A 1155463

Single-cell suspensions from the spleen or BM were cultured with A-1210477 (catalog number CT-A121, ChemieTek, Indianapolis, IN, USA) at a concentration range of 0, 1, 5 or 20 μM, A-1155463 (catalog number CT-A115, ChemieTek) at 0, 10, 100 or 100 nM, or ABT-199 (catalog number A0776, LKT Laboratories, St Paul, MN, USA) at 0, 10, 100 or 1000 nM for 20 h at 37ºC. Cell viability after treatment was assessed by flow cytometry using the TO-PRO-3 dye (catalog number T3605, ThermoFisher, Waltham, MA, USA) after gating on specific B-cell subsets using the gating strategy as shown in Figures 1a and b or as previously published for PC.^{16 (link)} Specific apoptosis was calculated by measuring the altered percentage of TOPRO3⁻ (live) cells within indicated B-cell populations, compared with untreated cells. LC50 values were subsequently calculated using Excel and Graphpad Prism software, and specific apoptosis values after incubation with a concentration range of the inhibitors.

ABT-199 and WEHI-539 were purchased from Selleck Chemicals (Breda, The Netherlands), A-1155463 and AZD5991 from Chemietek (Indianapolis, USA). 5-Aza-2′-deoxycytidine (Decitabine) and Oxaliplatin were purchased from Sigma Aldrich. All compounds were dissolved in DMSO to a stock of 20 mM, except Oxaliplatin, which was dissolved in water to a stock of 12.5 mM.

ABT-199, ABT-263, and Q-VD-OPh were purchased from Selleckchem, Houston, TX, US. A-1155463 and AZD5991 were purchased from Chemietek, Indianapolis, IN, US. All compounds were dissolved in dimethyl sulfoxide (DMSO) at a 10 mM stock concentration.