Business & Portfolio Development Unit of SIFI SpA, Aci Sant'Antonio, Italy.
Cornea. 2020 Jul;39(7):892-897. doi: 10.1097/ICO.0000000000002308.
Acanthamoeba keratitis is a rare ocular disease caused by Acanthamoeba spp. The current Acanthamoeba keratitis treatments consist of unlicensed drugs, most commonly polihexanide (PHMB), a biguanide derivative, either as monotherapy or combined with a diamidine. The main purpose of this study was to develop an in vitro bioluminescence assay able to differentiate the antiamoebic activity of PHMB concentrations (0.02%, 0.04%, 0.06%, and 0.08%). Another objective of this study was to evaluate the antiamoebic activity of equal PHMB concentrations at different molecular weights (MWs).
Adenosine triphosphate (ATP) bioluminescence assay applicability was established by a linear correlation between amoebae number and relative light unit emitted from cysts. The protocol consisted in a series of sequential tests, the first of which compared relative light unit emission and the hemocytometric method after cysts exposure to the different PHMB concentrations. In this assay, dilutions of the initial PHMB concentrations were used as working solutions. Usually, only 5% or less of the instilled dose into the eye is distributed through the cornea; thus, the antiamoebic activity of initial PHMB concentrations 1/10 and 1/100 diluted can reasonably be considered as representative of their undiluted dose administered in vivo.
There was no significant difference between the 2 methods. The ATP bioluminescence assay ranked the antiamoebic activity of the working PHMB solutions and showed that the viability of the cysts was equally reduced at fixed PHMB concentration in a wide range of molecular weight.
The ATP bioluminescence assay showed to be a simple and rapid way for characterizing the in vitro antiamoebic activity of working PHMB solutions at different concentrations.
棘阿米巴角膜炎是一种由棘阿米巴属引起的罕见眼部疾病。目前的棘阿米巴角膜炎治疗方法包括未获许可的药物，最常见的是聚己双胍（PHMB），一种双胍衍生物，无论是单独使用还是与脒类联合使用。本研究的主要目的是开发一种能够区分 PHMB 浓度（0.02%、0.04%、0.06%和 0.08%）抗阿米巴活性的体外生物发光检测法。本研究的另一个目的是评估不同分子量（MW）的等 PHMB 浓度的抗阿米巴活性。
通过囊泡内的腺苷三磷酸（ATP）生物发光检测法与相对光单位（RLU）之间的线性相关性来建立可行性。该方案由一系列连续的测试组成，第一个测试比较了暴露于不同 PHMB 浓度后的相对光单位（RLU）发射和血球计数法。在该检测中，初始 PHMB 浓度的稀释液被用作工作溶液。通常，只有 5%或更少的滴眼剂量分布在角膜中；因此，初始 PHMB 浓度的 1/10 和 1/100 稀释液的抗阿米巴活性可以合理地被认为是代表其在体内给药的未稀释剂量。
两种方法之间没有显著差异。ATP 生物发光检测法对工作 PHMB 溶液的抗阿米巴活性进行了排序，并显示在固定 PHMB 浓度范围内，广泛的分子量范围内，囊泡的活力都同样降低。
ATP 生物发光检测法是一种简单、快速的方法，可以用于表征不同浓度的工作 PHMB 溶液的体外抗阿米巴活性。
