Chromosomal aberrations in Bloom syndrome patients with myeloid malignancies

Cancer Genetics and Cytogenetics 128 (2001) 39–42

Short communication

Chromosomal aberrations in Bloom syndrome patients with myeloid malignancies Bruce Poppea, Heidi Van Limbergena, Nadine Van Roya, Els Vandecruysb, Anne De Paepea, Yves Benoitb, Frank Spelemana,* a Center for Medical Genetics, De Pintelaan 185, B-9000 Gent, Belgium Pediatric Hematology and Oncology, Ghent University Hospital, De Pintelaan 185, B-9000 Gent, Belgium Received 25 September 2000; accepted 13 December 2000

b

Abstract

Bloom syndrome (BS) predisposes affected individuals to a wide variety of neoplasms including hematological malignancies. Thus far, cytogenetic findings in hematological neoplasms have been reported in only a few BS patients. We present the karyotypic findings in a BS patient diagnosed with acute myeloid leukemia (AML), FAB subtype M1, and a review of the literature, showing the preferential occurrence of total or partial loss of chromosome 7 in BS patients with AML or myelodysplastic syndromes (MDS). © 2001 Elsevier Science Inc. All rights reserved.

1. Introduction Bloom syndrome (BS) (MIM Number: 210900 [1]) is a rare, autosomal recessive, genetic disorder, clinically characterized by proportional dwarfism, sun-sensitive erythematous skin lesions, a characteristics facies and head configuration and immunodeficiency (for review, see ref. [2]). Affected individuals are predisposed to a wide variety of neoplasms. Hematological malignancies are frequent in BS patients and have occurred in 40 out of 168 individuals registered in the Bloom’s Syndrome Registry, accounting for 44 of the first 100 cancers diagnosed [3]. Although the number of previously published cases was small, it was assumed thus far that the chromosomal rearrangements occurring in leukemias which develop in BS patients are similar in type and incidence than those observed in other patients [4]. However, the present report on the cytogenetic findings in a BS patient with AML and a review of the literature suggest a preferential occurrence of monosomy 7 (7) and deletions of the long arm of chromosome 7 [7q, del(7q)] in BS patients with myeloid neoplasms. 2. Case report MS, an 8-year-old male BS patient born from consanguineous parents, was admitted to the Department of Pediat-

* Corresponding author. Tel: +32-9-240-24-51; fax: +32-9-240-49-70. E-mail address: [email protected] (F. Speleman).

ric Hematology and Oncology for leukemia. He, as well as one of his sisters, are listed in the BS Registry as 165(MuSaf) and 149(SeSaf), respectively (J. L. German, personal communication). Some weeks before admittance, there had been an episode of pain in both legs. Clinical examination did not reveal any abnormalities, clinical features of BS left aside. Peripheral blood analysis showed normochromic, normocytic anemia, normal leucocyte count, with blasts (47%) and lymphocytes (40%) but no trombocytopenia. Morphological examination of the bone marrow aspirate revealed a rich marrow with 84.3% myeloblasts, 10% of which were monoblasts, aplasia of the erythroid lineage and normal megakaryopoiesis. Twelve percent of the blasts were peroxidase-positive. An acute myeloid leukemia, subtype M1, was diagnosed [5]. Karyotypic analysis was performed on bone marrow cells and revealed the presence of clonal rearrangements (see subsequent text). Radiological examinations were normal, in particular no osteolytic lesions, no splenomegaly, no lymphadenopathy or infectious foci could be demonstrated. Induction therapy with an appropriate dose reduction was initiated: AraC: 50 mg/m2, twice a day for 4 days, VP16 50 mg/m2 once a day for 4 days, thioguanine 50 mg/m2 once a day for 5 days, and AraC: 10 mg intrathecally. The supportive treatment was comprehensive (transfusions, immunoglobulins, total parenteral nutrition, intestinal decontamination and antibiotics). The leukemic cells in the blood disappeared very rapidly but during the bone marrow reconstitution blasts reappeared in peripheral blood (12%) and bone marrow (9%

