Browsing by Subject "TLR"

Now showing 1 - 2 of 2
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    Antibodies In Cancer Therapy: New Targets, Applications And Combination Strategies
    (2019-07) Khanna, Vidhi Devendra
    Over the last decade, antibodies have become an important component in the arsenal of cancer therapeutics. Their high-specificity, low-off target effects, desirable pharmacokinetics and high success rate are a few of the many attributes that make them amenable for development as drugs. The work presented here explores the targeting, mechanisms and use of antibody-based cancer therapy. In the first chapter, we used a phage display-based cell panning procedure to develop two fully humanized antibodies, Tw1S4_6 and Tw1S4_AM6, that bind specifically to HSPG2/perlecan, a protein found to be overexpressed on tumor cells. Immunohistochemistry studies revealed high HSPG2 expression across various tumor sub-types including melanoma, bladder cancer, glioblastoma and ovarian cancer. There was significant correlation between high HSPG2 expression and poor survival in triple negative breast cancer, bladder and ovarian cancers. The data presented here points towards the relevance of HSPG2 as a novel target for not only triple negative breast cancer but other malignancies as well. Based on its over-expression in different solid tumors, we evaluated HSPG2 as a therapeutic target in the second chapter. We observed significant tumor growth inhibition with Tw1S4_AM6 in the triple negative MDA-MB-231-LM2 breast cancer xenograft model. This efficacy was reduced in NSG mice, suggesting NK cell-mediated antibody dependent cellular cytotoxicity (ADCC) as a potential mechanism of action. In vitro studies using human PBMCs confirmed induction of ADCC with anti-HSPG2 antibodies. In addition, conjugation of Tw1S4_AM6 on the surface of polymeric nanoparticles enabled increased tumor cell uptake of nanoparticles, suggesting Tw1S4_AM6 could be valuable as a targeting ligand for drug delivery systems. There is a significant interest in designing therapeutic agents that can enhance ADCC and thereby improve clinical responses with approved antibodies. We have developed a suite of highly substituted imidazoquinolines, which activate TLR 7 and/or 8 and induce significantly higher levels of cytokines compared to the FDA-approved TLR7 agonist, imiquimod. In the third chapter, we evaluated our series of TLR7/8 agonists for their ability to improve ADCC. Our studies show that the second generation TLR 7/8 agonists induce robust pro-inflammatory cytokine secretion and activate NK cells. These agonists also enhanced ADCC in vitro. Finally, we found that these agonists significantly improved the anticancer efficacy of two monoclonal antibodies in vivo. Thus, the work presented here encompasses the three critical aspects of antibody therapeutics: identifying the target, understanding their mechanisms, and leveraging these mechanisms to improve their efficacy.
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    Identifying Mechanisms And Biomarkers Predictive Of Efficacy Of Vaccines Against Opioid Use Disorders And Overdose
    (2022-08) Crouse, Bethany
    Opioid use disorders (OUD) and overdose are public health crises that are worsening despite the availability of approved pharmacotherapies. Active immunization with anti-opioid conjugate vaccines is a novel therapeutic strategy to treat OUD and prevent overdose. To date, clinical studies suggest that efficacy of anti-drug conjugate vaccines is limited to a subset of individuals who can produce optimal antibody responses. To increase positive treatment outcomes and clinical success, this research program investigated several complementary strategies to increase OUD vaccine efficacy. First, mechanisms of optimal anti-opioid vaccine response are investigated by elucidating the immunological mechanisms behind a previously established interleukin-4 (IL-4) mediated increase in vaccine efficacy. These studies found that depletion of IL-4 resulted in a Type I IL-4R mediated increase in germinal center formation and germinal center T cell response which leads to increased opioid-specific antibody secreting cells, and that vaccine efficacy is dependent on a balanced Th1/Th2 T cell response in mice. Next, these results provided a blueprint for next generation anti-fentanyl vaccine formulations incorporating novel adjuvants targeting toll-like receptors (TLRs). These data show that a TLR7/8 agonist adjuvant increases vaccine efficacy in rodent and porcine models of fentanyl misuse and overdose. Third, vaccine design and immunization paradigms were assessed to optimize the efficacy of a novel carfentanil vaccine alone and in combination with a lead fentanyl vaccine. Longer linker lengths and a co-administered bivalent immunization strategy were associated with increased vaccine efficacy. Then, environmental factors contributing to the immune response are investigated by testing whether changes in the gastrointestinal microbiome would affect vaccine efficacy and whether these specific changes in the microbiome could be utilized as biomarkers. These studies revealed that changes in the microbiome in specific pathogen free or immune-experienced rodents did not affect efficacy of anti-oxycodone or anti-fentanyl vaccines. Finally, exploratory studies were performed to identify putative biomarkers that may be predictive of anti-opioid vaccine response in preclinical and clinical investigations. These studies indicate that pre-immunization concentration of IL-4 is correlated with vaccine efficacy in genetically diverse mice, and that specific cytokines may be of interest as indicators of immune response in human patients. Overall, the findings outlined in this research program support the use of novel adjuvants and predictive biomarkers to increase clinical efficacy of vaccines to treat OUD and overdose.