Journal: Frontiers in Pharmacology

Article Title: Visomitin as a differentiation-inducing therapeutic agent through SYK inhibition in AML

doi: 10.3389/fphar.2026.1741351

Figure Lengend Snippet: Visomitin induces cell differentiation in AML. (A) CD14 expression was detected by FACS analysis after the Visomitin treatment (500 nM) for 72 h in U937 and HL 60 cell lines. (B) Giemsa staining results showing cell differentiation in AML cell lines in response to a 500 nM Visomitin treatment for 72 h in HL60 and U937 cell lines. (C) Phagocytosis was assessed by flow cytometry and fluorescence microscopy in U937 cells, with representative images shown from three independent experiments. (D) Oxidative burst was assessed by FACS analysis in U937 cells following 72-h treatment with 500 nM Visomitin using Dihydrorhodamine-123 staining. (E) Relative expression levels of myeloid differentiation-related genes were measured by qRT-PCR in U937 cells at 72-h intervals post 500 nM Visomitin treatment. (F) Relative expression levels of macrophage gene signatures were measured by qRT-PCR in U937 cells after 72-h treatment with 500 nM Visomitin. The data are presented as mean ± SD (n = 3) for each group and statistical analysis was performed using the Student’s t-test and two-tailed Mann–Whitney U test (*p < 0.05).

Article Snippet: The human AML cell lines HL60, U937 and mouse AML cell line M1 were purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Cell Differentiation, Expressing, Staining, Flow Cytometry, Fluorescence, Microscopy, Quantitative RT-PCR, Two Tailed Test, MANN-WHITNEY

Journal: Frontiers in Pharmacology

Article Title: Visomitin as a differentiation-inducing therapeutic agent through SYK inhibition in AML

doi: 10.3389/fphar.2026.1741351

Figure Lengend Snippet: Visomitin induces cell cycle arrest in AML cells. (A) HL60 and U937 cells were treated with Visomitin (1 μM) for 24 h. Cell cycle was analyzed by FACS after PI staining. Expression levels of Cyclin D1 and CDK4 (B) , as well as p21 and p16 (C) , were analyzed by Western blotting after treatment with 500 nM Visomitin for 24 h in U937 cells. β-actin was used as a loading control. All data are presented as mean ± SD and representative of three independent experiments and statistical analysis was performed using the two-tailed Mann–Whitney U test (*p < 0.05).

Article Snippet: The human AML cell lines HL60, U937 and mouse AML cell line M1 were purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Staining, Expressing, Western Blot, Control, Two Tailed Test, MANN-WHITNEY

Journal: Frontiers in Pharmacology

Article Title: Visomitin as a differentiation-inducing therapeutic agent through SYK inhibition in AML

doi: 10.3389/fphar.2026.1741351

Figure Lengend Snippet: Visomitin induces cell apoptosis in AML. (A) HL60 and U937 cells were treated with Visomitin at indicated concentrations for 48 h. Cell viability was measured using MTS assay. (B) Trypan blue staining was used to count cell numbers after treatment with Visomitin in HL60 and U937 cells at the indicated times and concentrations. (C) The apoptotic rate was assessed by FACS after PI/Annexin V staining in U937 cells treated with 500 nM Visomitin for 72 h. Statistical analysis was performed using the one-way ANOVA (*p < 0.05). (D) A colony forming assay was performed and quantified in U937 and HL60 cells treated with Visomitin (500 nM) for 7 days. (E) Western blot analysis of Bcl-2 family members in U937 cells treated with 500 nM Visomitin. MCL-1 and Bcl-xL were analyzed after 24 h treatment; Bax and Bak after 1 h β-actin was used as a loading control. (F) U937 cells were treated with 500 nM Visomitin for 24 h. To monitor mitochondrial membrane potential (MMP), the cells were stained with JC-1, and the JC-1 fluorescence ratio was calculated using flow cytometry. Intact and disrupted MMP were represented by red (J-aggregates) and green (J-monomers) fluorescence, respectively. (G) Caspase-3/7 activity was measured by ELISA-based bioluminescence assays following 24-h treatment with 500 nM Visomitin in HL60 cells. (H) Trypan blue staining was used to assess cell viability after treatment with 500 nM Visomitin for 24 h in primary AML patient bone marrow cells. Statistical analysis was performed using the two-tailed Mann–Whitney U test (*p < 0.05). All data are presented as mean ± SD (n = 3) for each group.