0165-4608/01/$ – see front matter © 2001 Elsevier Science Inc. All rights reserved. PII: S0165-4608(01)00 3 9 2 - 2

40

B. Poppe et al. / Cancer Genetics and Cytogenetics 128 (2001) 39–42

blasts). A second but higher dosed induction was started and was followed by a complete remission. Because he had a healthy HLA identical sister, bone marrow transplantation was started. The same drugs as used in the reinduction were used for the conditioning-regimen. Graft versus host prophylaxis consisted of ciclosporin only. After a period of prolonged cytopenia, however, the bone marrow recovery was autologous. Still in remission, a maintenance therapy with low dose thioguanine and low dose AraC was started. Six months after transplant he is still in treatment and in first remission.

3. Materials and methods Short-term (24 h) in vitro culture of bone marrow cells, harvesting and staining (Wright stain) was performed according to standard procedures. The constitutional karyotype was performed on metaphases obtained from PHA stimulated and methotrexate synchronized peripheral blood lymphocytes. Karyotypic findings were described according to the International System for Chromosome Nomenclature [6]. FISH was performed as previously described [7]. The following probes were selected for FISH: LSI D7S522/CEP 7 and LSI CBFB (Vysis, Downers Grove, IL, USA). Total RNA was isolated from frozen bone marrow cells using the Trizol reagent (GIBCO BRL, Grand Island, NY, USA). The cDNA synthesis and RT-PCR for the detection of the CBFB-MYH11 fusion transcript were performed as previously described [8].

4. Results Cytogenetic analysis was performed on bone marrow samples taken at diagnosis (day 0) and at days 1, 5, 15 and after 5 months. In the first three samples, the same clonal rearrangements were observed in all metaphases analyzed: 46,XY,del(7)(q21.2),inv(16)(p13q21–22) (Fig. 1). Karyotypic analysis of the last two bone marrow samples did not reveal clonal aberrations. The constitutional karyotype was normal except for the typical increased frequency of quadriradial figures and chromosomal breaks. Partial loss of chromosome 7 long arm was confirmed by FISH, whereas the chromosome 16 inversion did not result in a rearrangement of the CBFB locus. In agreement with this result, a chimeric CBFB-MYH11 fusion transcript could not be detected by RT-PCR.

5. Discussion Recently, the genetic defect causative in BS has been identified. Homozygosity or compound heterozygosity for mutations in the BLM gene causes somatic cells from affected individuals to accumulate excessive numbers of mutations [9]. This leads to a characteristic clinical picture including a high predisposition to the development of cancer. In contrast to the new insights in the structure and function of the causative gene for BS, little is known about the genetic abnormalities, which characterize the various malignancies occurring in these patients. The question remained whether these genetic changes resemble the non-random

Fig. 1. Partial G-banded karyotype of unstimulated bone marrow showing a del(7)(q21.2) and a inv(16)(p13q21q22) (arrow heads).

B. Poppe et al. / Cancer Genetics and Cytogenetics 128 (2001) 39–42

changes which are known to be associated with certain neoplasms or whether a specific subset of known or previously unsuspected non-random genetic aberrations preferentially occur in these patients. In a BS patient, diagnosed with AML, FAB M1 subtype, we observed a del(7)(q21.2) and an inv(16)(p13q21q22) in unstimulated bone marrow cells. Both FISH and PCR analyses showed that the chromosomal aberration was different from the classical inv(16). A review of the literature revealed that cytogenetic aberrations in BS patients with leukemia had been published on four occasions [10–13]. Karyotypes of four further cases have been reported partially [4] (Table 1). These patients suffered from acute leukemia (AL) or MDS, in 7 out of 8 cases de novo. Strikingly, complete or partial loss of chromosome 7 is a consistent finding in all but two of the BS patients with these hematological malignancies and in 4 out of 5 BS patients with myeloid malignancies. Thus, the present observation together with the literature data suggests that complete or partial loss of chromosome 7 occurs at a much higher frequency in BS patients than expected as compared to de novo leukemias occurring in patients without BS or other cancer predisposing syndromes, in which 7/7q occur in less than 5% [14]. Interestingly, 7/7q is also more frequently observed in other cancer predisposing constitutional disorders (including Fanconi’s anemia, neurofibromatosis type 1, congenital neutropenia, Schwachman-Diamond and Down syndrome) and in AML/MDS (t-AML/t-MDS) following therapy with alkylating agents or secondary AML/MDS after occupational or environmental exposure to chemical mutagens (for review see ref. [15]). Thus, although also present in de novo leukemias, 7/7q seems particularly frequent in specific patient groups which are either predisposed to cancer or who have been exposed to genotoxic agents. A growing body of evidence implicates genetic instabil-

