Regulation of breast cancer growth by activation peptide

激活肽调节乳腺癌生长

• 批准号: 6942739
• 负责人: VACLAV VETVICKA
• 金额: $ 20.59万
• 依托单位: UNIVERSITY OF LOUISVILLE
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-01 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6942739
• 关键词: affinity chromatography; aspartic endopeptidases; athymic mouse; biological signal transduction; breast neoplasms; cathepsin D; cell growth regulation; complementary DNA; enzyme biosynthesis; enzyme mechanism; growth factor; growth factor receptors; metastasis; mitogens; neoplasm /cancer chemotherapy; neoplasm /cancer pharmacology; nucleotides; prognosis; ribozymes; synthetic peptide; tissue /cell culture; transfection; yeast two hybrid system; zymogens

DESCRIPTION (provided by applicant): The long-term objective is to develop a new treatment for breast cancer based on blockade of the autocrine growth factor activity of procathepsin D. Breast cancer cells secrete procathepsin D, the zymogen from which the aspartic proteinase cathepsin D is generated by removal of an activation peptide (APpCD). Procathepsin D has been identified as an independent prognostic factor in breast cancer. In preliminary experiments, procathepsin D was found to act as a specific autocrine growth factor for breast cancer-derived cells, but not for any other cell type tested. These effects were mediated through a new, previously unknown specific receptor moiety expressed on breast cancer cell lines. The region of procathepsin D responsible for its mitogenic activity was localized in position 36-44 of the APpCD sequence. No growth factor activity could be shown with the mature enzyme cathepsin D. The proposed specific aims are based on the central hypothesis that procathepsin D is involved in breast cancer via a specific receptor that mediates autocrine activation for increased metastatic growth. For Aim lit is hypothesized that the overproduction of procathepsin D results in an increase in the metastatic potential of breast tumor cells. A low metastatic human breast cancer cell line will be transfected with human procathepsin D cDNA such that the cells will secrete constitutively varying amounts of procathepsin D. The metastatic potential of each transfected cell line will be evaluated both in vitro and in vivo in relationship to the amount of procathepsin D secretion. In addition, the synthesis of pCD will be inhibited using specifically constructed ribozymes. Attempts will be made to determine the exact site in procathepsin D responsible for breast cancer cell growth factor activity. Synthetic peptides representing fragments of APpCD will be prepared. Amino acid substitutions in the most active peptide fragment will be used to map the essential amino acid contact sites for the receptor. For Aim 2 attempts will be made to identify the membrane receptor for procathepsin D. A synthetic peptide representing the binding site domain of procathepsin D will be used to isolate candidate receptor molecules. For Aim 3 it is hypothesized that inhibition of the APpCD interaction with its receptor will result in inhibition of cancer cell growth. Peptide analogs or complementary peptides will be prepared with D-amino acids to block the growth and malignancy of cancer cells both in vitro and in vivo. The overall goal is to generate a pharmacological agent for breast cancer based on blockage of the autocrine growth factor activity of pCD.

描述(申请人提供)：长期目标是开发一种新的治疗乳腺癌的方法，其基础是阻断原蛋白D的自分泌生长因子活性。乳腺癌细胞分泌原蛋白D，通过去除激活肽(APCD)而产生天冬氨酸蛋白酶组织蛋白D的酶原。Procathepsin D已被认为是乳腺癌的独立预后因素。在初步实验中，原蛋白D被发现作为乳腺癌来源细胞的特异性自分泌生长因子，但对任何其他被测试的细胞类型都不起作用。这些作用是通过在乳腺癌细胞系上表达的一种以前未知的新的特定受体部分来介导的。负责其有丝分裂活性的原athepsin D区域位于APpCD序列的36-44位。成熟的组织蛋白酶D不能显示生长因子的活性。提出的特定目的是基于一个中心假设，即原组织蛋白酶D通过一种特定的受体参与乳腺癌的发生，该受体介导自分泌激活以促进转移生长。AIM LIT假设过量生产原蛋白D会导致乳腺肿瘤细胞转移潜能的增加。一种低转移的人乳腺癌细胞系将被人原蛋白D基因导入，以使该细胞分泌不同量的原蛋白D。将在体外和体内评估每个细胞系的转移潜能与原蛋白D分泌量的关系。此外，使用特殊构建的核酶将抑制PCD的合成。将尝试确定负责乳腺癌细胞生长因子活性的Proathepsin D的确切位置。将制备代表APpCD片段的合成肽。最活跃的多肽片段中的氨基酸替换将被用来绘制受体的必需氨基酸接触位点。为了达到这个目的，将进行2次尝试来鉴定原蛋白D的膜受体，一个代表原蛋白D结合部位结构域的合成肽将被用来分离候选受体分子。对于目标3，假设抑制APpCD与其受体的相互作用将导致抑制癌细胞生长。用D-氨基酸制备多肽类似物或互补多肽，在体外和体内都能阻断癌细胞的生长和恶性。总体目标是基于阻断PCD的自分泌生长因子活性来产生一种治疗乳腺癌的药物。