Complement receptor 1 (CR1) is a membrane glycoprotein with a highly duplicated domain structure able to bind multiple ligands such as C3b and C4b, the activated fragments of complement components C3 and C4, respectively. We have previously used our knowledge of this domain structure to identify CSL040, a soluble extracellular fragment of CR1 containing the long homologous repeat (LHR) domains A, B, and C. CSL040 retains the ability to bind both C3b and C4b but is also a more potent complement inhibitor than other recombinant CR1-based therapeutics. To generate soluble CR1 variants with increased inhibitory potential across all three complement pathways, or variants with activity skewed to specific pathways, we exploited the domain structure of CR1 further by generating LHR domain duplications. We identified LHR-ABCC, a soluble CR1 variant containing a duplicated C3b-binding C-terminal LHR-C domain that exhibited significantly enhanced alternative pathway inhibitory activity compared to CSL040. Another variant, LHR-BBCC, containing duplications of both LHR-B and LHR-C with four C3b binding sites, was shown to have reduced classical/lectin pathway inhibitory activity compared to CSL040, but comparable alternative pathway activity. Interestingly, multiplication of the C4b-binding LHR-A domain resulted in only minor increases in classical/lectin pathway inhibitory activity. The CR1 duplication variants characterized in these potency assays, as well as in affinity in solution C3b and C4b binding assays, not only provides an opportunity to identify new therapeutic molecules but also additional mechanistic insights to the multiple interactions between CR1 and C3b/C4b.

中文翻译：

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CR1 结构域复制抑制补体途径的机制见解


补体受体 1 (CR1) 是一种膜糖蛋白，具有高度重复的结构域结构，能够结合多个配体，例如 C3b 和 C4b，分别是补体成分 C3 和 C4 的激活片段。我们之前利用对此结构域结构的了解来鉴定 CSL040，它是 CR1 的可溶性胞外片段，包含长同源重复 (LHR) 结构域 A、B 和 C。CSL040 保留了结合 C3b 和 C4b 的能力，但也是一种比其他基于 CR1 的重组疗法更有效的补体抑制剂。为了生成在所有三个补体途径中具有增强的抑制潜力的可溶性 CR1 变体，或具有偏向于特定途径的活性的变体，我们通过生成 LHR 结构域重复来进一步利用 CR1 的结构域结构。我们鉴定出 LHR-ABCC，一种可溶性 CR1 变体，含有重复的 C3b 结合 C 端 LHR-C 结构域，与 CSL040 相比，其表现出显着增强的旁路途径抑制活性。另一种变体 LHR-BBCC 含有 LHR-B 和 LHR-C 的重复，具有四个 C3b 结合位点，与 CSL040 相比，经典/凝集素途径抑制活性降低，但替代途径活性相当。有趣的是，C4b 结合 LHR-A 结构域的倍增仅导致经典/凝集素途径抑制活性的轻微增加。这些效力测定中表征的 CR1 重复变体，以及溶液 C3b 和 C4b 结合测定中的亲和力，不仅提供了鉴定新治疗分子的机会，而且还为 CR1 和 C3b/C4b 之间的多重相互作用提供了额外的机制见解。