Visualizing lipid nanoparticle trafficking for mRNA vaccine delivery in non-human primates.

mRNA delivered using lipid nanoparticles (LNPs) has become an important subunit vaccine modality, but mechanisms of action for mRNA vaccines remain incompletely understood. Here, we synthesized a metal chelator-lipid conjugate enabling positron emission tomography (PET) tracer labeling of LNP/mRNA vaccines for quantitative visualization of vaccine trafficking in live mice and non-human primates (NHPs). Following intramuscular injection, we observed LNPs distributing through injected muscle tissue, simultaneous with rapid trafficking to draining lymph nodes (dLNs).

Modulation of antigen delivery and lymph node activation in nonhuman primates by saponin adjuvant saponin/monophosphoryl lipid A nanoparticle.

Saponin-based vaccine adjuvants are potent in preclinical animal models and humans, but their mechanisms of action remain poorly understood. Here, using a stabilized HIV envelope trimer immunogen, we carried out studies in nonhuman primates (NHPs) comparing the most common clinical adjuvant aluminum hydroxide (alum) with saponin/monophosphoryl lipid A nanoparticles (SMNP), an immune-stimulating complex-like adjuvant. SMNP elicited substantially stronger humoral immune responses than alum, including 7-fold higher peak antigen-specific germinal center B-cell responses, 18-fold higher autologous neutralizing antibody titers, and higher levels of antigen-specific plasma and memory B cells.

Immunofocusing on the conserved fusion peptide of HIV envelope glycoprotein in rhesus macaques.

During infection, the fusion peptide (FP) of HIV envelope glycoprotein (Env) serves a central role in viral fusion with the host cell. As such, the FP is highly conserved and therefore an attractive epitope for vaccine design. Here, we describe a vaccination study in non-human primates (NHPs) where glycan deletions were made on soluble HIV Env to increase FP epitope exposure.

Diverse priming outcomes under conditions of very rare precursor B cells.

Rare B cells can have special pathogen-recognition features giving them the potential to make outsized contributions to protective immunity. However, rare naive B cells infrequently participate in immune responses. We investigated how germline-targeting vaccine antigen delivery and adjuvant selection affect priming of exceptionally rare BG18-like HIV broadly neutralizing antibody-precursor B cells (~1 in 50 million) in non-human primates.

Charge-Stabilized Nanodiscs as a New Class of Lipid Nanoparticles.

Nanoparticles have the potential to improve disease treatment and diagnosis due to their ability to incorporate drugs, alter pharmacokinetics, and enable tissue targeting. While considerable effort is placed on developing spherical lipid-based nanocarriers, recent evidence suggests that high aspect ratio lipid nanocarriers can exhibit enhanced disease site targeting and altered cellular interactions. However, the assembly of lipid-based nanoparticles into non-spherical morphologies has typically required incorporating additional agents such as synthetic polymers, proteins, lipid-polymer conjugates, or detergents.

Neoantigen architectures define immunogenicity and drive immune evasion of tumors with heterogenous neoantigen expression.

Background: Intratumoral heterogeneity (ITH) and subclonal antigen expression blunt antitumor immunity and are associated with poor responses to immune-checkpoint blockade immunotherapy (ICB) in patients with cancer. The underlying mechanisms however thus far remained elusive, preventing the design of novel treatment approaches for patients with high ITH tumors.

Methods: We developed a mouse model of lung adenocarcinoma with defined expression of different neoantigens (NeoAg), enabling us to analyze how these impact antitumor T-cell immunity and to study underlying mechanisms.

Fungal Attachment-Resistant Polymers for the Additive Manufacture of Medical Devices.

This study reports the development of the first copolymer material that (i) is resistant to fungal attachment and hence biofilm formation, (ii) operates via a nonkilling mechanism, i.e., avoids the use of antifungal actives and the emergence of fungal resistance, (iii) exhibits sufficient elasticity for use in flexible medical devices, and (iv) is suitable for 3D printing (3DP), enabling the production of safer, personalized medical devices.

Two-dose priming immunization amplifies humoral immunity by synchronizing vaccine delivery with the germinal center response.

Prolonging exposure to subunit vaccines during the primary immune response enhances humoral immunity. Escalating-dose immunization (EDI), administering vaccines every other day in an increasing pattern over 2 weeks, is particularly effective but challenging to implement clinically. Here, using an HIV Env trimer/saponin adjuvant vaccine, we explored simplified EDI regimens and found that a two-shot regimen administering 20% of the vaccine followed by the remaining 80% of the dose 7 days later increased T responses 6-fold, antigen-specific germinal center (GC) B cells 10-fold, and serum antibody titers 10-fold compared with bolus immunization.

Modulation of antigen delivery and lymph node activation in non-human primates by saponin adjuvant SMNP.

Saponin-based vaccine adjuvants are potent in preclinical animal models and humans, but their mechanisms of action remain poorly understood. Here, using a stabilized HIV envelope trimer immunogen, we carried out studies in non-human primates (NHPs) comparing the most common clinical adjuvant alum with Saponin/MPLA Nanoparticles (SMNP), a novel ISCOMs-like adjuvant. SMNP elicited substantially stronger humoral immune responses than alum, including 7-fold higher peak antigen-specific germinal center B cell responses, 18-fold higher autologous neutralizing antibody titers, and higher levels of antigen-specific plasma and memory B cells.

