Plasminogen activator inhibitor-1 genotype 4G/5G associates with skin involvement in Armenian familial Mediterranean fever patients.

There is little and conflicting data on the role of the plasminogen activator inhibitor-1 (PAI-1, SERPINE1) 4G/5G polymorphism in familial Mediterranean fever (FMF). Therefore this study aimed at evaluating the impact of this polymorphism on the disease course in a cohort of 303 Armenian FMF patients. Genotyping for 12 Mediterranean fever (MEFV) gene mutations and the PAI-1 4G/5G (rs1799762) polymorphism were performed by PCR/reverse-hybridization (StripAssay) and real-time PCR, respectively. PAI-1 genotypes 4G/4G, 4G/5G, and 5G/5G could be identified in 4 (5.88%), 30 (18.63%) and 9 (12.16%) patients with erysipelas-like erythema (ELE), while this was the case for 64 (94.12%), 131 (81.37%), and 65 (87.84%) patients without ELE, respectively (P < 0.033). We have identified a significant relationship between the PAI-1 4G/5G genotype and the occurence of ELE in a relatively large cohort of Armenian FMF patients. Because of conflicting results concerning the impact of this polymorphism on the clinical course of FMF in different populations, further studies are desirable to substantiate the findings reported here.

A Population-Oriented Genetic Scoring System to Predict Phenotype: A Pathway to Personalized Medicine in Iraqis With ß-Thalassemia.

To assess the roles of genetic modifiers in Iraqi ß-thalassemia patients, and determine whether a genotype-based scoring system could be used to predict phenotype, a total of 224 Iraqi patients with molecularly characterized homozygous or compound heterozygous ß-thalassemia were further investigated for α-thalassemia deletions as well as five polymorphisms namely: rs7482144 C > T at HBG2, rs1427407 G > T and rs10189857 A > G at BCL11A, and rs28384513 A > C and rs9399137 T > C at HMIP. The enrolled patients had a median age of 14 years, with 96 males and 128 females. They included 144 thalassemia major, and 80 thalassemia intermedia patients. Multivariate logistic regression analysis revealed that a model including sex and four of these genetic modifiers, namely: ß+ alleles, HBG2 rs7482144, α-thalassemia deletions, and BCL11A rs1427407 could significantly predict phenotype (major versus intermedia) with an overall accuracy of 83.9%. Furthermore, a log odds genetic score based on these significant predictors had a highly significant area under curve of 0.917 (95% CI 0.882-0.953). This study underscores the notion that genetic scoring systems should be tailored to populations in question, since genetic modifiers (and/or their relative weight) vary between populations. The population-oriented genetic scoring system created by the current study to predict ß-thalassemia phenotype among Iraqis may pave the way to personalized medicine in this patient's group.

Very low frequency of the lactase persistence allele LCT-13910T in the Armenian population.

Primary lactose malabsorption is characterised by a down-regulation of lactase activity after weaning and inability to digest lactose in adulthood. It has been suggested that the historical introduction of dairying led to a positive selection for lactase persistence variants in a regulatory region upstream of the LCT gene. Here, we genotyped 202 Armenian subjects for LCT-13910T, a lactase persistence variant which is widespread in Europeans. The homozygous C/C genotype associated with primary hypolactasia, the heterozygous C/T and the homozygous T/T lactase persistence genotypes were found in 191 (94.6%), 11 (5.4%), and 0 (0.0%) samples, respectively. The frequency for the LCT-13910*T allele was 2.7%. The observed allele frequency of 2.7% for LCT-13910T is even lower than previously reported and supports current phenotypic data about lactose malabsorption in Armenia.

Serum amyloid A1 genotype associates with adult-onset familial Mediterranean fever in patients homozygous for mutation M694V.

