tRNA/miRNA (sephadex aptamer tagged methionyl tRNA fusion pre-mir) 1/5/10/20/30 nM in 96-well plate tRNA/miR34a prodrug (nine mice) formulated in in vivo-JetPEI (Polyplus-transfection SA) according to the manufacturer instructions, tRNA/MSA (nine mice) formulated in in vivo-JetPEI and vehicle control group (six mice) receiving only in vivo-JetPEI. Mice received treatments via tail vein injection (100 μL) consisting of a 50 μg initial loading dose on Day 1 followed by 25 μg maintenance doses three times per week for 5 doses.

Osteosarcoma (OSA) represents the most common primary bone tumor in humans and pet dogs. Little progress has been made with regard to viable treatment options in the past three decades and patients presenting with metastatic disease continue to have a poor prognosis. Recent mouse studies have suggested that microRNA-34a (miR-34a) may have anti-tumor activities in human OSA models. Due to the conservation of microRNA across species, we hypothesized that a bioengineered miR-34a prodrug (tRNA/miR-34a) would have similar effects in canine OSA, providing a valuable preclinical model for development of this therapeutic modality. Using a panel of canine OSA cell lines, we found that tRNA/miR-34a reduced viability, clonogenic growth, and migration and invasion while increasing tumor cell apoptosis. Furthermore, canine OSA cells successfully process the tRNA/miR-34a into mature miR-34a which reduces expression of target proteins such as platelet derived growth factor receptor alpha (PDGFRalpha), Notch1 and vascular endothelial growth factor (VEGF). Additionally, our subcutaneous OSA xenograft model demonstrated in vivo tumor growth delay, increased necrosis and apoptosis by tRNA/miR-34a, and decreased cellular proliferation ability. Taken together, these data support that this novel microRNA-based therapy may possess clinical utility in a spontaneously-occurring large animal model of OSA, which can then serve to inform the clinical development of this therapy for human OSA patients.