{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,11,5]],"date-time":"2026-11-05T18:51:36Z","timestamp":1793904696761,"version":"4.8.0"},"reference-count":37,"publisher":"MDPI AG","issue":"6","license":[{"start":{"date-parts":[[2022,5,28]],"date-time":"2022-05-28T00:00:00Z","timestamp":1653696000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100004681","name":"Higher Education Commission of Pakistan","doi-asserted-by":"publisher","award":["NRPU 2093"],"award-info":[{"award-number":["NRPU 2093"]}],"id":[{"id":"10.13039\/501100004681","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Genes"],"abstract":"<jats:p>Type III von Willebrand disease is present in the Punjab province of Pakistan along with other inherited bleeding disorders like hemophilia. Cousin marriages are very common in Pakistan so genetic studies help to establish protocols for screening, especially at the antenatal level. Factors behind the phenotypic variation of the severity of bleeding in type III vWD are largely unknown. The study was conducted to determine Mutations\/genetic alterations in type III von Willebrand disease and also to determine the association of different mutations, methylation status, ITGA2B\/B3 mutations and alloimmunization with the severity of type III vWD. After informed consent and detailed history of the patients, routine tests and DNA extraction from blood, mutational analysis was performed by Next Generation Sequencing on Ion Torrent PGM. DNA methylation status was also checked with the help of PCR. In our cohort, 55 cases were detected with pathogenic mutations. A total of 27 different mutations were identified in 55 solved cases; 16 (59.2%) were novel. The mean bleeding score in truncating mutations and essential splice site mutations was relatively higher than weak and strong missense mutations. The mean bleeding score showed insignificant variation for different DNA methylation statuses of the VWF gene at the cg23551979 CpG site. Mutations in exons 7,10, 25, 28, 31, 43, and intron 41 splice site account for 75% of the mutations.<\/jats:p>","DOI":"10.3390\/genes13060971","type":"journal-article","created":{"date-parts":[[2022,5,29]],"date-time":"2022-05-29T08:50:51Z","timestamp":1653814251000},"page":"971","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":5,"title":["Genetic Alterations, DNA Methylation, Alloantibodies and Phenotypic Heterogeneity in Type III von Willebrand Disease"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-4941-0153","authenticated-orcid":false,"given":"Muhammad Asif","family":"Naveed","sequence":"first","affiliation":[{"name":"Department of Haematology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8977-7315","authenticated-orcid":false,"given":"Aiysha","family":"Abid","sequence":"additional","affiliation":[{"name":"Centre for Human Genetics and Molecular Medicine, Sindh Institute of Urology Transplantation, Karachi 42000, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nadir","family":"Ali","sequence":"additional","affiliation":[{"name":"Kulsoom International Hospital, 2020 Jinnah Ave, G 6\/2 Blue Area, Islamabad 53000, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yaqoob","family":"Hassan","sequence":"additional","affiliation":[{"name":"Chughtai\u2019s Lahore Lab, 42300 Jail Road, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3240-0741","authenticated-orcid":false,"given":"Ali","family":"Amar","sequence":"additional","affiliation":[{"name":"Human Genetics and Molecular Biology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Aymen","family":"Javed","sequence":"additional","affiliation":[{"name":"Department of Obstetrics and Gynaecology Services Hospital Jail Road, Lahore 42500, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Khansa","family":"Qamar","sequence":"additional","affiliation":[{"name":"Department of Haematology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ghulam","family":"Mustafa","sequence":"additional","affiliation":[{"name":"Department of Haematology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9765-8818","authenticated-orcid":false,"given":"Ali","family":"Raza","sequence":"additional","affiliation":[{"name":"Centre