Secondary Acute Myeloid Leukemia (sAML) Immediately after Intensive Chemotherapy of Acute Lymphoblastic Leukemia (ALL)

A 20-year-old woman was admitted to our unit, because of pre-B acute lymphoblastic leukemia (ALL). Classical cytogenetics revealed a normal karyotype. In FISH analysis no TEL-AML1 fusion, rearranged MLL gene, or BCRABL were found. The induction and consolidation chemotherapy was conducted according to protocol PALG 5-2007. In the assessment of minimal residual disease using flow cytometry after consolidation 0.02% of the baseline phenotype of lymphoblasts were found. After two weeks of the completion of intensive chemotherapy, the patient was admitted in emergency to hematology inpatient unit, because of secondary acute myeloid leukemia (sAML). The conventional cytogenetic karyotype was complex: 46, XX, der (7) t (5;7;10;?)del(7)(q22), t(11;17)(p11;q21),del(14)(q24q32). After the diagnosis of sAML, induction chemotherapy was given according to the program PALG DAC: cytarabine 329 mg/d 1-7, daunorubicin 90 mg iv/d 1-3, cladribine 8 mg/d 1-5. The patient died due to progression of refractory leukemia in 45 days. The risk of developing of the sAML after treatment of ALL in adults is about 0.5-1% after few years. Our case was unique, because sAML developed rapidly, just after five months of ALL diagnosis, three months after CR1 and two weeks after completion of consolidation chemotherapy.

A 20-year-old Caucasian woman, previously healthy, was admitted to our department because of the growing weakness within four weeks that had begun with a viral infection of the upper respiratory tract. On admission, no signs of infection were stated. In physical examination, splenomegaly (3 cm from the left costal arch) was found. There were no hepatomegaly, lymphadenopathy, gingival hypertrophy, or mucocutaneous petechiae found in the physical examination. The complete blood count on admission was: WBC 44.87×10 9 /L (91% lymphoblasts in smear), Hgb 8.6 g/L, platelets 49×10 9 /L, 95% small homogeneous lymphoblasts infiltration of bone marrow. Flow cytometry performed on the bone marrow revealed pre-B acute lymphoblastic leukemia (ALL) with expression of CD19, CD10, CD34, TdT and cytoplasmatic IgM without co-expression of myeloid antigens (table 1). Conventional cytogenetics of bone marrow revealed a normal karyotype, 46, XX ( Figure 1). Twenty one metaphases were explored. In FISH analysis of bone marrow cells no TEL-AML1 fusion, rearranged MLL gene, or BCR-ABL was found ( Figure 2). The RT-PCR also did not confirm the presence of BCR-ABL and MLL/AF4.
After the diagnosis, induction chemotherapy was conducted according to the Polish Adult Leukemia Group protocol PALG 5-2007. The patient received prednisone 90 mg a day as pretreatment on days 1-7, then a total of 3 doses vincristine 2 mg/d iv on days 8, 15, 22, epirubicin 80 iv/d days 8, 15, 22, 4 times native L-asparaginase 10,000 IU/d, prednisone 90 mg p.o daily on days 1-28. During the induction chemotherapy period, intrathecal chemotherapy with pegylated cytarabine (DepoCyte) 50 mg was given only once due to thrombocytopenia. In the assessment of cerebrospinal fluid no deviations were found. Induction was reduced because of the profound neutropenia, febrile conditions, and coagulation disorders in the form of low levels of fibrinogen. During neutropenia related to the chemotherapy, granulocyte colony stimulating factor (G-CSF) 300 ug/d for 9 days was given. ALL was sensitive to steroids with reduction of WBC from 16 G/l to WBC 0.79 G/l after 7 days of pretreatment. On recovery, a subsequent bone marrow examination showed no evidence of residual disease (CR1) in accordance with WHO criteria.  Consolidation chemotherapy was also given consistently with the protocol PALG 5-2007. The patient received methotrexate 2.4 g iv/d for 2 days, etoposide 2×160 mg iv/d for 2 days, then cyclophosphamide 1600 mg iv/d for 4 days, cytarabine 4.8g/d for 2 days, the native L-asparaginase 10,000 IU iv/d 2 times were given. Afterwards, she received 3 time's triple intrathecal chemotherapy cytarabine 50 mg + methotrexate 15 mg + dexamethasone 4 mg. There were no deviations in the assessment of cerebrospinal fluid. Agranulocytosis period after chemotherapy was complicated by febrile states of the ongoing three days with symptoms of gastritis. In the assessment of minimal residual disease (MRD) using flow cytometry after consolidation 0.02% of the baseline phenotype of lymphoblasts, and besides, neutrophils 67.7%, myeloblasts 0.07%, lymphocytes 4.3%, erythroblasts 22.3% and monocytes 4.9% were found.
Retrospective studies of bone marrow cytogenetic preparations from the time of diagnosis of ALL using FISH, did not reveal the presence of above described aberrations.
In none of the analyzed 100 interphasal cells MLL, CBFB, RARA, IGH, gene rearrangements and deletion of TP53 gene locus were found. In the bone marrow studies by RT-PCR the presence of translocations AML1-ETO and CBFB-MYH11, as well as NPM1 mutations were excluded. The whole picture of genetic changes of myeloblasts indicates  the likely origin of secondary acute myeloid leukemia (sAML), which qualifies the patient to the group of a worse prognosis. However, the disease appeared extremely rapidly. LDH level was high and amounted to 601 U/L.
After the diagnosis of sAML, induction chemotherapy was given according to the program PALG DAC: cytarabine 329 mg c/d on days 1-7, daunorubicin 90 mg iv/d days 1-3, cladribine 8 mg iv/d days 1-5 [2,3]. Tolerability of treatment was good. Nadir of WBC 0.03×10 9 /L with neutrophils 0×10 9 /L was noted in 15 induction day. The period was complicated with post therapeutic agranulocytosis febrile states, without explicit signs of infection. In addition, coagulation abnormalities in the form of chronic low-level of fibrinogen and elevated D-dimers were observed. Patient required intensive substitution treatment. She received 10 units of PRBC, 6 units of FFP, 1 g of Fibrinogen, 40 units of the platelets. In 28 day of induction, the resistance of the type of regrowth of leukemic cells in the peripheral blood smear was shown. Supportive treatment was then carried out. The patient died due to progression of refractory leukemia in 45 days from induction beginning with symptoms of multi-organ failure.

