Article Information

Nakanobu Hayashi^1*, Chikako Abe¹, Jiro Kikuchi³, Momoko Hayashi¹, Sakura Hayashi⁹, Masahiro Ueda², Koyu Suzuki⁴, Masahiko Sugitani^5,6, Hiroaki Taniguchi^7,8, Toru Wake⁹, Yusuke Furukawa^3*

¹GeneTry, Inc., Tokyo, Japan

²Promega K K, Tokyo, Japan

³Division of Stem Cell Regulation, Center for Molecular Medicine, Jichi Medical University, Tochigi, Japan

⁴Department of Pathology, St. Luke’s International Hospital, Tokyo, Japan

⁵Department of Pathology, Nihon University School of Medicine, Tokyo, Japan

⁶Department of Diagnostic Pathology, Ageo Central General Hospital, Saitama, Japan

⁷Keio Cancer Center, Keio University School of Medicine, Tokyo, Japan

⁸Keio University Hospital Clinical and Translational Research Center, Keio University School of Medicine, Tokyo, Japan

⁹Okinawa Prefectural Nanbu Medical Center & Children's Medical Center, Okinawa, Japan

*Corresponding authors: Nakanobu Hayashi, GeneTry, Inc., Tokyo, Japan

Yusuke Furukawa, Division of Stem Cell Regulation, Center for Molecular Medicine, Jichi Medical University, Tochigi, Japan

Received: 26 November 2024; Accepted: 03 December 2024; Published: 2 December 2024

Citation: Nakanobu Hayashi, Chikako Abe, Jiro Kikuchi, Momoko Hayashi, Sakura Hayashi, Masahiro Ueda, Koyu Suzuki, Masahiko Sugitani, Hiroaki Taniguchi, Toru Wake, Yusuke Furukawa. Rapid Screening Methods for Universal Binding Peptide Aptamers Against SARS-CoV-2 Variant Spikes, Including Omicron Variants, and their Application to Diagnostic and Therapeutic Agents. Archives of Clinical and Biomedical Research. 8 (2024): 420-432.

Abstract

Keywords

SARS-CoV-2; Variant; Omicron; Universal; Binding; Peptide aptamers; 7-amino acid random peptide ribosome display library screening system

Article Details

1. Introduction

The Covid-19 pandemic is caused by SARS-COV-2. To date, more than 664 million people have been affected and 6.71 million have died [1-27].

An mRNA vaccine has been developed and has shown remarkable immunopreventive effects.

However, the emergence of SARS-COV-2 variants with increased infectivity, immune evasion, and altered virulence has continued since then in many parts of the world, and pandemics of these variants continue to occur [1-27].

With continued mRNA vaccination, neutralizing antibody titers against the SARS-COV-2 variant could be maintained, but with the emergence of new variants appearing one after another, it becomes difficult to maintain neutralizing antibody titers.

The Omicron variant of the highly infectious SARS-COV-2 mutant, which emerged at the
end of 2021, began as BA.1 and has mutated to BA.2 and BA.5. In January 2023, a BQ.1.1 strain derived from BA.5 and an XBB strain derived from BA.2 have emerged, although BA.5 is still the mainstream strain [1-31].
Furthermore, with the elimination of the zero-corona policy in China beginning in December 2022, it is estimated that there is an explosion of more than 900 million more infections at this point, and it is feared that new SARS-COV-2 variants, especially new Omicron lineages, may emerge in the future. Currently, the majority of cases are of the omicron type, which mainly infects the nasopharynx, but it is feared that the emergence of a new type, such as the delta type, with added lung invasiveness, would be a worldwide catastrophe.

Currently, the Omicron strain-compatible bivalent vaccine is used to address prevention of infection and severe disease. And in addition to the mRNA vaccine, the oral drugs Molnupiravir (Lagevrio, Merck) and nirmatrelvir+ritonavir (Paxlovid, Pfizer) are used for treatment [32-40].

As for antibody drugs, some of them have high therapeutic efficacy, but with the emergence of each new variant, the neutralizing antibody activity against the spike decreases, resulting in a short efficacy period, and it is necessary to constantly develop antibody drugs tailored to the new variant [4,41-54].

Antibody drugs are very effective in the treatment of immunocompromised patients with mild disease and risk factors such as severe cardiac disease, chronic respiratory disease, and obesity.

Antibody drugs are also expected to be effective in reducing the incidence of the disease in those who cannot be expected to acquire sufficient immunity through vaccination and in facilities with many elderly people with low basic physical ability.

Antibody drugs, by their nature as macromolecular biopharmaceuticals, take a long time to develop, and have problems such as difficulties in production and purification and their cost.

In addition, the constant use of antibody drugs for treatment is difficult because of the decreasing efficacy of antibody drugs with each new variant.

In recent years, attention has focused on the development of universal antibodies with neutralizing activity against several variants, but the emergence of new variants has weakened the neutralizing activity [55-70].

The omicron variant in particular is very strongly immuno evasive, and it is assumed that the spike mutant has an altered structure that makes it difficult for neutralizing antibodies to bind to it.

Antibody drugs bind easily to the convex surface of a molecule, but lose their binding ability when a conformational change occurs due to amino acid mutation on the surface of the molecule.

Molecular concavities, grooves, and clefts are well conserved and sometimes have enzyme active centers. However, molecular concavities, grooves, and clefts are places where amino acid mutations are less likely to occur, but also where macromolecular antibodies cannot penetrate and bind.

Therefore, in this study, we attempted in vitro screening of "7-amino acid peptide aptamers" that can bind not only to convex surfaces of molecules but also to concave surfaces, grooves, and clefts.

For screening, the ribosomal display method [71-81], which links genes and protein molecules on a one-to-one basis, was employed, allowing for larger library sizes.

In this ribosome display method [71-81], the stop codon of the gene DNA is omitted and in vitro transcription and translation are performed in a cell-free protein synthesis system to create a protein-mRNA-ribosome complex (PRM complex, ternary complex), followed by affinity selection against the target antigen, and clones that bind to the target antigen are purified.

Affinity selection (panning) (71, 72, 81–84) can be done once a day, and even if panning is repeated six times, rapid screening can be done in about a week's process.

A 7-amino acid random peptide ribosome display library [71-81] was constructed and screened for peptide aptamers that bind to the SARS-CoV-2 wild-type spike RBDs. Among them, we selected peptide aptamers that can bind not only wild-type spike RBDs but also delta variant, omicron variant, BA.1, BA.2, and BA.5 spike RBDs.

The peptide aptamers selected in this screening bind to the common epitope of the spike RBD of SARS-CoV-2 variants, including the omicron variant, and are potential universal binding peptide aptamers for all SARS-CoV-2 variants.

These peptide aptamers are approximately 30 amino acids in length, including a 7 amino acid binding site and a surrounding scaffold sequence. They can be easily obtained by peptide synthesis and purification. In addition, peptide aptamer proteins can be easily synthesized and purified in bacteria such as E. coli. These peptide aptamers can be diagnostic and therapeutic agents.

