The +33 C>G variant NM_000518.5(HBB):c.-18C>G in the 5' untranslated region (UTR) of the β-globin gene is described in the literature as both mild and silent, while it causes a phenotype of ...thalassemia intermedia in the presence of a severe β-thalassemia allele. Despite its potential clinical significance, the determination of its pathogenicity according to established standards requires a greater number of published cases and co-segregation evidence than what is currently available. The present study provides an extensive phenotypic characterization of +33 C>G using 26 heterozygous and 11 compound heterozygous novel cases detected in Cyprus and employs computational predictors (CADD, RegulomeDB) to better understand its impact on clinical severity. Genotype identification of globin gene variants, including α- and δ-thalassemia determinants, and rs7482144 (XmnI) was carried out using Sanger sequencing, gap-PCR, and restriction enzyme digestion methods. The heterozygous state of +33 C>G had a silent phenotype without apparent microcytosis or hypochromia, while compound heterozygosity with a β+ or β0 allele had a spectrum of clinical phenotypes. Awareness of the +33 C>G is required across Mediterranean populations where β-thalassemia is frequent, particularly in Cyprus, with significant relevance in population screening and fetal diagnostic applications.
The +33 C>G variant NM_000518.5(HBB):c.-18C>G in the 5′ untranslated region (UTR) of the β-globin gene is described in the literature as both mild and silent, while it causes a phenotype of ...thalassemia intermedia in the presence of a severe β-thalassemia allele. Despite its potential clinical significance, the determination of its pathogenicity according to established standards requires a greater number of published cases and co-segregation evidence than what is currently available. The present study provides an extensive phenotypic characterization of +33 C>G using 26 heterozygous and 11 compound heterozygous novel cases detected in Cyprus and employs computational predictors (CADD, RegulomeDB) to better understand its impact on clinical severity. Genotype identification of globin gene variants, including α- and δ-thalassemia determinants, and rs7482144 (XmnI) was carried out using Sanger sequencing, gap-PCR, and restriction enzyme digestion methods. The heterozygous state of +33 C>G had a silent phenotype without apparent microcytosis or hypochromia, while compound heterozygosity with a β+ or β0 allele had a spectrum of clinical phenotypes. Awareness of the +33 C>G is required across Mediterranean populations where β-thalassemia is frequent, particularly in Cyprus, with significant relevance in population screening and fetal diagnostic applications.
Haemoglobinopathies are the most common monogenic diseases, posing a major public health challenge worldwide. Cyprus has one the highest prevalences of thalassaemia in the world and has been the ...first country to introduce a successful population-wide prevention programme, based on premarital screening. In this study, we report the most significant and comprehensive update on the status of haemoglobinopathies in Cyprus for at least two decades. First, we identified and analysed all known 592 β-thalassaemia patients and 595 Hb H disease patients in Cyprus. Moreover, we report the molecular spectrum of α-, β- and δ-globin gene mutations in the population and their geographic distribution, using a set of 13824 carriers genotyped from 1995 to 2015, and estimate relative allele frequencies in carriers of β- and δ-globin gene mutations. Notably, several mutations are reported for the first time in the Cypriot population, whereas important differences are observed in the distribution of mutations across different districts of the island.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
β thalassemia carriers are usually symptom free with microcytic hypochromic red cells and a raised HbA2 level. However, an increased output of α globin through co-inheritance of extra α globin genes, ...converts a typically asymptomatic β thalassemia carrier state to that of thalassemia intermedia. We describe 3 families with 3 unique head-to-tail duplications of the a-globin cluster in which all the probands have thalassemia intermedia ranging from moderately severe anemia with splenomegaly, to transfusion-dependence.
