1. Molecular Biology and Biotechnology Department, Human Genetics Div., Chromosomes lab., Atomic Energy Commission of Syria, P.O. Box 6091, Damascus, Syria
2. Jena University Hospital, Institute of Human Genetics, Am Klinikum 1, D-07747 Jena, Germany
3. Molecular Biology and Biotechnology Department, Mammalians Biology Div., Flow-cytometry Lab., Atomic Energy Commission of Syria, P.O. Box 6091, Damascus, Syria
Academic Editor: Joep Geraedts
Special Issue: Applications of Fluorescence in Situ Hybridization
Received: August 14, 2018 | Accepted: May 31, 2019 | Published: June 20, 2019
OBM Genetics 2019, Volume 3, Issue 2, doi:10.21926/obm.genet.1902081
Recommended citation: AL-ACHKAR W, As’sad M, Liehr T, Ikhtiar A, Wafa A. A De Novo Childhood Case of T-cell Lymphoblastic Leukemia with High Hyperdiploid Karyotype Carrying an Unreported Balanced Translocation t(X;5)(q26;q31.3~32) in A Male Patient. OBM Genetics 2019;3(2):9; doi:10.21926/obm.genet.1902081.
© 2019 by the authors. This is an open access article distributed under the conditions of the Creative Commons by Attribution License, which permits unrestricted use, distribution, and reproduction in any medium or format, provided the original work is correctly cited.
T-cell acute lymphoblastic leukemia (T-ALL) is a highly aggressive malignancy derived from T-cells and is known to present multiple chromosomal abnormalities. It is observed in children as well as in adults as 15% and 25% of the new diagnosed ALL cases, respectively. Interestingly, a 3-fold higher incidence of T-ALL has been observed in males. Compared to the precursor B-ALL, T-ALL is often associated with unfavorable clinical outcomes, such as high white blood cell (WBC) count, central nervous system infiltration, and bulky adenopathy [1,2]. The current intensive therapy that may be combined with hematopoietic stem cell transplantation improves the outcome at a rate of nearly 10% in children with T-ALL [3,4,5,6] but 37% of adult T-ALL patients show relapse, leading to death in due course . Thus, in adult cases, the approach is not much promising.
Structural and/or numerical chromosomal aberrations with prognostic significance are observed in nearly 50% of ALL cases [8,9,10]. High hyperdiploidy (HeH) (51-65 chromosomes) is an established subtype of B-cell (B-ALL) and associated with good survival and excellent outcome [10,11,12,13]. HeH has been seen in 40% of children (young, 1-4 years) with B-ALL, which decreases with age to a frequency of 10% among adults (25-59 years) . Furthermore, the chromosomes acquired most commonly are +4, +6, +10, +14, +17, +18, +21, and +X . However, neither the mechanism for chromosomal gains in ALL nor their roles in leukemogenesis are yet fully understood [15,16].
Most frequently observable chromosomal abnormalities in T-ALL are deletions del (6) (q) and balanced translocation t(10;14). An involvement of the T-cell receptor (TCR) gene, localized in 14q11, in rearrangements such as translocations t(1;14) (p31;q11), t(10;14)(q24;q11), or t(8;14)(q24;q11) are frequently reported. Terminal deletions like del (9) (p21) and del (1) (p32) have also often been repeated [11,17,18,19,20]. Better survival rates have been observed in childhood T-ALL with normal karyotype or with translocation t (10; 11) (q24; q11.2), than in cases with the presence of any derivative chromosome . Nonetheless, cryptic chromosomal aberrations are still a big challenge in the cytogenetic characterization of T-ALL and the most important step for accurate diagnosis is karyotyping. Additionally, new insights into T-ALL pathogenesis may also be derived from new molecular (cyto) genetic data.
Here, we first time report a new case of cortical-T-ALL in a male patient with HeH karyotype including a balanced translocation t (X;5).