OBJECTIVES: FMF shows considerable variability in severity and type of clinical manifestations by geographic region, which are attributed to Mediterranean fever (MEFV) gene allelic heterogeneity, additional genetic modifiers and environmental factors. Considering the severe impact of MEFV mutation M694V on the FMF phenotype, this work aimed at investigating a possible disease modifying role of the serum amyloid A1 (SAA1) genotype in a cohort of 386 Armenian FMF patients homozygous for MEFV mutation M694V. METHODS: A cohort of 386 Armenian patients diagnosed with FMF based on the Tel-Hashomer criteria and carrying two MEFV M694V mutant alleles were included in this study. Fifty-two (13.40%) of these patients experienced their first attack at the age of ≥20 years (i.e. adult-onset FMF). MEFV and SAA1 analyses were performed by a commercial reverse-hybridization assay, and resulting genotypes were matched against the patients' clinicodemographic profiles. RESULTS: Genotypic distribution of SAA1 alleles was significantly different between patients with an age of onset <20 and ≥20 years. SAA1 genotypes α/α, α/ß and ß/ß could be identified in 8 (15.38%), 12 (23.08%) and 32 (61.54%) adult-onset patients while this was the case for 47 (14.07%), 172 (51.50%) and 115 (34.43%) patients with a disease onset <20 years, respectively (P < 0.001). Furthermore, adult-onset disease was associated with a less severe FMF phenotype (P < 0.001). CONCLUSION: We have identified a significant relationship between the SAA1ß/ß genotype and the age of disease onset in M694V homozygous FMF patients.

Association between serum amyloid A1 genotype and age of onset restricts to M694 homozygote familial Mediterranean fever patients in Armenia.

OBJECTIVES: Familial Mediterranean fever (FMF) is an autosomal-recessive, inflammatory disorder characterised by short, recurrent attacks of fever, accompanied by pain in the abdomen, chest, or joints and complications of amyloidosis. Recently, we observed a significant association between the serum amyloid A1 (SAA1) ß/ß genotype and a delayed disease onset in 386 M694V homozygous FMF patients. This follow-up study was conducted to additionally analyse MEFV genotypes other than M694V/M694V for a possible influence of the SAA1 genotype on the age of disease onset. METHODS: A total of 700 Armenian patients diagnosed with FMF based on the Tel-Hashomer criteria and carrying two MEFV mutant alleles were included in this study. Patients were divided into three MEFV genotypic subgroups: M694V homozygotes (M694V/M694V), M694V compound heterozygotes (M694V/Other), and patients with genotypes excluding M694V (Other/Other). MEFV and SAA1 analyses were performed by a commercial reverse-hybridisation assay, and resulting genotypes were matched against the demographic and clinical characteristics of the patients. RESULTS: Within the subgroup of M694/M694 homozygotes, SAA1 genotype ß/ß could be identified in 115 (34.43%) and 32 (61.54%) patients with an age of onset <20 and ≥20 years, respectively(p<0.001). However, no such relationship could be observed for MEFV genotypic subgroups M694V/Other (p=0.465) and Other/Other (p=0.697). CONCLUSIONS: Our data suggest, that the influence of SAA1 genotypic variation on the age of disease onset restricts to FMF patients homozygous for MEFV mutation M694V.

ISSAID/EMQN Best Practice Guidelines for the Genetic Diagnosis of Monogenic Autoinflammatory Diseases in the Next-Generation Sequencing Era.

BACKGROUND: Monogenic autoinflammatory diseases are caused by pathogenic variants in genes that regulate innate immune responses, and are characterized by sterile systemic inflammatory episodes. Since symptoms can overlap within this rapidly expanding disease category, accurate genetic diagnosis is of the utmost importance to initiate early inflammation-targeted treatment and prevent clinically significant or life-threatening complications. Initial recommendations for the genetic diagnosis of autoinflammatory diseases were limited to a gene-by-gene diagnosis strategy based on the Sanger method, and restricted to the 4 prototypic recurrent fevers (MEFV, MVK, TNFRSF1A, and NLRP3 genes). The development of best practices guidelines integrating critical recent discoveries has become essential. METHODS: The preparatory steps included 2 online surveys and pathogenicity annotation of newly recommended genes. The current guidelines were drafted by European Molecular Genetics Quality Network members, then discussed by a panel of experts of the International Society for Systemic Autoinflammatory Diseases during a consensus meeting. RESULTS: In these guidelines, we combine the diagnostic strength of next-generation sequencing and recommendations to 4 more recently identified genes (ADA2, NOD2, PSTPIP1, and TNFAIP3), nonclassical pathogenic genetic alterations, and atypical phenotypes. We present a referral-based decision tree for test scope and method (Sanger versus next-generation sequencing) and recommend on complementary explorations for mosaicism, copy-number variants, and gene dose. A genotype table based on the 5-category variant pathogenicity classification provides the clinical significance of prototypic genotypes per gene and disease. CONCLUSIONS: These guidelines will orient and assist geneticists and health practitioners in providing up-to-date and appropriate diagnosis to their patients.

