About this Journal Submit a Manuscript Table of Contents
Journal of Biomedicine and Biotechnology
Volume 2011 (2011), Article ID 230481, 8 pages
http://dx.doi.org/10.1155/2011/230481
Review Article

Cytogenetic Instability in Childhood Acute Lymphoblastic Leukemia Survivors

1Departamento de Puericultura e Pediatria, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, 14040-901 São Paulo, Brazil
2Departamento de Genética, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, 14040-901 São Paulo, Brazil
3Departamento de Biologia, Faculdade de Filosofia Ciências e Letras de Ribeirão Preto (FFCLRP-USP), Universidade de São Paulo, Avenida Bandeirantes 3900, 14040-901 Ribeirão Preto, SP, Brazil

Received 2 June 2010; Accepted 11 August 2010

Academic Editor: Hans Konrad Muller

Copyright © 2011 María Sol Brassesco et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Abstract

Contemporary anticancer therapies have largely improved the outcome for children with cancer, especially for Acute Lymphoblastic Leukemia (ALL). Actually, between 78% and 85% of patients achieve complete remission and are alive after 5 years of therapy completion. However, as cure rates increase, new concerns about the late effects of genotoxic treatment emerge, being the risk of developing secondary neoplasias, the most serious life-threatening rising problem. In the present paper, we describe and review the cytogenetic findings in peripheral lymphocytes from ALL survivors, and discuss aspects associated to the occurrence of increased chromosome rearrangements in this growing cohort.

1. Introduction

Acute lymphoblastic leukemia (ALL) is the most common malignancy in childhood and is associated with excellent outcomes [1]. Currently, 80% of children with ALL treated in modern centers are alive and disease-free at 5 years [2]. The major contributors to this long-term survival are the improvements in anticancer therapies. However, they have implied the exposure of patients to combinations of chemotherapy and, in some instances, preventive cranial irradiation, which are well-recognized DNA-damaging agents [3, 4]. Although most ALL survivors are at a lower risk of developing a late effect of therapy when compared with survivors of other pediatric cancers, these children are almost four times more likely than their siblings to develop a severe or life-threatening chronic medical condition [5]. Recently, the number of patients with second malignancies has increased among long-term survivors of pediatric ALL [68]. There is compelling evidence that specific exposure to radiation and chemotherapy are etiologic agents of secondary neoplasia [9].

Chromosomal aberrations, sister chromatid exchanges and micronuclei, which can be detected by cytogenetic analysis, have been used as important biomarkers of genotoxic exposure; moreover, the relevance of increased frequency of chromosome alterations as indicator of malignancy risk is supported by epidemiological studies suggesting that a high frequency of chromosomal aberrations is predictive of an increased likelihood of cancer [10, 11]. The genotoxicity of anticancer treatment has been evaluated by somatic cell mutation assays, immediately or shortly after completion of therapy. However, second malignancies in children usually occur several years after treatment [12]. Thus, the cytogenetic analysis in peripheral lymphocytes from cancer survivors, who have been exposed to chemotherapy, provides an opportunity to investigate the in vivo induction and persistence of genomic instability in humans, as well as to evaluate the long-term effects of cancer therapy.

In general, hematological malignancies are characterized by recurrent chromosomal aberrations that lead to the formation of gene fusions and the subsequent expression of chimeric proteins with unique properties [13]. Various gene fusions thought to be solely associated with leukemias and lymphomas, such as t(12;21) ETV6/RUNX1, t(9;22) BCR/ABL, t(14;18) IGH/BCL2, t(2;5) NPM-ALK, and MLL duplications have been detected in normal individuals with negative history of hematologic disorders [14, 15]. These and other findings suggested that the measurement of gene fusions in peripheral blood lymphocytes within a study group may be used as a sensitive assay for the detection of genomic instability, and may contribute to risk estimation for the development of lymphoid malignancies [16, 17].

During the last years, our group has investigated the existence of chromosome instability in peripheral lymphocytes from ALL survivors by means of fluorescent in situ hybridization (FISH) and molecular biology using leukemia/lymphoma associated gene fusions as putative markers. The proband group comprised 49 individuals aged 5 to 22 years (average = 12), diagnosed and previously treated for childhood ALL at the University Hospital (Faculty of Medicine, Ribeirão Preto, University of São Paulo, Brazil) with combined modality treatment according to the Brazilian Group of Pediatric Leukemia Treatment (GBTLI) [18]. In this protocol, children were treated with a polychemotherapeutic regimen that includes vincristine, dexamethasone, daunorrubicin, L-asparaginase, prednisone, methotrexate, cytosine arabinoside, cyclophosphamide, folinic acid, etoposide (VP-16), teniposide (VM-26), and 6 mercaptopurine. In some cases prophylactic cranial and/or neuroaxis irradiation was also included. The median event-free survival was 3.8 years (range 5 months to 16 years) and there were no relapses. The characterization of patients was described in the work by Brassesco et al. [19].

Frequency of chromosomal aberrations were compared to that observed for control nonsmoking healthy young subjects, aged 18 to 22 years (average = 19.9), who were not occupationally exposed, and had no history of prior or concurrent malignancy. The study was approved by the local Ethics Committee (HC-FMRP Ethics committee Proc. 1135/2000).

The results obtained for the different markers are pooled and described in the following subsections.

