IGFBP7 participates in the reciprocal interaction between acute lymphoblastic leukemia and BM stromal cells and in leukemia resistance to asparaginase.

The interaction of acute lymphoblastic leukemia (ALL) blasts with bone marrow (BM) stromal cells (BMSCs) has a positive impact on ALL resistance to chemotherapy. We investigated the modulation of a series of putative asparaginase-resistance/sensitivity genes in B-precursor ALL cells upon coculture with BMSCs. Coculture with stromal cells resulted in increased insulin-like growth factor (IGF)-binding protein 7 (IGFBP7) expression by ALL cells. Assays with IGFBP7 knockdown ALL and stromal cell lines, or with addition of recombinant rIGFBP7 (rIGFBP7) to the culture medium, showed that IGFBP7 acts as a positive regulator of ALL and stromal cells growth, and significantly enhances in-vitro resistance of ALL to asparaginase. In these assays, IGFBP7 function occurred mainly in an insulin- and stromal-dependent manner. ALL cells were found to contribute substantially to extracellular IGFBP7 levels in the conditioned coculture medium. Diagnostic BM plasma from children with ALL had higher levels of IGFBP7 than controls. IGFBP7, in an insulin/IGF-dependent manner, enhanced asparagine synthetase expression and asparagine secretion by BMSCs, thus providing a stromal-dependent mechanism by which IGFBP7 protects ALL cells against asparaginase in this coculture system. Importantly, higher IGFBP7 mRNA levels were associated with lower leukemia-free survival (Cox regression model, P=0.003) in precursor B-cell Ph(-) ALL patients (n=147) treated with a contemporary polychemotherapy protocol.

Colony stimulating factors for prevention of myelosupressive therapy induced febrile neutropenia in children with acute lymphoblastic leukaemia.

BACKGROUND: Acute lymphoblastic leukaemia (ALL) is the most common cancer in childhood and febrile neutropenia is a potentially life-threatening side effect of its treatment. Current treatment consists of supportive care plus antibiotics. Clinical trials have attempted to evaluate the use of colony-stimulating factors (CSF) as additional therapy to prevent febrile neutropenia in children with ALL. The individual trials do not show whether there is significant benefit or not. Systematic review provides the most reliable assessment and the best recommendations for practice. OBJECTIVES: To evaluate the safety and effectiveness of the addition of G-CSF or GM-CSF to myelosuppressive chemotherapy in children with ALL, in an effort to prevent the development of febrile neutropenia. Evaluation of number of febrile neutropenia episodes, length to neutrophil count recovery, incidence and length of hospitalisation, number of infectious disease episodes, incidence and length of treatment delays, side effects (flu-like syndrome, bone pain and allergic reaction), relapse and overall mortality (death). SEARCH STRATEGY: The search covered the Cochrane Central Register of Controlled Trials, MEDLINE, EMBASE, CANCERLIT, LILACS, and SciElo. We manually searched records of conference proceedings of ASCO and ASH from 1985 to 2003 as well as databases of ongoing trials. We consulted experts and scanned references from the relevant articles. SELECTION CRITERIA: We looked for randomised controlled trials (RCTs) comparing CSF with placebo or no treatment as primary or secondary prophylaxis to prevent febrile neutropenia in children with ALL. DATA COLLECTION AND ANALYSIS: Two authors independently selected, critically appraised studies and extracted relevant data. The end points of interest were:* Primary end points: number of febrile neutropenia episodes and overall mortality (death) * Secondary end points: time to neutrophil count recovery, incidence and length of hospitalisation, number of infectious diseases episodes, incidence and length of treatment delays, side effects (flu-like syndrome, bone pain and allergic reaction) and relapse. We conducted a meta-analysis of these end points and expressed the results as Peto odds ratios. For continuous outcomes we calculated a weighted mean difference and a standardised mean difference. For count data, meta-analysis of the logarithms of the rate ratios using generic inverse variance was employed. MAIN RESULTS: We scanned more than 5500 citations and included six studies with a total of 332 participants in the analysis. There were insufficient data to assess the effect on survival. The use of CSF significantly reduced the number of episodes of febrile neutropenia episodes (Rate Ratio = 0.63; 95% confidence interval (CI) 0.46 to 0.85; p =0.003, with substantial heterogeneity), the length of hospitalisation (weighted mean difference (WMD) = -1.58; 95% CI -3.00 to -0.15; p = 0.03), and number of infectious diseases episodes (Rate Ratio=0.44; 95%CI 0.24 to 0.80; p=0.002). In spite of these results, CSF did not influence the length of episodes of neutropenia (WMD = -1.11; 95% CI -3.55 to 1.32; p = 0.4) or delays in chemotherapy courses (Rate Ratio=0.77; 95%CI 0.49 to 1,23; p=0.28) . AUTHORS' CONCLUSIONS: Children with ALL treated with CSF benefit from shorter hospitalisation and fewer infections. However, there was no evidence for a shortened duration of neutropenia nor fewer treatment delays, and no useful information about survival. The role of CSF regarding febrile neutropenia episodes is still uncertain. Although current data shows statistical benefit for CSF use, substantial heterogeneity between included trials does not allow this conclusion.