2. Materials and Methods
2.1 Case Report
A 19-year-old male patient without any family history of malignancies presented with a one-month history of fatigue, weight loss, fever, and pleural effusion in his left lung. On physical examination, hepatomegaly and several skin nodules (1-1.5 cm) in different locations such as neck and armpit were diagnosed (data not shown). Initial complete blood count revealed WBC at 74.2 x 109/L (62% lymphocytes and 10% of cells were blasts), red blood cells (RBC) 2.48 x106/mm3, hemoglobin (Hgb) level of 7.6 g/dL, and platelet count of 171x109/L. Lymphobalsts percentage in BM were = 80% of all cells in BM. Biochemical analyses revealed the serum lactate dehydrogenase (LDH) value of 766 U/L (normal value is nearly 480 U/L), and serum alkaline phosphates value of 390 U/L (normal value nearly 128 U/L). The patient received chemotherapy treatment after cytogenetic analysis. He relapsed twice during DHAP/ABVD treatment (ABVD included doxorubicin, bleomycin, vinblastine, dacarbazine; DHAP included: dexamethasone, cytarabine, and cisplatin) and his hematological parameters were WBC 3.9x109/L, RBC 3.22x106/mm3, Hgb level of 10.6 g/dL, platelet count of 84x109/L, and LDH value of 642. He additionally suffered from fatigue, facial seventh cranial nerve palsy, and pain in the lower extremities.
2.2 Chromosomal Analysis
The GTG-banding analysis, according to standard procedures , was done prior to chemotherapy. At least 20 metaphases derived from unstimulated and cultivated bone marrow cells were studied. Karyotype was described using the International System for Human Cytogenetic Nomenclature (ISCN 2016) .
2.3 Molecular Cytogenetics
Fluorescence in situ hybridization (FISH) using whole chromosome painting (WCP) probes for chromosomes 5, 8, 11, 13, 19, 20, and X (MetaSystems, Altlussheim, Germany) was carried out following the manufacturer's instructions . Array-proven multicolor banding (aMCB) probes based on microdissection derived from region-specific libraries for chromosomes 5 and X were performed as described earlier . A minimum of 10 metaphase spreads were studied, using a fluorescence microscope (AxioImager.Z1 mot, Carl Zeiss Ltd., Hertfordshire, UK) and appropriate filters to discriminate between a maximum of five fluorochromes plus the counterstain DAPI (4',6- diamino–2-phenylindole). Image capture and FISH processing were performed using an ISIS imaging system (MetaSystems, Altlussheim, Germany).
2.4 Flow Cytometric Immunophenotyping
Immunophenotyping using a general panel of fluorescent antibodies against the following antigens distinctive for different cell lineages and cell types was performed: CD1a, CD2, , CD4, CD3, CD5, CD8, CD10, CD11c, CD11b, CD13, CD14, CD16, CD15, CD19, CD20, CD22, CD23, CD32, CD33, CD34, CD38, CD41a, CD45, CD56, CD57, CD64, CD103, CD117, CD123, CD138, CD209, CD235a, and CD243. In addition to Kappa and Lambda light chain antibodies, IgD, IgM, and HLADr were tested. All antibodies were purchased from BD Biosciences. Samples were scored using a BD FACSCalibur™ flow cytometer. Autofluorescence, isotype controls, and viability were included. Data acquisition and Flow cytometric analysis were conducted by BD Cellquest™ Pro software.
Figure 1 GTG-banding result shows a high hyperdiploidy karyotype with a translocation 51,XY,t (X;5)(q26;q31.3~32),+8,+11,+13,+19,+20. Derivative chromosomes are indicated by arrowheads.
Dual-color-FISH (D-FISH) using specific WCP probes for chromosomes 5, 8, 11, 13, 19, 20, and X confirmed the gain of chromosomes +8, +11, +13, +19, +20, and translocation t(5;X) (data not shown). aMCB (Figure 2) together with WCP data revealed a final karyotype 51,XY,t (X;5)(q26;q31.3~32),+8,+11,+13,+19,+20 .
