![Figure 4. CK1δ/ε inhibitor PF-670462 delays development of leukemia in the Eµ-TCL1 AT model of CLL. (A) In vivo CK1ε activity was assayed in primary PB WBCs isolated from Eµ-TCL1 mice with advanced spontaneous leukemia, which had been injected intraperitoneally by indicated doses of CK1 inhibitor. PB samples were taken 2 hours after the administration and processed by red blood cell (RBC) lysis and subsequent stimulation by 50 nM Calyculin A (10 minutes, 37°C), followed by cell lysis and western blotting analysis. (B) Activity of the CK1 inhibitor in vivo at time points 3 to 14 hours after administration was assessed by analysis of DVL3 phosphorylation status in the colon tissue of wild-type (WT) mice. The inhibitor was administered as 30 mg/kg (i.p., intraperitoneal; p.o., peroral = oral gavage), solvent only was used as a control (i.p.). DVL3 phosphorylation is a sensitive marker of CK1ε activity (see supplemental Figure 4B). (C) Typical disease progression in the AT leukemia model (N = 23). Day 0 = WT reference values (N = 16). Values are presented as means ± SEM. (Di) Scheme of experiment. (Dii) PF-670462 [21 days of treatment, administered by i.p. (N = 9) or by oral gavage (N = 6), control N = 8] reduces progression of CLL presented as lower WBC counts in the CK1 inhibitor treated group (unpaired Student t test, P = .0003). Merged data from 3 (i.p.) or 2 (oral gavage) independent experiments are presented. (Diii) The effect is evident also after 20 days treatment-free follow-up (P = .0435). Black lines indicate median. (E) Scheme of experiment analyzed in panels F through H. (F) Results of regular PB sampling of the experiment schematized in panel E. Values represent means ± SEM. Unpaired Student t test was used to compare CK1 inhibitor treated (N = 9) and control (N = 11) groups at indicated time points. (G) Spleen infiltration was measured by ultrasound after the end of inhibitor treatment (day 50; unpaired Student t test, P = .0183). Columns represent median value. (H) Mice were scored for overall survival (P = .027, Gehan-Breslow-Wilcoxon test). The x-axis presents the treatment-free period of the experiment, starting by day 50 (end of treatment).](https://ash.silverchair-cdn.com/ash/content_public/journal/blood/131/11/10.1182_blood-2017-05-786947/4/blood786947f4.jpeg?Expires=1735566717&Signature=Q-KfJxpELgR3tQIbXMSqhl6JTLDZPb8~ofH9caE9NwExXQhtrSBysPz1kqeqmmQnTmdD7CzVOxzUDAQQA-fACJpLnWxBGp-63aem5UH-7Z~wrdCVlKHEyfkX0c0j669Plr7rEo6X2F25MiPWxIIkZbl7MRxaXcPrZ2Jrfdvch7KyyzLGkM6uL0SPtkXiYd~ldOOHcX9C7uaSOuakke3TbaTux7YHdQkrsEnWZGze-nbyWmItDYehgYfBhgBhVBmSxJNq~pmxxVdC9mdkbhc9oUnswzaSDCZ9IBdUhj~nb5~Fv7tFxu617lGt35uPK~m3znDp~CWYhVuXvAoG~y5grA__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
CK1δ/ε inhibitor PF-670462 delays development of leukemia in the Eµ-TCL1 AT model of CLL. (A) In vivo CK1ε activity was assayed in primary PB WBCs isolated from Eµ-TCL1 mice with advanced spontaneous leukemia, which had been injected intraperitoneally by indicated doses of CK1 inhibitor. PB samples were taken 2 hours after the administration and processed by red blood cell (RBC) lysis and subsequent stimulation by 50 nM Calyculin A (10 minutes, 37°C), followed by cell lysis and western blotting analysis. (B) Activity of the CK1 inhibitor in vivo at time points 3 to 14 hours after administration was assessed by analysis of DVL3 phosphorylation status in the colon tissue of wild-type (WT) mice. The inhibitor was administered as 30 mg/kg (i.p., intraperitoneal; p.o., peroral = oral gavage), solvent only was used as a control (i.p.). DVL3 phosphorylation is a sensitive marker of CK1ε activity (see supplemental Figure 4B). (C) Typical disease progression in the AT leukemia model (N = 23). Day 0 = WT reference values (N = 16). Values are presented as means ± SEM. (Di) Scheme of experiment. (Dii) PF-670462 [21 days of treatment, administered by i.p. (N = 9) or by oral gavage (N = 6), control N = 8] reduces progression of CLL presented as lower WBC counts in the CK1 inhibitor treated group (unpaired Student t test, P = .0003). Merged data from 3 (i.p.) or 2 (oral gavage) independent experiments are presented. (Diii) The effect is evident also after 20 days treatment-free follow-up (P = .0435). Black lines indicate median. (E) Scheme of experiment analyzed in panels F through H. (F) Results of regular PB sampling of the experiment schematized in panel E. Values represent means ± SEM. Unpaired Student t test was used to compare CK1 inhibitor treated (N = 9) and control (N = 11) groups at indicated time points. (G) Spleen infiltration was measured by ultrasound after the end of inhibitor treatment (day 50; unpaired Student t test, P = .0183). Columns represent median value. (H) Mice were scored for overall survival (P = .027, Gehan-Breslow-Wilcoxon test). The x-axis presents the treatment-free period of the experiment, starting by day 50 (end of treatment).
