Available online at www.annclinlabsci.org Annals of Clinical & Laboratory Science, vol. 39, no. 3, 2009 307 Case Report and Review of the Literature: Chronic Myelomonocytic with der(9)t(1;9)(q11;q34) as a Sole Abnormality

Borum Suh,1 Tae Sung Park,1* Jin Seok Kim,2 Jaewoo Song,1 Juwon Kim,1 Jong-Ha Yoo,1,3 and Jong Rak Choi1 Departments of 1Laboratory Medicine and 2Internal Medicine, Yonsei University College of Medicine, Seoul, Korea; 3Department of Laboratory Medicine, National Health Insurance Corporation Ilsan Hospital, Goyang-si, Kyonggi-do, Korea

Abstract. The chromosomal abnormality der(9)t(1;9)(q11;q34) is a rare occurrence in patients with hematologic malignancies. As far as we know, only 3 cases of acute , 1 case of , and 1 case of multiple myeloma with this derivative have been reported in the literature. Here we report the first case of der(9)t(1;9)(q11;q34) in a patient with chronic myelomonocytic leukemia (CMML). A 45-yr-old man was brought to our hospital for evaluation of pancytopenia and . The patient’s persistent monocytosis in peripheral blood and his findings were consistent with the diagnosis of CMML. Chromosome study results repeatedly showed 46,XY,der(9)t(1;9)(q11;q34). In addition, the BCR/ABL fluorescent in situ hybridization (FISH) pattern of the interphase cells was interpreted as: “nuc ish(ABL, BCR) x 2[292/300],” consistent with the normal signal patterns found in 97% of the nuclei examined. For further evaluation, multi-color FISH (mFISH) analysis was performed and it showed the distinct unbalanced derivative chromosome der(9)t(1;9)(q11;q34) in 5 metaphase cells analyzed. Not only does this show an extraordinary type of trisomy 1q, but it reveals a rare recurrent case of der(9)t(1;9)(q11;q34) in patients with monocytic-lineage leukemia. Further studies are needed to evaluate the prognosis, survival, and treatment response of such patients with der(9)t(1;9)(q11;q34).

Keywords: der(9)t(1;9)(q11;q34), CMML, mFISH, monocytic-lineage leukemia

Introduction myeloproliferative (MPN) by the presence of both dysplastic and proliferative features at the Chronic myelomonocytic leukemia (CMML) is a time of initial presentation [1,2]. The most common clonal disorder of bone marrow stem cells in which chromosomal abnormalities in MDS or CMML monocytosis is a major defining feature [1]. CMML are deletion 5q, monosomy 7, and trisomy 8. was initially classified in the category of myelodys- Although a predominance of trisomy 1q in Korean plastic syndrome (MDS), but it is now categorized patients with MDS has been reported [3], complete by the 2001 World Health Organization (WHO) trisomy 1q is rare, even in Korean CMML patients. classification in a separate nosological group of In addition, der(9)t(1;9)(q11;q34) has been reported MDS/MPN, which is distinguished from MDS or in only 5 patients, including a case of acute myelomonocytic leukemia (AML-M4), a case of * Dr Tae Sung Park’s current address is Department of Laboratory Medicine, Kyung Hee Univiversity Medical acute (AML-M5a), and an School, Seoul, Korea; tel 822 958 8673; fax 822 958 8609. unspecified AML with , but not in Address correspondence to: Jong Rak Choi, M.D., Ph.D., CMML patients [4-9]. We describe a novel case of Department of Laboratory Medicine, Yonsei University der(9)t(1;9)(q11;q34) in a patient with CMML, College of Medicine, 250 Seongsanno, Seodaemun-gu, Seoul 120-752, Korea; tel 822 2228 2445; fax 822 313 0956; e-mail and review the medical and laboratory data of the [email protected]. patient and the relevant literature. 0091-7370/09/0300-0307. $2.10. © 2009 by the Association of Clinical Scientists, Inc. 308 Annals of Clinical & Laboratory Science, vol. 39, no. 3, 2009

