Aspergillosis in immunocompromised children acute myeloid leukemia and bone marrow aplasia.: Report of two cases

Two cases of Aspergillosis in immunocompromised children are reported. Both were caused by Aspergillns flavus. Early diagnosis and treatment led to the remission of the process. One patient had acute myeloid leukemia; the fungus was isolated from the blood. The other patient with bone marrow aplasia, presented an invasive aspergillosis of the paranasal sinuses with dissemination of fungal infection; the diagnosis was obtained by histology and culture of biopsied tissue from a palatal ulceration.

Acute myeloid leukemia in elderly patients: experience of a single center

Acute myeloid leukemia (AML) is a disease predominantly of older adults. Treatment of AML in the elderly is complicated not only by comorbidities but also by the high prevalence of poor prognosis markers. Thirty-one consecutive unselected patients with AML older than 60 years (representing 33% of all AML cases diagnosed at our institution during the same period) were followed over a period of 5 years (1997-2002). A high incidence of AML with multilineage dysplasia (45%) and no favorable cytogenetic abnormalities but 62% intermediate and 38% unfavorable karyotypes were found. Sixteen patients (52%) were selected for induction of intensive cytotoxic treatment and complete remission was achieved only by some of these intensively treated patients (7 of 16). Of these, 3 remained alive without disease (median: 11 months), 1 patient died shortly after complete remission, and 3 patients relapsed and died from refractory disease. Only 1 patient that was refractory to intensive cytotoxic treatment remained alive with disease under supportive care. Fifteen patients (48%) were managed with palliative/supportive care: 7 received palliative treatment and supportive care, 8 received supportive care only, and 4 patients remained alive with disease under supportive care (median: 9 months). Mortality rate was 74% and overall survival at two years was 12%. To the best of our knowledge, there is no previous report regarding elderly patients with AML in Brazilian subsets. The present data are similar to previously reported studies showing that elderly AML patients are not only older but also biologically distinct from younger AML patients, particularly in terms of the high incidence of poor prognostic karyotypes and resistance to therapy.

Low expression of APAF-1XL in acute myeloid leukemia may be associated with the failure of remission induction therapy

Apoptotic protease activating factor 1 (APAF-1) has a critical role in the regulation of apoptosis. In the present study, the mRNA expression analysis of different APAF-1 transcripts (APAF-1S, APAF-1LC, APAF-1LN, and APAF-1XL) was analyzed in bone marrow samples from 37 patients with acute myeloid leukemia (newly diagnosed, with no previous treatment). APAF-1XL and APAF-1LN transcripts (with and without an extra WD-40 repeat region, respectively) were detected in all samples, although the major form expressed was APAF-1XL in 65% of the samples (group 1), while 35% of the samples expressed primarily APAF-1LN (group 2). Only 46% of the patients presented complete remission in response to remission induction therapy (represented by less than 5% marrow blasts and hematological recovery), all but 2 cases being from group 1, 21.6% did not attain complete remission (only 1 case from group 1), and 32.4% of the patients died early. Lower expression of APAF-1XL (APAF-1XL/APAF-1LN ratio <1.2) was associated with a poor response to therapy (P = 0.0005, Fisher exact test). Both groups showed similar characteristics regarding white blood cell counts, cytogenetic data or presence of gene rearrangements associated with good prognosis as AML1-ETO, CBFB-MYH11 and PML/RARA. Since it has been shown that only the isoforms with the extra WD-40 repeat region activate procaspase-9, we suggest that low procaspase-9 activation may also be involved in the deregulation of apoptosis and chemotherapy resistance in acute myeloid leukemia.

Association of loss of heterozygosity with cytogenetic abnormalities in acute myeloid leukemia and myelodysplastic syndrome

