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Complex translocation involving Ph chromosome in a patient with typical chronic myelogenous leukemia
Complex Translocation Involving Ph Chromosome in a Patient With Typical Chronic Myelogenous Leukemia G. Calabrese, G. Palka, C. A. Westbrook, and D. Sheer
ABSTRACT:
We report a cytogenetic study of a patient with chronic myelogenous leukemia (CML) who, while displaying a Philadelphia (Ph) chromosome, resultingfrom a standard t(9;22J at diagnosis, during the chronic phase (CP) showed disappearance of the Ph and occurrence of new chromosome changes, including a marker probably arising from a translocation involving chromosome 17 and the Ph. In situ hybridization confirmed the cytogenetic appearance and demonstrated that the breakpoint on the Ph marker occurred below the BCR-ABL fusion gene
INTRODUCTION
More than 90% of patients with chronic myelogenous leukemia (CML) have a Philadelphia (Ph) chromosome in their bone marrow (BM) cells. This marker chromosome arises from a balanced translocation between chromosomes 9 and 22. As a consequence, the ABL protooncogene, normally located in band 9q34, moves to the BCR gene in band 22ql1, producing a BCR-ABL fusion gene believed to be involved in the pathogenesis of CML [l]. A few patients with typical clinical and hematologic signs of the disease show either no evidence of the Ph chromosome or a complex rearrangement involving chromosomes other than 9 and 22 [2-41. Each of these cases that has been subjected to molecular analysis has been shown to have the BCR-ABL fusion gene [5-91. We describe a patient with typical CML who developed a new translocation involving the Ph chromosome during the chronic phase (CP). CASE REPORT AND RESULTS
D. 0. G., a 50-year-old woman, was admitted to the division of hematology in November 1984 with complaints of asthenia, fever, pain below the left costal margin, and weight loss. Physical examination showed hepatosplenomegaly (3 and 6 cm, respectively, below the costal margins). Hematologic and biochemical investigations performed on peripheral blood (PB) showed a hemoglobin level 12 gidl, a red
From the Institute of Biology and Genetics, University ofchieti, Italy (G. C., G. P.), Department ofMedicine, Section ofHematology/ Oncology, University of Chicago Medical Center, Illinois (C. A. W.), and Human Cytogenetics Laboratory, Imperial Cancer Research Fund, London, England [D. S.J. Address reprint requests to: Giandomenico Palka, Via B. Buozzi 93, 65121 Pescara, Italy. Received December 27, 1991; accepted April 14, 1992.
blood cell (RBC) count of 4.5 x 1012/l, a white blood cell (WBC) count of 120 x log/L with 50% neutrophils, 2% eosinophils, 2% basophils, 12% myelocytes, 10% promyelocytes, 1% blasts, 3% monocytes, and 20% lymphocytes, and a platelet count of 600 x log/l; the leukocyte alkaline phosphatase score was 2. BM aspirate showed myeloid hyperplasia. A cytogenetic investigation performed on BM cells showed the karyotype 46,XX,t(9;22) (q34;qll) in 50 metaphases studied by GTG-banding. Induction treatment, started soon after diagnosis, comprised cytosine arabinoside and myleran. During CP, the patient was submitted to radiotherapy three times to reduce her splenomegaly. She remained in CP for 48 months, and all cytogenetic investigations performed on her BM cells in this period [every 6 months) showed the t(9;22)(q34;qll) as the only chromosome rearrangement. In November 1988, when the patient was still in clinical and hematologic CP, cytogenetic analysis showed the presence of a new clone containing a marker chromosome apparently derived from a translocation between the Ph and chromosome 17, with the breakpoints in the distal long arm of the Ph and at centromeric level of chromosome 17 (Fig. 1). No Ph was present in this clone, but other new aberrations observed were a t(5;14)(q35;q22), a de1(7)(p13), and monosomies 17 and 18. The 9q+ chromosome was also present in this clone. Fluorescence in situ hybridization [FISH) performed with a biotinylated 3’-ABL probe, c-H-abl probe [lo], and a cosmid probe, D178163, mapping to band 17qll-12 [ 1 l] demonstrated the presence of both probes on the new marker chromosome, confirming the involvement of chromosome 17 and indicating that the breakpoint on the Ph occurred below the BCR-ABL fusion gene (Fig. 2). Thus, according to the new Internation System for Human Cytogenetic Nomenclature [l2], the karyotype was defined as 44, XX,t(5;14)(q35;q22),de1(7)(p13),der(9)t(9;22)~q34;qll),-l7, - lB,der(22)t(9;22)(q34;qll)t(9;17)(q34;qlO). Twelve months later, BM blasts had increased by 30%.