Clinical and genetic heterogeneity in a large cohort of Armenian patients with late-onset familial Mediterranean fever.

PURPOSE: This work aimed at investigating demographic, clinical, and genetic characteristics of individuals experiencing their first familial Mediterranean fever (FMF) attack at age ≥40 years in a very large cohort of Armenian FMF patients. METHODS: In total, 10,370 Armenian patients diagnosed with FMF based on the Tel Hashomer criteria and carrying at least one MEFV mutant allele were included in this study. RESULTS: A total of 354 (3.40%) patients had late-onset FMF. Of these, 194 (54.80%) were female and 160 (45.20%) were male. The following genotypes were significantly associated with the late-onset variant: M680I/E148Q (P = 0.004), M694V/E148Q (P < 0.001), and V726A/V726A (P< 0.001). Of note, 12/354 (3.40%) patients were found to be homozygous for the M694V mutation. Individuals with late-onset FMF had a milder disease phenotype presenting significantly less frequent fever, skin manifestation, and chest pain compared to individuals with a disease onset before 40 years of age. Abdominal pain was found more often in the late-onset FMF group, whereas arthritis, proteinuria, and amyloidosis did not differ significantly between the two groups. CONCLUSION: Our data suggest that late-onset FMF is more prevalent in women and is of greater clinical as well as genetic heterogeneity than previously reported.

SLCO1B1 c.521T>C Genotyping in the Austrian Population Using 2 Commercial Real-Time Polymerase Chain Reaction Assays: An Implementation Study.

Statin-induced myopathy is reported to be significantly associated with the SCLO1B1 c.521T>C polymorphism. To date, SLCO1B1 c.521T>C epidemiologic data for the Austrian population is still lacking. Therefore, this study aimed at assessing the genotype and allele frequencies of the SLCO1B1 c.521T>C variant in Austria and evaluating the clinical performance of 2 commercial real-time polymerase chain reaction (PCR) assays. Genomic DNA isolated from 181 healthy individuals was analyzed for the SLCO1B1 c.521T>C polymorphism in a comparative manner using the SLCO1B1 c.521T>C RealFastTM Assay and the BioPro SLCO1B1 Genotyping real-time PCR Kit. A total of 10 (5.5%) and 44 (24.3%) out of 181 individuals were SLCO1B1 c.521T>C C/C-homo- and -C/T-heterozygotes, the genotypes indicative of high and increased risk of statin-induced myopathy, respectively. The SLCO1B1 c.521C allele frequency rate was 17.7%. In conclusion, the genetic predisposition of elevated statin-induced myopathy risk in the Austrian population is frequent. Both real-time PCR assays under investigation here are reliable and robust SLCO1B1 c.521T>C genotyping tools in clinical routine.

Comparing BRAF mutation status in matched primary and metastatic cutaneous melanomas: implications on optimized targeted therapy.

BACKGROUND: Selective BRAF inhibitors have shown dramatic results with regard to improving outcome in patients with melanoma. Testing the BRAF status in matched primary and metastatic melanomas to optimize individual targeted therapy is not well investigated. METHODS: Extended BRAF testing using PCR for 9 mutations and VE1 immunohistochemistry for BRAF V600E detection on 95 lesions including 40 primary melanomas with their matched metastases (n = 42), recurrences (n = 9) and second primaries (n = 4) was performed. Nine patients had multiple metastases. RESULTS: V600E was the only identified mutation type; 35.4% of primary vs. 18.9% of metastatic melanomas. The overall primary-metastatic BRAF status discordance rate was 32.3% using PCR and 27.5% with immunohistochemistry, and was significantly more frequent in primary lesions with mutant BRAF (67%). Males and patients with metastasis to lymph nodes were less likely to be discordant compared to females and those with metastasis to other sites (p = 0.023). Discordant BRAF mutation status was predicted by multivariate binary logistic regression: the presence of a mutant BRAF in the primary melanoma [OR (95% C.I.) = 23.4 (2.4-229.7)] and female gender [OR = 10.6 (1.08-95)]. Inter-metastases BRAF concordance was 100% (6 comparisons). CONCLUSION: A high discordant rate implies the need for clinical trials addressing the response to targeted therapy in patients with discordant BRAF statuses between their primary and metastatic lesions.

