A Child's Nutrition and Epigenetics.

Studies have shown a dramatic increase in the incidence and the prevalence of chronic diseases such as type 2 diabetes mellitus and cardiovascular disorders over the last several decades. Environmental triggers and nutrition are considered major contributors to this increase. The first 1,000 days of life, which is the period between conception and the first 2 years of age, is considered the time for environmental factors, such as nutrition, to exert their positive and most crucial effects on a child's health. Nutrigenomics, the study of how genes and food components interact, looks into diet-altering disease development by modulating processes involved with the onset, progression, and severity of disease. These factors affecting the development of these chronic diseases are thought to be mediated by epigenetic mechanisms, which are heritable and reversible, and carry genetic information without changing the nucleotide sequence of the genome and are also mediated by maternal and postnatal nutrition.

Global Globin Network Consensus Paper: Classification and Stratified Roadmaps for Improved Thalassaemia Care and Prevention in 32 Countries.

The Global Globin Network (GGN) is a project-wide initiative of the Human Variome/Global Variome Project (HVP) focusing on haemoglobinopathies to build the capacity for genomic diagnosis, clinical services, and research in low- and middle-income countries. At present, there is no framework to evaluate the improvement of care, treatment, and prevention of thalassaemia and other haemoglobinopathies globally, despite thalassaemia being one of the most common monogenic diseases worldwide. Here, we propose a universally applicable system for evaluating and grouping countries based on qualitative indicators according to the quality of care, treatment, and prevention of haemoglobinopathies. We also apply this system to GGN countries as proof of principle. To this end, qualitative indicators were extracted from the IthaMaps database of the ITHANET portal, which allowed four groups of countries (A, B, C, and D) to be defined based on major qualitative indicators, supported by minor qualitative indicators for countries with limited resource settings and by the overall haemoglobinopathy carrier frequency for the target countries of immigration. The proposed rubrics and accumulative scores will help analyse the performance and improvement of care, treatment, and prevention of haemoglobinopathies in the GGN and beyond. Our proposed criteria complement future data collection from GGN countries to help monitor the quality of services for haemoglobinopathies, provide ongoing estimates for services and epidemiology in GGN countries, and note the contribution of the GGN to a local and global reduction of disease burden.

Functional analysis of GALT variants found in classic galactosemia patients using a novel cell-free translation method.

Classic galactosemia is an autosomal recessive disorder caused by deleterious variants in the galactose-1-phosphate uridylyltransferase (GALT) gene. GALT enzyme deficiency leads to an increase in the levels of galactose and its metabolites in the blood causing neurodevelopmental and other clinical complications in affected individuals. Two GALT variants NM_000155.3:c.347T>C (p.Leu116Pro) and NM_000155.3:c.533T>G (p.Met178Arg) were previously detected in Filipino patients. Here, we determine their functional effects on the GALT enzyme through in silico analysis and a novel experimental approach using a HeLa-based cell-free protein expression system. Enzyme activity was not detected for the p.Leu116Pro protein variant, while only 4.5% of wild-type activity was detected for the p.Met178Arg protein variant. Computational analysis of the variants revealed destabilizing structural effects and suggested protein misfolding as the potential mechanism of enzymological impairment. Biochemical and computational data support the classification of p.Leu116Pro and p.Met178Arg variants as pathogenic. Moreover, the protein expression method developed has utility for future studies of GALT variants.

Clinical, biochemical and molecular characteristics of Filipino patients with mucopolysaccharidosis type II - Hunter syndrome.

