Methods and compositions for assessing antibody specificities
Inventors
Daugherty, Patrick Sean • Kamath, Kathryn Vinaya Louise • Reifert, Jack Ryan
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-11828762-B2
Publication Date
2023-11-28
Expiration Date
Abstract
The present invention provides compositions and methods that can be used to determine a peptide signature for an antibody repertoire in a sample comprising multiple antibodies. The method can be used to characterize a phenotype in a sample, such as providing a diagnosis, prognosis or theranosis of a medical condition.
Core Innovation
The invention relates to an array that includes an array surface and at least two peptide probes that extend from the array surface, where each peptide probe comprises a binding motif selected from a specified group of motif patterns. The peptide probes are defined by motif selection using the listed motif sequences or patterns such that the array surface is presented with multiple peptide binding motifs in a coordinated format.
The disclosure additionally describes peptide display library co-incubation with a patient sample, enrichment of peptide-antibody complexes, and next-generation sequencing of nucleic acids encoding bound peptides to obtain peptide signatures and epitope repertoires. It further describes QC and bioinformatic processing of NGS epitope repertoire data, including amplicon QC, barcode-based filtering, sequencing and unique-read metrics, and mapping motif and enrichment results for diagnostic scoring.
The disclosure describes construction of disease-specific diagnostic motif panels and computational motif discovery and enrichment analysis, including IMUNE versus MEME motif discovery. Motif enrichment, specificity thresholds, and z-score-based diagnostic scoring rules are used for diagnosis, prognosis, and theranosis, and the same motif-discovery framework is described as applied to multiple infectious and autoimmune diseases with corresponding diagnostic motif panels.
The disclosure also describes a depletion reagent strategy for reducing common antibody specificities, including generating peptide libraries, iterative MACS/FACS selection, quantifying cross-reactive motif overlap between parallel tracks, and validating depletion effectiveness by reduced motif enrichment, diversity reduction, and improved detection of non-depletion motifs.
Claims Coverage
The provided set includes one independent claim on an array with peptide probes and dependent claims that refine diagnostic use, detection format, optional antibody-removal, diagnostic performance thresholds, scaffold specificity, and additional array refinements. Across the identified claims, the inventive scope centers on a motif-defined peptide-probe array and diagnostic workflows that measure peptide-to-antibody binding.
Peptide-probe array with motif-defined peptide probes
An array comprising an array surface and at least two peptide probes extending from the array surface, where each peptide probe comprises a binding motif selected from a specified group of motif patterns.
Trypanosoma cruzi antibody-binding measurement using the motif array
A diagnostic method in which a biological sample containing antibodies is contacted with the peptide-probe array, incubated under conditions sufficient for binding, and binding of the at least two peptide probes to its target antibody is measured.
ELISA-based measuring of peptide-to-antibody binding
In the diagnostic method, the measuring step uses an ELISA assay.
Antibody-removing reagent to remove non-specific antibodies
The diagnostic method further includes contacting the biological sample with at least one antibody-removing reagent so that antibodies binding to array components other than the specified peptide probes are removed before applying the biological sample to the peptide-probe array.
High diagnostic performance thresholds
The diagnostic method is specified to have at least 99% sensitivity and at least 99% specificity.
Escherichia coli eCPX scaffold content in peptide probes
The claimed array includes at least two peptide probes that comprise at least a portion of an Escherichia coli eCPX scaffold.
The inventive features center on a motif-defined peptide-probe array and diagnostic workflows that measure peptide-to-antibody binding, optionally using an ELISA readout and an antibody-removal step, with specified sensitivity and specificity targets and scaffold content in the peptide probes.
Stated Advantages
Computational efficiency of IMUNE compared with MEME motif discovery.
Reduced common antibody specificities and improved detection of non-depletion motifs.
Motif enrichment, diversity reduction, and improved diagnostic scoring based on motif and enrichment mapping.
Improves selectivity of binding measurement by removing antibodies that bind to array components other than the specified peptide probes.
Provides quantified diagnostic performance with at least 99% sensitivity and at least 99% specificity.
Documented Applications
Diagnostic use for Trypanosoma cruzi infection using a peptide-probe array and measurement of peptide-to-antibody binding.
NGS epitope repertoire processing to identify motifs, construct disease-specific motif panels, and support phenotype diagnosis, including Lyme disease and multiple other infectious and autoimmune diseases, based on motif scoring.
Depletion reagent use to reduce common antibody specificities and improve detection of non-depletion motifs in motif discovery and diagnostic contexts.
Multiplex phenotype characterization for diagnosis, prognosis, and theranosis using motif panels and motif enrichment analysis [procedural detail omitted for safety].
Diagnostic motif panels are documented for celiac disease, Chagas (Trypanosoma cruzi), Lyme (Borrelia burgdorferi), acute/chronic Toxoplasma gondii, cysticercosis (Taenia solium), EBV mononucleosis/latent EBV, Zika virus (IgG/IgM), HIV, and Sjogren’s syndrome, including performance metrics [procedural detail omitted for safety].
Depletion reagent effectiveness is documented in the context of enriching peptide–antibody complexes in complex antibody repertoires [procedural detail omitted for safety].
Interested in licensing this patent?