Vaccine-induced immune thrombotic thrombocytopaenia (VITT) is a rare adverse effect of COVID-19 adenoviral vector vaccines^1,2,3. VITT resembles heparin-induced thrombocytopaenia (HIT) in that it is associated with platelet-activating antibodies against platelet factor 4 (PF4)⁴; however, patients with VITT develop thrombocytopaenia and thrombosis without exposure to heparin. Here we sought to determine the binding site on PF4 of antibodies from patients with VITT. Using alanine-scanning mutagenesis⁵, we found that the binding of anti-PF4 antibodies from patients with VITT (n = 5) was restricted to eight surface amino acids on PF4, all of which were located within the heparin-binding site, and that the binding was inhibited by heparin. By contrast, antibodies from patients with HIT (n = 10) bound to amino acids that corresponded to two different sites on PF4. Biolayer interferometry experiments also revealed that VITT anti-PF4 antibodies had a stronger binding response to PF4 and PF4–heparin complexes than did HIT anti-PF4 antibodies, albeit with similar dissociation rates. Our data indicate that VITT antibodies can mimic the effect of heparin by binding to a similar site on PF4; this allows PF4 tetramers to cluster and form immune complexes, which in turn causes Fcγ receptor IIa (FcγRIIa; also known as CD32a)-dependent platelet activation. These results provide an explanation for VITT-antibody-induced platelet activation that could contribute to thrombosis.

疫苗诱导的免疫血栓性血小板减少症 (VITT) 是 COVID-19 腺病毒载体疫苗^1,2,3的罕见不良反应。VITT 与肝素诱导的血小板减少症 (HIT) 相似，因为它与抗血小板因子 4 (PF4) ⁴的血小板活化抗体有关；然而，VITT 患者在未接触肝素的情况下会出现血小板减少和血栓形成。在这里，我们试图确定来自 VITT 患者的抗体在 PF4 上的结合位点。使用丙氨酸扫描诱变⁵，我们发现来自 VITT ( n = 5) 仅限于 PF4 上的 8 个表面氨基酸，所有这些氨基酸都位于肝素结合位点内，并且结合被肝素抑制。相比之下，来自 HIT 患者的抗体 ( n = 10) 与对应于 PF4 上两个不同位点的氨基酸结合。生物层干涉测量实验还表明，VITT 抗 PF4 抗体对 PF4 和 PF4-肝素复合物的结合反应比 HIT 抗 PF4 抗体更强，尽管解离率相似。我们的数据表明，VITT 抗体可以通过与 PF4 上的相似位点结合来模拟肝素的作用。这使得 PF4 四聚体聚集并形成免疫复合物，进而导致 Fcγ 受体 IIa（FcγRIIa；也称为 CD32a）依赖性血小板活化。这些结果为可能导致血栓形成的 VITT 抗体诱导的血小板活化提供了解释。