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Biomedical subjects

George P Patrinos

Publications and source records attributed to George P Patrinos.

At least 19 recordsLinked to original sources
PubMed2025

Integrating advanced analytical methods to assess epigenetic marks affecting response to hypomethylating agents in higher risk myelodysplastic syndrome.

BACKGROUND: Patients with higher-risk (HR) myelodysplastic syndrome (MDS), ineligible for allogeneic hematopoietic stem cell transplantation (alloHSCT), require prompt therapeutic interventions, such as treatment with hypomethylating agents (HMAs) to restore normal DNA methylation patterns, mainly of oncosuppressor genes, and consequently to delay disease progression and increase overall survival (OS). However, response assessment to HMA treatment relies on conventional methods with limited capacity to uncover a wide spectrum of underlying molecular events. METHODS: We implemented liquid chromatography-tandem mass spectrometry (LC-MS/MS) to assess 5' methyl-2' deoxycytidine (5mdC), 5' hydroxy-methyl-2'-deoxycytidine (5hmdC) levels and global adenosine/thymidine ([dA]/[T]) ratio in bone marrow aspirates from twenty-one HR MDS patients, pre- and post-HMA treatment. Additionally, targeted methylation analysis was performed by interpretation of NGS-methylation (MeD-seq) data obtained from the same patient cohort. RESULTS: LC/MS-MS analysis revealed a significant hypomethylation status in responders (Rs), already established at baseline and a trend for further DNA methylation reduction post-HMA treatment. Non-responders (NRs) reached statistical significance for DNA hypomethylation only post-HMA treatment. The 5hmdC epigenetic mark was approximately detected at 37.5-40% among NRs and Rs, implying the impairment of the natural active demethylation pathway, mediated by the ten-eleven (TET) 5mdC dioxygenases. R and NR subgroups displayed a [dA]/[T] ratio&#x2009;<&#x2009;1 (0.727&#x2009;-&#x2009;0.633), supporting high frequences of 5mdC transition to thymidine. Response to treatment, according to whole genome MeD-seq data analysis, was associated with specific, scattered hypomethylated DMRs, rather than presenting a global effect across genome. MeD-seq analysis identified divergent epigenetic effects along chromosomes 7, 9, 12, 16, 18, 21, 22, X and Y. Within statistically significant selected chromosomal bins, genes encoding for proteins and non-coding RNAs with reversed methylation profiles between Rs and NRs, were highlighted. CONCLUSIONS: Implementation of powerful analytical tools to identify the dynamic DNA methylation changes in HR MDS patients undergoing HMA therapy demonstrated that LC-MS/MS exerts high efficiency as a broad-based but rapid and cost-effective methodology (compared to MeD-seq) to decode different perspectives of the epigenetic background of HR MDS patients and possess discriminative efficacy of the response phenotype to HMA treatment.

Humans↗
PubMed2007

A versatile denaturing HPLC approach for human beta-globin gene mutation screening.

Hemoglobinopathies represent the most common genetic disorder worldwide, with a higher prevalence among populations with a history of malaria endemicity. More than 690 mutations in the human beta-globin gene are usually the cause of beta-type hemoglobinopathies. Here, we report a rapid and highly sensitive beta-globin gene mutation screening approach based on denaturing high-performance liquid chromatography (DHPLC), which contrary to the previously described ones can be used in every HPLC apparatus. The sensitivity and specificity of the method were tested in 120 healthy Greek subjects and 25 beta-thalassemia heterozygotes and homozygotes, in which 11 different beta-globin sequence variations had been previously characterized by denaturing gradient gel electrophoresis. Using this method, we were able to rapidly identify the commonest beta-globin gene mutations, accounting for more than 90% of the mutant beta-globin alleles reported for the Hellenic population. Compared to classical mutation screening approaches, our DHPLC approach provides the means for rapid, highly sensitive, cost-effective, and semi-automated simultaneous mutational scanning of a large number of samples.

Chromatography, High Pressure Liquid↗
PubMed2007

HbVar database of human hemoglobin variants and thalassemia mutations: 2007 update.

