Engineering an HIV-resistant immune system for HIV cure

? DESCRIPTION (provided by applicant): The candidate, Hind Fadel, M.D., Ph.D., is an Assistant Professor at the Mayo Clinic College of Medicine. She is a member of the Division of Infectious Diseases in the Department of Medicine, and of the Department of Molecular Medicine at the Mayo Clinic. Dr. Fadel is a physician-scientist committed to a career in basic and translational HIV-1 research and gene therapy. She has gathered an experienced team of mentors, collaborators and advisors and is proposing a comprehensive research, course work, and career development plan that will position her to be a successful independent investigator. The environment provided by the Mayo Clinic is ideally suited for the success of both the research and career development components of the award. She has also maintained and identified advisors and collaborators outside Mayo Clinic. The career development plan will allow her to hone her skills in genome editing with engineered site-specific nucleases, to gain insights into host factors role in HIV-1 biology, and to develop new skills to manipulate human genome for HIV-1 cure applications and gene therapy. The research plan focuses on developing HIV cure strategies that use targeting/editing of HIV-1 dependency and restriction factor genes with novel engineered nucleases in order to create HIV-1 resistance. The methods will be applied to primary CD34+ hematopoietic stem cells (HSCs) and studied in humanized mice. While antiretroviral drugs have made treatment of HIV disease possible, they in many ways represent a halfway technology. They leave existing patients vulnerable to major problems that include resistance, persistent immune dysfunction and immunosenescence, complex metabolic disturbances, and accelerated aging phenomena. The second problem is that while drug treatment can in theory prevent further transmission, the many problems associated with supplying the pill combinations life-long and with maintaining adherence have meant that the epidemic continues to expand. A definitive cure could solve both problems. Cure was until recently considered futuristic, but this has now changed. The recent cure of a patient with both HIV and leukemia -- the so-called Berlin patient -- by bone marrow transplantation from a donor lacking a main HIV cellular cofactor (the entry co-receptor CCR5) was energizing to the field. The specific approach (allogeneic BMT) is far too toxic for anyone without cancer. However, the value of targeting a gene that HIV needs (a host cofactor) has been made clear. Along with the very recent emergence of highly promising gene targeting technologies, and advances in our understanding of the disease process itself, the Berlin patient helped catalyze making cure a central priority in the field. Modification of a patient's own HSCs to regenerate an HIV- resistant immune system in subjects with HIV/AIDS, coupled with appropriately mild conditioning, could be an effective cure strategy. It is essential to recruit and develop young investigators to populate what is certain to be an important, expanding field. Dr. Fadel is proposing to target two vulnerabilities of HIV-1 as she develops curative gene therapy approaches: (i) eliminating host dependency factors HIV-1 requires, and (ii) editing host restriction factors that HIV-1 has evolved to evade in order to restore their ability to restrict HV-1. Her central hypothesis is that engineered nucleases will efficiently target HIV-1 dependency and restriction factors to generate resistance to HIV-1 infection for therapeutic application and t further our understanding of these factors in HIV-1 pathogenesis. The main aims are: 1) Knockout HIV-1 dependency factors in human CD4+ T-cell lines to simultaneously determine their virological roles and establish their potential for therapeutic targets for HIV cure applications; 2) Edit human restriction factors to restore HIV-1 resistance and determine effects on viral replication; 3) Create dependency factor knockouts and restriction factor knock-ins in CD34+ HSC cells ex-vivo and characterize them in pre-clinical animal studies in humanized mice. The proposed research has the potential to provide insights into the roles of multiple host factors in HIV-1 biology and to lead to translational applications for curative HIV-1 therapies. Developing HSC-mediated therapeutics in this disease can also advance related applications to other infectious, autoimmune and hematologic diseases.