Cytogenetic response to imatinib treatment in Southern Brazilian patients with chronic myelogenous leukemia and variant Philadelphia chromosome

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1 Ann Hematol (2013) 92: DOI /s ORIGINAL ARTICLE Cytogenetic response to imatinib treatment in Southern Brazilian patients with chronic myelogenous leukemia and variant Philadelphia chromosome Dayane B. Koshiyama & Marcelo E. Z. Capra & Giorgio A. Paskulin & Rafael F. M. Rosa & Ceres A. V. Oliveira & Tito Vanelli & Laura M. Fogliatto & Paulo R. G. Zen Received: 16 April 2012 / Accepted: 5 October 2012 / Published online: 14 October 2012 # Springer-Verlag Berlin Heidelberg 2012 Abstract Variant Philadelphia (Ph) chromosome can be observed in 5 10 % of chronic myelogenous leukemia (CML) patients. However, there are only a few studies which have analyzed the prognostic implications of these complex translocations in CML patients after the advent of imatinib mesylate and the results found are conflicting. We investigated the clinical features and cytogenetic response of Brazilian chronic phase (CP) CML patients with variant Ph treated with imatinib mesylate. Among 93 CP CML patients, eight (8.6 %) exhibited complex translocations, involving one (n06), two (n01), or three (n01) additional chromosomes. At 6, 12, and 18 months, a complete cytogenetic response was observed in 100 % of variant Ph patients, respectively. No significant difference was found between variant Ph and standard D. B. Koshiyama : G. A. Paskulin : R. F. M. Rosa : P. R. G. Zen (*) Clinical Genetics and Graduate Program of Pathology, Universidade Federal de Ciências da Saúde de Porto Alegre, Rua Sarmento Leite 245/403, Porto Alegre , RS, Brazil paulozen@ufcspa.edu.br M. E. Z. Capra : T. Vanelli Hematology Oncology Division, Grupo Hospitalar Conceição, C. A. V. Oliveira Institute of Education and Research, Hospital Moinhos de Vento, L. M. Fogliatto Hematology, Universidade Federal de Ciências da Saúde de Porto Alegre, translocation patients regarding the response to IM treatment at 6, 12, and 18 months. Likewise, there was no statistically significant difference between the two groups concerning the overall survival, failure-free survival, progression-free survival, and event-free survival. The results obtained in our study, despite our sample size, suggest, in agreement to other data found in the literature, that the presence of variant Philadelphia chromosome does not bestow a prognostic disadvantage when compared to the group with classic Ph. This observation does not suggest the need to adjust the treatment protocol due to the presence of variant Ph. However, further studies with larger sample sizes and evaluating both the cytogenetic and molecular response to IM treatment should be conducted to confirm our findings. Keywords Variant Ph. CML. Complex translocation. Imatinib mesylate Introduction Chronic myelogenous leukemia (CML) is a myeloproliferative disorder of hematopoietic pluripotent stem cells [1]. CML presents an estimated incidence of 1/100,000 cases per year, which accounts for % of leukemias. Although it might occur in all age groups, it is extremely rare in children and the age at diagnosis usually ranges between 53 and 65 years [2, 3]. In more than 90 % of CML cases, the presence of the Philadelphia chromosome (Ph), which is the product of a balanced translocation between the long arms of