BACKGROUND: Mucopolysaccharidosis type II, an X-linked recessive disorder is the most common lysosomal storage disease detected among Filipinos. This is a case series involving 23 male Filipino patients confirmed to have Hunter syndrome. The clinical and biochemical characteristics were obtained and mutation testing of the IDS gene was done on the probands and their female relatives. RESULTS: The mean age of the patients was 11.28 (SD 4.10) years with an average symptom onset at 1.2 (SD 1.4) years. The mean age at biochemical diagnosis was 8 (SD 3.2) years. The early clinical characteristics were developmental delay, joint stiffness, coarse facies, recurrent respiratory tract infections, abdominal distention and hernia. Majority of the patients had joint contractures, severe intellectual disability, error of refraction, hearing loss and valvular regurgitation on subspecialists' evaluation. The mean GAG concentration was 506.5 mg (SD 191.3)/grams creatinine while the mean plasma iduronate-2-sulfatase activity was 0.86 (SD 0.79) nmol/mg plasma/4 h. Fourteen (14) mutations were found: 6 missense (42.9%), 4 nonsense (28.6%), 2 frameshift (14.3%), 1 exon skipping at the cDNA level (7.1%), and 1 gross insertion (7.1%). Six (6) novel mutations were observed (43%): p.C422F, p.P86Rfs*44, p.Q121*, p.L209Wfs*4, p.T409R, and c.1461_1462insN[710]. CONCLUSION: The age at diagnosis in this series was much delayed and majority of the patients presented with severe neurologic impairment. The results of the biochemical tests did not contribute to the phenotypic classification of patients. The effects of the mutations were consistent with the severe phenotype seen in the majority of the patients.

Association of glutathione-S-transferase M1,P1, and T1 gene polymorphisms with treatment response in childhood acute lymphocytic leukemia

@#<p style="text-align: justify;">There is still a strong need for new treatment strategies that will maintain remission and prolong survival in patients with acute lymphoblastic leukemia (ALL). The glutathione-S-transferase (GST) enzymes, which are coded by highly polymorphic genes, have been associated with the risk of developing cancer and were found to regulate effect of cancer treatment drugs.<br /><strong>OBJECTIVES:</strong> The present study determines the association of GSTM1, GSTP1 and GSTT1 polymorphisms and treatment response in terms of occurrence of adverse events and relapse in ALL in Filipino children.<br /><strong>METHODS:</strong> This is a follow up study on the 2007 investigation done by Alcausin et al. which determined the association of the GST P1, M1, and T1 polymorphisms and occurrence of ALL. Four-year follow-up data were available for 46 out of the 50 patients from January 2007 to May 2011. Odds ratios (OR) as measures of association of GST M1, P1 and T1 gene polymorphisms with treatment outcomes were estimated at 95% confidence interval.<br /><strong>RESULTS:</strong> Results show a trend towards predisposition to elevation of liver enzymes in patients with GSTT1 and GSTP1 mutant genotypes showing an OR (95% Cl) of 2.0 (0.62-6.49). The presence of GSTM1 null genotype showed a trend towards protection from occurrence of relapse basing on both crude and adjusted ORs, 0.58 (0.16-2.07) and 0.23 (0.05-1.20), respectively. However, these results are not statistically significant.<br /><strong>CONCLUSION: </strong>The GSTP1 heterozygous genotype conferred increased predisposition to elevation of liver enzymes while the GSTT1 null genotype was shown to be a possible risk factor towards the occurrence of both infection and elevation of liver enzymes during chemotherapy. Furthermore, the GSTM1 null genotype appears to be protective from occurrence of relapse. It is recommended to do similar large-scale studies in the future to obtain more conclusive results.</p>

Novel cystathionine ß-synthase gene mutations in a Filipino patient with classic homocystinuria.

BACKGROUND: Classic homocystinuria due to cystathionine ß-synthase (CBS) deficiency is an autosomal recessive disorder of sulfur metabolism. Clinical manifestations include mental retardation, dislocation of the optic lens (ectopia lentis), skeletal abnormalities and a tendency to thromboembolic episodes. We present the first mutational analysis of CBS in a Filipino patient with classic homocystinuria. METHODS: Genomic DNA was extracted from peripheral blood collected from a diagnosed Filipino patient with classic homocystinuria. The entire coding region of CBS (17 exons) was amplified using polymerase chain reaction and bidirectionally sequenced using standard protocols. RESULTS: The patient was found to be compound heterozygous for two novel mutations, g.13995G>A [c.982G>A; p.D328K] and g.15860-15868dupGCAGGAGCT [c.1083-1091dupGCAGGAGCT; p. Q362-L364dupQEL]. Four known single-nucleotide polymorphisms (rs234706, rs1801181, rs706208 and rs706209) were also detected in the present patient's CBS. The patient was heterozygous for all the identified alleles. CONCLUSIONS: This is the first mutational analysis of CBS done in a Filipino patient with classic homocystinuria who presented with a novel duplication mutation and a novel missense mutation. Homocystinuria due to CBS deficiency is a heterogeneous disorder at the molecular level.

