Ve 822

Cells were treated with inhibitors: pladienolide B (EMD Millipore), E7107 (H3 Biomedicine), ATR inhibitors (VE-821, VE-822, SelleckChem; AZ-20), ATMi (KU55933, SelleckChem). Antibodies used in this study include AQR (A302–547A-T, Bethyl), pChk2 (2661, Cell Signaling), Flag (F7425, Sigma), GAPDH, (ABS16, Millipore), GFP (A11122, ThermoFisher Scientific), γH2AX (9718S, Cell Signaling), Ku70 (GTX70271, Genetex), PCNA (sc-56, Santa Cruz Biotechnology), pRPA (A300–246A, Bethyl), U2AF1 (ab197591, abcam), and S9.6 (Protein A purified from hybridoma S9.6 by Antibodies Incorporated).

ω-Pentadecalactone (PDL), poly(ethylene glycol) methyl ether (M_n = 2000 Da, MeO-PEG2K-OH), and Novozym 435 catalyst (Candida antarctica lipase B or CALB supported on acrylic resin) were obtained from Sigma-Aldrich Chemical. The enzyme catalyst was dried at 40 °C under 2.0 mmHg overnight prior to use. p-Dioxanone (DO) was acquired from Leap Labchem Scientific Co. in China. Aceonitrile and dimethylsulfoxide were obtained from J.T. Baker (Avantor Performance Materials, Central Valley, PA, USA). VE822 was obtained from Selleck (Houston, TX, USA). DiI dye was purchased from Thermo Fisher Scientific (Waltham, MA, USA). RG2 cells were purchased from ATCC (Manassas, VA). Normal Human Astrocytes were obtained from Tim Chan at Memorial Sloan Kettering Cancer Center. All cells were cultured in 5% CO2 and air humidified in a 37 °C incubator. Each cell line was cultured up to passage number 20.

All reagents were obtained from Sigma Aldrich if not otherwise specified. Synthetic oligonucleotides were obtained from Eurofins (Eurofins MWG Operon, Huntsville, AL, USA) and purified by HPLC. All small molecule inhibitors used were dissolved in dimethyl sulfoxide at 10 mM (Fisher Scientific, Watham, MA, USA) and stored at −20 °C until further use: AZD6738 (Astra Zeneca, London, UK), KU60019 (Selleck Chemicals LLC, Munich, Germany), AZ20 (Tocris, Bristol, UK), VE822 (Selleck Chemicals LLC), AZD7762 (Selleck Chemicals LLC).