Recently, in the very early stages of an emerging infectious disease outbreak with pathogenic potential, the identification of the pathogen by Next-generation sequencing is used to identify the pathogen and its component proteins.

While the development of diagnostics can be done quickly with PCR and other genetic diagnostics, the development of diagnostics and therapeutics for pathogen proteins requires the complicated, expensive, and time-consuming process of antibody production.

In such cases, antibody production becomes even more difficult for pathogens that repeatedly mutate, such as the SARS-CoV-2 variant in this case.

In this study, we screened universal binding peptide aptamers against SARS-CoV-2 variants using a 7-amino acid random peptide ribosome display library screening system, demonstrating the utility of this method for screening peptide aptamers as antibody alternatives.

2. Materials and Methods

2.1 The structure of 7-amino acid random peptide ribosome display library (GW019PAL); ( Figure 1)

The 7-amino acid random peptide ribosome display library (GW019PAL) shown in Figure 1 was provided by GeneWorld Inc. The structure is T7 promoter, SD sequence, GS linker, scafold1 sequence (SF1), 7 amino acid random peptide, scafold2 sequence (SF2), GS linker, followed by human fibronectin (2177-2250) lacking stop codon [71-81.

2.2 Immobilization of SARS-CoV-2 spike RBD antigens on Magnetic Beads

SARS-CoV-2 spike RBD; wild type (TAIYO NIPPON SANSO Corporation) was immobilized on NHS-Activated Magnetic Beads (Pierce™ NHS-Activated Magnetic Beads, 88826 [85]). (according to the product manual).

2.3 Affinity selection(Panning) [71,72,81-84]; ( Figure 2)

Protein synthesis was performed by in vitro transcription and translation reactions using an E. coli cell-free protein synthesis system (TAIYO NIPPON SANSO Corporation, Cell-free Musaibou-Kun; N Mini SS, A238-0303) (according to the product manual).

The 7-amino acid random peptide ribosome display library (GW019PAL) is lacking a stop codon, resulting in the formation of a protein (peptide)-ribosome-mRNA complex (PRM complex, ternary complex).

The PRM complex is mixed with the above SARS-CoV-2 wild-type spike RBD (Taiyo Nippon Sanso) immobilized on Magnetic Beads and reacted at 4°C for 1 hour. After the reaction, the Magnetic Beads are washed 10 times with ice-cold PBS, the mRNA is dissociated and purified, and RT-PCR (30 cycles) is performed to produce a purified peptide ribosome display library construct. (This process is called the affinity selection cycle, or Panning).

This affinity selection cycle (Panning) was repeated for 6 cycles to purify peptide aptamers that bind to SARS-CoV-2 spike RBD; wild type (TAIYO NIPPON SANSO Corporation).

2.4 Subcloning into the E. coli expression vector pT7-FLAG-1 (Sigma-Aldrich;P1118) [86,87] and peptide aptamer protein synthesis and purification

The region flanking the peptide aptamer of the 6th panning RT-PCR product peptide ribosome display library construct is PCR amplified with primers containing NotI, XmaI restriction enzyme sites and subcloned into NotI, XmaI restriction site of the E. coli expression vector pT7-FLAG-1.

After sonication and centrifugation of the bacterial resuspension, the supernatant was purified for peptide aptamers (proteins) using an anti-FLAG M2 antibody affinity gel (Sigma-Aldrich; A2220) [88]. ( Refer to the product manual.)

2.5 ELISA [89-102]

SARS-CoV-2 spike RBD protein at a concentration of 1ug/ml; wild type, delta (TAIYO NIPPON SANSO Corporation), BA.1 (Sino Biological,40592-V08H121), BA.2 (AcroBioSystems, SPD-C522g), BA.5 (Sino Biological,40592-V08H131) are immobilized on an ELISA separator plate (Sumitomo Bakelite, MS-8508M).

ELISA was performed by direct method. Plates are immobilized with each of SARS-CoV-2 wild type, delta, BA.1, BA.2, and BA.5 spike RBD proteins, and peptide aptamers after flag-tag purification (cloned proteins purified by panning and protein synthesized) are reacted at 37°C for 30 minutes and then washed.

Further, Anti-DDDDK-tag pAb-HRP-DirecT (MBL,PM020-7) is reacted at 37°C for 30 minutes and washed. After that, SureBlue TMB 1-Component Microwell Peroxidase Substrate (KPL, 5120-0075) is added and the reaction is chromogenized. Then, add a reaction stopper (1N H₂SO₄) to stop the color development, and determine the color by visual inspection.

It is important to select peptide aptamer clones that are only chromogenic with the spike RBD protein, but not almost or completely chromogenic with the control BSA-containing blocking buffer, as a positive ELISA reaction. This will eliminate peptide clones that bind non-specifically to the plate as much as possible.

2.6 Nucleotide Sequence and Amino Acid Sequence Determination of Peptide Aptamers [103-107]

Sequencing primers (for pT7-FLAG-1) and Applied Biosystems BigDye Terminator v3.1 were added to the pT7-FLAG-1 plasmid subcloned with peptide aptamers, and cycle sequencing reactions were performed and analyzed with SeqStudio Genetic Analyzer.

The peptide aptamer sequence was analyzed with the genetic analysis software Genetyx and translated into an amino acid sequence.

Figure 1: The structure of 7-amino acid random peptide ribosome display library (GW019PAL) [71-81].

The structure of the 7-amino acid random peptide ribosome display library (GW019PAL) consists of a 7 promoter, SD sequence, GS linker, scafold1 sequence (SF1),7 amino acid random peptide, scafold2 sequence (SF2), GS linker, human fibronectin lacking stop ( 2177-2250).

Figure 2: Principle for screening peptide aptamers that bind to target antigens from a 7-amino acid random peptide ribosome display library [71,72,81-84].

The Affinity Selection (panning) step consists of the following five steps.

Step 1: The 7-amino acid random peptide ribosome display library (Figure 1) was T7 promoter, SD sequence, GS linker, scafold1 sequence (SF1), 7 amino acid random peptide, scafold2 sequence (SF2), GS linker, followed by human fibronectin (2177-2250) lacking the stop codon.

Step 2: 7-amino acid random peptide ribosome display library constructs are transcribed and translated in an E. coli S-30 cell-free protein synthesis system, then stopped by cooling on ice, and the ribosomal complex is stabilized by increasing magnesium concentration.

Step 3: Perform affinity selection on immobilized target antigens with a random peptide library. Peptide-ribosome-mRNA complexes that bind to the target antigen are purified and selected. In this process, nonspecific peptide-ribosome-mRNA complexes are removed by washing.

Step 4: The bound peptide-ribosome-mRNA complex (PRM complex) is dissociated and the mRNA is extracted.

