Phytohemagglutinin-L (PHA-L) lectin surface binding of N-linked β1-6 carbohydrate and its relationship to activated mutant ras in human pancreatic cancer cell-lines

Roderich E. Schwarz, Donald C. Wojciechowicz, Patrick Y. Park, Philip B. Paty

Research output: Contribution to journalArticle

17 Citations (Scopus)

Abstract

Alterations of the N-linked carbohydrate core structure of cell surface glycoproteins (β1-6 branching) can be detected by phytohemagglutinin (PHA-L) lectin binding and has been linked to tumor progression and K-ras activation in colon cancer. The purpose of this study was to determine the prevalence of this carbohydrate alteration and its relationship to K-ras activation in pancreatic cancer. Nine human pancreatic cancer cell lines and 4 colon lines as controls were grown under standard tissue culture conditions. K-ras genome analysis was performed by polymerase chain reaction amplification and sequencing. The proportion of cellular p21-ras bound to GTP (ras-GTP level) was determined using immunoprecipitation of 32P-labeled cell lysates followed by thin layer chromatography and phosphoimaging analysis. Lectin blot analysis was performed on crude membrane preparations. Sensitivity to lectins was assessed with cell culture thymidine incorporation. Of 9 pancreatic cancer lines tested, 3 had wild type K-ras, 2 had heterozygous and 4 had homozygous mutations in codon 12 of K-ras. These genotypes correlated strongly with the level of ras-GTP measured. K-ras mutants had increased levels of ras-GTP compared to wild-type cell lines. PHA-L binding to cell membranes correlated positively with ras-GTP levels in 7 out of 9 cell lines. PHA-L toxicity was greatest in cells with positive PHA-L reactivity on Western blotting. A positive correlation between the presence of K-ras mutation, increased ras-GTP level, and increased cell surface β1-6 N-linked carbohydrate exists in pancreatic cancer cell lines.

Original languageEnglish (US)
Pages (from-to)285-291
Number of pages7
JournalCancer Letters
Volume107
Issue number2
DOIs
StatePublished - Oct 22 1996

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Phytohemagglutinins
Guanosine Triphosphate
Pancreatic Neoplasms
Lectins
Carbohydrates
Cell Line
Proto-Oncogene Proteins p21(ras)
Mutation
Membrane Glycoproteins
Thin Layer Chromatography
Immunoprecipitation
Codon
Colonic Neoplasms
Thymidine
Colon
Cell Culture Techniques
Western Blotting
Genotype
Cell Membrane
Genome

Keywords

  • β1-6 N-linked carbohydrate
  • K-ras oncogen
  • Lectin
  • Pancreatic cancer
  • Phytohemagglutinin

ASJC Scopus subject areas

  • Cancer Research
  • Molecular Biology
  • Oncology

Cite this

Phytohemagglutinin-L (PHA-L) lectin surface binding of N-linked β1-6 carbohydrate and its relationship to activated mutant ras in human pancreatic cancer cell-lines. / Schwarz, Roderich E.; Wojciechowicz, Donald C.; Park, Patrick Y.; Paty, Philip B.

In: Cancer Letters, Vol. 107, No. 2, 22.10.1996, p. 285-291.

Research output: Contribution to journalArticle

Schwarz, Roderich E. ; Wojciechowicz, Donald C. ; Park, Patrick Y. ; Paty, Philip B. / Phytohemagglutinin-L (PHA-L) lectin surface binding of N-linked β1-6 carbohydrate and its relationship to activated mutant ras in human pancreatic cancer cell-lines. In: Cancer Letters. 1996 ; Vol. 107, No. 2. pp. 285-291.
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