1.1. ETV6/RUNX1 Fusions

FISH for t(12;21) ETV6/RUNX1 was carried out using the LSI TEL/AML1 ES probe (Vysis, Abbott Diagnostics, Maidenhead, United Kingdom) according to the manufacturers’ instructions: 1000 interphase nuclei were scored for each sample to determine the degree of positivity for the fusion and the interphase signal patterns of different cell populations. Numbers of red-green fusion signals were recorded corresponding to numbers of copies ETV6/RUNX1, while numerous red and green signals corresponding to RUNX1 and ETV6, respectively, were collectively considered as “extra signals.” Our study showed that ALL survivors ( ) presented significantly higher frequency of ETV6/RUNX1 fusions ( ) than those obtained for the control group ( ) ( ). Extra signals were also detected for both groups although frequencies were similar [20]. Gene fusions incidence in lymphocytes from the group of patients collected months or years after completion of therapy were then compared with those obtained retrospectively for fixed bone marrow samples collected at the time of diagnosis. Two out of ten samples studied were positive for the t(12;21) at diagnosis, however, when these samples were excluded from the statistical analysis, it was observed that the frequency of aberrations obtained for treated patients were significantly higher than those in the untreated bone marrow samples ( ) (Table 1). More interesting, the fusion gene frequency in bone marrow samples was similar to that observed for the control samples, suggesting an influence of previous exposure to anticancer drugs [20].

tab1
Table 1: Mean frequency of ETV6/RUNX1 translocations and extra signals obtained for ALL survivors, healthy individuals, and patients at the time of initial diagnosis [20].

Also, sequential FISH was performed on lymphocytes from ALL survivors, using chromosome specific library probes for chromosome 12. For one thousand metaphases scored, the frequency of translocations varied between zero and 7.6 ( ) for the group of patients while for the control group, the frequency was between zero and 5.5/1000 metaphases ( ) (Table 2). The statistical analysis revealed a significant difference ( ) between the patients and the control group, in accordance with the previous results using locus specific probes for ETV6/RUNX1, providing further evidence that chemotherapy had induced an increase in genomic instability manifested as de novo chromosome aberrations in peripheral blood lymphocytes sampled over years (most cases) after therapy completion.

tab2
Table 2: Translocation frequency for chromosome 12 obtained for controls and childhood ALL survivors. Peripheral blood lymphocyte samples were collected from the patients at variable periods after therapy conclusion.

For both types of probes the role of other covariates such as age at initial diagnosis, inclusion of prophylactic irradiation, and time passed after therapy conclusion were statistically analyzed, showing that neither of these parameters influenced the incidence of chromosomal aberrations.

1.2. MLL Fusions

FISH assessment for the presence of aberrations involving the MLL gene (11q23) was performed using the commercially available probes LSI MLL Break Apart Rearrangement according to the protocol of the manufacturer (Vysis). The expected signal pattern for a normal cell nucleus is two green(yellow)orange signals. In cells harboring MLL translocations, the green and orange signals appear separated without the yellow intersection allowing detection of any translocation at 11q23 irrespectively of the partner involved. One thousand metaphases from 49 ALL long-term survivors and the same number from control individuals were analyzed. Since MLL aberrations are characteristic of secondary leukemias associated with therapy with topoisomerase II inhibitors [21], the group of patients was divided into two subgroups depending on the inclusion or omission of VP-16 and/or VM-26 in the treatment protocol. Our results showed significant ( ) differences between the frequency of translocations observed for the groups of patients and controls. These differences were also significant ( ) when the groups of patients (independently of the inclusion of VP-16 and/or VM-26) and controls were compared, indicating that higher frequency in the patients cohort might be associated with the therapy with antitumoral drugs, independently of the inclusion of topoisomerase II inhibitors. Extra signals were also scored, however, their frequency did not differ between groups of patients and controls (Table 3) [19].

tab3
Table 3: Mean frequency of MLL translocations and extra signals obtained for ALL survivors and healthy individuals (taken from [19]).

In parallel, MLL translocations were analyzed by Inverse-PCR and identified by subsequent cloning and sequencing. This methodology is specifically applicable for the identification of rearrangements with known sequence, but whose end could be represented by a myriad of translocation partners as is the case of MLL fusions. Basically, DNA is digested by proper restriction enzymes, circularized and then amplified with divergent primers, detecting any segment flanking a known DNA sequence [22]. Individual PCR products were cut, extracted from gels, and subsequently cloned into the pGEM-T vector, further sequencing of PCR products was achieved in order to confirm the veracity of the rearrangements. Eighty-one percent of patients (35 out of 43) presented putative translocations; from those, 91% corresponded with t(4;11)(q21;q23), while the other 9% corresponded with t(X;11), t(8;11)(q23;q23), and t(11;16). The control group was also analyzed by this technique (   =  100), showing putative MLL rearrangements in 49% of individuals. The presence of two or three putative translocations was statistically higher in the group of patients ( ) when compared with the control group. The results were also analyzed regarding the time in complete clinical remission, which did not influence the results. Similarly, the analysis considering other variables (sex, risk of relapse, and inclusion of radiotherapy) did not influence the number of translocations between patients [19].

1.3. IGH/CCND1 Fusions

Locus-specific identifier dual-color directly labeled IGH(spectrum Green)/CCND1(Spectrum-Orange) (Vysis) probes were used for detecting the t(11;14)(q13;q32) originated by the juxtaposition of the immunoglobulin gene (IGH) at 14q32 with the CCND1 gene (cyclin D1, BCL1, or PRAD1), localized at 11q13. The expected signal pattern for a nucleus with t(11;14) is one green (normal IGH), one red (normal CCND1), and two red(yellow)green signals (1G1R2Y), due to the fact that the break usually occurs in the middle of the probe marked region, resulting in two fragments of each gene, however, these fragments can be lost, or extra signals of each gene can be detected, resulting in different patterns of signals (2G1R1Y, 1G2R1Y; or 3G2R, 2G3R, in the case of an extra signal). At least 1000 interphase nuclei were analyzed per individual. The group of patients in complete remission ( ) presented frequency of fusions varying between 0.09 and 0.49/100 cells. These values were significantly higher ( ) compared with those observed for the control group ( ), which varied from 0 to 0.29 fusions/100 cells. The probes also allowed the detection of extra signals for both genes, with frequency varying from 0.09 to 0.69 signals/100 cells for patients and from 0 to 0.48 signals/100 cells for controls. As seen for IGH/CCND1 fusions, the frequency of extra signals presented by the group of patients in complete remission were significantly higher ( ) than those presented by healthy individuals. When the inclusion of preventive cranial irradiation was considered no differences in IGH/CCND1 fusions frequency and extra signals were observed between of patients in complete remission and the control group.

