口服卡介苗研發之體內及體外分析評估   全文下載

In Vitro and In Vivo Assessments for Developing an Oral BCG Vaccine Formulation

本研究目的是建立體外分析方法以評估所研製的口服卡介苗（BCG）疫苗配方。此外，所研究的配方也經由天竺鼠實驗進行動物免疫反應之評估，以測定在結核病之可能保護效果。BCA（bicinchoninic acid）測定BCG之蛋白質方法也被發展出應用於快速測定BCG疫苗之含量。此法使用了含8% SDS（sodium dodecyl sulfate）及0.08 N NaOH之BCA試劑在45°C測定，於150~1000 μg/mL濃度所得到之精密度（變異係數 < 3.5%）及準確度（誤差 -3.6~2.6%）皆在可接受範圍。體外試驗模擬藥物在腸胃道生理狀況下，顯示所研究之口服配方能在低pH（0.1 N HCl）提供保護，但在較高的pH（磷酸鹽緩衝液pH 6.8）則可以釋出活性成分。所研製之口服BCG疫苗製劑在誘發IgG之免疫反應較弱。但在結核菌素PPD（purified protein derivative）之天竺鼠試驗，顯示口服BCG疫苗製劑4 mg可達到與皮下注射0.5 mg相同或較佳的反應。本研究之體外及體內測定法也可應用於口服BCG疫苗製劑之進一步開發。

The aim of this study was to establish in vitro analytical methods for assessing an investigated oral BCG vaccine formulation. In addition, in vivo immune response was also evaluated in guinea pigs by determining the potentially protective effect of the investigated formulation on tuberculosis. The bicinchoninic acid (BCA) method for BCG protein quantitation was developed for rapid measuring the content of BCG vaccine in the investigated formulation. The analytical method was conducted at 45°C to offer good precision (coefficient of variation <3.5%) and accuracy (error: -3.6~2.6%) in the range of 150~1000 μg/mL of BCG vaccine concentrations. In the simulation study of drug release from the investigated formulation in the gastrointestinal tract under physiological conditions, the investigated oral formulation could protect active component at lower pH (0.1 N HCl) and completely release active component at higher pH (phosphate buffer, pH 6.8). Although the investigated oral BCG vaccine formulation showed weak immune response to elicit IgG titer, the PPD (purified protein derivative) skin test in guinea pigs by oral dosing of 4 mg BCG vaccine formulation acquired about equal or higher response than that by 0.5 mg dose by subcutaneous vaccination. The analytical method and in vivo approach could be further applied in the development of oral BCG vaccine formulations.