Mutational analysis of the GALT gene in Filipino patients.

Classic galactosemia is an inherited metabolic disorder due to mutations in the galactose-1-phosphate uridyltransferase (GALT) gene. This study describes the results of the GALT gene analysis of four unrelated Filipino patients with Classic Galactosemia. DNA extracted from dried blood spots and peripheral blood of the patients, age one month to two and a half years, underwent PCR-amplification with subsequent bidirectional sequencing of all eleven exons with their flanking intronic regions following standard protocols. Clinical data of these patients were reviewed. The patients presented with jaundice, hepatomegaly, diarrhea, vomiting, poor feeding and seizures during their neonatal period. They were diagnosed with elevated blood galactose and galactose-1-phosphate and absent GALT activity. Four missense mutations were found wherein two were previously reported (p.V168L and p.A345D) and two were novel (p.L116P and p.M178R). The most frequent mutation in our cohort is p.V168L. This study suggests that GALT mutations are ethnic-specific and that galactosemia is a heterogeneous disorder at the molecular level. The importance of early detection, immediate and proper medical management and genetic counseling of the patients and their families cannot be overemphasized.

Genetic polymorphisms in CYP1A1, GSTM1, GSTP1, GSTT1, NAT1 and NAT2 and oral cavity cancer risk among Filipinos

Polymorphisms in metabolic genes have been shown to modulate susceptibility to oral cavity cancer. Cases (n=176) and controls (n=317) from the Filipino population were genotyped for selected polymorphisms in CYP1A1, GSTM1, GSTP1, GSTT1, NAT1 and NAT2. Medical and diet histories, occupational exposure and demographic data were also collected for all subjects. The CYP1A1m1/m1 genotype is protective against oral cancer, while being homozygous for the GSTP1 c.313G genotype and heterozygous for the NAT1*10 homozygotes and non-homozygotes for the CYP1A1 m1 allele. The risk from heterozygosity for the NAT1*10 allele was limited to subjects who were not homozygous for the GSTP1 c.313G genotype remained a significant oral cancer risk modifier, together with environmental variables, the homozygous GSTP1 c.313G genotype remained a significant oral cancer risk modifier, together with environmental risk factors, such as smoking, passive smoking, inverted smoking and tobacco chewing, and environmental protective factors, i.e. moderate consumption of fish sauce (patis) and shrimp paste (bagoong). The GSTP1 c.313G polymorphism increases susceptibility for oral cavity cancer in the Filipino population.

Diversity of approaches to classic galactosemia around the world: a comparison of diagnosis, intervention, and outcomes.

Without intervention, classic galactosemia is a potentially fatal disorder in infancy. With the benefit of early diagnosis and dietary restriction of galactose, the acute sequelae of classic galactosemia can be prevented or reversed. However, despite early and lifelong dietary treatment, many galactosemic patients go on to experience serious long-term complications including cognitive disability, speech problems, neurological and/or movement disorders and, in girls and women, ovarian dysfunction. Further, there remains uncertainty surrounding what constitutes a 'best practice' for treating this disorder. To explore the extent and implications of this uncertainty, we conducted a small but global survey of healthcare providers who follow patients with classic galactosemia, seeking to compare established protocols for diagnosis, intervention, and follow-up, as well as the outcomes and outcome frequencies seen in the patient populations cared for by these providers. We received 13 survey responses representing five continents and 11 countries. Respondents underscored disparities in approaches to diagnosis, management and follow-up care. Notably, we saw no clear relationship between differing approaches to care and long-term outcomes in the populations studied. Negative outcomes occurred in the majority of cases regardless of when treatment was initiated, how tightly galactose intake was restricted, or how closely patients were monitored. We document here what is, to our knowledge, the first global comparison of healthcare approaches to classic galactosemia. These data reinforce the idea that there is currently no one best practice for treating patients with classic galactosemia, and underscore the need for more extensive and statistically powerful comparative studies to reveal potential positive or negative impacts of differing approaches.