A retrospective analysis was also carried out with sequential bone marrow samples collected at the moment of diagnosis from nine of the fourteen ALL patients, with the aim of determining whether the fusion IGH/CCND1 was already present before treatment. The fusion appeared with frequency varying from 0 to 0.41 IGH/CCND1 fusions/100 cells and from 0 to 2.6 extra signals/100 cells. Interestingly, among these samples, only one patient showed high frequency of extra signals (2.6/100 cells), which might represent rearrangements of the IGH gene with another partner (Table 4).

tab4
Table 4: Mean frequency of IGH/CCND1 translocations and extra signals obtained for ALL survivors, healthy individuals, and patients at the time of initial diagnosis.

The analysis of other variables such as risk of relapse and inclusion of radiotherapy did not show differences in the frequency of IGH/CCND1 translocations between patients.

1.4. BCR/ABL1 Fusions

FISH for t(9;22) that juxtaposes the ABL gene at 9q34 and the BCR gene at 22q11.2 was carried out using the LSI BCR/ABL ES probe (Vysis) according to the manufacturers’ instructions. The hybridization pattern for these probes is two orange and two green signals for normal cells (2O2G). Cells with t(9;22) involving the M-bcr show one large orange, one smaller orange (ES), and one fused orange/green signal (2O1G1F), while for translocations involving the minor breakpoint (m-bcr) appear as one orange, one green, and two fusion signals (1O1G2F) are observed. In this case, samples from 8 ALL survivors and the same number of control individuals were analyzed. The cytogenetic analysis of 1000 metaphases showed translocation frequency varying from 0.236 to 1.627 for treated patients ( ) and between zero and 1.30 for the control group ( ). Extra signals were also detected with frequency varying from 0.59 to 3.69 and zero to 0.9 for patients and controls, respectively. However, the statistical analysis was unable to detect differences for translocations and extra signals between the studied groups ( ). A retrospective analysis with sequential samples obtained at the time of diagnosis was also performed ( ). Translocation incidence varied from 0.8 to 1.54, while extra signals varied from 1.10 to 15.56; however, this group was not big enough to establish comparisons with the frequency observed the other groups (Table 5).

tab5
Table 5: Mean frequency of BCR/ABL translocations and extra signals obtained for ALL survivors, healthy individuals, and patients at the time of initial diagnosis.

2. Discussion

Since 1970, the number of cancer survivors in developed countries has tripled and is growing by 2% each year [23]. Cancer treatments have become much more aggressive as new agents and better supportive therapies are in constant development. The long-term consequences of cancer cure may be significant for the patients who often experience physical or physiological late effects secondary to their previous cancer treatment [4]. Many of these effects may not occur or become clinically symptomatic until many years after completion of therapy, though, the development of a second malignant neoplasm can be the most devastating late effect of cancer treatment.

Childhood-cancer survivors present a higher risk of developing secondary malignancies [24] which characterize the leading cause of death among this cohort [25]. The time of occurrence has not been established, but in general between 3 and 12 percent of the patients develop a subsequent cancer within the first 20 years after therapy conclusion. The most widely recognized treatment-related risk factors are alkylating agents and topoisomerase II inhibitors (epipodophyllotoxins and anthracyclines). The magnitude of the risk associated with these factors depends on several variables such as initial diagnosis, age at diagnosis, the administration schedule, concomitant medications, and genetic predisposition among others [26]. Exposure to alkylating agents or topoisomerase inhibitors during therapy has also been associated with higher risk of developing secondary leukemia among ALL cured patients [27].

Treatment of pediatric ALL consists of several doses of anticancer drugs given weekly or daily for 2-3 years [28]. These drugs cause DNA breaks, facilitating the formation of chromosome rearrangements [4, 29]. Thus, like de novo leukemias, therapy-related hematological disorders are commonly associated with recurrent aberrations [7]. Chromosome(s) 7 and/or 5 monosomies or deletions are typical of alkylating agent-induced AML, while balanced translocations involving chromosome bands 11q23 and 21q22 are associated to previous therapy with DNA-topoisomerase II inhibitors [23] which usually lead to the formation of gene fusions involving MLL [21] and RUNX1 [30], respectively. Translocations involving the latter have been identified in therapy-related secondary leukemias [31] with partners such as ETO (Eight Twenty One) and EVI1 (Ecotropic Viral Integration site 1 oncogene) [32]. Other numerical and structural rearrangements have also been described in different secondary therapy-related neoplasms although at lower frequency, including t(3;21) [33], t(9;22) BCR-ABL1 [34], t(1;21), t(3;21), t(14;21), t(15;21), and t(17;21) [35].

In the present paper, we pooled a screening for structural aberrations (ETV6/RUNX1, MLL fusions, IGH/CCND1, and BCR/ABL1) in ALL long-term survivors, compared with control individuals, as evaluated by FISH on interphase nuclei using a specific commercial DNA probes. As shown in Figure 1, all the markers studied showed higher frequency when compared to the control group, thus providing evidence that chemotherapy could have induced an increase in genomic instability manifested as de novo chromosome aberrations in peripheral blood lymphocytes sampled over years (most cases) after therapy completion. The differences between groups were statistically different for the ETV6/RUNX1, MLL fusions, and IGH/CCND1 markers. In the case of BCR/ABL1, although the rearrangement appeared at higher frequency in ALL survivors, the study groups analyzed were not big enough for the proper comparison.