Figure 2 Karyotype and chromosomal aberrations were confirmed using molecular cytogenetic approaches. aMCB results are shown here. The normal chromosomes (if present) are mentioned in the second column and their derivatives on the right side of normal chromosomes. The unstained regions, when using chromosome-specific aMCB-probesets on the derivative chromosomes, are shown in gray. Abbreviations: der = derivative chromosome.
As per the WHO classification, the immunophenotyping of bone marrow specimen characterized this case as Cortical-T-ALL. The blast cell population was positive for CD45dim, CD10, TdT, CD7, CD1a, CD2, CD4, CD8, CD5dim, and cCD3. This cell population was negative for CD19, CD34, HLA-DR, CD33, TCR α/β, TCR γ/δ, and sCD3.
A de novo childhood cortical-T-ALL male patient with HEH karyotype including a balanced translocation t (X;5)(q26;q31.3~q32) was characterized in detail by (molecular) cytogenetics. According to the literature, HeH (>51 chromosomes) is commonly found in B-ALL patients [10–13] and is an uncommon event in T-ALL. To date, approximately 117 ALL cases with HeH have been reported in the Mitelman Database . Trisomy for chromosome 8 has been reported in 36 ALL with HeH cases (see ). Also, trisomy of chromosomes 11, 13, 19, and 20 was reported previously in 5, 9, 5 and 11 ALL cases, respectively with HeH . In addition, chromosomal bands such as Xq26 and 5q31 were reported to be involved in chromosomal rearrangements in 16 and 109 cases, respectively . However, translocation t(X;?) including short and/or long arms of these chromosomes has been seen previously only in two B-ALL cases with HeH . Interestingly, while all such reported ALL cases involving Xq26 (16 cases) were in females, the present case was observed in a male patient; the majority of these cases were of B-ALL (7 out 16 cases), four of these cases were of T-ALL, and the remaining cases were unclassified. To the best of our knowledge, this is the first report to observe HeH with translocation t(X;5)(q26;q31.3~q32) in a male T-ALL patient .
As a specific cytogenetic subgroup of ALL, HeH has a reported frequency of 5–10% [25,26]. The pattern observed is nonrandom; however, chromosomes X, 4, 6, 10, 14, 17, 18 and 21 are found to be the most frequently gained ones [11,14]. Thus, this subgroup has a cytogenetic profile very similar to that observed in pediatric ALL [14,27].
Univariate analysis revealed moderately strong evidence of heterogeneity in patient’s outcome with respect to the gain of the chromosomes 4, 5, 10, 14, 17, 20, 21, and X . Interestingly, all chromosome acquisitions, apart from chromosome 20, have prognostic significance in pediatric ALL [14,27,28]. On the other hand, adverse prognostic effects even in the presence of extra chromosome 20 in our case is a novel finding. In HeH, the gain of chromosome 20 is less frequently seen in children (only in 6%), thus, an adverse effect may have been hidden. However, in an adult cohort and in the case of the adolescent reported here, +20 had an extremely poor outcome, which was mainly caused by early relapse .
Interestingly, in Ewing’s Sarcoma as well, +20 has been associated with adverse outcomes, where it is present along with the gain of other chromosomes . Overall, the mechanism and consequences for tumor development in terms of simultaneous gain of multiple chromosomes is not yet understood [15-16]. The overexpression of one or more genes on each of the gained chromosomes may potentially be the cause of the disease . This idea is supported by the fact that the accumulation of methotrexate polyglutamate is higher in blast cells of HeH patients when compared to that in non-HeH patients  and may thus explain the favorable outcome of pediatric HeH patients .
