Late effects in survivors of childhood acute lymphoblastic leukemia: a study from Thai Pediatric Oncology Group.

Survivors of childhood acute lymphoblastic leukemia (ALL) are at risk of long-term late effects. Therefore, systematic screenings of the late complications are essential. The objective of this study was to determine the prevalence of late effects of Thai children and adolescents after completion of ALL therapy. We performed a cross-sectional study for evaluation of the late effects in ALL survivors who came for follow-up at 10 pediatric oncology centers in Thailand. We evaluated the treatment-related late complications of children and adolescents who had finished ALL treatment for at least 2 years. Demographic data, treatment modalities, and late effects were recorded and analyzed. There were 258 survivors with a median age of 12.2 years (range 3.6-23.3 years). The median follow-up time was 7.2 years (range 2-17.5 years). Forty-seven percent (122 cases) suffered from at least one late effect. Overweight/obesity was the most common late effect. Radiation of central nervous system was a significant risk factor for overweight/obesity (OR 1.97, 95% CI 1.02-3.81) and educational problems (OR 4.3, 95% CI 1.32-14.02). Our data have demonstrated a significant prevalence of late effects after childhood ALL therapy. A long-term follow-up program for survivors of childhood cancer is therefore needed in our country.

Natural history of Southeast Asian Ovalocytosis during the first 3 years of life.

Southeast Asian Ovalocytosis (SAO), the most common red cell membrane disorder found in the Far-East and Pacific rim, appears to be innocuous in man since it has been identified mostly in non-anemic healthy individuals. To further substantiate our previous observation that this condition might be symptomatic particularly in the neonatal period, we studied 1567 newborns from Southern Thailand where SAO is prevalent. Thirty-one babies (1: 50 with allele frequency of 0.01) have been identified with SAO and confirmed molecularly to carry a single defective AE-1 (band 3) allele. These babies had significant anemia at birth due to hemolysis with 51.6% of them developing neonatal hyperbilirubinemia. Co-inheritance of common UGT1A1 variants in such cases was not associated with their degree of jaundice. Interestingly, hematology data of these SAO babies became "normalized" in the first 3 years of life without further evidence of on-going and/or even "compensated" hemolysis.

Clinical features and molecular analysis in Thai patients with HbH disease.

We studied the alpha-globin gene abnormalities, the clinical features, hematologic values, growth assessment, transfusion therapy, and serum ferritin levels of patients with hemoglobin H (HbH) disease in southern Thailand. HbH disease in 83 of the 147 patients was the deletional type of HbH. The remaining 64 patients was the nondeletional type of HbH disease. All 83 patients with the deletional type were double heterozygotes of alpha(0)-thalassemia and alpha(+)-thalassemia. The Southeast Asian type of alpha(0)-thalassemia accounted for 98% of the Thai patients with HbH disease and the Thai type of alpha(0)-thalassemia made up the rest. A 3.7-kb deletion accounted for 91% of alpha(+)-thalassemia, and a 4.2-kb deletion made up the rest of the deletional type. In patients with nondeletional type of HbH disease, the Constant Spring variant was the majority of the disease. Newborns with a nondeletional genotype had higher mean corpuscular volume, had higher mean corpuscular hemoglobin, had higher red blood cell distribution width, had lower mean corpuscular hemoglobin concentration, and had higher proportions of Hb Bart's than those with a deletional genotype. Twenty-one percent of children with HbH disease had growth deficiency. A genotype-phenotype correlation was found; patients with the nondeletional type of HbH disease had more symptoms at a younger age, more severe hemolytic anemia, more growth deficiency, more dysmorphic facial features, larger spleens, larger livers, and higher serum ferritin levels and required more transfusions than patients with deletional HbH disease.

Prevalence of impaired glucose metabolism in beta-thalassemic children receiving hypertransfusions with a suboptimal dosage of iron-chelating therapy.

