|Year : 2016 | Volume
| Issue : 1 | Page : 9-14
Clinicopathologic features and prognostic impact of isochromosome 17q in chronic myeloid leukemia patients
Hoda M El Gendi1, Dina A Fouad1, Amal A Mohamed1, Doaa G Eissa MD 1, Nevine N Mostafa2
1 Department of Clinical Pathology, Ain Shams University, Cairo, Egypt
2 Department of Internal Medicine, Ain Shams University, Cairo, Egypt
|Date of Submission||01-Sep-2015|
|Date of Acceptance||06-Sep-2015|
|Date of Web Publication||10-Mar-2016|
Doaa G Eissa
Clinical Pathology Department, Ain Shams University, 20 Nagaty Sarag Street, Nasr City, Cairo
Source of Support: None, Conflict of Interest: None
Background Isochromosome 17q (i17q) is a well-known nonrandom secondary anomaly in chronic myeloid leukemia (CML), which occurs either solely or with other additional anomalies.
Objectives The aim of the study was to explore the influence of i17q in CML patients in different phases of the disease and the prognostic impact of acquiring such an anomaly on disease progression, outcome, and response to therapy.
Materials and methods Cytogenetic analysis was carried out on 100 CML patients by G-banding and fluorescence in-situ hybridization using LSI BCR/ABL, LSI p53(17p13)/MPO (17q22) i(17q), CEP 8, and CEP Y probes.
Results Isochromosome 17q was detected in 16% of cases. All examined bone marrow smears of i(17q)-positive patients were hypercellular and showed variable degrees of dysplastic changes mainly in myeloid lineage, in the form of hyposegmentation and hypogranulation, together with dysplastic features of megakaryocytes in 70% of them. A highly significant association of i(17q) with poor prognosis was confirmed statistically (P = 0.002) compared with the prognosis in negative patients. The event-free survival of the i(17q)-positive group was 1.6 months compared with 11.5 months in negative patients. However, no statistically significant association was revealed with standard prognostic factors (P>0.05).
Conclusion Isochromosome 17q identifies a subgroup of CML with distinct clinicopathologic features and with high risk for aggressive disease progression.
Keywords: chronic myeloid leukemia, fluorescence in-situ hybridization, isochromosome 17
|How to cite this article:|
El Gendi HM, Fouad DA, Mohamed AA, Eissa DG, Mostafa NN. Clinicopathologic features and prognostic impact of isochromosome 17q in chronic myeloid leukemia patients. Egypt J Haematol 2016;41:9-14
|How to cite this URL:|
El Gendi HM, Fouad DA, Mohamed AA, Eissa DG, Mostafa NN. Clinicopathologic features and prognostic impact of isochromosome 17q in chronic myeloid leukemia patients. Egypt J Haematol [serial online] 2016 [cited 2020 Apr 10];41:9-14. Available from: http://www.ehj.eg.net/text.asp?2016/41/1/9/178464
| Introduction|| |
Chronic myeloid leukemia (CML) is a myeloproliferative disorder characterized by the presence of the Philadelphia chromosome, which is the cytogenetic hallmark of CML. It is characterized by a reciprocal translocation t(9;22)(q34;q11). The resulting molecular event is the creation of the BCR/ABL fusion gene  . In CML, 30-50% of resistant cases are associated with additional chromosomal abnormalities (ACAs). Nonrandom, extra Ph, trisomy 8 (+8), isochromosome 17 i(17q), and trisomy 19 (19+) are the most common secondary changes (present in ~13-34% of cases with additional abnormalities)  .
Recent interest in ACAs in CML patients is now gaining more importance, particularly in progressive disease  . The appearance of these ACAs during treatment is commonly known as clonal evolution and seems to play an important role in imatinib mesylate resistance. The WHO classification suggests that those patients showing ACAs emerging during treatment should be considered in accelerated phase (AP). The European Leukemia Net recommendations suggest that the presence of ACAs at diagnosis may represent a 'warning' feature, requiring careful monitoring of the patient  .
