The reliability of immunohistochemistry as a prescreening method for the diagnosis of hereditary nonpolyposis colorectal cancer (HNPCC)--results of an international collaborative study.

Hereditary nonpolyposis colorectal cancer syndrome (HNPCC) is an autosomal dominant condition accounting for 2-5% of all colorectal carcinomas as well as a small subset of endometrial, upper urinary tract and other gastrointestinal cancers. An assay to detect the underlying defect in HNPCC, inactivation of a DNA mismatch repair enzyme, would be useful in identifying HNPCC probands. Monoclonal antibodies against hMLH1 and hMSH2, two DNA mismatch repair proteins which account for most HNPCC cancers, are commercially available. This study sought to investigate the potential utility of these antibodies in determining the expression status of these proteins in paraffin-embedded formalin-fixed tissue and to identify key technical protocol components associated with successful staining. A set of 20 colorectal carcinoma cases of known hMLH1 and hMSH2 mutation and expression status underwent immunoperoxidase staining at multiple institutions, each of which used their own technical protocol. Staining for hMSH2 was successful in most laboratories while staining for hMLH1 proved problematic in multiple labs. However, a significant minority of laboratories demonstrated excellent results including high discriminatory power with both monoclonal antibodies. These laboratories appropriately identified hMLH1 or hMSH2 inactivation with high sensitivity and specificity. The key protocol point associated with successful staining was an antigen retrieval step involving heat treatment and either EDTA or citrate buffer. This study demonstrates the potential utility of immunohistochemistry in detecting HNPCC probands and identifies key technical components for successful staining.

In vitro invasiveness of human epithelioid-sarcoma cell lines: association with cell motility and inverse correlation with the expression of tissue inhibitor of metalloproteinases.

Epithelioid sarcoma (ES) is a very aggressive soft-tissue tumor in vivo, but no experimental data on its invasive and metastatic behavior have been reported. In the present study, 3 different clonal sub-populations (GRU-1A, GRU-1B and GRU-1C), derived from the same human ES cell line, GRU-1, were investigated for in vitro invasiveness in relation to migration, adhesion and the expression of different invasion- and metastasis-related genes. Tumor spheroids of GRU-1A were markedly more invasive in the chick-heart invasion assay (CHIA) than spheroids of GRU-1B and GRU-1C. These results were paralleled by a significantly higher cell motility of GRU-1A than GRU-1B and GRU-1C (p < 0.05) on distinct substrates, suggesting that the observed differences in invasion result at least in part from differences in motility. When invasion was assayed with suspended tumor cells in the Matrigel assay, differences between the 3 cell lines were much more pronounced than in the CHIA, where cell-cell contacts are established. These results indicate that interclonal differences in ES invasion result mainly from differences in motility, but also partly depend on differences in cell-cell adhesion. On the molecular level, low invasive potential was associated with over-expression of distinct tissue inhibitor of metalloproteinases (TIMPs) relative to matrix metalloproteinase-2 and -9. However, no association was found between invasion and the expression of CD44 splicing variants or nm23 isoforms. Our results suggest that differences in invasion between GRU-1A, GRU-1B and GRU-1C are caused mainly by interclonal differences in migration, and might result from differences in the expression of distinct TIMPs.