OC 41.3 - Single-cell sequencing of PBMCs from hemophilia A and hemophilia B patients with inhibitors reveals different immune responses to FVIII and FIX after replacement therapy

Monday, June 24, 2024

15:15 – 15:30 ICT

Room: Ballroom B1

Presenting Author(s)

Enhao Li

Ph.D. candidate
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)

Co-Author(s)

JD

Jing Dai

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
Zhenping Chen

Associate professor
Hematologic Diseases Laboratory, Hematology Center, Beijing Pediatric Research Institute, Beijing Children's Hospital, Capital Medical University, National Center for Children’s Health, Beijing, 100045, ChinaChildren’s Health, Beijing, 100045, China
Beijing, Beijing, China (People's Republic)
YX

Yilun Xue

Research Assistant
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
Haoyang Wu

Ph.D. student
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
Zekun Li, PhD

Ph.D.
Department II of Hematology Center, Beijing Children’s Hospital, Capital Medical University, National Center for Children’s Health, Beijing, 100045, China
Beijing, Beijing, China (People's Republic)
CL

Can Lou

Research Assistant
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
Runhui Wu, PhD, MD

Professor
Department of Hematology, Beijing Children’s Hospital, Capital Medical University, National Center for Children’s Health, Beijing, 100045, China
Beijing, Beijing, China (People's Republic)
XW

Xuefeng Wang

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)
WW

Wenman Wu

Professor
Department of Laboratory Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, 200025, China
Shanghai, Shanghai, China (People's Republic)

Background: About 30~40% severe hemophilia A (HA) patients are complicated with FVIII inhibitors after replacement therapy due to alloimmune reactions, while only ~10% hemophilia B (HB) produce alloantibodies against FIX after receiving FIX agents. The mechanism under the apparent difference of immune reactions upon exposure to administered FVIII and FIX has not been fully appreciated.

Aims: We aim to explore different immune responses to FVIII and FIX in HA and HB patients and reveal different mechanisms leading to apparently different clinical manifestations of HA and HB patients with inhibitors.

Methods: We performed single-cell sequencing on PBMCs collected from five HA and five HB patients with inhibitors. After pre-processing the data with “CellRanger”. “Seurat”, “doubletfinder” and “harmony” R packages were applied for quality control and to cluster cell subsets and calculate differentially expressed genes in R 4.2.2 (Fig. 1A). Statistical analyses were performed in R 4.2.2.

Results: After quality control of the data, totally 75,051 cells, including 32,679 cells from HA and 42,372 cells from HB, were clustered to 19 subsets (Fig. 1B) according to the markers of each cluster shown in Fig. 1C. The cell ratio of each subset in each sample was shown in Fig. 1D. Student’s t test on cell ratios of different types of immune cells in lymphocytes between HA and HB groups showed that HB patients with inhibitors had statistically significantly higher T cell ratio, while HA patients with inhibitors had higher B cell ratio (Fig. 1E).

Conclusion(s): Our results support the notion proposed by previous studies suggesting that the immune systems might recognize and initiate alloimmune response to exogenous FVIII and FIX in HA and HB patients through different pathways. The structural difference, tissue distribution and metabolism of intravenously administered FVIII and FIX might be responsible for the apparently different immune reactions underlying inhibitor development in HA and HB patients.