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MET tyrosine kinase domain modeling location of the F1200 residue

Demographics of the METex14-mutated and EGFR-mutated cohorts.

Purpose:Although patients with advanced-stage non–small cell lung cancers (NSCLC) harboring MET exon 14 skipping mutations (METex14) often benefit from MET tyrosine kinase inhibitor (TKI) treatment, clinical benefit is limited by primary and acquired drug resistance. The molecular basis for this resistance remains incompletely understood.Experimental Design:Targeted sequencing analysis was performed on cell-free circulating tumor DNA obtained from 289 patients with advanced-stage METex14-mutated NSCLC.Results:Prominent co-occurring RAS–MAPK pathway gene alterations (e.g., in KRAS, NF1) were detected in NSCLCs with METex14 skipping alterations as compared with EGFR-mutated NSCLCs. There was an association between decreased MET TKI treatment response and RAS–MAPK pathway co-occurring alterations. In a preclinical model expressing a canonical METex14 mutation, KRAS overexpression or NF1 downregulation hyperactivated MAPK signaling to promote MET TKI resistance. This resistance was overcome by cotreatment with crizotinib and the MEK inhibitor trametinib.Conclusions:Our study provides a genomic landscape of co-occurring alterations in advanced-stage METex14-mutated NSCLC and suggests a potential combination therapy strategy targeting MAPK pathway signaling to enhance clinical outcomes.

The MET F1200 residue is conserved across multiple tyrosine kinases

Genes included in NGS Panels

OBJECTIVES: The purpose of our study was to establish breast epithelial cell cultures and cell lines from peoples of African origin (blacks). It is presumed that the biology of breast cancer in women of African origin has unique aspects that can be explored using cultured breast epithelial cells. STUDY DESIGN: Biopsy specimens were obtained from black women undergoing radical or modified mastectomies. Normal cell cultures were established using tissue from reduction mammaplasties or the milk of lactating mothers. The tissue specimens were lacerated, digested with collagenase solution, and plated on tissue culture plates. To extend the life of the epithelial cells in culture, they are transformed with SV40 virus. RESULTS: We have maintained breast tumor cells in culture from a 27-year-old black woman for more than 1 month. CONCLUSION: Despite the difficulty of establishing epithelial cell cultures, we have maintained breast tumor cells from blacks in culture for an extended period to allow characterization. (Am J Obstet Gynecol 1997;176:S240-5.)

MS-based proteomics has been the method of choice for biomarker discovery in the field of traumatic brain injury (TBI). Due to its high sensitivity and specificity, MS is now being explored for biomarker quantitative validation in tissue and biofluids. In this study, we demonstrate the use of MS in both qualitative protein identification and targeted detection of acute TBI biomarkers released from degenerating cultured rat cortical mixed neuronal cells, mimicking intracellular fluid in the central nervous system after TBI. Calpain activation was induced by cell treatment with maitotoxin (MTX), a known calcium channel opener. Separate plates of mixed neuronal-glial culture were subjected to excitotoxin N-methyl-D-aspartate (NMDA) and apoptotic inducer staurosporine. Acute TBI biomarkers, GFAP and UCH-L1, were first detected and assessed in the culture media by Western blot. The cell-conditioned media were then trypsinized and subjected to bottom up proteomic analysis. GFAP was readily detected by data-dependent scanning but not UCH-L1. As a proof-of-principle study, rat glia-enriched cell cultures treated with MTX were used to investigate the time-dependent release of GFAP breakdown product by Western blot and for isotope dilution MS absolute quantitation method development. Absolute quantitation of the GFAP release was conducted using the three cortical mixed neuronal cell cultures treated with different agents. Other differentially expressed proteins identified in the glial-enriched and cortical mixed neuronal cell culture models were further analyzed by bioinformatic tools. In summary, this study demonstrates the use of MS in both protein identification and targeted quantitation of acute TBI biomarkers and is the preliminary step toward development of TBI biomarker validation by targeted MS.