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The clinically approved drugs dasatinib and bosutinib induce anti-inflammatory macrophages by inhibiting the salt-inducible kinases.

Ozanne J, Prescott AR, Clark K - Biochem. J. (2015)

Bottom Line: Bosutinib and dasatinib were originally developed as inhibitors of the protein tyrosine kinases Bcr-Abl and Src but we show that, surprisingly, the effects of bosutinib and dasatinib on macrophage polarization are the result of the inhibition of the salt-inducible kinases.Importantly, these effects of bosutinib and dasatinib on IL-10 gene expression are lost in macrophages expressing a drug-resistant mutant of salt-inducible kinase 2 (SIK2).In conclusion, our study identifies the salt-inducible kinases as major targets of bosutinib and dasatinib that mediate the effects of these drugs on the innate immune system and provides novel mechanistic insights into the anti-inflammatory properties of these drugs.

View Article: PubMed Central - PubMed

Affiliation: *MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, Sir James Black Centre, University of Dundee, Dundee DD1 5EH, Scotland, U.K.

ABSTRACT
Macrophages switch to an anti-inflammatory, 'regulatory'-like phenotype characterized by the production of high levels of interleukin (IL)-10 and low levels of pro-inflammatory cytokines to promote the resolution of inflammation. A potential therapeutic strategy for the treatment of chronic inflammatory diseases would be to administer drugs that could induce the formation of 'regulatory'-like macrophages at sites of inflammation. In the present study, we demonstrate that the clinically approved cancer drugs bosutinib and dasatinib induce several hallmark features of 'regulatory'-like macrophages. Treatment of macrophages with bosutinib or dasatinib elevates the production of IL-10 while suppressing the production of IL-6, IL-12p40 and tumour necrosis factor α (TNFα) in response to Toll-like receptor (TLR) stimulation. Moreover, macrophages treated with bosutinib or dasatinib express higher levels of markers of 'regulatory'-like macrophages including LIGHT, SPHK1 and arginase 1. Bosutinib and dasatinib were originally developed as inhibitors of the protein tyrosine kinases Bcr-Abl and Src but we show that, surprisingly, the effects of bosutinib and dasatinib on macrophage polarization are the result of the inhibition of the salt-inducible kinases. Consistent with the present finding, bosutinib and dasatinib induce the dephosphorylation of CREB-regulated transcription co-activator 3 (CRTC3) and its nuclear translocation where it induces a cAMP-response-element-binding protein (CREB)-dependent gene transcription programme including that of IL-10. Importantly, these effects of bosutinib and dasatinib on IL-10 gene expression are lost in macrophages expressing a drug-resistant mutant of salt-inducible kinase 2 (SIK2). In conclusion, our study identifies the salt-inducible kinases as major targets of bosutinib and dasatinib that mediate the effects of these drugs on the innate immune system and provides novel mechanistic insights into the anti-inflammatory properties of these drugs.

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Bosutinib and dasatinib induce key features of ‘regulatory’ macrophages(A) Effect of bosutinib and dasatinib on cytokine secretion. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for 8 h. The concentration of the different cytokines in the culture supernatant was measured using the BIOPLEX system. The data are depicted as the fold-change in cytokine secretion in the presence of the drug (n=4, mean±S.E.M.). Statistical significance was determined by comparing each dataset to 0 using a one-sample Student's t test. (B) Effect of bosutinib and dasatinib on the expression of markers of ‘regulatory’ macrophages. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for the indicated times. Expression of LIGHT, SPHK1 and arginase 1 were measured by qPCR. mRNA levels were normalized to 1 in unstimulated cells (mean±S.E.M., n=4). (C) Effect of bosutinib and dasatinib on the expression of markers of alternatively activated macrophages. Experiment was performed as described in (B) but measuring the mRNA levels of FIZZ, Ym1 and Mgl2 (mean±S.E.M., n=4).
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Figure 7: Bosutinib and dasatinib induce key features of ‘regulatory’ macrophages(A) Effect of bosutinib and dasatinib on cytokine secretion. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for 8 h. The concentration of the different cytokines in the culture supernatant was measured using the BIOPLEX system. The data are depicted as the fold-change in cytokine secretion in the presence of the drug (n=4, mean±S.E.M.). Statistical significance was determined by comparing each dataset to 0 using a one-sample Student's t test. (B) Effect of bosutinib and dasatinib on the expression of markers of ‘regulatory’ macrophages. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for the indicated times. Expression of LIGHT, SPHK1 and arginase 1 were measured by qPCR. mRNA levels were normalized to 1 in unstimulated cells (mean±S.E.M., n=4). (C) Effect of bosutinib and dasatinib on the expression of markers of alternatively activated macrophages. Experiment was performed as described in (B) but measuring the mRNA levels of FIZZ, Ym1 and Mgl2 (mean±S.E.M., n=4).

