PROTEASE INHIBITORS IN HUMAN BLADDER CANCER INVASION

本建议的目的是确定蛋白酶的影响 抑制剂在消除人膀胱肿瘤细胞的侵袭中的作用。 为了研究膀胱肿瘤侵袭的生化机制，我们 先前分析的侵袭性移行细胞癌细胞系EJ和 非侵袭性移行细胞癌细胞系RT4， 植入裸鼠膀胱中。 我们的研究结果表明， 组织蛋白酶B，一种半胱氨酸蛋白酶，具有降解 基底膜层粘连蛋白，存在于侵袭性组织的质膜中， 而非侵袭性肿瘤细胞具有B限于 它们的溶酶体 这表明组织蛋白酶B被重新分布， 被侵入性细胞用来降解基底膜成分。 我们 这一发现进一步表明，蛋白酶抑制剂可能发挥重要作用， 在限制人膀胱肿瘤细胞的侵袭潜力中的作用。 在这个建议中，我们将确定半胱氨酸蛋白酶的影响， 抑制质膜结合组织蛋白酶活性的抑制剂 B在生化水平上使用酶覆盖膜和等电 聚焦亚细胞组分。 此外，我们将确定 蛋白酶抑制剂在抑制降解中的作用 层粘连蛋白，一种基底膜成分，通过膜结合的组织蛋白酶B。 在我们初步的生化分析之后，我们将确定 蛋白酶抑制剂对膀胱肿瘤侵袭的抑制作用 细胞通过体外人工基底膜。 最后我们 将开发一个体内模型，以确定蛋白酶的影响， 膀胱肿瘤侵袭的抑制剂。 在这些研究中，我们不仅要确定 这种干预在生物化学、细胞和免疫学方面的有效性 体内水平，而且还解决了基本的生化机制， 肿瘤细胞侵袭。

The purpose of this proposal is to determine the effects of protease inhibitors in the abrogation of invasion by human bladder tumor cells. To study the biochemical mechanisms of bladder tumor invasion, we have previously analyzed invasive transitional cell carcinoma cell line EJ and non-invasive transitional cell carcinoma cell line RT4 which had been implanted into the bladders of nude mice. Our results demonstrated that cathepsin B, a cysteine proteinase which has the ability to degrade basement membrane laminin, is found in the plasma membrane of invasive tumor cells whereas non-invasive tumor cells have cathepsin B confined to their lysosomes. This suggests that cathepsin B is redistributed and may be used by invasive cells to degrade basement membrane components. Our finding further suggests that protease inhibitors may play a significant role in limiting the invasive potential of human bladder tumor cells. In this proposal, we will determine the effects of cysteine proteinase inhibitors in inhibiting the activity of plasma membrane bound cathepsin B at a biochemical level using enzyme overlay membranes and isoelectric focusing of subcellular fractions. Furthermore, we will determine the effects of proteinase inhibitors in inhibiting the degradation of laminin, a basement membrane component, by membrane bound cathepsin B. After our initial biochemical analyses, we will determine the effects of proteinase inhibitors in inhibiting the invasion of human bladder tumor cells through an in vitro artificial basement membrane. And finally, we will develop an in vivo model to determine the effects of protease inhibitors in bladder tumor invasion. We will attempt in these studies to determine not only the possible effectiveness of such intervention at a biochemical, cellular, and in vivo level, but also address the fundamental biochemical mechanisms of tumor cell invasion.