BRAF analysis on a spectrum of melanocytic neoplasms: an epidemiological study across differing UV regions.

BRAF mutation has been linked to the development of melanocytic tumors in homogeneous Caucasian cohorts. The role of solar UV radiation (UVR) in BRAF mutation status is poorly understood. We studied the epidemiology of BRAF mutation across a spectrum of melanocytic neoplasms in populations with differing UVR rates. Extended testing for 9 mutation types was attempted on 600 melanocytic neoplasms including banal nevi (n = 225), dysplastic nevi (n = 113), primary (n = 172), and metastatic melanomas (n = 90). Specimens were collected from 4 countries with increasing UVR rates (in kJ/m/yr): Syria (n = 45; UVR = 93.5), Lebanon (n = 225; UVR = 110), Pakistan (n = 122; UVR = 128), and Saudi Arabia (n = 208; UVR = 139). UVR was estimated from 21-year averages from The National Center for Atmospheric Research database. The overall BRAF mutation rate was 49% (268 of 545) and differed significantly by the geographic location [34% Pakistan, 49% Lebanon, 67% Syria, and 54% Saudi Arabia; P = 0.001], neoplasm type (P < 0.001), and anatomical location (P < 0.001) but not with age (P = 0.07) and gender (P = 1.0). V600E was the predominant mutation type, found in 96.3% of the cases. Incidence of melanoma was significantly greater in BRAF-negative (39%) versus BRAF-positive (17%) groups. For BRAF-positive cases, less severe lesions were systematically more frequent (P < 0.001). Multivariate analysis indicated that BRAF mutation is predicted by neoplasm type, anatomical site, and geographic location. In our Near East cohort, BRAF mutation rates varied by geographic location but not based on UVR. BRAF-positive status was associated with less severe lesions.

Characterizing a cohort of α-thalassemia couples collected during screening for hemoglobinopathies: 14 years of an Iranian experience.

Our study aimed to determine the number of couples with normal hemoglobin (Hb) electrophoresis and low-borderline hematological values, which may come up with a clinically critical status in their offspring. The number of couples at risk for severe α-thalassemia (α-thal) needed to be estimated before recommending genetic counseling and prenatal diagnosis (PND). During the past 14 years, from at least 7000 referrals, 754 couples were investigated for α-thal by direct mutation detection methods followed by reverse strip assay and α-globin gene sequencing for inconclusive cases. Detection of silent ß-thalassemia (ß-thal) mutations was done in suspected cases by complete ß-globin gene sequencing. We were able to provide a molecular diagnosis in 87.3% (658/754) of couples. A total of 9.1% (60/658) may have a clinically significant hemoglobinopathy in their offspring. Significant conditions included hydrops fetalis (20.0%; 12/60), certain Hb H (ß4) genotypes (78.3%; 47/60) and ß-thal intermedia (ß-TI) (1.7%; 1/60). The diagnostic flowchart for couples with microcytic hypochromic anemia in countries with a high prevalence of hemoglobinopathies should include α and ß gene sequencing. As our results indicate, every nine out of 100 of these couples will face significant hemoglobinopathies and every two out of 100 can carry Hb Bart's (γ4) hydrops fetalis in their pregnancies. For such cases, PND should be utilized to allow the carrier couples to decide whether or not to abort the fetus.

Allelic frequencies of polymorphism c.521T>C (rs4149056) favor preemptive SLCO1B1 genotyping in Armenia.