2 186 Ann Hematol (2013) 92: Table 1 Clinical features and cytogenetic results of CP variant Ph patients Patient Sex Age WBC PLT Hb Sokal score Karyotype 4 F Low 46,XX,t(9;22;17)(q34;q11.2;q21) [20] 9 M High 46,XY,t(4;9;22)(q35;q34;q11.2) [20] 28 M Int 46,XY,t(3;7;9;22;5)(p21;q22;q34;q11.2;q22) [24] 51 F Low 46,X,t(X;9;22)(q28;q34;q11.2) [20] 84 M Low 46,XY,t(9;22;19)(q34;q11.2;q13.3) [22] 109 M Int 46,XY,t(6;6;9;22)(q21;p21.3;q34;q11.2) [32] 114 M Low 46,XY,t(9;22;14)(q34;q11.2;q22) [45] 136 F Low 46,XX,t(9;22;11)(q34;q11.2;q13) [20] All karyotypes were described according to ISCN 2009 WBC white blood cells count, PLT platelet count, Hb hemoglobin concentration, F female, M male, Int intermediate chromosomes 9 and 22, can be detected during a conventional cytogenetic analysis [4 7]. This rearrangement involves ABL1 (Abelson) oncogene, localized at the long arm of chromosome 9, and breakpoint cluster region (BCR) gene, located at the long arm of chromosome 22 [8, 9]. Its fusion results in the formation of a BCR-ABL1 hybrid gene, which encodes a constitutively active tyrosine kinase protein [10 12]. This protein plays a crucial role in CML pathogenesis, since it is responsible for the activation of proteins involved in cell cycle regulation, therefore accelerating cell division and affecting DNA repair [13, 14]. In 5 10 % of CML cases, a complex translocation leads to the formation of a variant Ph chromosome [15, 16] and involves not only chromosomes 9 and 22, but also a third one, and, less frequently, more chromosomes [17]. The advent of imatinib mesylate (IM), a potent selective inhibitor of BCR-ABL tyrosine kinase, represents an important milestone in CML treatment because this drug is able to induce a complete cytogenetic response in most patients [7, 18]. However, there are only a few studies which have analyzed the prognostic implications of these complex translocations in CML patients and the results found are conflicting. We herein present data from a study conducted with 93 chronic phase (CP) CML patients who attended at Hospital Nossa Senhora da Conceição aiming to correlate the clinical importance of variant Ph with cytogenetic response to IM treatment. Patients and methods The Hematology Oncology Division of Grupo Hospitalar Conceição (GHC), located in Porto Alegre, Rio Grande do Sul, is a reference center for the care of CML patients and, between 1998 and 2011, 158 patients were followed up. This study included only CP CML patients who underwent cytogenetic evaluation at the time of diagnosis and after the IM treatment and who had complete medical records available. Thus, the final sample consisted of 93 patients, who were treated either with a dose of 400 or 600 mg of IM. In this retrospective study, a systematic review of patients' records was conducted in order to fill in a standard clinical protocol, which contained information regarding their clinical features, such as phase at diagnosis, hemoglobin level, platelet and white blood cell (WBC) counts, as well as the results from their bone marrow cytogenetic evaluation through the GTG-banding karyotype. Standard International System for Human Cytogenetic Nomenclature Table 2 Comparison between clinical characteristics of CP CML patients with variant Ph and standard translocation Characteristics Variant Ph Standard translocation Median age, years (range) 38 (13 75) 48 (12 86) Sex, male/female, n (%) 5/3 (62.5/37.5) 48/37 (56.5/44.5) Median hemoglobin level, g/dl (range) ( ) 11.4 ( ) Median platelet count, 10 9 /L (range) ( ) (91 890) Median WBC count, 10 9 /L (range) ( ) ( ) Sokal score Low, n (%) 5 (62.5) 59 (69.4) Intermediate, n (%) 2 (25) 14 (16.5) High, n (%) 1 (12.5) 12 (14.1)

3 Ann Hematol (2013) 92: (2009) was used to report cytogenetic results [19]. Cytogenetic response was defined according to the European LeukemiaNet criteria [20]. The statistical analysis was carried out using the Kaplan Meier method to estimate overall survival (OS), failure-free survival (FFS), progression-free survival (PFS), and eventfree survival (EFS), which were calculated from the beginning of the IM treatment until death (OS), until failure or death (FFS), until progression to accelerated phase or blast crisis or death (PFS), and until any event (EFS), respectively. Log-rank test was applied to compare survival curves. Fisher's exact test was used to compare the cytogenetic response between the groups. For the sample size of our study, and considering a maximum difference in survival between the two groups of 20 %, the power of the test is 40.3 %. In order to perform