Mentions: ‘Regulatory’-like macrophages are also characterized by the production of low levels of pro-inflammatory cytokines and the expression of markers such as SPHK1, LIGHT and arginase 1 [4]. Interestingly, treatment of macrophages with bosutinib and dasatinib inhibited the production of IL-6, IL-12p40 and TNFα induced by LPS stimulation (Figure 7A; Supplementary Figure S3). Moreover, bosutinib and dasatinib elevated the expression of SPHK1, LIGHT and arginase 1 (Figure 7B) without affecting the expression of FIZZ, YM1 and Mgl2 (Figure 7C), which are markers of alternatively activated (M2a) macrophages generated in response to IL-4. Similar results were obtained when macrophages were stimulated with Pam3CSK4, a TLR1/2 agonist (Supplementary Figure S4). Our results demonstrate that bosutinib and dasatinib, like other SIK inhibitors such as MRT199665 and HG-9-91-01 [13], drive the polarization of macrophages towards a ‘regulatory’-like phenotype.


The clinically approved drugs dasatinib and bosutinib induce anti-inflammatory macrophages by inhibiting the salt-inducible kinases.

Ozanne J, Prescott AR, Clark K - Biochem. J. (2015)

Bosutinib and dasatinib induce key features of ‘regulatory’ macrophages(A) Effect of bosutinib and dasatinib on cytokine secretion. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for 8 h. The concentration of the different cytokines in the culture supernatant was measured using the BIOPLEX system. The data are depicted as the fold-change in cytokine secretion in the presence of the drug (n=4, mean±S.E.M.). Statistical significance was determined by comparing each dataset to 0 using a one-sample Student's t test. (B) Effect of bosutinib and dasatinib on the expression of markers of ‘regulatory’ macrophages. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for the indicated times. Expression of LIGHT, SPHK1 and arginase 1 were measured by qPCR. mRNA levels were normalized to 1 in unstimulated cells (mean±S.E.M., n=4). (C) Effect of bosutinib and dasatinib on the expression of markers of alternatively activated macrophages. Experiment was performed as described in (B) but measuring the mRNA levels of FIZZ, Ym1 and Mgl2 (mean±S.E.M., n=4).
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Figure 7: Bosutinib and dasatinib induce key features of ‘regulatory’ macrophages(A) Effect of bosutinib and dasatinib on cytokine secretion. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for 8 h. The concentration of the different cytokines in the culture supernatant was measured using the BIOPLEX system. The data are depicted as the fold-change in cytokine secretion in the presence of the drug (n=4, mean±S.E.M.). Statistical significance was determined by comparing each dataset to 0 using a one-sample Student's t test. (B) Effect of bosutinib and dasatinib on the expression of markers of ‘regulatory’ macrophages. Bone-marrow-derived macrophages were treated with vehicle control, 3 μM bosutinib or 0.3 μM dasatinib for 1 h and then stimulated with LPS for the indicated times. Expression of LIGHT, SPHK1 and arginase 1 were measured by qPCR. mRNA levels were normalized to 1 in unstimulated cells (mean±S.E.M., n=4). (C) Effect of bosutinib and dasatinib on the expression of markers of alternatively activated macrophages. Experiment was performed as described in (B) but measuring the mRNA levels of FIZZ, Ym1 and Mgl2 (mean±S.E.M., n=4).
Mentions: ‘Regulatory’-like macrophages are also characterized by the production of low levels of pro-inflammatory cytokines and the expression of markers such as SPHK1, LIGHT and arginase 1 [4]. Interestingly, treatment of macrophages with bosutinib and dasatinib inhibited the production of IL-6, IL-12p40 and TNFα induced by LPS stimulation (Figure 7A; Supplementary Figure S3). Moreover, bosutinib and dasatinib elevated the expression of SPHK1, LIGHT and arginase 1 (Figure 7B) without affecting the expression of FIZZ, YM1 and Mgl2 (Figure 7C), which are markers of alternatively activated (M2a) macrophages generated in response to IL-4. Similar results were obtained when macrophages were stimulated with Pam3CSK4, a TLR1/2 agonist (Supplementary Figure S4). Our results demonstrate that bosutinib and dasatinib, like other SIK inhibitors such as MRT199665 and HG-9-91-01 [13], drive the polarization of macrophages towards a ‘regulatory’-like phenotype.

