Extended Data Figure 10: Ibrutinib increases AID expression and the frequency of translocations to AID on- and off-target sites in human B cells.
From: Phosphatidylinositol 3-kinase δ blockade increases genomic instability in B cells
a, AICDA mRNA levels analysed by RT–qPCR in MEC1, Mino, JeKo-1 and GM06990 cell lines after treatment with DMSO or ibrutinib. Data are expressed as mean ± s.d. (n = 3 technical replicates, n = 3 biological replicates). *P < 0.05, **P ≤ 0.01, ***P ≤ 0.001, two-tailed Student’s t-test. b, Western blot for AID protein expression in MEC1, Mino, JeKo-1 and GM06990 B-cell lines treated with the indicated inhibitors (1 μM) for 48 h. For comparison, Mino and GM06990 panels correspond to the panel shown in Fig. 4c and Extended Data Fig. 6d, respectively, with the addition of the ibrutinib lane (n = 3 biological replicates). For gel source data, see Supplementary Fig. 1. c, Histograms showing translocation junctions frequency to AID on-target and off-target sites in MEC1 and JeKo-1 B-cell lines treated with ibrutinib. For each treatment, data are from pooled HTGTS libraries of similar size from three independent experiments (Supplementary Tables 1, 4). Significance is calculated as FDR in ibrutinib over DMSO-treated human B cells. Statistical analysis is indicated in the Method. ***FDR ≤ 0.001. d, Mutation frequency of control non AID off-targets: intragenic regions for PAX5, MTFR1, BCAT1, CLMN, UBXN7, ERICH6B and random intergenic regions on chromosome 1, chromosome 8, and chromosome 18 were targeted re-sequenced in patients with CLL untreated or treated with idelalisib and ibrutinib. Box plots indicate cumulative frequencies of C-to-T or G-to-A transition mutation in DNA samples collected before (pre) and after (post) treatment in each patient (control, n = 8; idelalisib, n = 10; ibrutinib, n = 10; Supplementary Table 6). Whiskers extend to a maximum of 1.5× interquartile range beyond the box. P values calculated by paired samples two-tailed Student’s t-test.
