Protein expressions in del11q and non-del11q cells, with and without irradiation, and the interaction of ravoxertinib with AZD1390 and ibrutinib. (A) Western analysis of human B cells derived from individuals without (control) and with ataxia-telangiectasia (A-T; compound inactivating mutations in ATM exon 4) cotreated with AZD1390 and 8 Gy irradiation. AZD1390 reduced pATMS1981 and pKAP1S824 formation in control cells, whereas AZD1390 did not affect other B-cell–specific signaling pathways (mTOR, BTK, ERK, AKT, SYK, and LYN) in control/wild-type (WT) and A-T cells. Interestingly, genetic loss of ATM resulted in alterations to mTOR, SYK, and ERK activity, independent of AZD1390 treatment. (B) Western analysis of CLL cells from a representative non-del11 case and a del11q case (>50% cells effected) following incubation ex vivo for 18 hours in SFM or with anti-IgM, 3-hour drug treatment + 8 Gy irradiation with 30 minute recovery at 37°C. (C) Expression of pERK in 7 non-del11q/del17p cases and 5 del11q cases obtained by densitometry from western analysis in panel B, showing a trend toward decreased pERK in del11q cases coinciding with decreased pATM and pKAP1 (supplemental Figure 6). (D) Ibrutinib (synergism) or AZD1390 (antagonism) cotreatment with ravoxertinib on CLL cell survival. Combenefit plots represent data combined from 9 independent cells from patients with CLL. (E) Postulated mechanisms to explain the chemotherapy resistance, more rapid disease progression, and sensitivity to ibrutinib in del11q CLL. DMSO, dimethyl sulfoxide; IR, irradiation; unt, untreated.
