{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,15]],"date-time":"2026-09-15T12:03:15Z","timestamp":1789473795087,"version":"build-2803163510"},"reference-count":42,"publisher":"Georg Thieme Verlag KG","issue":"06","content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Thromb Haemost"],"published-print":{"date-parts":[[2026,6]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>With exon-focused approaches, a proportion of protein C (PC) deficiencies remains genetically unresolved. In these cases, deep intronic variation or structural variation could be causal.<\/jats:p>\n                  <jats:p>To identify the causal variation in unrelated patients presenting a PC deficiency in whom no genetic variation was found using conventional exploration.<\/jats:p>\n                  <jats:p>Whole PROC gene was analyzed using next generation sequencing (NGS) in probands with unexplained confirmed decreased PC clot activity after genetic exploration in our centres since 2000. The pathogenic impact of identified candidate variants was assessed using both in silico analysis (MaxEntScan and SpliceAI) and in vitro splicing assay in HeLa and Huh7 cell lines.<\/jats:p>\n                  <jats:p>Among 2,263 probands, 53 remained unexplained after conventional genetic exploration. Whole PROC gene sequencing was performed in 38\/53 probands for which a DNA sample was available. In total, 9 candidate variants from 11 probands (29%) were identified. They corresponded to 7 single nucleotide variants, one 541\u2009bp deletion, and one 1.298\u2009kb balanced inversion. Both the 541\u2009bp deletion and the large balanced inversion disrupted PROC and were considered as pathogenic. Among the seven deep intronic substitutions, splicing functional assay found a deleterious impact (intron retention in mature mRNA or pseudo-exon activation) for four of them: c.237\u2009+\u200975G\u2009&gt;\u2009A, c.535\u2009+\u2009936C\u2009&gt;\u2009T, c.536\u201395G\u2009&gt;\u2009A, and c.796\u2009+\u200949G\u2009&gt;\u2009T. Thus, these variants were classified as likely pathogenic. Finally, NGS allowed the identification of causal variants in 8\/38 patients previously unsolved protein C deficiency (21%).<\/jats:p>\n                  <jats:p>This study highlights the value of NGS for whole PROC gene sequencing in unexplained protein C deficiency.<\/jats:p>","DOI":"10.1055\/a-2668-5003","type":"journal-article","created":{"date-parts":[[2025,8,11]],"date-time":"2025-08-11T22:50:47Z","timestamp":1754952647000},"page":"582-594","source":"Crossref","is-referenced-by-count":1,"title":["Whole PROC Gene Sequencing to Explain Genetically Unresolved Protein C Deficiencies"],"prefix":"10.1055","volume":"126","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-3946-2312","authenticated-orcid":false,"given":"Laetitia","family":"Mauge","sequence":"additional","affiliation":[{"name":"Service d'H\u00e9matologie biologique, AP HP, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, France"},{"name":"Universit\u00e9 Paris Cit\u00e9, Inserm, PARCC, Paris, France"},{"name":"Universit\u00e9 Claude Bernard Lyon 1, UR 4609 H\u00e9mostase et thrombose, Lyon, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Carla","family":"Rial","sequence":"additional","affiliation":[{"name":"Service d'H\u00e9matologie biologique, AP HP, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, France"},{"name":"Universit\u00e9 Paris Cit\u00e9, Inserm, PARCC, Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Philippe De","family":"Mazancourt","sequence":"additional","affiliation":[{"name":"Service d'H\u00e9matologie biologique, AP HP, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, France"},{"name":"Universit\u00e9 Versailles-St-Quentin, Inserm, UMR 1179, St-Quentin en Yvelines, France"},{"name":"Laboratoire de Biologie Mol\u00e9culaire, AP HP, H\u00f4pital Ambroise Par\u00e9, Boulogne-Billancourt, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dominique","family":"Helley","sequence":"additional","affiliation":[{"name":"Service d'H\u00e9matologie biologique, AP HP, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, France"},{"name":"Universit\u00e9 Paris Cit\u00e9, Inserm, PARCC, Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Isabelle","family":"Pr\u00e9sot","sequence":"additional","affiliation":[{"name":"Service d'H\u00e9matologie biologique, AP