CLINICAL CORRELATIVE STUDIES IN GERM CELL TUMORS

描述：(申请人摘要)在这份竞争性续签申请中， 申请者将对他的观察结果进行扩展，这些观察表明TP53基因发生了突变， 广泛变化的p53蛋白表达，bcl2蛋白表达，以及 有缺陷的细胞凋亡。所有这些都表明生殖细胞中的耐药性 肿瘤(GCTS)部分与细胞死亡途径缺陷有关。GCTS 对化疗表现出不同寻常的敏感性。只有20%到30%的肿瘤是 对以顺铂为基础的化疗耐药。GCTS还显示体细胞 分化(畸胎瘤，通常为组织学良性)。因此，畸胎瘤 也有“抗药性”，但化疗后出现畸胎瘤 很少会导致病人死亡。因此，两种形式的耐药性 都在现场。因此，申请者将研究相关的遗传事件。 对药物敏感(治愈)的患者未分化的耐药 GCTS，耐药(患者复发或死于疾病)，未分化 GCTS和耐药分化型GCTS(畸胎瘤)。以特定的目标 1，估计GCTS中TP53突变的频率。他的 初步数据显示，部分耐药的GCTS中存在TP53突变。 在特定的目标2中，细胞凋亡反应和表达 依赖于P53的细胞凋亡或细胞周期调节蛋白(例如，bax，bcl2， 将对P21)进行评估。他在两个GCT细胞系中的初步数据(一个 药物敏感(TP53野生型)和1例耐药(TP53突变体) 未能上调p21，TP53突变体的凋亡率没有变化 与野生型相比，当接触顺铂时，并标记 P53、Bax和bcl2蛋白在敏感型和非敏感型肺癌中的表达差异 耐药GCTS。在特定目标3，比较基因组杂交(CGH) 将用于评估基因组过剩或丢失受限区域的GCTS 到耐药肿瘤。他的初步数据确定了六个 五个不同染色体臂上的扩增区域。在其中的每一个中 具体目标，他将进一步确定是否存在关联 遗传事件(标记物)与临床终点的单因素分析 (例如，治愈与死亡)，最初使用赔率比作为衡量 关联性，其次是多变量分析，以建立更好的方法 治疗计划。对于每个肿瘤，将进行SSCP和测序 Tp53突变；bax、bcl2和p21的免疫组化检测 细胞凋亡率的表达和原位末端标记法，以及CGH的搜索区域 基因组失衡的原因。从这些数据中，申请者将开始剖析 抗性途径(S)，可能发现与GCT相关的新基因 耐药性，并确定他们是否有独立的临床 预测值。

DESCRIPTION: (Applicant's Abstract) In this competing renewal application, the applicant will expand on his observations which show mutations in TP53, widely variable p53 protein expression, bcl-2 protein expression, and defective apoptosis. All of these suggest that drug resistance in germ cell tumors (GCTs) is partly related to defects in the cell death pathway. GCTs display unusual sensitivity to chemotherapy. Only 20% to 30% of tumors are resistant to cisplatin-based chemotherapy. GCTs also display somatic differentiation (teratoma, usually histologically benign). Thus, teratomas are also "drug resistant", but the presence of teratoma after chemotherapy rarely leads to a patient's death. Therefore, two forms of drug resistance are present. Thus, the applicant will study the genetic events associated with drug resistance in drug-sensitive (patient cured) undifferentiated GCTs, drug-resistant (patient relapsed or dead of disease) undifferentiated GCTs, and drug-resistant differentiated GCTs (teratomas). In Specific Aim 1, the frequency of TP53 mutations in GCTs will be estimated. His preliminary data show that TP53 mutations exist in some drug-resistant GCTs. In Specific Aim 2, the apoptotic response and the expression of p53-dependent apoptosis or cell cycle regulatory proteins (e.g., bax, bcl-2, p21) will be assessed. His preliminary data in two GCT cell lines (one drug-sensitive (TP53 wild-type) and one drug-resistant (TP53 mutant)) show failure to upregulate p21, no change in apoptotic rate in the TP53 mutant line compared to the wild-type when exposed to cisplatin, and marked differences in p53, bax, and bcl-2 protein expression in sensitive and resistant GCTs. In Specific Aim 3, comparative genomic hybridization (CGH) will be used to evaluate GCTs for regions of genomic excess or loss limited to drug-resistant tumors. His preliminary data have identified six amplified regions on five different chromosome arms. In each of these specific aims, he will further determine whether an association exists in univariate analysis between genetic events (markers) and clinical endpoints (e.g., cured vs. dead), initially using the odds-ratio as the measure of association, followed by multivariable analysis to establish better methods of treatment planning. On each tumor, SSCP and sequencing will be performed for TP53 mutations; immunohistochemistry performed for bax, bcl-2, and p21 expression and TUNEL for apoptotic rate; and CGH performed seeking regions of genomic imbalance. From these data, the applicant will begin to dissect the resistance pathway(s), possibly identify new genes associated with GCT drug resistance, and determine whether they have independent clinical predictor value.