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Effects of hapten density on the induced antibody repertoire.

ABSTRACT: Small peptides and oligosaccharides are important antigens for the development of vaccines and the production of monoclonal antibodies. Because of their small size, peptides and oligosaccharides are non-immunogenic on their own and typically must be conjugated to a larger carrier protein to elicit an immune response. Selection of a suitable carrier protein, conjugation method, and hapten density are critical for generating an optimal immune response. We used a glycan array to compare the repertoire of antibodies induced after immunizing with either low or high-density conjugates of the tumor-associated Tn antigen. At high hapten density, a broader range of antibodies was induced, and reactivity to the clustered Tn antigen was observed. In contrast, antibodies induced by the low-density conjugate had narrower reactivity and did not bind the clustered Tn antigen.

SUBMITTER: Li Q

PROVIDER: S-EPMC3462448 | biostudies-literature | 2010 Aug

REPOSITORIES: biostudies-literature

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Effects of hapten density on the induced antibody repertoire.

Li Qian Q Rodriguez Luis G LG Farnsworth David F DF Gildersleeve Jeffrey C JC

Chembiochem : a European journal of chemical biology 20100801 12

Small peptides and oligosaccharides are important antigens for the development of vaccines and the production of monoclonal antibodies. Because of their small size, peptides and oligosaccharides are non-immunogenic on their own and typically must be conjugated to a larger carrier protein to elicit an immune response. Selection of a suitable carrier protein, conjugation method, and hapten density are critical for generating an optimal immune response. We used a glycan array to compare the reperto ...[more]

PMID: 20602400

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Project description:Immunotherapy is becoming increasingly important in the fight against cancers, using and manipulating the body's immune response to treat tumors. Understanding the immune repertoire-the collection of immunological proteins-of treated and untreated cells is possible at the genomic, but technically difficult at the protein level. Standard protein databases do not include the highly divergent sequences of somatic rearranged immunoglobulin genes, and may lead to miss identifications in a mass spectrometry search. We introduce a novel proteogenomic approach, AbScan, to identify these highly variable antibody peptides, by developing a customized antibody database construction method using RNA-seq reads aligned to immunoglobulin (Ig) genes.AbScan starts by filtering transcript (RNA-seq) reads that match the template for Ig genes. The retained reads are used to construct a repertoire graph using the "split" de Bruijn graph: a graph structure that improves on the standard de Bruijn graph to capture the high diversity of Ig genes in a compact manner. AbScan corrects for sequencing errors, and converts the graph to a format suitable for searching with MS/MS search tools. We used AbScan to create an antibody database from 90 RNA-seq colorectal tumor samples. Next, we used proteogenomic analysis to search MS/MS spectra of matched colorectal samples from the Clinical Proteomic Tumor Analysis Consortium (CPTAC) against the AbScan generated database. AbScan identified 1,940 distinct antibody peptides. Correlating with previously identified Single Amino-Acid Variants (SAAVs) in the tumor samples, we identified 163 pairs (antibody peptide, SAAV) with significant cooccurrence pattern in the 90 samples. The presence of coexpressed antibody and mutated peptides was correlated with survival time of the individuals. Our results suggest that AbScan (https://github.com/csw407/AbScan.git) is an effective tool for a proteomic exploration of the immune response in cancers.

Project description:Vaccines against drugs of abuse have induced antibodies in animals that blocked the biological effects of the drug by sequestering the drug in the blood and preventing it from crossing the blood-brain barrier. Drugs of abuse are too small to induce antibodies and, therefore, require conjugation of drug hapten analogs to a carrier protein. The efficacy of these conjugate vaccines depends on several factors including hapten design, coupling strategy, hapten density, carrier protein selection, and vaccine adjuvant. Previously, we have shown that 1 (MorHap), a heroin/morphine hapten, conjugated to tetanus toxoid (TT) and mixed with liposomes containing monophosphoryl lipid A [L(MPLA)] as adjuvant, partially blocked the antinociceptive effects of heroin in mice. Herein, we extended those findings, demonstrating greatly improved vaccine induced antinociceptive effects up to 3% mean maximal potential effect (%MPE). This was obtained by evaluating the effects of vaccine efficacy of hapten 1 vaccine conjugates with varying hapten densities using two different commonly used carrier proteins, TT and cross-reactive material 197 (CRM197). Immunization of mice with these conjugates mixed with L(MPLA) induced very high anti-1 IgG peak levels of 400-1500 ?g/mL that bound to both heroin and its metabolites, 6-acetylmorphine and morphine. Except for the lowest hapten density for each carrier, the antibody titers and affinity were independent of hapten density. The TT carrier based vaccines induced long-lived inhibition of heroin-induced antinociception that correlated with increasing hapten density. The best formulation contained TT with the highest hapten density of ?30 haptens/TT molecule and induced %MPE of approximately 3% after heroin challenge. In contrast, the best formulation using CRM197 was with intermediate 1 densities (10-15 haptens/CRM197 molecule), but the %MPE was approximately 13%. In addition, the chemical synthesis of 1, the optimization of the conjugation method, and the methods for the accurate quantification of hapten density are described.

Dataset Information

Effects of hapten density on the induced antibody repertoire.

Publications

Effects of hapten density on the induced antibody repertoire.

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