Ibrutinib does not prevent kidney fibrosis following acute and chronic injury

Abstract

Recent studies suggested that ibrutinib, a Bruton tyrosine kinase (BTK) inhibitor, developed for the treatment of chronic lymphocytic leukemia, may prevent NLRP3 inflammasome activation in macrophages, IL-1β secretion and subsequent development of inflammation and organ fibrosis. The role of NLRP3 has been underlined in the various causes of acute kidney injury (AKI), a pathology characterized by high morbimortality and risk of transition toward chronic kidney disease (CKD). We therefore hypothesized that the BTK-inhibitor ibrutinib could be a candidate drug for AKI treatment. Here, we observed in both an AKI model (glycerol-induced rhabdomyolysis) and a model of rapidly progressive kidney fibrosis (unilateral ureteral obstruction), that ibrutinib did not prevent inflammatory cell recruitment in the kidney and fibrosis. Moreover, ibrutinib pre-exposure led to high mortality rate owing to severer rhabdomyolysis and AKI. In vitro, ibrutinib potentiated or had no effect on the secretion of IL-1β by monocytes exposed to uromodulin or myoglobin, two danger-associated molecule patterns proteins involved in the AKI to CKD transition. According to these results, ibrutinib should not be considered a candidate drug for patients developing AKI.

Figure 1

Ibrutinib promotes renal fibrosis after unilateral ureteric obstruction (UUO). (A) Design of the study. (B–E) Representatives images and quantifications of Fibronectin (B), Sirius Red (C), B220 (D) and F4/80 (E) staining in sham and UUO mice receiving ibrutinib (Ibru) or vehicle (Veh). Data are represented as means ± SEM. *p < 0.05 (Mann–Whitney test; n = 6 mice per group); ns, not significant.

Figure 2

Prophylactic treatment with ibrutinib exacerbates rhabdomyolysis with subsequent severer acute kidney injury and higher mortality. (A) Design of the study. (B) Plasma level of CPK (6 h) and BUN (day 2 (D2) and 20 (D20)) after glycerol injection. (Mann–Whitney test and ANOVA, Tukey’s test; n = 3–15 mice per group). (C) Kaplan–Meier survival curves after glycerol injection in mice receiving ibrutinib (Ibru) or vehicle (Veh). Kaplan–Meier curves significance was calculated according to the Log-rank test. Values are expressed as mean ± SEM. *p < 0.05, ***p < 0.001, compared with the vehicle group; ns, not significant.

Figure 3

Ibrutinib promotes rhabdomyolysis-induced renal fibrosis. (A) Design of the study. (B) Kaplan–Meier survival curves after glycerol injection in mice receiving ibrutinib (Ibru) or vehicle (Veh) from day 3. (C) Blood urea nitrogen (BUN) 20 days after glycerol injection. (D–G) Representatives images and quantification of Sirius Red (D), Fibronectin (E), F4/80 (F) and B220 (G) staining 20 days after glycerol injection in mice receiving ibrutinib (Ibru) or vehicle (Veh) from day 3. Values are expressed as mean ± SEM. * p < 0.05, compared with the vehicle group (Mann–Whitney test, n = 5–10 mice); ns, not significant.

Figure 4

Kidney expression of BTK in sham and after glycerol-induced rhabdomyolysis (day 2; GLY) or unilateral ureteric obstruction (day 7, UUO).

Figure 5

Effect of Ibrutinib on human PBMC treated with myoglobin or uromodulin. Concentration (pg/ml) of IL-1β assessed by ELISA in culture supernatant of human PBMC treated with Uromodulin (Umod, A) or Myoglobin (Myo, B) and with ibrutinib (Ibru) or vehicle (Veh). Data expressed as mean ± SEM, n = 4–12, *p < 0.05, ****p < 0.001 compared with the Veh or Ctl group (ANOVA Tukey’s test; n = 3–15 samples per group); ns, not significant.