SBIR-STTR Award

The development of a small animal model to study human immunodeficiency virus type-1 (HIV-1) infection would significantly facilitate studies on disease pathogenesis, as well as vaccine and anti-viral drug development and testing. However, HIV-1 replication is subjected to a number of species-specific restrictions at the level of cellular entry and/or post-entry. To date, no satisfactory small animal model for HIV-1 infection has been identified. The cotton rat has been a superb model for human infectious diseases. This animal is susceptible to an extraordinary spectrum of human pathogens, particularly viruses. Inspired by these observations, it has been cloned and characterized a battery of more than 20 cotton rat genes of immunological and inflammatory importance, and reagents for their detection has been developed. Human immunodeficiency virus (HIV-1) was shown to infect two species of cotton rats, Sigmodon hispidus and S. fulviventer and infectious virus was transmitted from animal to animal by blood. In new studies it was found that cotton rat cells (primary macrophages and a cotton rat osteosarcoma cell line) after transfection with a plasmid containing the backbone genome of HIV-1 support levels of HIV transcription analogous to those observed in human monocytes, indicating the absence of transcription blockage. Additionally, cotton rat cells became permissive to a HIV-1 pseudotyped infection when they transiently co-expressed human CD4 and CCR5 or CXCR4 chemokine receptors. The overall goal of this proposal is to generate a small animal model for HIV-1 infection by generating transgenic cotton rats expressing HIV-1 co-receptors. An HIV-1-permissive cotton rat could be widely used by the research community. At the completion of the exploratory experiments proposed for phase I of this SBIR, we will know the potential of the cotton rat as a transgenic model for HIV-1 studies, and whether is worth the time and expense to develop transgenic animals in a phase II study.

Thesaurus Terms:
HIV infection, disease /disorder model, human immunodeficiency virus 1, laboratory rat, model design /development, transgenic animal, virus receptor cytokine receptor, gene targeting, genetic transcription, virus infection mechanism, virus replication

The development of a small animal model to study human immunodeficiency virus type-1 (HIV-1) infection would significantly facilitate studies of disease pathogenesis, as well as vaccine and anti-viral drug development and testing. However, HIV-1 replication is subjected to a number of species-specific restrictions at the level of cellular entry and/or post-entry. To date, no satisfactory small animal model for HIV-1 infection has been identified. The cotton rat has been a superb model for human infectious diseases. This animal is susceptible to an extraordinary spectrum of human pathogens, particularly viruses. Inspired by these observations, we have cloned and characterized a battery of more than 270 cotton rat genes of immunological and inflammatory importance, and reagents for their detection have been developed. Human immunodeficiency virus (HIV-1) was shown to infect cotton rats and infectious virus was transmitted from animal to animal by blood with low efficiency. In new studies developed during the Phase I of this SBIR, we have demonstrated that cotton rat cells expressing human co-receptors for HIV-1 (hCD4 with hCXCR4 or hCD4 with hCCR5) support HIV-1 infection (by kinetic experiments that measured the production of p24gag in cotton rat infected cells) and DNA integration (by cloning experiments that isolated chimeric DNA containing HIV-1 DNA sequences linked to cotton rat DNA sequences from infected cotton rat cells). Furthermore, we have shown that cotton rat cells are able to produce infective particles that can infect fresh human PBMCs. The data presented in our Progress Report indicate that the main blockage for HIV replication in cotton rat cells occurs during viral entrance and that it could be circumvented by the expression of HIV-1 co-receptors. Due to our success in demonstrating the feasibility of the model in vitro during phase I, our goal in phase II is to produce line(s) of transgenic cotton rats expressing hCD4 with hCXCR4 and hCD4 with hCCR5 molecules and test them for their infectivity to HIV-1. An HIV-1-permissive cotton rat could be widely used by the research community. The hypothesis to be tested is that cotton rats (S. hispidus) after engineered to produce HIV-1 co-receptors will be permissive to productive HIV-1 infection. We will test our hypothesis by producing lines of transgenic animals expressing hCD4 with hCCR5 and hCD4 with hCXCR4. Finaly, we will Test these transgenic lines of cotton rats in infection protocols using HIV-1. At the completion of the work proposed for phase II of this SBIR, we will have produced several lines of transgenic cotton rat expressing human co-receptors for HIV-1. Most importantly, we will have characterized these lines of animals in ex vivo and in vivo experiments An estimated 5 million people became infected with HIV worldwide in 2003, and as many as 3 million died from AIDS, according to the Joint United Nations Program on HIV/AIDS (UNAIDS). The total number of infected people worldwide is estimated at 40 million (and 600 new infections per hour). Preventive vaccine and prophylactic therapies against HIV infection and more efficient drugs for AIDS are not only urgently needed but they are also scientifically possible. However, one of the major obstacles in translational HIV research rests in the absence of inexpensive and efficient pre-clinical trial models. This is reflected in the fact that the 30 vaccine candidates that are currently being tested in clinical trials in 21 countries (only 2 have advanced to clinical phase III efficacy trials) are very similar to each other with nearly all based on only one hypothesis (protection by eliciting a cell mediated immune response), while other strong alternative hypotheses have been largely neglected (vaccines that induce neutralizing antibodies and live attenuated vaccines) due to monetary and logistic impediment to test a greater number of candidates in preclinical settings. Thus, several potential vaccines and therapeutics turn out to be buried in laboratory notebooks because the institutions where they were developed cannot afford to continue with non-human primate trials. The lack of a small animal model for HIV infection that can be used for screening a larger variety of candidates in preclinical studies is one of the most evident obstacles to speed up the process of developing HIV vaccines and AIDS therapies