改性anti-BMP2抗体协同激活成骨破骨与预防双磷酸盐颌骨坏死的作用及机制研究

Bisphosphonate-related osteonecrosis of jaw (BRONJ) is a serious and intractable complication. Coordinated mobilization of osteoclasts and osteoblasts to promote physiological new bone formation is key to prevent and treat BRONJ. The applicant found that anti-BMP2 antibody can mobilize osteoblasts and osteoclasts through its Fab and Fc segments respectively. However, how to screen and optimize the anti-BMP2 antibody with the best osteoclast/ osteoblast inducing efficiency so as to prevent BRONJ remains to be studied. According to cell affinity, osteoblast and osteoclast inducing potential, the applicant has screened out the 3G7 anti-BMP2 antibody as the basic type for further study. In the present project, we will try to establish anti-BMP2 antibody with different osteoclast/osteoblast inducing efficiency by modifying the sialic acid and galactose components of 3G7 antibody. Then the potential to prevent BRONJ would be analyzed so as to determine the best osteoclast/osteoblast inducing efficiency. Finally, with an emphasis on osteoclast activation, the mechanism of anti-BMP2 antibody in preventing BRONJ was analyzed from two aspects: the type of effector Fc receptor and the phenotypic change of Fc receptors of cells in environment with bisphosphonates. This study is of great significance for optimizing the clinical treatment protocol of BRONJ and exploring the pathomechanism of BRONJ, and may provide new methods to treatment disorders associated with osteoclast-osteoblast imbalance.

双磷酸盐颌骨坏死（BRONJ）是口腔相关的严重并发症，目前疗效欠佳。破骨成骨共抑制是BRONJ发生的根源，协调并进地调动破骨与成骨是防治的关键。申请人发现anti-BMP2抗体可通过其Fab与Fc结构分别调动成骨与破骨，但如何筛选并优化生成破骨/成骨效能最佳的anti-BMP2抗体并达到预防BRONJ的目的仍有待研究。申请人已根据亲和度与成/破骨诱导功能筛选出具备基本条件的anti-BMP2抗体克隆，本研究拟改变该抗体的唾液酸与半乳糖成分，建立具有不等破骨/成骨效能的改性抗体，分析各改性抗体预防BRONJ的生物潜能并明确最佳改性方案，最终以破骨激活为线索，从Fc的效应受体类型以及双磷酸盐环境下细胞Fc受体表型改变两方面出发，分析anti-BMP2抗体预防BRONJ的作用机制。本研究对于优化BRONJ临床处理方案与探索BRONJ机制有重要意义，并可能为其他破骨/成骨失调疾病提供新治疗方法。

双磷酸盐颌骨坏死（BRONJ）是口腔相关的严重并发症，目前疗效欠佳。破骨成骨共抑制是BRONJ发生的根源，协调并进地调动破骨与成骨是防治的关键。申请人发现anti-BMP2抗体可通过其Fab与Fc结构分别调动成骨与破骨，但如何筛选并优化生成破骨/成骨效能最佳的anti-BMP2抗体并达到预防BRONJ的目的仍有待研究。申请人通过糖基改性建立具有不等破骨/成骨效能的改性抗体，从中筛选出能够预防BRONJ的最佳类型，实现了BMP2抗体的优化。继而针对anti-BMP2抗体的体内成骨机制进行了深入研究，发现anti-BMP2抗体的体内具有一定的破骨抑制效应，而在体外具有破骨激活效应，提示破骨效应与体内缓解有关。接下来以破骨激活为线索，从Fc的效应受体类型以及双磷酸盐环境下细胞Fc受体表型改变两方面出发，通过成骨-破骨共培养，发现anti-BMP2抗体在体内形成的免疫复合物是通过上调成骨细胞与破骨细胞的直接接触率，启动接触依赖的EphrinB2-EphB4双向信号通路而实现成骨诱导与破骨抑制的。而当破骨细胞与成骨表明失偶联，成为游离破骨细胞后，BMP2 IC则仅发挥破骨诱导作用。本研究以BRONJ为模型，探讨BMP2抗体在双磷酸盐导致成骨破骨共抑制时，调动破骨与成骨，预防BRONJ发生的生物学功能及作用机制，并从FcγR表型改变这一新角度探讨BRONJ的致病机制，可为其他成骨破骨失调相关疾病提供新的治疗策略。在验证糖基改性anti-BMP2抗体的BRONJ防治效应时，课题组引入了ACPA作为阴性对照组。针对BRONJ与自身免疫疾病的关系进行了深入观察，以期揭示糖基改性ACPA可防治BRONJ的原因。最终基于预实验数据，申请人发现了BRONJ可能具有自身免疫特点。