Background: This attractive and intriguing Ribonucleic acid (RNA) nanotechnology has been conceptualized over the last two decades and with our increasing understanding of RNA structure and function and improvements of RNA nanotechnology it is now possible to use this in clinical settings. Methods: Here we review the unique properties and the recent advances in RNA nanotechnology and then look at its scientific and preclinical applications for tumor diagnosis and targeted delivery and RNA-based therapy using RNA nanoparticles with diverse structures and functions. Finally, we discuss the future perspectives and challenges to RNA nanotechnology. Results: RNA can be designed and manipulated in a similar way to DNA while having different rules for base-pairing and displaying functions similar to proteins. Rationally designed RNA nanoparticles based on the three-way junction (3WJ) motif as the core scaffold have been extensively explored in the field of nanomedicine and targeted cancer diagnosis and therapy. Conclusions: RNA nanostructures based on 3WJs demonstrate promising future applications due to their thermal stability, molecular-level plasticity, multifunctional chemotherapeutic drug delivery and other intrinsic characteristics, which will greatly improve the treatment of cancer and promote further major breakthroughs in this field.