Multiplexed single‐cell mass cytometry reveals distinct inhibitory effects on intracellular phosphoproteins by midostaurin in combination with chemotherapy in AML cells

Experimental Hematology & Oncology

Thomas Gustafsson¹  Kourosh Lotfi²  Jan-Ingvar Jönsson³  Erik Hultin³  Emma Rörby³  Jörg Cammenga⁴  Jörgen Adolfsson⁵ 

[1]Department of Hematology, Linköping University Hospital, Linköping, Sweden

[2]Department of Hematology, Linköping University Hospital, Linköping, Sweden

[3]Clinical Pharmacology, Division of Drug Research, Department of Biomedical and Clinical Sciences, Linköping University, Linköping, Sweden

[4]Experimental Hematology Unit, Department of Biomedical and Clinical Sciences, Linköping University, 58185, Linköping, Sweden

[5]Experimental Hematology Unit, Department of Biomedical and Clinical Sciences, Linköping University, 58185, Linköping, Sweden

[6]Department of Hematology, Linköping University Hospital, Linköping, Sweden

[7]Experimental Hematology Unit, Department of Biomedical and Clinical Sciences, Linköping University, 58185, Linköping, Sweden

[8]Science for Life Laboratory, National Mass Cytometry Facility, Linköping University, Linköping, Sweden

关键词: Acute myeloid leukemia;    Intracellular therapy response;    Midostaurin;    Chemotherapy;    FLT3;    Signaling proteins;    Proteomics;    Mass cytometry;   

DOI  :  10.1186/s40164-021-00201-w

来源: Springer

【 摘 要 】

BackgroundFms-related tyrosine kinase 3 (FLT3) receptor serves as a prognostic marker and therapeutic target in acute myeloid leukemia (AML). Approximately one-third of AML patients carry mutation in FLT3, associated with unfavourable prognosis and high relapse rate. The multitargeted kinase inhibitor midostaurin (PKC412) in combination with standard chemotherapy (daunorubicin and cytarabine) was recently shown to increase overall survival of AML patients. For that reason, PKC412 has been approved for treatment of AML patients with FLT3-mutation. PKC412 synergizes with standard chemotherapy, but the mechanism involved is not fully understood and the risk of relapse is still highly problematic.MethodsBy utilizing the unique nature of mass cytometry for single cell multiparameter analysis, we have explored the proteomic effect and intracellular signaling response in individual leukemic cells with internal tandem duplication of FLT3 (FLT3-ITD) after midostaurin treatment in combination with daunorubicin or cytarabine.ResultsWe have identified a synergistic inhibition of intracellular signaling proteins after PKC412 treatment in combination with daunorubicin. In contrast, cytarabine antagonized phosphorylation inhibition of PKC412. Moreover, we found elevated levels of FLT3 surface expression after cytarabine treatment. Interestingly, the surface localization of FLT3 receptor increased in vivo on the blast cell population of two AML patients during day 3 of induction therapy (daunorubicin; once/day from day 1–3 and cytarabine; twice/day from day 1–7). We found FLT3 receptor expression to correlate with intracellular cytarabine (AraC) response. AML cell line cultured with AraC with or without PKC412 had an antagonizing phosphorylation inhibition of pAKT (p = 0.042 and 0.0261, respectively) and pERK1/2 (0.0134 and 0.0096, respectively) in FLT3high compared to FLT3low expressing cell populations.ConclusionsOur study provides insights into how conventional chemotherapy affects protein phosphorylation of vital signaling proteins in human leukemia cells. The results presented here support further investigation of novel strategies to treat FLT3-mutated AML patients with PKC412 in combination with chemotherapy agents and the potential development of novel treatment strategies.

【 授权许可】

CC BY