title: SP174, NRAS Q61R Mutant-Specific Antibody, Cross-Reacts With KRAS Q61R Mutant Protein in Colorectal Carcinoma.
abstract: BACKGROUND
  - NRAS is a member of the RAS family oncoproteins implicated in cancer. Gain-of-function NRAS mutations were reported in a subset of colorectal cancers. These mutations occur at codons 12, 13, and 61 and are detected by molecular genetic testing. Recently, an antibody (clone SP174) became available to immunohistochemically pinpoint NRAS Q61R mutant protein. In malignant melanoma, NRAS Q61R mutant-specific immunohistochemistry was shown to be a valuable supplement to traditional genetic testing.
  
  
  OBJECTIVE
  - To evaluate the significance of NRAS Q61R mutant-specific immunohistochemistry in a cohort of colorectal carcinomas.
  
  
  METHODS
  - A total of 1185 colorectal carcinomas were immunohistochemically evaluated with SP174 antibody. NRAS Q61R mutant-specific immunohistochemistry was validated by molecular genetic testing including Sanger sequencing, quantitative polymerase chain reaction (qPCR), and next-generation sequencing.
  
  
  RESULTS
  - Twelve tumors showed strong SP174 immunoreactivity. Sanger sequencing detected an identical c.182A>G substitution, causing NRAS Q61R mutation at the protein level, only in 8 SP174-positive cases. These results were confirmed by qPCR study. Subsequently, NRAS wild-type tumors with strong SP174 staining were evaluated by next-generation sequencing and revealed KRAS c.182A>G substitutions predicted to cause KRAS Q61R mutation. Review of colorectal carcinomas with known KRAS and NRAS genotype revealed that none of 62 wild-type tumors or 47 mutants other than Q61R were SP174 positive.
  
  
  CONCLUSIONS
  - SP174 immunohistochemistry allows sensitive detection of NRAS and KRAS Q61R mutants. However, molecular genetic testing is necessary to determine specifically which RAS gene is mutated.
authors.forename: Jerzy
authors.initials: J
authors.lastname: Lasota
authors.forename: Artur
authors.initials: A
authors.lastname: Kowalik
authors.forename: Anna
authors.initials: A
authors.lastname: Felisiak-Golabek
authors.forename: Shingo
authors.initials: S
authors.lastname: Inaguma
authors.forename: Zeng-Feng
authors.initials: ZF
authors.lastname: Wang
authors.forename: Liliana
authors.initials: L
authors.lastname: Pięciak
authors.forename: Sebastian
authors.initials: S
authors.lastname: Zięba
authors.forename: Rafał
authors.initials: R
authors.lastname: Pęksa
authors.forename: Janusz
authors.initials: J
authors.lastname: Kopczynski
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Okoń
authors.forename: Piotr
authors.initials: P
authors.lastname: Waloszczyk
authors.forename: Stanislaw
authors.initials: S
authors.lastname: Gozdz
authors.forename: Wojciech
authors.initials: W
authors.lastname: Biernat
authors.affiliation: From the Laboratory of Pathology, National Cancer Institute, Bethesda, Maryland (Drs Lasota, Felisiak-Golabek, Inaguma, Wang, and Miettinen); the Departments of Molecular Diagnostics (Dr Kowalik, Ms Pięciak, and Mr Zięba), Surgical Pathology (Dr Kopczynski), and Clinical Oncology (Dr Gozdz), Holycross Cancer Center, Kielce, Poland; the Department of Pathology, Aichi Medical University School of Medicine, Nagakute, Japan (Dr Inaguma); the Department of Pathomorphology, Medical University of Gdansk, Poland (Drs Pęksa and Biernat); the Department of Pathomorphology, Jagiellonian University, Krakow, Poland (Dr Okoń); ZDUNOMED/Histopathology, Szczecin, Poland (Dr Waloszczyk); and Faculty of Health Sciences, The Jan Kochanowski University, Kielce, Poland (Dr Gozdz). Drs Lasota and Kowalik contributed equally to this work.
authors.forename: Markku
authors.initials: M
authors.lastname: Miettinen
mesh_terms: D000906:Antibodies / Q000276:immunology*; D001483:Base Sequence; D015331:Cohort Studies; D015179:Colorectal Neoplasms / Q000175:diagnosis / Q000235:genetics / Q000276:immunology*; D003429:Cross Reactions / Q000276:immunology*; D020558:GTP Phosphohydrolases / Q000235:genetics / Q000276:immunology*; D005820:Genetic Testing / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D008565:Membrane Proteins / Q000235:genetics / Q000276:immunology*; D050505:Mutant Proteins / Q000276:immunology*; D020125:Mutation, Missense / Q000276:immunology*; D016133:Polymerase Chain Reaction; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics / Q000276:immunology; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D017422:Sequence Analysis, DNA
chemical_list: D000906:Antibodies; C117307:KRAS protein, human; D008565:Membrane Proteins; D050505:Mutant Proteins; D020558:GTP Phosphohydrolases; C579846:NRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)