Tobacco smoking has consistently been the leading cause of preventable diseases and premature deaths. Currently, pharmacological interventions have only shown limited smoking cessation efficacy and sometimes are associated with severe side effects. As an alternative, nicotine vaccines have emerged as a promising strategy to combating nicotine addiction. However, conventional conjugate nicotine vaccines have shown limited ability to induce a sufficiently strong immune response due to their intrinsic shortfalls. In this study, a lipid-poly(lactic-co-glycolic acid) (PLGA) nanoparticle-based next-generation nicotine vaccine has been developed to overcome the drawbacks of conjugate nicotine vaccines. Also, the influence of multiple factors, including nanoparticle size, hapten density, hapten localization, carrier protein, and molecular adjuvants, on its immunogenicity has been investigated. Results indicated that all these studied factors significantly affected the immunological efficacy of the nicotine nanovaccine. First, 100 nm nanovaccine was found to elicit a significantly higher anti-nicotine antibody titer than the 500 nm nanovaccine. Secondly, the high-density nanovaccine exhibited a better immunological efficacy than the low- and medium-density counterparts. Thirdly, the nanovaccine with hapten localized on both carrier protein and nanoparticle surface induced a significantly higher anti-nicotine antibody titer and had a considerably better ability to block nicotine from entering the brain of mice than the nanovaccines with hapten localized only on carrier protein or nanoparticle surface. Fourthly, the nanovaccines carrying cross reactive materials 197 (CRM197) or tetanus toxoid (TT) showed a better immunological efficacy than the nanovaccines using keyhole limpet hemocyanin (KLH) or KLH subunit as carrier proteins. Finally, the co-delivery of monophosphoryl lipid A (MPLA) and Resiquimod (R848) achieved a considerably higher antibody titer and brain nicotine reduction than only using MPLA or R848 alone as adjuvants. Collectively, the findings from this study may lead to a better understanding of the impact of multiple factors on the immunological efficacy of the hybrid nanoparticle-based nicotine nanovaccine. The findings may also provide significant guidance for the development of other drug abuse and nanoparticle-based vaccines. In addition, the optimized lipid-PLGA hybrid nanoparticle-based nicotine nanovaccine obtained by modulating the studied factors can be a promising candidate as the next-generation nicotine vaccine for treating nicotine addiction. / PHD / Tobacco smoking is prevalent and represents one of the largest public health concerns in the world. Tobacco use remains to be the leading cause of preventable diseases and premature deaths. Typically, smokers without medications experience huge difficulty in quitting smoking due to the addictiveness of nicotine. Although several pharmacological interventions are available to smokers, they have only shown limited smoking cessation efficacy. In recent years, nicotine vaccines that can induce the production of nicotine specific antibodies have been proposed as a promising strategy for smoking cessation. Currently, almost all the already-developed nicotine vaccines are conjugate nicotine vaccines in which nicotine haptens are attached to a carrier protein for presentation. However, conventional conjugate nicotine vaccines suffer from many innate shortcomings that largely limit their immunological efficacy. This study aimed to develop a nanoparticle-based next-generation nicotine vaccine to overcome the drawbacks of conventional conjugate nicotine vaccines. Because many factors may potentially affect the immunological efficacy of a nicotine vaccine, this study focused on investigating the influence of multiple factors, including nanoparticle size, hapten density, hapten localization, carrier protein, and molecular adjuvants, on the immunological efficacy of the developed hybrid nanoparticle-based nicotine vaccine. Results from this study revealed that all these studied factors significantly influenced the immunological efficacy of the nicotine nanovaccine. By modulating v these factors, enhanced immune responses that can result in higher titers of anti-nicotine antibodies and better ability in blocking nicotine from entering the brain of mice could be achieved. The findings from this study would lead to a better understanding of how multiple factors influence the immunological efficacy of a nanoparticle-based nicotine vaccine. In addition, the findings from this study can also provide significant guidance for the development of other drug abuse and nanoparticle-based vaccines. More importantly, the hybrid nanoparticle-based nicotine nanovaccine can be a promising candidate for smoking cessation.