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Bioengineering
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Bioengineering of Biotherapeutics
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Bioengineering of Biotherapeutics

A special issue of Bioengineering (ISSN 2306-5354).

Deadline for manuscript submissions: closed (25 November 2022) | Viewed by 23874

Special Issue Editor

Dr. Hanieh Khalili

E-Mail Website
Guest Editor
School of biomedical science, University of West London, London W5 5RF, UK
Interests: biopharmaceutical; ocular drug delivery; antibody-based medicine; protein conjugation and characterization; binding assays; antibody formulation; pharmacokinetics
Special Issues, Collections and Topics in MDPI journals

Special Issue Information

Dear Colleagues,

Bioengineering is currently soliciting manuscripts for a Special Issue focusing on novel manufacturing methods (recombinant or chemical methods) to develop antibody-based therapeutics. Manuscripts can include literature reviews as well as primary research papers. Topics of particular interest include platforms for discovery/development of antibodies, antibody fragments, bispecific antibodies, and antibody–drug conjugates. Different strategies for manufacturing, characterization, and formulation of biotherapeutic with focus on purity, yield, and binding are particularly encouraged.

Dr. Hanieh Khalili
Guest Editor

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All submissions that pass pre-check are peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Bioengineering is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 2700 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • biotherapeutics
  • antibody fragments
  • bioengineering
  • bispecific antibodies
  • antibody–drug conjugates
  • chemical–recombinant method

Published Papers (5 papers)

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Research

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15 pages, 2326 KiB  
Article
An Integrated In Vivo/In Vitro Protein Production Platform for Site-Specific Antibody Drug Conjugates
by Jeffrey Hanson, Dan Groff, Abi Carlos, Hans Usman, Kevin Fong, Abigail Yu, Stephanie Armstrong, Allison Dwyer, Mary Rose Masikat, Dawei Yuan, Cuong Tran, Tyler Heibeck, James Zawada, Rishard Chen, Trevor Hallam and Gang Yin
Bioengineering 2023, 10(3), 304; https://doi.org/10.3390/bioengineering10030304 - 28 Feb 2023
Cited by 3 | Viewed by 2517
Abstract
The XpressCF+® cell-free protein synthesis system is a robust platform for the production of non-natural amino acids containing antibodies, which enable the site-specific conjugation of homogeneous antibody drug conjugates (ADCs) via click chemistry. Here, we present a robust and scalable means of [...] Read more.
The XpressCF+® cell-free protein synthesis system is a robust platform for the production of non-natural amino acids containing antibodies, which enable the site-specific conjugation of homogeneous antibody drug conjugates (ADCs) via click chemistry. Here, we present a robust and scalable means of achieving a 50–100% increase in IgG titers by combining the high productivity of cell-based protein synthesis with the unique ability of XpressCF+® reactions to produce correctly folded and assembled IgGs containing multiple non-natural amino acids at defined positions. This hybrid technology involves the pre-expression of an IgG light-chain (LC) protein in a conventional recombinant E. coli expression system, engineered to have an oxidizing cytoplasm. The prefabricated LC subunit is then added as a reagent to the cell-free protein synthesis reaction. Prefabricated LC increases IgG titers primarily by reducing the protein synthesis burden per IgG since the cell free translation machinery is only responsible for synthesizing the HC protein. Titer increases were demonstrated in four IgG products in scales ranging from 100-µL microplate reactions to 0.25-L stirred tank bioreactors. Similar titer increases with prefabricated LC were also demonstrated for a bispecific antibody in the scFvFc-FabFc format, demonstrating the generality of this approach. Prefabricated LC also increases robustness in cell-free reactions since it eliminates the need to fine-tune the HC-to-LC plasmid ratio, a critical parameter influencing IgG assembly and quality when the two IgG subunits are co-expressed in a single reaction. ADCs produced using prefabricated LC were shown to be identical to IgGs produced in cell-free alone by comparing product quality, in vitro cell killing, and FcRn receptor binding assays. This approach represents a significant step towards improving IgG titers and the robustness of cell-free protein synthesis reactions by integrating in vivo and in vitro protein production platforms. Full article
(This article belongs to the Special Issue Bioengineering of Biotherapeutics)
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14 pages, 2255 KiB  
Article
Challenges and Strategies for a Thorough Characterization of Antibody Acidic Charge Variants
by Y. Diana Liu, Lance Cadang, Karenna Bol, Xiao Pan, Katherine Tschudi, Mansour Jazayri, Julien Camperi, David Michels, John Stults, Reed J. Harris and Feng Yang
Bioengineering 2022, 9(11), 641; https://doi.org/10.3390/bioengineering9110641 - 03 Nov 2022
Cited by 9 | Viewed by 3534
Abstract
Heterogeneity of therapeutic Monoclonal antibody (mAb) drugs are due to protein variants generated during the manufacturing process. These protein variants can be critical quality attributes (CQAs) depending on their potential impact on drug safety and/or efficacy. To identify CQAs and ensure the drug [...] Read more.
Heterogeneity of therapeutic Monoclonal antibody (mAb) drugs are due to protein variants generated during the manufacturing process. These protein variants can be critical quality attributes (CQAs) depending on their potential impact on drug safety and/or efficacy. To identify CQAs and ensure the drug product qualities, a thorough characterization is required but challenging due to the complex structure of biotherapeutics. Past characterization studies for basic and acidic variants revealed that full characterizations were limited to the basic charge variants, while the quantitative measurements of acidic variants left gaps. Consequently, the characterization and quantitation of acidic variants are more challenging. A case study of a therapeutic mAb1 accounted for two-thirds of the enriched acidic variants in the initial characterization study. This led to additional investigations, closing the quantification gaps of mAb1 acidic variants. This work demonstrates that a well-designed study with the right choices of analytical methods can play a key role in characterization studies. Thus, the updated strategies for more complete antibody charge variant characterization are recommended. Full article
(This article belongs to the Special Issue Bioengineering of Biotherapeutics)
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10 pages, 3073 KiB  
Article
Soluble Papain to Digest Monoclonal Antibodies; Time and Cost-Effective Method to Obtain Fab Fragment
by Matthew Collins and Hanieh Khalili
Bioengineering 2022, 9(5), 209; https://doi.org/10.3390/bioengineering9050209 - 12 May 2022
Cited by 4 | Viewed by 8203
Abstract
Antigen binding fragments (Fabs) used in research (e.g., antibody mimetics, antibody-drug conjugate, bispecific antibodies) are frequently obtained by enzymatic digestion of monoclonal antibodies using immobilised papain. Despite obtaining pure Fab, using immobilised papain to digest IgG has limitations, most notably slow digestion time [...] Read more.
Antigen binding fragments (Fabs) used in research (e.g., antibody mimetics, antibody-drug conjugate, bispecific antibodies) are frequently obtained by enzymatic digestion of monoclonal antibodies using immobilised papain. Despite obtaining pure Fab, using immobilised papain to digest IgG has limitations, most notably slow digestion time (more than 8 h), high cost and limited scalability. Here we report a time and cost-effective method to produce pure, active and stable Fab using soluble papain. Large laboratory scale digestion of an antibody (100 mg) was achieved using soluble papain with a digestion time of 30 min and isolated yields of 55–60%. The obtained Fabs displayed similar binding activity as Fabs prepared via immobilised papain digestion. Site-specific conjugation between Fabs and polyethylene glycol (PEG) was carried out to obtain antibody mimetics FpF (Fab-PEG-Fab) indicating that the native disulphide bond had been preserved. Surface-plasmon resonance (SPR) of prepared FpFs showed that binding activity towards the intended antigen was maintained. We anticipate that this work will provide a fast and less costly method for researchers to produce antibody fragments at large scale from whole IgG suitable for use in research. Full article
(This article belongs to the Special Issue Bioengineering of Biotherapeutics)
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Review

