对头颈部鳞状细胞癌中CDKN2B（p15INK4b）、CDKN2A（p14ARF、p16INK4a）和MTAP基因的基因结构精细定位缺失。

Fine-mapping loss of gene architecture at the CDKN2B (p15INK4b), CDKN2A (p14ARF, p16INK4a), and MTAP genes in head and neck squamous cell carcinoma.

作者信息

Worsham Maria J, Chen Kang Mei, Tiwari Nivedita, Pals Gerard, Schouten Jan P, Sethi Seema, Benninger Michael S

机构信息

Department of Otolaryngology-Head and Neck Surgery and Research Division, Henry Ford Hospital, Detroit, MI 48202, USA.

出版信息

Arch Otolaryngol Head Neck Surg. 2006 Apr;132(4):409-15. doi: 10.1001/archotol.132.4.409.

DOI:10.1001/archotol.132.4.409

PMID:16618910

Abstract

OBJECTIVE

To identify the extent and the smallest region of loss for CDKN2B(INK4b), CDKN2A(ARF,INK4a), and MTAP. Homozygous deletions of human chromosome 9p21 occur frequently in malignant cell lines and are common in squamous cell carcinoma of the head and neck (HNSCC). This complex region encodes the tumor suppressor genes cyclin-dependent kinase 2B (CDKN2B) (p15(INK4b)) and CDKN2A (p14(ARF), p16(INK4a)) and the housekeeping gene methylthioadenosine phosphorylase (MTAP).

DESIGN

A targeted probe panel designed to finely map the region of 9p21 loss comprised 3 probes for CDKN2B(INK4b), 7 for CDKN2A(ARF, INK4a), and 3 for MTAP and was interrogated using the multiplex ligation-dependent probe amplification assay (MLPA). The MLPA genomic copy number alterations for CDKN2A were validated using real-time polymerase chain reaction.

SUBJECTS

Six HNSCC primary (A) and recurrent or metastatic (B) cell lines were examined: UMSCC-11A/11B, UMSCC-17A/17B, and UMSCC-81A/81B.

RESULTS

Cell line UMSCC-11B retained all 9p loci tested in the region. Cell lines UMSCC-17A/B indicated homozygous deletion of CDKN2A(ARF, INK4a) starting at p16(INK4) exon 1alpha to include exons 2 and 3. Homozygous loss was indicated for CDKN2B(INK4b) and CDKN2A(ARF,INK4a) in UMSCC-11A, and UMSCC-81A. Cell line UMSCC-81B indicated retention of all 9p loci except for exon 1alpha (p16(INK4a)). Selective loss of the 3' end of MTAP was observed in UMSCC-11A. Genomic alterations by fine-mapping MLPA were validated at the DNA level for CDKN2A.

CONCLUSIONS

We identified exon 1alpha (p16(INK4a)) as the smallest region of loss in the CDKN2A(ARF, INK4a) gene. The frequency and precise loss of CDKN2B(INK4b), CDKN2A(ARF, INK4a), and MTAP in the prognosis of 9p21-deleted HNSCC may provide impetus for use of these targets as therapeutic biomarkers in head and neck cancer.

摘要

目的

确定细胞周期蛋白依赖性激酶2B（CDKN2B，即INK4b）、细胞周期蛋白依赖性激酶2A（CDKN2A，即ARF、INK4a）和甲硫腺苷磷酸化酶（MTAP）缺失的范围及最小缺失区域。人染色体9p21的纯合缺失在恶性细胞系中频繁发生，在头颈部鳞状细胞癌（HNSCC）中也很常见。该复杂区域编码肿瘤抑制基因细胞周期蛋白依赖性激酶2B（CDKN2B）（p15INK4b）和细胞周期蛋白依赖性激酶2A（p14ARF、p16INK4a）以及管家基因甲硫腺苷磷酸化酶（MTAP）。

设计

一个旨在精细定位9p21缺失区域的靶向探针组，包含用于CDKN2B（INK4b）的3个探针、用于CDKN2A（ARF、INK4a）的7个探针和用于MTAP的3个探针，并使用多重连接依赖探针扩增法（MLPA）进行检测。使用实时聚合酶链反应验证CDKN2A的MLPA基因组拷贝数改变。

研究对象

检测了6种HNSCC原代（A）及复发或转移（B）细胞系：UMSCC-11A/11B、UMSCC-17A/17B和UMSCC-81A/81B。

结果

细胞系UMSCC-11B保留了该区域检测的所有9p位点。细胞系UMSCC-17A/B显示CDKN2A（ARF、INK4a）从p16INK4外显子1α开始纯合缺失，包括外显子2和3。UMSCC-11A和UMSCC-81A中CDKN2B（INK4b）和CDKN2A（ARF、INK4a）显示纯合缺失。细胞系UMSCC-81B显示除外显子1α（p16INK4a）外所有9p位点均保留。在UMSCC-11A中观察到MTAP 3'端的选择性缺失。通过精细定位MLPA检测到的基因组改变在DNA水平上对CDKN2A进行了验证。