RNA Nanotechnolgy in Cancer Therapy

(1) We have discovered unusual properties of pRNA’s 3WJ (3-Way Junction) domain. This 3WJ domain can be assembled from three pieces of RNA oligos with unusually high affinity in absence of metal salts; displays thermodynamically stable properties; resistant to denaturation even in presence of 8M urea, and remains intact without dissociating at ultra-low concentrations. Melting experiments demonstrate that the three components of the 3WJ-pRNA core (TM: 58.0 ± 0.5 °C) have a much higher affinity to interact favorably in comparison with any of the two components, indicating cooperative simultaneous folding of the three helical stems. Comparing 3WJ-pRNA with 25 other 3WJ motifs in biological RNA, pRNA displayed the highest TM with the steepest slope. The discovery of this unusual RNA motif will open a new avenue in RNA nanotechnology. (2) We have constructed varieties of RNA nanoparticles using the 3WJ-pRNA as scaffold and demonstrated that the resulting RNA nanoparticles harboring different functional modules retained their folding and independent functionalities for specific cell binding, cell entry, gene silencing, catalytic function, and cancer targeting both in vitro and in animal trials. The particles have been tested for the delivery to the cells of leukemia, neuron, and liver hepatocytes. (3) The 3WJ-pRNA based RNA nanoparticles display favorable pharmacological profiles concerning biodistribution, pharmacokinetics (stability, half life and clearance rate), immune response (antibody induction, á and â interferon, toll-like and innate immunity, PKR effect, and cytokine induction). These findings offer a platform for the construction of therapeutic RNA drugs. (4) We have constructed stable 4WJ (4-WayJunction) motif that can serve as scaffold to assemble RNA nanoparticles in the absence of metal salts. It displays similar stable attributes as the 3WJ-pRNA with regards to thermodynamic stability, resistance to urea denaturation and dissociation at ultra-low concentrations. Furthermore, tetrameric RNA nanoparticles can be constructed by self assembly of functional modules (aptamers, two different siRNAs and Folate) using the 4WJ scaffold. (5) We have demonstrated that anti-BaffR1 aptamer evolved via in vitro SELEX procedure binds to the B-cell specific BaffR1 receptor and can be used to block Baff ligand signaling and deliver an anti-STAT3 siRNA into B-cell lymphomas, as demonstrated in a NOG-SCID hu mouse model.