Uniform-sized Biodegradable Polymer-based Microparticles As Adjuvants For Subunit Vaccines

Biodegradable polymer-based microparticles (MPs) are promising vaccine adjuvants. Understanding how various physicochemical properties affect the adjuvant effects of MPs is a crucial requirement for rational adjuvant design. In most studies concerning MPs adjuvants, antigen was entrapped within MPs. Little attention was focused on other antigen-MPs formulation methods, such as adsorption and mixture. Therefore, the overall goal of this thesis is to clarify the effects of various parameters on MPs adjuvanticity and the underlying mechanisms, when antigen is formulated with MPs by adsorbing or simply mixing. In detail, this thesis mainly included the following issues:1) Uniform-sized poly(lactic acid) (PLA) MPs with different size were prepared by membrane emulsification technique to clarify the effects of size on MPs adjuvanticity. Compared to MPs with other size (500 nm,2.1μm, and 4.9 μm), MPs900nm were more effective in inducing macrophages activation and improving antigen uptake by macrophages, both of which were essential for antigen presentation and initiation of adaptive immune response. In vivo study revealed that MPS900 nm elicited more potent antigen-specific immune responses, showing superior adjuvant effects to other MPs.2) Based on the aforementioned investigations, PLA MPs (900 nm) were further explored as adjuvant for H5N1 influenza split vaccine. Compared to traditional alum adjuvant, MPs more efficiently augmented antigen-specific immune response. Action mode analysis suggested that alum induced severe local inflammation, whereas MPs not. Considering the adjuvanticity and side effects (local inflammation) of both adjuvants, it’s concluded that PLA MPs are promising alternative adjuvant for influenza split vaccine.3) It was investigated that how surface charge affected antigen-MPs interaction, and subsequent antigen-specific immune response. With the MPs surface charge increasing, the adsorption efficiency of antigen onto MPs increased. In vitro cell experiments suggested that enhanced positive surface charge of MPs favored antigen uptake by antigen-presenting cells. Both humoral and cellular immune response elicited by MPs-adjuvanted vaccine could be improved by regulating MPs surface charges.4) The adjuvanticity of immunopotentiator-loaded polymeric MPs was further evaluated. Incorporating imiquimod (IMQ) into MPs significantly improved the efficacy of MPs in activating antigen-presenting cells (BMDCs and pMΦs), and improving cellular antigen uptake. IMQ-loaded MPs showed superior adjuvanticity to blank MPs. Considering adjuvanticity and safety profiles (polymer and IMQ, both approved by FDA), we conclude that IMQ-loaded polymeric microparticles are promising robust vaccine adjuvant.5) It was investigated that how various antigen-MPs formulations impacted antigen kenitics and the resultant antigen-specific immune responses. The combined formulation (antigen encapsulated in MPs+antigen mixed with MPs) induced more powerful antigen-specific immune responses than each single-component formulation. Enhanced efficacy of combined formulation might be attributed to effective provision of both adequate initial antigen exposure and long-term antigen persistence, which further efficiently induced DCs activation and follicular Th cell differentiation in lymph nodes.In conclusion, these results indicated that MPs adjuvant with superior activity could be prepared by optimizing MPs size, surface charge, and antigen-MPs formulation patterns, and combining molecular immunopotentiators with MPs. Understanding how various parameters affect the efficacy of MPs-adjuvanted vaccine might have significant implications for rational vaccine design.