the statistical analyses, we used the SPSS 12.0 program. The results were considered as significant if P<0.05. This study was approved by the ethics committees of GHC and of Universidade Federal de Ciências da Saúde de Porto Alegre. Results Among the 93 patients who met the inclusion criteria, eight (8.6 %) displayed the presence of variant Ph chromosome, and their main features are shown in Table 1. Five patients were male, their median age was 38 years (ranged between 13 and 75 years), median WBC count was L (ranged between 35.2 and 409.9), median hemoglobin level was g/dl (ranged between 6.6 and 14.4), and median platelet count was L (ranged between 134 and 620). Five patients presented a low Sokal risk score, whilst two exhibited an intermediate risk and one a high Sokal risk [21]. In regard to the cytogenetic characteristics, all complex translocations involving the Ph chromosome were present at diagnosis. In six cases, three chromosomes were implicated (three-way translocation), whereas the two remaining cases concerned four and five chromosomes, in a four-way translocation and a five-way translocation, respectively. Rearrangements involving chromosomes X, 3, 4, 5, 6, 7, 11, 14, 17, and 19 were detected. Currently, seven patients are still alive with a median follow-up time of 43 months (ranges between 6 and 97). One patient died at 24 months due to sepsis. From the 85 patients who presented standard translocation at diagnosis, 48 were males and 37 females and their main clinical characteristics are listed in Table 2. The median age was 48 years (ranged between 12 and 86). Median WBC count was L (ranged between 3.8 and 424), median hemoglobin level was 11.4 g/dl (ranged between 6.5 and 14.4), and median platelet count was L (ranged between 91 and 890). In this group, 59 patients (69.4 %) presented a low Sokal risk score, whereas 14 (16.5 %) exhibited an intermediate risk and 12 (14.1 %) a high Sokal risk [21]. Currently, 80 are alive and the median follow-up time is 57 months (ranges between 2 and 172). The main cause of death among the five deceased patients was sepsis. After an average of 6 months of IM treatment, 100 % of variant Ph patients presented a complete cytogenetic response (CCgR), whereas in the standard t(9;22) group, the CCgR was observed in 87.5 % of the patients (P01.000). At 12 months, a CCgR was obtained in 100 and 86.7 % of patients with and without complex translocations, respectively (P01.000). At Table 3 Description of studies that evaluated the role of variant Ph at CML patients' outcome El-Zimaity et al. [22] Stagno et al. [25] Marzocchi et al. [23] Fabarius et al. [24] Present study Variant Ph, n (%) 44 (6.1) 10 (6.5) 30 (5.4) 69 (6) 8(8.6) Sex, male/female, n (%) 24/20 (54.5/45.5) 9/1 (90/10) 19/11 (63/37) 38/31 (55/45) 5/3 (62.5/37.5) Median age, years (range) 48 (27 75) 57 (29 74) 52 (33 84) 54 (16 88) 38 (13 75) Median hemoglobin level, ND 10.9 ( ) 11.8 ( ) ND ( ) g/dl (range) Median platelet count, 10 9 /L ND 252 ( ) 317 (124 1,364) ND ( ) (range) Median WBC count, 10 9 /L ND ( ) ND ND ( ) (range) Stage, AP/BC/CP, n (%) 19/1/24 (44/2/55) 0/0/10 (0/0/100) 0/0/30 (0/0/100) 0/0/69 (0/0/100) 0/0/8 (0/0/100) CCgR (%) a ND 100 MCgR (%) a ND 100 Classic Ph WBC white blood cells, AP accelerated phase, BC blast crisis, CP chronic phase, CCgR complete cytogenetic response, MCgR major cytogenetic response, ND not described a Last cytogenetic response cited in the study

