The development of therapeutic monoclonal antibodies through mouse immunization often originates drug candidates that are not cross-reactive to the mouse ortholog. In such cases, and particularly in oncology, drug efficacy studies are performed on human tumor xenografts or with “surrogate” anti-mouse ortholog antibodies if targeting tumor host cells. Safety assessment of drug candidate(s) is performed at a later development stage in healthy non-human primates. While the latter remains necessary before a drug advances into human subjects, it precludes evaluation of safety in disease conditions and drug de-risking during early development. Therefore, mouse models that allow concomitant evaluation of drug efficacy and safety are highly desirable. The C-X-C motif chemokine receptor 4 (CXCR4) is an attractive target for tumor-targeted and immuno-oncology therapeutics, with multiple mouse immunization-derived antibodies undergoing clinical trials. Given the pleiotropic role of CXCR4 in cancer biology, we anticipate continuous interest in this target, particularly in the testing of therapeutic combinations for immuno-oncology. Here, we describe the generation and validation of the first mouse knock-in of the whole coding region of human CXCR4. Homozygous human CXCR4 knock-in (hereafter designated as HuCXCR4KI) mice were viable and outwardly healthy, reproduced normally and nursed their young. The expression pattern of human CXCR4 in this model was similar to that of CXCR4 expression in normal human tissues. The human CXCR4 knock-in gene was expressed as a biologically active protein, thereby allowing normal animal development and adequate”homing” of leukocytes to the bone marrow. To further validate our model, we used an in vivo functional assay of leukocyte mobilization from bone marrow to peripheral blood by blocking CXCR4 signaling. Both an anti-human CXCR4 -specific blocking antibody and the small molecule CXCR4 inhibitor AMD3100 induced increased leukocyte counts in peripheral blood, whereas an anti-mouse CXCR4 –specific blocking antibody had no effect. This new mouse model is useful to evaluate efficacy and safety of anti-human CXCR4 -specific drugs as single agents or in combination therapies, particularly in the oncology, immuno-oncology, wound healing and chronic inflammation therapeutic areas.

通过小鼠免疫开发治疗性单克隆抗体通常会产生与小鼠直系同源蛋白没有交叉反应性的候选药物。在这种情况下，特别是在肿瘤学中，如果靶向肿瘤宿主细胞，则对人肿瘤异种移植物或使用“替代”抗小鼠直系同源抗体进行药物功效研究。在健康的非人类灵长类动物的发育后期，进行候选药物的安全性评估。尽管后者在药物进入人体之前仍然是必需的，但它无法评估疾病状况下的安全性以及在早期开发过程中降低药物的风险。因此，非常需要能够同时评估药物功效和安全性的小鼠模型。CXC基序趋化因子受体4（CXCR4）是针对肿瘤靶向和免疫肿瘤疗法的引人注目的靶标，多种源自小鼠免疫的抗体正在接受临床试验。鉴于CXCR4在癌症生物学中的多效性作用，我们预计将对该目标产生持续的兴趣，尤其是在免疫肿瘤治疗组合的测试中。在这里，我们描述了人类CXCR4整个编码区的第一个小鼠敲入的生成和验证。纯合的人类CXCR4敲入小鼠（以下称为HuCXCR4KI）是活的且向外健康的小鼠，可正常繁殖并哺育其幼仔。该模型中人CXCR4的表达模式与正常人组织中CXCR4的表达模式相似。人类CXCR4敲入基因被表达为具有生物活性的蛋白质，从而使动物正常发育，并使白细胞充分“归巢”到骨髓。为了进一步验证我们的模型，我们使用了阻断CXCR4信号转导从骨髓到外周血的白细胞动员的体内功能测定。抗人CXCR4特异性阻断抗体和小分子CXCR4抑制剂AMD3100均可诱导外周血白细胞计数增加，而抗小鼠CXCR4特异性阻断抗体则无作用。这种新的小鼠模型可用于评估作为单药或联合疗法的抗人CXCR4特异性药物的功效和安全性，尤其是在肿瘤学，免疫肿瘤学，伤口愈合和慢性炎症治疗领域。