# Complement and CR2/CD21 in the Immune Pathogenesis of HIT

> **NIH NIH R01** · DUKE UNIVERSITY · 2021 · $464,233

## Abstract

ABSTRACT
Heparin-induced thrombocytopenia (HIT) is a life-threatening immune-mediated thrombotic disorder caused by
antibodies to PF4/heparin complexes. The immune response is common and yet little is understood about its
occurrence. In the last funding period, we made fundamental observations related to early events surrounding
the host’s encounter with the PF4/heparin antigenic complex. Specifically, we showed: 1) preferential binding
of PF4/heparin to circulating B-cells, compared with other leukocytes or platelets, 2) heparin-dependent
binding of PF4/heparin to B-cells in patients receiving heparin, 3) complement fixation by PF4/heparin antigen,
4) complement-mediated PF4/heparin binding to circulating B-cells in patients receiving heparin, and 5) binding
of complement-coated antigen to B-cells via complement receptor 2 (CR2/ CD21). In the following aims, we will
test the hypothesis that complement activation by PF4/heparin complexes and deposition of antigen on B-cells
via CD21 is essential for development of HIT autoantibodies. Specific Aim 1: Mechanism of complement
activation by PF4/heparin complexes. In preliminary data, we show that mannose-binding lectin and ficolin-2
bind PF4/heparin complexes. Based on these findings, we will test the hypothesis that PF4/heparin complexes
activate complement via the lectin pathway. We will define the structural basis of PF4/heparin-lectin
interactions, study functional interactions of lectins with PF4/heparin complexes, and examine the role of
complement inhibition in HIT. Specific Aim 2: Cellular consequences of CD21 engagement by
PF4/heparin and complement. Binding of complement-coated antigen to CD21 augments humoral immunity
by 103 to 104 fold. We hypothesize that binding of complement-coated multivalent PF4/heparin to the CD21
complex facilitates recruitment and signaling of antigen-specific B-cells. We will examine downstream
signaling, activation and proliferation of cognate and non-cognate B-cells, perform in vivo studies in mice with a
fixed B-cell receptor and characterize the contribution of CD21-expressing follicular dendritic cells to Ab
formation. Specific Aim 3: Examine host predictors of PF4/heparin seroconversion. Complement-coated
antigen can be detected on B-cells in some, but not all, patients receiving heparin therapy. Using a novel C3
capture immunoassay, we also show significant donor to donor variation in healthy donor plasma incubated
with a fixed dose of PF4/heparin. Based on these observations, we will examine host susceptibility to anti-
PF4/heparin seroconversions by characterizing the complement proteome and using antigen-positive B-cells
as a marker for seroconversions in heparinized patients. By defining the cellular pathways that initiate
formation of PF4/heparin Abs, we hope to uncover mechanisms relevant to the immunogenicity of other auto-
or exogenous antigens.

## Key facts

- **NIH application ID:** 10077790
- **Project number:** 5R01HL136512-04
- **Recipient organization:** DUKE UNIVERSITY
- **Principal Investigator:** Gowthami M Arepally
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2021
- **Award amount:** $464,233
- **Award type:** 5
- **Project period:** 2018-01-15 → 2022-02-28

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/10077790

## Citation

> US National Institutes of Health, RePORTER application 10077790, Complement and CR2/CD21 in the Immune Pathogenesis of HIT (5R01HL136512-04). Retrieved via AI Analytics 2026-05-25 from https://api.ai-analytics.org/grant/nih/10077790. Licensed CC0.

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