Association Between the Polymorphism of Angiotensin-Converting Enzyme Gene and Interleukin-1 Beta Gene and the Response to Erythropoietin Therapy in Dialysis Patients with Anemia.

Introduction: The polymorphism of the angiotensin-converting enzyme (ACE) gene and interleukin-1 beta (IL-1b) gene could be associated with resistance in the treatment of anemia in dialysis patients with recombinant human erythropoietin (rHuEPO). The aim of the study was to evaluate the association between the polymorphism of the ACE and IL-1b genes and the response to rHuEPO therapy in dialysis patients with anemia. Material and methods: The study investigated 69 patients on dialysis with anemia treated with recombinant human erythropoietin for 12 months. Genotyping of ACE and IL-1b polymorphism was done in all study patients at the initiation of the study. The patient's demographic characteristics, dialysis vintage, and laboratory parameters were also evaluated as factors associated with rHuEPO resistance. The erythropoietin resistance index (ERI) was calculated as the weekly rHuEPO dose per kg of body weight, divided by the hemoglobin (Hb) concentration in g/dl. Results: The Hb ≥ 110 g/l was registered in 37 (53.6%) patients. Patients with Hb ≥ 110 g/l were characterized by significantly higher serum levels of albumin, cholesterol, and iron than those with Hb < 110 g/l. The serum level of the CRP, the weekly dose of rHuEPO, and ERI were significantly higher in patients with Hb < 110 g/l compared to patients with Hb ≥ 110 g/l. The ERI value of ≥ 10 IUkg/weekly/g/dl was present in 27 (39.1%) patients. The serum levels of ferritin and CRP, and weekly dose of rHuEPO were significantly higher in patients with ERI value ≥ 10 IU kg/weekly/g/dl compared with the patients with ERI value < 10 IUkg/weekly/g/dl. There was no significant association between the ERI and polymorphism of the ACE and IL-1b genes in study patients. Conclusion: The polymorphism of the ACE and IL-1b genes was not significantly associated with the response to erythropoietin therapy in dialysis patients with anemia. Iron deficiency, malnutrition, and inflammation were factors associated with anemia and resistance to erythropoietin therapy in dialysis patients.

Co-Existence of CYP2C19*1/*2 and ABCB1c.3435 CT Genotype has a Potential Impact on Clinical Outcome in CAD Patients Treated with Clopidogrel.

Clopidogrel, is a standard treatment in the prevention of major adverse cardiovascular events (MACE) in patients with coronary artery disease (CAD). Clopidogrel response is highly variable, mainly due to the presence of polymorphisms in the genes involved in drug metabolism. The aim of this study was to evaluate the association between the presence of the ABCB1 C3435T and CYP2C19*2 polymorphism and the clinical outcome in patients with CAD treated with clopidogrel. A total of 96 patients with CAD were included in the study. Genomic DNA from peripheral blood was extracted from all patients with standard phenol/chloroform protocol. The genotyping was performed by Real-Time PCR using TagMan assays. The frequency of the reduced-function allele, in both genes, was higher in patients with negative outcome (36.36% vs 21.15%). A negative clinical outcome and an increased risk for MACE was observed in patients with concomitant inheritance of the CYP2C19 *1/*2 and ABCB1 CT genotype vs patients with other genotypes (22.73% vs 9.62%; OR 3.455; 95% CI= [0.936-12.743], p=0.05722. A trend towards higher risk of MACE was also noted in carriers of the CYP2C19*1/*1 and ABCB1 CC/CT genotype. Our results support the data on the association of the CYP2C19 *2 alone, or in combination with the ABCB1 C polymorphism with the increased risk of MACE. The results also indicate that the presence of ABCB1 C343T polymorphism might be potentially considered as independent predictor of MACE in patients on clopidogrel. However, these results are preliminary and should be confirmed on a larger number of patients.

Molecular Basis of Inherited Colorectal Carcinomas in the Macedonian Population: An Update.

