DETECTION OF CIRCULATING CANCER-CELLS BY NESTED REVERSE TRANSCRIPTION-POLYMERASE CHAIN-REACTION OF CYTOKERATIN-19 (K19)-POSSIBLE CLINICAL-SIGNIFICANCE IN ADVANCED GASTRIC-CANCER

Intermediate filament cytokeratin-19 (K19) protein is expressed in nor mal and malignant gastrointestinal epithelial cells, but not in periph eral blood (PB). Small amount of circulating gastric cancer cells can be detected by a sensitive nested reverse transcription-polymerase cha in reaction (RT-PCR) with primers specific for K19 mRNA. Thirty-four P B samples obtained from patients with inoperable/metastatic gastric ca ncer were examined. The mononuclear cell (MNC) fraction was collected by Ficoll centrifugation, and followed by total RNA extraction by acid guanidinium thiocyanate-phenol-chloroform method. RNA fr om 8 gastric cancer cell lines and the mononuclear cells of 33 healthy adults were used as positive and negative controls, respectively. DNA fragment of 774 bp amplified by the internal primers was found to be a highly rel iable marker for K19 mRNA expression The sensitivity of detection was between 1 and 10 cells/10(6) normal MNCs. The K19 transcripts were det ected in 20.6% (7/34; 8-37%, 95% C.l) of PB samples. None of the other pertinent clinicopathological features, including the disease extent and the histopathologic types of the tumors, were related to the expre ssion of K19 in PB. All 34 patients had been a treated by systemic che motherapy. Among the 17 non-responders to chemotherapy, the survival o f the 4 patients with detectable K19 was significantly shorter than th at of 13 patients without detectable K19 in their circulating blood (p =0.014). However, the survival impact of K19 was less significant in t he other 17 patients whose tumors had responded to systemic chemothera py. Of the whole group of patients, the median survival of the 7 and 2 7 patients with and without detectable K19 in their circulating blood was 1 and 3.5 months, respectively (p=0.368). We concluded that detect ing circulating cancer cells by K19 nested RT-PCR is associated with p oor prognosis of gastric cancer, particularly in those patients who ar e not responsive to systemic chemotherapy.