Adam Grippin

Effects of Relative Humidity and Spraying Medium on UV Decontamination of Filters Loaded with Viral Aerosols

ABSTRACT Although respirators and filters are designed to prevent the spread of pathogenic aerosols, a stockpile shortage is anticipated during the next flu pandemic. Contact transfer and reaerosolization of collected microbes from used respirators are also a concern. An option to address these potential problems is UV irradiation, which inactivates microbes by dimerizing thymine/uracil in nucleic acids. The objective of this study was to determine the effects of transmission mode and environmental conditions on decontamination efficiency by UV. In this study, filters were contaminated by different transmission pathways (droplet and aerosol) using three spraying media (deionized water [DI], beef extract [BE], and artificial saliva [AS]) under different humidity levels (30% [low relative humidity {LRH}], 60% [MRH], and 90% [HRH]). UV irradiation at constant intensity was applied for two time intervals at each relative humidity condition. The highest inactivation efficiency (IE), around 5.8 logs, was seen for DI aerosols containing MS2 on filters at LRH after applying a UV intensity of 1.0 mW/cm2 for 30 min. The IE of droplets containing MS2 was lower than that of aerosols containing MS2. Absorption of UV by high water content and shielding of viruses near the center of the aggregate are considered responsible for this trend. Across the different media, IEs in AS and in BE were much lower than in DI for both aerosol and droplet transmission, indicating that solids present in AS and BE exhibited a protective effect. For particles sprayed in a protective medium, RH is not a significant parameter.

Use of dialdehyde starch treated filters for protection against airborne viruses

Abstract In the event of a pandemic, the general public would use filters as protective devices. However, most commercial filters only remove airborne viruses physically without inactivating them, allowing reproduction on the surface and yielding the mask as a fomite. The objective of this study was to investigate the inactivation performance of dialdehyde starch (DAS) treated filters against airborne viruses. The viable removal efficiency by and relative survivability on the biocidal filters prepared with dialdehyde starch compared to untreated filters were investigated using MS2 bacteriophage at high relative humidity (80–90%) and room temperature. Experimental results showed no significant difference in viable removal efficiency and pressure drop between the treated and untreated filters for polypropylene filtering facepiece respirators. The pressure drop of DAS treated cellulose filters significantly decreased although there was no significant change in viable removal efficiency; the combination of these two factors resulted in an increase of filter quality. All biocidal filters showed a significantly lower relative survivability than untreated filters, and the relative survivability decreased as the concentration of DAS increased. The biocidal filter treated with 4% DAS presented an average of 30% survivability compared to the baseline of untreated filters. The results demonstrate that dialdehyde starch can be incorporated onto filters to provide an effective means for inactivating MS2 viruses through surface contact.

Systemic activation of antigen-presenting cells via RNA-loaded nanoparticles

ABSTRACT While RNA-pulsed dendritic cell (DC) vaccines have shown promise, the advancement of cellular therapeutics is fraught with developmental challenges. To circumvent the challenges of cellular immunotherapeutics, we developed clinically translatable nanoliposomes that can be combined with tumor-derived RNA to generate personalized tumor RNA-nanoparticles (NPs) with considerable scale-up capacity. RNA-NPs bypass MHC restriction, are amenable to central distribution, and can provide near immediate immune induction. We screened commercially available nanoliposomal preparations and identified the cationic lipid 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP) as an efficient mRNA courier to antigen-presenting cells (APCs). When administered intravenously, RNA-NPs mediate systemic activation of APCs in reticuloendothelial organs such as the spleen, liver, and bone marrow. RNA-NPs increase percent expression of MHC class I/II, B7 co-stimulatory molecules, and maturation markers on APCs (all vital for T-cell activation). RNA-NPs also increase activation markers on tumor APCs and elicit potent expansion of antigen-specific T-cells superior to peptide vaccines formulated in complete Freunds adjuvant. We demonstrate that both model antigen-encoding and physiologically-relevant tumor-derived RNA-NPs expand potent antitumor T-cell immunity. RNA-NPs were shown to induce antitumor efficacy in a vaccine model and functioned as a suitable alternative to DCs in a stringent cellular immunotherapy model for a radiation/temozolomide resistant invasive murine high-grade glioma. Although cancer vaccines have suffered from weak immunogenicity, we have advanced a RNA-NP formulation that systemically activates host APCs precipitating activated T-cell frequencies necessary to engender antitumor efficacy. RNA-NPs can thus be harnessed as a more feasible and effective immunotherapy to re-program host-immunity.

Cross-talk between T cells and hematopoietic stem cells during adoptive cellular therapy for malignant glioma