Hereditary factors are assumed to play a role in ~35.0-45.0% of all colorectal cancers (CRCs) with about 5.0-10.0% associated with high penetrant disease-causing mutations in genes correlated to hereditary polyposis (HP) or hereditary non polyposis syndromes (HNPCC). Although inherited germline mutations in mismatch repair (MMR) and the APC genes contribute significantly to CRC, genetic diagnosis cannot yet be obtained in more than 50.0% of familial cases. We present updated data of 107 probands from the Macedonian population with clinically diagnosed HP (n = 41) or HNPCC (n = 66) obtained by next generation sequencing (NGS) with three different gene panels covering the coding, flanking and promoter regions of 114 cancer predisposition genes. Using this approach, we were able to detect deleterious mutations in 65/107 (60.7%) patients, 50.4% of which were in known well-established CRC susceptibility genes and 10.2% in DNA repair genes (DRG). As expected, the highest frequencies of deleterious variants were detected in familial adenomatous polyposis (FAP) and in HNPCC patients with microsatellite instability (MSI) tumors (93.8 and 87.1%, respectively). Variants of unknown significance (VUS) were detected in 24/107 (22.4%) patients, mainly in HNPCC patients with microsatellite stable (MSS) tumors or patients with oligopolyposis. The majority of VUS were also found in DRG genes, indicating the potential role of a doble-strand brake DNA repair pathway deficiency in colorectal cancerogenesis. We could not detect any variant in 18/107 (16.8%) patients, which supports the genetic heterogeneity of hereditary CRC, particularly in HNPCC families with MSS tumors and in families with oligopolyposis.

Evaluation of the JAK2V617F Mutational Burden in Patients with Philadelphia Chromosome Negative Myeloproliferative Neoplasms: A Single-center Experience.

The identification of the JAK2V617F mutation in several distinct myeloproliferative neoplasms (MPNs) raised the question how one single mutation incites expression of at least three different clinical phenotypes, i.e., polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF). In order to further evaluate already published data on the correlation between mutant JAK2V617F allele burden and specific hematological and clinical parameters, we tested the level of the JAK2 mutation in 134 JAK2+ patients with different MPNs. The patients were diagnosed according to the 2008 WHO criteria and followed for a median of 48 months. The JAK2 V617F quantification was done with a real time polymerase chain reaction (real time-PCR) method. The median allele burden was lowest in ET (25.8%), followed by 34.6% in PV and 51.8% in PMF patients (p<0.01). There was statistically significant association between the mutational load of 10.0-50.0% and blood count parameters in the PV patients (p<0.05). In PMF patients the mutational load was in correlation with older age and leukocyte count that were higher in patients with the mutational load of 10.0-50.0% and >50.0% compared to those with a mutational load of <10.0%. There were no statistically significant associations between the allele burden and blood counts in the ET cohort. Our study confirmed an association between the JAK2V617F allele burden and the distinct MPN phenotypes, indicating unfavorable prognosis in patients with a higher JAK2 allele burden. Our results suggest that JAK2 quantification should be incorporated in the diagnostic work-up of MPN patients as a useful tool for optimal treatment decision.

Influence of SLCO1B1 polymorphisms on atorvastatin efficacy and safety in Macedonian subjects.

