Induction of human endometrial cancer cell senescence through modulation of HIF‐1α activity by EGLN1

Abstract

Previous observations indicate that transfer of human chromosome (chr.) 1 induces senescence of endometrial cancer cells. To identify the gene(s) responsible for the senescence, we first analyzed the structural integrity of the introduced chr. 1 in immortal revertant from chr.1-transferred HHUA cells. The data demonstrated a correlation between nonrandom deletions within the 1q31-qter region and reversion to immortality. Next, by using a panel of 12 microsatellite markers, we found high frequencies of loss of heterozygosity in the particular 1q region (1q41-42), in surgically removed samples. Then, we screened the genetic mutation of the genes involved in this region, with endometrial cancer panel. Among them, EGLN1, that is a member of prolyl hydroxylase and can facilitate HIF-1 degradation by ubiquitination through the hydroxylation of HIF-1, was mutated at significantly higher frequencies (12/20, 60%). Introduction of wild-type EGLN1 into endometrial cancer cell lines (HHUA, Ishikawa and HWCA), that carry EGLN1 gene mutations induced senescence. This was invoked through the negative regulation of HIF-1 expression. In addition, alternative way of negative regulation of HIF-1 by Factor inhibiting HIF-1(FIH), SiRNA against HIF-1, and HIF-1 inhibitor, YC-1, could also induce senescence. Thus, EGLN1 can be considered as a candidate tumor suppressor on chr. 1q, and our observation could open the new aspect in exploring the machinery of senescence induction associated with HIF-1 signal transduction. These results also suggested the availability of negative regulation of HIF-1 signals for uterine cancer treatment, especially for uterine sarcomas that have worse prognosis and show a high frequency of EGLN1 gene abnormality. ' 2005 Wiley-Liss, Inc.Key words: HIF-1; EGLN1; endometrial cancer; prolyl hydroxylase; Chromosome 1Endometrial cancer is one of the most common gynecologic malignancies. Considerable evidence has suggested a number of genetic events associated with this tumor. However, the molecular pathogenesis of endometrial cancers remains poorly understood. Genetic alterations that represent gross chromosomal alterations are known to occur during endometrial carcinogenesis. Among them, nonrandom chromosome changes have been found in almost 80% of endometrial cancers and are consistent with rearrangements and deletion in a centromeric or paracentromeric area, leading to an excess of chromosome (chr.) 1q.1-3 Previously, we transferred a single human chromosome into endometrial cancer cells (HHUA and Ishikawa cells) via microcell fusion to investigate the role of chr.1 alterations in the carcinogenesis. As a result, we found that the tumorigenicity of microcell hybrid clones with an introduced chr.1, 6, or 9 is completely suppressed. However, the hybrids with an introduced chr.1 differed from those with an introduced chr. 6 or 9.4,5 A large proportion of the hybrids with chr. 1 had arrested cell growth and showed remarkable alterations in cell morphology. This finding is in contrast with the transfer of chr. 6 ...

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