230481.fig.001
Figure 1: Mean frequency of gene fusions in peripheral lymphocytes from ALL survivors analyzed months or year(s) after therapy completion (black boxes) and healthy donors (white boxes). In the case of MLL fusions the group of survivors was subdivided into patients treated with topoisomerase II inhibitors (grey box) and patients treated without the inclusion of such drugs in the treatment protocol (black box). . Data for ETV6/RUNX1 and MLL was taken from Brassesco et al. [20] and [19], respectively.

Mutational studies at the FMS proto-oncogene demonstrated the acquisition of mutation in patients treated with alkylating agents and radiation [3]. The absence of such mutations in biopsies at initial diagnosis suggested that they were somatically acquired after treatment and they appeared with increased frequency in relation to the normal population not exposed to cytotoxic drugs. Similarly, studies of HRPT (hypoxanthine phosphoribosyltransferase) mutations demonstrated significantly elevated frequency in patients treated for pre-B ALL, even two years after therapy completion [12].

Furthermore, rearrangements at 11q23 have been found during treatment of primary cancer, as observed in case of t(11;17) MLL/GASP [36]. Other chromosomal rearrangements such as PML-RARA, CBFB-MYH11, MLL-MLLT1, and BCR-ABL1 have also been observed in Non-Hodgkin Lymphoma patients after high-dose therapy [37] demonstrating the susceptibility of specific genes to mutagenic or carcinogenic agents [38].

Indeed, although translocations might be formed by different mechanisms, several studies have mapped specific chromatin structural elements, such as in vivo topoisomerase II (topo II) cleavage sites, DNAse I hypersensitive sites and scaffold attachment regions (SARs) in the breakpoint regions of relevant genes (including MLL, AF4, AF9, AML1, ETO, CBFB, MYHI1, PML, RARA, TEL, E2A, PBX1, BCR, and ABL) that are probably associated with preferential breakage after exposure to damage including topoisomerase II inhibitors [39]. In most cases, the breakpoints do not colocalize exactly with topoisomerase II cleavage sites, but map close to or at a variable distance from them. After the induction of DNA cleavage at these sites, the religation of the double strand breaks occurs by the action of complex DNA repair mechanisms, mainly nonhomologous end joining (NHEJ) [39].

Other mechanisms such as illegitimate recombination mediated by Alu repeats and long interspersed nuclear elements (LINEs) have been proposed to participate in the formation of gene fusions [40, 41]. Illegitimate V(D)J recombination have also been described, with site-specific DNA cleavage at heptamer/nonamer signal sequences with the IGH variable region or the TCR binding region arranged in juxtaposition with other tumor suppressors genes or oncogenes like MYC or CCND1 in lymphoid malignancies [42, 43]; besides several studies have demonstrated that the exposure to DNA-damaging agents in vitro increase V(D)J rearrangements frequency at cryptic signal sequences other than IGH and TCR loci [44, 45].

In addition, the induction of leukemia-associated fusion genes such as t(8;21) AML1/ETO, t(9;22) BCR/ABL, and t(6;9) DEK/CAM (associated with AML and CML) by high doses of ionizing radiation has been reported in vitro [46] and in patients accidentally exposed to ionizing radiation [47, 48].

According to the literature, the inclusion of radiotherapy in different treatment regimens is associated with an increase (2-3 times) in the risk of developing solid tumors [49]. Among the ALL survivors analyzed in the present study, 13 (28%) were submitted to different doses of cranial or neuroaxis -radiation. However, these patients did not show increased translocation frequency (for any marker) compared with patients who were only treated with chemotherapy.

Several authors have demonstrated that the production of unstable aberrations (rings and dicentrics) after radiotherapy decreases with time, and sequential cytogenetic studies performed on peripheral lymphocytes taken from patients treated for different cancers showed a constant decline in chromosomal aberrations [50]. M’kacher et al. [51] demonstrated by chromosome painting that after an initial decrease for up to 6 months after treatment, the frequency of complex rearrangements remained constant for up to 2 years during the follow-up of Hodgkin’s lymphoma survivors. In our study, the period after therapy conclusion varied between 5 and 192 months (median event-free survival was 3.8 years), however, the statistical analysis did not show any significant relationship between the frequency of translocations for any of the markers and time passed after therapy conclusion.

In the last years, the treatment directed to the central nervous system has suffered modifications with a tendency of lowering doses and the irradiated area, using dose rationing regimens [52]. In general, protocols have eliminated radiation to the spine, decreased cranial irradiation doses or replaced them with intrathecal and/or systemic drugs such as high-dose methotrexate [5]. Currently, ALL patients at our service are submitted to doses of 18 Gy (classified as standard risk of relapse) and 24 Gy (higher risk of relapse), according to the GBTLI protocol [18]. Nonetheless, several epidemiological studies have demonstrated that the risk of t-LMAs or t-MDSs is predominantly associated with chemotherapy and not to radiotherapy, principally emphasizing the relationship between specific drugs and chromosome rearrangements [53], as verified for topoisomerase II inhibitors associated to 11q23 aberrations.

When considering the MLL aberrations, the group of patients presented significantly higher aberration frequency when compared to control individuals, although such differences were independent of the inclusion of VP-16 or VM-26 in the treatment protocol. Some studies have demonstrated that concomitant adjuvant chemotherapy (such as asparaginase, doxorubicin, cyclophosphamide) or antimetabolites (such as mercaptopurine or methotrexate) administration, increase the risk of secondary leukemia [26].