Atorvastatin, as 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, is a widely prescribed medication for the treatment of dyslipidemia. However, despite its clinical efficacy in reducing major cardiovascular events, a wide inter-individual variability in its response exists. Several studies in this area point to the effect of polymorphisms in the solute carrier organic anion transporter 1B1 (SLCO1B1) gene encoding the multiple organic anion-transporting polypeptide 1B1 (OATP1B1) involved in hepatic uptake of atorvastatin. Hence, the aim of this study was to analyze the association between the SLCO1B1 c.388A>G, c.521T>C, c.571T>C, c.597C>T, c.1086C>T, c.1463G>C and c.*439T>G polymorphisms and lipid-lowering effect and safety of atorvastatin. A hundred and fifty six patients with hyperlipidemia IIa and IIb, all of Macedonian origin, were included in the study receiving atorvastatin 20 - 80 mg/day for 3 months. SLCO1B1 single nucleotide polymorphisms (SNPs) were genotyped using the TaqMan allelic discrimination assay. As parameters of atorvastatin response, total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglycerides (TG), apolipoprotein A (ApoAI), apolipoprotein B (ApoB), lipoprotein(a) (Lp(a)), creatine phosphokinase (CPK), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were measured, using standard laboratory methods, at baseline and after 3 months of treatment. No statistically significant association between the different SLCO1B1 SNPs and atorvastatin response was observed. However, the carriers of c.521CC manifested a lower decrease in plasma levels of TG, TC, LDL-C and Lp(a), with percentage difference being 16%, 7%, 29% and 149%, respectively, compared to the carriers of c.521TT variant. Lower increase in HDL-C (271%) and ApoAI (293%) and higher increase in CPK (69%) in c.521CC carriers were also observed, confirming the lower OATP1B1 activity in carriers of the variant c.521 C allele. Similar results were obtained when a comparison between the percentage of biochemical parameter change was made between *15/*16/*17 heterozygotes and *15/*16/*17 non-carriers. The lack of a statistically significant association between the SLCO1B1 polymorphism and atorvastatin response can be explained dominantly by the low number of individuals homozygous for the rare c.521C variant allele. Despite this limitation, the study offers valuable information on the influence of the genetic determinant SLCO1B1 on atorvastatin response in the Macedonian population.

T-lymphoblastic leukemia/lymphoma in macedonian patients with Nijmegen breakage syndrome.

Nijmegen breakage syndrome (NBS) is a rare autosomal recessive chromosomal instability disorder characterized by microcephaly, immunodeficiency, radiosensitivity and a very high predisposition to malignancy. The gene responsible for the disease, NBS1, is located on chromosome 8q21 and encodes a protein called nibrin. After identification of the gene, a truncating 5 bp deletion, 657-661delACAAA, was identified as the disease-causing mutation in patients with the NBS. In this report, we describe two patients with NBS and T-lymphoblastic leukemia/lymphoma in a Macedonian family. To the best of our knowledge, this is the first family with NBS reported from Macedonia. Both children presented with microcephaly, syndactyly and the development of T cell lymphoblastic lekemia/lymphoma at the age of 7 and 10 years, respectively. The molecular analysis of NBS1 genes in our patients showed homozygosity for the 657del5 mutation in the NBS1 gene. The parents were heterozygotes for the 657del5 mutation and they had no knowledge of a consanguineous relationship. The first child was treated with the International Berlin-Frankfurt-Münster (BFM)-Non Hodgkin lymphoma (NHL) protocol and achieved a complete remission that lasted for 21 months. Subsequently, he developed a medullar relapse with hyperleukocytosis and died due to lethal central nervous system (CNS) complications. The second child was treated according to the International Collaborative Treatment Protocol for Children and Adolescents with Acute Lymphoblastic Leukemia 2009 (AIOP-BFM ALL 2009) protocol. Unfortunately, remission was not achieved.

Effects of single nucleotide polymorphisms and haplotypes of the SLCO1B1 gene on the pharmacokinetic profile of atorvastatin in healthy Macedonian volunteers.

