kras mutant cancer cell lines  (Human Protein Atlas)

Journal: Cancer Research Communications

Article Title: Constitutive EGFR Activation Induced by PTPRR Downregulation Confers Resistance to KRAS Inhibitors

doi: 10.1158/2767-9764.CRC-25-0489

Figure Lengend Snippet: H2122AR14 and H2122AR30 cells are resistant to sotorasib in vitro and show EGFR activation. A, Cell viability assay for KRAS G12C -mutant parental H2122 cells and H2122AR14 and H2122AR30 sotorasib-resistant clones exposed to the indicated concentrations of sotorasib for 72 hours. Data are means ± SD ( n = 3 independent experiments). B, Immunoblot analysis of KRAS, phosphorylated (p) and total (t) forms of AKT and ERK, and actin (loading control) in H2122, H2122AR14, and H2122AR30 cells treated with sotorasib (1 μmol/L) for 0 or 24 hours. C, Human phospho-RTK assay for H2122, H2122AR14, and H2122AR30 cells treated with 1 μmol/L sotorasib for 72 hours. D, Immunoblot analysis of Y845-, Y1068-, or Y1173-phosphorylated and total forms of EGFR in H2122 and H2122AR30 cells. Data in B–D are representative of at least three independent experiments.

Article Snippet: The KRAS G12C -mutant NSCLC cell lines NCI-H2122 (RRID: CVCL_1531), SW1573 (RRID: CVCL_1720), and NCI-H358 (RRID: CVCL_1559); the EGFR -mutated NSCLC cell line NCI-H1975 (RRID: CVCL_1511); and the KRAS G12C -mutant colorectal cancer cell line SW837 (RRID: CVCL_1729) were obtained from the ATCC.

Techniques: In Vitro, Activation Assay, Viability Assay, Mutagenesis, Clone Assay, Western Blot, Control

Journal: Cancer Research Communications

Article Title: Constitutive EGFR Activation Induced by PTPRR Downregulation Confers Resistance to KRAS Inhibitors

doi: 10.1158/2767-9764.CRC-25-0489

Figure Lengend Snippet: Combination treatment with EGFR and KRAS G12C inhibitors overcomes sotorasib resistance due to PTPRR downregulation. A, Cell viability assay for H2122 cells transfected with nonspecific control (siSCR) or PTPRR-specific (siPTPRR) siRNAs and then exposed to the indicated concentrations of sotorasib for 72 hours. B, Immunoblot analysis of PTPRR as well as total and phosphorylated forms of AKT and ERK in H2122 cells transfected with siRNAs as in ( A ) and then incubated with or without sotorasib (1 μmol/L) for 4 hours. C, Cell viability assay for H2122AR30 cells stably transfected with control (GFP) or PTPRR expression plasmids and treated with the indicated concentrations of sotorasib for 72 hours. D, Immunoblot analysis of total or phosphorylated forms of EGFR, AKT, and ERK in H2122AR30 cells treated with sotorasib (1 μmol/L), cetuximab (10 μg/mL), or the combination of both drugs for 4 hours. E, Colony formation assay for H2122 and H2122AR30 cells incubated with or without sotorasib (1 μmol/L), cetuximab (10 μg/mL), or both drugs for 4 weeks. Surviving cells were stained with crystal violet. Data in A and C are means ± SD ( n = 3 independent experiments), and those in B , D , and E are representative of at least three independent experiments.

Article Snippet: The KRAS G12C -mutant NSCLC cell lines NCI-H2122 (RRID: CVCL_1531), SW1573 (RRID: CVCL_1720), and NCI-H358 (RRID: CVCL_1559); the EGFR -mutated NSCLC cell line NCI-H1975 (RRID: CVCL_1511); and the KRAS G12C -mutant colorectal cancer cell line SW837 (RRID: CVCL_1729) were obtained from the ATCC.

Techniques: Viability Assay, Transfection, Control, Western Blot, Incubation, Stable Transfection, Expressing, Colony Assay, Staining

Journal: Cancer Research Communications

Article Title: Constitutive EGFR Activation Induced by PTPRR Downregulation Confers Resistance to KRAS Inhibitors

doi: 10.1158/2767-9764.CRC-25-0489

Figure Lengend Snippet: Expression of PTPRR is regulated by promoter DNA methylation in KRAS G12C -mutant NSCLC cell lines. A, The human PTPRR gene including the promoter regions for the long and short isoforms of the encoded protein is shown. The positions of CpG islands as well as the histone marks H3K4me1, H3K4me3, and H3K27ac are indicated. B, Abundance of transcripts for the long and short isoforms of PTPRR in tumor specimens from patients with NSCLC in the dbGaP database ( n = 73). Data are presented as dot plots with means ± SD indicated by the dashed and solid lines for the short isoform, and they were analyzed with the Wilcoxon ranked-sum test. C, RT-qPCR analysis of mRNA abundance for the short form of PTPRR in the indicated KRAS G12C -mutant cell lines. Data are presented as dot plots, with means + SD being indicated by the dashed and solid lines ( n = 3 independent experiments) and analyzed by one-way ANOVA and Dunnett’s test. D, Bisulfite genomic sequencing of the promoter region for the short variant of PTPRR in the indicated KRAS G12C -mutant cell lines. The percentage of methylated CpG sites among total CpG sites is shown. Data are presented as violin plots, with the median and upper quartile values indicated by the dashed and dotted lines and analyzed by one-way ANOVA and Dunnett’s test. Data are representative of three independent experiments.

