Small-interfering RNA (siRNA)-mediated silencing of genes implicated in disease by the process of RNA interference offers a novel genetic medicine approach. Polymeric nanoparticles (or polyplexes) formed by self-assembly of polycations with siRNA can be used for site-specific delivery, cellular uptake and intracellular trafficking as a strategy to improve the therapeutic potential of siRNA. This review describes the application of polyplexes for in vivo delivery of synthetic siRNA with focus given to systemic and mucosal routes and in vivo requirements. Issues including use of stimuli-responsive systems for intracellular trafficking of siRNA are discussed as part of necessary future directives towards the development of RNA-based clinical therapeutics.