LAUSR

BACKGROUND & AIM. Interleukin-1 receptor-associated kinase 4 (IRAK4) plays a critical role in Toll-like receptor (TLR) signal transduction and the innate immune response. Recruitment of IRAK4 to these receptors and its subsequent activation is facilitated by the MYD88 adaptor protein. Approximately 2-5% of chronic lymphocytic leukemia (CLL) patients have activating mutations of MYD88, a driver mutation in this disease. Here we study the pharmacological inhibition of IRAK4 with ND2158, an IRAK4 competitive inhibitor, as a therapeutic approach in CLL. METHODS. TLR stimulation was performed with an agonist mix of Pam3CSK4, HKLM, FSL1 and ODN2006. Cytokine secretion and CLL proliferation were assessed by Luminex system and carboxyfluorescein succinimidyl ester staining, respectively. For in vivo studies, the Eμ-TCL1 adoptive transfer mouse model was used. RESULTS. TLR repertoire analysis in CLL cells revealed high expression levels of TLR1, TLR7 and TLR10. Gene expression profiling identified enrichment of gene sets related to cytokines and inflammation in the subgroup carrying mutations in MYD88. Accordingly, CLL cells from individuals with MYD88 mutations showed higher p65 NF-κB activity and secreted higher basal levels of CCL2, CCL3 and CCL4 cytokines compared with unmutated cases. ND2158 induced a selective dose-dependent cytotoxic effect in CLL cells, without differences regarding their MYD88 mutational status. In the context of TLR stimulation, ND2158 was able to efficiently decrease the secretion of CCL2, CCL3, CCL4, TNFα, IL1β and IL6 cytokines. NF-κB signaling was also modulated by the drug, as observed by the downregulation of IκBα phosphorylation protein level, p65 translocation to the nucleus and decreased p65 DNA binding activity. ND2158 also impaired CLL proliferation and migration towards CXCL12. In vitro experiments with splenocytes from leukemic Eμ-TCL1 mice corroborated the ND2158 antitumor effects. In vivo studies showed a significant reduction in tumor load in peripheral blood, lymph node, spleen and peritoneal cavity in mice treated with 100 mg/kg/day of the IRAK4 inhibitor. A decrease in spleen weight and size was also detected after the treatment. As monocytes have been shown to support CLL survival, we investigated ND2158 impact on this cell type. A significant reduction in monocyte absolute counts in spleen and skewing to inflammatory Ly6C+ monocytes in peritoneal cavity was detected after ND2158 treatment, thereby disrupting the support for tumor cells. CONCLUSION. Our findings support pharmacological inhibition of IRAK4 as a potential therapeutic strategy in CLL cells regardless of their MYD88 mutational status, indicating that TLR signaling plays an important role in CLL cells development and CLL tumor microenvironment. Citation Format: Laia Rosich, Neus Gimenez, Ralph Schulz, Morihiro Higashi, Marta Aymerich, Monica Lopez, Manel Juan, Martina Seiffert, Elias Campo, Dolors Colomer. Targeting IRAK4 disrupts inflammatory pathways and tumor microenvironment in chronic lymphocytic leukemia regardless MYD88 mutational status [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr LB-083.