Somatic deletions and other inactivating mutations of the plant homeodomain (PHD)-like finger 6 (PHF6) gene in Xq26.2 has been identified in 16% (14 of 89) of pediatric and in 38% (16 of 42) of adult T-ALL samples. Interestingly, most of them were derived from males . Immunohistochemical analysis showed a complete loss of the PHF6 protein in all T-ALLs with a mutation. Loss of PHF6 was observed in leukemia driven by aberrant expression of the homeobox transcription factor oncogenes TLX1 and TLX3. Altogether this indicates PHF6 as an X-linked tumor suppressor in T-ALL. Also, Kraszewska et al.  proposed that PHF6 could be a tumor suppressor gene and that mutations result in loss of its function. This would be consistent with silencing of the PHF6 promoter by DNA methylation in some T-ALL cases.
Overall, prognostic factors have been clearer for patients with B-ALL than for patients with T-ALL. HeH is an established genetic subtype of B-ALL, which is associated with good survival and excellent outcome [10,11,12,13]. Adverse prognosis in T-ALL is accompanied with hyperleukocytosis, enhanced mediastinal mass, male gender, central neural system involvement, and advanced age . Also, T-ALL patients who were CD1a positive in the trial of Medical Research Council (MRC)/Eastern Cooperative Oncology Group (ECOG) had an overall five years survival of 64% (95% CI, 48-80%), which was better than that for CD1a-negative patients [39%(26-52%) in (P=0.01)]. This may possibly be caused by a higher risk of relapse in CD1a-negative patients (50%; 36-65%) in five years, compared to that (23%; 8–38%) in CD1a-positive patients (P = 0.02) .
In conclusion, we presented here a de novo case of childhood cortical-T-ALL in a male patient with HeH karyotype, which included a balanced t(X;5)(q26;q31.3~q32), trisomy 20 and CD1a positivity. These may be indicators for poor prognosis in T-ALL with adverse effects and relapse during chemotherapy.
We thank Prof. I. Othman, the Director General of Atomic Energy Commission of SYRIA (AECS) and Dr. N. Mirali, Head of Molecular Biology and Biotechnology Department for their support. This work was supported by the AECS.
AW, MA and WA-A did (molecular) cytogenetic analyses and drafted the paper, AW provided the clinical case and collected all clinical data, AA did the immunophenotyping, TL did final evaluation of (molecular) cytogenetic analyses and finalized the paper in agreement with all authors.
The authors have declared that no competing interests exist.
- Uckun FM, Gaynon PS, Sensel MG, Nachman J, Trigg ME, Steinherz PG, et al. Clinical features and treatment outcome of childhood T-lineage acute lymphoblastic leukemia according to the apparent maturational stage of T-lineage leukemic blasts: A Children’s Cancer Group study. J Clin Oncol. 1997; 15: 2214-2221. [CrossRef]
- Goldberg JM, Silverman LB, Levy DE, Dalton VK, Gelber RD, Lehmann L, et al. Childhood T-cell acute lymphoblastic leukemia: The Dana-Farber Cancer Institute acute lymphoblastic leukemia consortium experience. J Clin Oncol. 2003; 21: 3616-3622. [CrossRef]
- Kox C, Zimmermann M, Stanulla M, Leible S, Schrappe M, Ludwig WD, et al. The favorable effect of activating NOTCH1 receptor mutations on long-term outcome in T-ALL patients treated on the ALL-BFM 2000 protocol can be separated from FBXW7 loss of function. Leukemia. 2010; 24: 2005-2013. [CrossRef]
- Breit S, Stanulla M, Flohr T, Schrappe M, Ludwig WD, Tolle G, et al. Activating NOTCH1 mutations predict favorable early treatment response and long-term outcome in childhood precursor T-cell lymphoblastic leukemia. Blood. 2006; 108: 1151-1157. [CrossRef]