Isochromosome 17q, or i(17q), is one of the most frequent nonrandom changes occurring in human neoplasia. Most of the i(17q) breakpoints cluster within a ~240-kb interval located in the Smith-Magenis syndrome's common deletion region in 17p11.2  . It has been described as both a primary and a secondary aberration. i(17q) is a frequent secondary chromosomal aberration in the AP or blast crisis (BC) of CML, indicating that this abnormality plays an important role in disease progression , . The i(17q) is also found in 1.4-2.4% of acute myeloid leukemias (AML), chronic myeloproliferative disorders, myelodysplastic syndromes, acute lymphoblastic leukemias, and chronic lymphoproliferative disorders with clonal chromosomal aberrations  .
Isochromosome 17q is associated with the loss of the short arm (17p) and duplication of the long arm (17q) that causes the abnormality. As the p53 gene is located on the short arm of chromosome 17, isochromosome 17q is known to be the most common precedent in cancer. This rearrangement results in tumor progression and initiation. However, p53 mutations were not found in CML cases with the i(17q), raising the possibility that the relevant pathogenic mechanism in CML-BC patients with the i(17q) abnormality is loss of function of yet unidentified genes  .
The morphological presentation of bone marrow (BM) smears of i(17q)-positive CML patients includes severe hyposegmentation of neutrophil nuclei and a prominence of the monocyte/macrophage lineage. The patients have a high risk for progression to AML and a relatively short median survival  .
This work aimed to study the influence of the ACAs, particularly i17q, in CML patients at different phases of the disease and the impact of acquiring such anomalies on disease progression and the outcome regarding the response to tyrosine kinase inhibitor therapy.
| Materials and methods|| |
This study was carried out on 100 CML patients during the period from March 2010 to March 2014. Informed consent for inclusion in the study was obtained from all patients, and the study was approved by the ethical committee of Ain Shams University. Patients were diagnosed according to the WHO diagnostic criteria (2008). They were classified into three groups according to their CML phase at presentation.
Group I: This included 60 patients in the chronic phase (CP), consisting of 28 male and 32 female patients, with a mean age of 47.2 ± 16.7 years.
Group II: This included 16 patients in AP, consisting of 10 male and six female patients, with a mean age of 50.1 ± 7.3 years.
Group III: This included 24 patients in blastic phase (BP), consisting of 10 male and 14 female patients, with a mean age of 51.1 ± 8.9 years.
Fifteen BM samples of volunteers with nonspecific BM changes and normal karyotyping were included as controls for fluorescence in-situ hybridization (FISH) probes.
All patients were subjected to thorough history taking, clinical examination, and laboratory investigations, including complete blood count, examination of Leishman-stained peripheral blood (PB) and BM smears, and neutrophil alkaline phosphatase score  . Conventional cytogenetic analysis (CCA) using the Giemsa banding technique was applied following the techniques described by Verma and Babu  : FISH for detection of t(9;22)(q34;q11) using a dual-color dual-fusion locus-specific identifier probe (LSI) BCR/ABL; FISH for detection of chromosome 8 and the Y chromosome using a centromeric probe; FISH for detection of i(17q) using the locus-specific identifier LSI p53(17p13) and the MPO (17q22) probe.
Follow-up for all patients was done over a period of 12 months. All of them were on imatinib mesylate. Patient outcome was expressed according to the European Leukemia Net as per Baccarani et al.  , which classified patients according to hematological, cytogenetic, and molecular remission. According to this classification we stratified patient prognosis in our study into good and bad, where patients with good prognosis were those achieving complete hematological, complete cytogenetic, and complete molecular remission or major molecular remission, and those with bad prognosis were those with failure to achieve any of these.
Conventional cytogenetic and fluorescence in-situ hybridization analysis
Cytogenetic analysis involves the examination of spontaneously dividing cell populations at metaphase stage using colcemid. This is followed by hypotonic wash and fixation and then slide making and staining with Giemsa using trypsin to induce G-banding. Scanning and analysis of at least 20 metaphases was done using an image analyzer  .
The FISH technique was applied on culture-prepared cells in 89 patients and on archived direct BM smears in 11 patients with no PB or BM samples on lithium heparin, as per Charwudzi et al.  .