Deletions on chromosomes 5 and 7 are frequently seen in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). It is assumed that these deletions indicate loss of tumor suppressor genes on these chromosomes and until these tumor suppressor genes are identified, the functional consequences of these deletions and the molecular basis of these myeloid disorders cannot be completely understood. We evaluated loss of heterozygosity (LOH) in 44 patients (18 MDS and 26 AML, diagnosed according to WHO classification criteria) at diagnosis, using a four-microsatellite marker panel: an intragenic marker on the 7th intron of gene IRF-1 of the 5q31.1 region and three markers located inside the 7q31.1 region and correlated the LOH with karyotype abnormalities. The microsatellites chosen corresponded to chromosome regions frequently deleted in MDS/AML. The samples with Q (peak area) less than or equal to 0.50 were indicative of LOH. The percent of informative samples (i.e., heterozygous) for the intragenic microsatellite in gene IRF-1 and in loci D7S486, D7S515 and D7S522 were 66.6, 73.7, 75.5, and 48.8%, respectively. Cytogenetic abnormalities by G-banding were found in 36% (16/44) of the patients (2 of 18 MDS and 14 of 26 AML patients). We found a significantly positive association of the occurrence of LOH with abnormal karyotype (P < 0.05; chi-square test) and there were cases with LOH but the karyotype was normal (by G-banding). These data indicate that LOH in different microsatellite markers is possibly an event previous to chromosomal abnormalities in these myeloid neoplasias.

Characterization and analysis of the outcome of adults with acute myeloid leukemia treated in a Brazilian University hospital over three decades

We evaluated the outcome of 227 patients with acute myeloid leukemia during three decades (period 1 - 1980’s, N = 89; period 2 - 1990’s, N = 73; period 3 - 2000’s, N = 65) at a single institution. Major differences between the three groups included a higher median age, rates of multilineage dysplasia and co-morbidities, and a lower rate of clinical manifestations of advanced leukemia in recent years. The proportion of patients who received induction remission chemotherapy was 66, 75, and 85% for periods 1, 2, and 3, respectively (P = 0.04). The median survival was 40, 77, and 112 days, and the 5-year overall survival was 7, 13, and 22%, respectively (P = 0.01). The median disease-free survival was 266, 278, and 386 days (P = 0.049). Survival expectation for patients with acute myeloid leukemia has substantially improved during this 30-year period, due to a combination of lower tumor burden and a more efficient use of chemotherapy and supportive care.

Possible benefit of consolidation therapy with high-dose cytarabine on overall survival of adults with non-promyelocytic acute myeloid leukemia

In adults with non-promyelocytic acute myeloid leukemia (AML), high-dose cytarabine consolidation therapy has been shown to influence survival in selected patients, although the appropriate doses and schemes have not been defined. We evaluated survival after calculating the actual dose of cytarabine that patients received for consolidation therapy and divided them into 3 groups according to dose. We conducted a single-center, retrospective study involving 311 non-promyelocytic AML patients with a median age of 36 years (16-79 years) who received curative treatment between 1978 and 2007. The 131 patients who received cytarabine consolidation were assigned to study groups by their cytarabine dose protocol. Group 1 (n=69) received <1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles. The remaining patients received high-dose cytarabine (≥1.5 g/m2 every 12 h on 3 alternate days for up to 4 cycles). The actual dose received during the entire consolidation period in these patients was calculated, allowing us to divide these patients into 2 additional groups. Group 2 (n=27) received an intermediate-high-dose (<27 g/m2), and group 3 (n=35) received a very-high-dose (≥27 g/m2). Among the 311 patients receiving curative treatment, the 5-year survival rate was 20.2% (63 patients). The cytarabine consolidation dose was an independent determinant of survival in multivariate analysis; age, karyotype, induction protocol, French-American-British classification, and de novo leukemia were not. Comparisons showed that the risk of death was higher in the intermediate-high-dose group 2 (hazard ratio [HR]=4.51; 95% confidence interval [CI]: 1.81-11.21) and the low-dose group 1 (HR=4.43; 95% CI: 1.97-9.96) than in the very-high-dose group 3, with no significant difference between those two groups. Our findings indicated that very-high-dose cytarabine during consolidation in adults with non-promyelocytic AML may improve survival.

