Adjuvant-carrying synthetic vaccine particles augment the immune response to encapsulated antigen and exhibit strong local immune activation without inducing systemic cytokine release

Augmentation of immunogenicity can be achieved by particulate delivery of an antigen and by its co-administration with an adjuvant. However, many adjuvants initiate strong systemic inflammatory reactions in vivo, leading to potential adverse events and safety concerns. We have developed a synthetic vaccine particle (SVP) technology that enables co-encapsulation of antigen with potent adjuvants. We demonstrate that co-delivery of an antigen with a TLR7/8 or TLR9 agonist in synthetic polymer nanoparticles results in a strong augmentation of humoral and cellular immune responses with minimal systemic production of inflammatory cytokines. In contrast, antigen encapsulated into nanoparticles and admixed with free TLR7/8 agonist leads to lower immunogenicity and rapid induction of high levels of inflammatory cytokines in the serum (e.g., TNF-α and IL-6 levels are 50- to 200-fold higher upon injection of free resiquimod (R848) than of nanoparticle-encapsulated R848). Conversely, local immune stimulation as evidenced by cellular infiltration of draining lymph nodes and by intranodal cytokine production was more pronounced and persisted longer when SVP-encapsulated TLR agonists were used. The strong local immune activation achieved using a modular self-assembling nanoparticle platform markedly enhanced immunogenicity and was equally effective whether antigen and adjuvant were co-encapsulated in a single nanoparticle formulation or co-delivered in two separate nanoparticles. Moreover, particle encapsulation enabled the utilization of CpG oligonucleotides with the natural phosphodiester backbone, which are otherwise rapidly hydrolyzed by nucleases in vivo. The use of SVP may enable clinical use of potent TLR agonists as vaccine adjuvants for indications where cellular immunity or robust humoral responses are required

Development of polymeric nanoparticle vaccines for immunostimulation

Thesis (Ph. D. in Medical Engineering and Medical Physics)--Harvard-MIT Program in Health Sciences and Technology, 2013.Cataloged from PDF version of thesis.Includes bibliographical references (pages 93-96).Vaccines have revolutionized medicine by increasing the life expectancy of children and substantially decreasing the morbidity of multiple infectious diseases worldwide. Over several decades, we have acquired significant gains in the understanding of the underlying mechanisms involved in developing protective immunity, yet vaccine development has progressed comparatively slowly. This thesis serves to explore two polymeric nanoparticle platforms to demonstrate the therapeutic potential of synthetic nanocarriers as vaccines with the aim of 1) providing greater spatiotemporal release of small molecule adjuvant to secondary lymphoid sites and 2) providing a tunable surface for loading B cell antigen epitopes in a specific conformation to drive epitope-specific antibody response. In recent decades, TLR mechanisms have been elucidated and novel agonists have been developed, yet our generation still has not seen paramount progress in the clinical translation of these agonists due to risks of systemic toxicity and off target effects. In the first section, we synthesized 223±18 nm poly(lactic-co-glycolic acid)- poly(ethylene glycol)/ poly(lactic acid)-R848 (PLGA-PEG/PLA-R848) nanoparticle vaccine that is designed to deliver a combination of antigen and control release of a small molecule adjuvant R848 (tl/2= 42 hours) to drive a potent antigen-specific immune response. Using ovalbumin as a model protein, this vaccine is able to enhance antigen presentation and co-stimulatory molecules on dendritic cells and subsequently enhanced proliferation of antigen-specific naive CD8+ cells in vitro. Upon vaccination, our delivery system is able to increase cell-mediated and humoral response in comparison to its soluble form, thereby illustrating the potential to bring novel small molecule adjuvants to the clinics. In the second section, we developed a nanoparticle vaccine platform that allows selective orientation of peptide epitopes to enhance B cell response in an application that has therapeutic potential for treatment for cardiovascular disease (CVD). Utilizing epitopes discovered through in silico modeling for human PCSK9, a plasma protein that plays an important role in LDL cholesterol (LDL-c) levels in the blood, our nanoparticle allows selective orientation through biotin-streptavidin conjugation. Upon vaccination with CPG, selected synthetic epitopes conjugated to polymeric nanoparticles trended to reduce serum LDL-c and serum PCSK9 in murine models. Additionally, antibodies in the serum showed promise to increase LDL-receptor levels in HepG2 cells transfected in with WT-hPCSK9 and GOF-hPCSK9 separately suggesting that this vaccine has the potential to reduce risks of CVD. These studies demonstrate that designing polymeric nanoparticles for applications to stimulate the immune system can help define new, cost-effective treatment options in applications for prophylaxis against infectious diseases that are unresponsive to traditional routes of vaccination or for immunotherapy against cardiovascular disease and cancer.by Pamela A. Basto.Ph.D.in Medical Engineering and Medical Physic

