臺灣博碩士論文加值系統

利用螢光光譜作為疾病診斷的方法，可分成自體螢光及染料螢光法兩種。其中，自體螢光法可以根據其型態及內在光感物質的變化，直接反應生物體的病變情形，但是自體螢光法相當受限於儀器的靈敏度及解析度，因此目前發展螢光光譜作為疾病診斷多是利用靈敏度及解析度均較佳的染料螢光法。染料螢光法是使用外加方式投予光感物質，利用光感物質可以侷限在疾病部位的特性，經特殊波長的光激發而產生螢光，以進行臨床診斷。5-aminolevulinic acid （ALA）5-胺基酮戊酸是目前最常被用來作為光動力診斷（Photodynamic Diagnosis；PDD）的藥物之一，它是光感物質的前驅物，在細胞內代謝生成的主要螢光物質為Protoporphyrin IX （PpIX）。由文獻報告得知利用口服或注射的方法投予ALA到體內，進行光動力診斷，在臨床上已經有不錯的成果，但是相較於局部投予方式（local adminiatration），使用此種全身性的投予方法（systemic administration），所需的ALA劑量較高，相對在全身所產生的PpIX總量也較多，因此也比較容易引起光敏感甚至其他的副作用。
在本實驗中，我們利用ALA-水膠局部塗抹劑型，進行口腔癌前病變及口腔癌的螢光診斷。由於光動力藥物ALA必須要先經過細胞代謝成PpIX後才具有螢光，所以在本劑型中的賦形劑篩選過程，除了考量賦形劑本身的物化特性之外，我們也利用細胞培養方法來確保所選的賦形劑不會阻礙ALA的吸收與PpIX的生成。經過挑選之後，水膠劑型的組成除了ALA之外，還包括為Carbopol 971P （CP971P）及Pluronic F-127（PF-127）。CP971P在此水膠劑型中，主要對口腔提供了黏膜吸附性，而PF-127的作用是藉由溫度的轉變，使此劑型具有從溶液轉變成膠體的可逆性，方便藥劑的配製及塗抹。為了瞭解所選擇的賦形劑在混合後是否產生交互作用，我們利用示差掃瞄式熱卡量計、紅外光吸收光譜來進行評估。我們將此劑型用在經化學物質DMBA誘導癌化的倉鼠口頰內側黏膜組織上，發現使用低劑量的ALA局部塗抹所產生的螢光強度相當於以腹腔注射較高劑量的ALA所獲得之結果。我們並依據螢光光譜檢測及病理切片的結果，進行劑型的修飾。目前已有一個劑型正在進行學術臨床試驗。

Two kinds of spectrofluorometric methods are currently under investigation for disease diagnosis: autofluorescence and photosensitizer fluorescence. Autofluorescence may directly reveal the biochemical as well as morphological changes, and its application in cancer diagnosis is mainly restricted to the instrumental sensitivity and resolution. Photosensitizer fluorescence, on the other hand, generates better signal to noise ratio since more photosensitizer is located at the disease site. 5-aminolevulinic acid （ALA）is one of the most popular chemicals for photodynamic diagnosis （PDD）. ALA is a photosensitizer precursor, and the strongest fluorescent photosensitizer produced after dosing ALA is protoporphyrin IX （PpIX）. ALA is usually administered orally or by intravenous injection into the body. Based on the literatures, high ALA dosing will generate more PpIX, and higher risk of side effects such as photosensitivity.
In this study, we have developed a new topical dosage form containing ALA for the diagnosis of oral cancers and premalignant lesions. Excipients were selected using a cell culture system to ensure that they will not interfere with the PpIX production. Two excipients, Carbopol 971P （CP971P） and Pluronic F-127 （PF-127）, were used in the formulation of the ALA gel. CP971P provides the main bioadhesiveness of the formulation to the oral cavity, and PF-127 gives the formulation a solution-gelling phase-transition character. We use thermal analysis technique and IR spectroscopy to evaluate whether there are any unexpected interactions in the excipient mixture. The ALA gel has been tested in a hamster pouch model. Lower dose of topical ALA has shown to generate the fluorescence intensity comparable to or stronger than higher dose of ALA intraperitoneal injection in hamsters. The formulation modification was based on the fluorescence spectra and pathological results using various disease stages of hamsters. One of the formulations is currently under academic clinical trials.

縮寫表…………………………………………………………………..IV
圖目錄…………………………………………………………………..VI
表目錄…………………………………………………………………..IX
英文摘要………………………………………………………………..X
中文摘要 ……………………………………………………………..XII
壹、緒論…………………………………………………………………1
貳、實驗
第一部份--細胞實驗………………………………………….11第二部分─劑型設計 …………………………………….…16
第三部份－動物實驗…………………………………………21
第四部份－臨床試驗…………………………………………26
參、結果
第一部份--細胞實驗 ……………………………………….28
第二部分─劑型設計 …………………………………….…33
第三部份－動物實驗 ………………………………………39
第四部份－臨床試驗 ………………………………………54
肆、結論…………………………………………………………………57
伍、參考文獻……………………………………………………………58
附錄
I、 血紅素生合成途徑的補充說明…………………………72
II、ALA生成PpIX的細胞實驗中條件的建立……………75
III、以Rhodamin 123（R123）替代ALA，藉以探討賦形劑與P-glycoprotein的關係………………………………77
IV、以致癌物誘發倉鼠產生不同程度口腔病變的動物模式78
V、以示差掃瞄式熱卡量計測量Carbopol、PF-127及其混合物…………………………………………………………81
VI、Carbopol、PF-127及其混合物的紅外光吸收光譜……92
VII、ALA分析方法及水膠中ALA安定性的量測…………97

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