Expression profile of genes from 12p in testicular germ cell tumors of adolescents and adults associated with i(12p) and amplification at 12p11.2-p12.1.

Gain of 12p material is invariably associated with testicular germ cell tumors (TGCTs) of adolescents and adults, most usually as an isochromosome 12p. We analyzed TGCTs with i(12p) using a global approach to expression profiling targeting chromosomes (comparative expressed sequence hybridization, CESH). This indicated overexpression of genes from 12p11.2-p12.1 relative to testis tissue and fibroblasts. The nonseminoma subtype showed higher levels of expression than seminomas. Notably, 12p11.2-p12.1 is amplified in about 10% of TGCTs and CESH analysis of such amplicon cases showed high levels of overexpression from this region. Microarray analysis, including cDNA clones representing most UniGene clusters from 12p11.2-p12.1, was applied to DNA and RNA from 5 TGCTs with amplification of 12p11.2-p12.1 and seven TGCTs with gain of the entire short arm of chromosome 12. Expression profiles were consistent with the CESH data and overexpression of EST595078, MRPS35 and LDHB at 12p11.2-p12.1 was detected in most TGCTs. High-level overexpression of BCAT1 was specific to nonseminomas and overexpression of genes such as CMAS, EKI1, KRAS2, SURB7 and various ESTs correlated with their amplification. Genes such as CCND2, GLU3, LRP6 and HPH1 at 12p13 were also overexpressed. The overexpressed sequences identified, particularly those in the region amplified, represent candidate genes for involvement in TGCT development.

Definition of chromosome aberrations in testicular germ cell tumor cell lines by 24-color karyotyping and complementary molecular cytogenetic analyses.

Many of the reported karyotypes for adult testicular germ cell tumors (GCTs) are complex and incomplete, although the presence of an isochromosome 12p, i(12p), and gain of 12p material have consistently been found. Here, an accurate definition of the chromosome aberrations associated with four cell lines derived from GCTs (GCT27, H12.1, Tera1, and Tera2) has been produced using 24-color karyotyping by mulifluor in situ hybridization, comparative genomic hybridization analysis, and further fluorescence in situ hybridization analysis to confirm some chromosomal assignments and refine involvement of specific regions of 12p. There was karyotypic heterogeneity. Isochromosomes in addition to i(12p) were found, as were other rearrangements with breakpoints at or near centromeric regions. The most frequent non-centromeric breakpoints were at 1p31 approximately p32, 1p21 approximately p22, 11q13, and Xq22, although consistent partner chromosomes were not involved. One cell line (Tera1) showed a subtle dosage increase in the copy number of a 12p probe known to be within the smallest overlapping region of amplification that has been defined in a number of testicular GCTs with amplicons at 12p11 approximately p12. The chromosome rearrangements and associated imbalances may be significant in GCT progression and the characterized cell lines can be used to investigate these further.

Additional copies of 1q in sequential samples from a phyllodes tumor of the breast.

Cytogenetic and fluorescent in situ hybridization (FISH) analysis has been performed on consecutive samples, taken 4 weeks apart, from a phyllodes breast tumor. This revealed the presence of two different chromosome 1 derivatives, namely a dic(1;10)(q10;q24) in the first sample and an i(1) (q10) in the second. In one cell out of 25 from the second sample both derivative chromosomes were seen. A chromosome 21 was lost in both samples. These results are consistent with phyllodes tumors having a clonal origin.

Cytogenetic findings in a case of nodular fasciitis of the breast.

We report the cytogenetic findings in a case of nodular fasciitis of the breast. The abnormalities found in all 11 metaphases available for analysis were -2, -2, -13, der(15)t(2;15)(q31;q26), + der(?) t(?;2), + mar1, + mar2. Other consistent abnormalities were also identified. Fluorescence in situ hybridization (FISH) was used to confirm the origin of some of the chromosomes. A large acrocentric chromosome was confirmed to be derived from chromosome 15 with chromosome 2 material translocated onto the q arm. The metacentric der(?)t(?;2) was demonstrated to have part of chromosome 2 on the q arm. No other chromosome 2 material was found. Eight of 11 cells were tetraploid and had two copies of a del(6)(q16q24).