HP, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Clarisse","family":"Billon","sequence":"additional","affiliation":[{"name":"Service de M\u00e9decine G\u00e9nomique des Maladies Rares, Groupe Hospitalier Universitaire Centre, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, AP-HP, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sylvie","family":"Labrouche-Colomer","sequence":"additional","affiliation":[{"name":"Laboratoire d'H\u00e9mostase mol\u00e9culaire, Groupe hospitalier Pellegrin, CHU de Bordeaux, Bordeaux, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Annabelle","family":"Venisse","sequence":"additional","affiliation":[{"name":"Service de M\u00e9decine G\u00e9nomique des Maladies Rares, Groupe Hospitalier Universitaire Centre, H\u00f4pital Europ\u00e9en Georges Pompidou, Paris, AP-HP, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yesim","family":"Dargaud","sequence":"additional","affiliation":[{"name":"Universit\u00e9 Claude Bernard Lyon 1, UR 4609 H\u00e9mostase et thrombose, Lyon, France"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karine","family":"Auribault","sequence":"additional","affiliation":[{"name":"Plateforme de Biologie Mol\u00e9culaire, AP HP, 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Res"},{"issue":"01","key":"ref2","doi-asserted-by":"crossref","first-page":"87","DOI":"10.1055\/s-0038-1653730","article-title":"Prevalence of protein C deficiency in the healthy population","volume":"73","author":"R C Tait","year":"1995","journal-title":"Thromb Haemost"},{"issue":"22","key":"ref3","doi-asserted-by":"crossref","first-page":"1512","DOI":"10.1056\/NEJM199011293232202","article-title":"Deficiencies of coagulation-inhibiting and fibrinolytic proteins in outpatients with deep-vein thrombosis","volume":"323","author":"H Heijboer","year":"1990","journal-title":"N Engl J Med"},{"issue":"02","key":"ref4","doi-asserted-by":"crossref","first-page":"271","DOI":"10.1111\/jth.14667","article-title":"Recommendations for clinical laboratory testing for protein C deficiency, for the subcommittee on plasma coagulation inhibitors of the ISTH","volume":"18","author":"P C Cooper","year":"2020","journal-title":"J Thromb Haemost"},{"issue":"03","key":"ref5","first-page":"256","article-title":"Familial thrombophilia: a complex genetic disorder","volume":"34","author":"B P Koeleman","year":"1997","journal-title":"Semin Hematol"},{"issue":"07","key":"ref6","doi-asserted-by":"crossref","first-page":"2598","DOI":"10.1182\/blood.V86.7.2598.2598","article-title":"Identification of mutations in 90 of 121 consecutive symptomatic French patients with a type I protein C deficiency. The French INSERM Network on Molecular Abnormalities Responsible for Protein C and Protein S deficiencies","volume":"86","author":"S Gandrille","year":"1995","journal-title":"Blood"},{"issue":"02","key":"ref7","doi-asserted-by":"crossref","first-page":"247","DOI":"10.1160\/TH11-12-0875","article-title":"Deficiencies of antithrombin, protein C and protein S\u2014practical experience in genetic analysis of a large patient cohort","volume":"108","author":"M Caspers","year":"2012","journal-title":"Thromb Haemost"},{"issue":"09","key":"ref8","doi-asserted-by":"crossref","first-page":"1270","DOI":"10.1055\/s-0040-1714100","article-title":"Genotype-phenotype relationships in a large French cohort of subjects with inherited protein C deficiency","volume":"120","author":"M Alhenc-Gelas","year":"2020","journal-title":"Thromb Haemost"},{"issue":"09","key":"ref9","doi-asserted-by":"crossref","first-page":"1093","DOI":"10.1007\/s00439-017-1809-4","article-title":"Deep intronic mutations and human 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genetically unresolved hemophilia B patients","volume":"21","author":"A Dericquebourg","year":"2023","journal-title":"J Thromb Haemost"},{"issue":"03","key":"ref13","doi-asserted-by":"crossref","first-page":"829","DOI":"10.3324\/haematol.2019.217539","article-title":"Next-generation sequencing and recombinant expression characterized aberrant splicing mechanisms and provided correction strategies in factor VII deficiency","volume":"105","author":"P Ferraresi","year":"2020","journal-title":"Haematologica"},{"issue":"05","key":"ref15","doi-asserted-by":"crossref","first-page":"491","DOI":"10.1038\/ng.806","article-title":"A framework for variation discovery and genotyping using next-generation DNA sequencing data","volume":"43","author":"M A