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Hepatitis C virus (HCV) infection becomes a major public health problem and leading cause of chronic liver disease and liver failure. Pegylated interferon-alfa and ribavirin are currently the standard treatment for chronic hepatitis C (CHC). It is indicated that genes that trace the interferon signaling could be associated with the host response to the therapy. In order to investigate the influence of these genes on host related response, we have analyzed seven single nucleotide polymorphisms (rs59248852, rs74390571, rs12811390, rs1169279, rs3213545, rs1083129 and rs2859398) in 2-5-Oligoadenylate- Synthetase-Like (OASL) gene in CHC cases from Republic of Macedonia. A simple and easy to use SNaPshot method was developed. A cohort of 100 HCV RNA positive patients - non responders and 109 patients with achieved virological response after the standard treatment were included in this study. We have found significant association in five of the seven studied SNP` s: rs59248852 [p = 6.5x10-31, OR=55.7 (20.0-155.3)]; rs12811390 [p = 2.2x10-11, OR=4.3 (2.3-6.7)]; rs2859398 [p=1.34x10-9, OR=3.4 (2.2-5.0)]; rs74390571 [p=4.3x10-7, OR=2.9 (1.9-4.4)], and rs1083129 [p=0.0139, OR=2.0 (1.1-3.5)]. The results from this study can be used as a predictive factor of future patient’s selection for the standard therapy, having an important cost benefit for the health insurance system.

Clinical importance of the most common CHEK2 (IVS2+1 G>A, 1100delС, I157T and del5395) and NBN (R215W and 657del5) gene mutations for breast cancer development in Macedonian breast cancer patients is unknown. We performed a case-control study including 300 Macedonian breast cancer patients and 283 Macedonian healthy controls. Genotyping was done using a fast and highly accurate single-nucleotide primer extension method for the detection of five mutations in a single reaction. The detection of the del5395 was performed using an allele-specific duplex polymerase chain reaction (PCR) assay. We have found that mutations were more frequent in breast cancer patients (n = 13, 4.3%) than in controls (n = 5, 1.8%), although without statistical significance. Twelve patients were heterozygous for one of the analyzed mutations, while one patient had two mutations (NBN R215W and CHEK2 I157T). The most frequent variant was I157T, found in 10 patients and four controls (p = 0.176) and was found to be associated with familial breast cancer (p = 0.041). CHEK2 1100delC and NBN 657del5 were each found in one patient and not in the control group. CHEK2 IVS2+1G>A and del5395 were not found in our cohort. Frequencies of the studied mutations are low and they are not likely to represent alleles of clinical importance in the Macedonian population.

A high-quality hemoglobinopathy diagnosis is based on the results of a number of tests including assays for molecular identification of causative mutations. We describe the current diagnostic strategy for the identification of b-thalassemias and hemoglobin (Hb) variants at the International Reference Laboratory for Haemoglobinopathies, Research Centre for Genetic Engineering and Biotechnology (RCGEB) “Georgi D. Efremov,” Skopje, Republic of Macedonia. Our overall approach and most of the methods we use for detection of mutations are designed for the specific target population. We discuss new technical improvements that have allowed us to substantially reduce the average time necessary for reaching a conclusive diagnosis.

The most significant and well characterized genetic risk factors for breast and/or ovarian cancer are germline mutations in the BRCA1 and BRCA2 genes. The BRCA1 and BRCA2 gene mutations strikingly increase breast cancer risk, suggesting that polymorphisms in these genes are logical candidates in seeking to identify low penetrance susceptibility alleles. The aim of this study was to initiate a screen for BRCA1/2 gene mutations in order to identify the genetic variants in the Republic of Macedonia, and to evaluate the association of several single nucleotide polymorphisms (SNPs) in these genes with breast cancer risk. In this study, we included 100 patients with invasive breast cancer from the Republic of Macedonia, classified according to their family history and 100 controls. The methodology included direct sequencing, single nucleotide primer extension method and multiplex ligation probe amplification (MLPA) analysis, all followed by capillary electrophoresis (CE) on an ABI PRISM™ 3130 Genetic Analyzer. We identified a total of seven carriers of mutations in the BRCA1/2 genes. None of the tested polymorphisms was associated with sporadic breast cancer risk, however, polymorphism rs8176267 in BRCA1 and N372H in BRCA2 showed an association with breast cancer risk in patients with at least one family member with breast cancer.

Hepatitis C virus (HCV) is a major public health problem. It is a leading cause of chronic liver disease and the most common indication for liver transplantation. The therapy for eradication of HCV infection is successful in only 50.0-80.0% of patients and is highly dependent on the HCV genotype. Molecular detection and characterization of HCV in the Republic of Macedonia started in 1990. Since then, more than 4000 samples have been analyzed at the Research Centre for Genetic Engineering and Biotechnology (RCGEB) “Georgi D. Efremov,” Skopje, Republic of Macedonia. The prevalence of HCV infections in the healthy population of the Republic of Macedonia was found to be 0.4%, while it varies between 23.0 and 43.0% in different at-risk groups of patients. The prevalence of HCV genotypes, according to associated risk factors in HCV infected patients from the Republic of Macedonia, was analyzed. We found genotype 1 to be predominant in a group of hemodialysis patients, while genotype 3 was predominant in intravenous (IV) drug users. Association of six polymorphisms in the Oligoadenylate synthetase (OASL)-like interferonstimulated gene with a sustained virological response was also analyzed. Our preliminary results suggest that non ancestral alleles in four of the six studies polymorphisms in OASL gene are associated with sustained virological response among HCV infected patients in R. Macedonia.

MicroRNAs (miRNAs) are small [~21 nucleotide (nt)] non coding RNAs (ncRNAs) that regulate gene expression posttranscriptionally. About 3.0% of human genes encode for miRNAs, and up to 30.0% of human protein coding genes may be regulated by miRNAs. Currently, more than 2000 unique human mature microRNAs are known. MicroRNAs play a key role in diverse biological processes including development, cell proliferation, differentiation and apoptosis. These processes are commonly dysregulated in cancer, implicating miRNAs in carcinogenesis, where they act as tumor supressors or oncogenes. Several miRNAs are associated with breast cancer. Here we present our initial results of miRNA analyses of breast cancer tissues using quantitative real time-polymerase chain reaction (ReTi-PCR) (qPCR) involving stem-loop reverse transcriptase (RT) primers combined with TaqMan® PCR and miRNA microarray analysis