52 Cancer Genet Cytogenet 6352-55 0165-4606/92/$05.00
(1992)
0 1992 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York. NY 10010
In Situ Detection of a Ph Complex Translocation
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Figure 1 Karyotype in GTG-banding showing monosomy 18, a t(5;14)(q35;q22), a del(7)(p11), a chromosome 9q +, and a translocation involving chromosomes 17 and Ph (M). Inset: Two other partial karyotypes from the latter rearrangement.
Cytogenetic analysis showed the karyotype described above in all cells. After a short time, the patient developed clinical signs of blast crisis (BC) and died 3 months later: a survival time of 63 months from diagnosis.
DISCUSSION
Variant Ph translocations in CML were initially described mainly in patients in BC of CML, but more recently have been detected at all stages of the disease. Cytogenetically they appear to be either of the simple type, usually involving chromosome 22 and a different chromosome, or of the complex type, involving chromosomes 9, 22, and a third or more chromosomes. The findings of ABL sequences on the
Ph chromosome and of the presence of a BCR-ABL fusion even in apparently simple variant translocations strongly suggest that all variant translocations involve both chromosomes 9 and 22 and are therefore of the complex type [5-9]. Moreover, they can arise as secondary translocations during evolution of the disease, involving the 9q + chromosome or more rarely the Ph previously generated from the standard t(9;22)(q34;q11) [4, 6-9, 13, 14]. At diagnosis, the patient with typical clinical and hematologic characteristics of CML had a standard t(9;22) (q34;q11). During CP, other cytogenetic changes occurred, including replacement of the Ph with a new marker that appeared to derive from a translocation between the Ph and chromosome 17. FISH showed the 3'ABL and D17S163 probes on the new marker, confirming the cytogenetic appearance of a "masked" Ph [6] and demonstrating that the
54
G. Calabrese et al.
A
B Figure 2 (A) Metaphase prebanded with Wright stain. Arrows show 9q + chromosome and the marker chromosome. (B) Probe signals from c-H-abl (long arrows) and D17S163 (short arrows) on the marker chromosome and normal chromosomes 9 and 17 after in situ hybridization on the same metaphase.
secondary breakpoint on the Ph was below the B C R - A B L fusion gene. This case and others s h o w in g secondary translocations i nvolv i n g the 9 q + or Ph c h r o m o s o m e s raise the question of w het h er specific sequences on these chromosomes are susceptible to preferential breakage during evolution of CML. A c o m b i n e d approach of molecular analysis and FISH with appropriate probes on c h r o m o s o m e s 9 and 22 might be informative in this regard. Presumably the leukemia cells carrying these translocations have a selective advantage over cells having only the standard t(9;22)(q34;q11). The pathogenetic and clinical effects of these translocations still need to be established. The terminal region of 17p, w h i c h normally contains the p53 gene, appears to have been lost as a result of the secondary translocation we describe. Because p53 has been shown to be rearranged in CML in both CP and BC [15, 16], loss of this gene in particular may be related to subsequent development of BC in our patient. The authors thank Dr. Sarah V. Williams for technical advice.
