Methods of diagnosing, classifying and treating endometrial cancer and precancer
a technology applied in the field of precancer and endometrial cancer diagnostic and treatment methods, can solve problems such as poor survival
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example 1
Detection of Activating FGFR2 Mutations in Endometrial Cancer
[0132]Our findings show that activation and overexpression of FGFR2 plays a role in endometrial tumorigenesis. Exon 8 is three nucleotides longer than exon 9, hence the FGFR2b isoform is one codon longer than the FGFR2c isoform. Specificity of signaling is also provided by tissue specific expression of receptors, ligands and heparin sulphate proteoglycans (Allen et al., 2001; Fiore, 2001). Due to the differences in length of the FGFR2 “b” and “c” isoforms, all mutations will be numbered relative to the epithelially expressed FGFR2b isoform (SEQ ID NO:2; NP—075259.2). For those occurring downstream of exon 8 we provide herein the equivalent mutation numbered relative to the FGFR2c isoform (SEQ ID NO:3; NP—000132.1) in brackets and in Table 2. The N550K (N549K) variant identified in two of the endometrial cell lines was likely to result in receptor activation as identical or similar germline missense changes had been reporte...
example 2
Treatment of Endometrial Cancer by Inhibition of FGFR2
Materials and Methods
Sequencing Analysis
[0145]Mutation analysis was performed as previously described (8). PCR primer sequences were M13 tailed and sequencing performed in two directions. Primer sequences are available by request from the author.
Cell Culture and Reagents
[0146]The human endometrial MFE296 cell line was purchased from the European Collection of Cell Cultures (Salisbury, Wiltshire, UK). The human endometrial cell lines AN3CA, HEC1A, Ishikawa, RL952, and KLE were provided by Dr. Paul Goodfellow (Washington University, St. Louis, Mo.). MFE296 cells were grown in MEM supplemented with 10% fetal bovine serum (FBS), 2 mM L-glutamine, and penicillin-streptomycin. AN3CA cells were cultured in DMEM supplemented with 10% FBS, non-essential amino acids, 2 mM L-glutamine, and penicillin-streptomycin. HEC1A cells were cultured in 50% DMEM and 50% RPMI 1640, supplemented with 10% FBS and penicillin-streptomycin. Ishikawa and RL9...
example 3
Activating-FGFR2 Mutations Promote Endometrial Cell Migration
Materials and Methods
[0171]To determine the ability of FGFR2 mutations to alter the phenotype of primary endometrial epithelial cells, HPV / TERT immortalized primary human endometrial epithelial cells were transduced with wild type FGFR2b or mutant FGFR2b (N550K) and FGF-stimulated migration evaluated.
[0172]Briefly, 3×105 cells were seeded on top of 8 μm pore size polycarbonate membranes (Cell Biolabs, Inc) in full growth media containing 1 nM FGF10. 24 hours later, the media was aspirated from inside the insert and non-migratory cells were removed from the top of the insert using cotton-tipped swabs. Migratory cells (those that had passed through the membrane) were stained and the membranes were photographed (5 random fields per insert). The stained cells on the inserts were then solubilized and quantified by measuring absorbance at 570 nm.
Results
[0173]Each FGFR2 construct was assessed in quadruplicate and the results are ...
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