- Clappier E, Collette S, Grardel N, Girard S, Suarez L, Brunie G, et al. NOTCH1 an FBXW7 mutations have a favorable impact on early response to treatment, but not on outcome, in children with T-cell acute lymphoblastic leukemia (T-ALL) treated on EORTC trials 58881 and 58951. Leukemia. 2010; 24: 2023-2031. [CrossRef]
- Zuurbier L, Homminga I, Calvert V, te Winkel ML, Buijs-Gladdines JG, Kooi C, et al. NOTCH1 and/or FBXW7 mutations predict for initial good prednisone response but not for improved outcome in pediatric T-cell acute lymphoblastic leukemia patients treated on DCOG or COALL protocols. Leukemia. 2010; 24: 2014-2022. [CrossRef]
- Marks DI, Paietta EM, Moorman AV, Richards SM, Buck G, DeWald G, et al. T-cell acute lymphoblastic leukemia in adults: clinical features, immunophenotype, cytogenetics, and outcome from the large randomized prospective trial (UKALL XII/ECOG 2993). Blood. 2009; 114: 5136-5145. [CrossRef]
- Teitell MA, Pandolfi PP. Molecular genetics of acute lymphoblastic leukemia. Annu Rev Pathol. 2009; 4: 175-198. [CrossRef]
- Mrózek K, Harper DP, Aplan PD. Cytogenetics and molecular genetics of acute lymphoblastic leukemia. Hematol Oncol Clin North Am. 2009; 23: 991-1010. [CrossRef]
- Moorman AV, Richards SM, Martineau M, Cheung KL, Robinson HM, Jalali GR, et al. Outcome heterogeneity in childhood high-hyperdiploid acute lymphoblastic leukemia. Blood. 2003: 102: 2756-2762. [CrossRef]
- Chilton L, Buck G, Harrison CJ, Ketterling RP, Rowe JM, Tallman MS, et al. High hyperdiploidy among adolescents and adults with acute lymphoblastic leukaemia (ALL): Cytogenetic features, clinical characteristics and outcome. Leukemia. 2014; 28: 1511-1518. [CrossRef]
- Paulsson K, Johansson B. High hyperdiploid childhood acute lymphoblastic leukemia. Genes Chromosomes Cancer. 2009; 48: 637-660. [CrossRef]
- Moorman AV. The clinical relevance of chromosomal and genomic abnormalities in B-cell precursor acute lymphoblastic leukaemia. Blood Rev. 2012; 26: 123-135. [CrossRef]
- Paulsson K, Forestier E, Andersen MK, Autio K, Barbany G, Borgström G, et al. High modal number and triple trisomies are highly correlated favorable factors in childhood B-cell precursor high hyperdiploid acute lymphoblastic leukemia treated according to the NOPHO ALL 1992/2000 protocols. Haematologica. 2013; 98: 1424-1432. [CrossRef]
- Paulsson K, Panagopoulos I, Knuutila S, Jee KJ, Garwicz S, Fioretos T, et al. Formation of trisomies and their parental origin in hyperdiploid childhood acute lymphoblastic leukemia. Blood. 2003; 102: 3010-3015. [CrossRef]
- Gruszka-Westwood AM, Horsley SW, Martinez-Ramirez A, Harrison CJ, Kempski H, Moorman AV, et al. Comparative expressed sequence hybridization studies of high-hyperdiploid childhood acute lymphoblastic leukemia. Genes Chromosomes Cancer. 2004; 41: 191-202. [CrossRef]
- Kebriaei P, Anastasi J, Larson RA: Acute lymphoblastic leukaemia: Diagnosis and classification. Best Pract Res Clin Haematol. 2002; 15: 597-621. [CrossRef]
- Cauwelier B, Dastugue N, Cools J, Poppe B, Herens C, De Paepe A, et al. Molecular cytogenetic study of 126 unselected T-ALL cases reveals high incidence of TCRbeta locus rearrangements and putative new T-cell oncogenes. Leukemia. 2006; 20: 1238-1244. [CrossRef]
- Inaba H, Greaves M, Mullighan CG. Acute lymphoblastic leukaemia. Lancet. 2013; 381: 1943-1955. [CrossRef]
- Schneider NR, Carroll AJ, Shuster JJ, Pullen DJ, Link MP, Borowitz MJ, et al. New recurring cytogenetic abnormalities and association of blast cell karyotypes with prognosis in childhood T-cell acute lymphoblastic leukemia: A pediatric oncology group report of 343 cases. Blood. 2000; 96: 2543-2549.
