Up to 30% of patients with stage II (pN0)
colon cancer develop recurrences, suggesting that the presence of lymph node (LN)
metastases escaped detection at histopathologic staging. A simple way to overcome this limitation and to improve staging accuracy is to use reverse transcription-polymerase chain reaction (RT-PCR) to examine a larger fraction or an entire specimen. The
Guanylyl cyclase C (GCC) gene is uniquely expressed in apical cells of the gastrointestinal tract. Its expression in
colon cancer cells and
metastases is conserved. Therefore, detection of GCC
mRNA in LNs has been shown to be indicative of the presence of
colon cancer metastases. As the current processing of LNs involves
formalin fixation and
paraffin embedding, we developed a method for extracting
RNA from
formalin-fixed
paraffin-embedded LN specimens and detecting GCC
mRNA by quantitative RT-PCR. The assay has a dynamic range of 5 logs, an average amplification efficiency of 98.4% (95% confidence interval, 96.6-100.3), a reaction linearity of 0.998 (95% confidence interval, 0.997-0.999), and also intraplate and interplate CVs of <1% and <5%, respectively. The test specificity was 98% with LNs collected from patients affected by conditions other than
colon cancer (n=380). Sensitivity was 97% for patients with stage III
colon cancer (n=34), whereas 35% of patients with stages I and II disease (n=51) had at least 1 GCC
mRNA-positive LN. The high specificity of GCC
mRNA suggests that routine utilization of the quantitative RT-PCR test has the potential to improve the detection of
colon cancer metastases in LNs.