title: Detection of mutant BRAF alleles in the plasma of patients with metastatic melanoma.
abstract: Mutations in the BRAF oncogene at amino acid 600 have been reported in 40 to 70% of human metastatic melanoma tissues, and the critical role of BRAF in the biology of melanoma has been established. Sampling the blood compartment to detect the mutational status of a solid tumor represents a highly innovative advance in cancer medicine, and such an approach could have advantages over tissue-based techniques. We report the development of a fluorescence-based polymerase chain reaction (PCR) assay to detect mutant BRAF alleles in plasma. A mutant-specific PCR assay was optimized to specifically amplify the mutant BRAF allele without amplifying the wild-type allele. Experiments mixing DNA from a BRAF mutant melanoma cell line with wild-type human placental DNA in varying proportions were performed to determine the threshold of this assay and to compare it with routine DNA sequencing. The assay was then applied to tissue and plasma specimens from patients with metastatic melanoma. The assay detected 0.1 ng of mutant DNA mixed in 100 ng of wild-type DNA and was 500-fold more sensitive than DNA sequencing. The assay detected mutant BRAF alleles in plasma samples from 14 of 26 (54%) metastatic melanoma patients. These data demonstrate the feasibility of blood-based testing for BRAF mutations in metastatic melanoma patients.
authors: Department of Dermatology, New York University School of Medicine, 550 First Ave., H-100, New York, NY 10016, USA.
authors: Molly
authors: M
authors: Yancovitz
authors: Joanne
authors: J
authors: Yoon
authors: Maryann
authors: M
authors: Mikhail
authors: Weiming
authors: W
authors: Gai
authors: Richard L
authors: RL
authors: Shapiro
authors: Russell S
authors: RS
authors: Berman
authors: Anna C
authors: AC
authors: Pavlick
authors: Paul B
authors: PB
authors: Chapman
authors: Iman
authors: I
authors: Osman
authors: David
authors: D
authors: Polsky
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000483:Alleles; D001483:Base Sequence; D045744:Cell Line, Tumor; D004252:DNA Mutational Analysis; D004273:DNA, Neoplasm / Q000032:analysis; D005260:Female; D005820:Genetic Testing; D006801:Humans; D008297:Male; D008545:Melanoma / Q000097:blood* / Q000235:genetics*; D008875:Middle Aged; D008969:Molecular Sequence Data; D050505:Mutant Proteins / Q000097:blood*; D009362:Neoplasm Metastasis; D016133:Polymerase Chain Reaction; D048493:Proto-Oncogene Proteins B-raf / Q000097:blood* / Q000235:genetics*; D012680:Sensitivity and Specificity; D013696:Temperature
chemical_list: D004273:DNA, Neoplasm; D050505:Mutant Proteins; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf