Molecular staging of lung cancer: Real-time polymerase chain reaction estimation of lymph node micrometastatic tumor cell burden in stage I non-small cell lung cancer - Preliminary results of cancer and leukemia group B trial 9761

Objective: The 5-year survival for patients with surgically resected stage I non-small cell lung cancer is only 60% to 70%, probably because of undetected systemic occult micrometastases. Detection of occult micrometastases in lymph nodes by reverse-transcriptase polymerase chain reaction for carcinoembryonic antigen messenger RNA in non-small cell lung cancer has not been reported. Detection of occult micrometastases by standard reverse-transcriptase polymerase chain reaction provides only yes or no answers about their presence, whereas quantitative real-time reverse-transcriptase polymerase chain reaction permits reproducible quantitation of target molecules. This study evaluated the ability of quantitative reverse-transcriptase polymerase chain reaction to quantitate lymph node occult metastases with carcinoembryonic antigen messenger RNA as a tumor marker. Methods: Standard reverse-transcriptase polymerase chain reaction and quantitative reverse-transcriptase polymerase chain reaction for carcinoembryonic antigen messenger RNA were performed on 232 lymph nodes from 53 patients with stage I disease (node negative according to histologic examination). Quantitative reverse-transcriptase polymerase chain reaction determined carcinoembryonic antigen messenger RNA quantity by detecting fluorescence increase at a threshold polymerase chain reaction cycle. Threshold polymerase chain reaction cycle values were correlated with standard curves created from serially diluted carcinoembryonic antigen-positive HTB-174 tumor cells to estimate the number of micrometastatic tumor cells in a lymph node. Results: Detection rates of occult metastases were similar for standard reverse-transcriptase polymerase chain reaction and quantitative reverse-transcriptase polymerase chain reaction at 38 of 232 (16.4 %) and 59 of 232 (25.4 %), respectively. Upstaging rates among 53 cases of stage I non-small cell lung cancer were also similar for standard reverse-transcriptase polymerase chain reaction and quantitative reverse-transcriptase polymerase chain reaction at 23 of 53 (43.4 %) and 30 of 53 (56.6%), respectively. Comparison of positive lymph node stations according to quantitative reverse-transcriptase polymerase chain reaction (threshold polymerase chain reaction cycle <45) with HTB-174 tumor cell standard curves yielded estimates of metastatic tumor cell burden of 1.07 × 10³ to 3.24 × 10⁵ cells per lymph node station (median 7190 tumor cells per lymph node station). Conclusions: Standard and quantitative real-time reverse-transcriptase polymerase chain reaction for carcinoembryonic antigen detected occult metastases in patients with stage I non-small cell lung cancer at similar rates; both upstaged about 50% of cases. Quantitative reverse-transcriptase polymerase chain reaction allows estimation of the number of metastatic cells per lymph node, however, which potentially allows greater precision in predicting recurrence risk.