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Genetic studies have identified mutations in key regulators of the Wnt/β-catenin pathway in a variety of cancers, most frequently in colon cancers. However, whether the pathway is activated in clinical cancer samples is not easily determined, and therefore it is useful to find markers that could be surrogates to show activation of the Wnt/β-catenin pathway. Gene expression profiles were analyzed in SW620, a colon cancer cell line in which β-catenin levels are stabilized as a consequence of truncated adenomatous polyposis coli and were compared with profiles of the same cells transfected with antisense oligodeoxynucleotides. Treatment of cells with β-catenin antisense oligodeoxynucleotides resulted in a decrease in the levels of axin2 and human naked cuticle ( hnkd ) mRNAs. Interestingly, the proteins encoded by both of these mRNAs are known inhibitors of the β-catenin pathway. In 30 human cell lines derived from different origins, axin2 and hnkd were expressed only in human colon cancer cell lines that are known to have activating mutations in the Wnt/β-catenin pathway. Further, levels of both axin2 and hnkd mRNA were also found to be elevated in about 65% of laser microdissected cells from human colon tumors compared with laser microdissected cells of normal morphology from the same patient samples. The increased expression of axin2 and hnkd correlated with truncations in adenomatous polyposis coli in the same patient samples. These results reveal that it is possible to detect activation of a carcinogenic pathway in human cancer samples with specific markers.

The product of the HER2 protooncogene, p185HER2, represents an attractive target for cancer immunotherapies. We have prepared anti-p185HER2 immunoliposomes in which Fab' fragments of a humanized anti-p185HER2 monoclonal antibody with antiproliferative properties (rhuMAb-HER2) were conjugated to either conventional or sterically stabilized liposomes. These immunoliposomes bind specifically to p185HER2-overexpressing breast cancer cells (SK-BR-3 and BT-474). High-affinity binding of anti-p185HER2 immunoliposomes is comparable to that of free rhuMAbHER2-Fab' or the intact antibody. Empty immunoliposomes inhibit the culture growth of p185HER2-overexpressing breast cancer cells, and this antiproliferative effect is superior to that of free rhuMAbHER2-Fab', indicating that liposomal anchoring of these anti-p185HER2 Fab' fragments enhances their biological activity. Efficient internalization of anti-p185HER2 immunoliposomes, demonstrated by light and electron microscopy, occurs by receptor-mediated endocytosis via the coated pit pathway and also possibly by membrane fusion. Doxorubicin-loaded anti-p185HER2 immunoliposomes are markedly and specifically cytotoxic against p185HER2-overexpressing tumor cells in vitro. Anti-p185HER2 immunoliposomes administered in vivo in Scid mice bearing human breast tumor (BT-474) xenografts can deliver doxorubicin to tumors. These results indicate that anti-p185HER2 immunoliposomes are a promising therapeutic vehicle for the treatment of p185HER2-overexpressing human cancers.