k562 human cml cell line  (ATCC)

Journal: bioRxiv

Article Title: Targeting SOS1 synergistically enhances efficacy of BCR/ABL tyrosine kinase inhibitors and overcomes resistance in chronic myeloid leukemia

doi: 10.1101/2025.09.29.679122

Figure Lengend Snippet: (a–b) Time-course analysis of cell proliferation (a) and Apoptosis analysis (% Annexin V⁺ cells) (b) in K562 and KCL22 cells treated with increasing concentrations of imatinib (10–10000 nM, blue) or BI-3406 (10–10,000 nM, orange) for up to 48 h. Cell viability was measured by live object count normalized to time 0, and apoptosis was assessed by Annexin V staining. (c–d) Dose-response viability curves (c) and Annexin V apoptosis assays (d) after 48 h treatment as indicated with imatinib alone or in combination with fixed doses of BI-3406 (1, 10, or 100 nM). (e) Synergy analysis using ZIP scoring algorithm. 3D surface plots estimating synergy between BI-3406 and imatinib in both CML cell lines, with ZIP scores >20 indicating strong combinatorial effects. (f) Summary table of IC 50 values (mean ± SD) for imatinib calculated after 48 h co-treatment of both K562 and KCL22 cell lines. Data are mean ± SEM from at least three technical and biological replicates; ICDD values were calculated using non-linear regression (four-parameter logistic model) in GraphPad Prism v8.0. Statistical significance was assessed by one-way ANOVA with Tukey’s post-test; **p < 0.01; ***p<0.001.

Article Snippet: The human CML cell lines K562 (ATCC® CCL-243TM) and KCL22 (ATCC® CRL-3349TM), as well as murine Ba/F3-p210 BCR/ABL cells (control strain and derivatives carrying the T315I or the E255K BCR-ABL1 mutations), were maintained in RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS; Gibco), 1% penicillin–streptomycin, and 2 mM L-glutamine at 37°C in a humidified atmosphere with 5% COD.

Techniques: Staining

Journal: bioRxiv

Article Title: Targeting SOS1 synergistically enhances efficacy of BCR/ABL tyrosine kinase inhibitors and overcomes resistance in chronic myeloid leukemia

doi: 10.1101/2025.09.29.679122

Figure Lengend Snippet: (a) Dose–response viability curves of K562 and KCL22 cells treated with dasatinib (left panels), nilotinib (middle panels), or ponatinib (right panels), alone or in combination with fixed doses of BI-3406 (1 nM, 10 nM, or 100 nM). BI-3406 monotherapy had minimal effect on viability but markedly potentiated the cytotoxic activity of all three TKIs in both cell lines, resulting in a pronounced reduction in ICDD values (see table in panel c). (b) Synergy analysis of BI-3406 in combination with dasatinib, nilotinib, or ponatinib using the ZIP model. 3D surface plots show areas of synergy (red) across concentration matrices. ZIP synergy scores >10 denote strong combinatorial interaction. Synergistic effects were observed in all conditions, with particularly high scores for Nilotinib combinations. Data presented as mean ± SEM from three independent replicates. ICDD values were calculated after 48 h treatment.

Article Snippet: The human CML cell lines K562 (ATCC® CCL-243TM) and KCL22 (ATCC® CRL-3349TM), as well as murine Ba/F3-p210 BCR/ABL cells (control strain and derivatives carrying the T315I or the E255K BCR-ABL1 mutations), were maintained in RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS; Gibco), 1% penicillin–streptomycin, and 2 mM L-glutamine at 37°C in a humidified atmosphere with 5% COD.

Techniques: Activity Assay, Concentration Assay

Journal: bioRxiv

Article Title: Targeting SOS1 synergistically enhances efficacy of BCR/ABL tyrosine kinase inhibitors and overcomes resistance in chronic myeloid leukemia

doi: 10.1101/2025.09.29.679122

Figure Lengend Snippet: (a) Western blot analysis of serum-starved K562 and KCL22 cells stimulated with EGF (100 ng/mL) for 2, 5, or 10 minutes, either alone or following pre-treatment with BI-3406 (1 μM). Active GTP-bound RAS, RAC and MRAS were assessed by pull-down assays as described in Methods. Tubulin (TUB) was used as a loading control for all blots. Quantification of RAS-GTP, RAC-GTP, pERK, and pAKT levels normalized to total protein is shown on the right. Graphs represent mean ± SEM from at least three independent experiments. (b) Cells were treated for 24 hours with BI-3406 (1μM), imatinib (IMA, 500nM), or their combination, and phospho-ERK and phospho-AKT levels were assessed by Western blot. Tubulin was used as loading control. Representative of at least three independent experiments. Blue asterisks indicate significance vs. imatinib alone; orange asterisks indicate significance vs. BI-3406 alone and black asterisks indicate significance vs vehicle treated alone. Statistical significance was assessed by one-way ANOVA with Tukey’s post-test; * p<0.05; **p < 0.01; ***p<0.001.

Article Snippet: The human CML cell lines K562 (ATCC® CCL-243TM) and KCL22 (ATCC® CRL-3349TM), as well as murine Ba/F3-p210 BCR/ABL cells (control strain and derivatives carrying the T315I or the E255K BCR-ABL1 mutations), were maintained in RPMI-1640 medium (Gibco) supplemented with 10% fetal bovine serum (FBS; Gibco), 1% penicillin–streptomycin, and 2 mM L-glutamine at 37°C in a humidified atmosphere with 5% COD.

Techniques: Western Blot, Control