Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken together our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human T-cell signaling, proliferation and function unaffected. and quizartinib did not have any negative impact on T-cell reactivity compared to DMSO control. Dasatinib impaired T-cell reactivity to less than 5% of control. (B, C) ELISPOT assay to assess for HD-TC-responses directed against viral antigens (n=10). Lytic T-cell activity was determined based on the frequency of IFN- secreting cells. Autologous APC loaded with a viral peptide pool (CMVpp65; PepMixTM HCMVA(pp65), JPT Inc., Berlin, Germany) were incubated with T cells (of the same individual) in the absence or presence of the respective TKI (as indicated). Midostaurin (50nM) and quizartinib (50nM) did not impair lytic activity. 10nM dasatinib significantly inhibited the virus-specific T-cell response. APC loaded with control peptides (CEF) or without peptides served as positive and negative control. These results could be recapitulated using human influenza virus derived peptides as an antigen. (DCG) Allogeneic (un-matched) GvHD (graft-versus-host-disease) assay in vivo: 0.5 106 CD3+ T cells derived from BL/6 mice were transplanted along with 2 106 BALB/c WBMC into lethally irradiated (13Gy) BALB/c recipient mice. Animals developed severe GvHD of the gut within 3C4 weeks after transplantation. Midostaurin treatment was performed after engraftment (Days 14C19; 100mg/kg body weight by gavage, every 24h). Ileum and colon fixed in formalin and embedded in paraffin were analyzed for apoptosis of crypt cells on Day 25. Midostaurin treatment (n=3) did not affect severity of GvHD in the mouse gut. Both, severity of GvH-reaction and induction of apoptosis in the ileum or colon were comparable to vehicle treated animals (n=2). Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken together our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human T-cell signaling, proliferation and function unaffected. Although our study is limited by the use of HD-TC that may act differently to T cells derived from AML patients, our findings facilitate a pre-clinical assessment for the use of FLT3-TKI in the context of alloSCT. Without affecting T-cell function, AR-9281 midostaurin and quizartinib could be concomitantly used until discontinuation of immunosuppressive therapy and thereby prevent relapse prior to appearance of a sufficient GvL-response. Interestingly, differential effects of FLT3-TKI on dendritic cell function that may even stimulate GvHD cannot be excluded by our experiments and need to be considered.15 Footnotes Information on authorship, contributions, and financial & other disclosures was provided by the authors and is available with the online version of this article at www.haematologica.org. Funding: this work was supported by a grant from Novartis Inc. to F.H.H. and partially by a DFG-grant (FI405/5-1) to T.F. and F.H and the Collaborative Research Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. and F.H.H. All human samples were collected and stored in the Hematology Tissue-Bank Magdeburg (HTM) supported by a grant from the Jose-Carreras Foundation (SP12/04) to F.H.H. F.H.H. received research funding from Novartis Inc. All other authors have nothing to disclose..and F.H and the Collaborative Research Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. IFN- secreting cells. Autologous APC loaded with a viral peptide pool (CMVpp65; PepMixTM HCMVA(pp65), JPT Inc., Berlin, Germany) were incubated with T cells (of the same individual) in the absence or presence of the respective TKI (as indicated). Midostaurin (50nM) and quizartinib (50nM) did not impair lytic activity. 10nM dasatinib significantly inhibited the virus-specific T-cell response. APC loaded with control peptides (CEF) or without peptides served as positive and negative control. These results could be recapitulated using human influenza virus derived peptides as an antigen. (DCG) Allogeneic (un-matched) GvHD (graft-versus-host-disease) assay in vivo: 0.5 106 CD3+ T cells derived from BL/6 mice were transplanted along with 2 106 BALB/c WBMC into lethally irradiated (13Gy) BALB/c recipient AR-9281 mice. Animals developed severe GvHD of the gut within 3C4 weeks after transplantation. Midostaurin treatment was performed after engraftment (Days 14C19; 100mg/kg body weight by gavage, every 24h). Ileum and colon fixed in formalin and embedded in paraffin were analyzed for apoptosis of crypt cells on Day 25. Midostaurin treatment (n=3) did not affect severity of GvHD in the mouse gut. Both, severity of GvH-reaction and induction of apoptosis in the ileum or colon were comparable to vehicle treated animals (n=2). Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken together our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human T-cell signaling, proliferation and function unaffected. Although our study is limited by the use of HD-TC that may act differently to T cells derived from AML patients, our findings facilitate a pre-clinical assessment for the use of FLT3-TKI in the context of alloSCT. Without affecting T-cell function, midostaurin and quizartinib could be concomitantly used until discontinuation of immunosuppressive therapy and thereby prevent relapse prior to appearance of a sufficient GvL-response. Interestingly, differential effects of FLT3-TKI on dendritic cell function that may even stimulate GvHD cannot be excluded by our experiments and need to be considered.15 Footnotes Information on authorship, contributions, and financial & other disclosures was provided by the authors and is available with the online version of this article at www.haematologica.org. Funding: this work was supported by a grant from Novartis Inc. to F.H.H. and partially by a DFG-grant (FI405/5-1) to T.F. and F.H and the Collaborative Research Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. and F.H.H. All human samples were collected and stored in the Hematology Tissue-Bank Magdeburg (HTM) supported by a grant from the Jose-Carreras Foundation (SP12/04) to F.H.H. F.H.H. received research funding from Novartis Inc. All other authors have nothing to disclose..Macroscopic analysis (soreness, Number 2D) as well as histology of ileum and colon was performed to assess for GvHD development. IFN- secreting cells. Autologous APC loaded with a viral peptide pool (CMVpp65; PepMixTM HCMVA(pp65), JPT Inc., Berlin, Germany) were incubated with T cells (of the same individual) in the absence or presence of the respective TKI (as indicated). Midostaurin (50nM) and quizartinib (50nM) did not impair lytic activity. 10nM dasatinib significantly inhibited the virus-specific T-cell response. APC loaded with control peptides (CEF) or without peptides served as positive and negative control. These results could be recapitulated using human being influenza virus derived peptides as an antigen. (DCG) Allogeneic (un-matched) GvHD (graft-versus-host-disease) assay in vivo: 0.5 106 CD3+ T cells derived from BL/6 mice were transplanted along with 2 106 BALB/c WBMC into lethally irradiated (13Gy) BALB/c recipient mice. Animals developed severe GvHD of the gut within 3C4 weeks after transplantation. Midostaurin treatment was performed after engraftment (Days 14C19; 100mg/kg body weight by gavage, every 24h). Ileum and colon fixed in formalin and inlayed in paraffin were analyzed for apoptosis of crypt cells on Day time 25. Midostaurin treatment (n=3) did not affect severity of GvHD in the mouse gut. Both, severity of GvH-reaction and induction of apoptosis in the ileum or colon were comparable to vehicle treated animals (n=2). Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken collectively our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human being T-cell signaling, proliferation and function unaffected. Although our study is limited by the use of HD-TC that may take action in a different way to T cells derived from AML individuals, our findings facilitate a pre-clinical assessment for the use of FLT3-TKI in the context of alloSCT. Without influencing T-cell function, midostaurin and quizartinib could be concomitantly used until discontinuation of immunosuppressive therapy and therefore prevent relapse prior to appearance of a sufficient GvL-response. Interestingly, differential effects of FLT3-TKI on dendritic cell function that may even stimulate GvHD cannot be excluded by our experiments and need to be regarded as.15 Footnotes Information on authorship, contributions, and financial & other disclosures was provided by the authors and is available with the online version of this article at www.haematologica.org. Funding: this work was supported by a give from Novartis Inc. to F.H.H. and partially by a DFG-grant (FI405/5-1) to T.F. and F.H and the Collaborative Study Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. and F.H.H. All human being samples were collected and stored in the Hematology Tissue-Bank Magdeburg (HTM) supported by a give from your Jose-Carreras Basis (SP12/04) to F.H.H. F.H.H. received study funding from Novartis Inc. All other authors have nothing to disclose..Midostaurin treatment (n=3) did not impact severity of GvHD in the mouse gut. control. Dasatinib impaired T-cell reactivity to less than 5% of control. (B, C) ELISPOT assay to assess for HD-TC-responses directed against viral antigens (n=10). Lytic T-cell activity was identified based on the rate of recurrence of IFN- secreting cells. Autologous APC loaded with a viral peptide pool (CMVpp65; PepMixTM HCMVA(pp65), JPT Inc., Berlin, Germany) were incubated with T cells (of the same individual) in the absence or presence of the respective TKI (as indicated). Midostaurin (50nM) and quizartinib (50nM) did not impair lytic activity. 