电压依赖性钙通道α2δ1亚基阳性肝肿瘤干细胞自我更新能力调控的分子机制

It is hypothesized that a rare subset of cancer cells, often operationally referred to as cancer stem cells (CSCs) or tumor-initiating cells, are responsible for sustaining tumor growth and recurrence. The self-renewal property of CSCs enables them to maintain at a constant population in a given tumor tissue and to sustain the growth of tumor, however, the molecular mechanisms underlying the self-renewal capability of CSCs reamin to be elucidated. We have previously identified a population of liver CSCs that expressing the isoform 5 of α2δ1, which is encoded by CACNA2D1, a composing subunit of a voltage-gated calcium channel. The α2δ1-positive liver CSC population was found to be a subset with the highest tumorigenic potential of known liver cancer stem cell population defined by the marker CD13, CD133, or EpCAM. Overexpression α2δ1 in non-liver CSCs could result in enhanced hepatosphere-forming ability of these cells with elevated intracellular calcium level, suggesting the acquirement of self-renewal property. Furthermore, α2δ1 is essential in Ca2+ signaling and maintenance of CSC properties including the self-renewal capability, suggesting that α2δ1 could serve as a therapeutic target in the treatment of hepatocellular carcinoma. Based on these findings, this project is designed to study further the molecular mechanisms underlying the roles of α2δ1 in the regulation of the self-renewal property of liver CSCs such as: What molecule(s) does α2δ1 interact with in its function? What are the downstream signaling cascades in the modulating of self-renewal property? Is there any cross-talk between the function of α2δ1 and the known signaling pathways that play roles in the self-renewal of CSCs? What are the molecular mechanisms to maintain the expression of α2δ1 in liver CSCs? Hopefully, we can find the signaling network and some key points involved in the determination and maintenance of the self-renewal property of these liver CSCs. This study would contribute to the understanding of the occurrence and progression of hepatocellular carcinoma at both the cellular and molecular levels and advance the development of prognostic and therapeutic strategies.

肿瘤干细胞（CSC）的存在可能是肿瘤产生、复发转移和治疗失败的根本原因。CSC自我更新是维持肿瘤组织内CSC持续存在和肿瘤生长的关键机制，但CSC自我更新调控的分子机制远未阐明。我们前期发现电压依赖性钙离子通道组成亚基α2δ1（亚型5）阳性肝癌细胞具有肿瘤干细胞特性，是多个已报道肝肿瘤干细胞群中共有的亚群。α2δ1（由基因CACNA2D1编码）可引起细胞内钙浓度升高而使肝癌细胞获得和维持自我更新等肿瘤干细胞特性，有望成为肝癌治疗的分子靶点。本课题拟进一步研究α2δ1参与调控肝肿瘤干细胞自我更新等特性的分子机制包括其作用的伴侣分子、下游信号通路、与其它信号通路的相互调控，以及肝肿瘤干细胞内维持α2δ1高表达的分子机制，寻找α2δ1（亚型5）阳性肝肿瘤干细胞自我更新等特性建立和维持的分子调控网络和关键节点，为阐明肝癌发生、发展的分子机理以及预后判断、治疗分子靶标研究提供新线索。

肿瘤干细胞（CSC）的存在可能是肿瘤产生、复发转移和治疗失败的根本原因，其自我更新是维持肿瘤组织内CSC持续存在和肿瘤生长的关键机制，但CSC自我更新调控的分子机制远未阐明。我们曾报道电压依赖性钙离子通道组成亚基α2δ1（亚型5）阳性肝癌细胞具有肿瘤干细胞特性，是多个已报道肝肿瘤干细胞群中共有的亚群。本课题基于这一发现，主要从α2δ1参与调控肝肿瘤干细胞自我更新涉及的信号通路、CSC维持α2δ1高表达的分子机制，寻找α2δ1（亚型5）阳性肝肿瘤干细胞自我更新等特性建立和维持的分子调控网络和关键节点等角度探讨了α2δ1（亚型5）阳性肝肿瘤干细胞自我更新的分子机制。研究结果表明，α2δ1是肝癌、胰腺癌、肺癌等多种肿瘤干细胞的功能性标志物和治疗靶点。α2δ1通过调控钙离子内流而IP3R2通过调控内质网钙释放导致CSC内钙振荡进而调控其自我更新和成瘤。进一步，α2δ1可以通过CaMKD2磷酸化PKM2参与α2δ1阳性癌干细胞自我更新能力的调控。更重要的是，我们发现，在α2δ1阳性肝癌干细胞中抑制性miRNA let-7c、hsa-mir222、hsa-mir 200b和hsa-mir424的低表达，导致其共同的靶基因PBX3处于高表达，激活了包括α2δ1的编码基因CACNA2D1、Notch3、Sall2等决定干性基因的表达，从而在CSC干细胞样特性的获得与维持中发挥重要作用。此外，我们还对α2δ1的单克隆抗体进行了人源化改造并构建了其CAR-T细胞，体内外实验均表明靶向α2δ1有可能为肝癌治疗提供新手段。这些发现为阐明CSC自我更新的分子调控网络、揭示肝癌等肿瘤发生、发展的分子机理以及预后判断、治疗分子靶标提供了新线索。