Characterization of mutations and polymorphisms in the G6PD among Filipino newborns with glucose-6-phosphate dehydrogenase deficiency

Background. Glucose-6-phosphate (G6PD) deficiency is the most prevalent enzyme deficiency to date. The global prevalence of G6PD deficiency is estimated at around 330 million people affected with the disease worldwide. This 4.9 percent prevalence, correlates highly with geographic areas endemic to malaria. It is the most common among the disorders in the Newborn Screening (NBS) panel in the Philippines, with one confirmed case for every 52 newborns (1:52). This paper determines the molecular background of G6PD deficiency among Filipino newborns detected by newborn screening. Methods. A total of 200 cases confirmed to have G6PD deficiency, 180 males and 20 females, were identified through the Philippine Newborn Screening Program from 2001-2003. Genomic DNA was extracted from dried blood spots followed by multiplex polymerase chain reaction using multiple tandem forward primers and a common reverse primer (MPTP) to detect previously reported common mutations and polymorphisms in exons 5, 6, 9, 11 and 12 of the G6PD gene. Results. Of the 200 samples analyzed, mutations and polymorphisms in the G6PD gene were identified in 148 cases (74%). The most common mutation was a G to A transition on nucleotide 871 (Viangchan) of exon 9 in combination with a silent mutation on exon 11, accounting for 32.9% of the cases. This was followed by a C to T transition on nucleotide 1360 (Union) in 21.1 % of the cases. Other mutations were Vanua Lava in 10%, Chatham in 9.4% and Canton in 3.5% of the newborns. The silent polymorphism on nucleotide 1311 was present in 12.9% of cases. There were combinations of these mutations and polymorphisms present in a minority of cases. Conclusion. Results of this study showed the molecular heterogeneity underlying G6PD deficiency among Filipino newborns.

Genotype-phenotype correlations in Filipino patients with type 3 gaucher disease

Gaucher disease is an inherited glycolipid storage disorder caused by a deficiency of the lysosomal enzyme glucocerebrosidase. Clinical manifestations include hepatosplenomegaly, skeletal abnormalities, anemia and thrombocytopenia. We present here the corresponding genotypes and the genotype-phenotype correlations of 3 Filipino patients. Clinical phenotypes and genotypes were documented by reviewing the charts of 3 Filipino patients with Gaucher disease. Clinical parameters such as liver and spleen sizes, hematologic variables, disease types and response to enzyme replacement therapy were compared. Likewise, quantitative enzyme assays and mutation analysis were reviewed. All have the type III neuronopathic Gaucher disease. Patients 1 and 2 are twin sisters who both have mild mental retardation with Patient 1 having a concomitant seizure disorder. They have the corresponding genotype of p.L444/p.P319A. Patient 3 has global developmental delay, oculomotor apraxia, pyramidal tract signs and carries the p.L444P/p.G202R/p.G202R genotype. Genotype-phenotype correlations for the 3 patients showed that their genotypes are compatible with the severe neuronopathic type of disease.

A master of science in genetic counseling program in the Philippines

In the Philippines, there is an urgent need to expand the clinical services for diagnosis, management and emotional support for patients with genetic conditions and their family members. Despite the lack of trained providers with specialization in genetics, public health related geneticsprograms are continuously being implemented. These address these current demands,strategic planning began in 2009 between local medical geneticists and international genetic professionals to develop the curriculum for an advanced degree in genetic counseling program. The board of regents at the University of the Philippines approved the proposed curriculum in January 2011, and training of the Philippines first cohort of genetic counseling students commenced in June 2011. The successful implementation of the MS of Genetic Counseling program will provide the opportunity to incorporate the match needed genetic counseling services in the country.

PD1 as a common candidate susceptibility gene of subacute sclerosing panencephalitis.

