Adenoviral Inciting Antigen and Somatic Hypermutation in VITT, 2026, Wang et al.

[SNT Gatchaman]

Adenoviral Inciting Antigen and Somatic Hypermutation in VITT

Spoiler: Authors

Jing Jing Wang; Linda Schönborn; Theodore E Warkentin; Luisa Müller; Thomas Thiele; Lena Ulm; Uwe Völker; Sabine Ameling; Sören Franzenburg; Lars Kaderali; Ana Tzvetkova; Alex Colella; Tim Chataway; Chee Wee Tan; Bridie Armour; Alexander Troelnikov; Lucy Rutten; James McCluskey; Roland Zahn; Tom P Gordon; Andreas Greinacher

BACKGROUND
Vaccine-induced immune thrombocytopenia and thrombosis (VITT) is a rare prothrombotic complication that occurs after adenoviral vector–based vaccination against coronavirus disease 2019; in rare cases, it can also occur after natural adenovirus infection. VITT is mediated by platelet-activating antibodies against the highly cationic protein platelet factor 4 (PF4). The underlying inciting antigen trigger and immunopathogenesis remain unknown.

METHODS
We used antibody proteomics to determine the amino acid sequences of anti-PF4 antibodies from 21 patients with VITT and sequenced the genes encoding the immunoglobulin light-chain hypervariable region from 100 patients with VITT. To identify an adenoviral trigger, we used the antigen-binding fingerprints of anti-PF4 and anti–adenovirus protein antibodies to identify a shared serum clonotype and subsequently used adenovirus protein peptides and recombinant anti-PF4 VITT antibodies to map the mimicking linear epitope.

RESULTS
Genomic and proteomic profiling of VITT antibodies revealed a shared immunoglobulin light-chain allele, IGLV3-21*02 or *03, harboring a critical somatic hypermutation, K31E. Only antibodies purified against adenoviral core protein VII (pVII) contained anti-PF4 species matching the VITT fingerprint; antibodies against intact virions or other adenoviral proteins did not. Cross-reactive IgGs were mapped to a basic linear epitope on pVII. A pathogenic anti-PF4 VITT antibody, back-mutated to germline (K31), lost its prothrombotic activity in vitro and in vivo and preferentially bound pVII, a finding that directly supported the role of the hypermutation in the antigenic shift from adenovirus pVII to PF4.

CONCLUSIONS
The results of our study indicate that VITT occurs when, in persons with immunoglobulin light-chain allele IGLV3-21*02 or *03, a specific somatic hypermutation develops that affects antibodies that recognize a specific epitope on the adenoviral core protein pVII, which results in misdirection of antibody targeting toward PF4.

(Funded by Deutsche Forschungsgemeinschaft and others; German Clinical Trials Register number, DRKS00025738; EU Post-Authorization Study Register number, EUPAS45098.)

Web | DOI | PDF | New England Journal of Medicine | Paywall

 

[SNT Gatchaman]

Molecular mimicry between the adenoviral core protein pVII and PF4, combined with somatic hypermutation transforming an anti-pVII immune response to a misdirected anti-PF4 immune response, is a fundamental pathobiologic mechanism of VITT. Immune cross-reactivity between pVII and PF4 results from shared, highly positively charged epitopes with structural similarity in the alpha helixes of both proteins. Nonetheless, production of pathogenic, high-avidity anti-PF4 reactive antibodies requires additional specific genetic features.

The first prerequisite is a genetic predisposition conferred by allele IGLV3-21*02 or *03 in the gene encoding the hypervariable region of the immunoglobulin light chain. This is supported by our finding that the allele was present in 99 of 100 patients with VITT (as compared with the expected background frequency of 20 to 60%, depending on the ethnic group).

The second prerequisite is a somatic hypermutation that changes a positively charged lysine to a negatively charged glutamic acid at position 31 of the light-chain hypervariable region of the anti-pVII antibodies. We confirmed this previously described typical fingerprint of the hypervariable IgG regions in all 21 patients with VITT in our study.

Many healthy persons with the genetic background of IGLV3-21*02 or *03 acquire antiadenovirus antibodies. However, induction of VITT requires B cells specific for the pVII RYARAKSRRRRIARR epitope and an additional, specific somatic hypermutation leading to a negatively charged amino acid at position 31 of the IgG light-chain hypervariable region.

The germline DNA of all 100 patients with VITT in our study encoded lysine at this position, which confirms that this mutation is somatic. This specific hypermutation probably occurs in few B cells, which explains why VITT anti-PF4 antibodies are monoclonal or oligoclonal.

In VITT antibodies, the key mutation at position 31 — from a positively charged lysine residue to a negatively charged glutamic or aspartic acid — results in a striking change in antibody avidity toward PF4 and pathogenicity, as shown in the in vitro and in vivo experiments involving our recombinant antibodies with and without the K31E mutation. This high avidity is the prerequisite for overcoming the positive charge cloud (zeta potential) that usually keeps PF4 molecules apart, thus allowing binding of one IgG molecule to two PF4 molecules, which leads to clustering of PF4 and to the formation of large, pathogenic, platelet-activating PF4–IgG immune complexes.

A novel aspect of this work is that we have used mass spectrometry–based proteomics to define molecular mimicry through clonal matching of anti-PF4 and anti-pVII repertoires. This method advances the conventional approach of immune cross-reactivity by providing unambiguous evidence of mimicry at a clonal level.

The mechanisms underlying pathologic VITT antibodies may help to unravel other poorly understood conditions in which a boosted immune response results in pathologic autoantibodies.

 

[Utsikt]

It might have gone over my head in the snippets, but do they show how the normal mechanisms for avoiding auto-immunity fail?

If the antibody has a high likelihood of binding to something in the body, wouldn’t it be dealt with long before it had a chance to get out into the body and do damage?

 

[Chandelier]

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