DePristo","year":"2011","journal-title":"Nat Genet"},{"issue":"11","key":"ref16","doi-asserted-by":"crossref","first-page":"1361","DOI":"10.1093\/bioinformatics\/btt172","article-title":"A support vector machine for identification of single-nucleotide polymorphisms from next-generation sequencing data","volume":"29","author":"B D O'Fallon","year":"2013","journal-title":"Bioinformatics"},{"issue":"16","key":"ref17","doi-asserted-by":"crossref","first-page":"2078","DOI":"10.1093\/bioinformatics\/btp352","article-title":"The sequence alignment\/map format and SAMtools","volume":"25","author":"H Li","year":"2009","journal-title":"Bioinformatics"},{"issue":"06","key":"ref18","doi-asserted-by":"crossref","first-page":"905","DOI":"10.1016\/j.jmoldx.2017.07.004","article-title":"Clinical validation of copy number variant detection from targeted next-generation sequencing panels","volume":"19","author":"J Kerkhof","year":"2017","journal-title":"J Mol Diagn"},{"issue":"04","key":"ref19","first-page":"656","article-title":"BLAT\u2014the BLAST-like alignment tool","volume":"12","author":"W J Kent","year":"2002","journal-title":"Genome Res"},{"issue":"02","key":"ref20","doi-asserted-by":"crossref","first-page":"306","DOI":"10.1111\/hae.13687","article-title":"Splicing analysis of 26 F8 nucleotide variations using a minigene assay","volume":"25","author":"Y Jourdy","year":"2019","journal-title":"Haemophilia"},{"issue":"10","key":"ref21","doi-asserted-by":"crossref","first-page":"1146","DOI":"10.1111\/jce.12763","article-title":"New family with catecholaminergic polymorphic ventricular tachycardia linked to the Triadin gene","volume":"26","author":"C Rooryck","year":"2015","journal-title":"J Cardiovasc Electrophysiol"},{"issue":"05","key":"ref22","doi-asserted-by":"crossref","first-page":"405","DOI":"10.1038\/gim.2015.30","article-title":"Standards and guidelines for the interpretation of sequence variants: a joint consensus recommendation of the American College of Medical Genetics and Genomics and the Association for Molecular Pathology","volume":"17","author":"S Richards","year":"2015","journal-title":"Genet Med"},{"issue":"01","key":"ref23","doi-asserted-by":"crossref","first-page":"73","DOI":"10.1186\/s13073-022-01073-3","article-title":"Recommendations for clinical interpretation of variants found in non-coding regions of the genome","volume":"14","author":"J M Ellingford","year":"2022","journal-title":"Genome Med"},{"key":"ref24","doi-asserted-by":"crossref","first-page":"82","DOI":"10.1016\/j.cca.2015.10.027","article-title":"Pitfalls in the detection of gross gene rearrangements using MLPA in Fabry disease","volume":"452","author":"L Ferri","year":"2016","journal-title":"Clin Chim Acta"},{"issue":"08","key":"ref25","doi-asserted-by":"crossref","first-page":"e22575","DOI":"10.1002\/jcla.22575","article-title":"A resolved discrepancy between multiplex PCR and multiplex ligation-dependent probe amplification by targeted next-generation sequencing discloses a novel partial exonic deletion in the Duchenne muscular dystrophy gene","volume":"32","author":"C Liu","year":"2018","journal-title":"J Clin Lab Anal"},{"issue":"04","key":"ref26","doi-asserted-by":"crossref","first-page":"224","DOI":"10.1038\/nrg.2015.25","article-title":"Mechanisms underlying structural variant formation in genomic disorders","volume":"17","author":"C MB Carvalho","year":"2016","journal-title":"Nat Rev Genet"},{"issue":"05","key":"ref27","doi-asserted-by":"crossref","first-page":"559","DOI":"10.1007\/s00439-001-0612-3","article-title":"A novel intronic mutation of the TAZ (G4.5) gene in a patient with Barth syndrome: creation of a 5\u2032 splice donor site with variant GC consensus and elongation of the upstream exon","volume":"109","author":"O Sakamoto","year":"2001","journal-title":"Hum Genet"},{"issue":"05","key":"ref28","doi-asserted-by":"crossref","first-page":"552","DOI":"10.1038\/ejhg.2011.223","article-title":"Splicing analysis of unclassified variants in COL2A1 and COL11A1 identifies deep intronic pathogenic mutations","volume":"20","author":"A J Richards","year":"2012","journal-title":"Eur J Hum Genet"},{"key":"ref29","doi-asserted-by":"crossref","first-page":"860","DOI":"10.1038\/35057062","article-title":"Initial sequencing and analysis of the human genome","volume":"409","author":"E S