OBJECTIVES: Statins represent an important pharmacological factor for the prevention of cardiovascular diseases but may also cause severe cases of myotoxicity. Numerous studies have described the association of the SLCO1B1 gene variant c.521C with statin-induced myopathy across different populations. This study aimed at evaluating the usefulness of preemptive SLCO1B1 genotyping in Armenia. METHODS: A total of 202 Armenian patients referred to the Center of Medical Genetics and Primary Health Care in Yerevan for upper respiratory tract infection between January and May 2022 were included in this study. Genotyping for SLCO1B1 c.521T>C (rs4149056) was performed using a commercially available real-time PCR assay (RealFast™). RESULTS: In total, 3/202 (1.5 %) samples were C/C homozygotes and 52/202 (25.7 %) were T/C heterozygotes, associated with a high and increased risk for statin-induced myopathy, respectively. The SLCO1B1 c.521C allelic frequency was 14.4 %. CONCLUSIONS: The observed allele frequency of 14.4 % for the c.521C variant is slightly lower than frequencies reported from Europe, but relatively high compared to Asian populations, suggesting that preemptive SLCO1B1 genotyping could be a useful approach for the reduction of statin-induced adverse effects in Armenia.

Predictors of BRAF mutation in melanocytic nevi: analysis across regions with different UV radiation exposure.

BACKGROUND: BRAF mutations have been implicated in initiating promutagenic cellular melanocytic proliferation mostly based on homogeneous Western-based cohorts. Data addressing the possible interaction between exposure to different solar ultraviolet radiation (UVR) magnitudes and BRAF mutation rate (BMR) in melanocytic nevi are limited. DESIGN: Extended BRAF testing for 9 mutations in 225 melanocytic nevus (MN) cases derived from 211 patients from 4 different UVR regions: Lebanon (n = 95; 110 kJ · m(-2) · yr), Syria (n = 23; 93.5 kJ · m(-2) · yr), Kingdom of Saudi Arabia (n = 70; 139 kJ · m(-2) · yr), and Pakistan (n = 37; 118 kJ · m(-2) · yr) was performed. Data collected included age, gender, anatomic location, and lesion size. Histological parameters recorded were MN type (junctional, compound, intradermal, classical blue, cellular blue, compound and intradermal spitz, and congenital) solar elastosis grade, and nevus pigmentation degree. Cumulative 21-year erythemally effective UV averages were derived from The National Center for Atmospheric Research. RESULTS: BRAF mutation status was obtained in 210 cases (6.7% failed polymerase chain reaction). Overall, BMR was 62.4% (131/210) with V600E mutation accounting for 98.5% of cases. Discordant mutation status was found in 2 of 10 patients with multiple nevi. BMR differed significantly, yet nonsystematically, among UVR regions; the highest was detected in nevi coming from Syria (18/23 cases, 78%), followed by Pakistan (21/30 cases, 70%), Kingdom of Saudi Arabia (47/70 cases, 67%), and Lebanon (45/87 cases, 52%). Mutation rates varied significantly across MN type (P < 0.001); the highest rate was recorded in the intradermal nevus type (33/39 cases, 84.6%), followed by the compound (26/32 cases, 81.2%) and congenital (60/74 cases 81.0%) nevi. Stratified by anatomic location, nevi occurring on the face (61/82, 74%) and trunk (58/78, 74%) had more frequent BMRs compared with those occurring on the upper (7/26, 27%) and lower extremities (5/24, 21%, P < 0.001). Severe pigmentation was less frequent in BRAF mutation-positive nevi [5/131 (4%) vs. 34/79 (43%); P < 0.001]. Multivariate independent predictors of BRAF mutation in MN were age [odds ratio (95% confidence interval ) = 1.43 (1.13-1.74) per 10 years; P = 0.004], anatomic location [P = 0.043 overall], and nevus type [P < 0.001 overall]. UVR region was not an independent predictor of BRAF mutation. CONCLUSIONS: Increased BRAF mutation with age along with the lack of a UVR magnitude-BRAF mutation association suggests that duration of exposure rather than UVR exposure dose is the more likely link to acquiring the mutation.

Risk for early pregnancy loss by factor XIII Val34Leu: the impact of fibrinogen concentration.

