靶向BAFFR和BCR信号通路干预SLE发病作用及机制的研究

Based on literature review and our previous findings, BAFF plays an important role in the proliferation and differentiation of autoreactive B cells in SLE. Belimumab, a specific monoclonal antibody of BAFF approved by FDA for SLE treatment, has successfully improved the SLE responsive index with definite therapeutic effect. But low efficacy and increased infection rates in its' long-term use are urgent problems need to be solved. We infer that the low concentration of BAFF after Belimumab administration would influence its binding with TACI and down-regulate TACI signaling pathway, which attenuated activation-induced cell death (AICD) of B cells and deregulated T cell independent innate immune responses. In addition, B cells could be abmormally activated by multiple signaling pathways, such as Ras-ERK signaling pathway involved in SLE pathogenesis. A multifunctional nanoparticle will be designed and synthesized to solve these problems by conjugating BAFFR specific monoclonal antibody on the surface while encapsulating RasGRP3 siRNA - Quantum dots complex inside the particle. The BAFFR antibody can be used to target the BAFF receptor on B cells with high specificity and further block the proliferation signaling of B cells mediated by BAFF-BAFFR interaction. In the meantime the RasGRP3 siRNA can be used to inhibit the proliferation and differentiation of B cells through Ras-ERK, a key downstream signaling pathway after BCR activation. Furthermore, the increased combination of BAFF with TACI can promote the innate immune responses and induce the apoptosis of autoreactive B cells through AICD. The SLE mice model and cell model will be established to verify the hypothesis and reveal the effect and mechanism of this dual-targeting therapy, that is, by targeting both the BAFFR and BCR activation signaling pathway. This will provide a promising strategy in the SLE treatment.

文献报道以及我们前期研究证明，BAFF在SLE自身反应性B细胞增殖分化中起重要作用，针对BAFF抗体Belimumab治疗SLE获得疗效，但长期使用疗效降低且感染率增加，推测其与抑制BAFF与TACI结合，使得TACI信号通路和AICD途径减弱，不能发挥固有免疫以及诱导自身反应性B细胞凋亡相关，且SLE可通过多条信号通路异常激活B细胞，如RAS/ERK信号通路。基于此，本课题设计，以BAFFR为靶点，以多功能纳米粒子作载体包被RasGRP3 siRNA量子点复合物，利用BAFFR抗体特异性靶向B细胞，阻断BAFF-BAFFR信号通路活化，沉默Ras/ERK信号通路阻止B细胞的增殖分化，同时增加BAFF与TACI结合，促进固有免疫应答，以及活化自身反应性B细胞凋亡。并通过细胞模型与动物模型研究验证上述假设与揭示其机制，从而为以BAFFR和BCR活化通路为靶点的SLE治疗提供新策略。

BAFF在系统性红斑狼疮的发生发展过程中发挥重要作用，主要通过BAFFR受体促进自身反应性B细胞存活及增殖分化。我们证实MRL-Faslpr/lpr 品系狼疮模型鼠血清BAFF水平明显升高，并在体内外实验中均发现BAFF可与TACI结合诱导MZ B细胞分泌IL-35。进一步研究发现该群产IL-35 B细胞具有CD5+CD1dhiFcgRIIbhi独特的细胞表型，能够抑制CD4+CD25-T细胞分泌炎性细胞因子IFN-γ及TNF-α，同时能够调节CD4+CD25+调节性T细胞的产生，促进CD4+T细胞进一步产生IL-35，形成正反馈环路，发挥免疫调节的作用。该研究不仅揭示了BAFF在自免疾病中的免疫调节作用，丰富了其参与免疫反应的机制，也为抗BAFF治疗长期疗效欠佳提供重要新依据。此外，体外实验证明封闭BAFF受体BAFFR后细胞凋亡增加，证明了抗BAFFR抗体的有效性，因此我们进一步制备了以Anti-BAFFR为靶向，包封BCR下游重要信号分子BTK小分子抑制剂Evobrutinib的纳米载药系统，联合抑制BCR、BAFFR信号通路，进而增强TACI调节性信号，探究其对SLE的治疗效应。发现该纳米载药体系，有效减缓了MRL-Faslpr/lpr 品系狼疮模型鼠体重的下降及尿蛋白的升高，降低了脾脏T细胞、DC、巨噬细胞比例，以及巨噬细胞上共刺激分子CD80的表达，该研究为SLE的靶向精准治疗提供了新的策略和依据。