REFERENCES 1. Heisterkamp N, Stephenson JR, Groffen J, Hansen PF, de Klein A, Bartram CR, Grosveld G (1983): Localization of c-ABL oncogene adjacent to a translocation break point in chronic myelocytic leukemia. Nature 308:239-242. 2. First International Workshop on Chromosomes in Leukemia (1979): Chromosomes in Ph+ chronic myelogenous leukemia. Br J Haematol 39:305-309.
3. Hagemeijer A, Bartram CR, Smit EME, van Agthoven AJ, Bootsma D (1984): Is the chromosomal region 9q34 always involved in variants of the Ph translocations? Cancer Genet Cytogenet 13:1-16. 4. Ishihara T and Minamihisamatsu M (1988): The Philadelphia chromosome. Considerations based on studies of variant Ph translocations. Cancer Genet Cytogenet 32:75-92. 5. Hagemeijer A, de Klein A, Godde-Salz E, Turc-Care| C, Smith EME, van Agthoven A, Grosveld G (1985): Translocation of cabl to "masked" Ph in chronic myeloid leukemia. Cancer Genet Cytogenet 18:95-104. 6. Ohyashiki K, Ohyashiki JH, Kinniburgh AJ, Rowe J, Miller KB, Raza A, Preisler HD, Sandberg AA (1987): Transposition of breakpoint cluster region (3'bcr) in CML ceils with variant Philadelphia translocations. Cancer Genet Cytogenet 26:105-115. 7. Zaccaria A, Testoni N, Tassinari A, Celso B, Rassool F, Saglio G, Guerrasio A, Rosti G, Tura S (1989): Cytogenetic and molecular studies in patients with chronic myeloid leukemia and variant Philadelphia translocations. Cancer Genet Cytogenet 42:191-201, 8. van der Plas DC, Hermans ABC, Soekarman D, Smit EME, de Klein A, Smadja M, Alimena G, Goudsmit R, Grosveld G, Hagemeijer A (1989): Cytogenetic and molecular analysis in Philadelphia negative CML. Blood 73:1038-1044. 9. Abe S, Minamihisamatsu M, Ishihara T, Sasaki M (1989): Chromosomal in situ hybridization and southern blot analyses using c-abl, c-sis, or bcr probe in chronic myelogenous leukemia ceils with variant Philadelphia translocations. Cancer Genet Cytogenet 38:61-74. 10. Tkachuk DC, Westbrook CA, Andreef M, Donlon TA, Cleary ML, Suryanarayan K, Homge M, Redner A, Gray J, Pinkel D (1990): Detection of bcr-abl fusion in chronic myelogenous leukemia by in situ hybridization. Science 250:559562. 11. Yagle MK, Parruti G, Xu W, Ponder BAJ, Solomon E (1990):
In Situ Detection of a Ph C o m p le x Translocation
Genetic and physical map of the von Recklinghausen neurofibromatosis (NF1) region on chromosome 17. Proc Natl Acad Sci USA 87:7255-7259. 12, ISCN (1991): Guidelines for Cancer Cytogenetics, Supplement to an International System for Human Cytogenetic Nomenclature, F Mitelman (ed.), Karger, Basel, 1991. 13. Zitzelsberger H, Bauchinger M, Wilmanns W, Strauss PG (1990): Cytogenetic and molecular analysis of a "masked" Philadelphia in chronic and blastic phases of chronic myeloid leukemia. Cancer Genet Cytogenet 47:219-225. 14. Melnyk A, Kowal-Vern A (1990): Complex translocations in
55
a patient with chronic myelocytic leukemia: t(9;22;17) and t(10;18;19) after two failed bone marrow transplantations. Cancer Genet Cytogenet 46:83-87. 15. Kelman Z, Prokocimer M, Peller S, Kahn Y, Rechavi G, Manor Y, Cohen A~ Rotter V (1989): Rearrangement in the p53 gene in Philadelphia chromosome positive chronic myelogenous leukemia. Blood 74:2318-2324. 16. Mashal R, Shaltrid M, Talpaz M, Kantarijan H, Deisseroth A (1990): Rearrangement and expression of p53 in the chronic phase and blast crisis of chronic myelogenous leukemia. Blood 75:180-189.
