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Updated: Jul 11, 2026
Generation of Escape Variants of Neutralizing Influenza Virus Monoclonal Antibodies
Published on: August 29, 2017
Human monoclonal antibody combination against SARS coronavirus: synergy and coverage of escape mutants
Jan ter Meulen1, Edward N van den Brink, Leo L M Poon
1Crucell Holland B.V., Leiden, Netherlands. j.termeulen@crucell.com
Background:
Experimental animal data show that protection against severe acute respiratory syndrome coronavirus (SARS-CoV) infection with human monoclonal antibodies (mAbs) is feasible. For an effective immune prophylaxis in humans, broad coverage of different strains of SARS-CoV and control of potential neutralization escape variants will be required. Combinations of virus-neutralizing, noncompeting mAbs may have these properties.
Methods And Findings:
Human mAb CR3014 has been shown to completely prevent lung pathology and abolish pharyngeal shedding of SARS-CoV in infected ferrets. We generated in vitro SARS-CoV variants escaping neutralization by CR3014, which all had a single P462L mutation in the glycoprotein spike (S) of the escape virus. In vitro experiments confirmed that binding of CR3014 to a recombinant S fragment (amino acid residues 318-510) harboring this mutation was abolished. We therefore screened an antibody-phage library derived from blood of a convalescent SARS patient for antibodies complementary to CR3014. A novel mAb, CR3022, was identified that neutralized CR3014 escape viruses, did not compete with CR3014 for binding to recombinant S1 fragments, and bound to S1 fragments derived from the civet cat SARS-CoV-like strain SZ3. No escape variants could be generated with CR3022. The mixture of both mAbs showed neutralization of SARS-CoV in a synergistic fashion by recognizing different epitopes on the receptor-binding domain. Dose reduction indices of 4.5 and 20.5 were observed for CR3014 and CR3022, respectively, at 100% neutralization. Because enhancement of SARS-CoV infection by subneutralizing antibody concentrations is of concern, we show here that anti-SARS-CoV antibodies do not convert the abortive infection of primary human macrophages by SARS-CoV into a productive one.
Conclusions:
The combination of two noncompeting human mAbs CR3014 and CR3022 potentially controls immune escape and extends the breadth of protection. At the same time, synergy between CR3014 and CR3022 may allow for a lower total antibody dose to be administered for passive immune prophylaxis of SARS-CoV infection.
Insights
Two human monoclonal antibodies (mAbs), CR3014 and CR3022, offer synergistic protection against SARS-CoV infection. This combination may control immune escape and allow for lower therapeutic antibody doses in passive immune prophylaxis.
Area of Science:
- Virology
- Immunology
- Drug Discovery
Background:
- Experimental data suggest human monoclonal antibodies (mAbs) can protect against SARS-CoV infection.
- Broad coverage against SARS-CoV strains and control of escape variants are crucial for effective human immune prophylaxis.
- Combinations of non-competing, virus-neutralizing mAbs may offer enhanced protection.
Purpose of the Study:
- To investigate the potential of combining two human mAbs, CR3014 and CR3022, for SARS-CoV immune prophylaxis.
- To assess if the antibody combination can overcome neutralization escape variants and provide synergistic protection.
- To evaluate the safety of anti-SARS-CoV antibodies in human macrophages.
Main Methods:
- Generated and characterized SARS-CoV variants that escape neutralization by mAb CR3014.
- Screened an antibody-phage library to identify a complementary mAb, CR3022.
- Assessed the binding epitopes and neutralization capabilities of individual mAbs and their combination.
- Investigated the effect of anti-SARS-CoV antibodies on SARS-CoV infection in human macrophages.
Main Results:
- mAb CR3014 neutralized SARS-CoV in ferrets, preventing lung pathology and shedding.
- CR3014 escape variants possessed a P462L mutation in the spike protein, abolishing CR3014 binding.
- mAb CR3022 neutralized CR3014 escape variants and bound to different epitopes, including those from civet cat SARS-CoV.
- No escape variants were generated with CR3022.
- The combination of CR3014 and CR3022 demonstrated synergistic neutralization of SARS-CoV.
- Subneutralizing antibody concentrations did not enhance SARS-CoV infection in human macrophages.
Conclusions:
- The combination of non-competing human mAbs CR3014 and CR3022 offers a strategy to control immune escape and broaden protection against SARS-CoV.
- Synergy between CR3014 and CR3022 may enable a reduced total antibody dose for passive immune prophylaxis.
- The findings support the development of antibody cocktails for SARS-CoV prevention and treatment.