- AL-achkar W, Wafa A, Nweder MS. A complex translocation t(5;9;22) in Philadelphia cells involving the short arm of chromosome 5 in a case of chronic myelogenous leukemia. J Exp Clin Cancer Res. 2007; 26: 411-415.
- McGowan-Joran J, Simons A, Schmid MS. ISCN 2016: An International System of Human Cytogenomic Nomenclature. Unionville, CT, USA: S. Karger Publications, Inc; 2016. [CrossRef]
- Liehr T, Heller A, Starke H, Rubtsov N, Trifonov V, Mrasek K, et al. Microdissection based high resolution multicolor banding for all 24 human chromosomes. Int J Mol Med. 2002; 9: 335-339. [CrossRef]
- Mitelman F, Johansson B, Mertens F, editors: Mitelman database of chromosome aberrations and gene fusions in cancer (2016). http://cgap.nci.nih.gov/Chromosomes/Mitelman [accessed 26.07.2018].
- Mi JQ, Wang X, Yao Y, Lu HJ, Jiang XX, Zhou JF, et al. Newly diagnosed acute lymphoblastic leukemia in China (II): Prognosis related to genetic abnormalities in a series of 1091 cases. Leukemia. 2012; 26: 1507-1516. [CrossRef]
- Gomez-Segui I, Cervera J, Such E, Martinez-Cuadron D, Luna I, Ibanez M, et al. Prognostic value of cytogenetics in adult patients with Philadelphia-negative acute lymphoblastic leukemia. Ann Hematol. 2012; 91: 19-25. [CrossRef]
- Dastugue N, Suciu S, Plat G, Speleman F, Cave H, Girard S, et al. Hyperdiploidy with 58-66 chromosomes in childhood B-acute lymphoblastic leukemia is highly curable: 58951 CLG-EORTC results. Blood. 2013; 121: 2415-2423. [CrossRef]
- Heerema NA, Raimondi SC, Anderson JR, Biegel J, Camitta BM, Cooley LD, et al. Specific extra chromosomes occur in a modal number dependent pattern in pediatric acute lymphoblastic leukemia. Genes Chromosomes Cancer. 2007; 46: 684-693. [CrossRef]
- Roberts P, Burchill SA, Brownhill S, Cullinane CJ, Johnston C, Griffiths MJ, et al. Ploidy and karyotype complexity are powerful prognostic indicators in the Ewing’s sarcoma family of tumors: A study by the United Kingdom Cancer Cytogenetics and the Children’s Cancer and Leukaemia Group. Genes Chromosomes Cancer. 2008; 47: 207-220. [CrossRef]
- Whitehead VM, Vuchich MJ, Lauer SJ, Mahoney D, Carroll AJ, Shuster JJ, et al. Accumulation of high levels of methotrexate polyglutamates in lymphoblasts from children with hyperdiploid (>50 chromosomes) B-lineage acute lymphoblastic leukemia: A Pediatric Oncology Group study. Blood. 1992; 80: 1316-1323.
- Van Vlierberghe P, Palomero T, Khiabanian H, Van der Meulen J, Castillo M, Van Roy N, et al. PHF6 mutations in T-cell acute lymphoblastic leukemia. Nat Genet. 2010; 42: 338-342. [CrossRef]
- Kraszewska MD, Dawidowska M, Larmonie NS, Kosmalska M, Sedek L, Szczepaniak M, et al. DNA methylation pattern is altered in childhood t-cell acute lymphoblastic leukemia patients as compared with normal thymic subsets: Insights into cpg island methylator phenotype in t-all. Leukemia. 2012; 26: 367-371. [CrossRef]