The Hexosamine Biosyntheic Pathway and Platinum-Ethylenedicysteine-Glucosamine (Pt-ECG) for Targeted Therapy in Diffuse Large B-Cell Lymphoma

Abstract 587

Aggressive non-Hodgkin lymphomas (NHL), such as diffuse large B cell lymphomas (DLBCL), are very common in the US with increasing incidences. Although these lymphomas are now potentially curable, almost half the treated patients still develop relapsed/refractory disease with poor survival outcomes, indicating an urgent need for better therapeutic approaches with improved efficacy.

The hexosamine signaling pathway terminating in O-linked N-acetyl glucosamine (O-GlcNAc) cycling has been implicated in cellular signaling cascades and regulation of transcription factors involved in cancer biology. Biological functions of the hexosamine biosynthetic signaling pathways need elucidation, to determine whether altered O-GlcNAc metabolism plays a significant role in hematologic tumors such as DLBCL, and utilize this bifunctional pathway as a targeted therapeutic strategy in DLBCL. We have identified key enzymes of the hexosamine biosynthetic pathways to be highly-expressed in DLBCL cell lines and patient tumor cells. In contrast to normal circulating and tonsillar B cells, DLBCL cells expressed high levels of the rate limiting enzyme glutamine: fructose-6-phosphate amidotransferase (GFAT) as well as terminating enzyme O-GlcNAc transferase (OGT). We discovered that several key growth and survival transcription factors, such as NF-kB and NFAT, known to be highly-activated in DLBCL, are linked to the hexoasmine biosynthetic pathway. We demonstrated that both NF-kB (p65) and NFATc1 directly associated with OGT, and down-regulation of OGT by siRNA inhibits these transcription factors activation, suggesting that both NF-kB-p65 and NFATc1 require O-GlcNAc glycosylation by OGT for their activation. ChiP on Chip analysis on NFATc1 indicated that this transcription factor regulates a set of genes involved in glucose metabolism, including hexokinase and GFAT. These results suggest that the hexosamine pathway is highly active and utilized in DLBCL, and that exploiting this bi-functional pathway(s) as a therapeutic approach is feasible. We have previously developed an imaging agent, ^99mTc-ethylenedicysteine-glucosamine (^99mTc-EC-G) because EC-G mimics phosphorylated N-acetylglucosamine. ECG treatment in DLBCL cells enhances p65 and NFATc1 nuclear translocation. For therapeutic strategies, we developed metallic unlabeled Platinum (Pt) derivatives-EC-G as potential therapeutic agents. Pre-clinical in vitro studies have shown that our two lead compounds, Pt- and Pt-(DACH)-EC-G effectively inhibit lymphoma cell growth and induce apoptosis. These lead compounds can also induce DNA damage in DLBCL cells, through the up-regulation of phosphorylated histone 2AX (pH2AX), leading to the disruption of p65 and NFATc1 binding to DNA.

This data importantly demonstrates that the hexosamine biosynthetic pathway is linked to key growth and survival pathways involved in the pathophysiology of DLBCL. Targeting these pathways with novel platinum EC-G compounds as a theranostic approach should lead to new, more effective treatments and diagnosis for DLBCL, particularly for relapsed/refractory DLBCL.