Detection of oncogenic human papillomavirus in sporadic retinoblastoma.

PURPOSE: To investigate the presence of human papillomavirus (HPV) DNA in tumour tissue from patients with unilateral retinoblastoma. METHODS: Samples of paraffin-embedded tumour tissue from 43 children with unilateral retinoblastoma were collected to investigate the presence of HPV DNA using polymerase chain reaction (PCR) and dot blot hybridization. RESULTS: Oncogenic HPV DNA types 16 and 35 were detected in 12 (27.9%) of 43 tumour specimens. A higher frequency of differentiated tumours (63.3%) was observed among the HPV-positive tumours. CONCLUSIONS: Future studies are necessary to demonstrate an association between HPV and sporadic retinoblastoma.

Transplacental chemical exposure and risk of infant leukemia with MLL gene fusion.

Infant acute leukemia (IAL) frequently involves breakage and recombination of the MLL gene with one of several potential partner genes. These gene fusions arise in utero and are similar to those found in leukemias secondary to chemotherapy with inhibitors of topoisomerase II (topo-II). This has led to the hypothesis that in utero exposures to chemicals may cause IAL via an effect on topo-II. We report a pilot case-control study of IAL across different countries and ethnic groups. Cases (n = 136) were population-based in most centers. Controls (n = 266) were selected from inpatients and outpatients at hospitals serving the same populations. MLL rearrangement status was derived by Southern blot analysis, and maternal exposure data were obtained by interviews using a structured questionnaire. Apart from the use of cigarettes and alcohol, very few mothers reported exposure to known topo-II inhibitors. Significant case-control differences were apparent for ingestion of several groups of drugs, including herbal medicines and drugs classified as "DNA-damaging," and for exposure to pesticides with the last two being largely attributable, respectively, to one nonsteroidal anti-inflammatory drug, dipyrone, and mosquitocidals (including Baygon). Elevated odds ratios were observed for MLL+ve (but not MLL-ve) leukemias (2.31 for DNA-damaging drugs, P = 0.03; 5.84 for dipyrone, P = 0.001; and 9.68 for mosquitocidals, P = 0.003). Although it is unclear at present whether these particular exposures operate via an effect on topo-II, the data suggest that specific chemical exposures of the fetus during pregnancy may cause MLL gene fusions. Given the widespread use of dipyrone, Baygon, and other carbamate-based insecticides in certain settings, confirmation of these apparent associations is urgently required.

Diamond-Blackfan anemia: report of seven further mutations in the RPS19 gene and evidence of mutation heterogeneity in the Italian population.

