The humanized mouse model of HIV latency as a preclinical tool for therapeutic vaccine and anti-latency agent development.

Combined antiretroviral therapy (cART) has dramatically improved the clinical outcome of HIV-1 infected patients. Effective cART leads to suppression of viral replication thereby preventing viral transmission, partial restoration of the immune function, improved quality of life and return to a near-normal life expectancy (Katlama C, Lancet 2013). Nevertheless, successful cART requires lifetime adherence, has no effect on the eradication of viral reservoirs and does not restore an effective virus-suppressive immune response against HIV (Siliciano et al., Nat Med 2003; Plana et al., Lancet 1998). Treatment resistance, adverse effects in the medium-long term and the high financial cost of cART are important limitations for lifelong adherence to this therapy. Therefore, an urgent need exists for developing an HIV cure overcoming the current limitations of cART. This could be achieved either by completely clearing the virus from the host (sterilizing cure) or by inducing host-mediated control of viral replication in the absence of cART (functional cure).

Research on HIV pathogenesis and development of new therapeutic strategies have long been hampered by the lack of robust, reproducible and reliable preclinical models of HIV infection. Since small animal models are easier to house and more cost-effective than primate models, considerable efforts have been made to reconstitute a functional human immune system in mice and thus make them permissive for HIV infection. In this regard, the Department of Infection and Immunity (DII) of the Luxembourg Institute of Health (LIH) has gained expertise in the generation and HIV infection of humanized NOD/LtszscidIL2null strain (NSG mice) in collaboration with Pr Moutschen, University of Liège, Belgium, to obtain a full human immune system including CD4 and CD8 T cells, CD19 B cells, CD56 CD16 NK cells, CD14 monocytes and CD11c dendritic cells.
Regarding HIV infection, this humanized mice model supports long-term viral replication after infection by both the intravenous and rectal routes and replicates many features of human infection and disease state. (Singh et al, plos one 2012). Besides human CD4 depletion and decrease of the human CD4/CD8 ratio in HIV-infected mice, we have shown an increased expression of CD70+ CD4+ T cells and homing properties of immature transitional B cells (IT B cells) similarly to lymphopenic or viremic individuals (Lantto et al., AIDS in press and Amu et al., in preparation).
174 to 195 days post-transplantation humanized mice were infected with HIV-1 (strain BaL-1, 10000 TCID50). Three months post-infection, tissues and blood samples were processed immediately upon sacrifice. Plasma was separated from blood cells by centrifugation and frozen for further quantification of the viral load using the Abbott RealTime™ HIV-1 assay. Lymph nodes, spleen and bone marrow were dissociated with syringes and passed through a nylon cell strainer to obtain single-cell suspensions for flow cytometry.
Our goal is now to optimize an HIV latency model for the design of therapeutic vaccines and therapies targeting the long-lived reservoir of HIV infection in resting CD4+T cells. The use of single tablet regimen for once-daily treatment containing an integrase inhibitor, highly potent in reducing viral load, would be highly beneficial to treat humanized mice when administrated in the food. Recent clinical studies have demonstrated that early cART can reduce the size of the viral reservoir and delay or reduce viral rebound following cART discontinuation in humans (Ananworanich J 2012, Hocqueloux 2013) and in SIV-infected rhesus monkeys (Whitney JB, 2014).
To validate this model of HIV latency we will
1) determine if HIV could establish a latent reservoir by confirming the presence of resting human T cells in cells from peripheral blood, bone marrow, spleen, lymph nodes, liver, lung, and thymic organoid of HIV+ ART suppressed mice. Replication-competent virus will be further recovered from pooled resting CD4 T cells in individual mice using the viral outgrowth assay.
2) compare the impact of cART administered during the primary phase of the infection and during the chronic phase of the infection on different parameters:
- establishment of viral reservoirs in resting CD4+T cells (total HIV DNA, episomal 2-LTR circles, un-integrated HIV and unspliced HIV RNA) using digital droplet PCR (Malatinkova et al., Kiselinova et al.) in collaboration with Pr. L.Vandekerkhove at the University Hospital of Ghent
- specific markers of HIV pathogenesis (immune activation, inflammation, microbial translocation, mucosal degradation)
- delayed/reduced viral rebound following cART interruption