SOX11 expression sensitizes MCL cells to ara-C through impairing ara-CTPase activity of SAMHD1. (A) Representative western blot showing SOX11 expression patterns in JVM-2^vector and JVM-2^iSOX11 in presence and absence of 0.1 μM doxycycline for 96 hours. One representative western blot out of 6 replicates is shown. SOX11 was detected at 74 kDa and GAPDH at 37 kDa. (B-C) Dose response curve for ara-C treatment for 72 hours in JVM-2^vector and JVM-2^iSOX11 in presence (B, DOX⁺), and absence (C, DOX⁻) of doxycycline (DOX). ara-C treatment was applied 24 hours after doxycycline (0.1 μM) treatment. Data are represented as mean ± SEM of 6 independent biological replicates. (D) Dose response curve for 72 hours of treatment with ara-C in JVM-2^vector and JVM-2^iSOX11 cultured in the indicated concentrations of doxycycline. ara-C treatment started 24 hours after doxycycline-induced SOX11 expression. Viability was measured using CellTiter-Glo Luminescent Cell Viability Assay after 72 hours of ara-C treatment. The values on the y-axis represent the relative viability values, which were calculated by normalizing luminescence value at each dose of ara-C for each condition to respective untreated controls. Data were represented as mean ± SEM of 4 independent biological replicates. (E) Dose response curve for ara-C determined in dX- or Vpx-treated JVM-2^vector and JVM-2^iSOX11 that were induced by 0.1 μM doxycycline. After 24 hours of culturing in doxycycline-supplemented media, cells were treated with dX or Vpx for 3 hours, followed by ara-C treatment for 72 hours. Viability was measured using CellTiter MTS assay after 72 hours of treatment. The values on the y-axis represent the relative viability values, which were calculated by normalizing absorbance values at each dose of ara-C for each condition to respective untreated controls. Data are represented as mean ± SEM for 5 independent biological replicates. (F) Western blot shows the effect of treatment of ara-C at sublethal dose (0.5 μM) on cleaved PARP1 was detected at 89 kDa, phospho-ChK2 (T68) (∼62 kDa), cleaved caspase-3 (17 kDa), and γ-H2A.X (14 kDa) in dX- or Vpx-treated JVM-2^vector and JVM-2^iSOX11. ara-C treatment was applied after 3 hours of treatment with either dX or Vpx and treated cells were harvested after 72 hours of ara-C treatment, as explained in panel A. One representative experiment out of 3 is shown. (G) Intracellular ara-CTP levels normalized to the canonical dTTP, determined using HPLC-MS/MS. Both JVM-2^vector and JVM-2^iSOX11 were treated with 10 μM ara-C for 24 hours. Circles and error bars correspond to individual values, mean ± SEM of at 3 independent experiments. Analyses were performed using unpaired 2-tailed t tests; ∗∗P < .01.