Acute leukemia in early childhood

Acute leukemia in early childhood is biologically and clinically distinct. The particular characteristics of this malignancy diagnosed during the first months of life have provided remarkable insights into the etiology of the disease. The pro-B, CD10 negative immunophenotype is typically found in infant acute leukemia, and the most common genetic alterations are the rearrangements of the MLL gene. In addition, the TEL/AML1 fusion gene is most frequently found in children older than 24 months. A molecular study on a Brazilian cohort (age range 0-23 months) has detected TEL/AML1+ve (N = 9), E2A/PBX1+ve (N = 4), PML/RARA+ve (N = 4), and AML1/ETO+ve (N = 2) cases. Undoubtedly, the great majority of genetic events occurring in these patients arise prenatally. The environmental exposure to damaging agents that give rise to genetic changes prenatally may be accurately determined in infants since the window of exposure is limited and known. Several studies have shown maternal exposures that may give rise to leukemogenic changes. The Brazilian Collaborative Study Group of Infant Acute Leukemia has found that mothers exposed to dipyrone, pesticides and hormones had an increased chance to give birth to babies with infant acute leukemia [OR = 1.48 (95%CI = 1.05-2.07), OR = 2.27 (95%CI = 1.56-3.31) and OR = 9.08 (95%CI = 2.95-27.96)], respectively. This review aims to summarize recent clues that have facilitated the elucidation of the biology of early childhood leukemias, with emphasis on infant acute leukemia in the Brazilian population.

Growth and puberty after treatment for acute lymphoblastic leukemia

Over the last 20 years, after combining treatment of chemotherapy and radiotherapy, there has been an improvement in the survival rate of acute lymphoblastic leukemia patients, with a current cure rate of around 70%. Children with the disease have been enrolled into international treatment protocols designed to improve survival and minimize the serious irreversible late effects. Our oncology unit uses the international protocol: GBTLI LLA-85 and 90, with the drugs methotrexate, cytosine, arabinoside, dexamethasone, and radiotherapy. However, these treatments can cause gonadal damage and growth impairment. PATIENTS AND METHOD: The authors analyzed 20 children off therapy in order to determine the role of the various doses of radiotherapy regarding endocrinological alterations. They were divided into 3 groups according to central nervous system prophylaxis: Group A underwent chemotherapy, group B underwent chemotherapy plus radiotherapy (18 Gy), and group C underwent chemotherapy plus radiotherapy (24 Gy). Serum concentrations of LH, FSH, GH, and testosterone were determined. Imaging studies included bone age, pelvic ultrasound and scrotum, and skull magnetic resonance imaging. RESULTS: Nine of the patients who received radiotherapy had decreased pituitary volume. There was a significant difference in the response to GH and loss of predicted final stature (Bayley-Pinneau) between the 2 irradiated groups and the group that was not irradiated, but there was no difference regarding the radiation doses used (18 or 24 Gy). The final predicted height (Bayley-Pinneau) was significantly less (P = 0.0071) in both groups treated with radiotherapy. Two girls had precocious puberty, and 1 boy with delayed puberty presented calcification of the epididymis. CONCLUSION: Radiotherapy was been responsible for late side effects, especially related to growth and puberty.

Effect of all-trans retinoic acid on newly diagnosed acute promyelocytic leukemia patients: results of a Brazilian center

Thirty-seven patients with acute promyelocytic leukemia (APL) were treated with all-trans retinoic acid (ATRA). Patients received 45 mg m-2 day-1 po of ATRA until complete remission (CR) was achieved, defined as: a) presence of less than 5% blasts in the bone marrow, with b) white blood cells >103/mm3, c) platelets >105/mm3 and d) hemoglobin concentration >8 g/dl, with no blood or platelet transfusions. Thirty-one (83.7%) patients achieved CR by day 50, and 75% of these before day 30. Correction of the coagulopathy, achieved between days 2 and 10 (mean, 3 days), was the first evidence of response to treatment. Only one patient had been previously treated with chemotherapy and three had the microgranular variant M3 form. Dryness of skin and mucosae was the most common side effect observed in 82% of the patients. Thrombosis, hepatotoxicity and retinoid acid syndrome (RAS) were observed in 7 (19%), 6 (16%) and 4 (11%) patients, respectively. Thirteen (35%) patients had to be submitted to chemotherapy due to hyperleukocytosis (above 40 x 103/mm3) and six of these presented with new signs of coagulopathy after chemotherapy. Four (11%) patients died secondarily to intracerebral hemorrhage (IH) and two (5.4%) dropped out of the protocol due to severe ATRA side effects (one RAS and one hepatotoxicity). RAS and IH were related strictly to hyperleukocytosis. The reduced use of platelets and fresh frozen plasma probably lowered the total cost of treatment. We conclude that ATRA is an effective agent for inducing complete remission in APL patients.

