ACTIVATION-SPECIFIC C-ERBB-2 DETECTION IN BREAST CANCER

大约50%的乳腺癌患者有淋巴 结节阴性肿瘤。尽管这类患者的总体预后 实质上好于那些具有正结节的患者。30%会 仍然死于转移性疾病。节点正向体验 疾病表明，这个假阴性患者队列可能源于 从辅助性全身治疗中获得显著的生存益处。 C-erbB-2(neu，HER-2)受体的表达被认为是一种 预测人类乳腺癌自然病程的标准，但 这种受体在早期疾病中的预测性仍然存在 不确定。我之前的工作证实了c-erbB-2受体的存在 以两种可相互转换的形式-活化的175 kDa(p(175))酪氨酸- 磷酸化形式，以及一个不活跃的185 kDa(P185)酪氨酸- 去磷酸化形式。这项工作导致了P175的表征 激活特异性抗体在人肿瘤细胞中的表达 通过复制合成的磷肽免疫动物而产生的 一个主要的c-erbB-2自动磷酸化位点的序列。现在 该提案旨在澄清该病的发病和预后意义。 利用该新基因研究人乳腺癌c-erb-B-2表型 技术该项目有三个目标： 第一个目标是确定信号的重要性 通过生成c-EBB-2受体的单个自动磷酸化位点 相关磷酸肽和脱磷肽的多克隆抗血清： 第二个目标是提高特定于激活和失活的 识别不同磷肽序列的单抗 C-EBB-2受体激活状态。 第三个目标是使用这些激活特异性的单抗 确定p175和p185在侵袭前的表达的抗体， 浸润性和转移性乳腺肿瘤及其相互关系 受体亚型与c-ebb-2配体(Heregrin)的表达 MRNA和蛋白质水平。 这些临床病理相关性将确定 辅助性全身治疗的前瞻性随机研究 P175、p185和hereglin在结节阴性乳腺肿瘤中的表达 鉴于激活的到来，这一目标既是可以实现的，也是及时的- 国家特有的磷蛋白免疫检测(APID)技术， CRC乳房小组在人类特性方面的记录 活体乳腺肿瘤与查令十字新近的融合 帝国理工学院医学院。最多的 然而，这个项目成功的重要因素是， CRC实验室提供对4000美元冻结银行的无限制访问 乳腺癌样本，连同计算机数据库提供 乳腺癌的临床表现，疾病分期， 治疗反应和生存结果。

Approximately 50% of patients presenting with breast cancer have lymph node-negative tumors. Although the overall prognosis for such patients is substantially better than for those with positive node. 30% will still succumb to metastatic disease. Experience with node-positive disease suggests that this false-negative patient cohort may derive significant survival benefit form adjuvant systemic therapy. Expression of the c-erbB-2 (neu, HER-2) receptor has been proposed as one criterion for predicting the natural history of human breast cancer, but the predictivity of this receptor in early-stage disease remains uncertain. My previous work established that c-erbB-2 receptors exist in two interconvertible forms- an activated 175kDa(p(175) tyrosine- phosphorylated form, and an inactive 185kDa(p185) tyrosine- dephosphorylated form. This work led to the characterization of p175 expression in human tumor cells using activation-specific antibodies created by immunizing animals with synthetic phosphopeptides duplicating the sequence of a major c-erbB-2 autophosphorylation site. The present proposal aims to clarify the pathogenetic and prognostic significance of the c-erb-B-2 phenotype in human breast cancer by exploiting this novel technology. The project has three objectives: The first objective is to determine the signalling significance of individual c-ebb-2 receptor autophosphorylation sites by generating polyclonal antisera to the relevant phospho- and dephosphopeptides: The second objective is to raise activation- and inactivation-specific monoclonal antibodies to phosphopeptide sequences identifying different c-ebb-2 receptor activation states. The third objective is to use these activation-specific monoclonal antibodies to define expression of p175 and p185 in pre-invasive, invasive and metastatic breast tumors, and to correlate presence of these receptor isoforms with expression of the c-ebb-2 ligand (heregulin) at the mRNA and protein levels. These clinicopathologic correlations will establish the feasibility of a prospective randomized study of adjuvant systemic treatment based on expression of p175, p185 and heregulin in node-negative breast tumors. This goal is both achievable and timely given the advent of activation- state-specific phosphoprotein immunodetection (APHID) technology, the track record of the CRC Breast Group in the characterization of human breast tumors in vivo, and the recent amalgamation of Charing Cross Medical School with Imperial College of Science and Technology. The most important factor favoring success in this project, however, is that the CRC Laboratories provided unlimited access to a bank of 4000 frozen breast cancer specimens, together with computerized databases supplying all details of breast cancer clinical presentation, disease stage, therapeutic response and survival outcome.