Human proteins repurposed as biologics for clinical use have been engineered through in vitro techniques that improve the affinity of the biologics for their ligands. However, the techniques do not select against properties, such as protease sensitivity or self-reactivity, that impair the biologics’ clinical efficacy. Here we show that the B-cell receptors of primary murine B cells can be engineered to affinity mature in vivo the human CD4 domains of the HIV-1-entry inhibitor CD4 immunoadhesin (CD4-Ig). Specifically, we introduced genes encoding the CD4 domains 1 and 2 (D1D2) of a half-life-enhanced form of CD4-Ig (CD4-Ig-v0) into the heavy-chain loci of murine B cells and adoptively transferred these cells into wild-type mice. After immunization, the B cells proliferated, class switched, affinity matured and produced D1D2-presenting antibodies. Somatic hypermutations in the D1D2-encoding region of the engrafted cells improved the binding affinity of CD4-Ig-v0 for the HIV-1 envelope glycoprotein and the inhibitor’s ability to neutralize a panel of HIV-1 isolates without impairing its pharmacokinetic properties. In vivo affinity maturation of non-antibody protein biologics may guide the development of more effective therapeutics.

重新用作临床用途的生物制剂的人类蛋白质是通过体外技术进行工程改造的，这些技术提高了生物制剂对其配体的亲和力。然而，这些技术不会选择损害生物制剂临床疗效的特性，例如蛋白酶敏感性或自身反应性。在这里，我们表明原代小鼠 B 细胞的 B 细胞受体可以被改造成在体内亲和成熟 HIV-1 进入抑制剂 CD4 免疫粘附素 （CD4-Ig） 的人类 CD4 结构域。具体来说，我们将编码半衰期增强形式的 CD4-Ig （CD4-Ig-v0） 的 CD4 结构域 1 和 2 （D1D2） 的基因引入小鼠 B 细胞的重链基因座，并将这些细胞过继转移到野生型小鼠中。免疫后，B 细胞增殖、类别转换、亲和力成熟并产生 D1D2 呈递抗体。移植细胞 D1D2 编码区的体细胞超突变提高了 CD4-Ig-v0 对 HIV-1 包膜糖蛋白的结合亲和力以及抑制剂在不损害其药代动力学特性的情况下中和一组 HIV-1 分离株的能力。非抗体蛋白生物制剂的体内亲和力成熟可能指导更有效疗法的开发。