Pilot Study on Mass Spectrometry–Based Analysis of the Proteome of CD34+CD123+ Progenitor Cells for the Identification of Potential Targets for Immunotherapy in Acute Myeloid Leukemia

Abstract

Targeting of leukemic stem cells with specific immunotherapy would be an ideal approach for the treatment of myeloid malignancies, but suitable epitopes are unknown. The comparative proteome-level characterization of hematopoietic stem and progenitor cells from healthy stem cell donors and patients with acute myeloid leukemia has the potential to reveal differentially expressed proteins which can be used as surface-markers or as proxies for affected molecular pathways. We employed mass spectrometry methods to analyze the proteome of the cytosolic and the membrane fraction of CD34 and CD123 co-expressing FACS-sorted leukemic progenitors from five patients with acute myeloid leukemia. As a reference, CD34+CD123+ normal hematopoietic progenitor cells from five healthy, granulocyte-colony stimulating factor (G-CSF) mobilized stem cell donors were analyzed. In this Tandem Mass Tag (TMT) 10-plex labelling–based approach, 2070 proteins were identified with 171 proteins differentially abundant in one or both cellular compartments. This proof-of-principle-study demonstrates the potential of mass spectrometry to detect differentially expressed proteins in two compartment fractions of the entire proteome of leukemic stem cells, compared to their non-malignant counterparts. This may contribute to future immunotherapeutic target discoveries and individualized AML patient characterization.

Landscape of Tumor Suppressor Mutations in Acute Myeloid Leukemia

“…With the advent of mass-spectrometry-based methods performed directly on human AML-sorted stem cells, a significant number of leukemia-specific proteins, especially membrane-associated, have been identified. The main objective of this approach has been the identification of novel AML stem cell biomarkers to exploit as immunotherapeutic targets, in order to eradicate the disease [182][183][184][185]. Moreover, patients' proteomic profiles could correlate with the mutational status and thus with the prognosis of AML patients, suggesting that proteogenomic approaches might become the main goal in the near future.…”

CD34 cells in somatic, regenerative and cancer stem cells: Developmental biology, cell therapy, and omics big data perspective

“…In an attempt to identify molecular markers that can be targeted for treating AML, a comparative proteomic analysis of hematopoietic stem and progenitor cells isolated from patients with AML and from that of healthy individuals was carried out . A density gradient centrifugation method was used for the isolation of mononuclear cells from peripheral blood.…”

Properties of Leukemic Stem Cells in Regulating Drug Resistance in Acute and Chronic Myeloid Leukemias

“…Comparative tandem mass tag (TMT) multiplex proteomics analysis of CD34 + CD123 + AML patient LSCs and control HSCs identified 171 significantly differentially expressed proteins. Subsequent enrichment pathway analysis linked these proteins to gene ontology clusters such as cellular response to endogenous stimulus and cellular response to metal ion, among many others [ 156 ]. The distinct methodologies adopted by the aforementioned studies make it difficult to consolidate their findings and draw definitive conclusions regarding the proteomic landscape of AML LSCs; obstacles include different surface markers used to identify LSCs as well as the uneven coverage and depth of protein identification and quantification based on the disparate techniques employed.…”