PB0176 - Gene expression profile in severe Hemophilia A patients developing anti-Factor VIII neutralizing alloantibodies

Sunday, July 10, 2022

6:30 PM – 7:30 PM

Location: Exhibition

Presentation Format: In-Person

Presenting Author(s)

Andrea Cairo

Biologist
Fondazione IRCCS Ca’Granda Ospedale Maggiore Policlinico, Angelo Bianchi Bonomi Hemophilia and Thrombosis Center
Milan, Lombardia, Italy

Co-Author(s)

Silvia Spena, PhD, board certified specialist in Medical Genetics

Angelo Bianchi Bonomi Haemophilia and Thrombosis Center, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, and Luigi Villa Foundation, Milan, Italy
Milano, Lombardia, Italy
EP

Emanuela Pappalardo

Lab Technician
Università degli Studi di Milano, Department of Pathophysiology and Transplantation and Fondazione Luigi Villa, Milan, Italy
Milan, Lombardia, Italy
MM

Mimosa Mortarino

Dr
Angelo Bianchi Bonomi Hemophilia and Thrombosis Center, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico and Fondazione Luigi Villa, Milan
Milan, Lombardia, Italy
IG

Isabella Garagiola

Dr
Angelo Bianchi Bonomi Hemophilia and Thrombosis Center, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico and Fondazione Luigi Villa, Milan
Milan, Lombardia, Italy
Flora Peyvandi, MD, PhD

Professor of Internal Medicine; Director of the Angelo Bianchi Bonomi Hemophilia & Thrombosis Center
Angelo Bianchi Bonomi Hemophilia and Thrombosis Center, Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, Italy; Department of Pathophysiology and Transplantation, Università degli Studi di Milano, Milan, Italy
Milan, Lombardia, Italy

Background: Approximately 30% of severe hemophilia A (HA) patients develop anti FactorVIII (FVIII) neutralizing alloantibodies in response to replacement therapy that decrease the efficacy of the treatment. Understanding the etiology of inhibitor development and the biological mechanisms involved is need.

Aims: To investigate differentially expressed genes in severe HA patients who developed anti-FVIII neutralizing alloantibodies after exposure to FVIII.

Methods: Twenty-nine severe HA patients from the SIPPET study (15 with and 14 without alloantibodies) were analyzed. All patients had a FVIII:C < 1% and were treated with recombinant FVIII products. RNA was collected after alloantibodies development and extracted using PAXgene Blood RNA Kit. Libraries were prepared with TruSeq Stranded Total RNA Library Prep Kit and sequenced on Illumina NovaSeq. Reads were aligned with STAR and counted with RSEM; DESeq2 and edgeR were used to perform differential expression analysis. Validation of identified genes was conducted by qRT-PCR (TaqMan assays). Pre-ranked pathway analyses were performed with GSEA on Hallmark dataset starting from DESeq2 and edgeR results.

Results: Three genes (OTOF, TNFAIP6 and IGLC7) were downregulated in HA patients with inhibitor (Table1). Technical validation confirmed a reduced expression of OTOF and TNAFIP6, whereas IGLC7 validation has not completed yet. TNFAIP6 plays an anti-inflammatory role and can be secreted after proinflammatory stimuli and it has already been reported to be upregulated in the spleen of HA mice in response to FVIII infusion. OTOF is an interferon inducible gene reported to be associated with systemic lupus erythematosus. IGLC7 product is an immunoglobulin involved in the activation of immune system. GSEA analyses revealed a downregulation of interferon gamma/alpha response pathways in patients with alloantibodies in both approaches used.

Conclusion(s): We identified three downregulated genes related to inflammatory response in severe HA patients with alloantibodies suggesting that inflammation induced by treatment could have a role in inhibitor’s development.