P16.01.B DOXORUBICIN GLYCOSYLATION AS A NOVEL APPROACH TO SPECIFICALLY TARGET GLIOBLASTOMA

Abstract BACKGROUND Despite recent advances in the treatment of cancer, malignant brain tumors such as Glioblastoma (GBM) are still a major challenge in oncology with a poor prognosis. This evidence can be partly explained by the inability of most drugs, such as Doxorubicin (DOX), to be delivered within the brain due to the presence of drug resistance mechanisms including the active efflux mediated by ATP-binding cassette transporters, which are overexpressed on the blood brain barrier (BBB).Malignant tumors like GBM also show a significant glycolytic rate and a high glucose request through glucose transporters (Warburg Effect). In this context, GLUT1 is reported to be implicated in glucose uptake through the BBB.In our study we have developed an innovative approach able to increase drug efficiency and selectivity, by conjugating the anticancer agent DOX to glucose. MATERIAL AND METHODS DOX glycoconjugates were obtained using oxime-based linkers with hydroxylamine-containing spacers between position number 6 of glucose and carbonyl group of DOX. Two different glycoconjugates (EB73 and EB74) were synthesized and characterized for their absorbance, fluorescence and binding affinity to ctDNA. MTT assay was performed to evaluate the cytotoxic effect of EB73 and EB74, compared to DOX, in two GBM cell lines (A172 and DOX resistant T98G). These cells were also used to evaluate the effect of glucose deprivation on the expression levels of GLUT1 and breast cancer resistance protein (BCRP) by Western Blot. Molecules uptake by cells was evaluated using an in vitro BBB model and immunofluorescence (IF) in glioblastoma and neuroglia cell lines. RESULTS Our results showed that EB73 and EB74 have kept Ka binding constants comparable to DOX itself, confirming that functionalization with glucose did not alter DOX intercalation capability and fluorescence.The in vitro cytotoxicity results demonstrated that EB74 has the major effect in A172 compared to DOX and revealed a high efficacy in T98G DOX-resistant cells starting from 24 hours of treatment. Moreover, glucose deprivation showed a GLUT1 overexpression and BCRP downregulation in both cell lines improving EB74 efficacy. According to MTT results, BBB in vitro model and IF experiments revealed that only EB74 was able to accumulate in cells nuclei as conventional DOX. CONCLUSION In conclusion, these results indicated that glycoconjugation of DOX and glucose deprivation may be helpful instruments to increase drugs uptake and effectiveness in aggressive and chemoresistant tumor cells.