Trisomy 8 as the sole chromosomal aberration in myelocytic malignancies: a multicolor and locus-specific fluorescence in situ hybridization study

Trisomy 8 is the most common chromosomal aberration in myelocytic malignancies, occurring both as a sole change as well as in addition to other abnormalities. In spite of this, next to nothing is known about its pathogenetic importance or its molecular genetic consequences. Possible mechanisms involved in the transformation process include dosage effects of genes mapping to chromosome 8 and presence of specific mutations or cryptic fusion genes on the duplicated chromosome. In the latter case, +8 would be secondary to a cryptic primary rearrangement and not involved in leukemogenesis as such, but rather in tumor evolution. Although hidden genetic changes have been found in some trisomies, for example, mutations in KIT in acute myelocytic leukemia (AML) with +4 and in MET in hereditary papillary kidney carcinoma with trisomy 7, none associated with +8 have so far been discovered. To address this issue, we have investigated a total of 13 cases of AML, myelodysplastic syndromes, and chronic myeloproliferative disorders with trisomy 8 as the sole chromosomal anomaly. All cases were studied by combined binary ratio multicolor fluorescence in situ hybridization (FISH) and with FISH using locus-specific probes for both arms of chromosome 8, the subtelomeric regions of 8p and 8q, and the leukemia-associated genes FGFR1, MOZ, ETO, and MYC. No cryptic changes were detected, thus excluding the possibility of gross genetic rearrangements or aberrations involving these loci on chromosome 8.     

Isodicentric 7p, idic(7)(q11.2), in acute myeloid leukemia associated with older age and favorable response to induction chemotherapy: a new clinical entity?

Three adult de novo acute myeloid leukemias (AML M1, M2, and M4) with an isochromosome 7p are presented. No additional abnormalities were detected by G-band and multicolor, using combined binary ratio labeling, fluorescence in situ hybridization (FISH) analyses, indicating that the i(7p) was the sole, i.e., the primary, chromosomal aberration. Although the patients were elderly--68, 72, and 78 years old--they all responded very well to chemotherapy, achieving complete remission lasting more than a year. Further FISH analyses, using painting, centromeric, as well as 7q11.2-specific YAC probes, revealed that the i(7p) contained two centromeres and that the breakpoints were located in 7q11.2. Thus, the abnormality should formally be designated idic(7)(q11.2). The detailed mapping disclosed a breakpoint heterogeneity, with the breaks in 7q11.2 varying among the cases, being at least 1,310 kb apart. Furthermore, the breakpoints also differed within one of the cases, being located on both the proximal and the distal side of the most centromeric probe used. Based on our three patients, as well as on a previously reported 82-year-old patient with AML M2 and idic(7)(q11) as the only chromosomal change, we suggest that this abnormality, as the sole anomaly, is associated with AML in elderly patients who display a good response to induction chemotherapy and, hence, have a favorable prognosis. Furthermore, the heterogeneous breakpoints in 7q11.2 suggest that the important functional outcome of the idic(7)(q11.2) is the genomic imbalance incurred, i.e., gain of 7p and loss of 7q material, rather than a rearrangement of a specific gene.     

Identification of a commonly amplified 4.3 Mb region with overexpression of C8FW, but not MYC in MYC-containing double minutes in myeloid malignancies

Double minutes (dmin), the cytogenetic hallmark of genomic amplification, are found in approximately 1% of karyotypically abnormal acute myeloid leukemias (AML) and myelodysplastic syndromes (MDS). The MYC gene at 8q24 has been reported to be amplified in the majority of the cases, and generally it has been assumed that MYC is the target gene. However, only a few studies have focused on the extent of the amplicon or on the expression patterns of the amplified genes. We have studied six cases (five AML and one MDS) with MYC-containing dmin. Detailed fluorescence in situ hybridization analyses identified a common 4.3 Mb amplicon, with clustered proximal and distal breakpoints, harboring eight known genes (C8FW, NSE2, POU5FLC20, MYC, PVT1, AK093424, MGC27434 and MLZE). The corresponding region was deleted in one of the chromosome 8 homologues in five of the six cases, suggesting that the dmin originated through extra replication (or loop-formation)--excision--amplification. Northern blot analysis revealed that MYC was not overexpressed. Instead, the C8FW gene, encoding a phosphoprotein regulated by mitogenic pathways, displayed increased expression. These results exclude MYC as the target gene and indicate that overexpression of C8FW may be the functionally important consequence of 8q24 amplicons in AML and MDS.     

Isochromosome 17q in blast crisis of chronic myeloid leukemia and in other hematologic malignancies is the result of clustered breakpoints in 17p11 and is not associated with coding TP53 mutations

