The naturally-occurring RNA interference (RNAi) pathway represents a powerful tool for the sequence-specific post-transcriptional silencing of gene expression. By exploiting the endogenous mammalian RNAi pathway, several expression-based strategies have been developed to inhibit human immunodeficiency virus (HIV) gene expression and replication. This approach potentially has utility as a protective 'therapeutic vaccine' of virus-susceptible lymphocytes. In this review we discuss new developments aimed at improving efficacy and delivery of novel RNAi-based gene expression antiviral strategies. Particular attention is given to advances in combinatorial gene expression systems that prevent the emergence of RNAi-resistant virus by simultaneously targeting multiple HIV targets. Potential usefulness of silencing host factors that are required for viral replication is also discussed. These approaches form the basis for a number of promising ongoing and future clinical trials aimed at providing an effective, safe and prolonged single-intervention therapy for HIV/AIDS.