14-3-3zeta in ErbB2 Mediated Mammary Carcinogenesis

描述(申请人提供)：ERBB2经常在高级别的人类DCIS中检测到(50%-60%)，并且在80%-85%的非侵袭性、癌前、粉刺样型DCIS中发现ErbB2基因扩增。有趣的是，ErbB2在25%-30%的浸润性乳腺癌中过度表达。目前尚不清楚一些过表达ErbB2的DCIS如何发展为浸润性导管癌(IDC)。在三维(3D)培养中生长的MCF10A乳腺上皮细胞(10A.B2MECs)中ErbB2的急性激活导致了缺乏与DCIS相关的结构特性的侵袭性结构的产生。因此，激活的ErbB2诱导MEC的早期转化，但不足以诱导侵袭行为。据推测，MEC需要更多的遗传/分子事件或更高水平的ErbB2才能获得侵袭性。然而，激活的ErbB2诱导微血管内皮细胞早期转化的机制以及与其获得侵袭性相关的事件尚不清楚。有趣的是，我们发现参与许多重要细胞过程的蛋白质14-3-3zeta与ErbB2在大约30%的乳腺癌患者的DCIS病变中高水平共表达，并与侵袭和转移疾病的复发有关。在10A.B2MEC中，逆转录病毒感染增加了14-3-3zeta的表达，导致3D培养中有侵袭性的多腺泡结构，而载体对照组没有侵袭性。另一方面，siRNA阻断10A.B2细胞中14-3-3zeta的表达，抑制其向多腺泡结构的发育，并用14-3-3zeta干扰突变体阻断14-3-3zeta的功能，从而抑制ErbB2介导的NIH3T3细胞转化。因此，我们推测14-3-3zeta参与了ErbB2介导的乳腺癌的发生，14-3-3zeta的高表达为ErbB2激活的MEC提供了侵袭潜能，可能促进了ErbB2过表达的DCIS向IDC的进展。在本研究中，我们将研究14-3-3zeta在ErbB2激活的微血管内皮细胞(AIMS 1和2)转化和获得侵袭力中的作用和机制，并确认14-3-3Zeta在患者从ErbB2过表达的DCIS向IDC进展中的作用(AIMS 3)。这些综合的方法将确定14-3-3zetain ErbB2介导的乳腺癌的发生和侵袭，并最终在ErbB2过表达的DCIS向IDC的进展中所起的作用。这项研究将为早期干预、诊断和治疗带来新的见解，可能使ErbB2和14-3-3zeta过度表达肿瘤的高危患者受益。

DESCRIPTION (provided by applicant): ErbB2 is frequently detected in high-grade human DCIS (50-60%) and ErbB2 gene amplification has been found in 80-85% of the non-invasive, premalignant, comedo-type DCIS tumors. Intriguingly, ErbB2 is overexpressed in 25-30% of invasive breast cancers. It is not clear how some of the ErbB2 overexpressing DCIS develop into invasive ductal carcinoma (IDC). Acute activation of ErbB2 in MCF10A mammary epithelial cells (10A.B2 MECs) grown in a three dimensional (3D) culture that simulates in vivo conditions of acini formation in the mammary gland has led to the generation of multi-acinar structures lacking invasive properties that share structural properties associated with DCIS. Thus, activated ErbB2 induces the early transformation of MEC but is not sufficient to induce invasive behavior. It was postulated that additional genetic/molecular events or much higher levels of ErbB2 are needed for MECs to acquire invasive properties. However, the mechanisms by which activated ErbB2 induces early transformation of MECs and the events involved in their acquisition of invasive properties are not clear. Interestingly, we found that 14-3-3zeta, a protein that participates in many important cellular processes, is co-expressed at high levels with ErbB2 in approximately 30% of DCIS lesions of breast cancer patients and is associated with recurrence of invasive and metastatic diseases. Increasing 14-3-3zeta expression by retroviral infection in the 10A.B2 MECs led to invasive multi-acinar structures in 3D culture in contrast to the lack of invasion in the vector controls. On the other hand, blocking 14-3-3zeta expression in the 10A.B2 cells by siRNA inhibited their development into multi-acinar structures and blocking 14-3-3zeta function with a 14 -3-3zeta interfering mutant led to the inhibition of ErbB2-mediated transformation in NIH3T3 cells. Therefore, we hypothesize that 14-3-3zeta contributes to ErbB2-mediated mammary carcinogenesis and 14-3-3zeta high expression confers invasive potential in ErbB2-activated MECs that may facilitate the progression of ErbB2-overexpressing DCIS to IDC. In this proposal, we will investigate the role and the mechanisms of 14-3-3zeta in the transformation and acquisition of invasiveness of ErbB2 activated MECs (Aims 1 and 2) and confirm the role of 14-3-3zeta in the progression of ErbB2-overexpressing DCIS to IDC in patients (Aim 3). These comprehensive approaches will define the role of 14 -3-3zetain ErbB2-mediated mammary carcinogenesis and invasion, and ultimately in the progression of ErbB2-overexpressing DCIS to IDC. The study will bring new insights into early intervention, diagnosis, and treatment that may benefit the high-risk group of patients with ErbB2 and 14-3-3zeta overexpressing tumors.