Diamond-Blackfan anemia (DBA) is a congenital disease characterized by defective erythroid progenitor maturation and physical malformations. Most cases are sporadic, but dominant or, more rarely, recessive inheritance is observed in 10% of patients. Mutations in the gene encoding ribosomal protein (RP) S19 have recently been found in 25% of patients with either the dominant or the sporadic form. DBA is the first human disease due to mutations in a ribosomal structural protein. Families unlinked to this locus have also been reported. In an investigation of 23 individuals, we identified eight different mutations in 9 patients. These include five missense, one frameshift, one splice site defect, and one 4-bp insertion in the regulatory sequence. Seven mutations are new; one has so far been found in 8 patients and is a relatively common de novo event. Two mutations are predicted to generate a truncated protein. We also report the prevalence of RPS 19 mutations in the Italian DBA population, as shown by an analysis of 56 patients. No genotype-phenotype correlation was found between patients with the same mutation. The main clinical applications for molecular analysis are clinical diagnosis of patients with an incomplete form of DBA and testing of siblings of a patient with a severe form so as to avoid using those who carry a mutation and a silent phenotype as allogeneic stem cell donors.

Selective surgical indication in the management of neutropenic children presenting with acute abdomen.

As the treatment of pediatric malignancies improves and survival increases, the diagnosis of acute abdomen in these patients also becomes more common. Nevertheless, the management of this condition is still controversial. The authors report their experience in treating 12 neutropenic children with acute abdomen. The charts of 12 neutropenic patients with a diagnosis of acute abdomen treated at Boldrini Children's Cancer Center in Campinas, Brazil, between 1991 and 1996, were reviewed. Therapeutic strategy included an initial period of bowel rest, general supportive measures, and broad-spectrum antibiotics while waiting for the neutrophil count to rise. Three patients recovered completely without surgery, 8 underwent late surgery without complications, and 1 died due to uncontrolled sepsis before surgery. The treatment of acute abdomen in neutropenic children remains controversial. As shown in the present series, an initial nonoperative approach with selective surgical indication appears to be safe and to yield good results. Supportive treatment, until the neutrophil count rises, followed by surgery, if necessary, appears to be a sound therapeutic approach for neutropenic children with acute abdomen.

[Development of puberty in girls treated for acute lymphocytic leukemia]. / Desenvolvimento puberal em meninas tratadas de LLA.

BACKGROUND: In order to evaluate the puberal development of girls treated by Acute Lymphocytic Leukaemia (ALL) a retrospective study was done at Campinas-SP, Brazil. MATERIAL AND METHODS: Forty two girls were treated by ALL with either 18 or 24 Grays of cranial irradiation. All patients were treated with chemotherapy including intrathecal methotrexate in similar dose regimens in either groups. RESULTS: The results showed lower mean ages at telarche, pubarche and menarche in the treated group, mainly in the group treated before five years old. No differences were observed in the 18 Grays or 24 Grays group. CONCLUSIONS: Our data suggest that girls treated by ALL have a precocious puberal development.

Desenvolvimento puberal em meninas tratadas de LLA / Development of puberty in girls treated by acute lymphocytic leukaemia

Objetivo. Com o objetivo de avaliar o desenvolvimento puberal após o tratamento de leucemia linfóide aguda (LLA) na infância, foi realizado um estudo retrospectivo, em meninas tratadas de janeiro de 1980 a janeiro de 1991, no Centro de Investigaçoes Hematológicas "Dr. Domingos A. Boldrini", em Campinas-SP. Casuística e Método. Foram selecionadas 42 meninas, tratadas antes da puberdade com quimioterapia sistêmica e intratecal e radioterapia cranial, utilizando doses de 18 a 24 Grays (Gy). Resultados. As idades médias da telarca, pubarca e menarca foram inferiores às do grupo-controle, embora com significância estatística apenas para a idade da telarca. Nao houve diferenças entre os grupos tratados com 18 ou 24Gy. As meninas tratadas antes dos cinco anos de idade apresentaram idade média da menarca estatisticamente inferior àquelas tratadas após cinco anos e em relaçao ao grupo-controle. Conclusao. Os resultados mostraram que o desenvolvimento puberbal em meninas tratadas de LLA na infância foi mais precoce que o de mininas saudáveis.

[Treatment of acute lymphocytic leukemia and growth]. / Tratamento de leucemia linfóide aguda e crescimento.

