Unfortunately, long-term outcomes for nicotine replacement therapies remain poor and achieve an abstinence rate of only 10–20% after the first year. Anti-addiction vaccines induce antibodies that block the pharmacological effects of drugs like nicotine. To date, vaccines for smoking cessation have shown considerable promise in preclinical animal models for their ability to diminish nicotine-mediated physiological and behavioral responses including nicotine craving. However, clinical studies in humans failed to measure significant differences in smoking abstinence between the intervention and placebo groups. Importantly, subgroup analyses from two Phase II studies indicated that subjects with the highest antibody titers showed increased abstinence at 12 months, and the non-abstaining subjects within the high Ab group reduced daily cigarette consumption by 50%. Encouraged by these findings, two subsequent Phase 3 studies attempted to increase Ab titers even further by modifying vaccine dose and immunization schedule. However, clinical responses were not improved and both studies failed to achieve the established efficacy endpoints. To investigate this result further, a follow-on study measured the nicotine binding capacity induced in vaccinated subjects and determined that the Abs could only prevent,12% of an infused dose of nicotine from reaching the brain. Thus, the resulting Ab responses induced by the vaccine were insufficient for providing clinical benefit. This has Temozolomide 85622-93-1 raised questions about the requirements for improved vaccine efficacy. Nicotine and other drugs of abuse are non-immunogenic and must be conjugated to a protein carrier to facilitate antigen presentation and the induction of T cell help, which regulates durable Ab responses and memory generation. To date, most hapten carriers are derived from microbial sources like keyhole limpet hemocyanin, tetanus toxoid, diphtheria toxoid, and pseudomonas exotoxin A, although they may be limited in important ways. For instance, epitope density is a critical factor influencing the magnitude and quality of the immune response. However, the maximum number of haptens that can be loaded on the clinical candidate vaccines, which is typically dictated by the number of lysines used for chemical conjugation, is less than 50. Also, hapten stoichiometry and spacing likely varies within each carrier and uncertainty remains about which linkages within the protein present the best epitope for stimulating high affinity Ab titers. Another potential problem is that these proteins are highly immunogenic and might induce anti-carrier antibodies that could limit booster immunizations over time. “Epitopic suppression” is a widely recognized phenomenon first observed with licensed polysaccharide conjugate vaccines, and experiments have shown that this effect is suppressed by increasing hapten density. Clinical findings suggest that the vaccines tested for smoking cessation failed to induce meaningful titers of functional Abs sufficient to block nicotine entry into brain. A number of variables are known to influence conjugate vaccine performance including the carrier, hapten structure, hapten density, and the choice of adjuvant.