Step 5: Isolated mRNA is reverse transcribed to cDNA and cDNA is amplified by PCR for 30 cycles. (RT-PCR). The amplified DNA is subjected to the next affinity selection cycle (panning) to further purify the peptide aptamer. This affinity selection cycle (panning) is performed for 6 cycles.

3. Results

A 7-amino acid random peptide ribosome display library was panned against SARS-CoV-2 wild-type spiked RBD protein and 30 clones were picked up.

Insert-check PCR was performed on these peptide aptamer clones, and 25 clones had a peptide aptamer insert of 7 amino acid length.

To introduce the flag tag at the N-terminus into these 25 peptide aptamer clones, PCR was performed using primers with restriction enzymes and These clones were subcloned into the E. coli expression vector pT7-FLAG-1.

The peptide aptamer was then protein synthesized in E. coli, and the E. coli bacteria were crushed and Flag-peptide aptamer was purified on an anti-FLAG M2 antibody affinity gel column.

25 peptide aptamer clones were examined by ELISA for binding to SARS-CoV-2 wild type as well as delta, BA.1, BA.2, and BA.5 spike RBD.

The results showed that 4 peptide aptamer clones showed coloration and binding in ELISA to all of the SARS-CoV-2 wild-type, delta-type, BA.1, BA.2, and BA.5 spike RBDs.

The start time and degree of coloration in ELISA were similar for all four clones.

In this experiment, the ELISA was determined strictly by visual inspection for the blocking protein containing BSA, which was the control. This means that only clones that were very weakly chromogenic or negative for the blocking protein containing BSA were selected.

Table 1 shows the amino acid sequences of the 7-amino acid peptide aptamer clones of these four clones.

These 4 clones are expected to bind to the common epitope of the spike RBD of SARS-CoV-2 wild type, Delta, and Omicron strains BA.1, BA.2, and BA.5.

3 of the 4 clones have two or more Cys residues within the peptide aptamer. The N-terminal side scaffold sequence SF1 of the peptide aptamer contains a Cys residue, which is expected to be a complex higher-order structure between this Cys residue and two or more Cys residues within the peptide aptamer [108-112].
The remaining one of the 4 clones is expected to have a linear structure. However, there is a Tyr residue within this peptide aptamer that is expected to contribute to binding to the target [113-118].

Table 1: Amino acid sequence of the SARS-CoV-2 variant universal binding peptide aptamer clone.

Three of the four clones have two or more Cys residues within the peptide aptamer; the Cys residues are located in the N-terminal scaffold sequence SF1 of the peptide aptamer, which is expected to be a complex higher order structure [108-112]. The remaining one clone is a linear structure. However, Tyr residues are present within this peptide aptamer and are expected to contribute to binding to the target [113-118].

4. Discussion

In this study, we show that a 7-amino acid random peptide ribosome display library screening system can be used to rapidly screen in vitro for peptide aptamers that exhibit universal binding to any of multiple SARS-CoV-2 variants [1-31].

The 7-amino acid random peptide ribosome display library [71-81] is a premade library, so it does not require immunization of animals, as is the case with antibody production.

Furthermore, screening for peptide aptamers can be easily performed in a very short process, just one week.

Specifically, affinity selection (panning) [71,72,81-84] is repeated six times against the target antigen, and binding to the target antigen can be easily determined by visual ELISA.

The spike RBD of SARS-CoV-2 is easily mutated, and if antibody drugs are developed for each spike variant of the spike RBD, further mutations will reduce the neutralizing activity of the developed antibody drugs.

Therefore, it is difficult to use antibody drugs as therapeutic agents.

Developing antibody drugs for each variant would be very complicated and costly, and would not make much sense.

Therefore, the development of a universal antibody is desired. Such antibodies have already been developed [4,41-70], but it is not known whether they have sufficient neutralizing activity against the continuously emerging variants of omicron strains [4,41-70].

While BA.5 has been the dominant strain since the end of 2022, variants such as BQ.1.1, strain derived from BA.5 and XBB strain derived from BA.2 have been emerging [119-125].

In particular, XBB.1.5 has immune escape ability against neutralizing antibodies produced by omicron-type bivalent vaccines [(conventional strain (strain of origin)/Omicron strain BA.1) or (conventional strain (strain of origin)/Omicron strain BA.4-5)]. There are currently no antibody drugs that exhibit effective neutralizing antibody activity against XBB.1.5 [99,121,125-144].

This is because mutations in the Spike molecule change the conformation of the Spike, and the neutralizing activity of the antibody is easily lost. Because antibodies are macromolecules, they are good at recognizing the convex surfaces of molecules, but have difficulty penetrating and binding to the concavities, grooves, and clefts of molecules.

Therefore, we wondered if peptide aptamers could be utilized as antibody-like drugs to replace antibody drugs.

To this purpose, we attempted an in vitro screening of "7-amino acid middle-molecule peptide aptamers" that can bind to molecular convexities as well as concavities, grooves, and clefts.

The 7-amino acid random peptide ribosome display library screening system was used to examine whether universal peptide aptamers that bind to a common epitope of the SARS-CoV-2 variant spike RBD, including variants of the Omicron lineage, could be screened.

Normally, when the target antigen for screening is the SARS-CoV-2 wild strain spiked RBD, the ELISA chromogenic binding reaction is performed with the SARS-CoV-2 wild strain spiked RBD, but at this time, ELISA screening of peptide aptamer clones that can bind to the Omicron lineage delta, BA.1, BA.2, and BA.5 spike RBDs as well as the SARS-CoV-2 wild strain spike RBD will be performed. In this way, we thought it would be possible to screen for universal peptide aptamers that can bind to multiple epitopes commonly present in the Omicron lineage spike RBDs, including the SARS-CoV-2 wild strain spike RBD.

The results of the experiment allowed screening of four peptide aptamers that showed universal binding to SARS-CoV-2 wild type, delta type, Omicron lineage BA.1, BA.2, and BA.5 spike RBD.

It will be necessary in the future to measure the neutralizing activity and binding affinity of these four peptide aptamer clones.

If these peptide aptamers have neutralizing activity, they could be used as a therapeutic alternative to antibody drugs.

In addition, the peptide aptamer is a medium-sized molecule about 30 amino acid length even with the surrounding scaffold sequence. Peptide synthesis and purification and protein synthesis and purification by bacteria are considered easy.

Moreover, multiple peptide aptamers are easily obtained through screening, and peptide aptamers are easily multivalent and multiplexed due to their small molecular weight of about 30 amino acids.
It is also easy to further increase the specificity and affinity of the peptide aptamer to the target antigen by making it multivalent and multiplexed.

Since each peptide aptamer can bind to a different epitope, multivalency and multiplexing may be effective against immune evasion.

In addition, recombination of monovalent, multivalent, or multiplexed peptide aptamers with the Fc region of IgG can easily add effector functions such as ADCC and CDC, and this higher-molecular weight can also extend the half-life in the blood.

