SYK inhibition targets acute myeloid leukemia stem cells by blocking their oxidative metabolism

Spleen tyrosine kinase (SYK) is an important oncogene and signaling mediator activated by cell surface receptors crucial for acute myeloid leukemia (AML) maintenance and progression. Genetic or pharmacologic inhibition of SYK in AML cells leads to increased differentiation, reduced proliferation, and cellular apoptosis. Herein, we addressed the consequences of SYK inhibition to leukemia stem-cell (LSC) function and assessed SYK-associated pathways in AML cell biology. Using gain-of-function MEK kinase mutant and constitutively active STAT5A, we demonstrate that R406, the active metabolite of a small-molecule SYK inhibitor fostamatinib, induces differentiation and blocks clonogenic potential of AML cells through the MEK/ERK1/2 pathway and STAT5A transcription factor, respectively. Pharmacological inhibition of SYK with R406 reduced LSC compartment defined as CD34+CD38-CD123+ and CD34+CD38-CD25+ in vitro, and decreased viability of LSCs identified by a low abundance of reactive oxygen species. Primary leukemic blasts treated ex vivo with R406 exhibited lower engraftment potential when xenotransplanted to immunodeficient NSG/J mice. Mechanistically, these effects are mediated by disturbed mitochondrial biogenesis and suppression of oxidative metabolism (OXPHOS) in LSCs. These mechanisms appear to be partially dependent on inhibition of STAT5 and its target gene MYC, a well-defined inducer of mitochondrial biogenesis. In addition, inhibition of SYK increases the sensitivity of LSCs to cytarabine (AraC), a standard of AML induction therapy. Taken together, our findings indicate that SYK fosters OXPHOS and participates in metabolic reprogramming of AML LSCs in a mechanism that at least partially involves STAT5, and that SYK inhibition targets LSCs in AML. Since active SYK is expressed in a majority of AML patients and confers inferior prognosis, the combination of SYK inhibitors with standard chemotherapeutics such as AraC constitutes a new therapeutic modality that should be evaluated in future clinical trials

Harmonization of Flow Cytometric Minimal Residual Disease Assessment in Multiple Myeloma in Centers of Polish Myeloma Consortium

Minimal residual disease (MRD) status is now considered as one of the most relevant prognostic factors in multiple myeloma (MM) while MRD negativity became an important endpoint in clinical trials. Here, we report the results of the first study evaluating the reproducibility of high-sensitivity flow cytometry MM MRD assessment in four laboratories in Poland. EuroFlow protocols for instrument setting standardization and sample preparation in MM MRD assessment were implemented in each laboratory. In the inter-laboratory reproducibility study, 12 bone marrow samples from MM patients were distributed and processed in participant laboratories. In the inter-operator concordance study, 13 raw data files from MM MRD measurements were analyzed by five independent operators. The inter-laboratory study showed high 95% overall concordance of results among laboratories. In the inter-operator study, 89% of MRD results reported were concordant, and the highest immunophenotype interpretation differences with regard to expression of CD27, CD45, CD81 were noticed. We confirmed the applicability and feasibility of the EuroFlow protocol as a highly sensitive method of MRD evaluation in MM. Results of our inter-center comparison study demonstrate that the standardization of MM MRD assessment protocols is highly desirable to improve quality and comparability of results within and between different clinical trials