OATP1B1 is an influx transporter known to mediate the uptake of various endogenous compounds and xenobiotics. Several sequence variations have been discovered in the SLCO1B1 gene encoding OATP1B1. The aim of this study was to investigate the effects of SLCO1B1 polymorphisms on the pharmacokinetics of atorvastatin in healthy volunteers of Macedonian origin. Twenty three participants, genotyped for SLCO1B1 c.388A > G, c.521T > C, c.571T > C, c.597C > T, c.1086C > T, c.1463G > C and c.*439T > G polymorphisms using TaqMan allelic discrimination assay, ingested a single 80 mg dose of atorvastatin. The plasma concentrations of atorvastatin were measured for 48 h using Tandem Liquid Chromatography-Mass Spectrometry, LC-MS-MS, and the peak plasma concentration (C(max)), time to peak plasma concentration (T(max)), elimination half-life (t1/2), constant rate of elimination (k(el)), mean residence time (MRT, expo), volume of distribution (Vd/kg), clearance (CL/kg), area under curve AUC(0.48h) and AUC(0-∞), were determined. Our data confirmed that the SLCO1B1 gene is highly polymorphic, with a frequency of the c.521T > C single-nucleotide polymorphism (SNP) being the lowest (app. 15%) and of all other SNPs alleles above 40%. Exceptions were c.1463G > C and c.1086C > T SNPs for which variant alleles were not identified. The strongest correlation was observed between the c.521T > C and c.571T > C SNPs pair. The haplotype analysis revealed 10 different haplotypes, with *1J/*1K/*1L being the dominant, with a frequency of app. 40%. The haplotype *15/*16/*17, containing both variant alleles of the functionally most distinguished SNPs, c.388A > G and c.521T > C, occurred with a frequency of 13%. However, *15/*16/*17 homozygotes were not identified in the study group. In this study, no significant differences in the k(el), t1/2, C(max), T(max), AUC(0-48h), AUC(0-∞), MRT expo, Vd and CL between the carriers of different c.388A > G, c.597C > T and c.*439T > G genotypes were observed. Subject with a variant allele C in the c.521T > C SNP, c.521CC genotype, had markedly higher values for C(max) and AUC(0.48h), 140% and 67%, respectively, in comparison with the carriers of the c.521TT genotype. Also, the carriers of the variant allele C at c.571T > C SNP, c.571 CC genotype, had 55% and 43% lower mean C(max) and AUC(0-48h) in comparison with the carrier of c.571TT. These differences lacked statistical significance due to the size of the sample. In addition, no significant differences in the pharmacokinetic parameters of atorvastatin between the *15/*16/*17 heterozygotes and *15/*16/*17 non-carriers were observed. In conclusion, this extensive analysis of the effect of SLCO1B1 polymorphisms on the pharmacokinetic profile of atorvastatin showed that c.521T > C and c.571T > C SNPs may affect the inter-individual response to atorvastatin. Additional studies, with a large sample size, are needed to confirm this finding.

Pharmacogenetics and antipsychotic treatment response.

Antipsychotic drugs are widely used in the treatment of schizophrenia and psychotic disorder. The lack of antipsychotic response and treatment-induced side-effects, such as neuroleptic syndrome, polydipsia, metabolic syndrome, weight gain, extrapyramidal symptoms, tardive dyskinesia or prolactin increase, are the two main reasons for non-compliance and increased morbidity in schizophrenic patients. During the past decades intensive research has been done in order to determine the influence of genetic variations on antipsychotics dosage, treatment efficacy and safety. The present work reviews the molecular basis of treatment response of schizophrenia. It highlights the most important findings about the impact of functional polymorphisms in genes coding the CYP450 metabolizing enzymes, ABCB1 transporter gene, dopaminergic and serotonergic drug targets (DRD2, DRD3, DRD4, 5-HT1, 5HT-2A, 5HT-2C, 5HT6) as well as genes responsible for metabolism of neurotransmitters and G signalling pathways (5-HTTLPR, BDNF, COMT, RGS4) and points their role as potential biomarkers in everyday clinical practice. Pharmacogenetic testing has predictive power in the selection of antipsychotic drugs and doses tailored according to the patient's genetic profile. In this perception pharmacogenetics could help in the improvement of treatment response by using different medicinal approaches that would avoid potential adverse effects, reduce stabilization time and will advance the prognosis of schizophrenic patients.

Frequencies of single-nucleotide polymorphisms and haplotypes of the SLCO1B1 gene in selected populations of the western balkans.