ABSTRACT:
We report a cytogenetic study of a patient with chronic myelogenous leukemia (CML) who, while displaying a Philadelphia (Ph) chromosome, resultingfrom a standard t(9;22J at diagnosis, during the chronic phase (CP) showed disappearance of the Ph and occurrence of new chromosome changes, including a marker probably arising from a translocation involving chromosome 17 and the Ph. In situ hybridization confirmed the cytogenetic appearance and demonstrated that the breakpoint on the Ph marker occurred below the BCR-ABL fusion gene
INTRODUCTION
More than 90% of patients with chronic myelogenous leukemia (CML) have a Philadelphia (Ph) chromosome in their bone marrow (BM) cells. This marker chromosome arises from a balanced translocation between chromosomes 9 and 22. As a consequence, the ABL protooncogene, normally located in band 9q34, moves to the BCR gene in band 22ql1, producing a BCR-ABL fusion gene believed to be involved in the pathogenesis of CML [l]. A few patients with typical clinical and hematologic signs of the disease show either no evidence of the Ph chromosome or a complex rearrangement involving chromosomes other than 9 and 22 [2-41. Each of these cases that has been subjected to molecular analysis has been shown to have the BCR-ABL fusion gene [5-91. We describe a patient with typical CML who developed a new translocation involving the Ph chromosome during the chronic phase (CP). CASE REPORT AND RESULTS
D. 0. G., a 50-year-old woman, was admitted to the division of hematology in November 1984 with complaints of asthenia, fever, pain below the left costal margin, and weight loss. Physical examination showed hepatosplenomegaly (3 and 6 cm, respectively, below the costal margins). Hematologic and biochemical investigations performed on peripheral blood (PB) showed a hemoglobin level 12 gidl, a red
From the Institute of Biology and Genetics, University ofchieti, Italy (G. C., G. P.), Department ofMedicine, Section ofHematology/ Oncology, University of Chicago Medical Center, Illinois (C. A. W.), and Human Cytogenetics Laboratory, Imperial Cancer Research Fund, London, England [D. S.J. Address reprint requests to: Giandomenico Palka, Via B. Buozzi 93, 65121 Pescara, Italy. Received December 27, 1991; accepted April 14, 1992.
blood cell (RBC) count of 4.5 x 1012/l, a white blood cell (WBC) count of 120 x log/L with 50% neutrophils, 2% eosinophils, 2% basophils, 12% myelocytes, 10% promyelocytes, 1% blasts, 3% monocytes, and 20% lymphocytes, and a platelet count of 600 x log/l; the leukocyte alkaline phosphatase score was 2. BM aspirate showed myeloid hyperplasia. A cytogenetic investigation performed on BM cells showed the karyotype 46,XX,t(9;22) (q34;qll) in 50 metaphases studied by GTG-banding. Induction treatment, started soon after diagnosis, comprised cytosine arabinoside and myleran. During CP, the patient was submitted to radiotherapy three times to reduce her splenomegaly. She remained in CP for 48 months, and all cytogenetic investigations performed on her BM cells in this period [every 6 months) showed the t(9;22)(q34;qll) as the only chromosome rearrangement. In November 1988, when the patient was still in clinical and hematologic CP, cytogenetic analysis showed the presence of a new clone containing a marker chromosome apparently derived from a translocation between the Ph and chromosome 17, with the breakpoints in the distal long arm of the Ph and at centromeric level of chromosome 17 (Fig. 1). No Ph was present in this clone, but other new aberrations observed were a t(5;14)(q35;q22), a de1(7)(p13), and monosomies 17 and 18. The 9q+ chromosome was also present in this clone. Fluorescence in situ hybridization [FISH) performed with a biotinylated 3’-ABL probe, c-H-abl probe [lo], and a cosmid probe, D178163, mapping to band 17qll-12 [ 1 l] demonstrated the presence of both probes on the new marker chromosome, confirming the involvement of chromosome 17 and indicating that the breakpoint on the Ph occurred below the BCR-ABL fusion gene (Fig. 2). Thus, according to the new Internation System for Human Cytogenetic Nomenclature [l2], the karyotype was defined as 44, XX,t(5;14)(q35;q22),de1(7)(p13),der(9)t(9;22)~q34;qll),-l7, - lB,der(22)t(9;22)(q34;qll)t(9;17)(q34;qlO). Twelve months later, BM blasts had increased by 30%.