for Human Genetics and Molecular Medicine, Sindh Institute of Urology Transplantation, Karachi 42000, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Umera","family":"Saleem","sequence":"additional","affiliation":[{"name":"Department of Pathology, Nishter Medical University, Nishter Road, Multan 32003, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shabbir","family":"Hussain","sequence":"additional","affiliation":[{"name":"Department of Biochemistry, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Madiha","family":"Shakoor","sequence":"additional","affiliation":[{"name":"Human Genetics and Molecular Biology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shagufta","family":"Khaliq","sequence":"additional","affiliation":[{"name":"Human Genetics and Molecular Biology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Shahida","family":"Mohsin","sequence":"additional","affiliation":[{"name":"Department of Haematology, University of Health Sciences, Khayaban Jamia Punjab, Lahore 52600, Pakistan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2022,5,28]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","unstructured":"Castaman, G., and Linari, S. (2017). Diagnosis and Treatment of von Willebrand Disease and Rare Bleeding Disorders. J. Clin. Med., 6.","DOI":"10.3390\/jcm6040045"},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"7","DOI":"10.4103\/0250-474X.89751","article-title":"Von Willebrand Disease: An Overview","volume":"73","author":"Bharati","year":"2011","journal-title":"Indian J. Pharm. Sci."},{"key":"ref_3","unstructured":"Adam, M.P., Ardinger, H.H., Pagon., R.A., Wallace, S.E., Bean, L.J.H., Gripp, K.W., Mirzaa., G.M., and Amemiya., A. (2011). von Willebrand Disease, University of Washington."},{"key":"ref_4","unstructured":"(2022, May 10). Orphanet: Von Willebrand Disease. Available online: https:\/\/www.orpha.net\/consor\/cgi-bin\/OC_Exp.php?lng=EN&Expert=903."},{"key":"ref_5","doi-asserted-by":"crossref","first-page":"40","DOI":"10.4274\/tjh.2011.0020","article-title":"The Prevalence of von Willebrand Disease and Significance of in Vitro Bleeding Time (PFA-100) in von Willebrand Disease Screening in the \u0130zmir Region","volume":"30","author":"Kavakli","year":"2013","journal-title":"Turk. J. Hematol."},{"key":"ref_6","doi-asserted-by":"crossref","first-page":"E102","DOI":"10.1177\/1076029610387125","article-title":"Clinical features and types of von Willebrand disease in Karachi","volume":"17","author":"Borhany","year":"2010","journal-title":"Clin. Appl. Thromb."},{"key":"ref_7","doi-asserted-by":"crossref","first-page":"19514","DOI":"10.1016\/S0021-9258(19)47144-5","article-title":"The Journal of Biological Chemistry Structure of the Gene for Human von Willebrand Factor*","volume":"264","author":"Mancum","year":"1989","journal-title":"J. Biol. Chem."},{"key":"ref_8","doi-asserted-by":"crossref","first-page":"4689","DOI":"10.1182\/blood.V126.23.4689.4689","article-title":"Molecular Genetics Analysis of Von Williebrand Disease: Studies in Cohort Pakistani Patients","volume":"126","author":"Ahmed","year":"2015","journal-title":"Blood"},{"key":"ref_9","doi-asserted-by":"crossref","unstructured":"Kasatkar, P., Shetty, S., and Ghosh, K. (2014). Genetic Heterogeneity in a Large Cohort of Indian Type 3 von Willebrand Disease Patients. PLoS ONE, 9.","DOI":"10.1371\/journal.pone.0092575"},{"key":"ref_10","doi-asserted-by":"crossref","first-page":"1048","DOI":"10.1111\/j.1365-2516.2009.02059.x","article-title":"The mutation spectrum associated with type 3 von Willebrand disease in a cohort of patients from the north west of England","volume":"15","author":"Sutherland","year":"2009","journal-title":"Haemophilia"},{"key":"ref_11","doi-asserted-by":"crossref","first-page":"5031","DOI":"10.1182\/blood-2018-99-112976","article-title":"A Unique Case of Type 3 Von Willebrand Disease","volume":"132","author":"Mandava","year":"2018","journal-title":"Blood"},{"key":"ref_12","doi-asserted-by":"crossref","first-page":"311","DOI":"10.5114\/pjp.2019.93135","article-title":"Acquired von Willebrand syndrome associated with plasma cell myeloma\u2014A rare histopathological bone marrow image","volume":"70","author":"Chraszczewska","year":"2020","journal-title":"Pol. J. Pathol."