Acute promyelocytic leukemia: the study of t(15;17) translocation by fluorescent in situ hybridization, reverse transcriptase-polymerase chain reaction and cytogenetic techniques

Acute promyelocytic leukemia (AML M3) is a well-defined subtype of leukemia with specific and peculiar characteristics. Immediate identification of t(15;17) or the PML/RARA gene rearrangement is fundamental for treatment. The objective of the present study was to compare fluorescent in situ hybridization (FISH), reverse transcriptase-polymerase chain reaction (RT-PCR) and karyotyping in 18 samples (12 at diagnosis and 6 after treatment) from 13 AML M3 patients. Bone marrow samples were submitted to karyotype G-banding, FISH and RT-PCR. At diagnosis, cytogenetics was successful in 10 of 12 samples, 8 with t(15;17) and 2 without. FISH was positive in 11/12 cases (one had no cells for analysis) and positivity varied from 25 to 93% (mean: 56%). RT-PCR was done in 6/12 cases and all were positive. Four of 8 patients with t(15;17) presented positive RT-PCR as well as 2 without metaphases. The lack of RT-PCR results in the other samples was due to poor quality RNA. When the three tests were compared at diagnosis, karyotyping presented the translocation in 80% of the tested samples while FISH and RT-PCR showed the PML/RARA rearrangement in 100% of them. Of 6 samples evaluated after treatment, 3 showed a normal karyotype, 1 persistence of an abnormal clone and 2 no metaphases. FISH was negative in 4 samples studied and 2 had no material for analysis. RT-PCR was positive in 4 (2 of which showed negative FISH, indicating residual disease) and negative in 2. When the three tests were compared after treatment, they showed concordance in 2 of 6 samples or, when there were not enough cells for all tests, concordance between karyotype and RT-PCR in one. At remission, RT-PCR was the most sensitive test in detecting residual disease, as expected (positive in 4/6 samples). An incidence of about 40% of 5' breaks and 60% of 3' breaks, i.e., bcr3 and bcr1/bcr2, respectively, was observed.

In vitro cytotoxicity of the LDE: daunorubicin complex in acute myelogenous leukemia blast cells

Acute myelogenous leukemia (AML) blast cells show high-affinity degradation of low-density lipoprotein (LDL), suggesting an increased expression of cellular LDL receptors. LDE is a lipid microemulsion easily synthesized in vitro which is known to mimic the metabolic pathway of LDL. We used LDE as a carrier for daunorubicin and assayed the cytotoxicity of the complex using AML blast cells since RT-PCR analysis showed that AML cells express LDL receptor mRNA. The LDE:daunorubicin complex killed 46.7% of blast cells and 20.2% of normal bone marrow cells (P<0.001; Student t-test). Moreover, this complex destroyed AML blast cells as efficiently as free daunorubicin. Thus, LDE might be a suitable carrier of chemotherapeutic agents targeting these drugs to neoplastic cells and protecting normal tissues.

Microsatellite instability and cytogenetic survey in myeloid leukemias

Microsatellites are short tandem repeat sequences dispersed throughout the genome. Their instability at multiple genetic loci may result from mismatch repair errors and it occurs in hereditary nonpolyposis colorectal cancer. This instability is also found in many sporadic cancers. In order to evaluate the importance of this process in myeloid leukemias, we studied five loci in different chromosomes of 43 patients, 22 with chronic myelocytic leukemia (CML) in the chronic phase, 7 with CML in blast crisis, and 14 with acute myeloid leukemia (AML), by comparing leukemic DNA extracted from bone marrow and constitutional DNA obtained from buccal epithelial cells. Only one of the 43 patients (2.1%), with relapsed AML, showed an alteration in the allele length at a single locus. Cytogenetic analysis was performed in order to improve the characterization of leukemic subtypes and to determine if specific chromosome aberrations were associated with the presence of microsatellite instability. Several chromosome aberrations were observed, most of them detected at diagnosis and during follow-up of the patients, according to current literature. These findings suggest that microsatellite instability is an infrequent genetic event in myeloid leukemias, adding support to the current view that the mechanisms of genomic instability in solid tumors differ from those observed in leukemias, where specific chromosome aberrations seem to play a major role.