52 Cancer Genet Cytogenet 6352-55 0165-4606/92/$05.00
(1992)
0 1992 Elsevier Science Publishing Co., Inc. 655 Avenue of the Americas, New York. NY 10010
In Situ Detection of a Ph Complex Translocation
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Figure 1 Karyotype in GTG-banding showing monosomy 18, a t(5;14)(q35;q22), a del(7)(p11), a chromosome 9q +, and a translocation involving chromosomes 17 and Ph (M). Inset: Two other partial karyotypes from the latter rearrangement.
Cytogenetic analysis showed the karyotype described above in all cells. After a short time, the patient developed clinical signs of blast crisis (BC) and died 3 months later: a survival time of 63 months from diagnosis.
DISCUSSION
Variant Ph translocations in CML were initially described mainly in patients in BC of CML, but more recently have been detected at all stages of the disease. Cytogenetically they appear to be either of the simple type, usually involving chromosome 22 and a different chromosome, or of the complex type, involving chromosomes 9, 22, and a third or more chromosomes. The findings of ABL sequences on the
Ph chromosome and of the presence of a BCR-ABL fusion even in apparently simple variant translocations strongly suggest that all variant translocations involve both chromosomes 9 and 22 and are therefore of the complex type [5-9]. Moreover, they can arise as secondary translocations during evolution of the disease, involving the 9q + chromosome or more rarely the Ph previously generated from the standard t(9;22)(q34;q11) [4, 6-9, 13, 14]. At diagnosis, the patient with typical clinical and hematologic characteristics of CML had a standard t(9;22) (q34;q11). During CP, other cytogenetic changes occurred, including replacement of the Ph with a new marker that appeared to derive from a translocation between the Ph and chromosome 17. FISH showed the 3'ABL and D17S163 probes on the new marker, confirming the cytogenetic appearance of a "masked" Ph [6] and demonstrating that the
54
G. Calabrese et al.
A
B Figure 2 (A) Metaphase prebanded with Wright stain. Arrows show 9q + chromosome and the marker chromosome. (B) Probe signals from c-H-abl (long arrows) and D17S163 (short arrows) on the marker chromosome and normal chromosomes 9 and 17 after in situ hybridization on the same metaphase.
secondary breakpoint on the Ph was below the B C R - A B L fusion gene. This case and others s h o w in g secondary translocations i nvolv i n g the 9 q + or Ph c h r o m o s o m e s raise the question of w het h er specific sequences on these chromosomes are susceptible to preferential breakage during evolution of CML. A c o m b i n e d approach of molecular analysis and FISH with appropriate probes on c h r o m o s o m e s 9 and 22 might be informative in this regard. Presumably the leukemia cells carrying these translocations have a selective advantage over cells having only the standard t(9;22)(q34;q11). The pathogenetic and clinical effects of these translocations still need to be established. The terminal region of 17p, w h i c h normally contains the p53 gene, appears to have been lost as a result of the secondary translocation we describe. Because p53 has been shown to be rearranged in CML in both CP and BC [15, 16], loss of this gene in particular may be related to subsequent development of BC in our patient. The authors thank Dr. Sarah V. Williams for technical advice.
REFERENCES 1. Heisterkamp N, Stephenson JR, Groffen J, Hansen PF, de Klein A, Bartram CR, Grosveld G (1983): Localization of c-ABL oncogene adjacent to a translocation break point in chronic myelocytic leukemia. Nature 308:239-242. 2. First International Workshop on Chromosomes in Leukemia (1979): Chromosomes in Ph+ chronic myelogenous leukemia. Br J Haematol 39:305-309.
