Effect of second-generation BTK inhibitors on the functionality of macrophages and neutrophils from CLL patients

ANA COLADO; JOSÉ LUIS MARÍN FRANCO; ESTEBAN ELÍAS; MARICEF VERGARA; GREGORIO CORDINI; ALEXIA VEREERTBRUGGHEN; HORACIO FERNANDEZ GRECCO; SANTIAGO CRANCO; CAROLINA MAHUAD; FLORENCIA AIZPURUA; MARTA ZERGA; LUCIANA BALBOA; PABLO MORANDE; FERNANDO BEZARES; MIRTA GIORDANO; ROMINA GAMBERALE ; MERCEDES BORGE

Congreso; XVIII International Workshop on Chronic Lymphocytic Leukemia; 2019

The development of BTK inhibitors (BTKi) has changed the treatment landscape in CLL. To date, ibrutinib is the only BTKi approved for CLL treatment, while second generation BTKi with greater selectivity for BKT, such as acalabrutinib, approved for MCL, and spebrutinib, are being evaluated in clinical trials. Besides its effects on leukemic B-cells, ibrutinib also affects key innate immune cells such as macrophages and neutrophils. In this regard we and others have reported that ibrutinib impairs macrophage-M1 polarization and macrophage response to TLR`s ligands, Mycobacterium tuberculosis (Mtb) and Aspergillus fumigatus (Colado A, et al. Blood Cancer Journal. 2018, Fiorcari, S et al.Oncotarget. 2016, Bercusson A, et al. Blood, 2018), while others have reported that ibrutinib affects neutrophil functionality (Stadler, N. et al. haematologica. 2017). Infections are a main cause of morbidity and mortality in CLL, and while rates of infections during ibrutinib treatment are similar or lower compared to other treatments (Ball S, et al. European journal of haematology. 2018 and O'Brien S, Clinical Lymphoma, Myeloma & Leukemia. 2018), they are still one of the most frequent adverse events (O'Brien S, et al. Blood. 2018), and they are among the most common toxicities leading to drug discontinuation in relapsed/refractory patients (Mato A, et al. Haematologica. 2018). Therefore, to better understand the impact of new agents on immune cells may help to improve CLL patients´ care. Our aim was to investigate how second generation BTKi affect macrophage and neutrophil functions. First we evaluated the effect of acalabrutinib and spebrutinib on macrophage polarization towards the IFN-γ-induced M1 profile, which is associated with an effective anti-microbial immune response. To this aim, human monocytes were cultured with GM-CSF and IFN-γ with or without BTKi at clinically relevant doses. We found that, similar to ibrutinib, acalabrutinib impaired M1 polarization as shown by the down-regulation in the expression of the M1-associated marker CD86, the decrease in TNF-α secretion, the up-regulation of the M2-associated markers CD16, CD14, CD163, CD206 and the increase in IL-10 secretion. In contrast, we found that spebrutinib did not modify M1/M2 markers or cytokine secretion (Figure 1A-G). When we analyzed the effect of the BTKi on macrophage response towards microbialstimulation we found that acalabrutinib, as reported for ibrutinib, significantly decreased TNF-α secretion by macrophages stimulated with irradiated-Mtb, heat inactivated Aspergillus fumigatus conidia or heat-inactivated Candida albicans yeast (Figure 1H-J). In line with this, we found that ibrutinib and acalabrutinib impaired TNF-α secretion when macrophages were stimulated with Pam3CSK4, a TLR2 ligand, or zymosan a TLR2 andDectin-1 ligand, which are receptors involved in macrophage-recognition of Mtb and fungi (n=8, p