The Nucleocapsid protein of SARS-CoV-2 is highly conserved and has been used as a diagnostic marker for antigen testing.

Although detailed data are not presented here, a number of anti-nucleocapsid peptide aptamer clones were also successfully screened using the 7 amino acid random peptide ribosome display library screening method.

In addition, multivalency and multiplexing of these multiple anti-nucleocapsid peptide aptamers will enable the development of more sensitive antigen test reagents that are comparable to the sensitivity of PCR test reagents.

5. Conclusions and Perspectives

In this study, we demonstrated that our 7-amino acid random peptide ribosome display library screening system can be used to rapidly screen for middle-molecule peptide aptamers that can bind to target antigens in a very short period of time (approximately one week).

In addition, multiple peptide aptamers obtained in screening can be multivalent, multiplexed, and recombined with molecules having effector functions at will.

In the present study, taking the SARS-CoV-2 variant as an example, we were able to obtain multiple peptide aptamers that can bind to several common epitopes present in the variant.

This 7-amino acid random peptide ribosome display library screening system is expected to be an effective tool for screening peptide aptamers for research, diagnosis, and therapy targeting various proteins of pathogens through rapid analysis of pathogen structures using next-generation sequencers in the event of new emerging infectious disease outbreaks.

As shown in this study, when an emerging infectious disease pathogen repeatedly makes mutations, such as SARS-CoV-2 variant, the peptide aptamer screening system is useful as a rapid screening method for universal binders to epitopes commonly present in variant strains.

The SARS-CoV-2 Omicron lineage has repeatedly made mutations and Omicron sub-lineages have emerged one after another, resulting in increased infectivity and immune evasion ability, and now the XBB.1.5 sub-lineage is increasing. The 7-amino acid random peptide ribosome display library screening system is a method for screening peptide aptamers as early as within a week, so it is easy to pan directly against the XBB.1.5 spike RBD to obtain peptide aptamers and a rapid way to combat the Omicron lineage.

The 7-amino acid random peptide ribosome display library screening system is a potential alternative to polyclonal and monoclonal antibody production and may be useful in antibody-based research, diagnostics, and therapy.

In particular, the 7-amino acid random peptide ribosome display library screening system we have developed can be applied to the field of cancer research, enabling rapid screening of peptide aptamers that target cancer antigens.

Furthermore, by making peptide aptamers multivalent and multiplexed, or by creating recombinant peptide aptamers with effector molecules that have cell-killing effects, they may be applied as diagnostic and therapeutic tools for various cancer cells.

Peptide aptamers are small, with a molecular weight of about 30 amino acids, and if peptide aptamer recombinants of monomeric, multimeric, and multiplexed forms with an added effector function are not utilized in the form of proteins, but are applied in medicine as mRNA drugs like the SARS-CoV-2 mRNA vaccine by Pfizer and Moderna, Inc., it would eliminate the need for protein synthesis and purification and open the way for the early development of therapeutic drugs.

Recently, Felicity Liew et al. reported that although IgG and IgA antibodies in the blood are maintained after mRNA vaccination, IgA antibodies in the nasal mucosa, which are necessary for protection against viral infection, rapidly decrease is one reason for the difficulty in infection protection [145]. Therefore, multivalent and multiplexed peptide aptamers that universally bind to SARS-CoV-2 variant spikes, including omicron variants, are expected to be used as mRNA agents for nasal delivery to protect against (prevent) infection and for post-infection treatment by neutralizing the virus [146-150].

Author Contributions

Nakanobu Hayashi designed the experiments. Nakanobu Hayashi, Chikako Abe, and Jiro Kikuchi were in charge of the experiments; Masahiro Ueda assisted in the selection and acquisition of reagents necessary for the research; Momoko Hayashi and Sakura Hayashi collected and organized the references for the papers; Koyu Suzuki, Masahiko Sugitani, Hiroaki Taniguchi, Toru Wake discussed and reviewed the experiments and results. Yusuke Furukawa reviewed and supervised the final contents.

Nakanobu Hayashi, Chikako Abe, Jiro Kikuchi and Yusuke Furukawa, these authors contributed equally. All agreed on the content of the paper.

Acknowledgments

We thank Geneworld, Inc. for providing the 7-amino acid random peptide ribosome display library (GW019PAL) for free.

We thank TAIYO NIPPON SANSO Corporation for providing the E. coli cell-free protein synthesis system (Cell-free Musaibou-Kun; N Mini SS, A238-0303) and SARS-CoV-2 wild type and delta type spike RBD proteins for free.

We thank Promega K.K. for free supply of some reagents necessary for this research and experiments.

Financial Support and Sponsorship

All research-related expenses were paid by GeneTry, Inc.

No financial support has been received from any public or private organizations.

Conflict of interest

Nakanobu Hayashi is the founder and shareholder of GeneTry, Inc. The other authors declare that no competing interests exist.