BACKGROUND: The treatment of the acute lymphocytic leukaemia can determine impaired growth. SUBJECTS AND METHOD: All the patients had length measurements at the time of the beginning of the treatment and, at least, one year after the end of it. CONCLUSIONS: There was impaired growth after the treatment according to the dose regimens (18 or 24 Grays). No relation was observed related to the age at the diagnosis.

Tratamento de leucemia linfóide aguda e crescimento / Treatment of acute lymphocytic leukemia and growth

Objetivo. Determinar alteraçoes no crescimento após o tratamento de leucemia linfóide aguda em meninas. Pacientes e Métodos. Realizou-se estudo restrospectivo com 59 meninas que apresentavam medidas de estatura antes e com no mínimo um ano do tratamento, subdivididas de acordo com a dose de radioterapia cranial utilizada [18 ou 24 Grays Gy] e com a idade no início do tratamento (antes e após os cinco anos de idade). Resultados. Observou-se deficiência do crescimento com um, dois e mais de dois anos do tratamento. O crescimento foi mais afetado no grupo tratado com 24Gy, quando comparado com 18Gy. Nao houve diferenças com relaçao à idade no início do tratamento. Conclusoes. Houve retardo no crescimento após o tratamento, independentemente da idade em que foi realizado, mas dependente da dose de radioterapia cranial utilizada.

Diamond-Blackfan anemia: expansion of erythroid progenitors in vitro by IL-9, but exclusion of a significant pathogenetic role for the IL-9 gene and the hematopoietic gene cluster on chromosome 5q.

Diamond-Blackfan anemia (DBA) is a congenital pure red blood cell aplasia that often requires lifelong transfusional therapy. Autosomal dominant and recessive inheritance have both been reported, suggesting genetic heterogeneity, but most cases occur sporadically. The origin of impaired erythropoiesis is unknown. Several erythroid growth factors have been thought to have a role in the pathogenesis of DBA. However, there is neither molecular nor clinical evidence for the involvement of erythropoietin (EPO), its receptor, stem cell factor (SCF), or interleukin (IL)-3, even if the addition of SCF to IL-3 and EPO does significantly increase the growth of erythroid progenitors in in vitro cultures in most patients. In this work we evaluated the possible role of another early-acting erythroid growth factor, IL-9. We found that the addition of IL-9 to SCF, IL-3, and EPO further increases burst-forming unit-erythroid growth in in vitro cultures of those DBA patients who responded to SCF. To investigate the role of the IL-9 gene, we evaluated its segregation in 22 families with members who have DBA by using a polymorphic microsatellite located within its intron 4. Lod score analysis ruled out any statistically significant involvement of the IL-9 gene in the pathogenesis of DBA. Moreover, linkage analysis with 11 highly polymorphic markers spanning 5q31.1-q33.2 excluded this region, which is included in the major cluster of genes active in hematopoiesis of the human genome.

Barriers to bone marrow transplantation for sickle cell anemia.

While allogeneic marrow transplantation is curative therapy for patients with sickle cell anemia, only a small fraction of patients in the United States receive this treatment. We surveyed participants in our multicenter study of marrow transplantation for sickle cell anemia to determine reasons for not proceeding to transplantation. Among the 4848 patients less than 16 years of age with sickle cell anemia that were followed in 22 collaborating centers, 315 (6.5%) patients were reported to meet protocol entry criteria for transplantation, although there was wide variation among the institutions (0.9-36%). Among the 315 patients eligible for transplantation, 128 (41%) had human leukocyte antigen (HLA) typing performed, and of these 44 (14% of those meeting entry criteria) had an HLA-identical sibling. Common reasons for not proceeding with HLA typing in the remaining 187 patients included lack of a candidate sibling donor (76 patients, 24% of those meeting criteria) and lack of financial or psychosocial support (33, 10.5%). Parental refusal (30, 9.5%), physician refusal (13, 4%), history of medical noncompliance (2, < 1%), and other reasons (33, 10.5%) were less frequently cited. To date, 25 patients have been transplanted. Of the remaining 19 patients with HLA-matched donors, seven did not proceed to transplantation because of parental refusal, while the others anticipate a future transplantation (6), have experienced symptomatic improvement (4), or have relocated abroad (2). We conclude that the major barrier to marrow transplantation for sickle cell anemia is lack of an HLA-identical donor. But since only 6.5% of all children with sickle cell disease were considered eligible for transplantation, it is possible that other significant obstacles remain to be identified. For patients reported to meet eligibility criteria, parental refusal and limited financial or psychosocial support were infrequent barriers to transplantation.

