Antizyme-mediated inhibition of polyamine transport

描述（由申请人提供）：该项目的长期目标是阐明抗酶（AZ）在细胞多胺运输反馈调节中的作用。肿瘤细胞的生长和转移潜能与异常的多胺积累密切相关。多胺转运到癌细胞的活性增强，为细胞毒性药物靶向肿瘤提供了一个理想的系统。因此，许多以多胺为基础的抗癌化合物正在进行测试，其中一些已进入临床试验阶段。最近的研究表明，大多数这些多胺类似物也刺激AZ的产生，AZ是一种调节蛋白，通过抑制多胺转运蛋白来防止多胺的过度积累。这种az介导的对潜在有用药物摄取的限制显然对这种转运系统的药理学使用提出了严重的潜在问题。不幸的是，这种反馈系统的机制甚至最基本的特征都不为人所知。该项目的四项调查旨在探索阿斯利康参与阻断这种转运体的基本方面。该研究将受益于最近发现的短n端序列，仅存在于次要的全长AZ异构体上，对最大的运输抑制至关重要。具体来说，这些研究将(a)确定两种阿斯利空同种异构体在抑制多胺摄取和刺激多胺出口方面的相对影响；(b)确定抗酶抑制剂（AZI）在逆转反馈抑制多胺运输中的有效性；(c)探索AZ亚型与潜在转运中间分选连接蛋白-5相互作用的可能差异；(d)确定对该反馈活动重要的n端序列。运输抑制研究将使用稳定转染的CHO克隆进行，以控制不同AZ亚型、AZ抑制剂和分选连接蛋白-5的表达。通过阐明AZ参与控制这一摄取系统，本研究将改进设计和利用利用这一有用转运体的化合物的努力。

DESCRIPTION (provided by applicant): The long-term goal of this project is to clarify the role of antizyme (AZ) in feedback regulation of cellular polyamine transport. It has been well established that cancer cell growth and metastatic potential are closely associated with abnormal polyamine accumulation. The enhanced activity of polyamine transport into cancer cells provides an ideal system for targeting cytotoxic drugs specifically to tumors. Accordingly, many polyamine-based anticancer compounds are being tested and some have progressed to clinical trials. Recent studies have shown that most of these polyamine analogs also stimulate the production of AZ, a regulatory protein that prevents excessive accumulation of polyamines by inhibiting the polyamine transporter. This AZ-mediated limitation of the uptake of potentially useful drugs obviously presents a serious potential problem to the pharmacological use of this transport system. Unfortunately, the mechanisms and even the most basic characteristics of this feedback system are not known. The four investigations in this project are designed to explore fundamental aspects of AZ's involvement in blocking this transporter. The investigations will benefit from the recent discovery that a short N-terminal sequence, present only on the minor, full-length AZ isoform, is critical for maximal transport inhibition. Specifically these studies will (a) establish the relative effects of the two AZ isoforms on both inhibiting polyamine uptake and stimulating export; (b) determine the effectiveness of antizyme inhibitor (AZI) in reversing feedback inhibition of polyamine transport by each of the AZ isoforms; (c) explore possible differences in the interactions of the AZ isoforms with the potential transport intermediate sorting nexin-5; and (d) identify the N-terminal sequences that are important for this feedback activity. The transport inhibition studies will be conducted using CHO clones that have been stably transfected for controlled expression of the distinct AZ isoforms, AZ inhibitor and sorting nexin-5. By elucidating AZ's involvement in controlling this uptake system, this investigation will improve efforts to design and utilize compounds exploiting this useful transporter.