10nM dasatinib significantly inhibited the virus-specific T-cell response. APC loaded with control peptides (CEF) or without peptides served as positive and negative control. These results could be recapitulated using human being influenza virus derived peptides as an antigen. (DCG) Allogeneic (un-matched) GvHD (graft-versus-host-disease) assay in vivo: 0.5 106 CD3+ T cells derived from BL/6 mice were transplanted along with 2 106 BALB/c WBMC into lethally irradiated (13Gy) BALB/c recipient mice. Animals developed severe GvHD of the gut within 3C4 weeks after transplantation. Midostaurin treatment was performed after engraftment (Days 14C19; 100mg/kg body weight by gavage, every 24h). Ileum and colon fixed in formalin and inlayed in paraffin were analyzed for apoptosis of crypt cells on Day time 25. Midostaurin treatment (n=3) did not affect severity of GvHD in the mouse gut. Both, severity of GvH-reaction and induction of apoptosis in the ileum or colon were comparable to vehicle treated animals (n=2). Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken collectively our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human being T-cell signaling, proliferation and function unaffected. Although our study is limited by the use of HD-TC that may take action in a different way to T cells derived from AML individuals, our findings facilitate a pre-clinical assessment for the use of FLT3-TKI in the context of alloSCT. Without influencing T-cell function, midostaurin and quizartinib could be concomitantly used until discontinuation of immunosuppressive therapy and therefore prevent relapse prior to appearance of a sufficient GvL-response. Interestingly, differential effects of FLT3-TKI on dendritic cell function that may even stimulate GvHD cannot be excluded by our experiments and need to be regarded as.15 Footnotes Information on authorship, contributions, and financial & other disclosures was provided by the authors and is available with the online version of this article at www.haematologica.org. Funding: this work was supported by a give from Novartis Inc. to F.H.H. and partially by a DFG-grant (FI405/5-1) to T.F. and F.H and the Collaborative Study Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. and F.H.H. All human being samples were collected and stored in the Hematology Tissue-Bank Magdeburg (HTM) supported by a give from your Jose-Carreras Basis (SP12/04) to F.H.H. F.H.H. received AR-9281 study funding from Novartis Inc. All other authors have nothing to disclose..and partially by a DFG-grant (FI405/5-1) to T.F. individual) in the absence or presence of the respective TKI (as indicated). Midostaurin (50nM) and quizartinib (50nM) did not impair lytic activity. 10nM dasatinib significantly inhibited the virus-specific T-cell response. APC loaded with control peptides (CEF) or without peptides served as positive and negative control. These results could be recapitulated using human being influenza virus derived Vav1 peptides as an antigen. (DCG) Allogeneic (un-matched) GvHD (graft-versus-host-disease) assay in vivo: 0.5 106 CD3+ T cells derived from BL/6 mice were transplanted along with 2 106 BALB/c WBMC into lethally irradiated (13Gy) BALB/c recipient mice. Animals developed severe GvHD of the gut within 3C4 weeks after transplantation. Midostaurin treatment was performed after engraftment (Days 14C19; 100mg/kg body weight by gavage, every 24h). Ileum and colon fixed in formalin and inlayed in paraffin were analyzed for apoptosis of crypt cells on Day time 25. Midostaurin treatment (n=3) did not affect severity of GvHD in the mouse gut. Both, severity of GvH-reaction and induction of apoptosis in the ileum or colon were comparable to vehicle treated animals (n=2). Dasatinib (2.5mg/kg) treated animals (n=3) revealed decreased induction of apoptosis and reduced severity of GvHD in both, ileum and colon. Taken collectively our results provide first evidence that clinically relevant doses of PKC412 and AC220 leave human being T-cell signaling, proliferation and function unaffected. Although our study is limited by the use of HD-TC that may take action in a different way to T cells derived from AML individuals, our findings facilitate a pre-clinical assessment for the use of FLT3-TKI in the context of alloSCT. Without influencing T-cell function, midostaurin and quizartinib could be concomitantly used until discontinuation of immunosuppressive therapy and therefore prevent relapse prior to appearance of a sufficient GvL-response. Interestingly, differential effects of FLT3-TKI on dendritic cell function that may even stimulate GvHD cannot be excluded by our experiments and need to be considered.15 Footnotes Information on authorship, contributions, and financial & other disclosures was provided by the authors and is available with the online version of this article at www.haematologica.org. Funding: this work was supported by a grant from Novartis Inc. to F.H.H. and partially by a DFG-grant (FI405/5-1) to T.F. and F.H and the Collaborative Research Cluster (CRC854) to T.M.S., S.K., B.S., M.B.W., B.I., T.F. and F.H.H. All human samples were collected and stored in the Hematology Tissue-Bank Magdeburg (HTM) supported by a grant from the Jose-Carreras Foundation (SP12/04) to F.H.H. F.H.H. received research funding from Novartis Inc. All other authors have nothing to disclose..