As a membrane influx transporter, organic anion-transporting polypeptide 1B1 (OATP1B1) regulates the cellular uptake of a number of endogenous compounds and drugs. The aim of this study was to characterize the diversity of the solute carrier organic anion transporter family member 1B1 (SLCO1B1) gene encoding this transporter in two ethnic groups populating the Western Balkans. The distribution of SCLO1B1 alleles was determined at seven variant sites (c.388A>G, c.521T>C, c.571T>C, c.597C>T, c.1086C>T, c.1463G>C and c.*439T>G) in 266 Macedonians and 94 Albanians using the TaqMan allelic discrimination assay. No significant difference in the frequencies of the single nucleotide polymorphisms (SNPs) was observed between these populations. The frequency of the c.521T>C SNP was the lowest (<13.7 and 12.2%, respectively), while the frequencies of all other SNP alleles were above 40.0%. Variant alleles of c.1463G>C and c.1086 C>T SNPs were not identified in either ethnic group. The haplotype analysis revealed 20 and 21 different haplotypes in the Macedonian and Albanian population, respectively. The most common haplotype in both ethnic groups, *1J/*1K/*1L, had a frequency of 39.0% and 26.6%, respectively. In both populations, the variant alleles of the functionally significant c.521T>C and c.388A>G SNPs existed in one major haplotype (*15/*16/*17), with a frequency of 8.6 and 2.4% in the Macedonian and Albanian subjects, respectively. In conclusion, sequence variations of the SLCO1B1 gene in the studied populations occur at high frequencies, which are similar to that of the Caucasian population. Further studies are needed to evaluate the clinical significance of these SNPs and/ or the major SLCO1B1 haplotypes they form for a large number of substrates and for susceptibility to certain diseases.

Rhabdomyolysis and Cardiomyopathy in a 20-Year-Old Patient with CPT II Deficiency.

Aim. To raise the awareness of adult-onset carnitite palmitoyltransferase II deficiency (CPT II) by describing clinical, biochemical, and genetic features of the disease occurring in early adulthood. Method. Review of the case characteristics and literature review. Results. We report on a 20-year-old man presenting with dyspnea, fatigue, fever, and myoglobinuria. This was the second episode with such symptoms (the previous one being three years earlier). The symptoms occurred after intense physical work, followed by a viral infection resulting in fever treated with NSAIDs. Massive rhabdomyolysis was diagnosed, resulting in acute renal failure necessitating plasmapheresis and hemodialysis, acute hepatic lesion, and respiratory insufficiency. Additionally, our patient had cardiomyopathy with volume overload. After a detailed workup, CPT II deficiency was suspected. We did a sequencing analysis for exons 1, 3, and 4 of the CPT II gene and found that the patient was homozygote for Ser 113 Leu mutation in exon 3 of the CPT II gene. The patient recovery was complete except for the cardiomiopathy with mildly impaired systolic function. Conclusion. Whenever a patient suffers recurrent episodes of myalgia, followed by myoglobinuria due to rhabdomyolysis, we should always consider the possibility of this rare condition. The definitive diagnose of this condition is achieved by genetic testing.

Distribution of the most Common Genetic Variants Associated with a Variable Drug Response in the Population of the Republic of Macedonia.

Genetic variation in the regulation, expression and activity of genes coding for Phase I, Phase II drug metabolizing enzymes (DMEs) and drug targets, can be defining factors for the variability in both the effectiveness and occurrence of drug therapy side effects. Information regarding the geographic structure and multi-ethnic distribution of clinically relevant genetic variations is becoming increasingly useful for improving drug therapy and explaining inter-individual and inter-ethnic differences in drug response. This study summarizes our current knowledge about the frequency distribution of the most common allelic variants in three broad gene categories: the Phase I oxidation-cytochrome P450 (CYP450) family (CYP2C9, CYP2C19, CYP3A5, CYP2D6); the Phase II conjugation (GSTT1, SULT1A1; UGT1A1) and drug target (TYMS-TSER, MTHFR and VKORC1) in the population of the Republic of Macedonia and compares the information obtained with data published for other indigenous European populations. Our findings define the population of the Republic of Macedonia as an ethnic group with a highly polymorphic genetic profile. These results add to the evidence regarding the distribution of clinically important variant alleles in DME and drug target genes in populations of European ancestry.

