The antigen-binding domain of a human IgG-anti-F(ab′)<sub>2</sub> autoantibody

Welschof, Martin; Terness, Peter; Kipriyanov, Sergey M.; Stanescu, Diana E.; Breitling, F.; Dörsam, Heinz; Dübel, Stefan; Little, Melvyn; Opelz, Gerhard

doi:10.1073/pnas.94.5.1902

Cited by 99 publications

(50 citation statements)

References 56 publications

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“…Mutation analysis performed with our phage Abs revealed that 40 -50% of the V H and V L genes coding for the Abs in both patients showed evidence of somatic hypermutation, supporting the idea that a significant part of the anti-␣ IIb ␤ 3 response developed in AITP or GT patients is driven by Ag. Our results confirm recent experiments where phage display was successfully used to isolate Abs from individuals with demonstrable serum Ab responses to a variety of Ags, including infectious agents such as HIV-1 (17), mutated protein in malignancy (18), and self Ags in autoimmune diseases (12)(13)(14)(15)(16)19). Isolated Abs have been shown to reflect the in vivo Ab response, and, even if theoretically, the combinatorial approach implies the possibility of pairings of L and H chains not present in the original source tissue.…”

Section: Discussionsupporting

confidence: 87%

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Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Jacobin

Laroche‐Traineau

Little

et al. 2002

The Journal of Immunology

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Previous studies of the immune response in polytransfused Glanzmann thrombasthenia (GT) patients and in autoimmune thrombocytopenic purpura (AITP) have relied on serum analysis and have shown the frequent development of Abs directed against the αIIbβ3 integrin. However, little is known about the molecular diversity of the humoral immune response to αIIbβ3 due to the paucity of mAbs issuing from these pathologies. We have isolated human IgG anti-αIIbβ3 binding fragments using combinatorial libraries of single-chain IgG created from the B cells of a GT and an AITP patient, both with serum Abs. Ab screening was performed using activated platelets or activated αIIbβ3-expressing Chinese hamster ovary cells. Sequencing of selected phage Abs showed that a broad selection of genes from virtually all V gene families had been used, indicating the diversity of the immune response. About one-half of the VH and VL segments of our IgG anti-αIIbβ3 fragments displayed extensive hypermutations in the complementarity-determining region, supporting the idea that an Ag-driven immune response was occurring in both patients. The H chain complementarity-determining region 3 analysis of phage Abs revealed motifs other than the well-known RGD and KQAGDV integrin-binding sequences. To our knowledge, our study is the first to illustrate multiple human IgG anti-αIIbβ3 reactivities and structural variations linked to the anti-platelet human immune response. Human αIIbβ3 Abs preferentially directed against the activated form of the integrin were further characterized because platelet αIIbβ3 inhibitors are potential therapeutic reagents for treating acute coronary syndromes. Currently available αIIbβ3 antagonists do not specifically recognize the activated form of the integrin.

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Section: Discussionsupporting

confidence: 87%

“…Combinatorial libraries from immune donors have been often used to determine the nature of the humoral immune response, for example, in patients with autoimmune or neoplastic diseases or with viral infection (12)(13)(14)(15)(16)(17)(18)(19). Nucleotide sequences of variable regions of such Abs may help us to understand the origin of the secreting B cells.…”

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confidence: 99%

Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Jacobin

Laroche‐Traineau

Little

et al. 2002

The Journal of Immunology

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“…The preferential autoantibody binding of IgG3 to peptides oriented with free C-termini differed from a previous and detailed epitope analysis (54). That work showed that two recombinant, human, single-chain Fvs with anti-hinge specificity were bound to multiple synthetic double-chain peptides (and variants) containing the core C-P-P-C hinge sequence with a free N terminus corresponding to threonine 223 in the upper IgG1 hinge (54,55). Nevertheless, when tested on actual fragments of IgG, those same two scFvs exhibited primarily anti-F(abЈ) 2 reactivity with little or no anti-Fc binding (in accord with our own findings with serum IgG3) (55).…”