Although the exact pathogenesis of subacute sclerosing panencephalitis (SSPE) remains to be determined, our previous data suggested a genetic contribution to the host susceptibility to SSPE. During chronic viral infection, virus-specific cytotoxic T lymphocytes display poor effector functions. Since co-inhibitory molecules are involved in the suppression of T lymphocytes, we investigated whether single nucleotide polymorphisms (SNPs) of genes encoding co-inhibitory molecules contributed to a susceptibility to SSPE. Association studies on a total of 20 SNPs in 8 genes (CTLA4, CD80, CD86, PD1, PDL1, PDL2, BTLA and HVEM) and subsequent haplotype analysis of 4 SNPs in the PD1 genes were performed in Japanese and Filipino SSPE patients and controls. Then, we investigated a functional difference in promoter activity of two haplotypes and compared the expression levels of PD1 between SSPE and controls. The frequency of GCG(C) haplotype of PD1 containing -606G allele was significantly higher in SSPE patients than in controls both in Japanese and in Filipinos. The promoter activity was significantly higher in the construct with -606G allele than in that with -606A allele. The expression levels of PD1 were significantly higher in SSPE patients than in the controls. Our results suggested that the PD1 gene contributed to a genetic susceptibility to SSPE.

Mutations of the phenylalanine hydroxylase (PAH) gene in Filipino patients with phenylketonuria

INTRODUCTION: Phenylketonuria (PKU), an autosomal recessive metabolic disorder caused by phenylalanine hydroxylase (PAH) deficiency, leads to hyperphenylalaninemia and neurological damage if untreated. This is the first study in the Philippines to identify the disease-causing mutations in the PAH gene of clinically diagnosed Filipino PKU patients. METHODS: The study included four unrelated PKU patients detected by the Philippine Newborn Screening Program from 1996 to 2008. Plasma amino acid analyses for all patients showed increased phenylalanine and low to normal tyrosine levels consistent with the diagnosis of PKU. Mutations in the PAH gene were identified by genomic DNA extraction from dried blood spots of the patients, PAH exon amplification by polymerase chain reaction and subsequent bi-directional DNA sequence analysis. RESULTS: All patients presented with significantly elevated phenylalanine levels on bacterial inhibition assay and thin layer chromatography. Urinary pterins confirmed the diagnosis of Tetrahydrobiopterin deficiency in two patients while the other 2 patients had the Classical PKU phenotype. Four previously identified mutations in the PAH gene (p.I65T, p.R413P, p.EX6-96A>G, p.R243Q) were identified in those with Classical PKU. CONCLUSION: The present results confirm the heterogeneity of mutations at the PAH locus in Filipinos. Neonatal screening and the use of molecular diagnosis significantly aid in the medical management and genetic counseling of patients and their families.

Mutations of the steroid 21-hydroxylase gene among Filipino patients with congenital adrenal hyperplasia

Congenital adrenal hyperplasia (CAH), an autosomal recessive disorder, is due to deficiency of the enzymes involved in adrenal steroidogenesis. Phenotypic manifestations vary as a result of the degree of glucocorticoid or mineralocorticoid deficiency and androgen excess present. Among Filipinos, the estimated crude incidence of CAH is approximately 1 in 7,000, which is higher than what is reported in most populations. More than 90% of all cases result from a 21-hydroxylase (21-OH) (cytochrome P450c21) enzyme deficiency involving two 21-OH genes, the active gene (CYP21) and a pseudogene (CYP21P). Studies have shown that mutations result from unequal crossover during meiosis which leads to complete deletion of the gene, gene conversion events or to point mutations. To date, there are no published data on the types of mutations present among Filipinos diagnosed with congenital adrenal hyperplasia. The objective of this study is to describe the profile of Filipino patients diagnosed with CAH and to determine the disease-causing alleles in the 21-OH gene of these patients. Using a method of combined differential polymerase chain reaction and amplification created restriction site approach, direct probing for the presence of known mutations in exons 1,3,4,6,7,8 and intron 2 of the CYP21 and CYP21P genes among Filipino patients with CAH was performed. A total of 12 unrelated CAH patients were examined. A majority of these cases had a premature splicing error mutation at nucleotide 656 of intron 2. The determination of the most frequent alleles in our population can facilitate rapid screening for mutations in the 21-OH gene and lead to a definitive diagnosis of CAH.

