In-silico mediated rapid process development of an orthogonally selective downstream polishing sequence for the purification of monoclonal antibodies from product-related impurities

This work focuses on the process development of membrane-assisted solvent extraction of hydrophobic compounds such as monoterpenes. Beginning with the choice of suitable solvents, quantum chemical calculations with the simulation tool COSMO-RS were carried out to predict the partition coefficient (log P ) of (S)-(+)-carvone and terpinen-4-ol in various solvent–water systems and validated afterwards with experimental data. COSMO-RS results show good prediction accuracy for non-polar solvents such as n-hexane, ethyl acetate and n-heptane even in the presence of salts and glycerol in an aqueous medium. Based on the high log P value, n-heptane was chosen for the extraction of (S)-(+)-carvone in a lab-scale hollow-fibre membrane contactor. Two operation modes are investigated where experimental and theoretical mass transfer values, based on their related partition coefficients, were compared. In addition, the process is evaluated in terms of extraction efficiency and overall product recovery, and its biotechnological application potential is discussed. Our work demonstrates that the combination of in silico prediction by COSMO-RS with membrane-assisted extraction is a promising approach for the recovery of hydrophobic compounds from aqueous solutions.

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Potentiating Antigen-Specific Antibody Production with Peptides Obtained from In Silico Screening for High-Affinity against MHC-II

Molecules ◽

10.3390/molecules24162949 ◽

2019 ◽

Vol 24 (16) ◽

pp. 2949

Author(s):

Yoshiro Hanyu ◽

Yuto Komeiji ◽

Mieko Kato

Keyword(s):

Monoclonal Antibodies ◽

Binding Affinity ◽

Antibody Production ◽

Specific Antibody ◽

In Silico ◽

Igg Antibody ◽

Mhc Ii ◽

High Affinity ◽

Specific Antibody Production

Monoclonal antibodies with high affinity and specificity are essential for research and clinical purposes, yet remain difficult to produce. Agretope peptides that can potentiate antigen-specific antibody production have been reported recently. Here, we screened in silico for peptides with higher affinity against the agretope binding pocket in the MHC-II. The screening was based on the 3D crystal structure of a complex between MHC-II and a 14-mer peptide consisting of ovalbumin residues 323–339. Using this 14-mer peptide as template, we constructed a library of candidate peptides and screened for those that bound tightly to MHC-II. Peptide sequences that exhibited a higher binding affinity than the original ovalbumin peptide were identified. The peptide with the highest binding affinity was synthesized and its ability to boost antigen-specific antibody production in vivo and in vitro was assessed. In both cases, antigen-specific IgG antibody production was potentiated. Monoclonal antibodies were established by in vitro immunization using this peptide as immunostimulant, confirming the usefulness of such screened peptides for monoclonal antibody production.

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A high-throughput 2D-analytical technique to obtain single protein parameters from complex cell lysates for in silico process development of ion exchange chromatography

Journal of Chromatography A ◽

10.1016/j.chroma.2013.09.043 ◽

2013 ◽

Vol 1318 ◽

pp. 84-91 ◽

Cited By ~ 7

Author(s):

Frieder Kröner ◽

Dennis Elsäßer ◽

Jürgen Hubbuch

Keyword(s):

Ion Exchange ◽

High Throughput ◽

In Silico ◽

Process Development ◽

Ion Exchange Chromatography ◽

Exchange Chromatography ◽

Complex Cell ◽

Analytical Technique ◽

Cell Lysates ◽

Single Protein

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Regulatory Approved Monoclonal Antibodies Contain Framework Mutations Predicted From Human Antibody Repertoires

Frontiers in Immunology ◽

10.3389/fimmu.2021.728694 ◽

2021 ◽

Vol 12 ◽

Author(s):

Brian M. Petersen ◽

Sophia A. Ulmer ◽

Emily R. Rhodes ◽

Matias F. Gutierrez-Gonzalez ◽

Brandon J. Dekosky ◽

...

Keyword(s):

Monoclonal Antibodies ◽

High Frequency ◽

In Silico ◽

Sequence Data ◽

Antibody Repertoire ◽

Human Antibody ◽

Therapeutic Antibodies ◽

T Cell Epitopes ◽

Antibody Repertoires ◽

Dynamics Simulations

Monoclonal antibodies (mAbs) are an important class of therapeutics used to treat cancer, inflammation, and infectious diseases. Identifying highly developable mAb sequences in silico could greatly reduce the time and cost required for therapeutic mAb development. Here, we present position-specific scoring matrices (PSSMs) for antibody framework mutations developed using baseline human antibody repertoire sequences. Our analysis shows that human antibody repertoire-based PSSMs are consistent across individuals and demonstrate high correlations between related germlines. We show that mutations in existing therapeutic antibodies can be accurately predicted solely from baseline human antibody sequence data. We find that mAbs developed using humanized mice had more human-like FR mutations than mAbs originally developed by hybridoma technology. A quantitative assessment of entire framework regions of therapeutic antibodies revealed that there may be potential for improving the properties of existing therapeutic antibodies by incorporating additional mutations of high frequency in baseline human antibody repertoires. In addition, high frequency mutations in baseline human antibody repertoires were predicted in silico to reduce immunogenicity in therapeutic mAbs due to the removal of T cell epitopes. Several therapeutic mAbs were identified to have common, universally high-scoring framework mutations, and molecular dynamics simulations revealed the mechanistic basis for the evolutionary selection of these mutations. Our results suggest that baseline human antibody repertoires may be useful as predictive tools to guide mAb development in the future.

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Molecular rationale for SARS-CoV-2 spike circulating mutations able to escape bamlanivimab and etesevimab monoclonal antibodies

Scientific Reports ◽

10.1038/s41598-021-99827-3 ◽

2021 ◽

Vol 11 (1) ◽

Author(s):

Erik Laurini ◽

Domenico Marson ◽

Suzana Aulic ◽

Alice Fermeglia ◽

Sabrina Pricl

Keyword(s):

Monoclonal Antibodies ◽

In Silico ◽

Structural Information ◽

Viral Evolution ◽

Protein A ◽

Immune Surveillance ◽

Protein S ◽

Spike Protein ◽

Wild Type ◽

Experimental Findings

AbstractThe purpose of this work is to provide an in silico molecular rationale of the role eventually played by currently circulating mutations in the receptor binding domain of the SARS-CoV-2 spike protein (S-RBDCoV‑2) in evading the immune surveillance effects elicited by the two Eli Lilly LY-CoV555/bamlanivimab and LY-CoV016/etesevimab monoclonal antibodies. The main findings from this study show that, compared to the wild-type SARS-CoV-2 spike protein, mutations E484A/G/K/Q/R/V, Q493K/L/R, S494A/P/R, L452R and F490S are predicted to be markedly resistant to neutralization by LY-CoV555, while mutations K417E/N/T, D420A/G/N, N460I/K/S/T, T415P, and Y489C/S are predicted to confer LY-CoV016 escaping advantage to the viral protein. A challenge of our global in silico results against relevant experimental data resulted in an overall 90% agreement. Thus, the results presented provide a molecular-based rationale for all relative experimental findings, constitute a fast and reliable tool for identifying and prioritizing all present and newly reported circulating spike SARS-CoV-2 variants with respect to antibody neutralization, and yield substantial structural information for the development of next-generation vaccines and monoclonal antibodies more resilient to viral evolution.

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