ORCID

Nakanobu Hayashi https://orcid.org/0000-0002-4849-1158

References

1. Karim SSA, Karim QA. Omicron SARS-CoV-2 variant: a new chapter in the COVID-19 pandemic. Lancet 398 (2021): 2126-2128.
2. Zuo F, Abolhassani H, Du L, et al. Heterologous immunization with inactivated vaccine followed by mRNA- booster elicits strong immunity against SARS-CoV-2 Omicron variant. Nat Commun13 (2022): 2670.
3. Cox M, Peacock PP, Harvey WT, et al. SARS-CoV-2 variant evasion of monoclonal antibodies based on in vitro studies. Nat. Rev. Microbiol 21 (2023): 112-124.
4. Wang Q, Li Z, Ho J, et al. Resistance of SARS-CoV-2 omicron subvariant BA.4.6 to antibody neutralisation. Lancet Infect Dis 22 (2022): 1666-1668.
5. Kuhlmann C, Mayer CK, Claassen M, et al. Breakthrough infections with SARS-CoV-2 omicron despite mRNA vaccine booster dose. Lancet 399 (2022): 625-626.
6. Zuo F, Abolhassani H, Du L, et al. Heterologous immunization with inactivated vaccine followed by mRNA booster elicits strong humoral and cellular immune responses against the SARS-CoV-2 Omicron variant. bioRxiv (2022).
7. Garcia-Beltran WF, St Denis KJ, Hoelzemer A, et al. mRNA-based COVID-19 vaccine boosters induce neutralizing immunity against SARS-CoV-2 Omicron variant. Cell 185 (2022): 457-466.e4.
8. Tan CW, Lima BE, Young BE, et al. Comparative neutralisation profile of SARS-CoV-2 omicron subvariants BA.2.75 and BA.5. Lancet Microbe 3 (2022): e898.
9. Hui DS, Hybrid immunity and strategies for COVID-19 vaccination. Lancet Infect Dis 23 (2023): 2-3.
10. Chen LL, Chu AWH, Zhang RRQ, et al. To, Serum neutralisation of the SARS-CoV-2 omicron sublineage BA.2. Lancet Microbe 3 (2022): e404.
11. Chen LL, Chua GT, Lu L, et al. Omicron variant susceptibility to neutralizing antibodies induced in children by natural SARS-CoV-2 infection or COVID-19 vaccine. Emerg. Microbes Infect. 11 (2022): 543-547.
12. Yang SL, The HS, Suah JL, et al. SARS-CoV-2 in Malaysia: A surge of reinfection during the predominantly Omicron period. Lancet Reg Health West Pac 26 (2022): 100572.
13. Leung K, Wu JT. Managing waning vaccine protection against SARS-CoV-2 variants. Lancet 399 (2022): 2-3.
14. Dejnirattisai W, Shaw RH, Supasa P, et al. Reduced neutralisation of SARS-CoV-2 omicron B.1.1.529 variant by post-immunisation serum. Lancet 399 (2022): 234-236.
15. Kirby T. Wolfgang Preiser-co-discoverer of SARS-CoV-2 variants. Lancet Infect Dis 22 (2022): 326.
16. Gruell H, Vanshylla K, Tober-Lau P, et al. Neutralisation sensitivity of the SARS-CoV-2 omicron BA.2.75 sublineage. Lancet Infect Dis 22 (2022): 1422-1423.
17. Arora P, Kempf A, Nehlmeier I, et al. Augmented neutralisation resistance of emerging omicron subvariants BA.2.12.1, BA.4, and BA.5. Lancet Infect Dis 22 (2022): 1117-1118.
18. Madhi SA, Kwatra G, Myers JE, et al. Population Immunity and Covid-19 Severity with Omicron Variant in South Africa N Engl J Med 386 (2022): 1314-1326.
19. Hentzien M, Autran B, Piroth L, et al. A monoclonal antibody stands out against omicron subvariants: a call to action for a wider access to bebtelovimab. Lancet Infect Dis 22 (2022): 1278.
20. Cloete J, Kruger A, Masha M, et al. Paediatric hospitalisations due to COVID-19 during the first SARS-CoV-2 omicron (B.1.1.529) variant wave in South Africa: a multicentre observational study. Lancet Child Adolesc Health 6 (2022): 294-302.
21. Wolter N, Jassat W, Walaza S, et al. Clinical severity of SARS-CoV-2 Omicron BA.4 and BA.5 lineages compared to BA.1 and Delta in South Africa. Nat Commun 13 (2022): 5860.
22. Wolter N, Jassat W, Walaza S, et al. Early assessment of the clinical severity of the SARS-CoV-2 omicron variant in South Africa: a data linkage study. Lancet 399 (2022): 437-446.
23. Chen Z, Denga X, Fang L, et al. Epidemiological characteristics and transmission dynamics of the outbreak caused by the SARS-CoV-2 Omicron variant in Shanghai, China: A descriptive study. The Lancet Regional Health - Western Pacific 29 (2022): 100592.
24. Cheng SMS, Mok CKP, Leung YWY, et al. Neutralizing antibodies against the SARS-CoV-2 Omicron variant BA.1 following homologous and heterologous CoronaVac or BNT162b2 vaccination. Nat. Med 28 (2022): 486-489.
25. Nyberg T, Ferguson NM, Nash SG, et al. Comparative analysis of the risks of hospitalisation and death associated with SARS-CoV-2 omicron (B. 1.1. 529) and delta (B. 1.617. 2) variants in England: a cohort study. Lancet 399 (2022): 1303-1312.
26. Menni C, Valdes AM, Polidori L, et al. Symptom prevalence, duration, and risk of hospital admission in individuals infected with SARS-CoV-2 during periods of omicron and delta variant dominance: a prospective observational study from the ZOE COVID Study. Lancet 399 (2022): 1618-1624.
27. Ettaboina SK, Nakkala K, Laddha KS. A Mini Review on SARS-COVID-19-2 Omicron Variant (B.1.1.529). SciMed J 3 (2021): 399-406.
28. Callaway E. Coronavirus variant XBB.1.5 rises in the United States - is it a global threat? Nature 613 (2023): 222-223.
29. Graham F. Daily briefing: Is subvariant XBB.1.5 a global threat? Nature (2023).
30. The rise of variant XBB.1.5, and more - this week’s best science graphics. Nature (2023). https:/doi.org/10.1038/d41586-023-00044-x.
31. Mahase E. Covid-19: What do we know about XBB.1.5 and should we be worried? BMJ 380 (2023): 153.
32. Wen W, Chen C, Tang J, et al. Efficacy and safety of three new oral antiviral treatment (molnupiravir, fluvoxamine and Paxlovid) for COVID-19: A meta-analysis. Ann Med 54 (2022): 516-523.
33. Saravolatz LD, Depcinski S, Sharma M. Molnupiravir and Nirmatrelvir-Ritonavir: Oral Coronavirus Disease 2019 Antiviral Drugs. Clin Infect Dis 76 (2023): 165-171.
34. Wang L, Berger NA, Davis PB, et al. COVID-19 rebound after Paxlovid and Molnupiravir during January-June 2022. medRxiv (2022).
35. Alteri C, Scutari R, Burastero GJ, et al. A proof-of-concept study on the genomic evolution of Sars-Cov-2 in molnupiravir-treated, paxlovid-treated and drug-naïve patients. Commun Biol 5 (2022): 1376.
36. Alteri C, Fox V, Scutari R, et al. Genomic evolution of SARS-CoV-2 in Molnupiravir-treated patients compared to Paxlovid-treated and drug-naïve patients: A proof-of-concept study. Research Square (2022).
