Alterations of DCC Gene in B-Cell Malignancies.

Abstract 4428

Many genetic alterations have been detected in various tumors to date, and these are concerned with oncogenesis closely. Oligonucleotide array analysis has contributed to detect genomic gains and losses implicated in the development of human malignant tumors, identifying either oncogenes or tumor suppressor genes. In our recent data, oligonucleotide array analysis showed recurrent genomic copy number variations at 18q21-22 in malignant lymphoma and multiple myeloma cell lines. There are a lot of genes related to oncogenesis at 18q21-22, such as SMAD4, DCC, MBD2, TCF4, TXNL1, WDR7, FECH, NARS, MALT1, and BCL2. We studied expression of these genes by reverse-transcriptase polymerase chain reaction (RT-PCR). Although most genes examined expressed in almost all cell lines examined, DCC (deleted in colorectal cancer) could be detected in a small number of malignant lymphoma cell lines.

DCC, which encodes a receptor for netrin-1, would control apoptosis and be concerned in oncogenesis. Loss of one DCC allele or decreased DCC expression occur in many solid tumors, suggesting that DCC inactivation constitutes a critical event in the development of these tumors. Similarly, the inactivation of DCC can have a role in the development of hematologic malignancies, including leukemia and malignant lymphoma. We analyzed DCC expression in 15 multiple myeloma and 9 malignant lymphoma cell lines, by RT-PCR using various primer pairs. However, no products could be detected using any primer pairs containing DCC exon 1 primer in three multiple myeloma cell lines, although other primer pairs could amplify products, suggesting the DCC transcripts lacking exon 1 in these cell lines. This result made us predict the abnormal form of DCC. Therefore, we tried to clone the DCC transcripts fused to unknown sequences instead of exon 1, using bubble PCR method for cDNA in these three cell lines. Some products were obtained and sequenceanalysis revealed that the bubble PCR products contained exon 2 of DCC fused to unknown sequences. BLAST search revealed that these unknown sequences were part of intron 1 of DCC, resulting in the lack of start codon in exon 1. Putative abnormal DCC protein lacks a part of N-terminus, suggesting that structural abnormalities of DCC play some roles in oncogenesis of hematologic malignancies. Although the previous studies described the abnormalities of DCC derived from allelic loss or decreased expression, structural abnormalities of DCC we found in this study might be another critical mechanism for DCC abnormalities even if the normal expression could be detected by RT-PCR which could amplify only limited portion of the gene.