Case Report Upon arrival, his CBC showed a Hb level of 9.0 g/dl, a platelet count of 130,000 /μl, and a WBC count of 2,090 /μl, with A 45-year old Korean man with a history of diabetes mellitus 14% segmental neutrophils, 21% lymphocytes, 3% atypical dating from November 2002 was brought to Ilsan Hospital lymphocytes, 2% immature cells, and 60% . His for evaluation of pancytopenia in August 2004. Complete third bone marrow examination, May 2008, showed normo- blood count (CBC) showed a hemoglobin (Hb) level of 8.5 cellularity with 3.3% blasts. Peripheral blood monocytosis g/dl, a platelet count of 120,000/μl, and a WBC count of (>1,000 /μl) has been consistently observed after the initial 1,900 /μl. Pancytopenia compelled us to perform a peripheral diagnosis of CMML. For further evaluation, cytogenetics, blood smear (PBS) and bone marrow examination, and the fluorescent in situ hybridization (FISH), and multi-color WBC differential counts were as follows: 24% segmental FISH (mFISH) analyses were conducted. neutrophils, 31% lymphocytes, and 45% monocytes. The first bone marrow aspiration (August 2004) showed 80% hyper- Materials and Methods cellular marrow with erythroid hyperplasia (the myeloid to erythroid ratio was 1.1:1). Although dyspoietic megakaryo- Standard cytogenetic techniques and nomenclature were cytes (microforms, separated nuclear lobules) were detected, followed [10]. no other distinct dysplasia was observed in the marrow. The FISH analysis was performed according to the PBS and bone marrow findings were consistent with the manufacturer’s instructions with commercially available diagnosis of CMML. FISH probes supplied by Abbott Molecular/Vysis (Des The patient was referred to Severance Hospital in March Plaines, IL). The analyses were performed with probes for 2005. His second bone marrow aspiration, July 2005, showed BCR/ABL (LSI BCR/ABL Dual Color, Dual Fusion a moderately hypercellular marrow with marked increase of Translocation Probe Set), IGH/MYC (LSI IGH(14q32)/ abnormal megakaryocytes. He was treated with oxymetholone MYC(8q24), CEP 8 Tri-Color, Dual Fusion Translocation for on an outpatient basis, and was admitted to Probe Set), p53 (LSI 17p13.1 SpectrumOrange Probe), and Severance Hospital for decitabin treatment in May 2008. probes for chromosome 5 (LSI EGR1 (5q31) SpectrumOrange/

Fig. 1. (a) A metaphase cell stained with Giemsa-banding before analysis. (b) Full karyogram of the bone marrow cells: 46,XY, der(9)t(1;9)(q11;q34). (c) Partial karyogram of der(9)t(1;9)(q11;q34) in this patient. (d) Diagrammatic representation of chromosome 9 and der(9)t(1;9)(q11;q34). The arrows indicate breakpoints of 1 and 9. A case of CMML with der(9)t(1;9)(q11;q34) 309

Fig. 2. Results of multi-color FISH (mFISH) analysis in this patient. (a) A metaphase cell stained with the human 24XCyte probe before analysis. (b) mFISH image (false color labeled) showing der(9)t(1;9)(q11;q34) as the sole abnormality. The arrows indicate der(9)t(1;9)(q11;q34). (c) Parital image of inverted DAPI showing der(9)t(1;9)(q11;q34). (d) Single color galleries showing normal chromosome 1, normal chromosome 9, and der(9)t(1;9)(q11;q34), respectively.

D5S23,D5S721 SpectrumGreen Probe Set), chromosome 7 studies were as follows: 46,XY,der(9)t(1;9)(q11;q3) (LSI D7S486 (7q31) SpectrumOrange/CEP 7 SpectrumGreen in 20 cells (August 2004), 14 cells (July 2005), and Probe Set), and chromosome 20 (LSI D20S108 (20q12) SpectrumOrange Probe). At least 300 bone marrow interphase 22 cells (May 2008) (Fig. 1). FISH signals from the cells were scored for signal patterns for each probe using a BCR/ABL probe showed the normal signal pattern fluorescence microscope (Axio Imager M1; Carl Zeiss Micro- in 97% of the nuclei examined, which was Imaging GmbH, Germany). interpreted as “nuc ish(ABL, BCR) x 2[292/300].” The mFISH analysis was performed with a human This is consistent with the karyotyping analysis, in 24XCyte (MetaSystems, Altlussheim, Germany) probe. The hybridization, post-hybridization washes, and signal detection which t(9;22)(q34;q11.2) was not detected, and were carried out according to the manufacturer’s protocol excludes the possibility of ABL gene involvement. (http://www.metasystems.de/downloads/mfish-manual.pdf). The results of the other FISH analyses with the Images were analyzed, captured, and processed with a IGH/MYC, p53, chromosome 5, chromosome 7, fluorescence microscope (Carl Zeiss MicroImaging GmbH) and Isis/mFISH imaging software (MetaSystems). and chromosome 20 probes were within reference ranges. In addition, mFISH analysis revealed the Results distinct unbalanced derivative chromosome der(9)t(1;9)(q11;q34) in 5 metaphase cells (May The patient repeatedly exhibited the same chromo- 2008) (Fig. 2), consistent with the cytogenetic somal abnormality. The results of the chromosome results of this patient (Fig. 1). 310 Table 1. Comparison of previous reports with der(9)t(1;9)(q11;q34) and this study. Annals of Clinical & Laboratory Science, vol. 39, no. 3, 2009 39, 3, no. vol. Science, &Laboratory of Clinical Annals Case 1 Case 2 Case 3

Sex/Age M/45 (yr) F/56 (yr) F/21 (mo)

Diagnosis CMML AML-M4 AML-M5a

Abnormal karyotype 46,XY,der(9)t(1;9)(q11;q34) 46,XX,der(9)t(1;9)(q11;q34) 46,XX,t(1;9)(q11;q34)/46,idem, ins(10;11)(p12;q23q13) N of analyzed cells (abnormal/total) 56/56 14/28 NA

Peripheral blood Initial WBC counts (/μl) 1,900 44,700 11,700 Dominant cells (%) 45 (monocytes) 19 (myelocytic precursors) 37 ()

