破骨细胞及其前体细胞在多发性骨髓瘤诱导的免疫抑制中的作用机制及逆转策略

The number and activity of osteoclasts are increased in multiple myeloma. Accumulating evidence suggests that osteoclasts mediate not only bone remodeling, but also may regulate the immune system, i.e. this relationship is usually termed as “osteoimmunology”. Specifically, during autoimmune condition, an association between osteoclastic bone resorption and T- cell immune activation has frequently been observed. Osteoclastogenesis is induced by activated T cells via production of potent osteoclastogenic cytokines RANKL and interleukin (IL)-1b . Moreover, activated T cells also inhibit osteoclast differentiation via IFN-γ, IL4, and IL10. Although, the reciprocal impact of osteoclasts on T cells is less well defined, but it is likely that osteoclasts effectively suppress T cell proliferation in a feedback loop mechanism and act as inducible immunosuppressive cells to prevent osteoporosis or osteosclerosis. We postulated that osteoclasts effectively suppress T cell proliferation in a feedback loop mechanism and act as inducible immunosuppressive cells to prevent osteoporosis or osteosclerosis. This leads to the hypothesis that this mechanism of osteoclast checkpoint may have been hijacked by MM cells to escape immune response. Our previous results have shown that osteoclasts rescued MM-specific cytotoxic T lymphocytes (CTLs) induced apoptosis of myeloma cells. In this proposal, we will analyze the role of osteoclasts and osteoclast precursors (OCPs) in the MM-induced immunosuppression on multiple levels, including phenotypic analysis of clinical specimens, in vitro cellular function/molecular mechanism studies, as well as mouse model in vivo. Immune-based approaches represent the most promising group of agents in the treatment of myeloma, and immunotherapy in MM come of Age. One of the major factors that limit the effect of cancer immune therapy is the persistence of tumor-associated immune suppression which arises in the tumor microenvironment. This study will provide a further strategy to improve the immunotherapy in MM and a new insight of the treatment of myeloma bone disease.

破骨细胞激活是多发性骨髓瘤(MM)的显著特征。骨免疫学(Osteoimmunology)研究表明破骨细胞不仅参与骨重塑，还与免疫系统紧密关联。自身免疫疾病中，T淋巴细胞异常激活后常伴有骨质破坏，提示T细胞对于破骨细胞具有重要的调节作用。为了避免发生骨质疏松或骨质硬化，我们推测机体存在“破骨细胞检查点”的负反馈机制，以调节T细胞。机体的这一负反馈机制可能被MM细胞所“绑架”，抑制T淋巴细胞，建立免疫抑制的微环境。我们前期工作表明，破骨细胞表达多种抑制分子，抑制T淋巴细胞反应，保护MM细胞免受特异细胞毒T淋巴细胞的杀伤。我们拟通过临床样本分析、细胞生物学功能/分子机制研究，研究破骨细胞及其前体细胞在MM诱导的免疫抑制微环境中的作用及逆转策略。MM即将进入免疫治疗时代，MM诱导的免疫抑制微环境严重制约疗效的提高。本课题将有助于发现提高免疫治疗疗效的策略，并为MM骨病治疗提供新的思路。

既往认为骨骼系统仅仅是一种支撑人体活动的组织器官。然而，越来越多的研究证明骨骼还是一个具有重要免疫调节功能的免疫器官。近年对骨与免疫系统之间的相互作用机制有了更进一步的认识，并且诞生了新的交叉学科骨免疫学（Osteoimmunology）多发性骨髓瘤（MM）肿瘤细胞的对于所处微环境的改造，最显著的特征是激活大量破骨细胞。破骨细胞在MM细胞促增殖、抗凋亡和耐药中的作用已经明确。但目前尚无研究从肿瘤免疫角度解释MM细胞激活数量众多破骨细胞的原因。为了维持骨代谢平衡，避免骨质疏松和骨质硬化，我们推测破骨细胞应该存在一种负反馈机制用于抑制T淋巴细胞，以实现双向调节功能，以免破骨细胞过度活化或者失活，称之为“破骨细胞检查点（Osteoclasts checkpoint）”。如果破骨细胞存在针对T淋巴细胞的负反馈机制，MM肿瘤细胞可能利用这些反馈机制达到抑制T淋巴细胞进行免疫逃逸。.我们的研究发现：1）破骨细胞可以显著抑制CD4+T和CD8+ T细胞的增殖，从而保护MM肿瘤细胞免受特异性细胞毒T淋巴细胞的杀伤；2）破骨细胞形成过程中，免疫检查点分子PD-L1、Galectin-9、HVEM和CD200，T细胞代谢调节分子indoleamine 2, 3-dioxygenase（IDO）和CD38均出现明显上调；除了CD38以外，这些分子在破骨细胞的表达均明显高于MM细胞；3）PD-L1抑制剂和IDO抑制剂可以部分抑制破骨细胞对于T淋巴细胞的抑制作用；5）破骨细胞分泌Galectin-9和APRIL；Galectin-9选择性的促进T淋巴细胞的凋亡，而对MM细胞和单核细胞没有影响；APPRIL可以通过pMEK信号通路促进MM细胞表达PDL1；5）破骨细胞形成过程中，CD38表达升高。抗CD38单克隆抗体不影响破骨细胞的形成，但是可以通过抑制HVEM和IDO的表达，减轻破骨细胞对于T淋巴细胞的抑制作用。.本研究从免疫角度解释了骨髓瘤骨病的发生机制，明确了破骨细胞在MM免疫微环境中的作用，对于多发性骨髓瘤的免疫治疗和骨病的治疗均具有重要的启示意义。