Article Snippet: The KRAS G12C -mutant NSCLC cell lines NCI-H2122 (RRID: CVCL_1531), SW1573 (RRID: CVCL_1720), and NCI-H358 (RRID: CVCL_1559); the EGFR -mutated NSCLC cell line NCI-H1975 (RRID: CVCL_1511); and the KRAS G12C -mutant colorectal cancer cell line SW837 (RRID: CVCL_1729) were obtained from the ATCC.

Techniques: Expressing, DNA Methylation Assay, Mutagenesis, Quantitative RT-PCR, Genomic Sequencing, Variant Assay, Methylation

Journal: Cancer Research Communications

Article Title: Constitutive EGFR Activation Induced by PTPRR Downregulation Confers Resistance to KRAS Inhibitors

doi: 10.1158/2767-9764.CRC-25-0489

Figure Lengend Snippet: Clinical relevance of PTPRR expression to sotorasib treatment in individuals with NSCLC. A, RT-qPCR analysis of PTPRR mRNA (short form) abundance in normal lung tissue ( n = 9) and NSCLC tumor tissue ( n = 10). Data are presented as dot plots with means ± SD indicated by the dashed and solid lines, and they were analyzed with the Wilcoxon ranked-sum test. B, Representative IHC images for specimens of normal lung tissue (negative) and NSCLC tumor tissue (low and high) stained with antibodies to PTPRR. Scale bars, 100 μm. C and D, Kaplan–Meier analysis of PFS ( C ) and OS ( D ) according to low ( n = 5) or high ( n = 8) status for cytoplasmic PTPRR staining in tumor cells for patients with KRAS G12C -mutant NSCLC treated with sotorasib.

Article Snippet: The KRAS G12C -mutant NSCLC cell lines NCI-H2122 (RRID: CVCL_1531), SW1573 (RRID: CVCL_1720), and NCI-H358 (RRID: CVCL_1559); the EGFR -mutated NSCLC cell line NCI-H1975 (RRID: CVCL_1511); and the KRAS G12C -mutant colorectal cancer cell line SW837 (RRID: CVCL_1729) were obtained from the ATCC.

Techniques: Expressing, Quantitative RT-PCR, Staining, Mutagenesis

Journal: ACS Chemical Biology

Article Title: Distal Covalent Targeting Suppresses Signaling of Oncogenic K‑Ras(G13C) in Cancer Cells

doi: 10.1021/acschembio.5c00249

Figure Lengend Snippet: (A) Alignment of K-Ras­(G13C) to adagrasib-bound K-Ras­(G12C). (B) Modular design of shift-register electrophiles to repurpose K-Ras­(G12C) covalent inhibitors for the G13C mutation. (C) Proximity scanning of Cys13 accessibility using unbranched alkyl linkers. (D) Labeling efficiency of G13C-targeting covalent inhibitors with various linker lengths. Conditions: K-Ras­(G13C)•GDP or •GppNHp (1 μM), compound (100 μM), RT incubation. Each box represents the mean of two independent runs. (E) Schematic summary of an expanded linker screening campaign. (F) Lead 1,3-disubstituted aromatic linked compounds and their covalent modification kinetics. Conditions: K-Ras­(G13C)•GDP or •GppNHp (200 nM), compound (10 μM), RT incubation.

Article Snippet: HCT116/KRAS G13C heterozygous and homozygous (SL753) cancer biomarker mutant cell lines were obtained from GeneCopoeia, Inc. (Rockville, MD).

Techniques: Mutagenesis, Labeling, Incubation, Modification

Journal: ACS Chemical Biology

Article Title: Distal Covalent Targeting Suppresses Signaling of Oncogenic K‑Ras(G13C) in Cancer Cells

doi: 10.1021/acschembio.5c00249

Figure Lengend Snippet: (A) Chemical structure of lead compound G13Ci-22 . (B) Aligned ligand poses of G13Ci-22 (CovDock) and MRTX1133 (PDB Entry 7RPZ ) in the corresponding Switch-II Pocket. (C) Covalent modification kinetics of K-Ras­(G13C) by inhibitors with the optimal 3-acrylamidobenzene carbonyl linker and alternative Switch-II Pocket binding moieties. Conditions: K-Ras­(G13C)•GDP or •GppNHp (200 nM), Compound (10 μL), RT incubation. (D) Chemical structures of the alternative Switch-II Pocket binding moieties. The optimal linker and Cys-warhead, same with those in G13Ci-22 , were omitted in the chemical structures.