4 188 Ann Hematol (2013) 92: months, 100 % of variant Ph patients exhibited CCgR, whereas 95.7 % of standard translocation patients presented a CCgR (P01.000). The estimated cumulative probabilities for OS, FFS, PFS, and EFS at 95 months were 80 and 92.6 %, 80 and 86.4 %, 80 and 84 %, and 80 and 73.2 % for patients with variant Ph and standard translocation, respectively. In the statistical analysis, using the Pearson chi-square test, no statistically significant difference was observed between the two groups concerning the Sokal risk score (P00.795). Fisher's exact test showed no significant difference between variant Ph and standard translocation patients regarding the response to IM treatment at 6 (P01.000), 12 (P01.000), and 18 months (P01.000). Likewise, using logrank test, there was no statistically significant difference between the two groups concerning OS (P00.111), FFS (P00.638), PFS (P00.436), and EFS (P00.926). Discussion In about 5 10 % of CML cases, a complex translocation leads to the formation of a variant Ph chromosome [15, 16]. The information available about survival rates and cytogenetic response concerning these patients is limited, and, to our knowledge, there are only four studies evaluating this aspect (Table 3). In our study, 8.6 % of CML patients exhibited variant Ph, which is in agreement with the data found in the literature [15 25]. The median age of our patients was 38 years, slightly lower than the one found in similar studies [22 25], which might be due to the fact that the present study included patients under 18 years of age. The male/ female ratio was 5:3, showing a higher proportion of male patients. Median hemoglobin level as well as median WBC count and median platelet count were similar to the other reports [23, 25]. In our study, the CCgR rates of variant Ph patients were 100, 100, and 100 % at 6, 12, and 18 months, respectively, and showed no statistically significant difference when compared to standard translocation individuals. In the first study conducted aiming to clarify the role of variant Ph in CML outcome, the authors identified 44 (6 %) patients with variant Ph among 721 CML individuals and the observed CCgR and major cytogenetic response (MCgR) rates were and %, respectively [22]. There were no significant statistical differences between patients with variant Ph and standard translocation concerning MCgR and CCgR rates, duration of the response, and survival rates, in agreement to the results obtained in the present study. A more recent report evaluated the variant Ph role in recently diagnosed CML patients in CP and who were treated with IM [23]. They identified 30 (5 %) variant Ph patients among 559 CML individuals. The CCgR at last contact for patients with variant Ph was 77.7 %, lower than the one found in our study (100 %). The estimated probabilities for OS, FFS, PFS, and EFS (85 and 90 %, 73 and 74 %, 83 and 87 %, and 70 and 64 % for patients with and without complex translocations, respectively), were similar to our results. In 2011, Fabarius et al. showed that the 69 patients with variantph(6%)includedintheirstudyhadthesame prognosis as patients with standard t(9;22). The OS and the PFS rates at 5 years were 92 and 87 % and 90 and 81 % for patients with standard or complex translocation, respectively. On the other hand, a recently conducted study with ten (6.5 %) patients presenting complex translocations at diagnosis and treated with IM or nilotinib demonstrated that these patients do exhibit a poor clinical outcome when compared to individuals with standard translocation [25]. The authors suggest that complex translocation involving the Ph chromosome is associated with a higher degree of genomic instability and, therefore, a more aggressive form of CML, hence conferring a less favorable clinical outcome. Nevertheless, the results of other studies [22 24], as well as our own, do not corroborate this hypothesis. Thus, the results obtained by Stagno et al. [25] might be attributed to sampling bias, as in their report there was only one female patient in the sample. In fact, it has been described that female CML patients have a longer median survival in comparison to male patients [26]. In conclusion, the results obtained in our study, despite our sample size, suggest, such as El-Zimaity et al. [22], Marzocchietal.[23], and Fabarius et al. [24], that the presence of variant Philadelphia chromosome does not bestow a prognostic disadvantage when compared to the group with classic Ph. 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