Akt and SHP-1 are DC-intrinsic checkpoints for tumor immunity

BM-derived DC (BMDC) are powerful antigen-presenting cells. When loaded with immune complexes (IC), consisting of tumor antigens bound to antitumor antibody, BMDC induce powerful antitumor immunity in mice. However, attempts to employ this strategy clinically with either tumor-associated DC (TADC) or monocyte-derived DC (MoDC) have been disappointing. To investigate the basis for this phenomenon, we compared the response of BMDC, TADC, and MoDC to tumor IgG-IC. Our findings revealed, in both mice and humans, that upon exposure to IgG-IC, BMDC internalized the IC, increased costimulatory molecule expression, and stimulated autologous T cells. In contrast, TADC and, surprisingly, MoDC remained inert upon contact with IC due to dysfunctional signaling following engagement of Fcγ receptors. Such dysfunction is associated with elevated levels of the Src homology region 2 domain-containing phosphatase-1 (SHP-1) and phosphatases regulating Akt activation. Indeed, concomitant inhibition of both SHP-1 and phosphatases that regulate Akt activation conferred upon TADC and MoDC the capacity to take up and process IC and induce antitumor immunity in vivo. This work identifies the molecular checkpoints that govern activation of MoDC and TADC and their capacity to elicit T cell immunity

Adjuvant-carrying synthetic vaccine particles augment the immune response to encapsulated antigen and exhibit strong local immune activation without inducing systemic cytokine release

Augmentation of immunogenicity can be achieved by particulate delivery of an antigen and by its co-administration with an adjuvant. However, many adjuvants initiate strong systemic inflammatory reactions in vivo, leading to potential adverse events and safety concerns. We have developed a synthetic vaccine particle (SVP) technology that enables co-encapsulation of antigen with potent adjuvants. We demonstrate that co-delivery of an antigen with a TLR7/8 or TLR9 agonist in synthetic polymer nanoparticles results in a strong augmentation of humoral and cellular immune responses with minimal systemic production of inflammatory cytokines. In contrast, antigen encapsulated into nanoparticles and admixed with free TLR7/8 agonist leads to lower immunogenicity and rapid induction of high levels of inflammatory cytokines in the serum (e.g., TNF-a and IL-6 levels are 50- to 200-fold higher upon injection of free resiquimod (R848) than of nanoparticle-encapsulated R848). Conversely, local immune stimulation as evidenced by cellular infiltration of draining lymph nodes and by intranodal cytokine production was more pronounced and persisted longer when SVP-encapsulated TLR agonists were used. The strong local immune activation achieved using a modular self-assembling nanoparticle platform markedly enhanced immunogenicity and was equally effective whether antigen and adjuvant were co-encapsulated in a single nanoparticle formulation or co-delivered in two separate nanoparticles. Moreover, particle encapsulation enabled the utilization of CpG oligonucleotides with the natural phosphodiester backbone, which are otherwise rapidly hydrolyzed by nucleases in vivo. The use of SVP may enable clinical use of potent TLR agonists as vaccine adjuvants for indications where cellular immunity or robust humoral responses are required