Idelalisib

TMD8 cells were obtained from the Tokyo Medical and Dental University, and OCI-LY10 cells were obtained from University Health Network. Both cell lines were cultured in RPMI-1640 medium supplemented with 20% FBS, 100 U/mL penicillin, and 100 μg/mL streptomycin (Life Technologies, Carlsbad, CA). idelalisib and ibrutinib resistant TMD8 were cultured in the presence of idelalisib (1 μM) or ibrutinib (10–20 nM), respectively, and grown in a humidified atmosphere of 5% CO₂ and 95% air at 37°C. Compounds used in this study include: idelalisib (Gilead Sciences, Inc., Foster City, CA), GS-649443 (Gilead Sciences) [11 ], BYL719, AZD-6482, GDC-0941, MK-2206 and GSK2334470 (Selleckchem, Houston, TX) [12 (link)–16 (link)], ibrutinib (Shanghai Medicilon Inc., Shanghai, China), and ONO/GS-4059 (Ono Pharmaceutical Co., Trenton, NJ).

B-ALL cell lines (NALM6 and RS4;11) and NALM6 phospho-GR mutants (GR-S203A and GR-S226A) were tested with combinations of prednisolone (Acros Organics, ThermoFisher, Waltham, MA, USA, #449470250) and the PI3Kδ inhibitor idelalisib (Gilead Sciences, Foster City, CA, USA). Viability was measured using PrestoBlue (ThermoFisher, Waltham, MA, USA, A13262). Synergy was evaluated using the Bliss synergy model in SynergyFinder 2.0 [19 (link)].
Primary specimens from children with newly diagnosed or relapsed B-ALL were obtained prior to initiating treatment after receiving informed consent (University of Iowa IRB protocol #201707711). Cells were obtained from either peripheral blood or bone marrow and isolated by Histopaque density gradient separation. Freshly isolated cells were used for all experiments involving primary specimens.
NALM6, SUP-B15, and RCH-ACV cells were tested with dexamethasone (Sigma-Alrich, St. Louis, MO, USA, D4902-1g) in combination with ERK1/2 inhibitor SCH772984 (SelleckChem, Houston, TX, USA, #S7101).

Idelalisib was synthesized at Gilead Sciences, dissolved in DMSO at 10 mM, and stored at −20°C. Rituximab and obinutuzumab were provided by Hoffmann–La Roche (Basel, Switzerland). Palivizumab was used as a negative control and was produced at Gilead Sciences.