In Family 1, both father (Chinese) and son (Chinese and Anglo-Saxon English) were heterozygous for the HBB:c.316-197C>T β thalassemia variant but had moderately severe anemia (Hb 67 to 91 g/L) with splenomegaly; they were both transfusion-independent. In Family 2, the father (Syrian) had normal hematology, while mother (Iraqi) had a hematologic profile (Hb 110 g/L, RBC 5.68x1012/L, MCH 19.4 pg, MCV 58.9 fL and HbA2 4.8%) typical of heterozygous β0 thalassemia. The proband presented at age 5 years with severe anemia precipitated by an infection, that needed a blood transfusion. She continued to need intermittent blood transfusion while an older sister with a hematologic profile of Hb 75 g/L, RBC 3.04x1012/L, MCH 19.6 pg, MCV 65.6 fL, and mild spenomegaly, remained transfusion-independent. In Family 3 (Greek Cypriot), both parents were asymptomatic; the father was heterozygous for the HBB:c.93-21G>A β thalassemia variant with a normal a globin genotype (aa/aa), and the mother had normal HbA2 levels. In contrast, both their daughter and son who had inherited father's β thalassemia, had moderately severe anemia and needed intermittent blood transfusion. In all 3 families, MLPA suggested duplication of the whole alpha globin cluster but could not differentiate the different duplications.
Next generation sequencing using Agilent SureSelect bait capture, targeted sequence analysis to the two globin loci. Sequence alignment to the reference sequence was performed using NextGene (SoftGenetits, USA). Analysis of the β globin gene sequence identified the β thalassemia-causing variant in each family. Comparison of the sequence coverage across the a loci on chromosome 16 between each case and normal controls, showed that where duplications had occurred, there was proportionally more sequence captured, similar to SNP or CGH array analysis. At the point where the sequence coverage increased, a duplication breakpoint was suspected, and the aligned sequence reads were examined in detail. In Family 1, individual sequence reads matched part of the reference sequence, BLAT query in UCSC showed that the two halves of the read aligned at either end of the duplication, indicating they were sequences that spanned the head-to-tail breakpoint. This was confirmed by gap-PCR and Sanger sequence analysis. In the other two families, breakpoints were identified within repetitive regions which could not be captured by the baits and were therefore not covered by the captured sequence. The high resolution of the coverage map allowed precise characterisation of the duplications by gap-PCR and Sanger sequencing analysis of the breakpoint amplicons.
Duplication of the a globin cluster was encompassed in 188.7 kb in family 1, 120.5 kb in family 2, and 274 kb in family 3 (Figure 1). Both father and son in Family 1 were heterozygous for the duplicated a globin cluster (aa/aa, aa) and HBB:c.316-197C>T mutation. Both siblings in Family 2 were heterozygous for mother's β thalassemia c.135delC variant and father's duplicated a globin cluster. In family 3, the mother had a 3.7 kb a deletion variant and a duplicated a globin cluster (-a3.7/aa, aa), a total of 5 a globin genes. Both the daughter and son had inherited mother's duplicated a globin cluster with father's β thalassemia variant.
These families clearly show that a duplicated a globin cluster does not have a discernible phenotype on its own but is readily detectable when co-inherited with a β thalassemia variant. In all 3 cases, the a globin cluster duplications are in a head-to-tail orientation and occurred in repeats. These have most likely formed by non-homologous recombination events involving repetitive Alu sequences interspersed throughout the region. Whether these events occur more commonly in this region or if the region tolerates these changes better, allowing them to accumulate, remains to be resolved.
