Abstract 4721: Glycosylation mediated cellular uptake and functional activity of conventional chemotherapy in acute lymphoblastic leukemia

Abstract Background: Acute lymphoblastic leukemia (ALL) is a heterogeneous hematologic disease characterized by the proliferation of immature lymphoid cells in the bone marrow, peripheral blood, and other organs. Conventional chemotherapeutic approaches are associated with toxicities leading to reduced efficacy over time due to its systemic approach to drug delivery. O-linked glycosylation patterns on lymphoid cells are common modifications on hydroxyl groups of serine or threonine residues that are associated with key functions including cell adhesion and migration. Aberrant expression of O-glycans have been associated with leukemic malignancies making it an attractive biomarker to explore in vitro. These studies aim to provide insights into the influence of O-glycosylation on cellular access of ALL target cells to model therapeutic agents, including antibody and relevant therapeutic agents currently employed for the treatment of ALL. Methods: The target cell line employed for use in this study was CCRF-CEM. Cells were plated at 20,000 cells/mL in 24-well plates or 500,000 cells/mL in 6-well plates. Cells were exposed to Benzyl-α-GalNAc, an O-linked glycosylation inhibitor, for 48 hours prior to exposure to FITC-tagged Anti-MUC1 antibodies or conventional chemotherapy at various time points. Conventional chemotherapeutic agents were selected based on the National Comprehensive Cancer Network guidelines for the treatment of ALL. Drug agents employed were, cyclophosphamide, imatinib, and doxorubicin hydrochloride. Cell viability was assessed via electronic cell count, Caspase 3, and SRB (Sulforhodamine) assay procedures. Results: Following inhibition of glycosylation a statistically significant decrease in the uptake of FITC-tagged Anti-MUC1 antibody by CCRF-CEM cells compared to control cells (P<0.001) was observed at 4 hours, with no difference reported at 24 hours. The finding suggests that the diminished glycosylation levels surrounding CCRF-CEM cells decreased the access of anti-MUC1 antibody to the membrane core protein. The studies showed that inhibition of glycosylation in CCRF-CEM cells decreased the overall effectiveness of chemotherapeutic agents compared to control in vitro. A diminished cytotoxic drug effect was reported for doxorubicin hydrochloride (P<0.0001), Imatinib (P<0.0001) and Cyclophosphamide (P<0.001), when compared to the effects of each agent alone. These findings were further supported by the cytotoxicity results from the SRB and Caspase 3 assay. Conclusions: Our preliminary findings support the notion that the inhibition of O-linked glycosylation in ALL cells decreased, rather than increased, the entry of drug agents into ALL cells. The data are supported by diminished general effects of conventional chemotherapeutic agents against the target cells. Additional confirmation studies are currently underway. Citation Format: Kenny D. Pham, Ghada Alhafez, Daniel Muteba, Robert B. Campbell. Glycosylation mediated cellular uptake and functional activity of conventional chemotherapy in acute lymphoblastic leukemia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 4721.