A cross-sectional study of impaired glucose metabolism was carried out in 48 beta-thalassemic patients receiving hypertransfusions. An oral glucose tolerance test (OGTT) was performed using the method and criteria of the American Diabetes Association (ADA). Diabetes mellitus was diagnosed in two patients, and impaired glucose tolerance was found in four patients, giving a prevalence of impaired glucose metabolism of 12.5% in our patient population. The significant clinical characteristics associated with the diagnosis of impaired glucose metabolism were wasting (-2.15/-0.86 SDS, p = 0.025), stunting (-2.69/-1.22 SDS, p = 0.03), higher ferritin levels (8679/4710 microg/L, p = 0.005), splenectomy (50/9.5%, p = 0.012), and lower area under curve (AUC) of insulin secretion after OGTT (40.0/77.7, p = 0.002). The significant decrease of AUC insulin in thalassemic patients with an impaired glucose tolerance test suggests that the pathogenesis may originate from pancreatic beta-cell damage rather than from insulin resistance. In conclusion, the prevalence of impaired glucose tolerance in our population of thalassemic patients receiving hypertransfusions with suboptimal iron chelating therapy was 12.5%. The clinical characteristics of thalassemic patients who developed impaired glucose tolerance were wasting, stunting, higher ferritin levels, splenectomy, and lower AUC insulin.

Clinical outcome of febrile neutropenia in children with cancer using ceftazidime and aminoglycosides.

To determine treatment outcome using ceftazidime-aminoglycosides in febrile neutropenic children with cancer, the authors conducted a prospective cohort study in 216 episodes. Early and complete responses to antibiotics were 108/216 (50.0%) and 133/216 (61.6%) episodes, respectively. Death, a modification of antibiotic(s), and resistance to ceftazidime were 2/118 (1.7%), 73/216 (33.8%), and 4/216 (1.9%) episodes, respectively. Primary bacteremia and emerging bacteremia during treatment were 20/216 (9.3%) and 5/216 (2.3%) episodes. Ceftazidime-aminoglycosides was found to be a reasonable initial treatment of febrile neutropenia in the authors' institution. Imipenem is considered in patients who have clinical sepsis and who fail to respond to initial treatment.

Thyroid function in beta-thalassemic children receiving hypertransfusions with suboptimal iron-chelating therapy.

A cross-sectional study of thyroid function, free thyroxine (FT) and thyrotropin (TSH) concentrations, was carried out in 51 transfusion-dependent beta-thalassemic patients receiving suboptimal iron-chelating therapy. Nine patients had normal FT4 levels with elevated TSH levels (5.9-15.6 mLU/L), consistent with the diagnosis of compensated primary hypothyroidism and giving a prevalence of abnormal thyroid function of 17.6%. All patients with abnormal thyroid function had negative thyroid antibodies. No particular risk factor for abnormal thyroid function could be identified. Of the nine patients with compensated primary hypothyroidism, one patient showed a further increase in TSH level after 1 year of follow-up. The results of the present study emphasize the importance of thyroid function monitoring in hypertransfused beta-thalassemic patients.

A low dose adrenocorticotropin test (1 microg ACTH) for the evaluation of adrenal function in children with beta-thalassemia receiving hypertransfusion with suboptimal iron-chelating therapy.

A cross-sectional study of adrenal function was carried out in 48 patients with beta-thalassemia who were receiving hypertransfusion with suboptimal desferoxamine. A low dose adrenocorticotropic hormone (1 microg ACTH) stimulation test was performed using the cut-off criteria of peak cortisol for adrenal sufficiency >18 microg/dl. Adrenal impairment was diagnosed in 22 patients, giving a prevalence of 45.8%. The peak cortisol concentrations in normal and impaired adrenal function groups were 26.22 +/- 2.84 and 14.03 +/- 3.12 microg/dl, respectively, and the mean basal morning cortisol was 8.93 +/- 2.97 and 6.52 +/- 2.45 microg/dl, respectively. There was no significant difference in any clinical characteristic between the patients with impaired adrenal function and those with normal adrenal function.

Glucose-6-phosphate dehydrogenase variants associated with favism in Thai children.

In a study conducted at Songklanagarind Hospital in the south of Thailand, the subjects were 225 patients (210 boys and 15 girls) with glucose-6-phosphate dehydrogenase (G6PD) deficiency. Favism was found in 3.6% of the G6PD-deficient children. Approximately one half of the G6PD-deficient patients with favism were younger than 2 years. Sudden onset of anemia was found within 1 to 3 days after ingestion of dried fava beans. The classic features of favism, which are pallor, hemoglobinuria, and jaundice, were detected in all cases. To characterize the known G6PD mutations in Thai children, molecular analysis was performed for 8 G6PD-deficient children with favism by a combination of polymerase chain reaction-restriction fragment length polymorphism analysis and amplification refractory mutation system analysis. The G6PD variants in these children were G6PD Kaiping 1388,G-->A; G6PD Mahidol 487,G-->A; G6PD Viangchan 871,G-->A; and uncharacterized mutation with silent mutation 1311,C-->T.

