A multi-specific, multi-affinity antibody (Multibody, MB) platform derived from the human apoferritin protomer was used by researchers to demonstrate how combining the avidity and specificity of conventional monoclonal antibodies (MAbs) could expand their neutralization breadth and resilience against viral diversity. This research was recently published in the Science Translational Medicine Journal.
Background
The MB platform combined increased affinity with multi-specificity, or the combination of several antibody fragments that recognize various antigen recognition epitopes and are unaffected by viral changes.
Additionally, they investigated the possibility of the MB platform providing in vivo defense against several low-dose coronaviruses (CoVs) including SARS-CoV-2 variants of concern (VOCs).
The United States Food and Drug Administration (U.S.-FDA) has given the REGEN-COV cocktail approval to treat coronavirus disease 2019 (COVID-19). However, the FDA canceled their authorization following the emergence of the Omicron BA.1.
The FDA canceled the authorizations of mAbs that successfully neutralized Omicron, notably bebtelovimab, with the appearance of Omicron subvariants, BQ.1.1 and XBB.1.
There is an urgent unmet medical need for innovative, more effective mAb therapeutics for SARS-CoV-2 with enhanced potency and broad activity but effective at the decreased therapeutic dose(s), as most FDA-authorized mAb therapies have gradually lost efficacy against the Omicron subvariants.
In order to meet demand on a global scale, SARS-CoV-2 mAbs should also become more reasonably priced. Researchers must therefore devise methods for alternative delivery systems for SARS-CoV-2 mAbs, such as intramuscular or subcutaneous injection.
In this context, using a strategy based on boosting antibody valency could aid in strengthening a mAb’s affinity for a target, thereby lowering its therapeutic dosage, broadening the range of neutralization, and allowing delivery via additional routes.
Numerous techniques for antibody engineering have been developed to take advantage of avidity and improve the functional responses of therapeutic mAbs. GEN3009, INBRX-106, and IGM-8444 are being evaluated in phase I/II clinical trial for their ability to treat hematological and solid malignancies, demonstrating the therapeutic utility and benefits of MB-like platforms.
Regarding this study
The MB platform was created by researchers to boost the SARS-CoV-2 targeting mAbs’ ability to neutralize the virus using the same theory; however, they used the human light-chain apoferritin protomer to promote the oligomerization of antibody fragments.
The main goal of the study was to confirm that the several powerful antibodies in the MB format were effective at combating viral variety.
The effectiveness was assessed utilizing in vitro neutralization assays using genuine SARS-CoV-2 or arboviruses or pseudoviruses (PsV). They looked examined whether an increase in the MB’s ability to neutralize SARS-CoV-2 in vitro translated into better in vivo protection at low doses.
Additionally, they evaluated whether MBs may increase their neutralization breadth to include other CoVs, such as arboviruses, and regain neutralizing potency against all VOCs.
The scientists created a self-assembled, oligomeric molecule with antibody-like biochemical capabilities called tri-specific 298-52-80 MB that is capable of ultrapotent neutralization against SARS-CoV-2 wild-type (WT) and Alpha, Beta, Gamma, Delta, and Omicron BA.1 VOCs.
It contained Fabs generated from three previously discovered, weakly potent mAbs (mAbs 298, 52, and 80), which raised its neutralizing power in comparison to the similar immunoglobulin G (IgG) cocktail, IgG4*, by about 1,000-fold.
Five mutations, namely S228P, F234A, L235A, G237A, and P238S, were added to the matching IgG cocktail with an IgG4 fragment, crystallizable (Fc), to prevent binding to Fc-gamma receptors (FcRs).
Cryogenic electron microscopy (cryo-EM) was employed by the researchers to validate the correct assembly of this MB. They also used biolayer interferometry (BLI) to investigate the molecular specifics of Fab interactions with the receptor-binding domain (RBD) and x-ray crystallography to determine antibody binding.
Results
The IgG4* antibody cocktail was only around 100 times as effective as the tri-specific 298-52-80 MB, which had an IC50 value of 0.0002 g/ml.
In line with other studies, the neutralization power of the IgG or the MB was unaffected by switching the Fc subtype from IgG1 to IgG4*.
Single chain (sc)Fab and scFc genetic fusions had a negligible influence, according to a cryo-EM study of the tri-specific 298-52-80 MB. The MB constructed on the apoferritin split design scaffold nevertheless accepted its intended structural disposition, proving the MB to be a uniform biologic.
When compared to an IgG4* cocktail, MB had a greater in vivo neutralizing potency against the deadly SARS-CoV-2 challenge and did so with 430 times less molar concentration.
Five mAbs—52, 80, 2-36, 11-11, and 10-40—exhibited 100% neutralization breadth at an IC50 of 5 g/ml. Only two mAbs, 11-11 and 10-40, demonstrated neutralizing efficacy against two SARS-CoV-2 VOCs after lowering the IC50 cutoff value to 0.01 g/ml to mimic the potency of REGEN-COV.
On the other hand, mAbs 2-7, 80, and 52 achieved 100% breadth as mono-specific MBs at an IC50 cutoff value of 0.01 g/ml. With the exception of 298 and 2-38, the remaining MBs lost their ability to neutralize Omicron BA.1 but retained an IC50 of less than 0.3 g/ml.
The density of spike (S) proteins on the surface of the virion may be encouraging the MB’s heightened avidity. As a result, while mAbs 80 and 2-7 with mutations in the RBD lost their effectiveness against VOCs, these changes had no effect on the apparent binding affinities and neutralization profiles of these antibody molecules when they were shown as MBs.
Years of study have led to the development of powerful, broadly neutralizing antibody therapies for influenza and human immunodeficiency virus 1 (HIV-1).
On the other hand, all SARS-CoV-2 VOCs, including the recently discovered Omicron subvariants BQ.1.1 and XBB.1, demonstrated strong in vitro neutralization when the tri-specific MB containing the specificities 2-7, 10-40, and 11-11 was used.
More significantly, it increased the range of CoVs to which it could neutralize at neutralization potencies that were compatible with FDA-approved SARS-CoV-2 therapies.
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The ability of MB to tolerate variations in viral sequence might speed up the development of antibodies. It might aid in extending the potency of already-known mAbs with the capacity to block newly-emerging SARS-CoV-2 VOCs without appreciable increases in self-reactivity.
Lack of multi-specificity caused mono-specific 80 MB to lose its ability to neutralize the Omicron BA.5 VOC. In light of the rapid SARS-CoV-2 development, combining several best-in-class antibodies into a multi-specific MB may assist (re)gain more resilient neutralizing potency.
Additionally, by combining numerous specificities into a single molecule, it will be possible to assure that all of the components will be bioavailable during the course of treatment, which previously hampered the efficacy of mAb cocktails.
A tri-specific MB has a distinct advantage over conventional mAbs in remaining resistant to future VOCs compared to mAbs alone because it covers a wider area of the RBD.
However, to establish the permanence of neutralization, ongoing monitoring and screening of new variations will be necessary.
Conclusions
In conclusion, the study findings provided “proof of concept” for the idea that the MB platform may be used to harness avidity and boost the effectiveness and breadth of antibody-based treatments against SARS-CoV-2 and other coronaviruses in vitro and in vivo.
The dose-sparing of mAb-based therapies was made possible by the application of avidity-based enhancements in neutralizing potency.
Even without effector activities, increases in neutralization potency in vivo were sufficient to give protection from a fatal challenge.
According to a key Fed indicator, inflation increased 0.4% in April and 4.7% from a year ago.
Future research should thus examine if MB potency may be further increased through effector functions, either by using wildtype or designed Fc to provide particular activity.
