Protein Kinase Inhibitor Dasatinib Potently Suppresses Immunostimulatory Activity of Human Plasmacytoid Dendritic Cells
Authors: Fujita Haruyuki , Kitawaki Toshio , Uchiyama Takashi , Takaori-Kondo Akifumi , Kadowaki Norimitsu 
Published: October 2019

Abstract
Abstract 3896

Background:
Plasmacytoid dendritic cells (pDCs) recognize endogenous nucleic acids derived from damaged cells through Toll-like receptor (TLR)7 and TLR9, and produce a vast amount of interferon (IFN)-α. These cells thereby play a pivotal role in the pathogenesis of inflammatory disorders such as lupus and psoriasis. Thus, development of a drug that suppresses IFN-α production by pDCs may lead to a novel therapy for such disorders. A tyrosine kinase inhibitor dasatinib developed as a drug for chronic myeloid leukemia inhibits not only abl tyrosine kinase but also other multiple protein kinases, including src family kinases (SFKs), which are important for immune regulation. Thus, we hypothesized that dasatinib may affect immune responses by modulating immunostimulatory activity of pDCs.

Methods:
We isolated pDCs from peripheral blood of healthy volunteers by cell sorting, and stimulated them with TLR7/8 ligands (influenza virus, an imidazoqinoline compound R-848) or TLR9 ligands CpG DNA (CpG-A: ODN2216, CpG-B: ODN2006) in the absence or presence of dasatinib. We used other abl tyrosine kinase inhibitors, imatinib and nilotinib, for comparison.

Results:
Dasatinib did not reduce the viability of pDCs. Low concentrations (10-30 nM) of dasatinib strongly suppressed (i) the production of IFN-α and tumor necrosis factor (TNF)-α and (ii) the upregulation of a costimulatory molecule CD86 by pDCs stimulated with CpG-A or influenza virus. In contrast, imatinib or nilotinib suppressed the production of cytokines and the upregulation of CD86 only at high concentrations. Dasatinib suppressed (i) the production of TNF-α and (ii) the upregulation of CD86 by pDCs stimulated with CpG-B or R-848 only weakly. SFK inhibitors, PP2 and SU6656, suppressed IFN-α production by pDCs, indicating that inhibition of SFKs is responsible for the suppressive effect of dasatinib at least in part. Studies by confocal microscopy revealed that dasatinib did not inhibit uptake of CpG-A by pDCs, and also did not block trafficking of TLR9 from endoplasmic reticulum to endosomes induced by stimulation with CpG-A. However, dasatinib inhibited nuclear translocation of IRF-7 and NF-kB in pDCs stimulated with CpG-A. These data indicate that dasatinib inhibits some points of the endosomal TLR9-MyD88 signaling pathway between early endosomes and activation of IRF-7. In the peripheral blood samples from the patients with chronic myeloid leukemia or Ph positive acute lymphoid leukemia, dasatinib suppressed IFN-α-producing capacity of pDC in vivo without reducing the numbers of pDC in blood.

Conclusion:
Dasatinib abrogates immunostimulatory activity of pDCs in vitro and in vivo at least in part by suppressing SFKs. As CpG-A and influenza virus are more prone to be retained in early endosomes in pDCs than CpG-B and R-848, the preferential suppression of CpG-A and influenza activity in pDCs suggests that dasatinib inhibits signaling components downstream of TLR7 and TLR9 closely connected with early rather than late endosomes. Dasatinib may alleviate inflammatory disorders that involve pDCs such as lupus and psoriasis. In addition, dasatinib might undermine antiviral immunity in which IFN-a derived from pDCs plays a part.

Disclosures:
No relevant conflicts of interest to declare.