37. Parums PV. Rebound COVID-19 and cessation of antiviral treatment for SARS-CoV-2 with paxlovid and molnupiravir. Med Sci Monit 28 (2022).
38. Kelleni M. Paxlovid and Molnupiravir Approved to Manage COVID-19: A Countdown for SARS CoV-2 Variant Apocalypse (2021).
39. Burki TK. The role of antiviral treatment in the COVID-19 pandemic. Lancet Respir Med 10 (2022): e18.
40. Zheng Q, Ma P, Wang M, et al. Efficacy and safety of Paxlovid for COVID-19: A meta-analysis. J Infect 86 (2023): 66-117.
41. Liu L, Iketani S, Guo Y, et al. Striking antibody evasion manifested by the Omicron variant of SARS-CoV-2. Nature 602 (2022): 676-681.
42. Guo Y, Huang L, Zhang G, et al. A SARS-CoV-2 neutralizing antibody with extensive Spike binding coverage and modified for optimal therapeutic outcomes. Nat. Commun 12 (2021): 2623.
43. Kim C, Ryu D, Lee J, et al. A therapeutic neutralizing antibody targeting receptor binding domain of SARS-CoV-2 spike protein. Nat. Commun 12 (2021): 288.
44. Yuan M, Chen X, Zhu Y, et al. A Bispecific Antibody Targeting RBD and S2 Potently Neutralizes SARS-CoV-2 Omicron and Other Variants of Concern. J Virol 96 (2022): e0077522.
45. Barnes CO, Jette CA, Abernathy ME, et al. SARS-CoV-2 neutralizing antibody structures inform therapeutic strategies. Nature 588 (2020): 682-687.
46. Lu RM, Liang KH, Chiang HL, et al. Broadly neutralizing antibodies against Omicron variants of SARS-CoV-2 derived from mRNA-lipid nanoparticle-immunized mice. bioRxiv, 2022.04.19.488843 (2022).
47. Wang P, Nair MS, Liu L, et al. Antibody resistance of SARS-CoV-2 variants B.1.351 and B.1.1.7. Nature 593 (2021): 130-135.
48. Tortorici MA, Czudnochowski N, Starr TN, et al. Structural basis for broad sarbecovirus neutralization by a human monoclonal antibody. bioRxiv (2021).
49. Lu J, Yin Q, Pei R, et al. Nasal delivery of broadly neutralizing antibodies protects mice from lethal challenge with SARS-CoV-2 delta and omicron variants. Virol Sin (2022).
50. Fu D, Zhang G, Wang Y, et al. Structural basis for SARS-CoV-2 neutralizing antibodies with novel binding epitopes. PLoS Biol 19 (2021): e3001209.
51. Dispinseri, Secchi M, Pirillo MF, et al. Neutralizing antibody responses to SARS-CoV-2 in symptomatic COVID-19 is persistent and critical for survival. Nat Commun 12 (2021): 2670.
52. Jones BE, Brown-Augsburger PL, Corbett KS, et al. The neutralizing antibody, LY-CoV555, protects against SARS-CoV-2 infection in nonhuman primates. Sci Transl Med 13 (2021).
53. Li T, Xue W, Zheng Q, et al. Cross-neutralizing antibodies bind a SARS-CoV-2 cryptic site and resist circulating variants. Nat Commun 12, 5652 (2021).
54. Yamin R, Jones AT, Hoffmann H, et al. Fc-engineered antibody therapeutics with improved anti-SARS-CoV-2 efficacy. Nature 599 (2021): 465-470.
55. Cameroni E, Bowen JE, Rosen LE, et al. Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift. Nature 602 (2021): 664-670.
56. Ng KW, Faulkner N, Finsterbusch K, et al. SARS-CoV-2 S2–targeted vaccination elicits broadly neutralizing antibodies. Sci Transl Med 14 (2022): eabn3715.
57. Chen Y, Zhao X, Zhou H, et al. Broadly neutralizing antibodies to SARS-CoV-2 and other human coronaviruses. Nat Rev Immunol (2022): 1-11.
58. Vangelista L, Secchi M. Prepare for the Future: Dissecting the Spike to Seek Broadly Neutralizing Antibodies and Universal Vaccine for Pandemic Coronaviruses. Front Mol Biosci 7 (2020): 226.
59. Vanshylla K, Fan C, Wunsch M, et al. Discovery of ultrapotent broadly neutralizing antibodies from SARS-CoV-2 elite neutralizers. Cell Host Microbe 30 (2022): 69-82. e10.
60. Schepens B, van Schie L, Nerinckx W, et al. An affinity-enhanced, broadly neutralizing heavy chain–only antibody protects against SARS-CoV-2 infection in animal models. Sci Transl Med 13 (2021): eabi7826.
61. Li W, Chen Y, Prévost J, et al. Structural basis and mode of action for two broadly neutralizing antibodies against SARS-CoV-2 emerging variants of concern. Cell Rep 38 (2022): 110210.
62. Du Y, Shi R, Zhang Y, et al. A broadly neutralizing humanized ACE2-targeting antibody against SARS-CoV-2 variants. Nat Commun 12 (2021): 5000.
63. Onodera T, Kita S, Adachi Y, et al. A SARS-CoV-2 antibody broadly neutralizes SARS-related coronaviruses and variants by coordinated recognition of a virus-vulnerable site. Immunity 54 (2021): 2385-2398.e10.
64. Chi X, Guo Y, Zhang G, et al. Broadly neutralizing antibodies against Omicron-included SARS-CoV-2 variants induced by vaccination. Signal Transduction and Targeted Therapy 7 (2022): 1-11.
65. Wec AZ, Wrapp D, Herbert AS, et al. Broad neutralization of SARS-related viruses by human monoclonal antibodies. Science 369 (2020):731-736.
66. Martinez DR, Schäfer A, Gobeil S, et al. A broadly cross-reactive antibody neutralizes and protects against sarbecovirus challenge in mice. Sci Transl Med 14 (2022): eabj7125.
67. Lee IJ, Sun C, Wu P, et al. A booster dose of Delta × Omicron hybrid mRNA vaccine produced broadly neutralizing antibody against Omicron and other SARS-CoV-2 variants. J Biomed Sci 29 (2022): 49.
68. Wang Y, Ye G, Shi K, et al. Structural basis for SARS-CoV-2 Delta variant recognition of ACE2 receptor and broadly neutralizing antibodies. Nat Commun 13 (2022): 871.
69. He WT, Yuan M, Callaghan S, et al. Broadly neutralizing antibodies to SARS-related viruses can be readily induced in rhesus macaques. Sci Transl Med 14 (2022): eabl9605.
70. Zhou B, Zhou R, Chan JF, et al. An elite broadly neutralizing antibody protects SARS-CoV-2 Omicron variant challenge. bioRxiv (2022).
71. Ministro J, Manuel AM, Goncalves J. Therapeutic Antibody Engineering and Selection Strategies. Adv Biochem Eng Biotechnol 171 (2020): 55-86.
72. Plückthun A, Ribosome display: a perspective. Methods Mol Biol 805 (2012): 3-28.
73. Lipovsek D, Plückthun A. In-vitro protein evolution by ribosome display and mRNA display. J Immunol Methods 290 (2004): 51-67.
74. Yan X, Xu Z. Ribosome-display technology: applications for directed evolution of functional proteins. Drug Discov Today 11 (2006): 911-916.