Cytogenetic studies in patients previously treated for Hodgkin's disease.

A cytogenetic study was made of bone marrow cells and lymphocytes from patients who had been successfully treated with various regimens for Hodgkin's disease. Most of the patients had been off treatment for at least 3 years before the study began. They were divided into three groups according to the intensity of the therapy received. The frequency of gaps and breaks in the chromosomes of lymphocytes was above normal limits and similar in the three treatment groups. In contrast, the frequency of both lymphocytes and bone marrow cells with rearranged karyotypes was correlated with the intensity of treatment. Clones of cells with an abnormal karyotype were found in only two patients, both of whom were in the group receiving the most intensive therapy, i.e., chemotherapy and total nodal irradiation.

Patterns of maturation in short-term culture of human acute myeloid leukaemic cells.

Leukaemic cells taken from the blood of patients with acute myelogenous leukaemia (AML) frequently proliferate in suspension culture without the addition of growth factors for a limited period only. After a 6--10-fold increase in total cells, cell numbers remain constant for a time and finally decline. The main cause for this limited growth in vitro is not, initially at least, cell death leading to a steady state, but maturation associated in its final stages with cessation of DNA synthesis. Two populations of AML cells from Patients St and Wi respectively were studied, and progressive maturation towards mature leucocytes was demonstrated by the gradual acquisition in culture by the growing blast cells of intracellular enzymes (lysozyme, arginase, acid phosphatase and esterase being measured), surface markers (Fc and C3 receptors), of lactoferrin by Wi cells and of colony-stimulating activity by St cells, as well as changes in Ia antigens, phagocytic properties, morphology and adhesiveness to plastic. With St cells, which carried a characteristic chromosome marker, maturation terminated in cells with the characteristic properties of macrophages. At an intermediate stage, non-adherent and still-dividing St cells acquired Fc and C3 receptors and enzymes characteristic of monocytes. Wi cells progressively became neutrophil-like, and again there was an intermediate population of dividing cells which had Fc and C3 receptors and proteins such as lactoferrin and esterases. characteristic of neutrophils.

Factors influencning mitotic yields from suspension cultures of normal and acute leukaemic bone marrows.

Cytogenetic studies of bone marrow are often hampered by a paucity of dividing cells. In an attempt to improve mitotic yields we have tried to promote mitotic activity in suspension cultures of bone marrow from both normal and leukaemic subjects. Enrichment of the media always increases yield and the addition of bone marrow stimulating factor (BMSF) may do so. The best prospect for improving the yield of mitoses in acute myeloid leukaemia is to culture the marrow in an enriched medium with BMSF.

Chromosome banding studies in acute leukaemia at diagnosis.

Cytogenetic study by a chromosome banding technique has been attempted in 93 cases of acute leukaemia at diagnosis. Banding patterns were difficult to visualise in the bone-marrow chromosomes of patients with acute leukaemia because of the fuzzy appearance of the fixed metaphases. The proportion of patients with abnormal chromosomes was higher in acute lymphoblastic (ALL) than in acute myeloid (AML) leukaemia. Abnormalities were present in all cases of other cytological types. Hyperdiploidy was the most commonly found numerical error in both ALL and AML but a larger proportion of patients with ALL had hyperdiploidy in more than 30% of the cells. In ALL it was generally found that the higher the frequency of hyperdiploidy the greater was the number of chromosomes per cell. Hypodiploidy not attributable to random losses was found in only 6 patients. Clones identified by rearranged or marker chromosomes were found in all types of leukaemia. Clones marked by a 7q-chromosome, in which the break point was the same, were identified in 1 adult with ALL and 2 children with AML. The high frequency of randomly disturbed chromosomal breakage found in the bone-marrow chromosomes of a high proportion of the patients may be related to the disease process.