Acute leukemias in Piauí: comparison with features observed in other regions of Brazil

Differences in age and sex distribution as well as FAB (French-American-British classification) types have been reported for acute leukemias in several countries. We studied the demographics and response to treatment of patients with acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) between 1989 and 2000 in Teresina, Piauí, and compared these results with reports from Brazil and other countries. Complete data concerning 345 patients (230 ALL, 115 AML) were reviewed. AML occurred predominantly in adults (77%), with a median age of 34 years, similar to that found in the southeast of Brazil but lower than the median age in the United States and Europe (52 years). FAB distribution was similar in children and adults and FAB-M2 was the most common type, as also found in Japan. The high frequency of FAB-M3 described in most Brazilian studies and for Hispanics in the United States was not observed. Overall survival for adults was 40%, similar to other studies in Brazil. A high mortality rate was observed during induction. No clinical or hematological parameter influenced survival in the Cox model. ALL presented the characteristic peak of incidence between 2-8 years. Most of the cases were CD10+ pre-B ALL. In 25%, abnormal expression of myeloid antigens was observed. Only 10% of the patients were older than 30 years. Overall survival was better for children. Age and leukocyte count were independent prognostic factors. These data demonstrate that, although there are regional peculiarities, the application of standardized treatments and good supportive care make it possible to achieve results observed in other countries for the same chemotherapy protocols.

Cytosine arabinoside-metabolizing enzyme genes are underexpressed in children with MLL gene-rearranged acute lymphoblastic leukemia

Infant acute lymphoblastic leukemia (IALL) is characterized by mixed lineage leukemia (MLL) gene rearrangements, unique gene expression profiles, poor prognosis, and drug resistance. One exception is cytosine arabinoside (Ara-C) to which IALL cells seem to be more sensitive. We quantified mRNA expression of Ara-C key enzymes in leukemic lymphoblasts from 64 Brazilian ALL children, 15 of them presenting MLL gene rearrangement, and correlated it with clinical and biological features. The diagnosis was based on morphological criteria and immunophenotyping using monoclonal antibodies. MLL gene rearrangements were detected by conventional cytogenetic analysis, RT-PCR and/or fluorescence in situ hybridization. The DCK and HENT1 expression levels were determined by real-time quantitative PCR using SYBR Green I. Relative quantification was made by the standard curve method. The results were analyzed by Mann-Whitney and Fisher exact tests. A P value of £0.05 was considered to be statistically significant. DCK and HENT1 expression levels were significantly lower in children with MLL gene-rearranged ALL compared to children with MLL germ line ALL (P = 0.0003 and 0.03, respectively). Our results differ from previous ones concerning HENT1 mRNA expression that observed a higher expression level in MLL gene-rearranged leukemias. In conclusion, the expression of the genes related to Ara-C metabolism was lower in MLL-positive children in the sample studied, suggesting the presence of population differences in the expression profile of these genes especially for HENT1.

Mesenchymal stem cells from patients with chronic myeloid leukemia do not express BCR-ABL and have absence of chimerism after allogeneic bone marrow transplant

Bone marrow is a heterogeneous cell population which includes hematopoietic and mesenchymal progenitor cells. Dysregulated hematopoiesis occurs in chronic myelogenous leukemia (CML), being caused at least in part by abnormalities in the hematopoietic progenitors. However, the role of mesenchymal stem cells (MSCs) in CML has not been well characterized. The objectives of the present study were to observe the biological characteristics of MSCs from CML patients and to determine if MSCs originate in part from donors in CML patients after bone marrow transplantation (BMT). We analyzed MSCs from 5 untreated patients and from 3 CML patients after sex-mismatched allogeneic BMT. Flow cytometry analysis revealed the typical MSC phenotype and in vitro assays showed ability to differentiate into adipocytes and osteoblasts. Moreover, although some RT-PCR data were contradictory, combined fluorescence in situ hybridization analysis showed that MSCs from CML patients do not express the bcr-abl gene. Regarding MSCs of donor origin, although it is possible to detect Y target sequence by nested PCR, the low frequency (0.14 and 0.34%) of XY cells in 2 MSC CML patients by fluorescence in situ hybridization analysis suggests the presence of contaminant hematopoietic cells and the absence of host-derived MSCs in CML patients. Therefore, we conclude that MSCs from CML patients express the typical MSC phenotype, can differentiate into osteogenic and adipogenic lineages and do not express the bcr-abl gene. MSCs cannot be found in recipients 12 to 20 months after BMT. The influence of MSCs on the dysregulation of hematopoiesis in CML patients deserves further investigation.