鑑定自體免疫腦下垂體炎之自體抗原，並建立血清檢驗法與探討其病理機制

Abstract

自體免疫腦下垂體炎(autoimmune hypophysitis)，是一種逐漸引起醫學界重視 的自體免疫疾病。然而目前並不了解它的自體抗原與病理機制。為了解決這些問 題，我們先前成功建立了一個與人類自體免疫腦下垂體炎十分相似的小鼠模式 (Experimental Autoimmune Hypophysitis, EAH)。我們進一步從發病的小鼠腦下 垂體，分離出 B 淋巴球，並建立了 14 株單株抗體。從實驗結果顯示，這些單株抗 體具有下列優勢: 1.同時辨識小鼠與人類腦下垂體特定蛋白，2.所辨識的蛋白是強 勢自體抗原(dominant autoantigen)，3.可將所 辨 識 的 蛋 白質 純 化 下來 ， 供 質 譜 分 析鑑定身分。在這個計畫中，我們的目標是找出自體免疫腦下垂體炎的自體抗原， 研發血清檢驗法，並探討這個疾病的病理機制。我們將由以下的策略來達成這些目 標: 一、 以我們建立的單株抗體來純化它們所辨識的蛋白，以質譜學技術(MALDI-TOF) 鑑定自體抗原，並以小鼠模式驗證所找出來自體抗原是否具有致病性。 二、檢驗人類病患的血清對自體抗原的辨識程度，建立自體免疫腦下垂體炎血清 檢驗法。 三、以小鼠模式研究自體抗原引發自體免疫腦下垂體炎的細胞與分子機制。

我們相信成功找出自體抗原，將可作為自體免疫腦下垂體炎的診斷標記，提升診斷 的準確度，更可以用來研究它的病理機制，以便將來設計出一個有效的治療方法， 來改善病患的醫療照護。

Autoimmune hypophysitis (AH) is an increasingly recognized disease of the pituitary. For example, several clinical trials of cancer immunotherapy have recently reported high incidences of AH in participating patients. A better understanding into the pathogenesis of AH will not only improve patient care, but also aid in designing optimal cancer immunotherapy to avoid adverse effects of AH. However the definitive autoantigen(s) and the pathogenesis in AH remain elusive despite many years of research. In order to solve these dilemmas, we have established a mouse model (called Experimental AH; EAH) that faithfully recapitulates the histopathology and the endocrine defects of the human disease. Using the mouse model, we have established 14 monoclonal hybridomas from pituitary-infiltrating B cells. Our preliminary data indicate that the antibodies produced by these monoclonal hybridomas have the following characteristics: (A) the monoclonal antibodies recognize a specific protein in both mouse and human pituitaries, suggesting the autoantigen is conserved between mouse and human, (B) the specific protein is a dominant autoantigen (capable of inducing most robust antibody response) in the mouse and human pituitaries, and (C) the monoclonal antibodies can immunoprecipitate the putative autoantigen from mouse pituitary lysate. Thus, we are confident that the monoclonal antibodies will lead us to identification of the autoantigen(s) involved in AH. Our goals in this study proposal are to identify the disease relevant autoantigen in AH, to develop a serum-based diagnostic test for AH, and to study the pathogenesis by which the autoantigen provokes AH in mouse. Over all we have the following strategy to achieve these goals: 1. Use the monoclonal antibodies to purify the autoantigen from pituitary and then identify the autoantigen by mass spectrometric technology such as MALDI-TOF. 2. Test recognition to the autoantigen in the sera of AH patients as the basis of a serum-based diagnostic test. 3. Study the pathogenesis of AH by which the autoantigen provokes cellular and molecular autoimmune responses.

We believe successful identification of the disease-relevant autoantigen can provide a sound basis for the development of an accurate diagnostic test and, for detailed studies into the pathogenesis in AH. These works are expected to improve the treatment of AH in patients.