An isochromosome of the long arm of chromosome 17, i(17q), is the most frequent genetic abnormality observed during the disease progression of Philadelphia chromosome-positive chronic myeloid leukemia (CML), and has been described as the sole anomaly in various other hematologic malignancies. The i(17q) hence plays a presumably important pathogenetic role both in leukemia development and progression. This notwithstanding, the molecular consequences of this abnormality have not been investigated in detail. We have analyzed 21 hematologic malignancies (8 CML in blast crisis, 8 myelodysplastic syndromes [MDS], 2 acute myeloid leukemias, 2 chronic lymphocytic leukemias, and 1 acute lymphoblastic leukemia) with i(17q) by fluorescence in situ hybridization (FISH). Using a yeast artificial chromosome (YAC) contig, derived from the short arm of chromosome 17, all cases were shown to have a breakpoint in 17p. In 12 cases, the breaks occurred within the Smith-Magenis Syndrome (SMS) common deletion region in 17p11, a gene-rich region which is genetically unstable. In 10 of these 12 cases, we were able to further map the breakpoints to specific markers localized within a single YAC clone. Six other cases showed breakpoints located proximally to the SMS common deletion region, but still within 17p11, and yet another case had a breakpoint distal to this region. Furthermore, using chromosome 17 centromere-specific probes, it could be shown that the majority of the i(17q) chromosomes (11 of 15 investigated cases) were dicentric, ie, they contained two centromeres, strongly suggesting that i(17q) is formed through an intrachromosomal recombination event, and also implicating that the i(17q), in a formal sense, should be designated idic(17)(p11). Because i(17q) formation results in loss of 17p material, potentially uncovering the effect of a tumor suppressor on the remaining 17p, the occurrence of TP53 mutations was studied in 17 cases by sequencing the entire coding region. In 16 cases, no TP53 mutations were found, whereas one MDS displayed a homozygous deletion of TP53. Thus, our data suggest that there is no association between i(17q) and coding TP53 mutations, and that another tumor suppressor gene(s), located in proximity of the SMS common deletion region, or in a more distal location, is of pathogenetic importance in i(17q)-associated leukemia.     

Evidence for a single-step mechanism in the origin of hyperdiploid childhood acute lymphoblastic leukemia

High hyperdiploidy (>50 chromosomes) in childhood acute lymphoblastic leukemia (ALL) is characterized by nonrandom multiple trisomies and tetrasomies involving in particular chromosomes X, 4, 6, 8, 10, 14, 17, 18, and 21. This characteristic karyotypic pattern, the most common in pediatric ALL, may arise via a tetraploid state with subsequent loss of chromosomes, by sequential gains of chromosomes in consecutive cell divisions, or by simultaneous gain of chromosomes in a single mitosis. These alternatives may be distinguished by investigation of the allelic ratios of loci on the tetrasomic and disomic chromosomes. Previous studies of tetrasomy 21 and of the occurrence of uniparental disomies (UPDs) have suggested that the most likely mechanism is simultaneous gain. However, the other pathways have not been definitely excluded because complete analyses of all disomies and tetrasomies have never been performed. In the present study, we investigated 27 hyperdiploid ALLs by using 58 polymorphic microsatellite markers mapped to 23 of the 24 human chromosomes. Twenty-six tetrasomies were analyzed (involving chromosomes X, 8, 10, 14, 18, and 21), and the frequency of UPDs was determined in 10 cases. In total, 200 chromosomes were studied. Equal allele dosage was observed in 24 of 26 tetrasomies, and only 7 UPDs were found. These data strongly suggest that hyperdiploidy in childhood ALL generally arises by a simultaneous gain of all additional chromosomes in a single abnormal mitosis.     

The seroprevalence of ten zoonoses in two villages of Crete,Greece

The seroprevalence and incidence of 10 zoonoses due toRickettsia typhi, R. conorii, Coxiella burnetii, Burcella sp.,Borrelia sp.,Toxoplasma sp.,Leishmania sp.,Entamoeba histolytica, Echinococcus granulosus andFasciola hepatica were studied in an animal husbandry and a farming village in Crete, Greece. The serum conversion incidence of each infectious agent was determined by testing 2 blood samples, collected in 1985 and in 1987. The surveillance was conducted using detailed transparent maps of the 2 villages studied, on which epidemiological data were interrelated to the results obtained from the serological tests. Thus the importance and spread of each infection were visualized.C burnetii, Toxoplasma sp.,R. conorii, andE. granulosus, were the most common infectious agents encountered during this study.     

Tiling resolution array comparative genomic hybridization, expression and methylation analyses of dup(1q) in Burkitt lymphomas and pediatric high hyperdiploid acute lymphoblastic leukemias reveal clustered near-centromeric breakpoints and overexpression of genes in 1q22-32.3

Although gain of 1q occurs in 25% of Burkitt lymphomas (BLs) and 10% of pediatric high hyperdiploid acute lymphoblastic leukemias (ALLs), little is known about the origin, molecular genetic characteristics and functional outcome of dup(1q) in these disorders. Ten dup(1q)-positive BLs/ALLs were investigated by tiling resolution (32k) array CGH analysis, which revealed that the proximal breakpoints in all cases were near-centromeric, in eight of them clustering within a 1.4 Mb segment in 1q12-21.1. The 1q distal breakpoints were heterogeneous, being more distal in the ALLs than in the BLs. The minimally gained segments in the ALLs and BLs were 57.4 Mb [dup(1)(q22q32.3)] and 35 Mb [dup(1)(q12q25.2)], respectively. Satellite II DNA on 1q was not hypomethylated, as ascertained by Southern blot analyses of 15 BLs/ALLs with and without gain of 1q, indicating that aberrant methylation was not involved in the origin of dup(1q), as previously suggested for other neoplasms with 1q rearrangements. Global gene expression analyses revealed that five genes in the minimally 57.4 Mb gained region--B4GALT3, DAP3, RGS16, TMEM183A and UCK2--were significantly overexpressed in dup(1q)-positive ALLs compared with high hyperdiploid ALLs without dup(1q). The DAP3 and UCK2 genes were among the most overexpressed genes in the BL case with gain of 1q investigated. The DAP3 protein has been reported to be highly expressed in invasive glioblastoma multiforme cells, whereas expression of the UCK2 protein has been correlated with sensitivity to anticancer drugs. However, involvement of these genes in dup(1q)-positive ALLs and BLs has previously not been reported.