Childhood Hodgkin's disease in Campinas, Brazil.

PURPOSE: Little clinical information about Hodgkin's disease in children is available from poor countries. The object of this study is to evaluate our data in Campinas, Brazil and hope "to make one dot on the geographic map of this disease more clear." PATIENTS AND METHODS: Between 1978 and 1988, 46 patients under the age of 17 years with biopsy-proven Hodgkin's Disease (HD) were referred for evaluation at Centro Boldrini in Campinas, São Paulo state, in Brazil. Thirty-seven of them were treated and followed-up only at this Center and are the subjects of this analysis. All the original histological slides were obtained, reviewed, and classified according to the Rye system. Staging procedures included exploratory laparotomy in 33 of 37 children, but none had lymphangiography. Treatment was individualized until January 1986 when the German protocol was adopted. RESULTS: Nineteen cases were classified as nodular sclerosis, 14 as mixed cellularity, and three as lymphocyte depleted. Mean age was 7 years; male/female ratio was 2:1. Fifty percent were advanced stages III and IV and 46% (17/37) had at least one of the systemic B symptoms. Mean follow-up was 81 months (range from 41 to 174 months). Five-year actuarial overall survival was 78%. Two children (5%) had acute myeloid leukemia at 25 and 49 months after diagnosis. CONCLUSIONS: Although distribution of histological subtypes of our cases is similar to other reports in developed countries, as well as percentage of advanced stages III/IV, our patients fared worse when compared to those reports. The reason for this continues to remain unclear but it does not seem to be related to histology subtypes.

Giant actin inclusions in hematopoietic cells associated with transfusion-dependent anemia and grey skin discoloration.

We evaluated a 13-month-old boy with cytoplasmic inclusions in hematopoietic cells, transfusion-dependent anemia, splenomegaly, and striking grey skin discoloration. Bright blue inclusions, 1 to 5 microns in diameter, were observed, primarily in the cytoplasm, of 30% to 40% of myeloid cells and in occasional monocytes, megakaryocytes, and lymphocytes on Wright Giemsa-stained bone marrow and blood smears. They occasionally involved the nucleus. The inclusions lacked lysosomes, polysaccharides, or lipids. Ultrastructurally, they lacked limiting membranes and consisted of tightly packed microfilaments averaging 7 nm in diameter, consistent with the size of actin monofilaments. On light microscopy, the inclusions stained with a monoclonal antibody to muscle-specific actin. Inclusion-positive cells contained increased F-actin content and were defective in chemotactic factor-activated actin polymerization; inclusion-negative cells polymerized actin normally. Neutrophil and platelet numbers and functional studies were mildly abnormal. Anemia and skin discoloration resolved spontaneously after 18 months, but the giant inclusions have persisted to the present. We conclude that this child has a previously unreported constellation of clinical and laboratory findings comprising severe anemia, intermittent neutropenia and thrombocytopenia, abnormal neutrophil migration and platelet aggregation, giant inclusions of actin in hematopoietic cells, and grey skin discoloration.

Treatment results of three consecutive Brazilian cooperative childhood ALL protocols: GBTLI-80, GBTLI-82 and -85. ALL Brazilian Group.

