IN VIVO INOCULATION OF GBM CELLS IN RATS AND TREATMENT WITH RUXOLITINIB

GlioStat (patent pending) denotes an invention that addresses the current deficiencies of glioblastoma (GBM) chemotherapy by developing a molecularly imprinted drug reservoir that is specifically designed for post-surgical recovery. The goal of our research was to guarantee the long-term release of the antitumor drug ruxolitinib (RUX), which is specifically designed to target residual infiltrative cancer cells, while simultaneously reducing the risk of adverse effects. In order to achieve this goal, we have successfully developed and characterized four distinctive molecularly imprinted polymers (MIPs), with one of them advancing to in vivo experimentation. The selection of one MIP for further in vivo investigations was guided by the Alamar Blue viability assay on C6 GBM cell cultures, which took into account both the efficacy and the potential toxicity of residual monomers. Over the course of 96 hours, MIP 2 demonstrated the most favorable risk-benefit profile, providing superior efficacy against GBM C6 cells. In contrast, its non-imprinted counterpart (NIP 2) exhibited minimal toxicity. The protocol involved anesthetising the male Wistar rats, immobilising them on a corkboard, without the use of a stereotactic headholder. After shaving the head and local disinfection with iodine solution, we made a linear sagittal incision and exposed the parietal bones. A 3x3 mm burr hole was made with a pneumatic drill in the right parietal bone, revealing the dura mater. Subsequently, we injected 20,000 C6 GBM cells suspended in 5 microliters of solution in the cerebral parenchyma at a depth of 3 mm. Clinical and imaging control was performed. For the animals that developed tumors, a second therapeutic intervention was performed. The rats were anesthetised, disinfected, and readied in the same manner as before. The burr hole was exposed, with the tumor being partially resected via a “microscooping” technique. Next, 5-10 microliters of MIP2 solution (4 mg/mL) were injected into the parenchyma respective of the tumor bed. Five microliters of thrombin solution were then added (100 UI/mL) to form the fibrin network. In comparison to the untreated controls and animals treated with free RUX, animals treated with MIP2 exhibited a substantial increase in survival time during the in vivo evaluation, extending from 20 to 50 days. As such, our research had yielded a potentially life-changing drug delivery system in GBM, with the capacity to significantly increase postoperative survival. More in vivo tests should be conducted to validate our findings.