Bottom Line: Bosutinib and dasatinib were originally developed as inhibitors of the protein tyrosine kinases Bcr-Abl and Src but we show that, surprisingly, the effects of bosutinib and dasatinib on macrophage polarization are the result of the inhibition of the salt-inducible kinases.Importantly, these effects of bosutinib and dasatinib on IL-10 gene expression are lost in macrophages expressing a drug-resistant mutant of salt-inducible kinase 2 (SIK2).In conclusion, our study identifies the salt-inducible kinases as major targets of bosutinib and dasatinib that mediate the effects of these drugs on the innate immune system and provides novel mechanistic insights into the anti-inflammatory properties of these drugs.

View Article: PubMed Central - PubMed

Affiliation: *MRC Protein Phosphorylation and Ubiquitylation Unit, College of Life Sciences, Sir James Black Centre, University of Dundee, Dundee DD1 5EH, Scotland, U.K.

ABSTRACT
Macrophages switch to an anti-inflammatory, 'regulatory'-like phenotype characterized by the production of high levels of interleukin (IL)-10 and low levels of pro-inflammatory cytokines to promote the resolution of inflammation. A potential therapeutic strategy for the treatment of chronic inflammatory diseases would be to administer drugs that could induce the formation of 'regulatory'-like macrophages at sites of inflammation. In the present study, we demonstrate that the clinically approved cancer drugs bosutinib and dasatinib induce several hallmark features of 'regulatory'-like macrophages. Treatment of macrophages with bosutinib or dasatinib elevates the production of IL-10 while suppressing the production of IL-6, IL-12p40 and tumour necrosis factor α (TNFα) in response to Toll-like receptor (TLR) stimulation. Moreover, macrophages treated with bosutinib or dasatinib express higher levels of markers of 'regulatory'-like macrophages including LIGHT, SPHK1 and arginase 1. Bosutinib and dasatinib were originally developed as inhibitors of the protein tyrosine kinases Bcr-Abl and Src but we show that, surprisingly, the effects of bosutinib and dasatinib on macrophage polarization are the result of the inhibition of the salt-inducible kinases. Consistent with the present finding, bosutinib and dasatinib induce the dephosphorylation of CREB-regulated transcription co-activator 3 (CRTC3) and its nuclear translocation where it induces a cAMP-response-element-binding protein (CREB)-dependent gene transcription programme including that of IL-10. Importantly, these effects of bosutinib and dasatinib on IL-10 gene expression are lost in macrophages expressing a drug-resistant mutant of salt-inducible kinase 2 (SIK2). In conclusion, our study identifies the salt-inducible kinases as major targets of bosutinib and dasatinib that mediate the effects of these drugs on the innate immune system and provides novel mechanistic insights into the anti-inflammatory properties of these drugs.

Show MeSH
Related in: MedlinePlus