Detection of nonmelanoma skin cancer micrometastases in lymph nodes by using reverse transcriptase-polymerase chain reaction for keratin 19 mRNA.

Abstract

A new sensitive method for the detection of skin cancer micrometastases in lymph nodes is based on amplification of keratin 19 (K19) mRNA by reverse transcriptase-polymerase chain reaction (RT-PCR). To compare results of RT-PCR with those of histological examination in terms of the detection rate of skin cancer micrometastases. Twenty-six lymph nodes obtained from 13 patients with squamous cell carcinoma (SCC), eccrine porocarcinoma and Paget's disease were investigated by histological examination (haematoxylin and eosin sections) and RT-PCR. RT-PCR was performed on extracted RNA by using K19 primer pairs. RT-PCR products were visualized by ethidium bromide staining and confirmed by non-radioactive hybridization with K19-specific probes. All of 10 histologically positive lymph nodes yielded the expected 460-bp band. Of the 16 histologically negative lymph nodes, one (6%) was found by RT-PCR to express K19 mRNA, indicating the presence of micrometastases which could not be detected by histological examination. A serial dilution study using RNA extracted from SCC cells mixed with RNA extracted from normal lymph node cells showed a detection sensitivity of K19 RT-PCR of 10-5 micro g cancer cell RNA in 1 micro g lymph node RNA. Nested RT-PCR showed a detection sensitivity of one tumour cell in 106 lymphocytes. These results demonstrate the usefulness of K19 RT-PCR for the detection of skin cancer micrometastases in lymph nodes.