},{"key":"ref_13","doi-asserted-by":"crossref","unstructured":"Sottilotta, G., Luise, F., Massara, E., Oriana, V., and Piromalli, A. (2017). Efficacy of Octocog Alfa (Advate) in a Child with Type 3 von Willebrand Disease and Alloantibodies. J. Clin. Med., 6.","DOI":"10.3390\/jcm6090085"},{"key":"ref_14","doi-asserted-by":"crossref","first-page":"280","DOI":"10.1182\/bloodadvances.2020003265","article-title":"ASH ISTH NHF WFH 2021 guidelines on the diagnosis of von Willebrand disease","volume":"5","author":"James","year":"2021","journal-title":"Blood Adv."},{"key":"ref_15","doi-asserted-by":"crossref","first-page":"171","DOI":"10.1111\/j.1365-2516.2007.01643.x","article-title":"von Willebrand disease (VWD): Evidence-based diagnosis and management guidelines, the National Heart, Lung, and Blood Institute (NHLBI) expert panel report (USA)","volume":"14","author":"Nichols","year":"2008","journal-title":"Haemophilia"},{"key":"ref_16","doi-asserted-by":"crossref","first-page":"952","DOI":"10.1002\/rth2.12392","article-title":"Classification of von Willebrand disease in the context of modern contemporary von Willebrand factor testing methodologies","volume":"4","author":"Favaloro","year":"2020","journal-title":"Res. Pract. Thromb. Haemost."},{"key":"ref_17","doi-asserted-by":"crossref","first-page":"2062","DOI":"10.1111\/j.1538-7836.2008.03182.x","article-title":"Generation and validation of the Condensed MCMDM-1VWD Bleeding Questionnaire for von Willebrand disease","volume":"6","author":"Bowman","year":"2008","journal-title":"J. Thromb. Haemost."},{"key":"ref_18","doi-asserted-by":"crossref","first-page":"pdb.prot4455","DOI":"10.1101\/pdb.prot4455","article-title":"Purification of nucleic acids by extraction with phenol: Chloroform","volume":"2006","author":"Sambrook","year":"2006","journal-title":"Cold Spring Harb. Protoc."},{"key":"ref_19","doi-asserted-by":"crossref","first-page":"3397","DOI":"10.1002\/elps.201200424","article-title":"Progress in ion torrent semiconductor chip based sequencing","volume":"33","author":"Merriman","year":"2012","journal-title":"Electrophoresis"},{"key":"ref_20","doi-asserted-by":"crossref","first-page":"348","DOI":"10.1038\/nature10242","article-title":"An integrated semiconductor device enabling non-optical genome sequencing","volume":"475","author":"Rothberg","year":"2011","journal-title":"Nature"},{"key":"ref_21","doi-asserted-by":"crossref","first-page":"211","DOI":"10.1007\/s00439-019-01978-x","article-title":"Gene panel sequencing identifies a likely monogenic cause in 7% of 235 Pakistani families with nephrolithiasis","volume":"138","author":"Amar","year":"2019","journal-title":"Hum. Genet."},{"key":"ref_22","doi-asserted-by":"crossref","first-page":"248","DOI":"10.1038\/nmeth0410-248","article-title":"A method and server for predicting damaging missense mutations","volume":"7","author":"Adzhubei","year":"2010","journal-title":"Nat. Methods"},{"key":"ref_23","doi-asserted-by":"crossref","first-page":"1073","DOI":"10.1038\/nprot.2009.86","article-title":"Predicting the effects of coding non-synonymous variants on protein function using the SIFT algorithm","volume":"4","author":"Kumar","year":"2009","journal-title":"Nat. Protoc."},{"key":"ref_24","doi-asserted-by":"crossref","first-page":"575","DOI":"10.1038\/nmeth0810-575","article-title":"Mutation Taster evaluates disease-causing potential of sequence alterations","volume":"7","author":"Schwarz","year":"2010","journal-title":"Nat. Methods"},{"key":"ref_25","doi-asserted-by":"crossref","first-page":"1729","DOI":"10.1111\/j.1538-7836.2008.03107.x","article-title":"An Alu-mediated novel large deletion is the most frequent cause of type 3 von Willebrand disease in Hungary","volume":"6","author":"Mohl","year":"2008","journal-title":"J. Thromb. Haemost."},{"key":"ref_26","doi-asserted-by":"crossref","first-page":"512","DOI":"10.1111\/jth.12130","article-title":"The genetics of Canadian type 3 von Willebrand disease: Further evidence for co-dominant inheritance of mutant alleles","volume":"11","author":"Bowman","year":"2013","journal-title":"J. Thromb. Haemost."