HbVar (http://globin.bx.psu.edu/hbvar) is a locus-specific database (LSDB) developed in 2001 by a multi-center academic effort to provide timely information on the genomic sequence changes leading to hemoglobin variants and all types of thalassemia and hemoglobinopathies. Database records include extensive phenotypic descriptions, biochemical and hematological effects, associated pathology, and ethnic occurrence, accompanied by mutation frequencies and references. In addition to the regular updates to entries, we report significant advances and updates, which can be useful not only for HbVar users but also for other LSDB development and curation in general. The query page provides more functionality but in a simpler, more user-friendly format and known single nucleotide polymorphisms in the human alpha- and beta-globin loci are provided automatically. Population-specific beta-thalassemia mutation frequencies for 31 population groups have been added and/or modified and the previously reported delta- and alpha-thalassemia mutation frequency data from 10 population groups have also been incorporated. In addition, an independent flat-file database, named XPRbase (http://www.goldenhelix.org/xprbase), has been developed and linked to the main HbVar web page to provide a succinct listing of 51 experimental protocols available for globin gene mutation screening. These updates significantly augment the database profile and quality of information provided, which should increase the already high impact of the HbVar database, while its combination with the UCSC powerful genome browser and the ITHANET web portal paves the way for drawing connections of clinical importance, that is from genome to function to phenotype.

DNA Mutational Analysis↗
PubMed2006

FINDbase: a relational database recording frequencies of genetic defects leading to inherited disorders worldwide.

Frequency of INherited Disorders database (FINDbase) (http://www.findbase.org) is a relational database, derived from the ETHNOS software, recording frequencies of causative mutations leading to inherited disorders worldwide. Database records include the population and ethnic group, the disorder name and the related gene, accompanied by links to any corresponding locus-specific mutation database, to the respective Online Mendelian Inheritance in Man entries and the mutation together with its frequency in that population. The initial information is derived from the published literature, locus-specific databases and genetic disease consortia. FINDbase offers a user-friendly query interface, providing instant access to the list and frequencies of the different mutations. Query outputs can be either in a table or graphical format, accompanied by reference(s) on the data source. Registered users from three different groups, namely administrator, national coordinator and curator, are responsible for database curation and/or data entry/correction online via a password-protected interface. Databaseaccess is free of charge and there are no registration requirements for data querying. FINDbase provides a simple, web-based system for population-based mutation data collection and retrieval and can serve not only as a valuable online tool for molecular genetic testing of inherited disorders but also as a non-profit model for sustainable database funding, in the form of a 'database-journal'.

Databases, Genetic↗
PubMed2006

Mutation screening in the human epsilon-globin gene using single-strand conformation polymorphism analysis.

The human epsilon-globin gene is necessary for primitive human erythropoiesis in the yolk sac. Herein we report a non-radioactive single-strand conformation polymorphism (SSCP) approach to screen the human epsilon-globin gene and its regulatory regions for possible mutations and single-nucleotide polymorphisms in normal adult subjects, in order to determine those genomic regions, which are not necessary for its proper regulation and function. We identified no sequence variations apart from the expected 5'epsilon /HincII polymorphism in the fragments analyzed, suggesting that genomic alterations in the epsilon-globin gene are most likely incompatible with normal erythropoiesis and proper embryonic development.

DNA Mutational Analysis↗
PubMed2006

National and ethnic mutation databases: recording populations' genography.

Genetic databases are gradually assuming an increasing importance in all areas of health care. The national and ethnic mutation databases (NEMDBs) are continuously updated mutation depositories, recording extensive information over the described genetic heterogeneity of an ethnic group or population. Together with the central and locus-specific databases, those resources not only enhance awareness of the various genetic disorders but also facilitate the provision of genetic services and provide useful insights into the genographic history of human populations. Fifteen independent NEMDBs devoted to the documentation of the extant genetic heterogeneity in various population groups within 57 different countries were assessed; 13 of the NEMDBs were fully functional. The contents of the 13 fully functional NEMDBs were thoroughly analyzed for the presence or absence of 39 criteria, pertaining database general information, operating platform, data source and submission, and querying capacity. This study provides a strong case for uniformity of data to make the NEMDBs content maximally useful. In this direction, a hypothetical content for the ideal NEMDB is derived, which is currently being incorporated in an upgraded version of the ETHNOS NEMDB development and curation software, as well as a community structure that would enhance the chances of mutation frequency capture and documentation in human populations. The ultimate goal is that interested parties and granting bodies will assist in achieving the vision of a comprehensive resource that collects and displays all population-specific genetic information discovered.