41

ity as one of the key steps in the multi-step process of cancer [16]. The mechanisms by which acquired and inherited genetic instability provoke or contribute to malignant transformation are poorly understood. However, the nonrandom association between genetic instability and chromosome 7 loss in leukemia prompts the hypothesis that genetic instability preferentially induces total or partial chromosome 7 loss. The mechanism by which total and partial chromosome 7 loss are induced in acquired and inherited genetic instability might even prove to be quite similar. In conclusion, we have shown that chromosome 7 sequences are preferentially lost in the leukemic cells of BS patients with myeloid neoplasms. Further data on leukemias in additional BS patients are needed to further establish a particular role for 7/7q in the biology of leukemias in these patients. Our observations add another constitutional syndrome to those that have already been associated with loss of chromosome 7, and confirms the association between 7/7q and genetic instability. Somatic hypermutability in BS patients might engage the same pathway(s) which is (are) involved in loss of chromosome 7 material in patients which have previously been exposed to genotoxic agents. Acknowledgments This work is supported by the Fund for Scientific Research-Flanders (FWO-Vlaanderen), grant nr. G.0028.00 and by the research fund of the University of Ghent (BOF, grant nr. 011D7699). BP is an Aspirant of the Fund for Scientific Research-Flanders (FWO-Vlaanderen). HVL is the recipient of a research grant of the University of Ghent. NVR is a postdoctoral researcher of the Fund for Scientific Research-Flanders (FWO-Vlaanderen). The authors thank Dr. J. L. German III for helpful discussions and for the communication of preliminary research results.

Table 1 Karyotypic aberrations in BS patients with acute leukemia or myelodysplastic syndromes Case

Rega

Sex

Age at diagnosis

FAB subtype

Karyotype

Ref.

1 2 3

41 103 132

M M M

39 7 1/2 34

BIP ALL L2 RAEBt

[10] [11] [12]

4 5 6

— 17 32

M M M

7 32 22

7 8 9

62 51 165

F M M

9 1/2 14 13 8

RAEBt AML M6 tMDS/AML nullALL AML M2/MDS AUL AML M1

44,X,del(3)(q26)?,t(4;22)(q21;q11),der(17)t(7;17)(p11;q11),del(8)(q22),del(20q) 4345,XY,?7,?9,12 marb 46,XY,7,der(13)t(1;13)(q11;p11),mar/ 46,XY,del(5)(q22q33),del(7)(q11q22),der(11)t(11;11) (p15;q13),der(13) 45,XY,7[60%]/45,X,Y[15%]/44,X,Y,7[15%]/46,XY[10%] 4,5,7, several rearrangements affecting 4,5,12 and 22 7, other

a

Hyperdiploid 46,XX,17,20,21,22,14p,15p,15p,der(1)t(1;17)(p11;q11),3mar Hypodiploid clone with monosomy 21 and duplication of a translocation between 1q and 8q 46,XY,del(7)(q21.2),inv(16)(p13q2122)

Case identification as in the Bloom Syndrome Registry. One of the markers probably being an i(7)(q10). Chromosomal rearrangements leading to total or partial chromosome 7 loss in bold. CR: current report Ref.: references b

[13] [4] [4]