},{"key":"ref_27","doi-asserted-by":"crossref","first-page":"1035","DOI":"10.1111\/hae.13841","article-title":"Characterization of the mutation spectrum in a Pakistani cohort of type 3 von Willebrand disease","volume":"25","author":"Ahmed","year":"2019","journal-title":"Haemophilia"},{"key":"ref_28","doi-asserted-by":"crossref","first-page":"2005","DOI":"10.3324\/haematol.2017.168765","article-title":"Molecular and clinical profile of von Willebrand disease in Spain (PCM-EVW-ES): Comprehensive genetic analysis by next-generation sequencing of 480 patients","volume":"102","author":"Batlle","year":"2017","journal-title":"Haematologica"},{"key":"ref_29","doi-asserted-by":"crossref","first-page":"365","DOI":"10.1097\/GIM.0b013e3182035931","article-title":"von Willebrand Disease","volume":"13","author":"James","year":"2011","journal-title":"Genet. Med."},{"key":"ref_30","doi-asserted-by":"crossref","first-page":"2103","DOI":"10.1111\/j.1538-7836.2006.02146.x","article-title":"Update on the pathophysiology and classification of von Willebrand disease: A report of the Subcommittee on von Willebrand Factor","volume":"4","author":"Sadler","year":"2006","journal-title":"J. Thromb. Haemost."},{"key":"ref_31","doi-asserted-by":"crossref","first-page":"264","DOI":"10.1016\/S1079-9796(03)00033-0","article-title":"Molecular defects in type 3 von Willebrand disease: Updated results from 40 multiethnic patients","volume":"30","author":"Baronciani","year":"2003","journal-title":"Blood Cells Mol. Dis."},{"key":"ref_32","doi-asserted-by":"crossref","first-page":"470","DOI":"10.1182\/blood.V124.21.470.470","article-title":"Epigenetic Regulation of Von Willebrand Factor Gene Expression May Contribute to Von Willebrand Disease Severity","volume":"124","author":"Kuldanek","year":"2014","journal-title":"Blood"},{"key":"ref_33","doi-asserted-by":"crossref","first-page":"301","DOI":"10.1182\/bloodadvances.2020003264","article-title":"ASH ISTH NHF WFH 2021 guidelines on the management of von Willebrand disease","volume":"5","author":"Connell","year":"2021","journal-title":"Blood Adv."},{"key":"ref_34","doi-asserted-by":"crossref","first-page":"731","DOI":"10.1002\/humu.24204","article-title":"von Willebrand factor propeptide missense variants affect anterograde transport to Golgi resulting in ER retention","volume":"42","author":"Yadegari","year":"2021","journal-title":"Hum. Mutat."},{"key":"ref_35","doi-asserted-by":"crossref","first-page":"15647","DOI":"10.1073\/pnas.0705175104","article-title":"Two Cys residues essential for von Willebrand factor multimer assembly in the Golgi","volume":"104","author":"Purvis","year":"2007","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"ref_36","doi-asserted-by":"crossref","first-page":"1523","DOI":"10.1182\/blood-2018-10-876300","article-title":"The von Willebrand factor D\u2032D3 assembly and structural principles for factor VIII binding and concatemer biogenesis","volume":"133","author":"Dong","year":"2019","journal-title":"Blood"},{"key":"ref_37","doi-asserted-by":"crossref","first-page":"11","DOI":"10.1097\/MBC.0b013e32834cb35d","article-title":"Structure and function of von Willebrand factor: The protein that is deficient and\/or abnormal in inherited von Willebrand disease","volume":"23","author":"Hassan","year":"2012","journal-title":"Blood Coagul. Fibrinolysis"}],"container-title":["Genes"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.mdpi.com\/2073-4425\/13\/6\/971\/pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,10,10]],"date-time":"2025-10-10T23:20:38Z","timestamp":1760138438000},"score":1,"resource":{"primary":{"URL":"https:\/\/www.mdpi.com\/2073-4425\/13\/6\/971"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2022,5,28]]},"references-count":37,"journal-issue":{"issue":"6","published-online":{"date-parts":[[2022,6]]}},"alternative-id":["genes13060971"],"URL":"https:\/\/doi.org\/10.3390\/genes13060971","relation":{},"ISSN":["2073-4425"],"issn-type":[{"value":"2073-4425","type":"electronic"}],"subject":[],"published":{"date-parts":[[2022,5,28]]}}}