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Poster Networking

Venous Thromboembolism

PB0949 - Whole PROC gene sequencing to explain genetically unresolved protein C deficiency.

Monday, June 24, 2024
13:45 – 14:45 ICT
Room: Exhibition Hall

    Co-Author(s)

  • LM

    Laetitia Mauge

    PharmD PhD
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges-Pompidou, F-75015, Université Paris Cité, Inserm, PARCC, F-75015 Paris, France, UR 4609 Hémostase et thrombose, Université Claude Bernard Lyon 1, Lyon, France
    Paris, Ile-de-France, France

  • CR

    Carla Rial

    PharmD
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges-Pompidou, F-75015, Université Paris Cité, Inserm, PARCC, F-75015 Paris, France
    Paris, Ile-de-France, France

  • PD

    Philippe De Mazancourt

    MD, PhD
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges Pompidou, F-75015, Laboratoire de biologie moléculaire, AP-HP, Hôpital Ambroise Paré, F-92100, Université Paris Saclay, Inserm, UMR 1179, F-92100 Boulogne-Billancourt, France
    Paris, Ile-de-France, France

  • DH

    Dominique Helley

    MD, PhD
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges-Pompidou, F-75015, Université Paris Cité, Inserm, PARCC, F-75015 Paris, France
    Paris, Ile-de-France, France

  • IP

    Isabelle Presot

    Laboratory technician
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges-Pompidou, F-75015
    Paris, Ile-de-France, France

  • CB

    Clarisse Billon

    PharmD
    Département de Génétique, Centre de Référence des Maladies Vasculaires Rares, AP HP, Hôpital Européen Georges Pompidou, F 75015 Paris, France
    Paris, Ile-de-France, France

  • SL

    Sylvie Labrouche-Colomer

    PhD
    Laboratoire d’Hémostase moléculaire, Groupe hospitalier Pellegrin, CHU de Bordeaux, 33000 Bordeaux, France
    Bordeaux, Aquitaine, France

  • AV

    Annabelle Venisse

    Laboratory technician
    Département de Génétique, Centre de Référence des Maladies Vasculaires Rares, AP HP, Hôpital Européen Georges Pompidou, F 75015 Paris, France
    Paris, Ile-de-France, France

  • Yesim G. Dargaud, MD,PhD

    Professor of Medicine
    UR 4609 Hemostase et Thrombose, Universite Lyon 1
    Lyon, Rhone-Alpes, France

  • KA

    Karine Auribault

    Engineer
    Plateforme de Biologie Moléculaire, AP HP, Hôpital Européen Georges Pompidou, F 75015 Paris, France
    Paris, Ile-de-France, France

  • SB

    Sebastien Bertil

    Engineer
    Service d’Hématologie biologique, AP HP, Hôpital Européen Georges Pompidou, F 75015 Paris, France
    Paris, Ile-de-France, France

  • CV

    Christine Vinciguerra

    PharmD PhD
    Service d’hématologie Biologique, Groupe Hospitalier Est, Hospices Civils de Lyon, Bron, France, Université Claude Bernard Lyon 1, UR 4609 Hémostase et thrombose, Lyon, France
    Lyon, Rhone-Alpes, France

  • SG

    Sophie Gandrille

    PharmD PhD
    Service d’Hématologie biologique, AP-HP, Hôpital Européen Georges-Pompidou, F-75015, Université Paris Cité, Inserm, F-75015 Paris, France
    Paris, Ile-de-France, France

    • Presenting Author(s)

  • YJ

    Yohann Jourdy, PharmD, phD

    PharmD PhD
    Hospices civils de Lyon, laboratoire d'hématologie biologique, Lyon, France
    Lyon, Rhone-Alpes, France

Background: With current molecular diagnosis, about 10% of protein C deficiencies remain genetically unresolved. In these cases, deep intronic variation or structural variation could be causal.

Aims: To identify the causal variation in unrelated patients presenting a protein C deficiency in whom no genetic variation was found using conventional genetic exon-focused approaches.

Methods: The whole PROC (NM_000312) was sequenced using Next Generation Sequencing capture method. All candidate variations were confirmed using Sanger sequencing. Breakpoint junctions of structural variants were characterized after specific polymerase chain reactions. Variant pathogenicity was determined using the American College of Medical Genetics and Genomics guidelines. The putative splicing impact of intronic variants of undetermined significance was studied using in silico analysis (MaxEntScan and Splice AI) and minigene assay.

Results: Among the 2248 cases of confirmed decreased protein C clot activity explored in our centers since 2000, 38 remained unexplained after conventional genetic exploration. Next generation sequencing data revealed 9 candidate variants in 11 patients (29%): seven deep intronic substitutions and two structural variants. No variant was found in the other 27 patients. The structural variants were a 1 298-kb inversion with a breakpoint in intron 8 of PROC identified in two patients, and a 541-bp deletion (c.797-382_955del). Both variants disrupted PROC and are classified as pathogenic. Among the seven deep intronic substitutions, four (c.237+75G>A, c.535+936C>T (n=2), c.536-95G>A and c.796+49C>T) showed an aberrant pattern of splicing with a retention of 43-93 bp intronic regions in the transcripts observed using minigene assay. These variants were thus classified as likely pathogenic. No impact on the splicing was observed in vitro for the three other intronic substitutions. Finally, variants causing protein C deficiency were identified in 8/38 patients (21%).

Conclusion(s): This study highlights the usefulness of whole PROC gene sequencing to improve the diagnosis of unexplained protein C deficiency.