Computational Biology↗
PubMed2006

The cypriot and Iranian National Mutation Frequency Databases.

The National Mutation Frequency Databases are continuously updated mutation depositories, which contain extensive information over the described genetic heterogeneity of an ethnic group or population. Here, we report the construction of the Cypriot (http://www.goldenhelix.org/cypriot) and Iranian National Mutation Frequency Databases (http://www.goldenhelix.org/iranian), both derived from an academic effort to provide high quality and up-to-date information on the underlying genetic heterogeneity of inherited disorders in the Cypriot and Iranian populations, respectively. Both databases have been built and maintained online using ETHNOS platform, a specialized software, which provides the means for national mutation database construction and curation. Each database contains brief summaries of the various genetic disorders studied for each population, and an easy-to-use query interface provides, both to specialist as well as to non-specialist users (i.e. patients and their families), instant access to the list and frequencies of the different mutations responsible for the inherited disorders in these populations. Furthermore, numerous links to the respective Online Mendelian Inheritance in Man (OMIM) entries and, when available, to the locus-specific databases fruitfully integrate the databases content into a single Web site. Both databases can serve as valuable online tools for molecular genetic testing of inherited disorders in these populations and could potentially motivate further investigations of yet unknown genetic diseases in the Cypriot and Iranian populations.

Cyprus↗
PubMed2005

Evidence for the molecular heterogeneity of sickle cell anemia chromosomes bearing the betaS/Benin haplotype.

There are at least four distinct African and one Asian chromosomal backgrounds (haplotypes) on which the sickle cell mutation has arisen. Additionally, previous data suggest that the beta(S)/Bantu haplotype is heterogeneous at the molecular level. Here, we report the presence of the (A)gamma -499 T-->A variation in sickle cell anemia chromosomes of Sicilian and North African origin bearing the beta(S)/Benin haplotype. Being absent from North American beta(S)/Benin chromosomes, which were studied previously, this variation is indicative for the molecular heterogeneity of the beta(S)/Benin haplotype.

Africa, Northern↗
PubMed2005

Hellenic National Mutation database: a prototype database for mutations leading to inherited disorders in the Hellenic population.

The exponential discovery rate of new genomic alterations, leading to inherited disorders, as well as the need for comparative studies of different population's mutation frequencies necessitates recording their population-wide spectrum in online mutation databases. We report the construction of the Hellenic National Mutation database (http://www.goldenhelix.org/hellenic), a prototype database derived from a multicenter academic initiative, aiming to provide high quality and up-to-date information on the underlying genetic heterogeneity of inherited disorders found in the Hellenic population. Database records include informative summaries of the various genetic disorders studied in the Hellenic population, focused in particular on their incidence in Greece, a comprehensive reference list, and a well-structured query interface, which provides easy access to the list of the different mutations responsible for the inherited disorders in the Hellenic population. Also, extensive links to the respective Online Mendelian Inheritance in Man (OMIM) entries and, when available, to the locus-specific databases are provided, so that the user can retrieve the maximum amount of information from a single website. Furthermore, the Hellenic National Mutation database design allows easy data entry and curation. Creation of the Hellenic National Mutation database will significantly facilitate molecular diagnosis of inherited disorders in Greece and will motivate further investigation of yet unknown genetic diseases in the Hellenic population.

Computational Biology↗
PubMed2005

Molecular diagnosis of inherited disorders: lessons from hemoglobinopathies.

Hemoglobinopathies constitute a major health problem worldwide, with a high carrier frequency, particularly in certain regions where malaria has been endemic. These disorders are characterized by a vast clinical and hematological phenotypic heterogeneity. Over 1,200 different genetic alterations that affect the DNA sequence of the human alpha-like (HBZ, HBA2, HBA1, and HBQ1) and beta-like (HBE1, HBG2, HBG1, HBD, and HBB) globin genes are mainly responsible for the observed clinical heterogeneity. These mutations, together with detailed information about the resulting phenotype, are documented in the globin locus-specific HbVar database. Family studies and comprehensive hematological analyses provide useful insights for accurately diagnosing thalassemia at the DNA level. For this purpose, numerous techniques can provide accurate, rapid, and cost-effective identification of the underlying genetic defect in affected individuals. The aim of this article is to review the diverse methodological and technical platforms available for the molecular diagnosis of inherited disorders, using thalassemia and hemoglobinopathies as a model. This article also attempts to shed light on issues closely related to thalassemia diagnostics, such as prenatal and preimplantation genetic diagnoses and genetic counseling, for better-quality disease management.

