Rapid identification of the intron 22 inversion in haemophilia A

Kloppers, Jean Frederick; Rensburg, Walter J. Janse van

doi:10.1111/hae.13142

Cited by 3 publications

(7 citation statements)

References 13 publications

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“…Inv22 carriers displayed both the mutant and wild-type PCR fragments. [10] The real-time inv22 assay detected 10 haemophilia A patients with inv22 (29.4%) and 3 inv22 carriers (16.7%). The remainder of the patients were all inv22-negative.…”

Section: Resultsmentioning

confidence: 97%

“…Inv22 mutation status is unknown in most SA haemophilia A patients, with very few studies published on patient genotypes. [10] Possible carriers were included in the study, as carrier detection plays an important role in counselling haemophilia A families with regard to family planning and the risk associated with having a child with a bleeding disorder. In our setting, only severe haemophilia A patients actively attend our haemophilia clinics, and the population group may therefore be misrepresented with regard to mild and moderate haemophilia A patients.…”

Section: Discussionmentioning

confidence: 99%

“…All the participants were screened with the published conventional inv22 PCR assay. [10] The Southern blot-confirmed inv22 patient samples were included in each run to serve as positive and negative inv22 controls. After gel electrophoresis, gel images were analysed to determine the inv22 genotypes of each study participant.…”

Section: Inv22 Screeningmentioning

confidence: 99%

“…Sanger sequencing was performed on all samples to confirm the PCR results and determine whether exon 23c is spliced to exon 22 in all positive inv22 cases. [10] Following cDNA synthesis, the amplification product of each sample was treated with ExoSAP-IT Express reagent (Affymetrix, USA). The purified PCR product consequently served as template for the sequencing reaction, using the BigDye Terminator v3.1 Kit (Applied Biosystems, USA), and impurities were removed using the ZR DNA Sequencing Clean-Up Kit (Zymo Research, USA).…”

Section: Sanger Sequence Analysismentioning

confidence: 99%

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Intron 22 inversion real-time polymerase chain reaction detection in haemophilia A families from central South Africa

2019

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Haemophilia A is an X-linked recessive bleeding disorder characterised by deficiency of coagulation factor VIII (FVIII). [1] The disorder can be classified according to procoagulant levels of FVIII: patients with severe haemophilia A have levels of <1 IU/dL, those with moderate disease 1-5 IU/dL, and those with mild disease 5-40 IU/ dL. The disorder mainly affects males, at a rate of 1 in 5 000-10 000 worldwide. [1,2] The 2017 World Federation of Hemophilia survey [3] reported that 149 764 individuals were affected by haemophilia A globally, with only 1 848 reported in South Africa (SA). Haemophilia A is underdiagnosed in SA, as in many other developing countries, owing to a lack of access to diagnostic expertise. [4] There is currently only one genetic testing facility, which services all the haemophilia A treatment centres in SA. [4] Genetic research on haemophilia is therefore limited in SA, [4] as evident from the FVIII gene (F8) variant database, to which SA has made no contribution. [5] Genetic testing is of paramount importance in the comprehensive treatment plan for haemophilia A, since mutations in F8 are the cause of the disorder. Furthermore, F8 mutations may be associated with the development of FVIII inhibitors, [6] and variant detection allows for prenatal and carrier detection to enable genetic counselling of haemophilia A families. [7] According to the literature, 45% of all severe haemophilia A cases are associated with an inversion in intron 22 (inv22) of F8. [8,9] Based on gene expression, an mRNA conventional inv22 polymerase chain reaction (PCR) screening method was developed to allow rapid detection of inv22. [10] This method assumes the presence of an additional exon, called exon 23c, contained in the inv22 mutant FVIII transcript. [11] Exon 23c was found to be spliced to exon 22 of F8 in unrelated haemophilia A patients. [10] Objectives To detect the presence of inv22 in the haemophilia A population of central SA and potential haemophilia A carriers. This open-access article is distributed under Creative Commons licence CC-BY-NC 4.0.

