TY - JOUR
T1 - A novel loss-of-function mutation in TP53 in an endometrial cancer cell line and uterine papillary serous carcinoma model
AU - Liu, Zhihe
AU - Wan, Guanghua
AU - Heaphy, Christopher
AU - Bisoffi, Marco
AU - Griffith, Jeffrey K.
AU - Hu, Chien An A.
N1 - Funding Information:
(GOG), which was funded by the National Cancer Institute, were used in this study. The use of this sample set was approved by the University of New Mexico Health Sciences Center Human Research Committee (#04-106). DNA was isolated from 25 mg of frozen endometrial tumors, He-co50co and control human fibroblast cell lines using the DNeasy genomic DNA extraction kit (Qiagen, Valencia, CA) and the manufacturer®s protocol.
PY - 2007/3
Y1 - 2007/3
N2 - The etiology of carcinoma of the uterine endometrium (ECa) is poorly understood. However, loss of apoptosis is one of the major factors that allow cancer cells to survive and progress. Hec50co, a poorly differentiated human ECa cell line, is widely used in the investigation of ECa. Previously, Hec50co xenograft tumor model in nude mice developed an advanced phenotype, similar to that of uterine papillary serous carcinoma (UPSC). Importantly, loss-of-function mutation in tumor suppressor TP53 was found in 20-30% of all ECa and >90% of UPSC. Thus, understanding the status of TP53 in Hec50co is essential for using Heco50co as a model for UPSC. To obtain an accurate genotype-phenotype status of TP53 in Hec50co, we performed mutation and functional analysis of TP53 gene of Hec50co by RT-PCR, genomic-PCR, and cloning and expression of mutant and wildtype TP53 alleles. We found a novel 42-bp deletion mutation in the exon6-intron6 splice junction of TP53 (TP53.del42bp) leading to a 113-bp exon6-deleted/skipped transcript was identified in Hec50co. In addition, the other TP53 allele in Hec50co is inactivated through a large deletion. Adenovirus (AD) harboring wildtype full-length TP53 cDNA induces caspase-dependent apoptosis; while the AD-TP53.del42bp allele does not. In addition, messenger RNA of TP53.del42bp allele is stable whereas the protein product of TP53.del42bp allele is made but not stable. Taken together, we demonstrate that Hec50co is a TP53-null cell line possessing one TP53.del42bp allele and the other lost allele and therefore provides an excellent model to dissect the molecular and cellular bases of UPSC and other p53-null cancers.
AB - The etiology of carcinoma of the uterine endometrium (ECa) is poorly understood. However, loss of apoptosis is one of the major factors that allow cancer cells to survive and progress. Hec50co, a poorly differentiated human ECa cell line, is widely used in the investigation of ECa. Previously, Hec50co xenograft tumor model in nude mice developed an advanced phenotype, similar to that of uterine papillary serous carcinoma (UPSC). Importantly, loss-of-function mutation in tumor suppressor TP53 was found in 20-30% of all ECa and >90% of UPSC. Thus, understanding the status of TP53 in Hec50co is essential for using Heco50co as a model for UPSC. To obtain an accurate genotype-phenotype status of TP53 in Hec50co, we performed mutation and functional analysis of TP53 gene of Hec50co by RT-PCR, genomic-PCR, and cloning and expression of mutant and wildtype TP53 alleles. We found a novel 42-bp deletion mutation in the exon6-intron6 splice junction of TP53 (TP53.del42bp) leading to a 113-bp exon6-deleted/skipped transcript was identified in Hec50co. In addition, the other TP53 allele in Hec50co is inactivated through a large deletion. Adenovirus (AD) harboring wildtype full-length TP53 cDNA induces caspase-dependent apoptosis; while the AD-TP53.del42bp allele does not. In addition, messenger RNA of TP53.del42bp allele is stable whereas the protein product of TP53.del42bp allele is made but not stable. Taken together, we demonstrate that Hec50co is a TP53-null cell line possessing one TP53.del42bp allele and the other lost allele and therefore provides an excellent model to dissect the molecular and cellular bases of UPSC and other p53-null cancers.
KW - Adenovirus
KW - Allelic loss
KW - Endometrial cancer
KW - Hec50co
KW - Loss of heterozygosity
KW - Loss-of-function splicing mutation
KW - TP53
KW - TP53-null cancer
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U2 - 10.1007/s11010-006-9345-x
DO - 10.1007/s11010-006-9345-x
M3 - Article
C2 - 17119852
AN - SCOPUS:33947379092
VL - 297
SP - 179
EP - 187
JO - Molecular and Cellular Biochemistry
JF - Molecular and Cellular Biochemistry
SN - 0300-8177
IS - 1-2
ER -