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Abundant human anti-Gal?3Gal antibodies display broad pathogen reactivity.

ABSTRACT: Antibodies of the IgG class to terminal Gal?3Gal (IgG anti-?Gal) is abundant in human plasma and are reported to bind most sepsis-causing Gram-negative bacteria. However, these seminal findings, made more than two decades ago, have not been reexamined. Our aim was to assess IgG anti-?Gal´s pathogen reactivity. We affinity purified IgG anti-?Gal from a therapeutic grade normal human IgG pool applying two rounds of positive selection with Gal?3Gal-coupled beads and included removal of column matrix reactive antibodies. The purified antibodies were rigorously characterized in terms of specificity and purity in various solid-phase immunoassays. We used flow cytometry to study reactivity against 100 consecutive clinical isolates diagnosed as cause of sepsis in humans. We found that the purified IgG anti-?Gal displays high specificity for Gal?3Gal. Also, IgG anti-?Gal at 5?mg/L bound 56 out of 100 pathogens with predilection for Gram-positive bacteria binding 39 out of 52 strains. We confirm that although IgG anti-?Gal comprise a small fraction of the human antibody pool (~0.1%), these antibodies targets an impressively large part of pathogens causing invasive disease.

SUBMITTER: Bernth Jensen JM

PROVIDER: S-EPMC7067764 | biostudies-literature | 2020 Mar

REPOSITORIES: biostudies-literature

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Abundant human anti-Galα3Gal antibodies display broad pathogen reactivity.

Bernth Jensen Jens Magnus JM Petersen Mikkel Steen MS Ellerman-Eriksen Svend S Møller Bjarne Kuno BK Jensenius Jens Christian JC Sørensen Uffe B Skov UBS Thiel Steffen S

Scientific reports 20200312 1

Antibodies of the IgG class to terminal Galα3Gal (IgG anti-αGal) is abundant in human plasma and are reported to bind most sepsis-causing Gram-negative bacteria. However, these seminal findings, made more than two decades ago, have not been reexamined. Our aim was to assess IgG anti-αGal´s pathogen reactivity. We affinity purified IgG anti-αGal from a therapeutic grade normal human IgG pool applying two rounds of positive selection with Galα3Gal-coupled beads and included removal of column matri ...[more]

PMID: 32165720

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Project description:BackgroundAnti-drug antibodies (ADAs) to bococizumab were detected in > 40% of subjects in the SPIRE lipid-lowering trials. The risk of cross-reactivity between anti-bococizumab antibodies and other approved anti-proprotein convertase subtilisin/kexin type-9 (PCSK9) monoclonal antibodies (mAbs) was investigated using a single-assay approach.MethodsBococizumab immunogenicity was assessed in SPIRE-HR, a 52-week study. The highest ADA titer sample from each ADA-positive subject (n = 155) was tested in vitro for cross-reactivity to alirocumab and evolocumab using a novel ADA assay approach. Additional specificity tiers within the bococizumab ADA assay against each drug were validated using recombinant PCSK9 as a surrogate cross-reactive positive control. If the highest ADA titer sample showed cross-reactivity, additional samples from that subject were analyzed. Cross-reactivity was determined by the ability of alirocumab or evolocumab to inhibit the sample signal greater than or equal to the cross-reactivity cut-points.ResultsADAs were detected in 44.0% (155/352) of bococizumab-treated subjects, and 27.0% also developed neutralizing antibodies (NAbs). Median ADA and NAb titers ranged from 276 to 526 and 8 to 12 over the course of the study, respectively. From 155 ADA-positive subjects tested for cross-reactivity, one (0.6%) subject showed weak cross-reactivity to both alirocumab and evolocumab. This cross-reactivity signal was transient (from Days 337 to 373) and undetectable at the last ADA-positive timepoint (Day 407).ConclusionA novel, single-assay approach was validated to assess the potential cross-reactivity of anti-bococizumab antibodies to alirocumab and evolocumab. In subjects who developed ADAs to bococizumab, the likelihood of clinically relevant cross-reactivity to marketed anti-PCSK9 mAbs is remote, based on the low frequency of cross-reactivity observed, which was weak in signal inhibition and transient in nature.Clinical trial registrationThe SPIRE-HR study is registered on ClinicalTrials.gov under the identifier NCT01968954.

Project description:Immunization of monkeys with the 82-kDa rhoptry protein (p82) of Plasmodium falciparum can protect them against a lethal blood stage challenge, and monoclonal antibodies to p82 inhibit parasite growth in vitro. The role that a p82-specific immune response might play in human immunity to the parasite is not known. To determine to what extent humans produce antibodies to p82 following infection with P. falciparum, sera from individuals believed to be hyperimmune, semi-immune, or never infected with the parasite were examined. Portions of the p82 gene were expressed separately as fusion proteins and used on immunoblots to test for antibodies to the recombinant proteins. All but 1 of the 30 immune sera possessed antibodies to p82, while nonimmune sera produced, at best, only a marginal signal to the fusion proteins. The signal intensity produced with the human immune sera depended on the region of p82 being assayed, with the N-terminal 37% of p82 producing stronger signals than more C-terminal parts of p82. This N-terminal domain contains a tandem octapeptide repeat (consensus KSSSPSXT/V) of the structure (repeat)2-Q-T-S-G-S/L-(repeat)3. It is shown here that the sequence of this repetitive motif is conserved among four parasite isolates at both the nucleotide and amino acid levels; the five-residue repeat interruption peptide QTSGS/L separating the two sets of repeats contains the only amino acid substitution (Ser or Leu) detected in this region to date. Despite their conservation of structure, the repeats do not appear to be the only epitope recognized by the human antibodies, since sera which recognize the N-terminal fusion protein containing the repeats also bind a related protein after truncation and removal of the repeats. These results indicate that the structurally conserved p82 molecule contains multiple B-cell epitopes and is likely to be immunogenic in most individuals during natural infections with P. falciparum. These observations are consistent with the idea that antibodies to p82 generated during parasite infection have a role in the development of immunity to the organism.

Dataset Information

Abundant human anti-Gal?3Gal antibodies display broad pathogen reactivity.

Publications

Abundant human anti-Galα3Gal antibodies display broad pathogen reactivity.

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