[4] [4] CR

42

B. Poppe et al. / Cancer Genetics and Cytogenetics 128 (2001) 39–42

References [1] Online Mendelian Inheritance in Man, OMIM (TM). Johns Hopkins University, Baltimore (MD). MIM Number: 210900, Date last edited: 01/08/2000. Available at: http:\\www.ncbi.nlm.nih.gov\omim. [2] German J, Ellis NA. Bloom syndrome. In: Vogelstein B, Kinzler KW, editors. The genetic basis of human cancer. New York: McGraw-Hill, 1998. p. 301–15. [3] German J. Bloom’s syndrome. XX. The first 100 cancers. Cancer Genet Cytogenet 1997;93:100–6. [4] German J. Bloom’s syndrome: incidence, age of onset, and types of leukemia in the Bloom’s syndrome registry. In: Bartsocas CS, Loukopoulos D, editors. Genetics of hematological disorders. Washington: Hemisphere Publications, 1992. p. 241–58. [5] Bennett JM, Catovsky D, Daniel MT, Flandrin G, Galton DA, Gralnick HR, Sultan C. Proposed revised criteria for the classification of acute myeloid leukemia. A report of the French-American-British Cooperative Group. Ann Intern Med 1985;103:620–5. [6] ISCN. An international system for human cytogenetic nomenclature. Mitelman F, editor. Basel: S. Karger, 1995. [7] Van Roy N, Laureys G, Cheng NC, Willem P, Opdenaker G, Versteeg R, Speleman F. 1;17 Translocations and other chromosome 17 rearrangements in human primary neuroblastoma tumors and cell lines. Genes Chromosomes Cancer 1994;10:103–14. [8] van der Reijden BA, Lombardo M, Dauwerse HG, Giles RH, Mühlematter D, Bellomo MJ, Wessels HW, Beverstock GC, van Ommen GJB, Hagemijer A, Breuning MH. RT-PCR diagnosis of patients with acute nolymphocytic leukemia and inv(16)(p13q22) and identi-

[9]

[10]

[11]

[12]

[13]

[14]

[15]

[16]

fication of new alternative splicing in CBFB-MYH11 transcripts. Blood 1995;86:277–82. Ellis N, Groden J, Ye TZ, Straughen J, Lennon DN, Ciocci S, Proytcheva M, German J. The Bloom’s syndrome gene product is homologous to RecQ helicases. Cell 1995;83:655–66. Shabtai F, Lewinski UH, Meroz A, Dvora Klar, Djaldetti M, Halbrecht I. Non-random chromosomal aberrations in a complex leukaemic clone of a Bloom’s syndrome patient. Hum Genet 1988;80: 311–14. Werner-Favre C, Wyss M, Cabrol C, Félix F, Guenin R, Laufer D, Engel E. Cytogenetic study in a mentally retarded child with Bloom Syndrome and acute lymphoblastic leukemia. Am J Med Genet 1983;18:215–21. Iwahara Y, Ishii K, Watanabe S, Taguchi H, Hara H, Miyoshi I. Bloom’s syndrome complicated by myelodysplastic syndrome and multiple neoplasia. Intern Med 1993;32:399–402. Aktas D, Koc A, Boduroglu K, Hicsonmez G, Tuncbilek E. Myelodysplastic syndrome associated with monosomy 7 in a child with Bloomsyndrome. Cancer Genet Cytogenet 2000;1:44–6. Berger R, Flandrin G, Bernheim A, Le Coniat M, Vecchione D, Pacot A, Derre J, Daniel MT, Valensi F, Sigaux F, Ochoa-Noguera ME. Cytogenetic studies on 519 consecutive de novo acute nonlymphocytic leukemias. Cancer Genet Cytogenet 1987;29:9–21. Luna-Fineman S, Shannon KM, Lange BJ. Childhood monosomy 7: epidemiology, biology, and mechanistic implications. Blood 1995; 85:1985–99. Wright EG. Inherited and inducible chromosomal instability: a fragile bridge between genome integrity mechanisms and tumourigenesis. J Pathol 1999;187:19–27.