Vaccines combining slow delivery and follicle targeting of antigens increase germinal center B cell clonal diversity and clonal expansion.

Vaccines incorporating slow delivery, multivalent antigen display, or immunomodulation through adjuvants have an important role to play in shaping the humoral immune response. Here we analyzed mechanisms of action of a clinically relevant combination adjuvant strategy, where phosphoserine (pSer)-tagged immunogens bound to aluminum hydroxide (alum) adjuvant (promoting prolonged antigen delivery to draining lymph nodes) are combined with a potent saponin nanoparticle adjuvant termed SMNP (which alters lymph flow and antigen entry into lymph nodes). When employed with a stabilized HIV Env trimer antigen in mice, this combined adjuvant approach promoted substantial enhancements in germinal center (GC) and antibody responses relative to either adjuvant alone.

Dose-dependent regulation of immune memory responses against HIV by saponin monophosphoryl lipid A nanoparticle adjuvant.

The induction of durable protective immune responses is the main goal of prophylactic vaccines, and adjuvants play an important role as drivers of such responses. Despite advances in vaccine strategies, a safe and effective HIV vaccine remains a significant challenge. The use of an appropriate adjuvant is crucial to the success of HIV vaccines.

Local delivery of cell surface-targeted immunocytokines programs systemic antitumor immunity.

Systemically administered cytokines are potent immunotherapeutics but can cause severe dose-limiting toxicities. To overcome this challenge, cytokines have been engineered for intratumoral retention after local delivery. However, despite inducing regression of treated lesions, tumor-localized cytokines often elicit only modest responses at distal untreated tumors.

Generation of an inflammatory niche in an injectable hydrogel depot through recruitment of key immune cells improves efficacy of mRNA vaccines.

Messenger RNA (mRNA) delivered in lipid nanoparticles (LNPs) rose to the forefront of vaccine candidates during the COVID-19 pandemic due in part to scalability, adaptability, and potency. Yet there remain critical areas for improvements of these vaccines in durability and breadth of humoral responses. In this work, we explore a modular strategy to target mRNA/LNPs to antigen presenting cells with an injectable polymer-nanoparticle (PNP) hydrogel depot technology which recruits key immune cells and forms an immunological niche in vivo.

Visualizing lipid nanoparticle trafficking for mRNA vaccine delivery in non-human primates.

mRNA delivered using lipid nanoparticles (LNPs) has become an important subunit vaccine modality, but mechanisms of action for mRNA vaccines remain incompletely understood. Here, we synthesized a metal chelator-lipid conjugate enabling positron emission tomography (PET) tracer labeling of LNP/mRNA vaccines for quantitative visualization of vaccine trafficking in live non-human primates (NHPs). Following i.

Vaccine targeting to mucosal lymphoid tissues promotes humoral immunity in the gastrointestinal tract.

Viruses, bacteria, and parasites frequently cause infections in the gastrointestinal tract, but traditional vaccination strategies typically elicit little or no mucosal antibody responses. Here, we report a strategy to effectively concentrate immunogens and adjuvants in gut-draining lymph nodes (LNs) to induce gut-associated mucosal immunity. We prepared nanoemulsions (NEs) based on biodegradable oils commonly used as vaccine adjuvants, which encapsulated a potent Toll-like receptor agonist and displayed antigen conjugated to their surface.

Vaccination with nanoparticles displaying gH/gL from Epstein-Barr virus elicits limited cross-protection against rhesus lymphocryptovirus.

Epstein-Barr virus (EBV) is associated with infectious mononucleosis, cancer, and multiple sclerosis. A vaccine that prevents infection and/or EBV-associated morbidity is an unmet need. The viral gH/gL glycoprotein complex is essential for infectivity, making it an attractive vaccine target.

Controlled Lipid Self-Assembly for Scalable Manufacturing of Next-Generation Immune Stimulating Complexes.

Immune stimulating complexes (ISCOMs) are safe and effective saponin-based adjuvants formed by the self-assembly of saponin, cholesterol, and phospholipids in water to form cage-like 30-40 nm diameter particles. Inclusion of the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA) in ISCOM particles yields a promising next-generation adjuvant termed Saponin-MPLA NanoParticles (SMNP). In this work, we detail protocols to produce ISCOMs or SMNP via a tangential flow filtration (TFF) process suitable for scalable synthesis and Good Manufacturing Practice (GMP) production of clinical-grade adjuvants.

Design of Cytotoxic T Cell Epitopes by Machine Learning of Human Degrons.

Antigen processing is critical for therapeutic vaccines to generate epitopes for priming cytotoxic T cell responses against cancer and pathogens, but insufficient processing often limits the quantity of epitopes released. We address this challenge using machine learning to ascribe a proteasomal degradation score to epitope sequences. Epitopes with varying scores were translocated into cells using nontoxic anthrax proteins.