Also, individual predisposing factors, including polymorphisms of detoxification and DNA-repair enzymes that, for instance, reduce the enzymatic activity of thiopurine methyltransferase, a variant of CYP3A that affects production of a DNA-damaging metabolite of epipodophyllotoxin. Other polymorphisms in glutathione S-transferase and NAD(P)H:quinine oxidoreductase (NQO1) are as well associated with increased risk of secondary leukemia after chemotherapy [23].

Since strategies in cancer treatment are continuously evolving, the investigation of rearrangements that could represent a risk factor for the development of secondary neoplasias is important for cancer survivors. Individuals with a history of childhood cancer have 10–20 fold greater risk of developing a secondary malignant neoplasm within the first 20 years after treatment [54] turning therapy-related neoplasias a worldwide problem of major proportions [55, 56].

Between 10% and 20% of acute leukemia patients reported in various series have developed a secondary cancer following previous therapy, and considering cured ALL patients, a greater frequency of neoplasias has been observed when compared to normal individuals [8, 57, 58]. None of our patients developed a secondary cancer over 5 years of clinical follow-up. Yet, even though the chromosome aberrations detected in this study appear at low frequency in ALL survivors, there were significant differences when compared to the control group.

Although some studies have demonstrated that some of these fusion genes can lead to a greater cellular proliferation, like BCR/ABL, ETV6/RUNX1, IGH/BCL-2, among others, such alterations by themselves are not enough for tumorigenesis, and secondary multiple genetic alterations are necessary [5961]. Recent studies have also demonstrated the presence of leukemia-related chromosomal translocations in normal individuals, such as t(14;18) IGH/BCL2 [6264] and t(9;22) [65], t(12;21) ETV6/RUNX1 [14], t(11;14)(p13;q11) LMO2/TCR and t(7;14)(q34;q11) TCR/TAL2 [42], t(15;17) PML/RARA [66], and MLL partial duplications [15, 67], indicating that the presence of such translocations per se does not define an apparent clinical disease.

Although, it cannot be assumed that the observed therapy-induced genetic damage in the group of ALL survivors is predictive of an increased risk of developing secondary neoplasias, cytogenetic surveillance should be considered to evaluate the occurrence of persistent genome instability in these patients, as well as to associate particular evolving genomic aberrations to the potential risk of secondary malignancies in this cohort.

Acknowledgment

This work was supported by FAPESP (Grant nos. 01/11225-3, 02/13317-8, and 03/01915-0), CAPES, and CNPq.