Section: Discussionmentioning

confidence: 49%

“…That work showed that two recombinant, human, single-chain Fvs with anti-hinge specificity were bound to multiple synthetic double-chain peptides (and variants) containing the core C-P-P-C hinge sequence with a free N terminus corresponding to threonine 223 in the upper IgG1 hinge (54,55). Nevertheless, when tested on actual fragments of IgG, those same two scFvs exhibited primarily anti-F(abЈ) 2 reactivity with little or no anti-Fc binding (in accord with our own findings with serum IgG3) (55). The authors concluded through additional peptide binding approaches that human serum IgG and IgA anti-hinge autoantibodies most likely bound to conformational epitopes in the hinge (26,34).…”

Section: Discussionmentioning

confidence: 99%

Human Anti-IgG1 Hinge Autoantibodies Reconstitute the Effector Functions of Proteolytically Inactivated IgGs

Brezski¹,

Luongo²,

Petrone³

et al. 2008

The Journal of Immunology

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A number of proteases of potential importance to human physiology possess the ability to selectively degrade and inactivate Igs. Proteolytic cleavage within and near the hinge domain of human IgG1 yielded products including Fab and F(ab′)2 possessing full Ag binding capability but absent several functions needed for immune destruction of cellular pathogens. In parallel experiments, we showed that the same proteolytically generated Fabs and F(ab′)2s become self-Ags that were widely recognized by autoantibodies in the human population. Binding analyses using various Fab and F(ab′)2, as well as single-chain peptide analogues, indicated that the autoantibodies targeted the newly exposed sequences where proteases cleave the hinge. The point of cleavage may be less of a determinant for autoantibody binding than the exposure of an otherwise cryptic stretch of hinge sequence. It was noted that the autoantibodies possessed an unusually high proportion of the IgG3 isotype in contrast to Abs induced against foreign immunogens in the same human subjects. In light of the recognized potency of IgG3 effector mechanisms, we adopted a functional approach to determine whether human anti-hinge (HAH) autoantibodies could reconstitute the (missing) Fc region effector functions to Fab and F(ab′)2. Indeed, in in vitro cellular assays, purified HAH autoantibodies restored effector functions to F(ab′)2 in both Ab-dependent cellular cytotoxicity and complement-dependent cytotoxicity assays. The results indicate that HAH autoantibodies selectively bind to proteolytically cleaved IgGs and can thereby provide a surrogate Fc domain to reconstitute cell lytic functions.

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“…(1) using the PCR primer set described by Welschof et al (2). The library, with a calculated complexity of 2 x 107 independent clones, was constructed in pSEX81 ( Fig.…”

Section: Bodymentioning

confidence: 99%

EGFR-Specific Single-Chain Antibody-Taxol Conjugate for Radiosensitizing Breast Carcinomas

Raisch¹

2001

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The antigen-binding domain of a human IgG-anti-F(ab′)₂ autoantibody

Abstract: Recent studies revealed an immunoregulatory role of natural IgG-anti-F(ab) 2

Cited by 99 publications

References 56 publications

Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Human Anti-IgG1 Hinge Autoantibodies Reconstitute the Effector Functions of Proteolytically Inactivated IgGs

EGFR-Specific Single-Chain Antibody-Taxol Conjugate for Radiosensitizing Breast Carcinomas

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The antigen-binding domain of a human IgG-anti-F(ab′)2 autoantibody

Abstract: Recent studies revealed an immunoregulatory role of natural IgG-anti-F(ab) 2

Cited by 99 publications

References 56 publications

Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Human IgG Monoclonal Anti-αIIbβ3-Binding Fragments Derived from Immunized Donors Using Phage Display

Human Anti-IgG1 Hinge Autoantibodies Reconstitute the Effector Functions of Proteolytically Inactivated IgGs

EGFR-Specific Single-Chain Antibody-Taxol Conjugate for Radiosensitizing Breast Carcinomas

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The antigen-binding domain of a human IgG-anti-F(ab′)₂ autoantibody