Lander","year":"2001","journal-title":"Nature"},{"issue":"02","key":"ref30","doi-asserted-by":"crossref","first-page":"135","DOI":"10.1007\/s00439-009-0752-4","article-title":"Transposable elements in disease-associated cryptic exons","volume":"127","author":"I Vorechovsky","year":"2010","journal-title":"Hum Genet"},{"issue":"02","key":"ref31","doi-asserted-by":"crossref","first-page":"221","DOI":"10.1016\/S1097-2765(04)00181-9","article-title":"Minimal conditions for exonization of intronic sequences: 5\u2032 splice site formation in alu exons","volume":"14","author":"R Sorek","year":"2004","journal-title":"Mol Cell"},{"issue":"05","key":"ref32","doi-asserted-by":"crossref","first-page":"847","DOI":"10.1111\/hae.14134","article-title":"Identification of new F8 deep intronic variations in patients with haemophilia A","volume":"26","author":"A Dericquebourg","year":"2020","journal-title":"Haemophilia"},{"issue":"10","key":"ref33","doi-asserted-by":"crossref","first-page":"921","DOI":"10.1038\/s10038-020-0786-y","article-title":"Identification of a deep intronic POLR3A variant causing inclusion of a pseudoexon derived from an Alu element in Pol III-related leukodystrophy","volume":"65","author":"T Hiraide","year":"2020","journal-title":"J Hum Genet"},{"issue":"04","key":"ref34","doi-asserted-by":"crossref","first-page":"313","DOI":"10.4161\/rna.28646","article-title":"Adventures in time and space: splicing efficiency and RNA polymerase II elongation rate","volume":"11","author":"E A Moehle","year":"2014","journal-title":"RNA Biol"},{"issue":"12","key":"ref35","doi-asserted-by":"crossref","first-page":"e14346","DOI":"10.1371\/journal.pone.0014346","article-title":"A rare myelin protein zero (MPZ) variant alters enhancer activity in vitro and in vivo","volume":"5","author":"A Antonellis","year":"2010","journal-title":"PLoS One"},{"issue":"11","key":"ref36","doi-asserted-by":"crossref","first-page":"1467","DOI":"10.1002\/humu.22395","article-title":"Novel FOXF1 deep intronic deletion causes lethal lung developmental disorder, alveolar capillary dysplasia with misalignment of pulmonary veins","volume":"34","author":"P Szafranski","year":"2013","journal-title":"Hum Mutat"},{"issue":"01","key":"ref37","doi-asserted-by":"crossref","first-page":"115","DOI":"10.1186\/s12864-018-4485-4","article-title":"A high throughput screen for active human transposable elements","volume":"19","author":"E M Kvikstad","year":"2018","journal-title":"BMC Genomics"},{"issue":"08","key":"ref38","doi-asserted-by":"crossref","first-page":"1369","DOI":"10.1055\/s-0042-1749345","article-title":"Long-read sequencing identifies the first retrotransposon insertion and resolves structural variants causing antithrombin deficiency","volume":"122","author":"B de la Morena-Barrio","year":"2022","journal-title":"Thromb Haemost"},{"issue":"05","key":"ref39","doi-asserted-by":"crossref","first-page":"387","DOI":"10.1532\/IJH97.06082","article-title":"Polymorphisms in vitamin K-dependent gamma-carboxylation-related genes influence interindividual variability in plasma protein C and protein S activities in the general population","volume":"84","author":"R Kimura","year":"2006","journal-title":"Int J Hematol"},{"issue":"07","key":"ref40","doi-asserted-by":"crossref","first-page":"1321","DOI":"10.1161\/01.ATV.0000132408.13064.09","article-title":"Protein C levels are regulated by a quantitative trait locus on chromosome 16: results from the Genetic Analysis of Idiopathic Thrombophilia (GAIT) Project","volume":"24","author":"A Buil","year":"2004","journal-title":"Arterioscler Thromb Vasc Biol"},{"issue":"07","key":"ref41","first-page":"e254","article-title":"Antithrombin, protein C, and protein S: genome and transcriptome-wide association studies identify 7 novel loci regulating plasma levels","volume":"43","author":"Y Ji","year":"2023","journal-title":"Arterioscler Thromb Vasc Biol"},{"issue":"23","key":"ref42","doi-asserted-by":"crossref","first-page":"5032","DOI":"10.1182\/blood-2010-05-283739","article-title":"Genome-wide association study identifies novel loci for plasma levels of protein C: the ARIC study","volume":"116","author":"W Tang","year":"2010","journal-title":"Blood"},{"issue":"02","key":"ref43","doi-asserted-by":"crossref","first-page":"230","DOI":"10.1111\/j.1365-2141.2011.09025.x","article-title":"Genome wide association study for plasma levels of natural anticoagulant inhibitors and protein C anticoagulant 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