Department of Biochemistry and Biophysics, 2128 Texas A&M University, College Station, TX 77843-2128, USA.
J Control Release. 2010 Sep 15;146(3):318-25. doi: 10.1016/j.jconrel.2010.06.003. Epub 2010 Jun 11.
A catalytic bioscavenger with broad substrate specificity for the therapeutic and prophylactic defense against recognized chemical threat agents has been a long standing objective of civilian and military research. A catalytic bioscavenger utilizing the bacterial enzyme organophosphorus hydrolase (OPH) is characterized in these studies, and has potential application for both military and civilian personnel in threat scenarios involving either nerve agents or OP pesticides. The present study examines the effects of PEGylation on the biochemical and pharmacological characteristics of OPH. The enzyme was conjugated with linear and branched methyl-PEO(n)-NHS esters of relatively small molecular mass from 333 to 2420Da. PEGylated OPH displayed a decreased maximal catalytic rate, though substantial activity was maintained against two tested substrates: up to 30% with paraoxon and up to 50-60% with demeton-S. The thermostability of the PEGylated enzymes ranged between 60 and 64 degrees C, compared to the unmodified OPH, which is approximately 67 degrees C. The enzyme conjugates revealed a significant improvement of pharmacokinetic properties in animal studies. The clearance from a guinea pig's blood stream significantly decreased relative to unmodified OPH, resulting in an increase of residence time and systemic availability. Evaluation of the humoral immune response indicated that the branched PEG-OPH conjugate significantly reduced production of anti-OPH antibodies, compared to the unmodified enzyme. The OPH-PEG conjugates with improved pharmacokinetic and immunogenicity properties, considerable catalytic activity and thermal stability provide a new opportunity for the in vivo detoxification of the neurotoxic OP compounds.
一种对已识别的化学威胁剂具有广泛底物特异性的催化生物清除剂，一直是民用和军用研究的长期目标。本研究中，利用细菌酶有机磷水解酶（OPH）的催化生物清除剂具有特征，并且在涉及神经毒剂或 OP 农药的威胁情况下，对军事和民用人员都具有潜在应用。本研究探讨了聚乙二醇化对 OPH 的生化和药理特性的影响。该酶与相对较小分子量的线性和支链甲基-PEO(n)-NHS 酯（333 至 2420Da）进行缀合。PEG 化 OPH 的最大催化速率降低，但对两种测试底物仍保持相当大的活性：对 paraoxon 高达 30%，对 demeton-S 高达 50-60%。与未修饰的 OPH 相比，PEG 化酶的热稳定性在 60 至 64°C 之间，而未修饰的 OPH 约为 67°C。酶缀合物在动物研究中显示出显著改善的药代动力学特性。与未修饰的 OPH 相比，从豚鼠血液清除的清除率显著降低，导致驻留时间和全身可用性增加。对体液免疫反应的评估表明，与未修饰的酶相比，支链 PEG-OPH 缀合物显著减少了抗 OPH 抗体的产生。具有改善的药代动力学和免疫原性特性、相当大的催化活性和热稳定性的 OPH-PEG 缀合物为体内解毒神经毒性 OP 化合物提供了新的机会。
