SOMATIC GENETIC ALTERATIONS IN GASTROINTESTINAL CANCER

我们采用了一种DNA指纹方法来检测肿瘤特异性 基因组差异。这种方法是基于对基因组的比较 同一个体的正常组织和肿瘤组织的指纹， 利用随机引发式聚合酶扩增DNA序列 链式反应(AP-PCR)。该方法能够检测出单一的和 简单的实验，包括数量和质量上的基因组差异 与肿瘤转化有关。它还允许克隆 这些改变的DNA序列的一个步骤，包括那些在 肿瘤细胞。 我们建议使用这种方法作为分子替代 实体瘤的细胞遗传学分析；评估肿瘤的程度和比较 发生在不同肿瘤中的基因改变的光谱 消化道在肿瘤进展的不同阶段；探讨其 结肠癌和结肠癌作为预后指标的潜在价值 直肠；以及在结直肠和其他组织中寻找未知的癌症基因 胃肠道肿瘤。 我们将检验这样一种假设，即遗传损伤的相对值 通过AP-PCR方法获得的非整倍体的程度反映了 癌细胞，这些信息可能对癌症有价值 诊断和预后。未定性的癌症基因的存在 参与明确的胃肠道疾病的发展和/或进展 肿瘤将通过分离和鉴定染色体来搜索。 代表反复遗传改变的DNA序列的定位 在这些肿瘤中。 基因改变将在一个由大约200个肿瘤组成的小组中被确定 胃肠道，包括结肠腺瘤和结肠癌 和直肠、胰腺和胃，通过比较AP-PCR图谱 肿瘤与正常组织的对比。特定于该基因的基因组改变 这些肿瘤中的任何一个的转移过程都将通过比较 原发肿瘤与转移性肿瘤采用相同的方法(特定目标1)。 我们将继续对这些肿瘤的一部分(约100例)进行随访 结直肠癌)，已经平均随访了 复发和存活3年以上。这些肿瘤已经 以ras癌基因突变和p53肿瘤为特征 抑制子基因。获得的遗传损伤的相对值 AP-PCR法检测的结直肠癌与ras和ras相关。 P53基因突变与复发率和生存率(特定目标2)。 复发性DNA片段的染色体定位 肿瘤特异性基因改变将通过AP-PCR来确定 人/仓鼠细胞杂交体的扩增。在适当的时候， 这些DNA片段将被克隆和测序，以进一步表征 用标准的聚合酶链式反应方法对它们的染色体进行定位 在其他肿瘤中的存在和出现频率(特定目标3)。

We have adapted a DNA fingerprinting approach to detect tumor specific genomic differences. The approach is based in the comparison of the genome fingerprints of normal and tumor tissues from the same individuals, after the amplification of DNA sequences by the Arbitrary Priming Polymerase Chain Reaction (AP-PCR). The method is able to detect in a single and simple experiment, both quantitative and qualitative genomic differences associated with neoplastic transformation. It also allows the cloning in a single step of these altered DNA sequences, including those deleted in the tumor cells. We propose to use this approach as a molecular alternative to the cytogenetic analysis of solid tumors; to estimate the extent and compare the spectrum of genetic alterations occurring in diverse tumors of the digestive tract at different stages of tumor progression; to explore its potential value as prognostic indicator for Cancer of the colon and rectum; and to search for unknown cancer genes in colorectal and other gastrointestinal tumors. We will test the hypothesis that the relative values of genetic damage obtained by the AP-PCR method reflect the degree of aneuploidy of the cancer cells, and that this information might have value for cancer diagnosis and prognosis. The existence of uncharacterized cancer genes involved in the development and/or progression of defined gastrointestinal neoplasms will be searched by isolating and characterizing the chromosomal localization of DNA sequences representing recurrent genetic alterations in these tumors. Genetic alterations will be identified in a panel of about 200 tumors of the gastrointestinal tract including adenomas and carcinomas of the colon and rectum, pancreas and stomach, by comparison of the AP-PCR patterns of tumor versus normal tissues. Genomic alterations specific to the metastatic process of any of these tumors will be searched by comparing primary versus metastatic tumors by the same approach (specific aim 1). We will continue the follow-up of a subset of these tumors (about 100 colorectal carcinomas), which have been already followed for an average of over 3 years for recurrence and survival. These tumors have been already characterized for mutations in the ras oncogene and in the p53 tumor suppressor gene. The relative values of genetic damage obtained for these colorectal carcinomas by the AP-PCR method will be correlated with ras and p53 gene mutations and recurrence and survival rates (specific aim 2). The chromosomal localization of DNA fragments corresponding to recurrent tumor specific genetic alterations will be determined by AP-PCR amplification of panels of human/hamster cell hybrids. When appropriate, these DNA fragments will be cloned and sequenced to further characterize by standard PCR approaches their chromosomal localization and their presence and frequency in other tumors (specific aim 3).