Purpose: Adoptive T-cell immunotherapy (ACT) has emerged as a viable therapeutic for peripheral and central nervous system (CNS) tumors. In peripheral cancers, optimal efficacy of ACT is reliant on dendritic cells (DCs) in the tumor microenvironment. However, the CNS is largely devoid of resident migratory DCs to function as antigen-presenting cells during immunotherapy. Herein, we demonstrate that cellular interactions between adoptively transferred tumor-reactive T cells and bone marrow–derived hematopoietic stem and progenitor cells (HSPCs) lead to the generation of potent intratumoral DCs within the CNS compartment. Experimental Design: We evaluated HSPC differentiation during ACT in vivo in glioma-bearing hosts and HSPC proliferation and differentiation in vitro using a T-cell coculture system. We utilized FACS, ELISAs, and gene expression profiling to study the phenotype and function of HSPC-derived cells ex vivo and in vivo. To demonstrate the impact of HSPC differentiation and function on antitumor efficacy, we performed survival experiments. Results: Transfer of HSPCs with concomitant ACT led to the production of activated CD86+CD11c+MHCII+ cells consistent with DC phenotype and function within the brain tumor microenvironment. These intratumoral DCs largely supplanted abundant host myeloid-derived suppressor cells. We determined that during ACT, HSPC-derived cells in gliomas rely on T-cell–released IFNγ to differentiate into DCs, activate T cells, and reject intracranial tumors. Conclusions: Our data support the use of HSPCs as a novel cellular therapy. Although DC vaccines induce robust immune responses in the periphery, our data demonstrate that HSPC transfer uniquely generates intratumoral DCs that potentiate T-cell responses and promote glioma rejection in situ. Clin Cancer Res; 24(16); 3955–66. ©2018 AACR.

Translational nanoparticle engineering for cancer vaccines

ABSTRACT Conventional cancer treatments remain insufficient to treat many therapy-resistant tumors.1 Cancer vaccines attempt to overcome this resistance by activating the patients immune system to eliminate tumor cells without the toxicity of systemic chemotherapy and radiation. Nanoparticles (NPs) are promising as customizable, immunostimulatory carriers to protect and deliver antigen. Although many NP vaccines have been investigated in preclinical settings, a few have advanced into clinical application, and still fewer have demonstrated clinical benefit. This review incorporates observations from NP vaccines that have been evaluated in early phase clinical trials to make recommendations for the next generation of NP-based cancer vaccines.

Detachment of Liquid-Water Droplets from Gas-Diffusion Layers

A critical issue for optimal water management in proton-exchange-membrane fuel cells at lower temperatures is the removal of liquid water from the cell. This pathway is intimately linked with the phenomena of liquid-water droplet removal from surface of the gas-diffusion layer and into the flow channel. Thus, a good understanding of liquid-water transport and droplet growth and detachment from the gas-diffusion layer is critical. In this study, liquid-water droplet growth and detachment on the gas-diffusion layer surfaces are investigated experimentally to improve the understating of water transport through and removal from gas-diffusion layers. An experiment using a sliding-angle measurement is designed and used to quantify and directly measure the adhesion force for liquid-water droplets, and to understand the droplets? growth and detachment from the gas-diffusion layers.

Personalized Tumor RNA Loaded Lipid-Nanoparticles Prime the Systemic and Intratumoral Milieu for Response to Cancer Immunotherapy

Translation of nanoparticles (NPs) into human clinical trials for patients with refractory cancers has lagged due to unknown biologic reactivities of novel NP designs. To overcome these limitations, simple well-characterized mRNA lipid-NPs have been developed as cancer immunotherapeutic vaccines. While the preponderance of RNA lipid-NPs encoding for tumor-associated antigens or neoepitopes have been designed to target lymphoid organs, they remain encumbered by the profound intratumoral and systemic immunosuppression that may stymie an activated T cell response. Herein, we show that systemic localization of untargeted tumor RNA (derived from whole transcriptome) encapsulated in lipid-NPs, with excess positive charge, primes the peripheral and intratumoral milieu for response to immunotherapy. In immunologically resistant tumor models, these RNA-NPs activate the preponderance of systemic and intratumoral myeloid cells (characterized by coexpression of PD-L1 and CD86). Addition of immune checkpoint inhibitors (ICIs) (to animals primed with RNA-NPs) augments peripheral/intratumoral PD-1+CD8+ cells and mediates synergistic antitumor efficacy in settings where ICIs alone do not confer therapeutic benefit. These synergistic effects are mediated by type I interferon released from plasmacytoid dendritic cells (pDCs). In translational studies, personalized mRNA-NPs were safe and active in a client-owned canine with a spontaneous malignant glioma. In summary, we demonstrate widespread immune activation from tumor loaded RNA-NPs concomitant with inducible PD-L1 expression that can be therapeutically exploited. While immunotherapy remains effective for only a subset of cancer patients, combination therapy with systemic immunomodulating RNA-NPs may broaden its therapeutic potency.

Poly(Lactic Acid) Magnetic Microparticle Synthesis and Surface Functionalization

Iron oxide nanoparticles have been employed for a variety of biomedical applications in both the diagnostic and therapeutic realm, and the number of applications continues to grow. In this letter, magnetic nanoparticles were synthesized via aqueous coprecipitation and subsequently coated in oleic acid to confer colloidal stability in organic solvents. The synthesized nanoparticles were encapsulated within poly(lactic acid) via a double-emulsion synthesis. Following the microparticle synthesis, surface functionalization experiments were performed to gain an understanding of the surface chemistry and the potential ability to conjugate biomacromolecules to the particle surface for use in biomedical applications. The ability to specifically bind to histidine-tagged proteins was assessed through the flow cytometry analysis of histidine-tagged green fluorescent protein after incubation with the particles. The magnetic separation of proinflammatory proteins was assessed using enzyme-linked immunosorbent assays to monitor cytokine depletion from known concentrations of tumor necrosis factor-alpha. Lastly, a fluorescently tagged protein molecule was bound to the surface, which allowed the particles to be observed visually under a fluorescent microscope. The particles described here have the potential for numerous biomedical applications including biomacromolecule isolation, cell separation, heavy metal toxin removal, and as a fluorescent imaging modality.