HG6-64-1, a chemical GCK inhibitor. (A) Six DLBCL cell lines and 2 control, non-DLBCL cell lines, were treated with serial dilutions of HG6-64-1. 3-(4,5-Dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl-2-(4-sulfophenyl)-2H-tetrazolium (MTS) readings were taken just prior to and after drug addition and compared with control, DMSO-treated cells. Cellular proliferation was calculated as the change in MTS reading for each experimental condition, divided by the increase in DMSO-treated cells. (B) The calculated EC₅₀ of HG6-64-1 for each listed cell line. (C) Asynchronously growing DLBCL cell lines were subject to media change followed by treatment with HG6-64-1 or the vehicle, DMSO. After treatment, cells were ethanol permeabilized and saturated with PI in PBS. Cell cycle was analyzed by flow cytometry, and results are depicted in a bar graph. (D) Time-dependent measurement of cellular death by flow cytometry in DLBCL cell lines. (E) Flow cytometry measurement of cellular death at 48 hours of tumor B cells isolated from fresh biopsies of DLBCL primary tumors, or normal B cells isolated from healthy tissue, and treated with HG6-64-1 or DMSO. In one primary sample, cells were derived from the leukemic phase (DLBCL2A) and from a spleenic tumor (DLBCL2B) of the same patient. (F) Measurement of cellular death, by flow cytometry, of DLBCL cells lines treated with HG6-64-1, doxorubicin, or both to measure potential additive effects of combination treatment. All cell-line experiments were performed at least 3 times.