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Coronavirus disease 2019 (COVID-19) is caused
by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection.
Over the past two years, many research groups have discovered and developed mAbs
, effective neutralizing monoclonal antibodies against SARS-CoV-2.
Monoclonal antibodies are effective treatments for the prevention of viral diseases by neutralizing viral particles, however cumulative variants of strains can reduce the neutralizing efficacy of host antibody responses against new variants, which requires the development of new monoclonal antibodies with potential synergistic effects to help reduce immune escape
from SARS-CoV-2.
Plants are a versatile recombinant protein production system that has made significant contributions
to developing countermeasures to the current coronavirus pandemic.
Plant cell expression of ACE2 has also been explored as a preventive/therapeutic approach
to prevent or treat SARS-CoV-2 infection.
Plants have also proven to be excellent systems
for mAb development and production.
Producing mAbs in plants has the potential to significantly reduce overall costs, allowing for rapid and high-level accumulation of recombinant mAbs
in a short period of time.
Recently, the internationally renowned magazine Plant Biotechnology Journal published an article entitled "A novel plant-made monoclonal antibody enhances the synergetic potency of an antibody cocktail against.
" The SARS-CoV-2 Omicron variant", which studies the expression of raw mAbs resistant to SARSCoV-2 RBD in plants and demonstrates that a monoclonal antibody called 11D7 neutralizes variant strains and shows neutralizing synergies with other therapeutic mAbs in neutralizing Omicron and other variants
.
(1) Expression of anti-SARS-CoV-2 RBD monoclonal antibody in tobacco
Mice were pretreated and immuno-enhanced with protocols, hybridoma was screened for immunogen (Wuhan-Hu-1 RBD) responsiveness and several strongly reactive antibodies were identified, and an 11D7 was selected for variable region gene rescue
。 Mouse light and heavy chain variable regions were transplanted into human κ and human γ constant regions, respectively, resulting in chimeric mAbs that were codons suitable for plant expression and expressed transiently in ΔXFT N.
benthamiana mutants, and 5 days after transgene delivery, plant-made 11D7 (p11D7) reached a peak expression of 131 μg mAb per gram of fresh leaf weight (FLW) (Figure 1).
。 When protein A affinity chromatography is performed on total plant protein extracts containing plant-made p11D7 mAbs, the recombinant mAbs are uniformly purified in a manner similar to mammalian cells purified by the same method expressing
mAbs.
The bands observed from the SDS-PAGE analysis confirmed to be the expected heterotetrameric IgG and its constituent light and heavy chains (Figure 2).
P11D7 was found by mass spectrometry to carry 91.
1% of the double antennae or single antennae N-acetylglucosamine (GnGn, MGn), a typical mammal-like glycan produced in ΔXFT plants (Table 1)
(2) p11D7 binds to RBD of the B.
1.
1.
529 (Omicron) variant
The study assessed the functionality
of the mAb by testing for binding to the antigen of interest, SARS-CoV-2 RBD.
A decrease in affinity for p11D7 was observed after specific binding to Omicron RBD at higher concentrations than the original WA1/2020 strain, but the new p11D7 monoclonal antibody still retained recognition of Omicron RBD, suggesting that it may be able to neutralize emerging variants
.
(Figure 3)
Figure 3
(3) Multiple variants of p11D7 and SARS-CoV-2
After confirming the binding of chimeric p11D7 to its target antigen, the neutralization potency
of the mAb is evaluated.
The continuous emergence of new variants has also prompted testing of p11D7
against the B.
1.
617.
2 (Delta) variant.
Overall neutralization data indicate that the neutralizing capacity of p11D7 decreases with mutation accumulation from variants derived from ancestral strains, but still neutralizes SARS-CoV-2 variants
.
(Figure 4)
Figure 4
(4) Synergistic effect of therapeutic monoclonal antibody and p11D7 in neutralizing Omicron
Competitive binding analysis of well-characterized monoclonal antibodies of all RBD-binding classes confirmed that p11D7 has at least partially overlapping epitopes with CR3022 (class 4 mAbs) on RBD, but not
with CB6 (also known as etesevim ab, class 1 monoclonal antibody).
The two monoclonal antibodies, tixagevimab (also known as COV2-2196, class 1/2 monoclonal antibody) and cilgavimab (COV2-2130, class 3 monoclonal antibody), do not compete with p11D7 in binding to RBD (Figure 5).
。 The unique binding site of p11D7 prompted the study to observe that the p11D7+tixagevimab combination and tixagevimab+cilgavimab combination have a strong neutralization synergy at their respective IC50s (Table 2), and demonstrated that there is a neutralization synergy between p11D7 and tixagevimab and cilgavimab, both of which are double combinations
.
(Table 3)
Figure 5
SARS-CoV-2 has prompted the development of new mAb therapies
using platforms with the potential for optimized utility and safety.
This study describes a novel SARS-CoV-2 neutralizing mAb that was discovered in traditional hybridoma systems and further developed
in plant expression systems.
Studies have shown that p11D7 is not an effective neutralizing mAb
by itself.
But combined with its unique binding site on RBD, it is speculated that p11D7 may neutralize viral particles
by interfering with mechanisms other than ACE2 binding.
Although p11D7 is not an effective neutralizing agent compared to other treatment-developed SARS-CoV-2 monoclonal antibodies, plant cells expressed mAbs in this study with a highly homogeneous (91.
1%) glycan population with no detectable core fucoschase, compared to 81.
9% of hybridoma monoclonal antibodies containing core fucose
.
This suggests that p11D7 may improve potency in vivo through pathways other than neutralizing potency, i.
e.
through Fc-mediated mechanisms such as ADCC, and will be the focus of
future research.
Plant Biotechnology Pbj Exchange Group
In order to help the majority of researchers obtain relevant information more effectively, Plant Biotechnology Pbj has established a WeChat group, Plant
Biotechnology
Journal submissions and literature-related questions, official account release content and public account submission questions will be answered in the group, and academic exchanges and collision thinking
will be encouraged in the group.
In order to ensure a good discussion environment in the group, please add Xiaobian WeChat first, scan the QR code to add, and then we will invite you to join the group
in time.
Tip: When adding Xiaobian WeChat and after joining the group, please be sure to note the school or + name, PI indicates at the end, we will invite you to enter the PI group
.