3. Hagemeijer A, Bartram CR, Smit EME, van Agthoven AJ, Bootsma D (1984): Is the chromosomal region 9q34 always involved in variants of the Ph translocations? Cancer Genet Cytogenet 13:1-16. 4. Ishihara T and Minamihisamatsu M (1988): The Philadelphia chromosome. Considerations based on studies of variant Ph translocations. Cancer Genet Cytogenet 32:75-92. 5. Hagemeijer A, de Klein A, Godde-Salz E, Turc-Care| C, Smith EME, van Agthoven A, Grosveld G (1985): Translocation of cabl to "masked" Ph in chronic myeloid leukemia. Cancer Genet Cytogenet 18:95-104. 6. Ohyashiki K, Ohyashiki JH, Kinniburgh AJ, Rowe J, Miller KB, Raza A, Preisler HD, Sandberg AA (1987): Transposition of breakpoint cluster region (3'bcr) in CML ceils with variant Philadelphia translocations. Cancer Genet Cytogenet 26:105-115. 7. Zaccaria A, Testoni N, Tassinari A, Celso B, Rassool F, Saglio G, Guerrasio A, Rosti G, Tura S (1989): Cytogenetic and molecular studies in patients with chronic myeloid leukemia and variant Philadelphia translocations. Cancer Genet Cytogenet 42:191-201, 8. van der Plas DC, Hermans ABC, Soekarman D, Smit EME, de Klein A, Smadja M, Alimena G, Goudsmit R, Grosveld G, Hagemeijer A (1989): Cytogenetic and molecular analysis in Philadelphia negative CML. Blood 73:1038-1044. 9. Abe S, Minamihisamatsu M, Ishihara T, Sasaki M (1989): Chromosomal in situ hybridization and southern blot analyses using c-abl, c-sis, or bcr probe in chronic myelogenous leukemia ceils with variant Philadelphia translocations. Cancer Genet Cytogenet 38:61-74. 10. Tkachuk DC, Westbrook CA, Andreef M, Donlon TA, Cleary ML, Suryanarayan K, Homge M, Redner A, Gray J, Pinkel D (1990): Detection of bcr-abl fusion in chronic myelogenous leukemia by in situ hybridization. Science 250:559562. 11. Yagle MK, Parruti G, Xu W, Ponder BAJ, Solomon E (1990):
In Situ Detection of a Ph C o m p le x Translocation
Genetic and physical map of the von Recklinghausen neurofibromatosis (NF1) region on chromosome 17. Proc Natl Acad Sci USA 87:7255-7259. 12, ISCN (1991): Guidelines for Cancer Cytogenetics, Supplement to an International System for Human Cytogenetic Nomenclature, F Mitelman (ed.), Karger, Basel, 1991. 13. Zitzelsberger H, Bauchinger M, Wilmanns W, Strauss PG (1990): Cytogenetic and molecular analysis of a "masked" Philadelphia in chronic and blastic phases of chronic myeloid leukemia. Cancer Genet Cytogenet 47:219-225. 14. Melnyk A, Kowal-Vern A (1990): Complex translocations in
55
a patient with chronic myelocytic leukemia: t(9;22;17) and t(10;18;19) after two failed bone marrow transplantations. Cancer Genet Cytogenet 46:83-87. 15. Kelman Z, Prokocimer M, Peller S, Kahn Y, Rechavi G, Manor Y, Cohen A~ Rotter V (1989): Rearrangement in the p53 gene in Philadelphia chromosome positive chronic myelogenous leukemia. Blood 74:2318-2324. 16. Mashal R, Shaltrid M, Talpaz M, Kantarijan H, Deisseroth A (1990): Rearrangement and expression of p53 in the chronic phase and blast crisis of chronic myelogenous leukemia. Blood 75:180-189.