Article Snippet: HCT116/KRAS G13C heterozygous and homozygous (SL753) cancer biomarker mutant cell lines were obtained from GeneCopoeia, Inc. (Rockville, MD).

Techniques: Modification, Binding Assay, Incubation

Journal: ACS Chemical Biology

Article Title: Distal Covalent Targeting Suppresses Signaling of Oncogenic K‑Ras(G13C) in Cancer Cells

doi: 10.1021/acschembio.5c00249

Figure Lengend Snippet: (A) Mutant selectivity of G13Ci-22 against relevant cyslight K-Ras proteins. (B) Thermal stabilization of K-Ras­(G13C) proteins due to covalent modification by G13Ci-22 . (C) Schematic description of guanine nucleotide cycles of K-Ras, K-Ras­(G13C), and covalently inhibited K-Ras­(G13C) GTPases. G13Ci-22 inhibited Ras-RafRBD binding (D), GTPase activity (E), and nucleotide exchange (F) of K-Ras­(G13C) oncoprotein.

Article Snippet: HCT116/KRAS G13C heterozygous and homozygous (SL753) cancer biomarker mutant cell lines were obtained from GeneCopoeia, Inc. (Rockville, MD).

Techniques: Mutagenesis, Modification, Binding Assay, Activity Assay

Journal: ACS Chemical Biology

Article Title: Distal Covalent Targeting Suppresses Signaling of Oncogenic K‑Ras(G13C) in Cancer Cells

doi: 10.1021/acschembio.5c00249

Figure Lengend Snippet: (A) Immunoblot of HCT116 cell and its genetically engineered derivative cells treated with G13Ci-22 at various concentrations. (B) Time-course immunoblot of the homozygous HCT116 (K-Ras­(G13C/G13C)) cell treated with 10 nM of G13Ci-22 . (C) Immunoblot of human cancer cell lines NCI-H1355 and NCI-H1734, which harbor K-Ras­(G13C) mutation, treated with G13Ci-22 .

Article Snippet: HCT116/KRAS G13C heterozygous and homozygous (SL753) cancer biomarker mutant cell lines were obtained from GeneCopoeia, Inc. (Rockville, MD).

Techniques: Western Blot, Mutagenesis

Journal: Journal of Clinical Oncology

Article Title: Onvansertib in Combination With Chemotherapy and Bevacizumab in Second-Line Treatment of KRAS -Mutant Metastatic Colorectal Cancer: A Single-Arm, Phase II Trial

doi: 10.1200/JCO-24-01266

Figure Lengend Snippet: PLK1 regulates the hypoxia pathway, related to . (A, B) KRAS -mutant CRC cells were treated with DMSO or Onv at the indicated doses for 20 hours and then exposed to hypoxia or kept in normoxia for 4 hours. (A) Heatmap of hypoxia-related genes significantly regulated by Onv in HCT116 and SW620 cells based on the RNA-seq analysis. (B) Expression of hypoxia-related genes in LoVo and DLD-1 cells, assessed by RT-qPCR and normalized to the housekeeping gene RPLP0 . Bar graphs represent expression relative to Normoxia_DMSO sample. (C, D) SW620 and HCT116 cells were transfected with nontargeting control siRNA (siNTC) or PLK1 targeting siRNA (siPLK1) for 20 hours and then exposed to hypoxia for 4 hours. (C) Left: Simple Western images of PLK1, HIF1α, and β-actin. Right: HIF1α and PLK1 protein expression normalized to β-actin. (D) Expression of hypoxia-related genes assessed by RT-qPCR and normalized to the housekeeping gene RPLP0 . Bar graphs represent expression relative to Normoxia_siNTC. (B-D) Data are shown as mean ± SEM of at least three independent biological replicates. HIF1α, hypoxia-inducible factor 1α; Hx, hypoxia; Nx, normoxia; Onv, onvansertib; PLK1, polo-like kinase 1; RT-qPCR, real-time quantitative polymerase chain reaction.

Article Snippet: Human KRAS -mutant colorectal cancer cell lines HCT116, SW620, LoVo, and DLD-1 (ATCC, Manassas, VA) were cultured in RPMI (Cat# 30-2001, ATCC) supplemented with 10% FBS (Cat#16000069, ThermoFisher Scientific, Waltham, MA) and 1× penicillin-streptomycin solution (Cat#30-2300, ATCC).

Techniques: Mutagenesis, RNA Sequencing, Expressing, Quantitative RT-PCR, Transfection, Control, Simple Western, Real-time Polymerase Chain Reaction