BACKGROUND: We have already described a significantly elevated overall risk for recurrent pregnancy loss (RPL) in women carrying the coagulation factor XIII (FXIII) Val34Leu and/or the plasminogen activator inhibitor-1 (PAI-1) 4G/5G polymorphism assuming that these polymorphisms contribute synergistically to RPL because of impaired hypofibrinolysis. Recent studies on FXIII indicate that the impact of the FXIII 34Leu genotype on fibrin structure and fibrinolysis is affected by fibrinogen concentration. Therefore, we reinvestigated the association between fibrinogen concentrations and FXIII Val34Leu with early RPL. MATERIALS AND METHODS: In this case-control study, we enrolled 49 women with a history of two consecutive or three to six nonconsecutive pregnancy losses between the 8th and 12th week of gestation and 48 healthy controls. The risk for RPL in carriers of FXIII 34Leu at fibrinogen levels above or below the median and first tertile of controls was evaluated. RESULTS: In carriers of the 34Leu allele, fibrinogen levels below the median (i.e., ≤ 300 mg/dl) and the first tertile (i.e., ≤ 284 mg/dl) of controls were associated with an increased risk for RPL [(2.9 (1.1-7.7), 3.9(1.0-15.0)]. CONCLUSIONS: The FXIII Val34Leu polymorphism may be associated with the development of early RPL in association with fibrinogen concentrations. At fibrinogen levels in the low normal range, FXIII 34Leu may modify fibrin structure toward an increased resistance to fibrinolysis.

Optimisation of individual cardiovascular risk assessment in a German coronary artery disease cohort using a commercial test for genetic polymorphisms - a pilot study.

BACKGROUND AND AIMS: Assessment of cardiovascular risk using established risk scores such as ESC SCORE2 or PROCAM insufficiently emphasise the role of genetic factors. We hypothesise that commercially available genetic assays may provide additional information on hereditary cardiovascular risk in a timely and cost-efficient manner. METHODS: In a cohort of 51 patients treated for coronary artery disease (CAD) at University Hospital Heidelberg, Germany, a subgroup of patients with "unstable" CAD (i.e. recurrent acute coronary syndrome) was identified and compared to patients with "stable" disease (i.e. chronic coronary syndrome). Gene array analysis using a commercial assay for 15 potentially pathogenic polymorphisms revealed our cohort's genetic risk profile regarding atherosclerotic/thromboembolic events. Improvement of cardiovascular risk assessment based on established risk scores was analysed using net reclassification, logistic regression and receiver operating characteristic (ROC) analysis. RESULTS: Discriminatory capacity of traditional risk scores such as SCORE2 or PROCAM with regard to stable and unstable CAD groups was poor (ROC AUC <0.5). Patients with "unstable" CAD exhibited a significantly increased frequency of pathogenic eNOS 894 T and MTHFR 1298 C polymorphisms compared to "stable" CAD patients, and information on these polymorphisms individually as well as combinations with additional polymorphisms included in the assay such as ACE D/D or PAI-1 5 G variants markedly improved risk prediction compared to SCORE2/PROCAM alone (ROC AUC ≥0.75). CONCLUSION: Commercially available assays for genetic polymorphisms may provide valuable information on individual genetic cardiovascular risk, potentially guiding future primary and/or secondary preventative therapies for coronary artery disease.

Analyzing 5'HS3 and 5'HS4 LCR core regions and NF-E2 in Iranian thalassemia intermedia patients with normal or carrier status for beta-globin mutations.

Our data on 114 Iranian individuals with thalassemia intermedia phenotype revealed homozygous or compound heterozygous beta-globin mutations to be the predominant disease factor in 86.2% of cases. However, 8.2% of these individuals were found to be heterozygous or wild type for beta-globin mutations. In search for determinants outside of the beta-globin gene, which could be responsible for the unexpected thalassemia intermedia phenotype in these subjects, we screened the alpha-globin genes, the 5'HS3 and 5'HS4 regions of the beta-globin LCR, and the NF-E2 transcription factor for sequence variations in selected individuals. The -3.7 deletion was the only alpha-globin mutation detected, and no alterations were found in 5'HS3 and NF-E2. Sequence analysis of the 5'HS4 LCR core region identified three known SNPs in a single patient, who required irregular blood transfusions. The A/G polymorphism in the 5'HS4 palindromic region was also observed to be variable. Family studies were carried out on a female G/G homozygous patient, who received irregular blood transfusions. Her father, who had the same heterozygous IVSII-1 beta-globin mutation but the A/G genotype at the 5'HS4 palindromic site, presented with mild anemia and no requirement for blood transfusions. This suggests an impact of SNPs in the 5'HS4 LCR core region on the thalassemia phenotype and offers an interesting subject for further investigations in the Iranian population.