Discussion
The risk of developing of the secondary cancer after treatment of acute lymphoblastic leukemia in children is estimated at 5-6% [4][5][6][7][8][9][10]. In adults, it is estimated at 9.4% after 15 years [11]. The risk of secondary hematological cancer is low and is estimated in adults at 2.36% after 30 years and 3.3% after 18 years of treatment.
Verma et al. [12] reports in the study of a total of 641 cases, the development of secondary acute myeloid leukemia in 6 patients (<1%) [12]. The disease evolved within a range of 13 to 74 (median 35.5) months since remission achievement after high dose chemotherapy hyper-CVAD. None of these patients had previous alloHSCT, one of them had radiotherapy. Only 1/16 patients (6%) achieved CR. OS was 6-16 months (median 8.25).
Tavernier, in the analysis of the ALL patients (treated within the studies of LALA group), found that the occurrence of secondary malignancies was 1.5% in patients after treatment of ALL, and AML accounted for 0.6%, that was the most common secondary neoplasm [11]. The development of secondary malignancies was estimated between 0.5 to 13.8 years (median 4.5 years) after diagnosis of ALL. Borgmann et al. [13] reports a similar incidence (0.5%) of the secondary AML in the group treated for ALL [13].
In our case, sAML developed rapidly, just after five months of ALL diagnosis, three months after CR1 and two weeks after completion of consolidation chemotherapy and after evaluation of MRD by flow cytometry at the amount of 0.02% and myeloblasts were calculated at 0.07%. The aberrations of the chromosome 5 and 7 long arms are quite typically found in sAML cases [1,[12][13][14][15]. However, in our patient only the aberrations involving short arms of these chromosomes were found. Together with balanced t(11;17) and 14q deletion, they formed complex, unbalanced karyotype with a very poor prognosis, which was confirmed by the absence of effect of DAC induction chemotherapy and the survival time was 45 days. Development of the secondary leukemia may be caused by many factors like biological nature of the primary tumor, type of chemotherapy and genetic predisposition. Etoposide and alkylating agents are indicated as possible causes of sAML, on account of leading to alterations within the chromosome 5 and 7 [1,[11][12][13][14]. In the described case, etoposide and cyclophosphamide were used, but in standard doses, which should not increase the risk of sAML. We suppose, there must have been a very deep defect in repair mechanisms.