75. Yang LM, Wang J, Kang L, et al. Construction and analysis of high-complexity ribosome display random peptide libraries. PLoS One 3 (2008): e2092.
76. Yang LM, Wang J, Kang L, et al. Correction: Construction and Analysis of High-Complexity Ribosome Display Random Peptide Libraries. PLoS ONE 7 (2012).
77. Schaffitzel C, Hanes J, Jermutus L, et al. Ribosome display: an in vitro method for selection and evolution of antibodies from libraries. J Immunol Methods 231 (1999): 119-135.
78. Hanes J, Jermutus L, Weber-Bornhauser S, et al. Ribosome display efficiently selects and evolves high-affinity antibodies in vitro from immune libraries. Proc Natl Acad Sci USA 95 (1998): 14130-14135.
79. Lamla T, Erdmann VA. Searching sequence space for high-affinity binding peptides using ribosome display. J Mol Biol 329 (2003): 381-388.
80. Hanes J, Plückthun A. In vitro selection and evolution of functional proteins by using ribosome display. PNAS USA 94 (2022): 4937-42.
81. Rothe A, Hosse RJ, Power BE. Ribosome display for improved biotherapeutic molecules. Expert Opin Biol Ther 6 (2006): 177-187.
82. Kunamneni A, Ogaugwu C, Bradfute S, et al. Ribosome Display Technology: Applications in Disease Diagnosis and Control. Antibodies (Basel) 9 (2020).
83. Yau KYF, Groves MAT, Li S, et al. Selection of hapten-specific single-domain antibodies from a non-immunized llama ribosome display library. J Immunol Methods 281 (2003): 161-175.
84. Hanes J, Schaffitzel C, Knappik A, et al. Picomolar affinity antibodies from a fully synthetic naive library selected and evolved by ribosome display. Nat Biotechnol 18 (2000): 1287-1292.
85. Bowen JE, Park YJ, Stewart C, et al. SARS-CoV-2 spike conformation determines plasma neutralizing activity. bioRxiv (2021).
86. Yagi-Utsumi M, Aoki K, Watanabe H, et al. Desiccation-induced fibrous condensation of CAHS protein from an anhydrobiotic tardigrade. Sci Rep 11 (2021): 21328.
87. Holmes B, Benavides-Serrato A, Saunders JT, et al. mTORC2-mediated direct phosphorylation regulates YAP activity promoting glioblastoma growth and invasive characteristics. Neoplasia 23 (2021): 951-965.
88. Rutkoski T, Huang M, Watson N. EZviewTM red protein A and ANTI-FLAG M2 affinity gels: Immunoprecipitation with enhanced visibility affinity beads (2023).
89. Crowther JR, The ELISA Guidebook (Humana Press, 2009).
90. Aydin S. A short history, principles, and types of ELISA, and our laboratory experience with peptide/protein analyses using ELISA. Peptides 72 (2015): 4-15.
91. Crook NE, Payne CC. Comparison of three methods of ELISA for baculoviruses. J Gen Virol 46 (1980): 29-37.
92. Schmidt SD, Nixon RA, Mathews PM. ELISA Method for Measurement of Amyloid-ß Levels. In Amyloid Proteins: Methods and Protocols Sigurdsson EM, Ed. (Humana Press, 2005), pp: 279-297.
93. Ma LN, Zhang J, Chen HT, et al. An overview on ELISA techniques for FMD. Virol J (2011).
94. Boonham N, Kreuze J, Winter S, et al. Methods in virus diagnostics: from ELISA to next generation sequencing. Virus Res 186 (2014): 20-31.
95. Clark MF, Lister RM, Bar-Joseph M. ELISA techniques” in Methods in Enzymology, (Academic Press, 1986) pp: 742-766.
96. Hamblin C, Barnett IT, Hedger RS. A new enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against foot-and-mouth disease virus. I. Development and method of ELISA. J Immunol Methods 93 (1986): 115-121.
97. Plested JS, Coull PA, Gidney MAJ. ELISA. Methods Mol Med 71 (2003): 243-261.
98. Petroková H, Mašek J, Kuchař M, et al. Targeting Human Thrombus by Liposomes Modified with Anti-Fibrin Protein Binders. Pharmaceutics 11 (2019).
99. Maeda R, Fujita J, Konishi Y, et al. A panel of nanobodies recognizing conserved hidden clefts of all SARS-CoV-2 spike variants including Omicron. Commun Biol 5 (2022): 669.
100. Norman A, Franck C, Christie M, et al. Discovery of Cyclic Peptide Ligands to the SARS-CoV-2 Spike Protein Using mRNA Display. ACS Cent Sci 7 (2021): 1001-1008.
101. Buss H, Chan TP, Sluis KB, et al. Protein carbonyl measurement by a sensitive ELISA method. Free Radic Biol Med 23 (1997): 361-366.
102. Shabani S, Moghadam MF, Gargari SLM. Isolation and characterization of a novel GRP78-specific single-chain variable fragment (scFv) using ribosome display method. Med Oncol 38 (2021): 115.
103. GETTING STARTED GUIDE, SeqStudio Genetic Analyzer (2019) (January 22, 2023).
104. Masciarelli SB, Tharp OM, Sansbury BM. An updated stepped elongation time (STeP-up) Sanger sequencing protocol to economize workflows and increase useability. Research Square (2022).
105. Thomas KM, Pelletier NJ, França CMB. Using metagenomics to detect West Nile virus in mosquitoes collected in Oklahoma. Bios 93 (2023): 139-148.
106. Ono LT, Silva JJ, Soto TS, et al. Fungal communities in Brazilian cassava tubers and food products. Int. J. Food Microbiol 384 (2023): 109909.
107. Kano R, Noguchi H, Hiruma M. A deletion mutation in the amino acid sequence of squalene epoxidase in terbinafine-resistant Trichophyton rubrum. J Infect Chemother 28 (2022): 741-744.
108. Chen G, Tao L, Li Z. Recent advancements in mass spectrometry for higher order structure characterization of protein therapeutics. Drug Discov Today 27 (2022): 196-206.
109. Kaltashov IA, Bobst CE, Pawlowski J, et al. Mass spectrometry-based methods in characterization of the higher order structure of protein therapeutics. J Pharm Biomed Anal 184 (2020): 113169.
110. McKenzie-Coe A, Montes NS, Jones LM. Hydroxyl Radical Protein Footprinting: A Mass Spectrometry-Based Structural Method for Studying the Higher Order Structure of Proteins. Chem Rev 122 (2022): 7532-7561.
111. Jahed Z, Domkam N, Ornowski J, et al. Molecular models of LINC complex assembly at the nuclear envelope. J Cell Sci 134 (2021).
112. Liu XR, Zhang MM, Gross ML. Mass Spectrometry-Based Protein Footprinting for Higher-Order Structure Analysis: Fundamentals and Applications. Chem Rev 120 (2020): 4355-4454.
113. Fellouse FA, Wiesmann C, Sidhu SS. Synthetic antibodies from a four-amino-acid code: a dominant role for tyrosine in antigen recognition. Proc Natl Acad Sci U. S. A. 101 (2004): 12467-12472.
