Abstract
摘要:免疫治療對於轉移性乳癌治療效果不佳。嘗試加上不同的免疫調節劑是目前試圖改善免疫療法的重要研究方向。癌細胞會演化出多種機制以逃避被免疫細胞識別或影響調節免疫細胞功能。最近研究顯示上皮-間質轉化(EMT)會降低免疫作用細胞對癌細胞的攻擊活性﹑改變腫瘤微環境,以利於腫瘤細胞存活。乳癌為一異質性疾病,EMT和間質-上皮轉化(MET)往往處在動態改變的狀態。我們透過培養metaplastic 乳癌患者的腫瘤細胞,建立了具有原發性(de novo)EMT/MET動態變化的細胞株模型。親代培養細胞中混合著E-cadherin(E-cad)陽性以及vimentin(vim)陽性的腫瘤細胞。透過篩選,我們進一步建立了一系列富含上皮分子生物表徵的HE細胞株(E-cad + / vim-)和富含間質分子生物表徵的HM細胞株(E-cad- / vim+)。功能測定顯示HM細胞的移行(migration)和侵犯能力增加, 機轉研究顯示ZEB1的表現是決定EMT變化的關鍵。透過該模型進行實驗,我們發現:1.促炎性巨噬細胞(proinflammatory macrophages)僅針對上皮型﹑而非間質型乳癌細胞具有細胞毒殺和抑制增殖作用。2.促炎性巨噬細胞對間質型乳癌細胞的無害性與ZEB1訊號路徑相關。3.相較於間質型癌細胞,上皮型癌細胞更易活化促炎性巨噬細胞,進而發生促炎性巨噬細胞誘導的凋亡式細胞死亡。4.間質型﹑而非上皮型癌細胞對Jurkat細胞(具細胞毒殺T細胞功能的白血病細胞株)具有較強的免疫抑制功能。5.上述實驗結果可藉以MCF7細胞作為上皮型﹑而MB231細胞代替為間質型乳癌細胞重複被驗證。依此,我們假說:1.與間質型乳癌細胞相比,上皮型乳癌細胞更易受先天性(innate)以及適應性(adaptive)免疫所影響;2.細胞的EMT狀態可被調整(polarized)以改變對先天性和適應性免疫的敏感性。具體目標:1.測試上皮型乳癌細胞(與間質型相比),是否更能活化促炎性巨噬細胞﹑NK細胞﹑和T淋巴細胞;2.了解使間質型乳腺癌細胞不受促炎性巨噬細胞﹑NK細胞﹑和T淋巴細胞影響的可能原因;3.測試我們是否可以通過操控乳癌細胞的EMT狀態來增強癌症免疫監控。此研究的結果將有助於我們了解免疫細胞和處於不同EMT狀態的乳癌細胞之間的相互作用,而這將可能是增進乳癌免疫治療效果的關鍵步驟。
Abstract: Metastatic breast cancer is generally associated with lower response rates to immunotherapy. In order to enhance the efficacy of immunotherapies on breast tumors, combinations of immunotherapy with various immune modulating agents are being pursued. Cancer cells evolved multiple mechanisms to escape immune recognition or to modulate immune cell function. Recently, it has been suggested that epithelial-mesenchymal transition (EMT) may decrease the susceptibility of cancer cells to immune effector cells, and enable cancer cells to refine the tumor microenvironment in favor of their survival. Breast cancer is a heterogeneous disease with dynamic EMT and mesenchymal-epithelial transition (MET) status. We have established a cell line model with de novo EMT/MET chronological changes through a primary tumor culture from a patient with metaplastic breast carcinoma. The parental primary culture cells are composed of mixture of E-cadherin-positive and vimentin-positive tumor cells. We have further established epithelia-enriched HE clones (predominant in E-cad+/vim- cells), and mesenchymal-enriched HM clones (predominant in E-cad-/vim+ cells). Functional assay demonstrated increased migration and invasion in HM cells. The detailed mechanical research demonstrated that ZEB1 expression is the key to determine the EMT change. By using this model, we found that: 1) THP-1-derived proinflammatory macrophages has cytotoxicity effect and proliferation inhibitory effect on the epithelial-type, but not the mesenchymal-type breast cancer cells; 2) the invulnerability of mesenchymal-type breast cancer cells to proinflammatory macrophages was correlated with ZEB1 signaling pathway; 3) compared to the mesenchymal-type breast cancer cells, the epithelial-type cells enhanced the activation of proinflammatory macrophages, and were more likely to go through proinflammatory macrophage-induced apoptotic cell death; 4) the mesenchymal-type, rather than the epithelial-type breast cancer cells, had stronger immunosuppressive functions towards Jurkat cells, a T cell leukemia cell line exerting cytotoxic T cell function; 5) aforementioned experiment could be repeated when using MCF7 cells as epithelial-type and MB231 cells as mesenchymal-type breast cancer cells. We hypothesized that: 1) Compared to the mesenchymal-type breast cancer cells, the epithelial-type breast cancer cells are more vulnerable to activate proinflammatory macrophage- and NK cell-associated innate immunity and T lymphocyte-oriented adaptive immunity; 2) The EMT states of breast cancer cells can be polarized into a state which are more susceptible to innate and adaptive immunity.Specific Aims: 1) to examine whether the epithelial-type, compared to the mesenchymal-type breast cancer cells, are more likely to activate proinflammatory macrophages, NK cells, and T lymphocytes in vitro and in vivo; 2) to understand what makes the mesenchymal-type breast cancer cells invulnerable to proinflammatory macrophages, NK cells, and T lymphocytes in vitro and in vivo; 3) to test whether we can enhance cancer immunosurveillance by manipulating EMT/MET states of breast cancer cells in vitro and in vivo.The result of this study will help us to decipher the interactions between immune cells and breast cancer cells in different EMT states, which may be a key step to improve immunotherapeutic effects on breast cancer.
Keyword(s)
上皮-間質轉化
乳癌
先天與後天免疫系統
免疫監控
免疫治療
epithelial-mesenchymal transition
breast cancer
innate and adaptive immunity
immunosurveillance
immunotherapy