Methylation assay for the diagnosis of lung cancer on bronchial aspirates: a cohort study

Clin Cancer Res. 2005 Nov 1;11(21):7728-34. doi: 10.1158/1078-0432.CCR-05-0999.


Purpose: Recent studies have detected aberrant promoter methylation of adenomatous polyposis coli promoter 1 A (APC), cyclin-dependent kinase inhibitor-2A (p16(INK4a)), retinoic acid receptor beta2, and RAS association domain family protein 1 (RASSF1A) in bronchial aspirates and suggested their use as biomarkers for lung cancer diagnostics. The purpose of this study was to validate these candidate marker genes in a retrospective cohort study.

Experimental design: Bronchial aspirates collected from a cohort comprising 247 patients with suspected lung cancer were investigated retrospectively regarding aberrant promoter methylation using a quantitative methylation-specific real-time PCR (QMSP).

Results: Eighty-nine patients were diagnosed with primary lung cancer, 102 had benign lung disease, and 56 showed miscellaneous other conditions. A panel consisting of APC, p16(INK4a), and RASSF1A emerged as useful combination. This panel detected aberrant methylation in bronchial aspirates of 22 of 35 (63%) and 21 of 44 (44%) centrally and peripherally located primary lung cancers, respectively. Bronchial aspirates also showed aberrant methylation in 5 of 7 (71%) patients with a recurrent lung cancer and in 8 of 30 (27%) cases without tumor recurrence. In contrast, only 1 of 102 patients with benign lung disease displayed a (false) positive test result. Rarely, aberrant methylation was found in patients with other malignancies (3 of 16). The QMSP assay correctly confirmed lung cancer in 8 of 12 (67%) cases with an ambiguous cytology. Moreover, it disclosed 9 of 26 (35%) of peripheral tumors lacking simultaneous cytologic or histologic diagnosis of malignancy.

Conclusions: Our findings suggest that the QMSP assay could be applied as a reflex test in cases of suspected lung cancer that defy a definite diagnosis by conventional methods. Thus, the assay could be a useful diagnostic adjunct especially regarding peripheral tumors.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adenomatous Polyposis Coli Protein / metabolism
  • Adult
  • Aged
  • Aged, 80 and over
  • Biopsy
  • Bronchi / metabolism*
  • Bronchoscopy
  • Case-Control Studies
  • Cell Line, Tumor
  • Cohort Studies
  • Cyclin-Dependent Kinase Inhibitor p16 / metabolism
  • DNA Methylation*
  • DNA Primers / chemistry
  • False Positive Reactions
  • Female
  • Humans
  • Lung / pathology
  • Lung Neoplasms / diagnosis*
  • Lung Neoplasms / genetics*
  • Lung Neoplasms / metabolism
  • Lung Neoplasms / pathology
  • Male
  • Middle Aged
  • Polymerase Chain Reaction
  • Receptors, Retinoic Acid / metabolism
  • Retrospective Studies
  • Reverse Transcriptase Polymerase Chain Reaction
  • Sulfites / pharmacology
  • Temperature
  • Tumor Suppressor Proteins / genetics


  • Adenomatous Polyposis Coli Protein
  • Cyclin-Dependent Kinase Inhibitor p16
  • DNA Primers
  • RASSF1 protein, human
  • Receptors, Retinoic Acid
  • Sulfites
  • Tumor Suppressor Proteins
  • retinoic acid receptor beta