Genotype-phenotype correlation in Iranian patients with Hb H disease.

Thalassemia is the most common genetic disorder in Iran. Some α-globin genotypes leading to Hb H disease may cause severe anemia and dependence on regular blood transfusions. In this study, 40 patients were analyzed for the molecular basis and the genotype-phenotype correlation of Hb H disease in Iran. α-Globin molecular analysis was performed by polymerase chain reaction (PCR) followed by agarose gel electrophoresis, reverse hybridization test strips or DNA sequencing. The most frequently observed α-globin genotypes were -α(3.7)/- -(MED) in 10 patients (25%), - -(20.5)/α(-5nt)α in six patients (15%) and - -(20.5)/-α(3.7) in four patients (10%). A subset of the identified Hb H genotypes, including - -(MED)/α(CS)α, - -(MED)/α(PolyA2)α and α(CS)α/α(CS)α, was associated with a need for regular or irregular blood transfusions. Our findings provide a basis for predicting phenotype severity by identifying the Hb H genotype and making more selective decisions for prenatal diagnosis.

VKORC1 -1639G>A and CYP2C9*3 are the major genetic predictors of phenprocoumon dose requirement.

PURPOSE: Phenprocoumon, similar to other coumarin-derived anticoagulants, is associated with a large variation in the individual dose requirement to achieve stable anticoagulation. Polymorphisms in the vitamin K epoxide reductase complex subunit 1 (VKORC1) and the liver enzyme cytochrome P450 2C9 (CYP2C9) effectively account for the variability in warfarin and acenocoumarol response but are less well-defined pharmacogenetic predictors in phenprocoumon therapy. METHODS: A retrospective study was performed on 185 outpatients attending anticoagulation clinics in Austria and Germany. These patients were genotyped for the VKORC1 -1639G>A and 3730G>A polymorphisms as well as for the CYP2C9 *2 and *3 polymorphisms using a reverse hybridisation-based teststrip assay. RESULTS: The VKORC1 -1639A allele, which was present at a frequency of 41.4% in the study cohort, significantly reduced the mean weekly phenprocoumon dose by 3 mg (19%) in the heterozygous and by 6.7 mg (43%) in the homozygous state compared to wild-type carriers (15.5 +/- 6.8 mg, p < 0.0001). A stepwise multiple regression analysis revealed that VKORC1 -1639G>A, age and CYP2C9*3 were the major independent determinants of phenprocoumon dose, accounting for 14.2, 9.1 and 4.7% of its variability, respectively (p A genotype had no additional predictive power for individual dose variability. CONCLUSION: Similar to warfarin and acenocoumarol, the VKORC1 -1639G>A polymorphism had the highest impact on the maintenance dose of phenprocoumon. The factor age was the second most important predictor and explained a greater percentage of the variability than CYP2C9 genotype.

alpha-Thalassemia mutation analyses in Mazandaran province, North Iran.

Two hundred and fifty-five patients from Mazandaran Province, Iran, all presenting with hypochromic and microcytic anemia, were selected for alpha-thalassemia (alpha-thal) mutation screening. We detected a total of 274 alpha-globin mutations in 227 (89%) of these patients. Among the 21 different alpha-globin alleles found, the -alpha(3.7) (44.9%), polyadenylation signal 2 (poly A2) (AATAAA>AATGAA) (18.2%), -alpha(4.2) (9.1%), alpha(IVS-I(-5 nt)) (6.5%), - -(MED) (4.3%), and alpha(codon 19 (-G)) (4%) were the most frequent. The other 15 mutations included variants that had not yet been observed in Iran, such as Hb Bleuland [alpha108(G15)ThrAsn, ACC>AAC (alpha2)], as well as a novel mutation on the alpha2 gene, also not described to date [3 ' untranslated region (3 'UTR) nucleotide (nt) 46 (C>A)]. These comprehensive new data are useful for establishing a screening strategy for the effective control of alpha-thal in Mazandaran Province.