Neutralization of Crimean-Congo hemorrhagic fever pseudotyped virions with heavy chain antibodies

Kewen Qian; Yu Zhang; Zhihao Li; Wei Ye; Yue Cui; Zheng Zhu; Zhengshan Chen; Jianrong Wang; Jin Han; Ping Huang; Pu Fan; Peng Lv; Ting Fang; Guanying Zhang; Changming Yu; Yunzhu Dong; Wujian Li; Fanglin Zhang; Xiangyang Chi

doi:10.1016/j.virs.2025.12.001

December 2025

Citation: Kewen Qian, Yu Zhang, Zhihao Li, Wei Ye, Yue Cui, Zheng Zhu, Zhengshan Chen, Jianrong Wang, Jin Han, Ping Huang, Pu Fan, Peng Lv, Ting Fang, Guanying Zhang, Changming Yu, Yunzhu Dong, Wujian Li, Fanglin Zhang, Xiangyang Chi. Neutralization of Crimean-Congo hemorrhagic fever pseudotyped virions with heavy chain antibodies .VIROLOGICA SINICA, 2025, 40(6) : 1021-1036. http://dx.doi.org/10.1016/j.virs.2025.12.001

Neutralization of Crimean-Congo hemorrhagic fever pseudotyped virions with heavy chain antibodies

Kewen Qian ^a ,
Yu Zhang ^a ,
Zhihao Li ^b,c ,
Wei Ye ^d ,
Yue Cui ^a ,
Zheng Zhu ^a ,
Zhengshan Chen ^a ,
Jianrong Wang ^a ,
Jin Han ^a ,
Ping Huang ^a ,
Pu Fan ^a ,
Peng Lv ^a ,
Ting Fang ^a ,
Guanying Zhang ^a ,
Changming Yu ^a ,
Yunzhu Dong ^{a
,,} ,
Wujian Li ^{b,c
,,} ,
Fanglin Zhang ^{d
,,} ,
Xiangyang Chi ^{a
,,}

a. Academy of Military Medical Sciences, Beijing 100071, China;
b. Division of Life Sciences and Medicine, University of Science and Technology of China, Hefei 230031, China;
c. Key Laboratory of Anhui Province for Emerging and Reemerging Infectious Diseases, Hefei 230026, China;
d. Department of Microbiology, Air Force Medical University:Fourth Military Medical University, Xi'an 710032, China

Corresponding author: Yunzhu Dong, 13811429044@163.com
Wujian Li, lwj636567@163.com
Fanglin Zhang, flzhang@fmmu.edu.cn
Xiangyang Chi, xiangyangchi@163.com
Received Date: 14 September 2025
Accepted Date: 01 December 2025
Available online: 04 December 2025

Abstract

Crimean-Congo hemorrhagic fever virus (CCHFV) is a highly pathogenic tick-borne virus that causes severe hemorrhagic fever with high mortality rates in humans. No licensed vaccines or efficacious antiviral therapies are currently available. Here, we identified seven heavy chain antibodies targeting CCHFV Gc, which consist of heavy-chain variable domain (VHH) fused to human IgG1 Fc region (VHH-Fc). These VHH-Fc antibodies exhibited neutralizing activity against both recombinant vesicular stomatitis virus (VSV)-vectored CCHFV pseudoviruses and CCHFV transcription- and entry-competent virus-like particles (tecVLPs). Among these, N025 achieved the most potent pseudovirus neutralization, while N013 showed remarkable efficacy in tecVLP systems, with IC₅₀ values of 22.7 ng/mL and 33.0 ng/mL, respectively. AlphaFold3 structural predictions revealed that all characterized VHH-Fc antibodies target epitopes within Domain II of the Gc protein, with partial or complete overlap with the fusion loop region. Alanine scanning mutagenesis confirmed the functional significance of these epitopes, with N013 showing the highest binding energy change (△△G = 25.36 kcal/mol) and moderate competition with a known fusion loop-targeting antibody. Sequence conservation analysis across representative CCHFV strains from different genetic lineages demonstrated complete conservation of the N013 and N025 epitopes, suggesting potential for broad-spectrum neutralizing activity. Together, our findings provide a novel strategy for developing CCHFV therapeutics and identify promising antibody candidates that could inform future broad-spectrum antiviral development efforts.
- Crimean-Congo hemorrhagic fever virus (CCHFV)
- , Heavy chain antibodies
- , VHH-Fc antibodies
- , Pseudovirus
- , Virus-like particles (VLPs)

Electronic Supplementary Material

10.1016j.virs.2025.12.001-ESM1.docx
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