References

  1. A. Baccichet, S. K. Qualman, and D. Sinnett, “Allelic loss in childhood acute lymphoblastic leukemia,” Leukemia Research, vol. 21, no. 9, pp. 817–823, 1997. View at Publisher · View at Google Scholar · View at Scopus
  2. C.-H. Pui, L. L. Robison, and A. T. Look, “Acute lymphoblastic leukaemia,” The Lancet, vol. 371, no. 9617, pp. 1030–1043, 2008. View at Publisher · View at Google Scholar · View at Scopus
  3. A. Baker, P. Cachia, S. Ridge et al., “FMS mutations in patients following cytotoxic therapy for lymphoma,” Leukemia Research, vol. 19, no. 5, pp. 309–318, 1995. View at Publisher · View at Google Scholar · View at Scopus
  4. J. Byrne, “Long-term genetic and reproductive effects of ionizing radiation and chemotherapeutic agents on cancer patients and their offspring,” Teratology, vol. 59, no. 4, pp. 210–215, 1999. View at Publisher · View at Google Scholar · View at Scopus
  5. P. C. Nathan, K. Wasilewski-Masker, and L. A. Janzen, “Long-term outcomes in survivors of childhood acute lymphoblastic leukemia,” Hematology/Oncology Clinics of North America, vol. 23, no. 5, pp. 1065–1082, 2009. View at Publisher · View at Google Scholar · View at Scopus
  6. M. A. Smith, L. Rubinstein, and R. S. Ungerleider, “Therapy-related acute myeloid leukemia following treatment with epipodophyllotoxins: estimating the risks,” Medical and Pediatric Oncology, vol. 23, no. 2, pp. 86–98, 1994. View at Publisher · View at Google Scholar · View at Scopus
  7. C. A. Felix, “Secondary leukemias induced by topoisomerase-targeted drugs,” Biochimica et Biophysica Acta, vol. 1400, no. 1–3, pp. 233–255, 1998. View at Publisher · View at Google Scholar · View at Scopus
  8. S. Bhatia, H. N. Sather, O. B. Pabustan, M. E. Trigg, P. S. Gaynon, and L. L. Robison, “Low incidence of second neoplasms among children diagnosed with acute lymphoblastic leukemia after 1983,” Blood, vol. 99, no. 12, pp. 4257–4264, 2002. View at Publisher · View at Google Scholar · View at Scopus
  9. E. J. Dann and J. M. Rowe, “Biology and therapy of secondary leukaemias,” Best Practice and Research, vol. 14, no. 1, pp. 119–137, 2001. View at Publisher · View at Google Scholar · View at Scopus
  10. S. Bonassi, L. Hagmar, U. Strömberg et al., “Chromosomal aberrations in lymphocytes predict human cancer independently of exposure to carcinogens,” Cancer Research, vol. 60, no. 6, pp. 1619–1625, 2000. View at Scopus
  11. H. Norppa, “Cytogenetic biomarkers and genetic polymorphisms,” Toxicology Letters, vol. 149, no. 1-3, pp. 309–334, 2004. View at Publisher · View at Google Scholar · View at Scopus
  12. S. Koishi, M. Kubota, M. Sawada et al., “Biomarkers in long survivors of pediatric acute lymphoblastic leukemia patients: late effects of cancer chemotherapy,” Mutation Research, vol. 422, no. 2, pp. 213–222, 1998. View at Publisher · View at Google Scholar · View at Scopus
  13. M. Greaves, “Molecular genetics, natural history and the demise of childhood leukaemia,” European Journal of Cancer, vol. 35, no. 2, pp. 173–185, 1999. View at Publisher · View at Google Scholar · View at Scopus
  14. M. Eguchi-Ishimae, M. Eguchi, E. Ishii et al., “Breakage and fusion of the TEL (ETV6) gene immature B lymphocytes induced by apoptogenic signals,” Blood, vol. 97, no. 3, pp. 737–743, 2001. View at Publisher · View at Google Scholar · View at Scopus
  15. J. Bäsecke, F. Griesinger, L. Trümper, and G. Brittinger, “Leukemia- and lymphoma-associated genetic aberrations in healthy individuals,” Annals of Hematology, vol. 81, no. 2, pp. 64–75, 2002. View at Publisher · View at Google Scholar · View at Scopus
  16. S. Lipkowitz, V. F. Garry, and I. R. Kirsch, “Interlocus V-J recombination measures genomic instability in agriculture workers at risk for lymphoid malignancies,” Proceedings of the National Academy of Sciences of the United States of America, vol. 89, no. 12, pp. 5301–5305, 1992. View at Publisher · View at Google Scholar · View at Scopus
  17. I. R. Kirsch and F. Lista, “Lymphocyte-specific genomic instability and risk of lymphoid malignancy,” Seminars in Immunology, vol. 9, no. 3, pp. 207–215, 1997. View at Publisher · View at Google Scholar · View at Scopus
  18. S. Brandalise, V. Odone, W. Pereira, M. Andrea, M. Zanichelli, and V. Aranega, “Treatment results of three consecutive Brazilian cooperative childhood ALL protocols: GBTLI-80, GBTLI-82 and -85,” Leukemia, vol. 7, supplement 2, pp. S142–S145, 1993. View at Scopus
  19. M. S. Brassesco, A. P. Montaldi, D. E. Gras et al., “Cytogenetic and molecular analysis of MLL rearrangements in acute lymphoblastic leukaemia survivors,” Mutagenesis, vol. 24, no. 2, pp. 153–160, 2009. View at Publisher · View at Google Scholar · View at Scopus
  20. M. S. Brassesco, M. L. Camparoto, L. G. Tone, and E. T. Sakamoto-Hojo, “Analysis of ETV6/RUNX1 fusions for evaluating the late effects of cancer therapy in ALL (acute lymphoblastic leukemia) cured patients,” Cytogenetic and Genome Research, vol. 104, no. 1-4, pp. 346–351, 2004. View at Publisher · View at Google Scholar · View at Scopus
  21. P. D. Aplan, D. S. Chervinsky, M. Stanulla, and W. C. Burhans, “Site-specific DNA cleavage within the MLL breakpoint cluster region induced by topoisomerase II inhibitors,” Blood, vol. 87, no. 7, pp. 2649–2658, 1996. View at Scopus
  22. C. J. Betti, M. J. Villalobos, M. O. Diaz, and A. T. M. Vaughan, “Apoptotic triggers initiate translocations within the MLL gene involving the nonhomologous end joining repair system,” Cancer Research, vol. 61, no. 11, pp. 4550–4555, 2001. View at Scopus
  23. G. Leone, L. Fianchi, L. Pagano, and M. T. Voso, “Incidence and susceptibility to therapy-related myeloid neoplasms,” Chemico-Biological Interactions, vol. 184, no. 1-2, pp. 39–45, 2010. View at Publisher · View at Google Scholar · View at Scopus