Development and validation of automated SPE-LC-MS/MS method for determination of indapamide in human whole blood and its application to real study samples.

A fast and simple liquid chromatography-electrospray ionization tandem mass spectrometry method for determination of indapamide in human whole blood was developed and validated. The sample extraction of indapamide from human whole blood was achieved using automated solid-phase extraction. Chromatographic separation was performed on Kinetex C18 column (100 × 2.1 mm, 1.7 µm particle size) using acetonitrile and 2 mm ammonium formate in ratio 90:10 (v/v) as a mobile phase. The mass spectrometer was operated in the multiple reaction monitoring mode using positive electrospray ionization for indapamide and the internal standard (zolpidem tartarate). The total run time was 2.5 min. The present method was found to be linear in the concentration range of 1-50 ng/mL with the coefficient of determination 0.9987. The absolute recoveries of indapamide were 90.51-93.90%. The method was validated according the recommendations for validation of bioanalytical methods of European Medicines Agency guideline and was successfully used to analyze human whole blood samples for application in a pharmacokinetic study.

Characterization of the most common CYP2C9 and CYP2C19 allelic variants in the population from the Republic of Macedonia.

The aim of this study was to evaluate the most common CYP2C9 and CYP2C19 polymorphisms in the population of Macedonia and compare them with the global geographic data reported from different ethnic populations. In total, 184 healthy volunteers from the general population were included. Genotypes for the CYP2C9 (*2 [rs1799853] and *3[rs1057910]) and CYP2C19 (*2-[rs4244285] and *17 [rs12248560]) polymorphisms were detected by Real-Time PCR using TaqMan SNP genotyping assay. The CYP2C9 wildtype allele (*1) was the most frequent (78.8%) and the non-functional alleles *2 and *3 had a frequency of 13.9% and 7, 3%, respectively. Seven subjects (2.97%) were poor metabolites (PMs) for CYP2C9 because of the *2/*2 and *3/*3 genotype. For CYP2C19, the frequencies of the*1 (wild-type) and the non-functional alleles (*2 and *17) were 65.4%, 14.4% and 20.1%, respectively. The *2/*2 genotype, corresponded to the predicted frequency of 2.7% for the CYP2C19 PM phenotype. The total of 59 out of 184 subjects (32.0%) was determined as UMs because of the *1/*17 and *17/*17 genotypes. The compound heterozygote (*2/*17), which is associated with a difficult-to-predict phenotype, was detected in 8 subjects (4.34%). The CYP2C9 and CYP2C19 are polymorphic in the population of the Republic of Macedonia. The frequencies of the most common CYP2C9 and CYP2C19 allelic variants are similar to those reported for Caucasians of European descendant, but differ from those of North America Caucasians. Our results suggest that the genetically determined capacity of CYP2C9 and CYP2C19 has to be taken into account in order to improve the individual risk / benefit ratio of the drug therapy in Macedonia.

Effectiveness of erlotinib as a second line therapy for patients with epidermal growth factor receptor (EGFR) mutation in non-small cell lung cancer (NSCLC): our clinical experience.

