Molecular Cytogenetics Research On Double Minutes

Double minute chromosomes (DM) are circular extrachromosomalautonomously-replicating DNA fragments lacking a centromere, which havebeen observed in a large number of human tumors including breast, lung, ovary,colon, and most notably, neuroblastoma. They are a manifestation of geneamplification during the development of tumors, which give the cells selectiveadvantages for growth and survival. They frequently harbor amplified oncog-enes and genes involved in drug resistance. Although much work had beendone for DMs, rare cases can see for the origin of DMs from different tumors.All tumor cell lines with DMs were ordered from ATCC, and get the cells ofpassage3,8and13. They were characterized by routine cytogenetics, FISHand array CGH. The results of the karyotype revealed the percentage and thenumber in one cell of DMs varied a lot among different cell lines. An excellentcorrelation between the copy number gains obviously detected by array CGHand the status of the amplification found by FISH. MYC gene was presented inMC-IXC cell line,COLO-320cell line as DMs and homogenous stainingregions(HSR), in SW480cell line and HL-60cell line as DMs only. PDGFRAgene, RAF1and SOX4gene, MYEOV and CCND1gene were amplicatedseparately in SW1783cell line,5637cell line and CAMA-1cell line as HSR.Only low percentage and small amount DMs can see in T84cell line, SNU-1cell line and NCI-N87cell line, and no obviously amplification region can seein array CGH, so no confirmation FISH。Conclusion1. The breakapart of DMs are different among different tumors with sameoncogene.2. DMs come not only from HSR.3. HSR can genomically originate from single oncogene or two oncogenes from different chromosomes.4. There are no differences for DMs among different passages of the samecell line.5. Different oncogenes are involved in the DMs and HSR in the same cell,and DMs in different patients with same pathology diagnosis are different.6. It's hard to tell the cytogenetic orinal with those methods for the cellswith low double minutes.Confirmed Conclution.1. DMs are common in solid tumor but rare in hematologic tumor.2. No obvious change of amplification among different passages.3. Cells with DMs always have complex karyotyping.4. DMs and HSR can bi-transform in some tumor cells.The main innovations:(1)Collect different tumors with DMs for amplification study.(2)Combine karyotype, array CGH and FISH together to make sure theamplification in tumor cell lines.(3)Breakpoint of amplified regions can get from array CGH.