Anti-Candida activity of CCL28 in oropharyngeal candidiasis

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PROJECT SUMMARY/ABSTRACT I have the ideal level of experience and institutional environment to maximally benefit from a Career Development Award. My research experiences started as an undergraduate when I was part of a research team who solved an RNA structure using NMR spectroscopy. I followed this project with a Cancer Research Training Award and an intensive research experience at the NIH. After focusing on clinical care during medical school and pediatric residency training, I returned to basic science research as a pediatric infectious diseases fellow. I was granted a prestigious Pediatric Infectious Diseases Society Fellowship Award which supported my fellowship research and a 4th year of mentored research as junior faculty. Mentored by Dr. Sarah Gaffen, I contributed to studies on the innate and adaptive host responses to mucosal Candida infections and initiated work on the relationship of neutrophils and IL-17 in oropharyngeal candidiasis. I also investigated the contribution of IL-17 to fungal burden and disease pathology in disseminated candidiasis. In all, I am an author on 9 manuscripts relating to candidiasis and/or IL-17 of which 4 are first authorships. My research projects have continued since starting as an assistant professor at the Medical College of Wisconsin (MCW) in August, 2014. The environment at MCW provides an outstanding venue for the proposed project. In particular, the vibrant campus Immunology group encourages collaboration and interaction. Although I continue to collaborate with Dr. Gaffen, I have established relevant mentorship for my current project with experts in chemokine structural biology and mucosal immunology (Drs. Brian Volkman and Mitchell Grayson, respectively). My laboratory participates in immunology, microbiology, and infectious diseases seminars. I have excellent lab space, ample support for animal studies, and access to all equipment and core resources required for the completion of this project including flow cytometers, histology services, and biostatistics support through the MCW Children’s Research Institute. My overall career goal is to be an independent, NIH-funded physician scientist focused on translational research in mucosal immunology as it relates to fungal infections. I am guided by my advisory Mentorship Committee, composed of local experts in mucosal immunology, structural biology, antimicrobial peptides, cellular immunology, chemokines, and pathogenesis (mentors Volkman and Grayson, as well as Drs. Nita Salzman, Bonnie Dittel, Michael Dwinell, and Jenifer Coburn). I plan training and mentorship activities to augment my knowledge of the responsible conduct of research, manuscript and grant reviews, and research- related topics including tissue culture, chemotaxis, rodent necropsy, histological techniques, mucosal immunology, and biostatistics. During the latter two years of the award period I intend to write and submit a successful NIH R01 application. In terms of ongoing guidance and evaluation, I will meet with each of my mentors independently biweekly and with my Mentorship Committee quarterly to ensure both achievement of research goals and growth towards independence. In the current proposal, we aim to investigate the function of the chemokine CCL28 in oral mucosal immunity from fungal infection. Oropharyngeal candidiasis (OPC) is a frequent and serious problem for immunocompromised individuals and a marker of risk for the development of high-mortality disseminated candidiasis in patients with barrier and immune deficits. CCL28 is a CC chemokine with in vitro antimicrobial peptide (AMP) activity, chemotactic activity for lymphocytes and eosinophils, and mucosal localization. Our approach and rationale for investigating CCL28 in mucosal host defense from infection is based on preliminary data on the anti-Candida motifs and gene expression in OPC. CCL28’s activities potentially fill gaps in the current model of OPC host defense which includes pro-inflammatory cytokines, cellular recruitment, and AMPs. We propose to study the function of CCL28 using constructed variants with abolished AMP or chemotactic activity. We hypothesize that CCL28 controls Candida at the mucosal interface directly as an AMP and indirectly through induction of neutrophil chemoattractants. The aims take an innovative approach of applying structural biology tools to determine the mechanisms of small immune proteins and their role in limiting infection. In Aim 1, we will characterize the regulation of chemokine and cytokine expression by CCL28 and determine the active motif for each distinct function. From the studies in Aim 2, we will determine the activity and potency of each CCL28 function in the mouse model of OPC. Using structure-function data to maximize CCL28’s potency in OPC while minimizing pathologic inflammatory activity will allow us to harness the biologic properties of CCL28 and expand understanding of immune mechanisms. Overall, this K award will allow me to gain independence in structure-function analysis of molecules relevant to mucosal host defense. I will be able to apply these skills to obtain future R01 funding as an independent investigator on the host immune response to fungal infections, including AMPs and chemokines, in order to improve recognition of risk factors for infection and develop new therapeutics.