抑癌蛋白SASH1调控Hippo/YAP通路在乳腺癌发生发展中的作用和机制

Hippo/YAP pathway plays a critical role in tumor initiation and progression. However, the mechanisms of regulating YAP expression and activity in breast cancer tumorigenesis remains to be further investigated. It has been reported that SAM and SH3 domain containing1 (SASH1), a tumor suppressor, is correlated with poor survival of breast cancer patients, although the underlying mechanism is largely unknown. Our preliminary data showed that SASH1 is a binding partner of LATS2 and ectopic expression of SASH1 in breast cancer cells reduces the expression of YAP. Based on these observations, we hypothesize that SASH1 regulates the Hippo/YAP pathway and contributes to breast cancer oncogenesis and progression. To further investigate the possibility, we will dissect the molecular mechanism underlying the interaction between SASH1 and LATS2 and the mechanism of the SASH1-LATS2 complex that regulates YAP expression. Furthermore, we will investigate the difference between the SASH1-LATS2 complex and the canonical LATS2 complex. In the functional assay, by knocking down SASH1 or overexpressing SASH1 and its mutants in breast cancer cells and mouse models, we will investigate the role of SASH1 regulating the Hippo/YAP pathway during breast cancer development. Finally, the expression of SASH1 and the Hippo/YAP pathway proteins will be examined in the breast cancer tissue samples and the correlation of SASH1/LATS2/YAP signaling and breast cancer grade will be also analyzed. Together, we aim to elucidate the function and molecular mechanism of tumor suppressor SASH1 in tumorigenesis with the implication of a new target in the diagnosis and treatment of breast cancer.

Hippo/YAP通路在肿瘤发生发展中起重要作用，但乳腺癌中YAP表达及活性的调控机制尚未充分阐明。已有报道提示抑癌蛋白SASH1与乳腺癌预后不良相关，然而分子机制不详。申请人的预研究发现: SASH1结合LATS2，在乳腺癌细胞中过表达SASH1下调YAP的蛋白水平。据此，申请人提出如下假说：SASH1通过调控Hippo/YAP通路发挥其抑癌作用。本研究拟阐明SASH1与LATS2相互作用及其调控YAP表达的的分子机制，探讨SASH1-LATS2复合体与Hippo/LATS2经典复合体的共性与区别；利用敲低或过表达方法并结合细胞和裸鼠乳腺癌模型研究SASH1调控Hippo/YAP通路在乳腺癌发生发展中的作用和细胞分子机理；并利用乳腺癌临床标本，结合临床资料，分析SASH1、Hippo/YAP等相关蛋白的表达和相关性，评价该通路的临床意义。该研究结果可能为乳腺癌的诊疗提供新思路和药物靶点。

乳腺癌严重危害妇女身心健康，全球乳腺癌发病率仍旧高居女性恶性肿瘤发病率第一位，死亡率排第二位。三阴性乳腺癌缺乏孕激素受体、雌激素受体和人表皮生长因子受体2的表达，目前尚无有效的靶向治疗药物。因此，寻觅用于治疗三阴性乳腺癌的新型靶点十分迫切。.SASH1（SAM和SH3结构域1）是SLY信号转接器蛋白家族的成员，抑制多种肿瘤细胞迁移和侵袭。SASH1在肝转移性结肠癌和侵袭性乳腺癌等多种肿瘤组织中表达较低，且低表达SASH1与肿瘤预后不良密切相关。有研究对30例三阴性乳腺癌的SNP标记筛选发现，SASH1是三阴性乳腺癌的一个重要遗传风险因子，但尚不清楚SASH1在三阴性乳腺癌中的抑癌作用机制。我们发现与其他类型乳腺癌相比，TNBC患者样本中的SASH1显著下调。过表达SASH1抑制三阴性乳腺癌细胞侵袭，而敲低SASH1促进三阴性乳腺癌细胞侵袭，同时伴有MMP2和MMP9的表达下调。在鸡胚绒毛尿囊膜体外侵袭模型和小鼠尾静脉肺转移模型中进一步证实了敲低SASH1的作用。在机制上，敲低SASH1激活Akt，促进CyclinD1的表达。此外，敲低SASH1下调Hippo激酶LATS1及其效应子YAP（Yes相关蛋白）的磷酸化水平，从而上调YAP及其下游靶基因CYR61的表达，而过表达SASH1则表现出相反的作用。重要的是，采用抑制剂抑制Akt或YAP活性，或者敲低YAP，可以显著抑制敲低SASH1引起的三阴性乳腺癌细胞侵袭。此外，SASH1调控YAP依赖于LATS1活性，且与MST1/2、LATS1/2和Ajuba相关。再者，SASH1可能被LATS1磷酸化，而SASH1 S407A突变体（磷酸化缺陷）不能回复敲低SASH1介导的YAP活性变化。综上所述，我们的研究发现SASH1可能通过Hippo/YAP通路调控三阴性乳腺癌细胞侵袭，这可以作为治疗干预的新靶点。