The Brazilian Cooperative Group for Treatment of Childhood Acute Lymphocytic Leukemia (GBTLI) has started clinical activities trials in 1980. Three consecutive multicenter studies in children with unprevious treated ALL have been completed including 994 patients. The first GBTLI-80 accrued 203 children from 1980 to 1982. It was delineated with the standard three drugs induction therapy, CNS protection for all pts comprised cranial irradiation and intrathecal Methotrexate. For low risk pts cranial irradiation with 18Gy was compared in a randomized trial with 24Gy. Maintenance therapy continued for 120 weeks. The 12 years of the event free survival rates for all risk groups is 50% (SD 5%). Regarding CNS relapses there was no significant statistical difference between pts that received 18 or 24Gy. The treatment strategy of GBTLI-82 (n = 360) from 1982 to 1985, consisted of the same previous induction, consolidation, CNS therapy with cranial irradiation 18 Gy (low risk) or 24Gy (high risk), followed by continuous maintenance for 2 years. The main question in this study was the comparison between sequential rotation or pulses of 3 pairs of drugs during maintenance. At a median follow-up of 10 years, the overall event free survival rates for all children is 58% (SD 4%). There was no significant difference between the two maintenance regimens. The successor GBTLI-85 ran from 1985 to 1988 and registered 431 pts. For the first time no cranial radiation was given to children with very good prognosis. For them, CNS protection was done with triple intrathecal therapy during all treatment. A consolidation therapy with high dose ARA-C was introduced for high risk pts and infants The 6.5 years event free survival for all children is 70% (SD 4%). Significant better results were achieved for high risk and infants pts (EFS 50%). Early intensification therapy and rotational combination chemotherapy improved the outcome in childhood ALL in Brazil.

Combination of rubidazone and cytosine arabinoside in the treatment of first relapse in acute myelocytic leukemia.

This study tested the efficacy of rubidazone and cytosine arabinoside in 35 patients (13 children and 22 adults) with acute myelocytic leukemia in first relapse. Induction consisted of 1-2 courses of rubidazone 200 mg/m2 days x 4 days plus cytosine arabinoside 100 mg/m2 x 7 days in CI followed by 2 consolidation courses of 3 days and 5 days. Nineteen patients (54%) achieved complete remission, 8 failed to respond, and 8 died. Twelve patients relapsed after 1 to 9 months, at a median of 4 months, 1 patient died of cardiac failure and 1 remains in complete remission at 12 months. Five patients underwent bone marrow transplantation, 3 of them autologous, 1 was still in complete remission at 29 months, 1 relapsed, and 1 died of sepsis. Two received allogeneic marrow transplants and died at 3 and 4 months afterwards of VOD and graft failure. The main toxicity was severe and prolonged myelosuppression.

Reduced phagocytic a activity mediated by C3b and Fc monocyte receptors from children with sckle-cell disease

The causes of high morbidity due to infection among children with sickle-cell disease (SD) are unknown. Immunlogical studies have focused on spleen function, on the alternative complement pathway, and recently on phagocytic activity. We evaluated Fc recptor-mediated phagocytic activity. We evaluated Fc receptor-mediated phagocytosis (sheep red cells opsonized with rabbit anti-E IgG, EA) and C3b receptor-mediated phagocytosis (zymosan particles incubated with fresh serum) in 27 children with SD. The control group consisted of 28 normal children matched by age and sex. The phagocytic indices obtained for cells from patients with SD were significantly lower than those for the controls (P < 0.001), both when EA and zymosan were used independent of whether the zymosan particles were incubated with patient serum or with a pool of normal sera. The results suggest the absence of abnormalities in the alternative complement pathway but do indicate an intrinsic cellular defect

Reduced phagocytic activity mediated by C3b and Fc monocyte receptors from children with sickle-cell disease.

The causes of high morbidity due to infection among children with sickle-cell disease (SD) are unknown. Immunological studies have focused on spleen function, on the alternative complement pathway, and recently on phagocytic activity. We evaluated Fc receptor-mediated phagocytosis (sheep red cells opsonized with rabbit anti-E IgG, EA) and C3b receptor-mediated phagocytosis (zymosan particles incubated with fresh serum) in 27 children with SD. The control group consisted of 28 normal children matched by age and sex. The phagocytic indices obtained for cells from patients with SD were significantly lower than those for the controls (P less than 0.001), both when EA and zymosan were used and independent of whether the zymosan particles were incubated with patient serum or with a pool of normal sera. The results suggest the absence of abnormalities in the alternative complement pathway but do indicate an intrinsic cellular defect.