Isolation of a novel tumor suppressor gene on the chromosome 13q14 associated with metastasis of oral cancer

我们利用18个不同的多态性位点，采用聚合酶链反应(PCR) -杂合性缺失（LOH）测定法研究了54例无血缘关系的口服SCC患者的肿瘤。这些结果使得在13q14.3定位的一个假定的最小缺失区域得以识别。通常被删除的区域位于RBI位点附近，但端粒较近。我们还通过免疫组织化学分析、pcr -单链构象多态性（SSCP）分析来检测RBI基因的失活和突变，以检测RBI基因的突变。然而，在我们的样本中既没有检测到RBI基因的突变也没有检测到RBI基因的失活。有趣的是，我们发现13q14.3的LOH与淋巴结转移有显著的相关性。这些结果提示在D13S273-D13S176位点附近存在一个新的抑制基因，该基因可能在口腔鳞状细胞癌的淋巴结转移中起作用。为了在D13S273-D13S176位点附近寻找可能在淋巴结转移中起作用的推定肿瘤抑制基因的候选基因，利用这些位点上有无LOH的肿瘤，采用差异显示法和代表性差异分析。分离出两个DNA片段，证实存在于染色体13q上。利用这些DNA片段作为探针，Northern blot分析发现，在有和没有淋巴结转移的肿瘤中，这些片段的表达水平不同，提示这些片段可能是推定的肿瘤抑制基因的一部分，参与了转移。

We studied tumors from 54 unrelated patients with oral SCC by the polymerase chain reaction (PCR) -loss of heterozygosity (LOH) assay, using 18 different polymorphic loci. The results enabled the identification of a putative minimal region of deletion mapped at 13q 14.3. The commonly deleted region is located close, but telomeric to the RBI locus. We also examined the same samples for inactivation and mutation of the RBI gene by immunohistochemical analysis, PCR-single-strand conformational polymorphism (SSCP) analysis to detect any mutation of the RBI gene. However, neither mutation nor inactivation of the RBI gene was detected in our samples. Interestingly, we found significant correlation between LOH of 13q14.3 and lymph node metastasis. These results suggest the existence of a new suppressor gene near D13S273-D13S176 loci, which may play a role in lymph node metastasis of oral SCC.To identify a candidate gene for the putative tumor suppressor gene near D13S273-D13S176 loci, which may play a role in lymph node metastasis, differential display method and representational difference analysis, using tumors with and without LOH at these loci. Two DNA fragments were isolated and confirmed the presence on chromosome 13q. Using these DNA fragments as probes, Northern blot analysis revealed the different expression levels between tumors with and without lymph node metastasis, suggesting that these fragments may be a part of putative tumor suppressor genes involved in metastasis.