Otx015

BRD4 inhibitors were purchased from ApexBio (JQ1) and Cayman Chemical (OTX015) and used at a concentration of 1 μM. ERK inhibitor SCH772984 was purchased from Active Biochem and used at a concentration range of 250 nM-1 μM. MEK inhibitor trametinib was purchased from Selleckchem and used at a concentration range of 25–100 nM. All drugs were reconstituted in sterile, dry dimethyl sulfoxide, aliquotted, and store at −80 °C until further use.

All live virus experiments were performed in the BSL3 laboratory at the University of Nebraska Medical Center (Omaha, NE, USA). Calu-3 cells were plated in a 96 well transparent bottom black color plate (15000 cells/well) and cultured for 48 hours before infection. On the day of infection, the cells were treated with different concentrations of JQ1 (Cayman Chemical, Cat # 11187) or OTX015 (Cayman Chemical, Cat # 15947) (0.1 μM to 0.001 μM) two hours before infection. The cells were infected with SARS-CoV-2 (Delta variant of concern (VOC); pangolin linage: B.1.617.2, obtained from BEI Resources, NIAID, NIH; cat # 55672) with 0.5 MOI of viral titer or left uninfected (negative control). One hour post-infection, virus inoculum was removed from the cells, washed 3 times with 1X PBS, replenished with fresh media and treated with respective compounds. The reaction was terminated 24 hours post-infection. The culture supernatant was collected for measuring the viral replication kinetics, and the cells were washed and fixed with 4% PFA for immunofluorescence analysis.

Primary mouse MPNST (mMPNST) cells were generated via a mouse MPNST model as described previously (34 (link), 50 (link), 51 (link)). Human S462 MPNST cells were a kind gift from Karen Cichowski (Harvard Medical School). Human MPNST cells were authenticated with human-specific PCR primers on 9/18/18 to confirm the absence of mouse tumor cell contamination. All leukemia cell lines were a kind gift from Dr. Chengcheng Zhang (UT Southwestern, Dallas, TX). Routine mycoplasma testing of the cell lines was not performed. All cells were cultured in DMEM (10% FBS, 1% L-glutamine, 1% sodium pyruvate, 1% penicillin-streptomycin). Drugs used: JQ1 (Cayman Chemical and MedChem Express), OTX-015 (Cayman Chemical), CPI-203 (Cayman Chemical), CPI-0610 (Axon Medchem), ARV-771 and ARV-825 (MedChem Express).