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17 pages, 1346 KiB  
Review
Bioengineering of Antibody Fragments: Challenges and Opportunities
by Sama Pirkalkhoran, Wiktoria Roksana Grabowska, Hamid Heidari Kashkoli, Reihaneh Mirhassani, David Guiliano, Colin Dolphin and Hanieh Khalili
Bioengineering 2023, 10(2), 122; https://doi.org/10.3390/bioengineering10020122 - 17 Jan 2023
Cited by 12 | Viewed by 4792
Abstract
Antibody fragments are used in the clinic as important therapeutic proteins for treatment of indications where better tissue penetration and less immunogenic molecules are needed. Several expression platforms have been employed for the production of these recombinant proteins, from which E. coli and [...] Read more.
Antibody fragments are used in the clinic as important therapeutic proteins for treatment of indications where better tissue penetration and less immunogenic molecules are needed. Several expression platforms have been employed for the production of these recombinant proteins, from which E. coli and CHO cell-based systems have emerged as the most promising hosts for higher expression. Because antibody fragments such as Fabs and scFvs are smaller than traditional antibody structures and do not require specific patterns of glycosylation decoration for therapeutic efficacy, it is possible to express them in systems with reduced post-translational modification capacity and high expression yield, for example, in plant and insect cell-based systems. In this review, we describe different bioengineering technologies along with their opportunities and difficulties to manufacture antibody fragments with consideration of stability, efficacy and safety for humans. There is still potential for a new production technology with a view of being simple, fast and cost-effective while maintaining the stability and efficacy of biotherapeutic fragments. Full article
(This article belongs to the Special Issue Bioengineering of Biotherapeutics)
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14 pages, 1308 KiB  
Review
Engineering CRISPR/Cas13 System against RNA Viruses: From Diagnostics to Therapeutics
by Yi Xue, Zhenzhen Chen, Wenxian Zhang and Jingjing Zhang
Bioengineering 2022, 9(7), 291; https://doi.org/10.3390/bioengineering9070291 - 29 Jun 2022
Cited by 2 | Viewed by 3980
Abstract
Over the past decades, RNA viruses have been threatened people’s health and led to global health emergencies. Significant progress has been made in diagnostic methods and antiviral therapeutics for combating RNA viruses. ELISA and RT-qPCR are reliable methods to detect RNA viruses, but [...] Read more.
Over the past decades, RNA viruses have been threatened people’s health and led to global health emergencies. Significant progress has been made in diagnostic methods and antiviral therapeutics for combating RNA viruses. ELISA and RT-qPCR are reliable methods to detect RNA viruses, but they suffer from time-consuming procedures and limited sensitivities. Vaccines are effective to prevent virus infection and drugs are useful for antiviral treatment, while both need a relatively long research and development cycle. In recent years, CRISPR-based gene editing and modifying tools have been expanded rapidly. In particular, the CRISPR-Cas13 system stands out from the CRISPR-Cas family due to its accurate RNA-targeting ability, which makes it a promising tool for RNA virus diagnosis and therapy. Here, we review the current applications of the CRISPR-Cas13 system against RNA viruses, from diagnostics to therapeutics, and use some medically important RNA viruses such as SARS-CoV-2, dengue virus, and HIV-1 as examples to demonstrate the great potential of the CRISPR-Cas13 system. Full article
(This article belongs to the Special Issue Bioengineering of Biotherapeutics)
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