PURPOSE: The aim of this study was to evaluate tumour response, QoL and adverse effects of erlotinib, EGFR tyrosine kinase inhibitor (TKI), as a second line therapy for patients with EGFR mutation in NSCLC, after failure of previous first-line therapy. METHODS: During the year 2010-11, 5 patients were enrolled in this study for testing EGFR mutations, after conditions for testing were created in Macedonia. We screened 5 patients for EGFR mutations by direct sequencing of exons 18 to 21, by retrospective analysis of their previous biopsy samples. Three of the patients were men and two of the patients were women. Two of the males were former smokers and one male and both females had never smoked. All the patients who were enrolled in the study had histologically proven adenocarcionoma. the patients started with erlotinib 150 mg, one tablet per day, after failure on previous first-line platinum-based chemotherapy, with or without surgery and radiotherapy. Assessment of tumour response was according to RECIST criteria at the follow-up visits every 4 weeks. We analysed tumour response from the beginning with erlotinib until tumuor progression or detection of severe toxicity. Assessment was performed only for those patients with EGFR mutations. Assessment of QoL was performed by patients' subjective answers, as subjective improvement and without subjective improvements. Adverse effects were applied according to WHO criteria. RESULTS: Tissue was available for all 5 cases, two (40%) of which were found to harbour an EGFR mutation, identified as exon 19 deletions. Two patients responded to therapy. A complete response was seen in a female patient for 37 months. Progressive disease was the reason for stopping erlotinib after 37 months and starting third-line therapy. A partial response in one male patient was assessed for 30 months and is still in follow up. This patient is still alive and in good condition. The two patients reported subjective improvements during treatment with erlotinib. Skin rash was grade 2-3, and diarrhoea was grade 1-2. Both patients complained of hair loss, but without complete alopecia. CONCLUSIONS: Considering our clinical results, we recommend target therapy with erlotinib for patients with NSCLC and EGFR mutations as a second-line treatment. Our excellent results encouraged us to require prospective tissue procurement for all patients in Macedonia. This may in fact require a shift in diagnostic practice, from the current emphasis on fine-needle aspiration, which often provides insufficient material for molecular analysis, to obtaining more substantial biopsies and to provide this treatment as first-line for selected patients.

Association of GPX1 polymorphism, GPX activity and prostate cancer risk.

Prostate cancer is the second most common cancer in men worldwide. Although the aetiology of this disease remains largely unclear, several lines of evidence suggest that oxidative stress plays a role in prostate carcinogenesis. The antioxidant enzyme glutathione peroxidase 1 (GPX1) is part of the enzymatic antioxidant defence, preventing oxidative damage to DNA, proteins and lipids by detoxifying hydrogen and lipid peroxides that may contribute to prostate cancer development. Some studies indicate an association between GPX1 Pro198Leu polymorphism and an increased risk of cancer. The purpose of the present study was to determine the possible association of GPX1 Pro198Leu polymorphism and erythrocyte GPX activity with the risk of developing prostate cancer and to clarify whether erythrocyte GPX activity levels were correlated with the GPX1 Pro198Leu genotype in the Turkish population. The GPX1 Pro198Leu genotype was determined in 33 prostate cancer patients and 91 control individuals. As evident from our results, there was no difference between genotype and/or allele frequencies in prostate cancer patients and controls. No significant difference was found in GPX1 genotype or allele frequency between aggressive and non-aggressive prostate cancer patients. It can be suggested with these findings that individual susceptibility of prostate cancer may be modulated by GPX1 polymorphism, but it needs further studies.

Cross-reactive epitopes present in campylobacter jejuni serotypes isolated from enteritis patients.

