Titrationanalysis: A Tool for High-throughput Analysis of Binding Kinetics Data for Multiple Label-Free Platforms

Li, Kan; Horn, Gillian Q.; Alam, S. Munir; Tomaras, Georgia D.; Dennison, S. Moses

doi:10.1016/j.bpj.2020.11.1701

Cited by 3 publications

(2 citation statements)

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“…The kinetics titration data collected were first preprocessed in NextGenKIT (Carterra) software, including reference subtraction, buffer subtraction and data smoothing. The data were then exported and analyzed using the TitrationAnalysis tool ( https://zenodo.org/record/7998652 ) developed in-house 82 ( https://gatesopenresearch.org/articles/7-107/v1 ). The specific binding time courses for each antibody construct immobilized on different spots were fitted to a 1:1 Langmuir model to derive k a (‘ k on ’, association rate constant), k d (‘ k off ’, dissociation rate constant) and K d (dissociation constant) values.…”

Section: Methodsmentioning

confidence: 99%

A candidate antibody drug for prevention of malaria

Williams,

Guerrero,

Flores-Garcia

et al. 2024

Nat Med

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Over 75% of malaria-attributable deaths occur in children under the age of 5 years. However, the first malaria vaccine recommended by the World Health Organization (WHO) for pediatric use, RTS,S/AS01 (Mosquirix), has modest efficacy. Complementary strategies, including monoclonal antibodies, will be important in efforts to eradicate malaria. Here we characterize the circulating B cell repertoires of 45 RTS,S/AS01 vaccinees and discover monoclonal antibodies for development as potential therapeutics. We generated >28,000 antibody sequences and tested 481 antibodies for binding activity and 125 antibodies for antimalaria activity in vivo. Through these analyses we identified correlations suggesting that sequences in Plasmodium falciparum circumsporozoite protein, the target antigen in RTS,S/AS01, may induce immunodominant antibody responses that limit more protective, but subdominant, responses. Using binding studies, mouse malaria models, biomanufacturing assessments and protein stability assays, we selected AB-000224 and AB-007088 for advancement as a clinical lead and backup. We engineered the variable domains (Fv) of both antibodies to enable low-cost manufacturing at scale for distribution to pediatric populations, in alignment with WHO’s preferred product guidelines. The engineered clone with the optimal manufacturing and drug property profile, MAM01, was advanced into clinical development.

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Section: Methodsmentioning

confidence: 99%

A candidate antibody drug for prevention of malaria

Williams,

Guerrero,

Flores-Garcia

et al. 2024

Nat Med

Self Cite

View full text Add to dashboard Cite

show abstract

“…The kinetics titration data collected were first pre-processed in the NextGenKIT (Carterra) software, including reference subtraction, buffer subtraction and data smoothing. The data were then exported and analyzed using the TitrationAnalysis tool developed in-house [ 41 ]. The specific binding time courses for each antibody construct immobilized on different spots were fitted to a 1:1 Langmuir model to derive k a , k d and K D values.…”

Section: Methodsmentioning

confidence: 99%

High-density binding to Plasmodium falciparum circumsporozoite protein repeats by inhibitory antibody elicited in mouse with human immunoglobulin repertoire

et al. 2022

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Antibodies targeting the human malaria parasite Plasmodium falciparum circumsporozoite protein (PfCSP) can prevent infection and disease. PfCSP contains multiple central repeating NANP motifs; some of the most potent anti-infective antibodies against malaria bind to these repeats. Multiple antibodies can bind the repeating epitopes concurrently by engaging into homotypic Fab-Fab interactions, which results in the ordering of the otherwise largely disordered central repeat into a spiral. Here, we characterize IGHV3-33/IGKV1-5-encoded monoclonal antibody (mAb) 850 elicited by immunization of transgenic mice with human immunoglobulin loci. mAb 850 binds repeating NANP motifs with picomolar affinity, potently inhibits Plasmodium falciparum (Pf) in vitro and, when passively administered in a mouse challenge model, reduces liver burden to a similar extent as some of the most potent anti-PfCSP mAbs yet described. Like other IGHV3-33/IGKV1-5-encoded anti-NANP antibodies, mAb 850 primarily utilizes its HCDR3 and germline-encoded aromatic residues to recognize its core NANP motif. Biophysical and cryo-electron microscopy analyses reveal that up to 19 copies of Fab 850 can bind the PfCSP repeat simultaneously, and extensive homotypic interactions are observed between densely-packed PfCSP-bound Fabs to indirectly improve affinity to the antigen. Together, our study expands on the molecular understanding of repeat-induced homotypic interactions in the B cell response against PfCSP for potently protective mAbs against Pf infection.

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