve a target total serum NicAb concentration known to be effective for blocking locomotor sensitization (LMS) to nicotine. Rats received vaccine, Nic311, both, or neither, followed by 0.3 mg/kg nicotine s.c. for 10 days to produce LMS. Combination immunotherapy completely blocked the development of LMS, while monotherapy with vaccine or Nic311 alone was only minimally effective. Lower brain nicotine levels were associated with reduced locomotor activity averaged over days 7-10. Despite its greater efficacy, combination immunotherapy did not reduce the variability in the resulting total serum NicAb concentrations. Variability in total serum NicAb concentrations was contributed to by both vaccine-generated antibody and by Nic311. These data show that combination immunotherapy, using a Nic311 dose that is by itself only minimally effective, can substantially enhance nicotine vaccine efficacy. However, variability in serum NicAb levels with combination immunotherapy may make translation of this approach challenging.

Tobacco extract but not nicotine impairs the mechanical strength of fracture healing in rats.

The influence of nicotine and tobacco extract (without nicotine) alone and in combination on and mechanical strength of closed femoral fractures in rats was investigated. One hundred four male Sprague-Dawley rats were divided into four groups receiving: nicotine, tobacco extract, tobacco extract plus nicotine, and saline. One week prior to fracture, osmotic pumps were implanted subcutaneously in all animals to administer nicotine equivalent to the serum level of nicotine observed in a smoker consuming one to two packs of cigarettes daily. An equivalent volume of saline was administered to the control animals. Tobacco extract was administered orally. A closed transverse femoral diaphysial fracture was performed, and stabilized with an intramedullary pin. The fractures were mechanically tested after 21 days of healing. Tobacco extract alone decreased the mechanical strength. Ultimate torque and torque at yield point of the tobacco extract group were decreased by 21% (p=0.010) and 23% (p=0.056), respectively, compared with the vehicle (saline) group, and by 20% (p=0.023) and 26% (p=0.004), respectively, compared with the nicotine group. No difference was found between the tobacco extract and tobacco extract plus nicotine groups. An 18% (p=0.013) reduction in torque at yield point was observed in the tobacco extract plus nicotine group compared with the nicotine group. No differences in ultimate stiffness, energy absorption, and callus bone mineral content at the fracture line were found between any of the groups. Serum levels of nicotine were between 40-50 ng/mL in the group given nicotine alone and the group given tobacco extract plus nicotine (equivalent to serum levels observed in persons smoking one to two packs of cigarettes per day).

Effects of nicotine-specific antibodies, Nic311 and Nic-IgG, on the transfer of nicotine across the human placenta.

The adverse effects of smoking during pregnancy on fetal development are, in part, due to nicotine. These effects may be due to the actions of nicotine in fetal circulation or on placental functions. In pregnant rats, vaccination with a nicotine immunogen reduces the transfer of nicotine from the maternal to fetal circulation. However, extrapolation of these results to pregnant women might not be valid due to the well-recognized differences between human and rat placentas. In the current investigation, the effects of nicotine-specific antibodies on the transfer of nicotine from the maternal to fetal circuit of the dually perfused human placental lobule were determined. Two types of nicotine-specific antibodies were investigated; nicotine-specific mouse monoclonal antibody (Nic311, K(d) for nicotine 60nM) and IgG from rabbits vaccinated with a nicotine immunogen (Nic-IgG, K(d) 1.6nM). Transfer of the antibodies from maternal to fetal circuits was negligible. Both rabbit Nic-IgG and, to a lesser extent, mouse monoclonal Nic311 significantly reduced nicotine transfer from the maternal to fetal circuit as well as the retention of the drug by placental tissue. These effects were mediated by a substantial increase in the protein binding of nicotine and a reduction in the unbound nicotine concentration. Therefore, the data cited in this report suggest that the use of nicotine-specific antibodies might reduce fetal exposure to the drug, and that antibody affinity for nicotine is a key determinant of the extent of nicotine transfer.

Continuous nicotine infusion reduces nicotine self-administration in rats with 23-h/day access to nicotine.

The effects of continuous nicotine infusion on nicotine self-administration (NSA) were studied in rats as a model of nicotine replacement therapy (NRT) in humans. A NSA model in which rats had 23-h/day access to nicotine was used to approximate nicotine access conditions in cigarette smokers. In order to estimate serum nicotine concentrations associated with NSA, arterial and venous serum nicotine concentrations were measured during a simulation of NSA. Nicotine was noncontingently administered as 30 doses/12 h of 0.03 mg/kg/i.n.f. or 60 doses/12 h of 0.01 mg/kg/i.n.f. daily. Venous serum nicotine concentrations were measured after the first nicotine dose of the day, and arterial and venous concentrations were measured after doses in the middle of the day. The range of mean concentrations measured was similar to those reported in cigarette smokers (venous concentrations 6-59 ng/ml, arterial concentrations 42-96 ng/ml). The effects of continuous nicotine infusion on NSA were studied by noncontingently administering nicotine at various rates via osmotic pump to animals self-administering nicotine (0.01 or 0.03 mg/kg/i.n.f.) during 23-h/day sessions. Continuous nicotine infusion at all infusion rates substantially suppressed NSA, but suppression was rate-related only for the 0.01-mg/kg/inf NSA unit dose. Nicotine infusion rates producing venous serum nicotine concentrations equaling or exceeding the peak venous levels associated with simulated NSA were more effective than lower infusion rates only at the lower NSA unit dose. The highest nicotine infusion rate had no sustained effect on food-maintained responding, demonstrating its specificity for suppression of NSA. These data provide a model for studying NRT in the rat.