Glucose-6-phosphate dehydrogenase deficiency in Filipino neonates with jaundice

OBJECTIVES: Glucose-6-phosphate dehydrogenase (G6PD) deficiency is the most common of all clinically significant enzyme defects of red blood cells. It has a high rate of prevalence in the Philippines. Concern about hemolytic anemia and jaundice due to unrecognized G6PD deficiency led us to determine the prevalence of G6PD deficiency among jaundiced neonates in the Philippine General Hospital, a tertiary referral hospital in the Philippines. It was hypothesized that G6PD deficiency was more prevalent in neonates with jaundice than in the normal population. We also compared the clinical presentation and course (hospital stay and days of phototherapy requirement) for G6PD deficient and G6PD normal neonates. MATERIALS AND METHODS: We studied 102 clinically jaundiced neonates admitted to the nursery of the Philippine General Hospital. Blood samples in individual microtainers were quantitatively tested for G6PD activity using a commercial G6PD assay kit. The clinical presentation and hospital courses of patients were statistically compared using the t-test for single proportions. RESULTS: G6PD deficiency was diagnosed in 17 of 102 cases[16.7% (95% CI: 10.0 to 25.3)], which is significantly higher than the normal population (p CONCLUSION: The prevalence of G6PD deficiency among jaundiced neonates was found to be higher than the normal population thus, early detection of this enzymopathy, regardless of sex, and close surveillance of the affected newborns is important in reducing the risk of severe hyperbilirubinemia.

Early diagnosis of maple syrup urine disease using polymerase chain reaction-based mutation detection.

BACKGROUND: Maple syrup urine disease (MSUD) is an autosomal recessive disorder caused by defective activity of the branched-chain alpha-ketoacid dehydrogenase enzyme complex. Early diagnosis and management of MSUD are imperative for preventing permanent neurological impairments. In the Philippines, a 4.7 kb deletion in the dihydrolipoamide branched-chain transacylase E2 (DBT) gene has been commonly identified in MSUD patients. Polymerase chain reaction (PCR) amplification of a junction fragment between intron 10 and exon 11 has been used to detect this deletion. The purpose of the present paper was to use PCR-based mutation detection of the deletion mutation to diagnose MSUD in neonates in order to provide proper diagnosis and effective treatment. METHODS: A region encompassing exon 11 and the junction fragment of the E2 (DBT) gene was PCR amplified from genomic DNA prepared from two neonates at risk for MSUD. RESULTS: PCR amplification of both exon 11 and the junction fragment from one of the neonates demonstrated that this case was a heterozygous carrier of the deletion. Thus, normal feeding was started. For the other neonate, PCR amplification of the junction fragment was successful, whereas the region encompassing exon 11 was not amplified. This neonate was genotyped as homozygous for the deletion, and treatment for MSUD was provided immediately. CONCLUSION: Examination of the deletion mutation in the E2 (DBT) gene facilitated early MSUD diagnosis and was beneficial for the determination of the proper course of treatment.

A novel deletion creating a new terminal exon of the dihydrolipoyl transacylase gene is a founder mutation of Filipino maple syrup urine disease.

Maple syrup urine disease (MSUD) is a rare, autosomal-recessive disorder of branched-chain amino-acid metabolism. In the Philippines, many MSUD cases have been diagnosed clinically. Here, molecular analysis of the dihydrolipoyl transacylase (E2) gene was done in 13 unrelated families from the Philippines. A novel deletion spanning 4.1 kb of intron 10 and 601 bp of exon 11, caused by non-homologous recombination between an L1 repeat in intron 10 and an Alu repeat in exon 11, was found in 8 out of 13 families, with 5 of them being homozygous for the mutation, implicating it as a founder mutation of Filipino MSUD. The resulting mutant E2 mRNA contains a 239-bp insertion after exon 10, thereby producing a new terminal exon. Large-scale population screening of the deletion revealed that one carrier of the mutation was identified in 100 normal Filipinos. These findings suggest that a limited number of mutations might underlie MSUD in the Filipino population, potentially facilitating prenatal diagnosis and carrier detection of MSUD in this group.