Databases, Nucleic Acid↗
PubMed2005

DNA, diseases and databases: disastrously deficient.

Recent progress in disease genetics and genome-related medicine has been substantial, with vast amounts of data being generated. However, this progress has not been matched by adequate database projects that gather and organize these data to enable their useful exploitation. This research area is complex, entailing core databases, locus-specific databases, national mutation databases, genotype-phenotype databases and patient databases--and much work is required to develop and properly integrate these various resources. To promote this, we present a timely overview of the field, emphasize its over-riding importance and discuss the disastrously deficient progress made so far. Many factors contribute to this slow progress (e.g. technological hurdles, publication requirements, the short-sighted and popularist research system). A lack of targeted funding is arguably the most fundamental problem, but one that can be solved.

Databases, Factual↗
PubMed2005

Isolation and characterization of hematopoietic transcription factor complexes by in vivo biotinylation tagging and mass spectrometry.

We have described the application of a simple biotinylation tagging approach for the direct purification of tagged transcription factor complexes, based on the use of artificial short peptide tags that are specifically and efficiently biotinylated by the bacterial BirA biotin ligase, which is co-expressed in cells with the tagged factor. We used this approach to initially characterize complexes formed by the hematopoietic transcription factor GATA-1 in erythroid cells. GATA-1 is essential for the erythroid differentiation, its functions encompassing upregulation of erythroid genes, repression of alternative transcription programs, and suppression of cell proliferation. However, it was not clear how all of these GATA-1 functions are mediated. Our work describes, for the first time, distinct GATA-1 interactions with the essential hematopoietic factor Gfi-1b, the repressive MeCP1 complex, and the chromatin remodeling ACF/WCRF complex, in addition to the known GATA-1/FOG-1 and GATA-1/TAL-1 complexes. We also provide evidence that distinct GATA-1 complexes are associated with specific GATA-1 functions in erythroid differentiation, for example, GATA-1/Gfi-1b with the suppression of cell proliferation and GATA-1/FOG-1/MeCP1 with the repression of other hematopoietic transcription programs. We next applied the biotinylation tag to Ldb-1, a known partner of GATA-1, and characterized a number of novel interaction partners that are essential in erythroid development, in particular, Eto-2, Lmo4, and CdK9. Last, we are in the process of applying the same technology to characterize the factors that are bound to the suppressed gamma-globin promoter in vivo.

Animals↗
PubMed2004

Multiple interactions between regulatory regions are required to stabilize an active chromatin hub.

The human beta-globin locus control region (LCR) is required for the maintenance of an open chromatin configuration of the locus. It interacts with the genes and the hypersensitive regions flanking the locus to form an active chromatin hub (ACH) transcribing the genes. Proper developmental control of globin genes is largely determined by gene proximal regulatory sequences. Here, we provide the first functional evidence of the role of the most active sites of the LCR and the promoter of the beta-globin gene in the maintenance of the ACH. When the human beta-globin gene promoter is deleted in the context of a full LCR, the ACH is maintained with the beta-globin gene remaining in proximity. Additional deletion of hypersensitive site HS3 or HS2 of the LCR shows that HS3, but not HS2, in combination with the beta-globin promoter is crucial for the maintenance of the ACH at the definitive stage. We conclude that multiple interactions between the LCR and the beta-globin gene are required to maintain the appropriate spatial configuration in vivo.

Animals↗
PubMed2004

Improvements in the HbVar database of human hemoglobin variants and thalassemia mutations for population and sequence variation studies.

