Self-Assembling Allergen Vaccine Platform Raises Therapeutic Allergen-Specific IgG Responses without Induction of Systemic Allergic Responses.

Allergen immunotherapies are often successful at desensitizing allergic patients but can require life-long dosing and suffer from frequent adverse events including instances of systemic anaphylaxis, leading to poor patient compliance and high cost. Allergen vaccines, in turn, can generate more durable immunological allergen desensitization with far fewer doses. However, like immunotherapies, allergen vaccines are often highly reactogenic in allergic patients, hampering their use in therapeutic settings. In this work, we utilize a peptide-based self-assembling nanofiber platform to design allergen vaccines against allergen B-cell epitopes that do not elicit systemic anaphylaxis when administered subcutaneously to allergic mice. We show that, in contrast to protein vaccines, nanofiber vaccines prevent leakage of allergen material into the vascular compartment, a feature that likely underpins their reduced systemic reactogenicity. Further, we show that our allergen vaccine platform elicits therapeutic IgG antibody responses capable of desensitizing allergic mice to allergen-induced Type I hypersensitivity reactions. Finally, we have demonstrated a proof-of-concept for the therapeutic potential of nanofiber-based peanut allergen vaccines directed against peanut allergen-derived epitopes.

A sublingual nanofiber vaccine to prevent urinary tract infections.

Urinary tract infections (UTIs) are a major public health problem affecting millions of individuals each year. Recurrent UTIs are managed by long-term antibiotic use, making the alarming rise of antibiotic resistance a substantial threat to future UTI treatment. Extended antibiotic regimens may also have adverse effects on the microbiome. Here, we report the use of a supramolecular vaccine to provide long-term protection against uropathogenic Escherichia coli, which cause 80% of uncomplicated UTIs. We designed mucus-penetrating peptide-polymer nanofibers to enable sublingual (under the tongue) vaccine delivery and elicit antibody responses systemically and in the urogenital tract. In a mouse model of UTI, we demonstrate equivalent efficacy to high-dose oral antibiotics but with significantly less perturbation of the gut microbiome. We also formulate our vaccine as a rapid-dissolving sublingual tablet that raises response in mice and rabbits. Our approach represents a promising alternative to antibiotics for the treatment and prevention of UTIs.

Combined PET and whole-tissue imaging of lymphatic-targeting vaccines in non-human primates.

Antigen accumulation in lymph nodes (LNs) is critical for vaccine efficacy, but understanding of vaccine biodistribution in humans or large animals remains limited. Using the rhesus macaque model, we employed a combination of positron emission tomography (PET) and fluorescence imaging to characterize the whole-animal to tissue-level biodistribution of a subunit vaccine comprised of an HIV envelope trimer protein nanoparticle (trimer-NP) and lipid-conjugated CpG adjuvant (amph-CpG). Following immunization in the thigh, PET imaging revealed vaccine uptake primarily in inguinal and iliac LNs, reaching distances up to 17 cm away from the injection site. Within LNs, trimer-NPs exhibited striking accumulation on the periphery of follicular dendritic cell (FDC) networks in B cell follicles. Comparative imaging of soluble Env trimers (not presented on nanoparticles) in naïve or previously-immunized animals revealed diffuse deposition of trimer antigens in LNs following primary immunization, but concentration on FDCs in pre-immunized animals with high levels of trimer-specific IgG. These data demonstrate the capacity of nanoparticle or "albumin hitchhiking" technologies to concentrate vaccines in genitourinary tract-draining LNs, which may be valuable for promoting mucosal immunity.

Titrating Polyarginine into Nanofibers Enhances Cyclic-Dinucleotide Adjuvanticity in Vitro and after Sublingual Immunization.

Effective sublingual peptide immunization requires overcoming challenges of both delivery and immunogenicity. Mucosal adjuvants, such as cyclic-dinucleotides (CDN), can promote sublingual immune responses but must be codelivered with the antigen to the epithelium for maximum effect. We designed peptide-polymer nanofibers (PEG-Q11) displaying nona-arginine (R9) at a high density to promote complexation with CDNs via bidentate hydrogen-bonding with arginine side chains. We coassembled PEG-Q11 and PEG-Q11R9 peptides to titrate the concentration of R9 within nanofibers. In vitro, PEG-Q11R9 fibers and cyclic-di-GMP or cyclic-di-AMP adjuvants had a synergistic effect on enhancing dendritic cell activation that was STING-dependent and increased monotonically with increasing R9 concentration. The polyvalent display of R9 on assembled nanofibers was significantly more effective at promoting CDN-mediated DC activation in vitro than mixing nanofibers with an equimolar concentration of unassembled R9 peptide. The sublingual administration of nanofibers revealed a bell-shaped trend between increasing R9 concentration and enhancements to antigen trafficking and the activation of DCs in the draining lymph nodes. Intermediate levels of R9 within sublingually administered PEG-Q11 fibers were optimal for immunization, suggesting a balance between polyarginine's ability to sequester CDNs along the nanofiber and its potentially detrimental mucoadhesive interactions. These findings present a potentially generalizable biomaterial strategy for enhancing the potency of CDN adjuvants and reveal important design considerations for the nascent field of sublingual biomaterial immunization.

Targeting HIV Env immunogens to B cell follicles in nonhuman primates through immune complex or protein nanoparticle formulations.

Following immunization, high-affinity antibody responses develop within germinal centers (GCs), specialized sites within follicles of the lymph node (LN) where B cells proliferate and undergo somatic hypermutation. Antigen availability within GCs is important, as B cells must acquire and present antigen to follicular helper T cells to drive this process. However, recombinant protein immunogens such as soluble human immunodeficiency virus (HIV) envelope (Env) trimers do not efficiently accumulate in follicles following traditional immunization. Here, we demonstrate two strategies to concentrate HIV Env immunogens in follicles, via the formation of immune complexes (ICs) or by employing self-assembling protein nanoparticles for multivalent display of Env antigens. Using rhesus macaques, we show that within a few days following immunization, free trimers were present in a diffuse pattern in draining LNs, while trimer ICs and Env nanoparticles accumulated in B cell follicles. Whole LN imaging strikingly revealed that ICs and trimer nanoparticles concentrated in as many as 500 follicles in a single LN within two days after immunization. Imaging of LNs collected seven days postimmunization showed that Env nanoparticles persisted on follicular dendritic cells in the light zone of nascent GCs. These findings suggest that the form of antigen administered in vaccination can dramatically impact localization in lymphoid tissues and provides a new rationale for the enhanced immune responses observed following immunization with ICs or nanoparticles.