  24. A. C. MacArthur, J. J. Spinelli, P. C. Rogers, K. J. Goddard, N. Phillips, and M. L. McBride, “Risk of a second malignant neoplasm among 5-year survivors of cancer in childhood and adolescence in British Columbia, Canada,” Pediatric Blood and Cancer, vol. 48, no. 4, pp. 453–459, 2007. View at Publisher · View at Google Scholar · View at Scopus
  25. S. C. W. Lawless, P. Verma, D. M. Green, and M. C. Mahoney, “Mortality experiences among 15+ year survivors of childhood and adolescent cancers,” Pediatric Blood and Cancer, vol. 48, no. 3, pp. 333–338, 2007. View at Publisher · View at Google Scholar · View at Scopus
  26. N. Hijiya, K. K. Ness, R. C. Ribeiro, and M. M. Hudson, “Acute leukemia as a secondary malignancy in children and adolescents: current findings and issues,” Cancer, vol. 115, no. 1, pp. 23–35, 2009. View at Publisher · View at Google Scholar · View at Scopus
  27. L. F. Lopes, B. Camargo, and A. Bianchi, “Late effects of childhood cancer treatment,” Revista da Associacao Medica Brasileira, vol. 46, no. 3, pp. 277–284, 2000. View at Scopus
  28. J. G. Blanco, T. Dervieux, M. J. Edick et al., “Molecular emergence of acute myeloid leukemia during treatment for acute lymphoblastic leukemia,” Proceedings of the National Academy of Sciences of the United States of America, vol. 98, no. 18, pp. 10338–10343, 2001. View at Publisher · View at Google Scholar · View at Scopus
  29. Y.-H. Han, M. J. F. Austin, Y. Pommier, and L. F. Povirk, “Small deletion and insertion mutations induced by the topoisomerase II inhibitor teniposide in CHO cells and comparison with sites of drug-stimulated DNA cleavage in vitro,” Journal of Molecular Biology, vol. 229, no. 1, pp. 52–66, 1993. View at Publisher · View at Google Scholar · View at Scopus
  30. M. Stanulla, J. Wang, D. S. Chervinsky, S. Thandla, and P. D. Aplan, “DNA cleavage within the MLL breakpoint cluster region is a specific event which occurs as part of higher-order chromatin fragmentation during the initial stages of apoptosis,” Molecular and Cellular Biology, vol. 17, no. 7, pp. 4070–4079, 1997. View at Scopus
  31. J. D. Rowley, “The role of chromosome translocations in leukemogenesis,” Seminars in Hematology, vol. 36, no. 4, pp. 59–72, 1999. View at Scopus
  32. M. L. Loh, L. B. Silverman, M. L. Young et al., “Incidence of TEL/AML1 fusion in children with relapsed acute lymphoblastic leukemia,” Blood, vol. 92, no. 12, pp. 4792–4797, 1998. View at Scopus
  33. S. W. Hiebert, W. Sun, J. N. Davis et al., “The t(12;21) translocation converts AML-1B from an activator to a repressor of transcription,” Molecular and Cellular Biology, vol. 16, no. 4, pp. 1349–1355, 1996. View at Scopus
  34. M. Hattori, M. Tanaka, Y. Yamazaki, Y. Nakahara, K. Tsushita, and M. Utumi, “Detection of major and minor bcr/abl fusion gene transcripts in a patient with acute undifferentiated leukemia secondary to treatment with an alkylating agent,” Leukemia Research, vol. 19, no. 6, pp. 389–396, 1995. View at Publisher · View at Google Scholar · View at Scopus
  35. D. Roulston, R. Espinosa III, G. Nucifora, R. A. Larson, M. M. Le Beau, and J. D. Rowley, “CBFA2(AML1) translocations with novel partner chromosomes in myeloid leukemias: association with prior therapy,” Blood, vol. 92, no. 8, pp. 2879–2885, 1998. View at Scopus
  36. M. D. Megonigal, N.-K. V. Cheung, E. F. Rappaport et al., “Detection of leukemia-associated MLL-GAS7 translocation early during chemotherapy with DNA topoisomerase II inhibitors,” Proceedings of the National Academy of Sciences of the United States of America, vol. 97, no. 6, pp. 2814–2819, 2000. View at Publisher · View at Google Scholar · View at Scopus
  37. J. Basecke, M. Podleschny, A. Becker et al., “Therapy-associated genetic aberrations in patients treated for non-Hodgkin lymphoma,” British Journal of Haematology, vol. 141, no. 1, pp. 52–59, 2008. View at Publisher · View at Google Scholar · View at Scopus
  38. K. Seeger, H.-P. Adams, D. Buchwald et al., “TEL-AML1 fusion transcript in relapsed childhood acute lymphoblastic leukemia,” Blood, vol. 91, no. 5, pp. 1716–1722, 1998. View at Scopus
  39. Y. Zhang and J. D. Rowley, “Chromatin structural elements and chromosomal translocations in leukemia,” DNA Repair, vol. 5, no. 9-10, pp. 1282–1297, 2006. View at Publisher · View at Google Scholar · View at Scopus
  40. M. P. Strout, G. Marcucci, C. D. Bloomfield, and M. A. Caligiuri, “The partial tandem duplication of ALL1 (MLL) is consistently generated by Alu-mediated homologous recombination in acute myeloid leukemia,” Proceedings of the National Academy of Sciences of the United States of America, vol. 95, no. 5, pp. 2390–2395, 1998. View at Publisher · View at Google Scholar · View at Scopus
  41. S. P. Whitman, M. P. Strout, M. Guido et al., “The partial nontandem duplication of the MLL (ALL1) gene is a novel rearrangement that generates three distinct fusion transcripts in B-cell acute lymphoblastic leukemia,” Cancer Research, vol. 61, no. 1, pp. 59–63, 2001. View at Scopus
  42. R. Marculescu, T. Le, P. Simon, U. Jaeger, and B. Nadel, “V(D)J-mediated translocations in lymphoid neoplasms: a functional assessment of genomic instability by cryptic sites,” Journal of Experimental Medicine, vol. 195, no. 1, pp. 85–98, 2002. View at Publisher · View at Google Scholar · View at Scopus
  43. A. K. Abbas, A. H. Lichtman, and J. S. Pober, Cellular and Molecular Immunology, W. B. Saunders, Philadelphia, Pa, USA, 5th edition, 2003.
  44. C.-L. Chen, J. C. Fuscoe, Q. Liu, and M. V. Relling, “Etoposide causes illegitimate V(D)J recombination in human lymphoid leukemic cells,” Blood, vol. 88, no. 6, pp. 2210–2218, 1996. View at Scopus
  45. R. L. Pinsonneault, P. M. Vacek, J. P. O'Neill, and B. A. Finette, “Induction of V(D)J-mediated recombination of an extrachromosomal substrate following exposure to DNA-damaging agents,” Environmental and Molecular Mutagenesis, vol. 48, no. 6, pp. 440–450, 2007. View at Publisher · View at Google Scholar · View at Scopus
