Molecular dissection of signaling pathways exerting bone anabolic and anti-tumor effects of physical stimuli in myeloma bone disease

多发性骨髓瘤（MM）是一种恶性浆细胞疾病，其中肿瘤细胞诱导溶骨性骨病。虽然恶性肿瘤中骨疾病的合成代谢治疗仍是一个讨论的问题，但预防或挽救骨质减少的非药物方法是使用控制良好的物理刺激。在临床前MM研究中，我们已经证明，机械负荷形式的物理刺激具有有希望的骨保护结果，甚至可以减轻肿瘤细胞的生长和扩散。皮质骨细胞的转录组分析揭示了由肿瘤细胞引起的一组细胞外基质相关基因的基因表达的实质性改变，这些基因被机械负荷拯救或刺激。细胞粘附是组织中机械载荷的重要组成部分，并且粘附机制的紧密性通过载荷来加强。在分析MM细胞在体外的亚群，坚持与骨骼前体不同的力量，我们发现了一组特征性的差异表达的基因在紧密粘附的MM细胞。我们确定了体内和体外基因组之间的部分重叠，代表了一个令人信服的相互作用的基因网络，其中候选人属于三个集群（1）骨骼发育，（2）脂质转运和（3）细胞外基质组织。此外，我们发现来自该网络的主要候选物（如低密度脂蛋白受体相关蛋白1（LRP 1））的低表达与使用大量临床来源的RNAseq数据的MM患者生存率较差相关。在这个项目中，我们将从分子水平上剖析粘附/机械传导和肿瘤归巢和扩散背后的网络，寻找有效类型的机械负载和创新的药物靶点，以解决骨合成代谢和抗肿瘤作用。我们假设LRP 1是协调机械反应性骨重建和介导MM和骨细胞中物理刺激的拯救作用的枢纽的主要候选者。在存在MM的情况下，骨中LRP 1的表达减少，并且其骨特异性敲除在过去显示严重破坏骨重建。类似地，丝氨酸蛋白酶抑制剂H1，一种对胶原蛋白折叠和完整性至关重要的分子伴侣，介导MM细胞紧密粘附并控制骨细胞中细胞外基质的产生。这将在体外通过骨骼前体、骨细胞系和MM细胞之间的相互作用实验以及在各自的小鼠模型中的体内实验来证明，其中使用全身低幅度高频振动、MM细胞的基因工程以及LRP 1和Serpin H1信号传导的药理学调节来调节粘附/负载条件。这将允许在机械负荷模型中识别目标，其可以被处理以重建健康的骨组织，同时在微环境中赋予抗肿瘤作用。

Multiple myeloma (MM) is a malignant plasma cell disorder, in which tumor cells induce osteolytic bone disease. While anabolic treatment of bone disease in malignancies is still a matter of discussion, a non-pharmacological approach to prevent or rescue osteopenia is the use of well-controlled physical stimuli. In preclinical MM studies we have shown that physical stimuli in the form of mechanical loading have promising bone-sparing outcomes and even mitigate tumor cell growth and dissemination. Transcriptome profiling of cortical osteocytes revealed substantial alterations of gene expression caused by tumor cells in a set of extracellular matrix-related genes which were rescued or stimulated by mechanical loading. Cell adhesion is an important component of mechanical loading in tissue and the tightness of adhesion mechanisms is enforced by loading. While analyzing subpopulations of MM cells in vitro that adhered with different forces to skeletal precursors we found a characteristic set of differentially expressed genes in tightly adhering MM cells. We identified a partial overlap between the in vivo and in vitro gene sets representing a convincing interactive gene network in which candidates belong to three clusters of (1) skeletal development, (2) lipid transport and (3) extracellular matrix organization. In addition, we found that low expression of lead candidates from this network like the low-density lipoprotein receptor-related protein 1 (LRP1) is associated with worse MM patient survival using large cohorts of clinically derived RNAseq data. In this project we will molecularly dissect the networks behind adhesion / mechanotransduction and tumor homing and spread, searching for effective types of mechanical loading and innovative druggable targets that address the bone anabolic and anti-tumor effects. We hypothesize that LRP1 is a lead candidate for a hub orchestrating mechanoresponsive bone remodeling and mediating rescue effects of physical stimulation in MM and bone cells. LRP1 expression is diminished in bone in the presence of MM and its bone specific knockout was shown in the past to severely disrupt bone remodeling. Similarly, Serpin H1, a chaperone essential for collagen folding and integrity, mediates tight MM cell adhesion and controls extracellular matrix production in bone cells. It will be proven both in vitro by interaction experiments between skeletal precursors, osteocyte cell lines and MM cells and in vivo in respective mouse models where adhesion / loading conditions are modulated using whole-body low-magnitude high frequency vibration, genetic engineering of MM cells and pharmacological modulation of LRP1 and Serpin H1 signaling. This will allow for identifying targets in models of mechanical loading that can be addressed to reconstitute healthy bone tissue and at the same time impart anti-tumor effects in the microenvironment.