Initial BM findings 80% hypercellular marrow with erythroid Hypercellular marrow 100% of leukemic cells hyperplasia, dyspoietic megakaryocytes with 40% of leukemic cells

FISH (or further study) No BCR/ABL rearrangement (mFISH) ABL was not involved NA

Treatment oxymetholone, decitabin fludarabine, cytarabine, idarubicine r ubidazone, cytarabine, teniposide

Follow up 4 yr NA >6 mo

(CR status) (no CR) (CR after one course of ) (CR)

References Park et al, 2009 [present study] Piccaluga et al, 2004 [5] Berger et al, 1982 [6]

Abbreviations: M, male; F, female; CMML, chronic myelomonocytic leukemia; AML-M4, acute myelomonocytic leukemia; AML-M5a, acute monoblastic leukemia (poorly differentiated); BM, bone marrow; NA, not available; FISH, fluorescence in situ hybridization; mFISH, multi-color FISH; CR, complete remission. A case of CMML with der(9)t(1;9)(q11;q34)

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Discussion The present study is interesting from several laboratory and clinical viewpoints. First, The French-American-British (FAB) classification der(9)t(1;9)(q11;q34) is an extraordinary type of system for MDS was first devised in 1982 [11], and chromosomal abnormality revealing trisomy 1q. In included the following 5 entities: refractory anemia general, unbalanced WATs between the entire long (RA), refractory anemia with ringed sideroblasts arm of chromosome 1 and the entire long (or short) (RARS), refractory anemia with excess blasts arm of another chromosome are created by fusion (RAEB), refractory anemia with excess blasts in in the centromeric region. However, in our case, transformation (RAEB-t), and CMML. Among the whole long arm of chromosome 1 was relocated these, CMML always seemed out of place when to a distant telomeric region of chromosome 9q34. compared with the other 4 entities [12]; the clinical, Second, the results of the FISH and mFISH hematologic and morphologic features of CMML analyses led us to conclude that der(9)t(1;9)(q11;q34) are heterogeneous, and vary along a spectrum from was the sole chromosomal abnormality in this predominantly myelodysplastic to mainly myelo- patient. This derivative chromosome involves a proliferative [1]. According to the diagnostic breakpoint at 9q34, which is also the target of criteria, CMML is characterized by persistent typical t(9;22)(q34;q11.2) translocation of chronic monocytosis (>1,000/μl) in the peripheral blood, myelogenous leukemia. However, the possible absence of and BCR/ involvement of the ABL gene is not likely due to the ABL fusion gene, fewer than 20% blasts in the normal signal pattern of BCR/ABL FISH probes blood or bone marrow, and dysplasia involving one observed in 97% of the interphase nuclei examined. or more myeloid lineages. In 2001, however, the Unfortunately, we did not employ a confirmatory WHO classification moved CMML from MDS to method, such as reverse transcriptase PCR analysis, a new category of mixed MDS/MPD, and also to detect BCR/ABL gene rearrangement, and this is defined CMML I and CMML II according to a limitation of our study. medullary and peripheral blast counts [12,13]. In Third, the patient in our study is still alive 4 yr 2008, the WHO again revised the classification for and 7 mo after his first diagnosis in August 2004. MPD, but this revision did not substantively change This greatly exceeds the reported median survival the MDS/MPD mixed category [12,14]. of 20-40 mo for CMML patients at initial diagnosis Whole arm translocations (WATs) involving [17-19]. Although this is worthy of note, it is the long arm of chromosome 1 usually imply the uncertain whether this longevity is independently formation of an extra copy of a part of or the entire associated with this derivative chromosome. 1q with its relocation to a nonhomologous chromo- Another limitation of our study is the lack of some, resulting in the generation of a derivative data regarding other cytogenetic abnormalities chromosome [11,15]. Among these WATs, certain such as alterations in the RAS pathway that have abnormalities, eg, der(1;7)(q10;p10), are frequently been frequently found in CMML [20,21]. Such found in MDS, acute myelogenous leukemia information may be helpful in further elucidating (AML), and MPD as the sole anomaly, and their the molecular biology of this disease. clinical implications have been reported [16]. In This is the first report of der(9)t(1;9)(q11;q34) contrast to other ethnic groups, a predominance of in a patient with CMML as a sole abnormality. trisomy 1q in Korean patients with MDS was Although more reports are needed to investigate recently reported by Lee et al [3]. However, complete the molecular pathogenesis of this rare WAT in trisomy 1q in CMML is very rare. In addition, hematologic malignancies, we suggest that der(9)t(1;9)(q11;q34) has been reported in 5 patients der(9)t(1;9)(q11;q34) is more likely than not a rare, with myeloid including a case of AML- recurrent chromosomal aberration in myeloid neo- M4 and a case of AML-M5a, but not in CMML plasms, particularly in monocytic-lineage leukemia. patients (Table 1). Due to the lack of clinical Further cases are needed to evaluate the prognosis, information provided [7-9], only two reported cases survival, and treatment response of such patients and the present study are included in Table 1. with der(9)t(1;9)(q11;q34). 312 Annals of Clinical & Laboratory Science, vol. 39, no. 3, 2009

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