The FISH technique was applied as per the manufacturer's protocol based on denaturation of target DNA and its hybridization to a complementary single-stranded nucleic acid sequence (fluorochrome-labeled) probe. Reaction conditions were adjusted so that hybridization occurred only between the probe and the target DNA sequence of high homology. Target DNA is made visible by counterstaining using a DNA-specific fluorescent dye  . Scanning and analysis of 500 interphases or 25 metaphases was done using the image analyzer. The cutoff value for positivity was 6% for all aberrations.
Interpretation of the results
Results were interpreted according to manufacturer instructions (Vysis, Stuttgart, Germany). Negativity for i(17q) was read as two red signals for MPO (17q22) and two green signals for p53 at 17p13. Positivity for i(17q) was read as a signal pattern of three red signals for MPO (17q22) and one green signal for p53 at 17p13 ([Figure 1]). Positive for heterozygous deletion of 17p13(p53) as a sole anomaly was read as two red signals for MPO (17q22) and one green signal for p53 at 17p13 ([Figure 2]). Positive for p53 deletion and i(17q) was read as three red signals for MPO (17q22) and no green signals for p53 at 17p13.
|Figure 1 Positive case for i17q showing three red signals for MPO (17q22) and one green signal for p53 at 17p13.|
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|Figure 2 Positive case for heterozygous deletion of 17p13 (p53) as a sole anomaly: two red signals for MPO (17q22) and one green signal for p53.|
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SPSS statistics (V. 21.0, 2012; IBM Corp., Chicago, USA) were used for data analysis. Data were expressed as mean ± SD for quantitative parametric measures in addition to number and percentage for categorized data. Comparative analysis for quantitative data was performed using the Student t-test for comparison between two independent groups with normal distribution. The χ2 -test was used for comparison between two independent groups as regards qualitative data. Probability or P value less than 0.05 was considered statistically significant, whereas a P value of 0.001 or less was considered highly statistically significant.
| Results|| |
The clinical, laboratory, and prognostic characteristics of all patients are summarized in [Table 1].
Conventional cytogenetics analysis in chronic myeloid leukemia patients
The metaphase banding was analyzed in 89 patients only as no PB or BM samples were sent to the laboratory for karyotyping in the remaining 11 patients. Successful karyotyping was analyzed in 82/89 (92%) patients, whereas failed mitosis was detected in seven patients. The karyotypic patterns are summarized in [Table 2]. The Ph was positive in all of them, and ACAs were encountered in 13/82 (15.8%), with the highest rate in BC. The i(17q) was detected in 5/82 (6%) analyzable metaphases ([Table 2]).
Fluorescence in-situ hybridization analysis in chronic myeloid leukemia patients
Re-evaluation by the used FISH probes improved the ability of detection of genetic aberrations, as the Ph chromosome was detected in 100% of patients compared with 92% by CCA, as FISH analysis was applied on direct BM smears in the 11 patients with no PB or BM samples on lithium heparin. Moreover, i(17q)(q10) was detected in 16% compared with 6% only by CCA.
i(17q)(q10) was detected alone in 8% of cases and in combination with other cytogenetic abnormalities in another 8% ([Table 3]).
FISH also improved the ability of detection of ACAs, with peaks of incidence in the AP and BC phases ([Table 3]). The presence of ACAs in association with Ph were in the form of i(17q)/+der (22)/+8/−y/del 9q34 (ABL)/amplification of ABL and BCR genes and t(9;22;13).
These ACAs were detected in 6/60 patients (10%) in CP, in 7/16 (43.8%) in AP, and in 13/24 (54.2%) in BC phase. The genes amplification was detected in BC phase only ([Table 3]).
Morphological, cytogenetic, and prognostic features of i(17q)-positive patients
Bone marrow examination
Hypercellular marrow showed variable degrees of dysplastic changes in myeloid and megakaryocytic lineages, in the form of hyposegmentation (pseudo-Pelger- Huet) and hypogranulation as constant features in all examined BM smears of i(17q)-positive patients and dysplastic megakaryocytes in the form of micromegakaryocytes in 70% of patients.
