Johnson M D, Torri J A, Lippman M E, Dickson R B
Lombardi Cancer Research Center, Georgetown University Medical Center, Washington, DC 20007.
Cancer Res. 1993 Feb 15;53(4):873-7.
The aspartyl protease cathepsin D has been shown to be a marker of poor prognosis when found at high levels in primary breast tumors. It has been suggested that this is because the production of cathepsin D increases the invasive potential of the tumor cells, thus increasing the probability of metastasis. We have therefore conducted experiments to determine if secreted cathepsin D makes a significant contribution to the invasive phenotype of breast cancer cells in the Boyden chamber assay of invasion, which measures the ability of a cell to invade through an artificial basement membrane. Cathepsin D secretion and Boyden chamber invasiveness were measured in nine clones of the breast cancer cell line MCF-7, and no correlation was found between cathepsin secretion and invasive behavior. Invasion assays were also conducted in the presence of the aspartyl protease inhibitor pepstatin A, and no inhibition of the invasive behavior of cells was seen. Since low-pH environments are required for both the activation of pro-cathepsin D and the activity of the mature enzyme, assays were also conducted in the presence of chloroquine to neutralize the pH in the acidic compartments of the cells. This treatment did not inhibit invasiveness. Cathepsin D secretion by the breast cancer cell lines MDA-MB-231, MDA-MB-435, MDA-MB-435s, MDA-MB-468, SK-Br-3, and MCF-7-ADRr was also measured. Again, there was no correlation with invasion. In fact, cathepsin D levels were inversely correlated with aggressive behavior in vivo and in vitro in previously reported studies. These data suggest that cathepsin D secretion by tumor cells is not an important determinant of the invasiveness of the tumor cells per se. These data also reinforce the view that the poor prognosis in clinical breast cancer linked to high tumor levels of cathepsin D is probably due to high levels of cathepsin D in the stromal components of the tumor such as infiltrating inflammatory cells.
天冬氨酸蛋白酶组织蛋白酶D在原发性乳腺肿瘤中高表达时，已被证明是预后不良的标志物。有人认为，这是因为组织蛋白酶D的产生增加了肿瘤细胞的侵袭潜能，从而增加了转移的可能性。因此，我们进行了实验，以确定在测量细胞穿过人工基底膜侵袭能力的Boyden室侵袭试验中，分泌的组织蛋白酶D是否对乳腺癌细胞的侵袭表型有显著贡献。在乳腺癌细胞系MCF-7的9个克隆中测量了组织蛋白酶D的分泌和Boyden室侵袭性，未发现组织蛋白酶分泌与侵袭行为之间存在相关性。在天冬氨酸蛋白酶抑制剂胃蛋白酶抑制剂A存在的情况下也进行了侵袭试验，未观察到细胞侵袭行为受到抑制。由于组织蛋白酶D原的激活和成熟酶的活性都需要低pH环境，因此也在氯喹存在的情况下进行了试验，以中和细胞酸性区室中的pH。这种处理并未抑制侵袭性。还测量了乳腺癌细胞系MDA-MB-231、MDA-MB-435、MDA-MB-435s、MDA-MB-468、SK-Br-3和MCF-7-ADRr的组织蛋白酶D分泌。同样，与侵袭没有相关性。事实上，在先前报道的研究中，组织蛋白酶D水平与体内外的侵袭性行为呈负相关。这些数据表明，肿瘤细胞分泌组织蛋白酶D本身并不是肿瘤细胞侵袭性的重要决定因素。这些数据也强化了这样一种观点，即临床乳腺癌中与组织蛋白酶D高肿瘤水平相关的预后不良可能是由于肿瘤基质成分（如浸润性炎症细胞）中组织蛋白酶D水平较高所致。
