The cysteine protease cathepsin B has been implicated in malignant progression of human colorectal carcinomas and gliomas and murine B16 melanoma. There are striking increases in expression of cathepsin B in a wide variety of human tumors (bladder, breast, colon, gastric, lung, prostate and thyroid carcinomas and glioma). This expression is not homogeneous, but occurs primarily at the invading margins. In tumor cells in this region, there are increases in cathepsin B transcripts, staining and activity. They have evidence that suggests that transcription of the cathepsin B gene may be induced in some cell types. Cathepsin B transcripts are heterogeneous due prinarily to alternative splicing in the 5'-untranslated region. Thus, there is the potential for regulation of cathepsin B expression at both the transcriptional and post-transcriptional levels. In normal cells, cathepsin B is localized in perinuclear lysosomes. In tumors/tumor cells, cathepsin B is: 1) concentrated in focal adhesions and invadopodia and at the basal membrane, 2) associated with the outer basal surface, 3) secreted, and/or 4) cytoplasmic. The experiments proposed in this application are designed to study a) the mechanisms responsible for altered localization of cathepsin B in malignant cells, b) the mechanisms responsible for increased expression of cathepsin B in malignant cells, and c) whether the observed changes in localization and expression of cathepsin B in human tumors can be functionally linked to tumor invasion and other phenotypic properties associated with malignant progression. In the first two aims of this proposal, they will determine a) whether localized concentrations of cathepsin B result in secretion and surface binding, b) the mechanisms responsible for surface binding, and c) whether these are functionally related to the malignant phenotype.
In aim three, they will begin to characterize the transcriptional and post-transcriptional mechanisms that regulate expression of the cathepsin B gene.
In aim four, they will a) determine what transcript species are present in a series of human tumors and correlate this with levels and patterns of expression of cathepsin B in specimens of those same tumors. Then, using this data, they will determine whether transfection with engineered cathepsin B transcript variants identified in some human tumors can reproduce the observed levels and patterns of expression. The eventual goal is to use the information obtained in these four aims to manipulate the expression and localization of cathepsin B and determine whether this has a direct effect on the malignant phenotype.
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