Background: The detection of micrometastatic cells in lymph nodes and bone marrow independendly predicts survival and metastatic relapse in patients with operable esophageal carcinoma. So far, little is known about the biology of these cells. A recently developed PCR-method allows the comprehensive genomic analysis of single micrometastatic tumor cells. We therefore investigated the genomic alterations of disseminated tumor cells in esopahgeal carcinoma patients.
Methods: Epithelial tumor cells from patients with operable esophageal carcinoma (pT1-3, pN10-1, pM0) were enriched from bone marrow and lymph nodes by density-gradient centrifugation, detected by immunocytochemistry (anti-cytokeratin antibody for bone marrow and anti-EpCAM antibody for lymph nodes) and subsequently isolated by micromanipulation. Areas of the primary tumor were microdissected. We globally amplified the genomic DNA of disseminated tumor cells and from the primary tumor using the Mse-adapter PCR method and subsequently performed comparative genomic hybridization (CGH) for the genome-wide screening of DNA-gains and -losses. Point mutations in the p53 tumor suppressor gene were investigated by single-strand conformation polymorphism (SSCP).
Results: Here we demonstrate the presence of chromosmal aberrations in isolated disseminated tumor cells of patients with esophageal carcinoma. The observed chromosomal alterations detected by CGH are typical for tumors of this entity (deletions at 5q, 13q, 17p and amplifications at 8p, 17q and 19). Compared to the primary tumors we observed many shared aberrations, but also clonal variation. Furthermore, SSCP revealed p53 point mutations in the disseminated tumor cells, which are considered as a key event in esophageal carcinogenesis.
Conclusion: Our study provides a description of genomic alterations in minimal residual tumor cells detected in bone marrow and lymph nodes of patients with operable esophageal carcinoma. The genomic data confirms the tumor cell identity of cells detected by cytokeratin antibodies in bone marrow and EpCAM antibodies in lymph nodes. The altered chromosomal regions of the micrometastatic tumor cells may help to identify genes that regulate the growth of metastases. Genomic aberrations detected in disseminated tumor cells may become new prognostic markers and may help to discover new therapeutic targets for patients with esophageal cancer.
