抑制SCD1促进神经酰胺聚集诱导肿瘤细胞凋亡的作用及机制

Stearoyl-CoA Desaturase 1(SCD1) has been shown to play an important role in lipid metabolism, which is highly expressed in many kinds of human tumors. However, mechanism underlying SCD1-mediated anti-tumor effect has maintained unclear. We have reported endo-lipid messenger ceramides played a critical role in tumor fate modulated by SCD1 inhibition. In vitro study in colorectal and ovary cancer cells demonstrated that inhibition of SCD1 activity promoted apoptosis events mediated by intracellular ceramide signals through induction of ceramide biosynthesis, rather than exclusive SFA accumulation. In vivo study in xenograft colorectal cancer mice showed pharmacologic administration of SCD1 inhibitor A939 significantly delayed tumor growth, which was reversed by L-cycloserine, an inhibitor of ceramide de novo biosynthesis. Herein, we propose that inhibition of SCD1 activity might promote the ceramide synthases SPT1 and CerS1 to induce endogenous ceramides accumulation and cancer cell apoptosis. This increase in ceramide production might also associate with the activation of TLR-4/IKKβ signaling or inhibition of ceramide metabolic pathway. In order to validate the hypothesis, LoVo, Colo205, SKOV3 cell lines and xenograft cancer mice will be used to investigate the mechanism in the cross-talk of SCD1-mediated lipid pathway and endo-ceramide biosynthesis pathway. This study will disclose the unambiguous roles of ceramide signals in SCD1-mediated anti-tumor property.

硬脂酰辅酶A去饱和酶(SCD1)是调节游离脂肪酸不饱和度的关键酶，在肿瘤组织中多见高表达。我们的研究显示，SCD1抑制剂对不同肿瘤细胞的抑制作用存在显著差异，单纯SFA/MUFA比例变化并不直接影响肿瘤细胞增殖；抑制SCD1引起促肿瘤细胞凋亡、抑制在体肿瘤增殖行为均由内源性神经酰胺介导，从头合成途径参与其中。但SCD1如何调控神经酰胺及诱导肿瘤细胞凋亡的机制并不明确。基于前期结果，我们提出假说：抑制SCD1后，通过调节神经酰胺从头合成酶SPT1和CerS1，导致神经酰胺普遍升高基础上，某些神经酰胺特异性聚集，引起肿瘤细胞凋亡，此过程可能与脂肪酸不饱和度变化从而激活TLR-4和/或IKKβ信号通路有关。本项目拟分别采用人结肠癌和卵巢癌细胞为工具细胞，通过体内外实验，研究SCD1抑制对内源性神经酰胺合成代谢途径的影响，为揭示脂代谢网络调控肿瘤发生发展的机制，寻找防治肿瘤的潜在药物靶标奠定基础。

我们曾经报导了在肠癌中，stearoyl-CoA-desaturase 1 (SCD1) 缺失可以通过促进内源性神经酰胺的从头合成而导致细胞凋亡。然而，SCD1和神经酰胺这两条通路之间的信号联接机制还并没有被阐明。本研究采用SCD1敏感的肠癌细胞Colo205和SCD1耐药的卵巢癌细胞SKOV3为研究对象，分别用抑制剂和基因敲低的方式使SCD1活性缺失，研究SCD1通过何种途径调控神经酰胺合成及肿瘤增殖。结果显示，SCD1缺失后，肿瘤细胞中脂肪酸首先发生变化，肠癌细胞中饱和脂肪酸（SFA）含量不变，单不饱和脂肪酸（MUFA）含量显著下降，细胞增殖受到抑制；而卵巢癌细胞中SFA含量增幅大于MUFA，细胞增殖不受影响。外源性补充OA可逆转SCD1缺失导致的神经酰胺升高和细胞增殖抑制。由此可以推论，肿瘤细胞受到SCD1调控后，MUFA的含量下降是影响神经酰胺从头合成和细胞增殖的重要因素。接下来对神经酰胺具体合成途径的研究过程中发现，SCD1缺失可提高肠癌细胞中神经酰胺从头合成途径相关酶CerS6和SPT1的表达，而OA可逆转此过程。对CerS6进行深入研究发现，同时敲低SCD1和CerS6，能够逆转SCD1缺失所导致的肠癌细胞凋亡，而P53可介导CerS6的表达量增加。为进一步研究SCD1通过何种途径调控神经酰胺合成，本课题将SCD1抑制后的肿瘤细胞系进行了RNAseq差异基因表达谱分析。结果显示，肠癌细胞中SCD1抑制前后的差异基因在信号转导通路NF-κB的富集程度最高，提示我们NF-κB可能是SCD1调控的重要靶点之一。体外实验研究发现，同为SCD1抑制，肠癌细胞中的NF-κB通路被激活，而卵巢癌中则无此现象。OA和NF-κB抑制剂PDTC均能抑制SCD1缺失所导致的NF-κB激活，且PDTC能够逆转SCD1缺失所引起的神经酰胺合成相关酶表达升高。裸鼠体内荷瘤实验中，抑制NF-κB可逆转SCD1抑制剂的抗肿瘤作用。综上所述，我们的研究阐明了在肠癌细胞中，NF-κB信号通路的激活在SCD1缺失所导致的神经酰胺从头合成过程中起到重要作用，CerS6和SPT1参与其中，而减少的MUFA是此过程的促发因素之一。