Evaluation of mutation effects on UGT1A1 activity toward 17beta-estradiol using liquid chromatography-tandem mass spectrometry.

Mutations in the gene encoding UDP-glucuronosyltransferase 1A1 (UGT1A1) may reduce the glucuronidation of estradiol, bilirubin, etc. In the present study, we used a liquid chromatography-tandem mass spectrometry (LC/MS/MS) method to assay the activities of recombinant mutated UGT1A1 toward 17beta-estradiol (E2), by determining its glucuronide (E2G) content. Direct evidence for glucuronide formation was provided by E2G-specific ion peaks. The UGT1A1 activities of G71R (exon 1), F83L (exon 1), I322V (exon 2) and G493R (exon 5) mutants were 24, 30, 18 and 0.6% of the normal UGT1A1 activity, respectively. In conclusion, our study showed that LC/MS/MS enabled accurate evaluation of the effects of mutations on recombinant UGT1A1 activity towards E2.

Neonatal anemia associated with Southeast Asian ovalocytosis.

The purposes of this study were to evaluate the reliability of the previously described diagnostic criteria for Southeast Asian ovalocytosis (SAO) in adults in the diagnosis of SAO in newborns and to describe the role of SAO in newborn infants presenting with pallor and jaundice. The inclusion criteria in this retrospective descriptive study were that the patient be a newborn with pallor or jaundice and with ovalocytes in the peripheral blood smear (PBS). The exclusion criteria were newborn status with other causes of neonatal hemolysis or anemia. Controls were age-matched newborn infants who did not have SAO or other causes of neonatal anemia or hemolysis. Hematological data were assessed with a hematology analyzer. DNA analysis for SAO band 3 was done by polymerase chain reaction. Among 107 newborn infants with SAO, 30 infants were excluded from the study. The exclusions were premature infants, an infant with congenital syphilis, low-birth-weight infants, infants with ABO blood group incompatibility, infants with 3-thalassemia, infants with hemoglobin E heterozygote or homozygotes, glucose-6-phosphate dehydrogenase-deficient infants, and infants with fetomaternal hemorrhage. The DNA analysis for SAO band 3 was done in 56 newborns, and 54 had positive results for SAO band 3 gene deletion. Approximately one half of the 54 newborn infants with SAO had hyperbilirubinemia, and 3 had severe hyperbilirubinemia. The mean hemoglobin concentration, packed cell volume, and red blood cell (RBC) count in the infants with SAO in the first week of life were significantly lower than those in control infants. The mean absolute number of reticulocytes, mean corpuscular hemoglobin, and red cell volume distribution width in infants with SAO band 3 in the first week of life were significantly higher than those in control infants. The neonatal diagnosis of SAO can be made by examination of RBC morphology in the PBS with the presence of stomatocytes, theta cells, and > or = 25% ovalocytes. SAO plays a role in anemia and hyperbilirubinemia in newborn infants.

Molecular heterogeneity of glucose-6-phosphate dehydrogenase (G6PD) variants in the south of Thailand and identification of a novel variant (G6PD Songklanagarind).

Two hundred and twenty-five G6PD-deficient subjects in Songklanagarind Hospital in the south of Thailand comprising 210 males and 15 females were studied. Neonatal jaundice was detected in 85% of these patients. Acute hemolysis related to infection was detected in 17.3% of the G6PD-deficient subjects. Drug-induced acute hemolysis was detected in 1.8% and favism was observed in 3.6% of G6PD-deficient patients. The molecular analysis was performed on 134 G6PD-deficient individuals by a combination of PCR-RFLP, multiplex polymerase chain reaction by multiple tandem forward primers and a common reverse primer assay (MPTP) and DNA sequencing to characterize the mutations of the samples with abnormal MPTP bands. We found 10 different missense G6PD mutations and the three most common variants were G6PD Viangchan 871,G-->A (31.3%), G6PD Kaiping 1388,G-->A (20.1%) and G6PD Mahidol 487,G-->A (17.2%) followed by G6PD Canton 1376,G-->T (9.7%), G6PD Union 1360,C-->T (2.2%), G6PD Gaohe 95,A-->G (1.5%), G6PD Quing Yuan 392,G-->T (0.7%), G6PD Mediterranean 563,C-->T (0.7%), G6PD Songklanagarind 196,T-->A (0.7%), silent mutation 1311,C-->T (6.7%), and uncharacterized variant (9%). A novel missense mutation at codon 196, TTC-->ATC in exon 4 of the G6PD gene predicting a single amino acid substitution, Phe66Ile was identified and we designated this novel class II variant as G6PD Songklanagarind. The G6PD variants among the Thais in the southern part are heterogeneous and G6PD Viangchan, Kaiping, Mahidol, and Canton variants account for about 78% of the cases. Our findings provide some evidence that G6PD Viangchan and Mahidol are common Southeast Asian variants and support the theory of genetic drifts throughout Southeast Asia.

