14-3-3zeta in ErbB2 Mediated Mammary Carcinogenesis

描述（由申请方提供）：ErbB 2经常在高级别人类DCIS中检测到（50-60%），并且在80-85%的非侵袭性、癌前病变、粉刺型DCIS肿瘤中发现ErbB 2基因扩增。 有趣的是，ErbB 2在25-30%的浸润性乳腺癌中过表达。 目前尚不清楚一些ErbB 2过表达的DCIS如何发展成浸润性导管癌（IDC）。 在模拟乳腺腺泡形成的体内条件的三维（3D）培养物中生长的MCF 10A乳腺上皮细胞（10A.B2 MEC）中ErbB 2的急性激活导致产生缺乏与DCIS相关的结构特性的侵入性特性的多腺泡结构。 因此，激活的ErbB 2诱导MEC的早期转化，但不足以诱导侵袭行为。 据推测，额外的遗传/分子事件或更高水平的ErbB 2需要MEC获得侵入性。 然而，激活的ErbB 2诱导MEC早期转化的机制以及它们获得侵袭性特性所涉及的事件尚不清楚。 有趣的是，我们发现14-3- 3 zeta是一种参与许多重要细胞过程的蛋白质，在大约30%的乳腺癌患者的DCIS病变中与ErbB 2高水平共表达，并且与浸润性和转移性疾病的复发相关。 在10A.B2 MEC中通过逆转录病毒感染增加14-3- 3 zeta表达导致3D培养中的侵袭性多腺泡结构，与载体对照中缺乏侵袭形成对比。 另一方面，通过siRNA阻断10A.B2细胞中的14-3- 3 zeta表达抑制了它们向多腺泡结构的发育，并且用14 - 3 - 3 zeta干扰突变体阻断14 - 3 - 3 zeta功能导致抑制NIH 3 T3细胞中ErbB 2介导的转化。 因此，我们假设14-3- 3 zeta有助于ErbB 2介导的乳腺癌发生，14-3- 3 zeta高表达赋予ErbB 2激活的MEC侵袭潜力，这可能促进ErbB 2过表达的DCIS向IDC的进展。 在本提案中，我们将研究14-3- 3 zeta在ErbB 2激活的MEC转化和获得侵袭性中的作用和机制（目的1和2），并确认14-3- 3 zeta在患者中ErbB 2过表达DCIS向IDC进展中的作用（目的3）。 这些全面的方法将确定14 - 3 - 3 zetain ErbB 2介导的乳腺癌发生和侵袭的作用，并最终在ErbB 2过表达DCIS向IDC的进展中的作用。 这项研究将为早期干预、诊断和治疗带来新的见解，可能使ErbB 2和14-3- 3 zeta过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.