Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: a , Western blot analysis of p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), and ERK1/2 in A549 and HCC827 cells treated for 24h with 1 µM axitinib, 1µM imatinib, 1 µM lenvatinib or 1 µM regorafenib. b, Western blot analysis of phosphorylated tyrosine kinases, p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), and ERK1/2 in A549 and CFPAC following treatment with trametinib alone or in combination with imatinib (1 μM) or erlotinib (1 μM). c, Dot plot showing log₂ fold changes in phosphatase gene expression between the combination-treated and untreated A549 cells. d, Colony formation assays demonstrating synergistic anti-cancer effects of trametinib and imatinib in KRAS mutant A549 and CFPAC cells, as well as KRAS -WT H1975 and HCC827 cells treated with indicated concentrations for 6 days. e, Colony formation assay of A549 and CFPAC cells treated for 6 days with sotorasib, adagrasib, imatinib (1 µM), trametinib (30 nM), or imatinib-trametinib combination at the indicated concentrations. Data represent mean ± SD from at least three independent experiments. ***p < 0.001. f, Colony formation assays of H23 and MiaPaCa2 cells treated as described in (e). Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Western Blot, Gene Expression, Mutagenesis, Colony Assay

Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: a , Dot plot of pathway analysis of the genes up-regulated in KRAS mutant cells treated with the combination of imatinib and trametinib compared to untreated cells using KEGG datasets. b, Western blot analysis of LC3I/II, NCOA4 and HIF-1α in A549, CFPAC, H1975 and HCC827 cells treated with imatinib and/or trametinib for 72 h. c, The levels of lipid peroxidation in A549 and CFPAC cells treated with 1 µM imatinib and/or 30 nM trametinib in the presence or absence of 1 µM ferrostatin for 72 h. Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001. d, The levels of lipid peroxidation in A549 and CFPAC cells treated with 1µM imatinib and/or 30nM trametinib in the presence or absence of 10µM hydroxychloroquine (HCQ) for 72 h. Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001. e, Confocal microscopy analysis of Fe²⁺ using iron sensing dye FerroOrange in A549, CFPAC, H1975, and HCC827 cells treated with 1µM imatinib, 30 nM trametinib, or the combination for 72 h. f, Colony formation assay of A549 and CFPAC cells treated with 1 µM imatinib and/or 30 nM trametinib in the presence/absence of 10 µM HCQ, 1 µM ferrostatin, or 2 µM liproxstatin for 6 days. Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001. g, Graphical summary of the mechanism of ferritinophagy and ferroptosis induction by the combination of imatinib and trametinib in KRAS mutant cells.

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Mutagenesis, Western Blot, Confocal Microscopy, Colony Assay

Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: a , Tumor growth curves of NPG mice bearing CFPAC KRAS G12V mutant pancreatic cancer xenografts treated with vehicle, imatinib (15 mg/kg), trametinib (0.3 mg/kg), or the combination. Data represent mean ± SEM; n = 6 mice/group. b, Luminescence signals representing xenograft tumor sizes on day 30 used in ( a ) (n=6 mice/group). Data represent mean ± SD from six mice. *p < 0.05, **p < 0.01, ***p < 0.001. c, Survival curve of the mice used in ( a ). d, Representative IHC images (20x) of Ki67, p-AKT (S473), p-MEK1/2 (S217/221), p-ERK1/2 (Y204/T202), NCOA4 and HIF-1⍺ in the tumors of the mice on day 30 used in ( a ). e, Quantification of IHC staining shown in ( d ). Data represent mean ± SD from three mice. *p < 0.05, **p < 0.01, ***p < 0.001. f, Tumor growth curves of ASID mice bearing A549 KRAS G12S mutant lung cancer xenografts treated orally with vehicle, imatinib (15 mg/kg), trametinib (0.3 mg/kg), or the combination. Data represent mean ± SEM; n = 5 mice/group. g, Luminescence signals representing xenograft tumor sizes on day 30 used in ( f ) (n=5 mice/group). Data represent mean ± SD from five mice. ***p < 0.001. h, Representative IHC images (20x) of Ki67, p-AKT (S473), p-MEK1/2 (S217/221), p-ERK1/2 (Y204/T202), NCOA4 and HIF-1⍺ in the tumors of the mice on day 30 used in ( f ). i, Quantification of IHC staining shown in ( h ). Data represent mean ± SD from three mice. *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Mutagenesis, Immunohistochemistry

Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: a , Primary KRAS G12D mutant pancreatic cancer cells were treated with imatinib, trametinib, or the combination at the indicated concentrations for 72 h, and their lysates were analyzed by western blot to detect p-AKT (S473), AKT, p-ERK1/2(Y204/T202), ERK1/2, LC3I/II, NCOA4 and HIF-1⍺ expression. b, Schematic diagram of generation of pancreatic patient-derived xenograft (PDX) mice. c, PDX tumor growth curves of NPG mice treated with vehicle, imatinib (15 mg/kg), trametinib (0.3 mg/kg), or the combination. Data represent mean ± SEM; n = 6 mice/group. d, Tumor size of the mice on day 30 used in ( c ). Data represent mean ± SD from six mice. *p < 0.05, **p < 0.01, ***p < 0.001. e, Survival curve of the mice used in ( c ). f, Western blot analysis of p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), ERK1/2, LC3I/II, ferroptoisis-associated genes, including NCOA4 and HIF-1⍺ in the tumors of the mice on day 30 used in ( c ). g, Representative IHC images (20×) of Ki67, p-AKT (S473), p-MEK1/2 (S217/221), p-ERK1/2 (Y204/T202), NCOA4 and HIF-1⍺ in the tumors of the mice on day 30 used in ( c ). h, Quantification of IHC staining shown in ( g ). Data represent mean ± SD from three mice. *p < 0.05, **p < 0.01, ***p < 0.001.

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Mutagenesis, Western Blot, Expressing, Derivative Assay, Immunohistochemistry

Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: a , Cell viability assay comparing parental H23 and sotorasib (AMG-510)-resistant H23R cells treated with the indicated concentrations of sotorasib. Data are presented as mean ± SD. b, Western blot analysis of phosphorylated tyrosine kinases, p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), and ERK1/2 in H23 and H23R cells. c, Western blot analysis of p-EGFR (Y1068), EGFR, p-PDGFR⍺ (Y849) /PDGFRβ (Y857) and PDGFR⍺/β in H23 and H23R cells. d, Western blot analysis of phosphorylated tyrosine kinases, p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), and ERK1/2 in H23R cells following treatment with trametinib alone or in combination with imatinib (1 μM) or erlotinib (1 μM). e, Colony formation assays of H23R cells treated with sotorasib, imatinib (1 μM), trametinib (30 nM), or the imatinib-trametinib combination at the indicated concentrations for 6 days. Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01,***p < 0.001. f, Western blot analysis of p-AKT (S473), AKT, p-ERK1/2 (Y204/T202), ERK1/2, LC3I/II, NCOA4 and HIF-1α in H23R cells treated with imatinib and trametinib for 24 h. g, The levels of lipid peroxidation in H23R cells treated with imatinib (1 μM) and/or trametinib (30 nM), in the presence/absence of ferrostatin (1 μM) for 2 days. Data represent mean ± SD from at least three independent experiments. *p < 0.05, **p < 0.01, ***p < 0.001. h, The levels of lipid peroxidation in H23R cells treated with imatinib (1 μM) and/or trametinib (30 nM), in the presence/absence of hydroxychloroquine (HCQ) 10 μM for 2 days. Data represent mean ± SD from at least three independent experiments. **p < 0.01. i, Colony formation assay of H23R cells treated with imatinib (1 μM) and/or trametinib (30 nM) in the presence/absence of hydroxychloroquine (HCQ, 10 μM), ferrostatin (1 μM), or liproxstatin (2 μM) for 6 days. Data represent mean ± SD from at least three independent experiments. ***p < 0.001.

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Viability Assay, Western Blot, Colony Assay

Journal: medRxiv

Article Title: Reciprocal Feedback Blockade with Trametinib and Imatinib Overcomes the Limitations of Current KRAS-targeted Therapy

doi: 10.64898/2026.07.01.26356985

Figure Lengend Snippet: Clinical evaluation of the combination of imatinib and trametinib in patient with KRAS non-G12C mutant, chemotherapy-refractory pancreatic cancer a, Change of carbohydrate antigen 19-9 (CA19-9) of four participants following start of therapy. b, Kaplan-Meier plots of progression-free survival (PFS). c, Kaplan-Meier plots of overall survival (OS).

Article Snippet: Imatinib, trametinib, and hydroxychloroquine were purchased from MedChemExpress.

Techniques: Mutagenesis