An intracellular form of cathepsin B contributes to invasiveness in cancer.
An intracellular form of cathepsin B contributes to invasiveness in cancer.
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发表时间：
2001-04
期刊：
影响因子：
11.2
通讯作者：
Anna M. Szpaderska;Allen Frankfater
中科院分区：
文献类型：
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作者：
Anna M. Szpaderska;Allen Frankfater
组织蛋白酶B是一种溶酶体半胱氨酸蛋白酶，其表达和运输在癌症中经常改变，并且质膜和分泌形式被认为有助于恶性肿瘤的侵袭性和转移性。我们已经操纵了组织蛋白酶B在几种肿瘤细胞系中的表达，并测量了它们通过重建的细胞外（Matrigel）基质侵入的能力。人组织蛋白酶B在转移性差的B16 F1鼠黑色素瘤变体中的瞬时表达产生了组织蛋白酶B活性的3-5倍增加和侵袭性的相当增加。高转移性人黑色素瘤A375 M和前列腺癌PC 3 M细胞系的稳定反义组织蛋白酶B表达克隆产生的组织蛋白酶B比对照细胞少40-50%，并且侵袭性相应降低。相比之下，操纵组织蛋白酶B水平对细胞穿过未涂覆膜的迁移没有影响。阴离子型组织蛋白酶B抑制剂（L-3-trans-propylcarbamoyloxirane-2-carbony）-L-异亮氨酰-L-脯氨酸（CA-074），在1 μ M的浓度下，引起细胞外组织蛋白酶B几乎定量的抑制，但对基质胶侵袭没有影响。相反，同样有效，但选择性较低的抑制剂，trans-epoxysuccinyl-L-leucamino（4-guanidino）butane（E-64）抑制75%的入侵。令人惊讶的是，在10 μ M的浓度下，CA-074缓慢地渗透细胞，在12小时（侵袭测定的持续时间）后引起细胞内组织蛋白酶B的80-95%抑制。膜渗透性组织蛋白酶B抑制剂CA-074甲酯和抑制细胞内组织蛋白酶B的较高浓度的CA-074均显著降低基质胶侵袭。总的来说，这些结果确定了组织蛋白酶B在基质降解中的细胞内作用。它们还表明，当CA-074用于抑制长时间生物过程时，应谨慎假设CA-074无法进入细胞。
Cathepsin B is a lysosomal cysteine proteinase whose expression and trafficking are frequently altered in cancer, and plasma membrane and secreted forms are thought to contribute to the invasive and metastatic properties of malignant tumors. We have manipulated the expression of cathepsin B in several tumor cell lines and measured their capacity to invade through a reconstituted extracellular (Matrigel) matrix. Transient expression of human cathepsin B in a poorly metastatic B16F1 murine melanoma variant produced a 3-5-fold increase in cathepsin B activity and a comparable increase in invasiveness. Stable antisense cathepsin B-expressing clones of the highly metastatic human melanoma A375M and prostate carcinoma PC3M cell lines produced 40-50% less cathepsin B than control cells and were proportionately less invasive. In contrast, manipulating cathepsin B levels had no effect on cell migration across an uncoated membrane. The anionic cathepsin B inhibitor (L-3-trans-propylcarbamoyloxirane-2-carbony)-L-isoleucyl-L-proline (CA-074), at a concentration of 1 microM, caused a nearly quantitative inhibition of extracellular cathepsin B but had no effect on Matrigel invasion. In contrast, the equally potent but less selective inhibitor, trans-epoxysuccinyl-L-leucylamino(4-guanidino)butane (E-64) inhibited invasion by 75%. Surprisingly, at a concentration of 10 microM, CA-074 slowly permeated the cells, causing an 80-95% inhibition of intracellular cathepsin B after 12 h, the duration of the invasion assay. The membrane-permeant cathepsin B inhibitor, CA-074 methyl ester, and the higher concentration of CA-074 that inhibited intracellular cathepsin B both significantly reduced Matrigel invasion. Collectively, these results identify an intracellular role for cathepsin B in matrix degradation. They also indicate that caution should be exercised in assuming that CA-074 is unable to enter cells when it is used to inhibit biological processes of long duration.