114. Koide S, Sidhu SS. The importance of being tyrosine: lessons in molecular recognition from minimalist synthetic binding proteins. ACS Chem Biol 4 (2009): 325-334.
115. Fellouse FA, Barthelemy PA, Kelley RF, et al. Tyrosine plays a dominant functional role in the paratope of a synthetic antibody derived from a four amino acid code. J Mol Biol 357 (2006): 100-114.
116. Hoschke A, László E, Holló J. A study of the role of tyrosine groups at the active centre of amylolytic enzymes. Carbohydr Res 81 (1980): 157-166.
117. Kiss L, Kóródi I, Nánási P. Study on the role of tyrosine side-chains at the active centre of emulsin β-d-glucosidase. Biochimica et Biophysica Acta (BBA) - Enzymology 662 (1981): 308-311.
118. Eberhardt ES, Wittmayer PK, Templer BM, et al. Contribution of a tyrosine side chain to ribonuclease A catalysis and stability. Protein Sci 5 (1996): 1697-1703.
119. Yue C, Song W, Wang L, et al. Enhanced transmissibility of XBB.1.5 is contributed by both strong ACE2 binding and antibody evasion. bioRxiv (2023).
120. Qu P, Faraone JN, Evans JP, et al. Extraordinary evasion of neutralizing antibody response by Omicron XBB.1.5, CH.1.1 and CA.3.1 variants. bioRxiv (2023).
121. European Centre for Disease Prevention and Control. Implications for the EU/EEA of the spread of the SARSCoV-2 Omicron XBB.1.5 sub-lineage for the EU/EEA – 13 January 2023. ECDC: Stockholm (2023).
122. Uriu K, Itoa J, Zahradnik J, et al. Enhanced transmissibility, infectivity and immune resistance of the SARS-CoV-2 Omicron XBB.1.5 variant. bioRxiv(2023).
123. Velavan TP, Ntoumi F, Kremsner PG, et al. Emergence and geographic dominance of Omicron subvariants XBB/XBB.1.5 and BF.7 – the public health challenges. Int J Infect Dis 159 (2023).
124. Chakraborty AK. The 249RWMD spike protein insertion in Omicron BQ.1 subvariant compensates the 24LPP and 69HV deletions and may cause severe disease than BF.7 and XBB.1 subvariants (2023).
125. Addetia A, Piccoli L, Case JB, et al. Therapeutic and vaccine-induced cross-reactive antibodies with effector function against emerging Omicron variants. bioRxiv (2023).
126. Urano E, Itoh Y, Suzuki T, et al. An engineered ACE2 decoy broadly neutralizes Omicron subvariants and shows therapeutic effect in SARS-CoV-2-infected cynomolgus macaques. bioRxiv (2023).
127. Wang B, et al. Human antibody BD-218 has broad neutralizing activity against concerning variants of SARS-CoV-2. Int J Biol Macromol 227 (2023): 896-902.
128. Witte L, Baharani VA, Schmidt F, et al. Epistasis lowers the genetic barrier to SARS-CoV-2 neutralizing antibody escape. Nat Commun 14 (2023): 302.
129. Vikse EL, Fossum E, Erdal MS, et al. Poor neutralizing antibody responses against SARS-CoV-2 Omicron BQ.1.1 and XBB in Norway in October 2022. bioRxiv (2023).
130. Chang A, Koff JL, Lai L, et al. Low neutralizing activity of AZD7442 against current SARS-CoV-2 Omicron variants in patients with B cell malignancies. Blood Adv (2023).
131. Jiang N, Wang L, Hatta M, et al. Bivalent mRNA vaccine improves antibody-mediated neutralization of many SARS-CoV-2 Omicron lineage variants. bioRxiv (2023).
132. Wang Y, Long Y, Wang F, et al. Characterization of SARS-CoV-2 recombinants and emerging Omicron sublineages. Int J Med Sci 20 (2023): 151-162.
133. Liu H, Wu L, Liu B, et al. Two pan-SARS-CoV-2 nanobodies and their multivalent derivatives effectively prevent Omicron infections in mice. Cell Reports Medicine 4 (2023): 100918.
134. Tada T, Dcosta BM, Minnee J, et al. Vectored Immunoprophylaxis and Treatment of SARS-CoV-2 Infection. bioRxiv (2023).
135. Brandolini M, Gatti G, Grumiro L, et al. Omicron Sub-Lineage BA.5 and Recombinant XBB Evasion from Antibody Neutralisation in BNT162b2 Vaccine Recipients. Microorganisms 11 (2023).
136. Coria LM, Rodriguez JM, Demaria A, et al. A Gamma-adapted recombinant subunit vaccine induces broadly neutralizing antibodies against SARS-CoV-2 variants and protects mice from infection. bioRxiv (2023).
137. Chatterjee S, Bhattacharya M, Nag S, et al. A Detailed Overview of SARS-CoV-2 Omicron: Its Sub-Variants, Mutations and Pathophysiology, Clinical Characteristics, Immunological Landscape, Immune Escape, and Therapies. Viruses 15 (2023).
138. Kotaki R, Moriyama S, Takahashi Y. Humoral immunity for durable control of SARS-CoV-2 and its variants. Inflamm. Regen 43 (2023).
139. Yue C, Song W, Wang L, et al. Enhanced transmissibility of XBB. 1.5 is contributed by both strong ACE2 binding and antibody evasion (preprint) (2023).
140. Carabelli AM, Peacock TP, Thorne LG, et al. SARS-CoV-2 variant biology: immune escape, transmission and fitness. Nat Rev Microbiol (2023): 1-16.
141. Wang Q, Guo Y, Iketani S, et al. Antibody evasion by SARS-CoV-2 Omicron subvariants BA. 2.12. 1, BA. 4 and BA. 5. Nature 608 (2022): 603-608.
142. Wang Q, Iketani S, Li Z, et al. Alarming antibody evasion properties of rising SARS-CoV-2 BQ and XBB subvariants. Cell 186 (2023): 279-286.e8.
143. Arora P, Cossmannc A, Schulz SR, et al. Neutralisation sensitivity of the SARS-CoV-2 XBB.1 lineage. Lancet Infect Dis (2023).
144. Tamura T, Ito J, Uriu K, et al. Virological characteristics of the SARS-CoV-2 XBB variant derived from recombination of two Omicron subvariants. bioRxiv (2022).
145. Liew L, et al. SARS-CoV-2-specific nasal IgA wanes 9 months after hospitalisation with COVID-19 and is not induced by subsequent vaccination. EBioMedicine 87 (2023): 104402.
146. Van Hoecke L, Roose K. How mRNA therapeutics are entering the monoclonal antibody field. J Transl Med 17 (2019): 54.
147. Di Trani CA, Fernandez-Sendin M, Cirella A, et al. Advances in mRNA-based drug discovery in cancer immunotherapy. Expert Opin Drug Discov 17 (2022): 41-53.
148. Sahin U, Karikó K, Türeci Ö. mRNA-based therapeutics—developing a new class of drugs. Nat Rev Drug Discov 13 (2014): 759-780.
149. Chen B, Chen Y, Li J, et al. A Single Dose of Anti-HBsAg Antibody-Encoding mRNA-LNPs Suppressed HBsAg Expression: a Potential Cure of Chronic Hepatitis B Virus Infection. MBio 13 (2022): e0161222.
150. Rybakova Y, Kowalski PS, Huang Y, et al. mRNA Delivery for Therapeutic Anti-HER2 Antibody Expression in Vivo Mol Ther 27 (2019): 1415-1423.