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Author ORCID Identifier

https://orcid.org/0000-0002-3188-7455

AccessType

Campus-Only Access for Five (5) Years

Document Type

dissertation

Degree Name

Doctor of Philosophy (PhD)

Degree Program

Animal Biotechnology & Biomedical Sciences

Year Degree Awarded

2022

Month Degree Awarded

September

First Advisor

Ashish Kulkarni

Subject Categories

Biochemical and Biomolecular Engineering | Biomaterials | Biotechnology | Immunotherapy

Abstract

Chronic inflammatory disorders such as colitis, Alzheimer’s, arthritis, cardiovascular diseases contribute to about 60% of deaths worldwide. Such lifelong debilitating illness degrades the quality of life for patients and their caregivers while contributing to enormous social and economic burdens often associated with high costs for therapy and care. According to CDC, about 90% of the nation’s total health care costs contribute to chronic illnesses. This suggests that there’s an unmet need for advanced tools and techniques for the improvement of current diagnostic and therapeutic procedures. Inflammasomes have been extensively shown to be associated with the pathogenesis of various inflammatory diseases. It’s a form of innate immune response which is triggered when any form of pathogenic or sterile insult is recognized by the cytosolic threat sensors. This further trigger the assembling of three different proteins including sensor, adaptor and zymogen into the multimeric inflammasome protein complex which immediately leads to proximity-induced cleavage of caspase-1 zymogen into its active form. Active caspase-1 enzyme cleaves the downstream modulators, pore-forming protein gasdermin D, and procytokines into their active forms which subsequently leads to a specialized form of cell death, called pyroptosis, causing enhanced inflammation. Inflammasome is a host defense mechanism that, upon dysregulation, leads to several disorders. Thus, inflammasome signaling components could act as a common target for devising efficient diagnostic and therapeutic approaches for various inflammatory disorders. This thesis is focused on developing nanoplatforms, offering inflammasome-specific diagnostics and therapeutic opportunities, with proof of principal applications demonstrated in inflammatory disease models like gouty arthritis, colitis, and sepsis. We first aimed at understanding the interactions of nanomaterial and immune cells to identify the inflammasome activating nanoparticle-associated-molecular-patterns. This study identified nanoparticle core hydrophobicity as a core and tunable determinant of inflammasome activation. We further demonstrated that nanoparticle core hydrophobicity can be tuned to obtain different degrees of inflammasome assembly via distinct signaling inclinations and kinetics. This knowledge could be utilized for a myriad of applications, viz, the particles that don’t activate inflammasome could be utilized for immunotherapy as well as diagnostics, and the particles activating inflammasome can be employed as vaccine adjuvants or for prophylactic responses where the controlled inflammasome activation is desired. In the two follow-up studies, we developed reporter and theranostic nanoplatforms that can monitor inflammasome activation in real-time and offer simultaneous therapeutic benefits. We engineered two pair of nanoparticles termed as MATIN I and MATIN II, where MATIN corresponds to Monitoring and Therapeutics of Inflammasome via Nanoparticles. Each pair included a reporter and a theranostic platform encapsulating either just the caspase-1 responsive probe or an additional inflammasome inhibiting drug. Both the MATIN I and II platforms offer robust monitoring as well as therapeutic potential in-vitro in nigericin-treated immortalized macrophages and in-vivo in inflammatory disease models of gouty arthritis and DSS-induced colitis. The final study aimed at developing an advanced synergistic therapeutic platform by codelivering rational combinations of inflammasome-inhibiting drugs via nanoparticles. Here we synthesized and characterized a nanoparticle co-encapsulating two potent inflammasome inhibitors, MCC-950 and disulfiram. These dual-drug nanoparticles exhibited a synergistic response and significantly higher efficiency in-vitro when compared to single-drug nanosystems as well as free drugs. Additionally, they offered complete protection against LPS-mediated sepsis model, proving their heightened in-vivo efficacy. Overall, this thesis aims to serve as a guide to developing platforms that can offer inflammasome-specific diagnostics and therapeutics as well as leverage them for prophylactic responses to obtain the desired level of inflammation for host defense.

DOI

https://doi.org/10.7275/30257773

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