ONCOLOGY REPORTS 12: 207-211, 2004 Ectopic expression ofzyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLKJIHGFEDCBA DICE1 suppresses tumor cell growth ILSEWIELAND 1 , CHRISTIAN SELL 2 , ULRICH H. WEIDLE 3 and P. WIEACKER 1 Institute of Human Genetics, Otto-von-Guericke-University, Magdeburg; 2 Lankenau Institute for Medical Research, Wynnewood, PA; Roche Diagnostics GmbH, Penzberg, Germany Received February 26, 2004; Accepted May 5, 2004 zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQPONMLK Abstract. The tumor suppressor gene D1CE1 is located within a previously reported critical region of loss of heterozygosity on chromosome 13ql4.3. Expression of the remaining DICE] allele is down-regulated in non-small cell lung carcinomas. Ectopic expression of D1CE1 cDNA by DICE 1-green fluor- escent protein fusion constructs resulted in inhibition of colony formation of human non-small cell lung carcinoma cell line SK- MES-1 and NCI-H520 and prostate carcinoma cell line DU145. In IGF-IR transformed Balb/c 3T3, DICE! substantially sup- pressed growth in soft agar. These results demonstrate that DICE1 has a growth-suppressing activity and interferes with anchorage-independent growth of IGF-IR transformed tumor cells dependent upon IGF-I signaling. Introduction In cancer cells, multiple chromosomal deletions often extending over large genomic distances are detected by loss of hetero- zygosity (LOH) and comparative genomic hybridization (CGH). The molecular search for genes in these regions that function as tumor suppressor genes has been particularly intensive in previous years. The 13ql4 region is one of the chromosomal regions frequently affected by allelic deletions in lung, oral and prostate cancer and other solid tumors, as well as in lymphoid malignancies (1-7). Of the known tumor suppressor genes, the retinoblastoma predisposing gene RBI is localized in this region. However, in non-small cell lung carcinomas, prostate and other solid tumors, RBI is not the main target of gene inactivation suggesting that additional tumor suppressor genes are located in 13ql4 (1-6). Extended allelic deletions at 13ql4 may indicate loss of more than a single tumor suppressor gene in particular tumor types as has been observed for the 3p21 chromosomal region in lung cancer Correspondence to: Dr Ilse Wieland, Institut flir Humangenetik, Otto-von-Guericke-Universität, Leipzigerstr. 44, D-39120 Magdeburg, Germany E-mail: ilse.wieland@medizin.uni-magdeburg.de Abbreviations: DICE], deleted in cancer 1; EGFP, enhanced green fluorescence protein; IGF-IR, insulin-like growth factor I receptor; VWFA, Von Willebrand factor A Key words: tumor suppressor gene, DICE], growth inhibition, 13ql4 (8). The 3p21.3 region is one of the genomic regions with highest rates of allelic deletions in lung cancer. This region harbors several genes that inhibit tumor cell growth and induce apoptosis. However, a single gene prone to mutations could not be found (9). Therefore, it was suggested that allelic loss of multiple contiguous genes in 3p21.3 contributes to tumori- genesis (8). For the 13ql4 region, also several candidate tumor suppressor genes have recently been proposed. We isolated the D1CE1/DDX26 gene (OMIM *604331) from 13ql4 as a candidate tumor suppressor gene in lung cancer (10). The DICE1 gene colocalizes with microsatellite marker D13S284, and LOH at this marker frequently occurs in non-small cell lung carcinoma, oral and prostate cancer indicating deletion of one DICE1 allele in the tumor cells (11,1-3,5). Loss or down-regulation of expression of D1CE1 was observed in non-small cell lung carcinoma cell lines and primary tumors (10). This reduced expression of DICE] is associated with promoter hypermethylation of the remaining allele (11). In esophageal squamous cell carcinoma allelic loss of the DICEI gene occurred in 73% of informative patients. However, somatic mutations in the DICE! gene were detected in only 5% of the patients, indicating that mutation of DICE] is an infrequent event in this tumor entity (12). Nevertheless, the study of Li et al (12) demonstrated for the first time that the DICEI gene fulfills the predictions of the Knudson's standard two-hit model of class I tumor suppressor genes. For prostate carcinogenesis, the chromosome condensation 1-like (CHC1-L) gene has been considered as a candidate gene at 13ql4.2 (13). This gene was mapped to a smallest commonly deleted region and showed reduced expression in prostate tumors. In B-cell malignancies, the candidate genes RBI, CLLD7, KPNA3, CLLD6, and RFP2 are located within and close to the minimally deleted region, and expression of these genes is down-regulated in B-cell chronic lymphocytic leukemia patients. Mutations or epigenetic changes were not detected in these genes, and it has been proposed that haplo-insufficiency may be the mechanism to abrogate tumor suppressor activity (14). At the proximal end of the B-cell chronic lymphocytic leukemia region the FAM10A4 gene, which is related to the ST13 putative tumor suppressor gene, has been discussed as a candidate tumor suppressor gene for B-cell malignancies and prostate cancer (15). So far, the true relevance of these candidate genes for the different tumor types has not been determined. In this study, we show that ectopic expression of DICEI by transfection suppressed clonogenic growth of non-small cell