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GEOZS, IJS, IMTLJ, KILJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBJE, UILJ, UL, UM, UPCLJ, UPUK, ZAGLJ, ZRSKP
α-Thalassemia (α-thal) is widely reported in the Arabian Peninsula as one of the main causes of asymptomatic microcytic hypochromic red blood cells with or without anemia in the pediatric population. ...This is the first study that provides information about the molecular basis of α-thal in the Qatari population. Qatari school children between the ages of 5 and 15, exhibiting laboratory findings suggestive of microcytic anemia were pooled, and those with a mean corpuscular volume (MCV) of <80.0 fL and a hemoglobin (Hb) electropherogram that ruled out β-thalassemia (β-thal), were narrowed down to a group of 127. This group was screened for the −α
3.7
(rightward) deletion, and the α
−5 nt
, α
polyA1
(α
T-Saudi
), α
polyA2
mutations. A second group of randomly selected Qatari individuals was also screened in order to determine the population's allele frequency for the −α
3.7
deletion. Thirty-nine point four percent of the individuals with microcytic hypochromic anemia were positive for the −α
3.7
deletion (heterozygotes 30.0%, homozygotes 9.4%), 2.6% were positive for the α
polyA1
deletion and 0.8% positive for the α
−5 nt
mutation. None of the children exhibited changes in α
polyA2
. Analysis of the random samples determined that 26.4% were heterozygous and 4.5% homozygous for the −α
3.7
deletion with a 17.7% allele frequency. Our results suggest that a significant number of the Qatari pediatric population with microcytic hypochromic anemia are carriers of α-thal mutations. However, 45.6% of the children failed to exhibit any of the above four mutations tested. This suggests the possibility of other mutations in the Qatari pediatric population that are yet to be elicited.
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DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, UILJ, UKNU, UL, UM, UPUK
Background: This study investigated alternative, non-invasive methods for human papillomavirus (HPV) detection in head and neck cancers (HNCs). We compared two approaches: analyzing computed ...tomography (CT) scans with a Deep Learning (DL) model and using radiomic features extracted from CT images with machine learning (ML) models. Methods: Fifty patients with histologically confirmed HNC were included. We first trained a modified ResNet-18 DL model on CT data to predict HPV status. Next, radiomic features were extracted from manually segmented regions of interest near the oropharynx and used to train four ML models (K-Nearest Neighbors, logistic regression, decision tree, random forest) for the same purpose. Results: The CT-based model achieved the highest accuracy (90%) in classifying HPV status. Among the ML models, K-Nearest Neighbors performed best (80% accuracy). Weighted Ensemble methods combining the CT-based model with each ML model resulted in moderate accuracy improvements (70–90%). Conclusions: Our findings suggest that CT scans analyzed by DL models hold promise for non-invasive HPV detection in HNC. Radiomic features, while less accurate in this study, offer a complementary approach. Future research should explore larger datasets and investigate the potential of combining DL and radiomic techniques.
Recent economic evaluations of alternative approaches to identifying and diagnosing chromosomal abnormalities in the fetus have focused on the relative merits of less invasive screening markers, such ...as maternal serum alpha-fetoprotein, unconjugated oestriol, and beta-human chorionic gonadotropin, with or without signs from early ultrasound scan, and have attempted to assess how many and which markers can be justified on cost-effectiveness grounds.1 This week in The Lancet, Ryan Harris and colleagues present a novel economic evaluation, which argues that prenatal risk assessment may not be required and that offering universal prenatal diagnostic testing can be justified on economic grounds. These investigators estimated the incremental cost-effectiveness of amniocentesis and chorionic villus sampling versus no invasive testing by synthesising epidemiological, clinical effectiveness, cost, and preference data from multiple sources within a decision-analytic framework. The most compelling rationale for the study is that (implied) clinical valuations of the risks (mainly, an increased risk of miscarriage) and benefits (mainly, information that can inform future pregnancy choices) of fetal karyotyping might not be shared by pregnant women themselves. Indeed, the study's results hinge on preference data from a sample of 534 sociodemographically diverse pregnant women from the San Francisco Bay area, for whom procedure-related miscarriage was considered preferable to having a baby with Down's syndrome.2 By combining these preference data with the likelihood of adverse outcomes, as well as clinical effectiveness, cost, and broader epidemiological evidence, Harris and colleagues estimate that amniocentesis results in an incremental cost per quality-adjusted life-year gained of less than US$15 000, regardless of women's ages or risk levels.
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DOBA, GEOZS, IJS, IMTLJ, IZUM, KILJ, KISLJ, NUK, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SIK, UILJ, UKNU, UL, UM, UPCLJ, UPUK, VSZLJ
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