Doxycycline containing food pellets

All experimental procedures described in this study were approved by, and adhered to guidelines of, the Memorial Sloan-Kettering Cancer Center institutional animal care and use committee. Doxycycline was administered to mice via 625 mg/kg doxycycline-containing food pellets (Harlan Teklad).

For induction of OIS in vivo, sleeping beauty (SB) transposase vector and two SB transposon vectors expressing N-Ras^G12D-IRES-GFP and a miR30-shRNA (against Renilla or Brd4; see Supplementary Methods for additional details) were co-delivered into C57BL/6J mice (purchased from Harlan-Teklad) via hydrodynamic injection. Mice were injected with a sterile 0.9 % NaCl solution of 12.5 μg DNA of each transposon vector together with CMV-SB13 Transposase (1:5 ratio) via the lateral tail vein. Doxycycline was administered to mice via 625 mg/kg doxycycline-containing food pellets (Harlan Teklad), starting 24 hr before injections. For pharmacologic inhibition of Brd4, i-BET-762 (Selleck Chemicals) was reconstituted in dimethylsulfoxide (DMSO) and further diluted in phosphate-buffered saline (PBS). Mice were treated with I-BET 762 or vehicle orally every day (15mg/kg). Mice were sacrificed at the indicated time points and chunks of liver tissue with either fixed in 10% neutral buffered formalin or fresh frozen in Optimal Cutting Temperature (OCT) compound for histological analyses performed as detailed in Supplementary Methods. All experiments with mice were performed in accordance with protocols approved by the Memorial Sloan-Kettering Cancer Center's Internal Animal Care and Use Committee (IACUC) and Federal Office of Laboratory Animal Welfare guidelines.