This proposal focuses on the therapeutic potential of RNA interference (RNAi) in preventing cervicovaginal HIV-1 transmission. RNAi refers to the sequence-specific degradation of RNA that follows the cellular introduction of homologous, short interfering (si) RNA. RNAi has emerged as a powerful tool to probe the function of genes of known sequence in vitro and in vivo. Advances in vector design permit the expression of siRNA in human cells by the transfer of DNA or viral vectors encoding short hairpin RNA (shRNA) cassettes. Investigators have proven the principle that RNAi can be used to inhibit the expression of a wide range of disease-specific genes. The majority of these investigations have been limited to in vitro systems. We hypothesize that short interfering RNA (siRNA) targeting human CCR5 can be effectively targeted to the cervicovaginal epithelia, specifically knock down chemokine expression and thereby prevent HIV-1 transmission. Our enthusiasm for this approach derives from preliminary data demonstrating that siRNA can be specifically targeted to murine cervicovaginal tissue in vivo using liposomal formulations. The siRNAs are efficiently taken up by cells-along the full thickness of the vagina and mediate durable and specific gene silencing. We wish to expand upon these promising results and apply them to models of HIV-1 transmission. Using a small animal model, we will pursue three goals:
Specific Aim 1 : To determine whether siRNA mediated knockdown of vaginal CCR5 expression prevents HIV-1 infection in the murine xenograft model of HIV-1 transmission.
Specific Aim 2 : To determine whether vaginal application of siRNA targeting HIV-1 reduces genital virus burden in vivo.
Specific Aim 3 : To evaluate the utility of adeno-associated virus as an HIV- RNAi delivery vector for cervicovaginal epithelia. In sum, our research will define the microbicidal potential of si/shRNA and may lead to the generation of new drugs that prevent the mucosal transmission of HIV-1. ? ? ? ?
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| Tang, Xiaoli; Li, Ming; Tucker, Lynne et al. (2011) Glycogen synthase kinase 3 beta (GSK3?) phosphorylates the RNAase III enzyme Drosha at S300 and S302. PLoS One 6:e20391 |
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| Tang, Xiaoli; Zhang, Yingjie; Tucker, Lynne et al. (2010) Phosphorylation of the RNase III enzyme Drosha at Serine300 or Serine302 is required for its nuclear localization. Nucleic Acids Res 38:6610-9 |
| Boden, Daniel; Pusch, Oliver; Ramratnam, Bharat (2007) Overcoming HIV-1 resistance to RNA interference. Front Biosci 12:3104-16 |
| Zhang, Yingjie; Cristofaro, Patricia; Silbermann, Rebecca et al. (2006) Engineering mucosal RNA interference in vivo. Mol Ther 14:336-42 |
