OC 78.1 - Study of missense changes involving cysteine residues in F11: implications on folding, dimerization, activity and FXI deficiency

Wednesday, July 13, 2022

10:15 AM – 10:30 AM

Location: ExCel Center, ICC Capital Suite Room 1

Presentation Format: Virtual

Presenting Author(s)

Carlos Bravo Perez, MD, PhD

MD, PhD
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
Murcia, Murcia, Spain

Co-Author(s)

AR

Ana María Rubio

Servicio de Hematología y Oncología Médica
Hospital Universitario Morales Meseguer-Centro Regional de Hemodonación, Universidad de Murcia, IMIB, CIBERER Spain
Murcia, Murcia, Spain
�P

Ángeles Palomo

MD, Servicio de Hematología y Hemoterapia
Centro Materno-Infantil del Hospital Regional Universitario Carlos de Haya, Málaga
Málaga, Andalucia, Spain
JE

Julio Esteban

MD, Servicio de Hematología
Hospital Virgen del Castillo de Yecla, Murcia
Yecla, Murcia, Spain
FB

Frederic Bauduer

MD, Service d'Hématologie
Centre Hospitalier de la Côte Basque, Bayonne
Bayonne, Aquitaine, France
AM

Antonia Miñano

Servicio de Hematología y Oncología Médica
Hospital Universitario Morales Meseguer-Centro Regional de Hemodonación, Universidad de Murcia, IMIB, CIBERER Spain
Murcia, Murcia, Spain
RC

Rosa Cifuentes

Predoctoral fellow
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
Murcia, Murcia, Spain
Belen de la morena-Barrio, PhD

Postdoctoral fellow
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
murcia, Murcia, Spain
Pedro Garrido-Rodríguez

PhD Student
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer-Centro Regional de Hemodonación, Universidad de Murcia, IMIB, CIBERER Spain
Murcia, Murcia, Spain
JR

Juan J Rojo

Servicio de Hematología y Oncología Médica
Hospital Universitario Morales Meseguer-Centro Regional de Hemodonación, Universidad de Murcia, IMIB, CIBERER Spain
Murcia, Murcia, Spain
VV

Vicente Vicente

Researcher and Medical Doctor
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
Murcia, Murcia, Spain
ML

María Luisa Lozano

Leader of Hematology Service in Murcia
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
Murcia, Murcia, Spain
JC

Javier Corral

Senior researcher
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Pascual Parrilla, CIBERER, Murcia, Spain
Murcia, Murcia, Spain
Maria Eugenia de la Morena-Barrio, PhD

Reseacher
Servicio de Hematología y Oncología Médica, Hospital Universitario Morales Meseguer, Centro Regional de Hemodonación, Universidad de Murcia, IMIB-Arrixaca, CIBERER, Murcia, Spain
Murcia, Murcia, Spain

Background: Factor XI (FXI) has 17 intramolecular disulfide bonds, and circulates as a disulfide-linked homodimer. F11 genetic variants affecting cysteines (Cys) have been identified in patients with FXI deficiency.

Aims: To study the impact of F11 variants affecting Cys on folding, dimerization and function of FXI.

Methods: Assessment of F11 missense variants removing/creating Cys, from our cohort of 105 unrelated FXI-deficient patients, and from literature review. Recombinant expression of wild-type FXI and selected variants was conducted in HEK293.

Results: Overall, 40 variants were identified involving Cys (Fig.1).
Twenty-five removed Cys. Only p.Cys599Tyr (a new variant from our cohort), on the catalytic domain, caused qualitative (CRM+) FXI deficiency. Considering cases with quantitative (CRM-) deficiency, mean FXI:C was < 50% (38.0±10.8%) in heterozygotes, suggesting an underlying dominant-negative effect. A CRM- case in our cohort eliminated two Cys in the same allele: p.Cys255Tyr and p.Cys339Phe, the latter implicated in FXI dimerization. Our recombinant model demonstrated that the rp.Cys339Phe precluded dimer formation, but not monomer secretion.
Fifteen missense variants introduced new Cys. Three of them caused CRM+ deficiency. We highlight a new variant from our cohort, p.Phe295Cys, identified in a hemizygous case due to concurrent de novo deletion of the wild-type allele. This variant allowed the secretion of two dimeric forms of FXI, one with an aberrant electrophoretic mobility. The recombinant model confirmed two alternative conformations by differential intramolecular disulfide architecture (Fig.2).

Conclusion(s): Our study confirms the importance of intramolecular disulfide bonds in FXI folding and function. Only one new variant, removing Cys from the catalytic domain, caused CRM+ deficiency. Missense change of p.Cys339, by preventing dimerization, provoked CRM- deficiency in vivo. Other mutations, such as p.Phe295Cys, create new Cys residues, that can result in aberrant dimer formation with impaired activity. Further research on the implications of FXI intramolecular disulfide bond exchanges is needed.

Funding: ISCIII&FEDER:PI21/00137;CM20/00094.