Growth patterns and final height of survivors of childhood leukemia.

Growth patterns of 85 survivors of childhood leukemia were analyzed retrospectively. All patients remained in first remission with no central nervous system involvement. The mean age at diagnosis was 5.8 +/- 3.6 years. The diagnoses were acute lymphoblastic leukemia (ALL) in 68 patients (80%) and acute non-lymphoblastic leukemia (ANLL) in 17 patients (20%). All except two patients received cranial irradiation: 51 patients with 1,800 cGy and 32 patients with 2,400 cGy. Mean height SDS was -0.7 +/- 1.36 at the time of diagnosis, which decreased to -0.92 +/- 1.31 by the end of treatment, and further decreased to -1.14 +/- 1.38 at 6 years after cessation of treatment. Mean weight SDS was -0.55 +/- 1.13 at the time of diagnosis, increasing slightly to -0.39 +/- 1.02 at the end of treatment, and decreasing to -0.46 +/- 1.65 at 6 years after cessation of treatment. Of these survivors, 51 patients (26 boys and 25 girls) reached a final height that was 1.04 SDS or 5.3 cm less than their target height. There was no difference of height and weight SDS between patients with ALL and ANLL. Girls and boys had different growth patterns. Girls had a slightly increased height SDS and gained more weight after cessation of treatment, resulting in less final height deficit and overweight for height, whereas boys had further height and weight reduction resulting in more deficit of final height.

Hb G-Makassar [beta6(A3)Glu-->Ala; codon 6 (GAG-->GCG)]: molecular characterization, clinical, and hematological effects.

We report a Thai family in which five members are Hb G-Makassar heterozygotes and one member is, in addition, a heterozygote for beta0-thalassemia (IVS-I-1, G-->T). We confirm that the previously presumed mutation at codon 6 of the beta-globin gene is GAG-->GCG. Hb G-Makassar heterozygotes are asymptomatic and hematologically normal. The Hb G-Makassar/beta0-thalassemia compound heterozygote has features of thalassemia minor. A simple and rapid polymerase chain reaction-restriction fragment length polymorphism for the detection of Hb G-Makassar is described.

Novel missense mutation of the UGT1A1 gene in Thai siblings with Gilbert's syndrome.

BACKGROUND: Gilbert's syndrome is a common inherited disorder of bilirubin metabolism contributing to the development of neonatal jaundice and causing recurrent jaundice after the neonatal period. In the patients with Gilbert's syndrome, mutations have been reported in the promoter and exons of the uridine diphosphate-glucuronosyl transferase 1 (UGT1A1) gene on chromsome 2q37, which encodes bilirubin uridine diphosphate-glucuronosyltransferase. However, the genetic basis of Gilbert's syndrome, including its inheritance trait, remains to be clarified. METHODS: Patients 1 and 2 were Thai sisters with Gilbert's syndrome. They had a history of prolonged neonatal jaundice and showed recurrent jaundice after their infancy, while the parents showed no symptoms. To search for the mutation in the patients, all exons of the UGT1A1 gene were amplified by polymerase chain reaction (PCR) and sequenced directly. The frequency of the mutation in controls was studied by PCR-restriction enzyme digestion method. RESULTS: The patients were homozygous for a novel single transition of T to C at nucleotide position 247 (exon 1), which would predict a substitution of leucine for phenylalanine at codon 83 of the enzyme protein. No other mutation was detected in any regions except exon 1. The parents with no symptoms showed heterozygosity for the mutation. Among the 110 Japanese controls, no homozygous individuals and three heterozygous individuals for the mutation were identified, giving a mutated allele frequency of 0.0136. CONCLUSIONS: A novel missense mutation in the UGT1A1 gene was identified in two Thai siblings with Gilbert's syndrome. The affected family showed that homozygosity for the mutation may lead to apparent symptoms and that the syndrome was inherited as an autosomal recessive trait. The mutation does not explain a high incidence of neonatal jaundice in Japan, because it is very rare in the Japanese population.