Campylobacter jejuni (C. jejuni) infection frequently triggers autoimmune-mediated neuropathies, especially the Guillain-Barre syndrome (GBS). The molecular mimicry between the core oligosaccharides of bacterial lipopolysaccharides (LPSs) and the human gangliosides presumably results in the production of anti-neural cross-reactive antibodies which are likely to be a contributory factor in the induction and pathogenesis of GBS. The aim of our study was to determine the presence of cross-reactive epitopes in C. jejuni LPSs isolated from enteritis patients and to determine their antigen reactivity. For that purpose we collected stool specimens from 21 patients with enteritis and without neurological symptoms. Seven different serotypes of C. jejuni (0:27; 0:6/0:7; 0:38; 0:3; 0:1/0:44; 0:19; 0:37) were detected using the Penner system. Unexpectedly, one serotype from this group was detected as 0:19, a serotype rarely isolated from enteritis patient and in close association with GBS. Binding studies using cholera toxin-B subunit and peanut agglutinin, showed the presence of ganglioside-like epitopes in C. jejuni strains 0:37, 0:19 and 0:27. Reactivity with sera from patient with GBS, with confirmed previous exposure to C. jejuni and with high a titre of anti-ganglioside antibodies, showed that the same three LPSs from C. jejuni serotypes 0:37, 0:19 and 0:27 bear cross-reactive epitopes in their LPSs structures. Our results confirm the results from previous studies that LPSs from certain C. jejuni serotypes bear cross-reactive ganglioside-like epitopes which might be involved in the induction of GBS after C. jejuni infection.

JAK2V617F mutations in myeloid malignancies: single center experience.

Recently, V617F mutation in JAK2 tyrosine kinase gene was established as a marker of myeloproliferation, useful for proving clonality and securing diagnosis in a considerable proportion of the myeloproliferative neoplasms (MPN) The discovery presents a major breakthrough in the understanding of the pathogenesis of the MPN. Moreover, some studies suggest a possible role of the JAK2V617F mutation in the pathogenesis of some specific acute myeloid leukemia (AML) subtypes. To further improve the understanding of the role of JAK2V617F mutations in the pathogenesis and the clinical course of the myeloid malignancies we screened 192 patients with various MPN and AML for the mutations and analyzed the possible association between JAK2V617F mutations and the clinical features of MPNs patients. The frequency of V617F JAK2 mutation was analyzed by theallele-specific PCR assay. Out of 153 cases with known or suspected diagnoses of MPNs, 100 (65.3%) were positive for the JAK2V617F mutation and 53 (34.7%) were negative. In 39 AML cases the mutant allele V617F was not expressed. Correlations of the clinical features at diagnosis and long-term prognosis between the two JAK2-V617F different MPNs groups revealed comparability regarding all tested parameters except for the incidence of thrombotic history. Patients with the mutation had significantly higher incidence of thrombotic complication (38.5%), compared to the group without the mutation (19.2%) (P < 0.005). Our results confirmed the diagnostic significance of JAK2V617F mutation in MPNs and supported the notion that patients with the mutation should be classified in a new entity of MPNs.

An Alu insert as the cause of a severe form of hemophilia A.

Alu sequences represent a specific human family of interspersed repetitive DNA, with a copy number in excess of 500,000 within the human genome. Alu repeats are rarely present in protein-coding regions of mature RNA, and only a few Alu insert mutations have been described so far. In this paper we present an Alu retroposition event in a family with a severe form of hemophilia A. The inserted Alu element belonging to the youngest Yb8 subfamily disrupts the reading frame at methionine 1224, exon 14 of the factor VIII gene, leading to a stop codon within the inserted sequence. This observation indicates that the retroposition of Alu elements is a continuing process possibly generating various human genetic defects.

A large beta-thalassemia deletion in a family of Indonesian-Malay descent.

The partial molecular characterization of a large deletion present in two members of an Indonesian-Malay family with beta-thalassemia trait is described. Polymerase chain reaction and sequencing analyses of the breakpoint identified a sequence which has previously been described in patients with the 45 kb Filipino beta 0-thalassemia deletion, i.e. a 5' breakpoint at position -4279 nucleotides 5' from the Cap site of the beta-globin gene. The 3' breakpoint is located in an L1 family of repetitive sequences at an unknown distance from the beta-globin gene. The hematological and hemoglobin data of the patients with this beta 0-thalassemia deletion further supports the concept that the unusually high Hb A2 levels are unique to deletions removing the 5' beta-globin gene region, and points to the importance of the 3' junction sequences for the regulation of Hb F levels in patients with deletional defects of the beta-globin gene cluster.