A novel cytogenetic abnormality r ( 7 ) ( : : p 11 . 2-> q 36 . 3 : : ) in a Philadelphia-positive chronic myeloid leukemia case

The so-called Philadelphia (Ph) chromosome is present in more than 90% of chronic myeloid leukemia (CML) cases. It results in juxtaposition of the 5' part of the BCR gene on chromosome 22 and the 3' part of the ABL1 gene on chromosome 9. An additional acquired monosomy 7 or deletion of 7q is associated with poor prognosis in a variety of myeloid disorders. Here we report a novel Ph chromosome positive CML case with a ring chromosome 7 [r(7)]. Immunophenotyping was compatible with CML, although 4.5% of total leucocytes appeared like acute myelogeneous leukemia (AML) subtype M2. The r(7) was characterized in detail by array-proven multicolor banding (aMCB), the latter being of enormous significance to characterize breakpoint regions in detail. Underlying mechanisms and prognostic are discussed, as ring chromosomes are rare cytogenetic abnormalities in hematopoietic malignancies.


INTRODUCTION
The so-called Philadelphia (Ph) chromosome is typical for by far over 95% of patients suffering form chronic myeloid leukemia (CML), a clonal malignant disorder of a pluripotent hematopoietic stem cell.A reciprocal translocation t(9;22)(q34;q11) leads to the formation of the Philadelphia (Ph) chromosome and a derivative of chromosome 9.The 3' portion of the ABL1 oncogene is translocated from 9q34 to the 5' portion of the BCR gene on 22q11.2.This leads to the formation of a chimeric BCR/ABL gene on the derivative chromosome 22 [1].It is known since years that the expression of the chimeric BCR/ABL protein has an increased tyrosine kinase activity and plays an essential role in the pathogenesis of CML [2].
Ring chromosomes are rare cytogenetic abnormalities that occur when the two ends of a chromosome fuse together and form a ring shape.Breaks in the chromosome arms and fusion of the proximal broken ends can lead to ring formation with loss of distal chromosomal material.Alternatively, rings can be formed by telomere dysfunction [3].
Complete or partial loss of chromosome 7, predominantly monosomy 7 or deletion of 7q, is associated with a variety of myeloid disorders, including de novo preleukemic myelodysplastic syndrome (MDS) and acute myelogenous leukemia (AML) in children and adults, as well as therapy-related ones in the latter [4,5].Also it has been reported in adult ALL, in which they frequently occur as secondary aberrations associated with a Ph chromosome and it is an adverse factor in both childhood and adult Ph + ALL [6].Additionally, deletions of 7p confer an inferior outcome in children with ALL, regardless of the presence of other poor prognostic features, whereas deletions of 7q are not associated with a worse outcome [6].
Here we reported a novel case of a Ph chromosome positive CML with r(7) and immunophenotyping consistent with CML, although 4.5% of total leucocytes showed AML M2 subtype.

Case Report
A 55-year-old male was diagnosed as suffering from CML in chronic phase (CP).In June 2010 the white blood cell count (WBC) was 93 × 10 9 /l with 49.3% neutrophils, 19.6% lymphocytes, 14.1% monocytes, 3.4% eosinophils, and 13.6% basophils.The platelets count was 181 × 10 9 /l and the hemoglobin level was 10.6 g/dl.A previous physical examination revealed splenomegaly.Serum lactate dehydrogenase (LDH) was 2057 U/l (normal up to 480 U/l), serum alanine aminotransferase (ALT) was 46 U/l (normal up to 41 U\l) and serum aspartate aminotransferase (AST) was 42 U/l (normal up to 40 U/l).Afterwards he was lost during follow-up.

Chromosome Analysis
Chromosome analysis using GTG-banding was performed according to standard procedures [7] before chemotherapeutic treatment.A total of 20 metaphase cells derived from unstimulated bone marrow culture were analyzed.Karyotypes were described according to the International System for Human Cytogenetic Nomenclature [8].

Molecular Cytogenetics
Fluorescence in situ hybridization (FISH) using LSI BCR/ABL dual color dual fusion translocation probe (Abbott Molecular/Vysis, USA) was applied according to manufacturer's instructions [9].FISH using a chromosome-7-specific aMCB probe set based on microdissection derived region-specific libraries was done as previously reported [10].A total of 20 metaphase spreads were analyzed, each using a fluorescence microscope (AxioImager.Z1 mot, Zeiss) equipped with appropriate filter sets to discriminate between a maximum of five fluorochromes plus the counterstain DAPI (4',6-diamino-2-phenylindole).Image capturing and processing were carried out using an ISIS imaging system (Meta-Systems, Altlussheim, Germany).

Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) for BCR/ABL Fusion Transcripts
RT-PCR was carried out as previously described [11].

Immunophenotyping
Immunophenotyping of leukemic blasts was performed as previously described [12].
The progression of CML from CP to blast crisis (BC) is frequently associated with nonrandom secondary chromosomal aberrations such as +8, i(17q), +19 and an extra Ph chromosome [16].
Ring chromosomes are rare cytogenetic abnormalities that occur in less than 10% of hematopoietic malignancies but have been reported in up to 70% of mesenchymal tumors [3].
Monosomy 7 or deletion of 7q in these disorders is associated with poor prognosis [17].It has been hypothesized that there is a tumor suppressor gene (TSG) on chromosome arm 7q that contributes to the pathogenesis of these diseases [18].However, monosomy 7 or structural abnormalities resulting in the deletion of 7p were infrequent in childhood ALL to confer increased risk of treatment failure in Ph-positive cases, and it has been hypothesized that a TSG on 7p may contribute to the poor outcome of these patients [6].
Both monosomy 7 and del (7p) are of the additional chromosomal abnormalities and they associated with adverse prognostic factors in CML patients treated with IM as a frontline therapy [19,20].
Recent studies have reported that major route abnormalities such as trisomy 8 at diagnosis were related to a worse outcome t (9,22) [20,21].
In conclusion, we reported here a novel case of a Ph chromosome positive CML in chronic phase with a new cytogenetic abnormality r (7) resulting in the deletion of 7p and immunophenotyping was consistent with CML although 4.5% of total leucocytes showed acute myelogeneous leukemia (AML) subtype M2.The r (7) and del 7p might be a marker for adverse prognosis in CML.