ImmTOR nanoparticles enhance AAV transgene expression after initial and repeat dosing in a mouse model of methylmalonic acidemia.

A major barrier to adeno-associated virus (AAV) gene therapy is the inability to re-dose patients due to formation of vector-induced neutralizing antibodies (Nabs). Tolerogenic nanoparticles encapsulating rapamycin (ImmTOR) provide long-term and specific suppression of adaptive immune responses, allowing for vector re-dosing. Moreover, co-administration of hepatotropic AAV vectors and ImmTOR leads to an increase of transgene expression even after the first dose. ImmTOR and AAV Anc80 encoding the methylmalonyl-coenzyme A (CoA) mutase (MMUT) combination was tested in a mouse model of methylmalonic acidemia, a disease caused by mutations in the MMUT gene. Repeated co-administration of Anc80 and ImmTOR was well tolerated and led to nearly complete inhibition of immunoglobulin (Ig)G antibodies to the Anc80 capsid. A more profound decrease of plasma levels of the key toxic metabolite, plasma methylmalonic acid (pMMA), and disease biomarker, fibroblast growth factor 21 (FGF21), was observed after treatment with the ImmTOR and Anc80-MMUT combination. In addition, there were higher numbers of viral genomes per cell (vg/cell) and increased transgene expression when ImmTOR was co-administered with Anc80-MMUT. These effects were dose-dependent, with the higher doses of ImmTOR providing higher vg/cell and mRNA levels, and an improved biomarker response. Combining of ImmTOR and AAV can not only block the IgG response against capsid, but it also appears to potentiate transduction and enhance therapeutic transgene expression in the mouse model.

Synthetic vaccine particles for durable cytolytic T lymphocyte responses and anti-tumor immunotherapy.

We previously reported that synthetic vaccine particles (SVP) encapsulating antigens and TLR agonists resulted in augmentation of immune responses with minimal production of systemic inflammatory cytokines. Here we evaluated two different polymer formulations of SVP-encapsulated antigens and tested their ability to induce cytolytic T lymphocytes (CTL) in combination with SVP-encapsulated adjuvants. One formulation led to efficient antigen processing and cross-presentation, rapid and sustained CTL activity, and expansion of CD8+ T cell effector memory cells locally and centrally, which persisted for at least 1-2 years after a single immunization. SVP therapeutic dosing resulted in suppression of tumor growth and a substantial delay in mortality in several syngeneic mouse cancer models. Treatment with checkpoint inhibitors and/or cytotoxic drugs, while suboptimal on their own, showed considerable synergy with SVP immunization. SVP encapsulation of endosomal TLR agonists provided superior CTL induction, therapeutic benefit and/or improved safety profile compared to free adjuvants. SVP vaccines encapsulating mutated HPV-16 E7 and E6/E7 recombinant proteins led to induction of broad CTL activity and strong inhibition of TC-1 tumor growth, even when administered therapeutically 13-14 days after tumor inoculation in animals bearing palpable tumors. A pilot study in non-human primates showed that SVP-encapsulated E7/E6 adjuvanted with SVP-encapsulated poly(I:C) led to robust induction of antigen-specific T and B cell responses.

Eliciting cytotoxic T-lymphocyte responses from synthetic vectors containing one or two epitopes in a C57BL/6 mouse model using peptide-containing biodegradable microspheres and adjuvants.

We describe a vaccine delivery mechanism consisting of a synthetic, non-living vector of large d,l poly(lactic-co-glycolic) acid (PLGA) microspheres that carry specific cytotoxic T lymphocyte (CTL) epitopes. We demonstrate in mice that it can be used to elicit substantial interferon gamma ELISPOT responses to more than one specific epitope in the same individual. Our data suggest that a superior adjuvant configuration for the formulation is to place a TLR-9 agonist CpG inside the microsphere and a TLR-4 agonist MPLA in the injectate solution. This finding contrasts with the observations of others. Our approach provides a means to elicit immune responses efficiently to select epitopes, which may be important for an effective vaccine against HIV.

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.

A comparison of the effects of two burs on endodontic access in all-ceramic high lucite crowns.

OBJECTIVE: The purpose of this study was to examine and compare the effects of diamond and tungsten carbide burs with respect to the preparation of access through all-ceramic crowns. STUDY DESIGN: Thirty extracted maxillary premolars were restored with all-ceramic crowns. Each specimen was assigned to one of two groups: (1) access opening prepared with a round diamond bur; (2) access opening prepared with a carbide fissure bur. Access preparations were scanned by means of scanning electron microscopy; observed were defects categorized as edge chipping, microcracks, and fractures. RESULTS: Edge chipping around the access was universal. Significant chipping (x > or = 0.1 m) was seen in 43% of access peripheries. Eleven percent of the crowns fractured. chi(2) analysis (P <.05) demonstrated no statistical difference between the bur groups. Results of a t test revealed no statistical difference in edge chipping between the two bur types. CONCLUSIONS: All-ceramic crowns bonded to extracted maxillary premolars may experience edge chipping, microcracks, and fractures at equal rates whether access is prepared with a round medium coarse diamond bur or a tungsten carbide fissure bur.

Complete-denture therapy for the geriatric patient.

Denture therapy for the geriatric patient will be in high demand for the decades ahead. An older adult's medical, functional, and psychological status should be considered in each phase of complete-denture treatment. Patient satisfaction is dependent on how well the dentist has restored facial appearance and rehabilitated chewing efficiency.

Fixed prosthodontics and esthetic considerations for the older adult.

A new generation of older adults, who are more educated, health-conscious, and economically independent than their predecessors is bringing a fresh perspective and poses unique opportunities and challenges to fixed prosthodontics. Subtle differences in technique, attention to detail, and innovative application of materials and procedures are the main ingredients of successful fixed prosthodontic care for seniors. The goals of fixed prosthodontics for the older adult are fundamentally different from those for a younger population. A practice that enhances the integrity of residual tooth structure while simultaneously creating an environment less prone to dental disease and disability should be emphasized.

The philosophies of dowel diameter preparation: a literature review.

This article reviewed the literature regarding the diameter of dowels and identified three distinct philosophies of dowel space preparation. One group advocated the narrowest diameter for fabrication of a dowel to a desired length. Another recommended a dowel space with an apical diameter equal to one third of the narrowest dimension of the root at the terminus of the dowel. A third group advised that at least 1 mm of sound dentin should surround the entire surface of the dowel. A combination of the one third and 1 mm minimal philosophies yielded a practical guideline for dowel space preparation, particularly in aged teeth. Requiring a definite amount of tooth structure surrounding the dowel, while adhering to the one third proportion, indicated upper limits on both the diameter and length of the dowel. These calculated limits served as convenient starting points in selecting a specific style of dowel and assisted in determining whether additional measures are warranted to enhance dowel retention.