Molecular and functional characterization of the normal and malignant plasma cell – bone interface – from single cell mutual interaction analysis to clinical implications in myeloma bone disease and survival

多发性骨髓瘤是一种恶性浆细胞在骨髓中扩散和积聚，继而导致骨疾病和血管生成的疾病。这些特征与其他恶性疾病是共同的。多发性骨髓瘤在两个方面不同：骨髓瘤细胞(MMC)扩散到并生活在与其生理对应物--骨髓浆细胞(BMPC)相同的“自然环境”中。此外，正如我们所显示的，当诱导血管生成和与骨转换相互作用时，MMC至少部分依赖于正常浆细胞功能来产生恶性行为。与正常浆细胞一样，恶性细胞依赖于通过与骨髓中其他类型的细胞相互作用而获得生存支持，统称为“生态位”。在体外，我们已经证明在骨髓瘤/间充质基质细胞(MSC)/成骨细胞共培养中，这种相互作用显著地通过物理相互作用而发挥，并导致两种细胞类型的表达和行为的变化。我们假设，这种物理相互作用创造了利基，并引发了个人签名，加强了粘连，支持了肿瘤细胞，扰乱了成骨，在临床上导致骨丢失和病理性骨折。小生境可能赋予骨髓瘤以休眠或抵抗(微小残留病)的特征。我们已经证明了这种分子串扰是可用药的，通过阻断骨细胞特异性WNT拮抗剂硬化素，观察到骨髓瘤荷瘤小鼠成骨细胞数量和骨形成的增加。我们还表明，接触诱导的KISS1R系统是一个潜在的靶点，作为治疗的PET示踪剂。在这个应用中，我们将重点关注正常和恶性浆细胞以及MSC/成骨细胞之间相互作用的表面分子。我们将使用包括单细胞RNA-seq的表达分析来解决群体异质性，并通过原子力显微镜和CRISPR/Cas9筛选对相互作用的功能评估，以及随后的体内协作验证。在第二步，我们的体外研究结果将与大量具有分子特征的恶性浆细胞样本以及全身成像评估骨骼疾病、浆细胞渗透和血管生成有关。我们已经证明，纳米/微到大规模临床数据集的数据关系可以作为相关性的过滤器，并将允许识别临床前适用的发现。第一个证据是硬化素仅在骨细胞中表达，而不是来自630名MM患者的恶性浆细胞；第二个证据是我们共同开发的针对BMPC/MMC细胞表面分子BCMA的T细胞双特异性抗体EM801/901于18年第一季度进入临床试验，为单独应用开发治疗用BCMA-PET-Tracer奠定了基础。我们将为微骨财团内骨髓瘤传播、骨疾病和血管生成相关研究问题的合作网络做出贡献。

Multiple Myeloma is a disease of malignant plasma cells spreading and accumulating in the bone marrow with subsequent induction of bone disease and angiogenesis. These features are in common with other malignant diseases. Multiple myeloma is different in two regards: Myeloma cells (MMC) spread to and live in the same “natural environment” as their physiological counterpart, bone marrow plasma cells (BMPC). Further, as shown by us, MMC rely at least partly on normal plasma cell functions for malignant behaviour when inducing angiogenesis and interacting with bone turnover. Malignant like normal plasma cells depend on receiving survival-support by interaction with other cell types in the bone marrow, collectively called “niche”. In vitro, we have shown in myeloma/ mesenchymal stromal cell (MSC)//osteoblast co-cultures that this interaction is prominently exerted by physical interaction and leads to changes in expression and behaviour in both cells types. We hypothesize that this physical interaction creates niches and elicits individual signatures that strengthen adhesion, support tumor cells, and disrupt osteogenesis, clinically leading to bone loss and pathological fractures. Niches may confer dormancy or resistance (minimal residual disease) features to myeloma. We have shown this molecular crosstalk being drugable exemplified by blocking the osteocyte-specific wnt antagonist sclerostin, observing increased osteoblast numbers and bone formation in myeloma bearing mice. We also showed that the contact-induced KISS1R system is a putative target as a theranostic PET tracer.In this application, we will focus on surface molecules featuring the interaction between normal and malignant plasma cells and MSC/osteoblasts. We will use expression analysis including single cell RNA-seq to address population heterogeneity, and functional assessment of interaction by atomic-force microscopy and CRISPR/CAS9 screen with subsequent collaborative in vivo validation in myeloma mouse models. In a second step, our in vitro findings will be related to a large cohort of molecularly characterized malignant plasma cell samples alongside whole-body imaging assessing bone disease, plasma cell infiltration and angiogenesis. We have shown that relation of data at nano-/micro- to large scale clinical data sets can act as a filter for relevance and will allow identification of pre-clinically applicable findings. The first is evidenced by sclerostin being expressed in osteocytes only, in contrast to malignant plasma cells from 630 MM patients; the second by our co-development of the T-cell bispecific antibody EM801/901 against the BMPC/MMC cell-surface molecule BCMA entering clinical trials in Q1/18, building the basis for a separate application for development of a theranostic BCMA-PET-Tracer. We will contribute to a collaborative network for myeloma dissemination, bone disease and angiogenesis related research questions within the µBone-consortium.