Development of image-guided drug delivery systems for real-time monitoring, quantifying and validating the delivery and therapeutic action is very important for both research and clinical applications. Various multifunctional nanoparticles of different materials have been investigated over the last decade as prospective imaging and therapeutic platforms; however, effective strategies to quantitatively evaluate delivery of therapeutic payloads to tumors and metastatic cancer cells in vivo are challenging due to low efficiency in specific cancer targeting and non-specific trapping in vital organs such as, liver an lungs. Our goal is to adopt an innovative RNA nanotechnology approach to construct ultrastable multifunctional RNA Beacons and RNA Dendrimers as image-guided agents for quantitative real-time assessment of tumor and metastatic cancer targeted therapeutic delivery, distribution, uptake and response. We have shown that our RNA nanoparticles are non-toxic, non-immunogenic and capable of penetrating across heterogeneous biological barriers to deliver high doses of therapeutics specifically to solid tumors and metastatic cells in mice with little accumulation in normal organs. This provides an ideal platform for non-invasive imaging of cancer therapeutics in vivo. Our multifunctional RNA nanoparticles will be designed to harbor: (1) imaging modules: NIR fluorophores for in vivo fluorescence imaging; radionuclides for PET/SPECT imaging; and gadolinium contrast agents for MRI; (2) targeting modules: such as RNA aptamers or chemical ligands for binding to cancer specific cell surface receptors resulting in internalization of RNA nanoparticles into cancer cells; and (3) therapeutic modules: siRNAs and anti-miRNA to silence the expression of oncogenic genes. The multifunctional RNA constructs will then be evaluated in our well established colon cancer animal models. We will employ multimodal imaging techniques (NIRF, PET/SPECT and/or MRI) for monitoring the therapeutic delivery process in real-time at various spatial and temporal resolution scales. This approach is based on the consideration that each of the three imaging modalities has advantages, and an integrated approach will lead to synergistic benefits with regards to image-guided therapy.