The current project is to study a novel matrix-degrading proteinase in breast cancer invasion and metastasis particularly in hormone dependent disease. We recently identified an apparently new matrix-degrading proteinase, of ubiquitous expression, specially in hormone-dependent breast cancer cells. This enzyme appears as a major gelatinase with an apparent molecular mass of 8OkDa. In addition to Ca2+, the enzyme activity can also be supported by Mn2+ and Mg2+. The enzyme can degrade type IV collagen. Unlike MMP-2, MMP-9 and other known MMPs, the 8OkDA proteinase is not activated by p-aminophenyl mercuric acetate and is not inhibited by TIMP2. The 8OkDa proteinase can also be detected in surgical human breast cancer samples. So far, the known inhibitors of the enzyme are EDTA, dithiothreitol, and leuptin. We hypothesize that the expression of 8OkDa proteinase contributes to breast cancer metastasis particularly in hormone-dependent disease. Currently we have purified and described the 8okDa enzyme. We will further establish its structure, define its regulation, range of expression, and study its relevance in breast cancer invasion and metastasis.
Aim 1 : We will obtain the full purification and internal sequence of the 8OkDA proteinase to confirm its novelty.
Aim 2. Monoclonal antibodies against the 8OkDA proteinase will be raised to facilitate its detection, inhibition and cloning.
Aim 3. To investigate whether the proteinase is regulated during malignant progression and during exposure to estrogenic and progestational hormones, we will also perform immunohistochemical assessment of the expression of the 8OkDA proteinase in clinical breast cancer specimens with different hormone receptor status and metastatic potentials.
Aim 4. To clone the cDNA.
Aim 5. The relevance of the 8OkDA proteinase to human breast cancer invasion and metastasis will be investigated by introducing its cDNA into lacZ expressing both hormone dependent and hormone independent human breast cancer cells. If the 8OkDA proteinase could be proved to play a role in the regulation of breast cancer metastasis, this enzyme can be used as diagnostic marker for evaluation of breast cancer malignant progression, and inhibition of this proteinase could open up new therapeutic approaches for the control of breast cancer.
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