Article Snippet: The human AML cell lines HL60, U937 and mouse AML cell line M1 were purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: MTS Assay, Staining, Western Blot, Control, Membrane, Fluorescence, Flow Cytometry, Activity Assay, Enzyme-linked Immunosorbent Assay, Two Tailed Test, MANN-WHITNEY

Journal: Frontiers in Pharmacology

Article Title: Visomitin as a differentiation-inducing therapeutic agent through SYK inhibition in AML

doi: 10.3389/fphar.2026.1741351

Figure Lengend Snippet: Visomitin did not increase mtROS level in normal myeloid cells. Mitochondrial ROS levels and apoptosis were assessed by flow cytometry following treatment with 500 nM Visomitin for 24 h. Analyses were performed on (A) normal mouse bone marrow cells, (B) myeloid cells (CD45 + /CD11b + double-positive cells isolated from bone marrow), and (C) the mouse AML cell line M1. Apoptotic cells were identified by Annexin V staining. Data are presented as mean ± SD (n = 3 per group). Statistical significance was determined using the two-tailed Mann–Whitney U test (*p < 0.05).

Article Snippet: The human AML cell lines HL60, U937 and mouse AML cell line M1 were purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Flow Cytometry, Isolation, Staining, Two Tailed Test, MANN-WHITNEY

Journal: Frontiers in Pharmacology

Article Title: Visomitin as a differentiation-inducing therapeutic agent through SYK inhibition in AML

doi: 10.3389/fphar.2026.1741351

Figure Lengend Snippet: Visomitin-mediated ROS accumulation inhibits SYK activation. (A) U937 cells were treated with Visomitin 500 nM for 24 h. The expression levels of phospho-SYK and total SYK were detected by Western blotting. (B) The expression levels of phospho-SYK and total SYK were detected by Western blotting in U937 cells treated with 100 nM hydrogen peroxide for 24 h. Statistical analysis was performed using the two-tailed Mann–Whitney U test (*p < 0.05). (C) CD14 expression was analyzed by FACS in U937 and HL60 cells following 72 h treatment with 10 μM SYK inhibitor, BAY 61-3606. (D) Apoptotic cells were detected by FACS analysis after Annexin V staining in U937 and HL60 cells treated with 10 μM BAY 61-3606 for 72 h. The statistical analysis was performed using the two-tailed Mann–Whitney U test (*p < 0.05). (E) U937 cells were treated with 2 μM BAY 61-3606 for 24 h, and cell viability was assessed using the MTS assay. (F) CD14 expression was analyzed by FACS in U937 cells following 72 h treatment with 500 nM Visomitin and transfection with 100 ng of SYK overexpression vector or control vector. (G) Apoptotic cells were detected by FACS after Annexin V staining. Statistical analysis was performed using one-way ANOVA (*p < 0.05). (H) Total SYK and phospho-SYK expression were detected by Western blotting in U937 cells treated with 100 nM Doxorubicin for 24 h β-actin was used as a loading control. Statistical analysis was performed using the two-tailed Mann–Whitney U test (*p < 0.05). All data are presented as mean ± SD (n = 3) for each group.

Article Snippet: The human AML cell lines HL60, U937 and mouse AML cell line M1 were purchased from Korean Cell Line Bank (Seoul, Korea).

Techniques: Activation Assay, Expressing, Western Blot, Two Tailed Test, MANN-WHITNEY, Staining, MTS Assay, Transfection, Over Expression, Plasmid Preparation, Control