Vaccination against nicotine is a potential treatment for tobacco smoking. Clinical trials show effect only in high antibody responders; therefore it is necessary to increase the effectiveness of nicotine vaccines. The use of a multivalent vaccine that activates several B cell populations is a possible approach to increase antibody response. The aim of this study was to investigate whether three different nicotine immunogens could be mixed to generate independent responses resulting in additive antibody titers, and whether this would alter nicotine distribution to a greater extent than antibodies generated by a monovalent vaccine. When immunogens were administered s.c. with alum adjuvant, the trivalent vaccine generated significantly higher titers and prevented the distribution of an i.v. nicotine dose to brain to a greater extent than an equivalent dose of a monovalent vaccine. The number of rats with antibody titers >1:10,000 was significantly increased in the trivalent group compared to the monovalent group. There were no correlations between the titers generated by the different nicotine immunogens in the trivalent vaccine, supporting the hypothesis that the immunogens generated independent responses from distinct populations of B cells. In contrast, when administered i.p. in Freund’s adjuvant, the trivalent nicotine vaccine was not more immunogenic than its component monovalent vaccine. Vaccine immunogenicity was suppressed if unconjugated protein was added to the monovalent vaccine formulated in Freund’s adjuvant, compared to monovalent vaccine alone. These data suggest a protein–protein interaction that affects titers negatively and is apparent when the vaccines are formulated with Freund’s adjuvant. In summary, a trivalent nicotine vaccine formulated with alum showed significantly higher efficacy than a dose-matched monovalent vaccine and may offer a strategy for increasing nicotine vaccine immunogenicity. This approach may be generalizable to other nicotine immunogens or vaccines for other addictive drugs.

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 were 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.

Structurally distinct nicotine immunogens can elicit independent antibody responses against nicotine when administered concurrently. Co-administering different nicotine immunogens together as a multivalent vaccine could be a useful way to generate higher antibody levels than with monovalent vaccines alone. The immunogenicity and additivity of monovalent and bivalent nicotine vaccines was studied across a range of immunogen doses, adjuvants, and routes to assess the generality of this approach. Rats were vaccinated with total immunogen doses of 12.5 - 100 μg of 3′-aminomethyl nicotine conjugated to recombinant Pseudomonas exoprotein A (3′-AmNic-rEPA), 6-carboxymethylureido nicotine conjugated to keyhole limpet hemocyanin (6-CMUNic-KLH), or both. Vaccines were administered s.c. in alum or i.p. in Freund’s adjuvant at matched total immunogen doses. When administered s.c. in alum, the contributions of the individual immunogens to total nicotine-specific antibody (NicAb) titers and concentrations were preserved across a range of doses. Antibody affinity for nicotine varied greatly among individuals but was similar for monovalent and bivalent vaccines. However when administered i.p. in Freund’s adjuvant the contributions of the individual immunogens to total NicAb titers and concentrations were compromised at some doses. These results support the possibility of co-administering structurally distinct nicotine immunogens to achieve a more robust immune response than can be obtained with monovalent immunogens alone. Choice of adjuvant was important for the preservation of immunogen component activity.