  46. M. W. N. Deininger, S. Bose, J. Gora-Tybor, X.-H. Yan, J. M. Goldman, and J. V. Melo, “Selective induction of leukemia-associated fusion genes by high-dose ionizing radiation,” Cancer Research, vol. 58, no. 3, pp. 421–425, 1998. View at Scopus
  47. R. Hromas, R. Shopnick, H. G. Jumean, C. Bowers, M. Varella-Garcia, and K. Richkind, “A novel syndrome of radiation-associated acute myeloid leukemia involving AML1 gene translocations,” Blood, vol. 95, no. 12, pp. 4011–4013, 2000. View at Scopus
  48. R. Hromas, T. Busse, A. Carroll et al., “Fusion AML1 transcript in a radiation-associated leukemia results in a truncated inhibitory AML1 protein,” Blood, vol. 97, no. 7, pp. 2168–2170, 2001. View at Publisher · View at Google Scholar · View at Scopus
  49. K. Kawai and H. Akaza, “Treatment of testicular and second cancer,” Gan to Kagaku Ryoho, vol. 26, no. 13, pp. 2021–2028, 1999. View at Scopus
  50. E. J. Tawn, C. A. Whitehouse, and F. A. Martin, “Sequential chromosome aberration analysis following radiotherapy—no evidence for enhanced genomic instability,” Mutation Research, vol. 465, no. 1-2, pp. 45–51, 2000. View at Publisher · View at Google Scholar · View at Scopus
  51. R. M'kacher, T. Girinsky, S. Koscielny et al., “Baseline and treatment-induced chromosomal abnormalities in peripheral blood lymphocytes of Hodgkin's lymphoma patients,” International Journal of Radiation Oncology Biology Physics, vol. 57, no. 2, pp. 321–326, 2003. View at Publisher · View at Google Scholar · View at Scopus
  52. J. Nachman, H. N. Sather, J. M. Cherlow et al., “Response of children with high-risk acute lymphoblastie leukemia treated with and without cranial irradiation: a report from the Children's Cancer Group,” Journal of Clinical Oncology, vol. 16, no. 3, pp. 920–930, 1998. View at Scopus
  53. M. K. Andersen, B. Johansson, S. O. Larsen, and J. Pedersen-Bjergaard, “Chromosomal abnormalities in secondary MDS and AML. Relationship to drugs and radiation with specific emphasis on the balanced rearrangements,” Haematologica, vol. 83, no. 6, pp. 483–488, 1998. View at Scopus
  54. L. Löning, M. Zimmermann, A. Reiter et al., “Secondary neoplasms subsequent to Berlin-Frankfurt-Munster therapy of acute lymphoblastic leukemia in childhood: significantly lower risk without cranial radiotherapy,” Blood, vol. 95, no. 9, pp. 2770–2775, 2000. View at Scopus
  55. L. L. Robison and S. Bhatia, “Late-effects among survivors of leukaemia and lymphoma during childhood and adolescence,” British Journal of Haematology, vol. 122, no. 3, pp. 345–359, 2003. View at Publisher · View at Google Scholar · View at Scopus
  56. J. Jazbec, P. Ećimović, and B. Jereb, “Second neoplasms after treatment of childhood cancer in Slovenia,” Pediatric Blood and Cancer, vol. 42, no. 7, pp. 574–581, 2004. View at Publisher · View at Google Scholar · View at Scopus
  57. S. Bhatia, L. L. Robison, O. Oberlin et al., “Breast cancer and other second neoplasms after childhood Hodgkin's disease,” New England Journal of Medicine, vol. 334, no. 12, pp. 745–751, 1996. View at Publisher · View at Google Scholar · View at Scopus
  58. R. Mody, S. Li, D. C. Dover et al., “Twenty-five-year follow-up among survivors of childhood acute lymphoblastic leukemia: a report from the Childhood Cancer Survivor Study,” Blood, vol. 111, no. 12, pp. 5515–5523, 2008. View at Publisher · View at Google Scholar · View at Scopus
  59. K. F. Wong, “11q13 is a cytogenetically promiscuous site in hematologic malignancies,” Cancer Genetics and Cytogenetics, vol. 113, no. 1, pp. 93–95, 1999. View at Publisher · View at Google Scholar · View at Scopus
  60. F. Bernardin, Y. Yang, R. Cleaves et al., “TEL-AML1, expressed from t(12;21) in human acute lymphocytic leukemia, induces acute leukemia in mice,” Cancer Research, vol. 62, no. 14, pp. 3904–3908, 2002. View at Scopus
  61. D. De Jong, “Molecular pathogenesis of follicular lymphoma: a cross talk of genetic and immunologic factors,” Journal of Clinical Oncology, vol. 23, no. 26, pp. 6358–6363, 2005. View at Publisher · View at Google Scholar · View at Scopus
  62. Y. Liu, A. M. Hernandez, D. Shibata, and G. A. Cortopassi, “BCL2 translocation frequency rises with age in humans,” Proceedings of the National Academy of Sciences of the United States of America, vol. 91, no. 19, pp. 8910–8914, 1994. View at Publisher · View at Google Scholar · View at Scopus
  63. M. Yasukawa, S. Bando, G. Dölken et al., “Low frequency of BCL-2/JH translocation in peripheral blood lymphocytes of healthy Japanese individuals,” Blood, vol. 98, no. 2, pp. 486–488, 2001. View at Publisher · View at Google Scholar · View at Scopus
  64. K. E. Summers, L. K. Goff, A. G. Wilson, R. K. Gupta, T. A. Lister, and J. Fitzgibbon, “Frequency of the Bcl-2/IgH rearrangement in normal individuals: implications for the monitoring of disease in patients with follicular lymphoma,” Journal of Clinical Oncology, vol. 19, no. 2, pp. 420–424, 2001. View at Scopus
  65. S. Bose, M. Deininger, J. Gora-Tybor, J. M. Goldman, and J. V. Melo, “The presence of typical and atypical BCR-ABL fusion genes in leukocytes of normal individuals: biologic significance and implications for the assessment of minimal residual disease,” Blood, vol. 92, no. 9, pp. 3362–3367, 1998. View at Scopus
  66. A. S. Quina, P. Gameiro, M. S. D. Costa, M. Telhada, and L. Parreira, “PML-RARA fusion transcripts in irradiated and normal hematopoietic cells,” Genes Chromosomes and Cancer, vol. 29, no. 3, pp. 266–275, 2000. View at Scopus
  67. S. Schnittger, B. Wörmann, W. Hiddemann, and F. Griesinger, “Partial tandem duplications of the MLL gene are detectable in peripheral blood and bone marrow of nearly all healthy donors,” Blood, vol. 92, no. 5, pp. 1728–1734, 1998. View at Scopus