HbVar (http://globin.cse.psu.edu/globin/hbvar/) is a relational database developed by a multi-center academic effort to provide up-to-date and high quality information on the genomic sequence changes leading to hemoglobin variants and all types of thalassemia and hemoglobinopathies. Extensive information is recorded for each variant and mutation, including sequence alterations, biochemical and hematological effects, associated pathology, ethnic occurrence and references. In addition to the regular updates to entries, we report two significant advances: (i) The frequencies for a large number of mutations causing beta-thalassemia in at-risk populations have been extracted from the published literature and made available for the user to query upon. (ii) HbVar has been linked with the GALA (Genome Alignment and Annotation database, available at http://globin.cse.psu.edu/gala/) so that users can combine information on hemoglobin variants and thalassemia mutations with a wide spectrum of genomic data. It also expands the capacity to view and analyze the data, using tools within GALA and the University of California at Santa Cruz (UCSC) Genome Browser.

Databases, Genetic↗
PubMed2004

Molecular characterization and diagnosis of Hb Crete [beta129(H7)Ala-->Pro].

We report the molecular characterization of Hb Crete [beta129(H7)Ala-->Pro] in a female subject from the Greek island of Crete. DNA sequence analysis revealed a 1368 GCC-->CCC base substitution in exon 3 of the beta-globin gene, leading to the Ala-->Pro amino acid change at codon 129. Both the proband and her mother, who were found to be heterozygotes for Hb Crete, presented with mild microcytic anemia and normal Hb A2 levels and iron metabolism indices. This is the first description of an heterozygous Hb Crete case, and also the first report on the molecular basis of Hb Crete. Moreover, the proposed NlaVI restriction enzyme-based detection of Hb Crete at the DNA level is a fast and accurate approach, useful for molecular diagnostics.

Adult↗
PubMed2003

High frequency of concomitant nm23-H1 and E-cadherin transcriptional inactivation in primary non-inheriting colorectal carcinomas.

This study evaluated the added significance of nm23-H1 to that of E-cadherin in determining metastatic proclivity in primary sporadic colorectal carcinomas (SCRCs). A clinical cohort of 52 SCRCs was examined for the significance of nm23-H1 and E-cadherin mRNA levels and E-cadherin protein expression levels into the progression of colorectal tumor invasion, determined by their relevance compared with conventional biological markers. A more than twofold decreased expression of nm23-H1 mRNA was reported in 28/52 (54%) of the carcinomas and was positively associated with the presence of nodal metastases and Astler-Coller stages B1 and B2 in 29% and 35% of the SCRCs, respectively. Reduced expression of E-cadherin mRNA was reported in 38.5% of the carcinomas and was similarly associated with stages Astler-Coller B1 and B2 in 27% of the SCRCs. Decreased E-cadherin immunohistochemical expression (grades II and III) was observed in 67% of the samples. E-cadherin mRNA and protein expression were significantly related to each other. The nm23-H1 (+)/E-cadherin (+) coexpression profile was observed in 31% and was significantly related to the absence of lymph node metastases in 31% and stages Astler-Coller B1 and B2 in 29% of the carcinomas examined. Furthermore, the nm23-H1 (-)/E-cadherin (+) coexpression profile was coupled to decreased E-cadherin immunohistochemical protein detection (grade II) in 21% of the cases examined. These findings suggest that impairment of nm23-H1 expression is an early event into the progression of colorectal metastasis that precedes E-cadherin transcriptional silencing in the majority of SCRCs examined. Nm23-H1 may therefore play an important role in suppressing the early steps of metastasis in sporadic cases of colorectal carcinomas.

Adult↗
PubMed2003

Glucose-6-phosphate dehydrogenase deficiency does not result from mutations in the promoter region of the G6PD gene.

In this study, we investigated whether glucose-6-phosphate dehydrogenase (G6PD) promoter mutations are responsible for G6PD deficiency. We analysed the G6PD proximal promoter and the 5' untranslated region (UTR) in 65 G6PD-deficient individuals, in which no mutations have been found in the G6PD gene coding sequences, using a nonradioactive polymerase chain reaction/single-strand conformation polymorphism (PCR/SSCP) analysis. We identified no sequence variations in the G6PD core promoter or in the 5' UTR of these G6PD-deficient individuals, which indicates that G6PD deficiency is not associated with promoter mutations in the G6PD locus.

DNA Mutational Analysis↗