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Section: Resultsmentioning

confidence: 97%

Section: Discussionmentioning

confidence: 99%

Section: Inv22 Screeningmentioning

confidence: 99%

Section: Sanger Sequence Analysismentioning

confidence: 99%

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Intron 22 inversion real-time polymerase chain reaction detection in haemophilia A families from central South Africa

2019

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“…In an individual with a positive Inv22, exon 22 of F8 is spliced to an alternative exon to exon 23, exon 23c, thus, clearly distinguishing Inv22 from the wild type. 4 Therefore, our local hemophilia treatment center (HTC) has started to screen all people with hemophilia A in our region for the Inv22 variation. It was previously reported that this variation is under-represented in people with severe hemophilia in our area (∼29%) when compared to the globally reported prevalence (∼45%).…”

Section: Introductionmentioning

confidence: 99%

De Novo Intron 22 Inversion Carrier Detection—The Emerging Value of Genetic Analysis in a Developing Country

Kloppers¹,

Plessis²,

Rensburg³

2021

Clin Med�Insights�Blood�Disord

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Genetic analysis of the F8 gene can play a vital role in the diagnosis and management of people with hemophilia A (PWH). Knowing the causative mutation can assist treating clinicians in predicting the risk of inhibitor formation and a person’s response to treatment. It is also a vital tool for genetic counselors in assisting PWH with family planning. Mutational analysis has been limited in developing countries, mainly due to resource constraints; however, with the development of more cost-effective screening tools, these assays are becoming more common in developing countries. Unfortunately, the clinical utility of these assays remain unclear in some of these underresourced countries, resulting in a lack of clinician buy-in. We aimed to highlight the wide-ranging impact mutational analysis potentially has on PWH and their extended families in a developing country. We used a gel-based in-house method to determine the inversion 22 (Inv22) mutation in a family where a young boy, without a family history, was recently diagnosed with hemophilia A. Inv22 was detected in the proband and his mother, but not in any other direct family member. Therefore, we concluded that the mutation was a de novo mutation in the mother. Subsequently, this result has bearing on at least 23 direct blood-relatives. Genetic screening extends far wider than the treatment of PWH and is vital in the management of PWH and their extended families. With the availability of cost-effective genetic screening assays this should also be the case in developing countries.

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Systematic Review of Genetic-Related Risk Factor and Inhibitor Epidemiology in People with Severe Hemophilia a from Africa: A 2023 Update

Baglo,

Zohoun,

Azonbakin

et al. 2023

IBRR

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Background and Aims: Prevalence of factor VIII inhibitors in patients with hemophilia A varies from study to study, ranging from 15% to 30%. The important risk of inhibitor development is factor VIII mutation responsible for hemophilia A. Few studies have reported factor VIII mutations in Africa. The aim of this study was to review on FVIII gène mutations of severe hemophilia A in Africa and those associated with inhibitor development. Study Design and Methodology: A systematic review was carried out using the electronic databases Pubmed, Science Direct, Index Medicus Global and African Journals online and the key words "hemophilia A", "inhibitor", "genetic" and "Africa". Studies written in French or English on the African continent and published between 2012 and 2023 were included. Publications relating to acquired hemophilia and duplicates were excluded. In the end, 17 articles were selected. Results: The factor VIII mutations involved in severe hemophilia A in Africa are variable, consisting of intron 22 inversion, large or point deletions, nonsense and missense mutations and splicing abnormalities. Among the latter, numerous previously unrecorded mutations have been identified, and a single case of intron 1 inversion has been found in Algeria. Prevalence of factor VIII inhibitors in severe hemophilia A in Africa varies between 7,8% and 30%. Genetic abnormalities associated with inhibitors include intron 22 inversion, large deletions such as exon 1-13 deletion, nonsense mutations and c.1010-2A>G mutation. Conclusion: A better knowledge of the factor VIII mutations involved in severe hemophilia A in Africa will help improve patient management.

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Rapid identification of the intron 22 inversion in haemophilia A

Cited by 3 publications

References 13 publications

Intron 22 inversion real-time polymerase chain reaction detection in haemophilia A families from central South Africa

Intron 22 inversion real-time polymerase chain reaction detection in haemophilia A families from central South Africa

De Novo Intron 22 Inversion Carrier Detection—The Emerging Value of Genetic Analysis in a Developing Country

Systematic Review of Genetic-Related Risk Factor and Inhibitor Epidemiology in People with Severe Hemophilia a from Africa: A 2023 Update

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