Additional cytogenetic aberrations associated with i(17q)(q10)
Isochromosome 17(q) was detected in association with ACAs in 8% of CML patients in different phases: in one patient in CP, in two in AP, and in 5% in BC phases ([Table 2] and [Table 3]).
Isochromosome 17q prognostic features
To evaluate the characteristic prognostic features of i(17q), patient outcome was assessed according to the European Leukemia Net recommendations at 12 months from presentation for hematological, cytogenetic, and molecular responses, and the 100 patients were categorized into two groups according to their positivity to i(17q) ([Table 4]). No statistically significant association was encountered between i(17q) and any of age, sex, spleen size, total leukocytic count, Hb, or platelet count (P>0.05).
|Table 4 Isochromosome 17 in relation to standard prognostic parameters and patient outcome|
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A significant association of i(17q) was encountered with poor outcome (P = 0.002) ([Table 4]). This was evident as all patients with i(17q) either alone or associated with other abnormalities failed to achieve complete cytogenetic response, with only one patient showing major cytogenetic response. In addition, the mean event-free survival in i(17q)-negative patients was 11.5 months, compared with 1.6 months in the i(17q)-positive group.
| Discussion|| |
In the present study CCA revealed successful mitosis in 92% of patients and failed mitosis in 8%, with the majority of cases being from patients of AP and BP. This is because blast cells are inert in culture and fail to enter the mitotic phase. The failure rate of mitosis was higher in our study than in that reported by Aoun et al.  and Reena et al.  (1.3%), and less than that shown by Patel et al.  (43.6%).
All patients who showed successful mitosis were positive for Ph chromosome by CCA, which agrees with the findings of Vardiman et al.  , who stated that the percentage of Ph in CML is 90-95% on CCA. ACAs were encountered in 13/82 cases (15.8%), with the highest rate in BP. This was consistent with the reports of Aoun et al.  and Reena et al.  (11% for both). Higher percentages of complex aberrations in AP and BP than in CP were previously documented by the well-established role of additional anomalies in clonal evolution and acceleration of CML  .
Regarding i(17q), it was detected in 5/82 (6%) analyzable metaphases by CCA, and in 16% of CML patients at different phases by means of the FISH technique. It was detected solely in 8% patients and associated with other anomalies in 8%. This was consistent with the findings of Lazarevic  , who reported that i(17q) occurs in CML either solely in 10% of cases, or with other additional anomalies in at least another 10% of cases. In contrast, Barbouti et al.  mentioned that i(17q) is one of the most frequent changes observed during disease progression of CML, occurring in ~21% of cases with chromosomal abnormalities in addition to the pathognomonic reciprocal translocation t(9;22).
A statistically significant association between the presence of i(17q) and poor prognosis was detected. This was evident as all patients with i(17q) either alone or associated with other abnormalities failed to achieve complete cytogenetic response, with only one that showed major cytogenetic response. This was in agreement with many previous studies ,, . Moreover, patients with i(17q) showed shorter event-free survival in all phases. Similarly, Barbouti et al.  reported that hematologic malignancies with i(17q) are characterized by adverse prognosis.
The most relevant finding in our study is the significant negative impact of major-route ACAs on the prognosis and response to treatment. Further, Verma et al.  reported that clonal evolution with chromosome 17, abnormalities and others have the worst outcome. They also added that the molecular events behind these abnormalities and potential therapeutic approaches directed at them need to be defined.
Moreover, Rashmi et al.  stated that Ph-negative myeloid neoplasms with isolated isochromosome 17q represent a clinicopathologic entity associated with a high risk for leukemic transformation.
More recently, Liu et al.  concluded that the genetic or functional inactivation of the p53 pathway plays an important role with regard to disease progression from the CP to BP and imatinib treatment response in CML. Finally, no correlation was found between i(17q) and known standard prognostic factors for CML. This was consistent with the findings of Seifert et al.  , who stated that p53 deletion due to loss of chromosome 17 short arm stands as an independent negative prognostic factor for disease-free survival, relapse risk, and overall survival in AML.
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Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2]
[Table 1], [Table 2], [Table 3], [Table 4]