A Novel Chromosome Abnormality in Human Neuroblastoma and Antifolate-Resistant Chinese Hamster Cell Lines in Culture 1 2

Abstract

Four cell lines, SK-N-SH, SK-N-MC, SK-N-BE(2), and IMR-32, established in vitro from tumor tissue of patients with neuroblastoma were analyzed by trypsin-Giemsa banding methods. In two of the lines a large, abnormally staining chromosome region was observed. This "homogeneously staining region" (HSR) was considerably longer than any of the bands present in normal human cells and, as revealed by both G- and Q-banding, stained with an intermediate intensity. It was located on chromosomes No 6, 10, 17, or 19 of the SK-N-BE(2) cell line and on chromosome No 1 of the IMR-32 line. In concurrent studies, long HSR's were also observed in Chinese hamster sublines that had been exposed to and had developed high levels of resistance to methotrexate or methasquin and high levels of activity of target enzyme dihydrofolate reductase. For several sublines with the highest levels of enzyme activity, approximately 2% of the total cell protein was dihydrofolate reductase. Of 13 independently derived sublines with acquired resistance to antifolate, only those 7 with greater than 100-fold increases in enzyme activity consistently exhibited HSR's. These regions comprised 2-5% of the total length of the chromosome complement and were specifically localized, as demonstrated by G-banding. Analysis of chromosome replication patterns of the HSR in human neuroblastoma and in drug-resistant Chinese hamster cells by tritiated thymidine radioautography indicated that the long, abnormally staining region replicated relatively rapidly and synchronously and terminated replication before the midpoint of the S phase. The HSR thus appeared to represent a novel chromosome abnormality that may be present in cells with specialized functions. Drug-resistant Chinese hamster cells were characterized by overproduction of target enzyme, whereas human neuroblastoma cells had phenotypes of normal neuronal cells. Whether the HSR is transcriptionally active was not elucidated.

Methotrexate-resistant Chinese hamster ovary cells have amplified a 135-kilobase-pair region that includes the dihydrofolate reductase gene.

“…Certain cell lines contain amplified sequences that appear to replicate synchronously with respect to one another and therefore behave as ifthey were composed of one or a few replicon types (1,2). These sequences are found in a variety of methotrexate (MTX)-resistant mouse and hamster cell lines, and arise through the amplification of a large segment which includes the gene for dihydrofolate reductase (DHFR; tetrahydrofolate dehydrogenase; 5,6,7,8-tetrahydrofolate:NADP' oxidoreductase, EC 1.5.1.3) (3-6) as well as unknown other genetic material.…”

“…These sequences are found in a variety of methotrexate (MTX)-resistant mouse and hamster cell lines, and arise through the amplification of a large segment which includes the gene for dihydrofolate reductase (DHFR; tetrahydrofolate dehydrogenase; 5,6,7,8-tetrahydrofolate:NADP' oxidoreductase, EC 1.5.1.3) (3-6) as well as unknown other genetic material. Several hundred copies of the amplified sequence are arranged in tandem on one or a few chromosome arms, and constitute homogeneously staining regions (HSRs) on mitotic chromosomes when subjected to G-and C-banding protocols (1,3,4,6). It has been shown that the HSRs in Chinese hamster lines begin to synthesize DNA over their length synchronously at many loci at the very beginning ofthe S period; in one ofthese lines, replication of the HSR is completed by the third hour of a 7-hr S period (2).…”

Substrate-specific effects of pirinixic acid derivatives on ABCB1-mediated drug transport

“…* P < 0.05 relative to untreated control. [18][19][20]. YS71 and YS80 displayed similar efficacy in the resistant neuroblastoma cell lines compared to UKF-NB-3 as indicated by fold changes IC 50 resistant cell line/ IC 50 UKF-NB-3 < 2.…”

“…In addition, we determined the effects of these compounds on the viability of Be(2)-C cells. Be(2)-C is a clonal subline of the neuroblastoma cell line SK-N-BE(2) that was isolated from a neuroblastoma patient after repeated courses of chemotherapy and radiotherapy [18] and that displays multi-drug resistance [19,20].…”

Determination of chromosomal area of double minute chromosomes and a homogeneously staining region in HL-60 human leukemia cells by the use of a color image analyzer

“…Homogeneously staining regions (HSR) are novel chromosome abnormalities which stain uniformly instead of the pattern of alternating pale and dark bands seen in normal chromosomes when stained with either trypsin-Giemsa or other banding procedures (Biedler and Spengler, 1976). Double minute chromosomes (drain), another type of unusual abnormality (Spriggs et al, 1962), are small, paired chromatin bodies whose number and size vary from one cell to another (Barker and Hsu, 1979;Cox et al, 1965).…”