开发检测体系，以高特异性测定单次吸入治疗后自身免疫性肺泡蛋白沉积症患者体内的沙莫司汀浓度。
Assay system development to measure the concentration of sargramostim with high specificity in patients with autoimmune pulmonary alveolar proteinosis after single-dose inhalation.
机构信息
Bioscience Medical Research Center, Niigata University Medical & Dental Hospital Niigata, Japan.
Clinical and Translational Research Center, Niigata University Medical & Dental Hospital Niigata, Japan.
出版信息
J Immunol Methods. 2018 Sep;460:1-9. doi: 10.1016/j.jim.2018.05.012. Epub 2018 Jul 9.
During a clinical trial of a Saccharomyces cerviciae-derived recombinant human granulocyte-macrophage colony stimulating factor (rhGM-CSF), sargramostim, in patients with autoimmune pulmonary alveolar proteinosis (aPAP), we conducted a pharmacokinetic study of single-dose sargramostim inhalation. Several problems were encountered whereby sargramostim formed an immune-complex with GM-CSF autoantibodies (GMAbs) immediately after entering the body; thus, we could not measure the concentration of sargramostim using a commercial high sensitivity enzyme-linked immunosorbent assay (ELISA). Moreover, the ELISA could not discriminate inhaled sargramostim from intrinsic GM-CSF. To solve these problems, we developed a novel ELISA system with a capture antibody that is specific for sargramostim and a detection antibody capable of binding with GM-CSF. This system quantified the serum sargramostim concentration, but not E. coli-, CHO-, or HEK293T-derived human recombinant GM-CSF. Using this system, serum pharmacokinetics were estimated in five patients after inhalation of 250 μg sargramostim, with a mean Cmax of 9.7 ± 2.85 pg/ml at a Tmax of 2 ± 1.22 h.
在一项针对自身免疫性肺泡蛋白沉积症（aPAP）患者使用来源于酿酒酵母重组人粒细胞-巨噬细胞集落刺激因子（rhGM-CSF），沙格司亭（sargramostim）的临床试验中，我们进行了单次吸入沙格司亭的药代动力学研究。在试验中发现了一些问题，即在进入人体后，沙格司亭会立即与 GM-CSF 自身抗体（GMAbs）形成免疫复合物；因此，我们无法使用商业高灵敏度酶联免疫吸附测定法（ELISA）来测量沙格司亭的浓度。此外，ELISA 无法区分吸入的沙格司亭和内源性 GM-CSF。为了解决这些问题，我们开发了一种新的 ELISA 系统，该系统使用一种针对沙格司亭的特异性捕获抗体和一种能够与 GM-CSF 结合的检测抗体。该系统可以定量血清中的沙格司亭浓度，但不能定量 E. coli、CHO 或 HEK293T 来源的人重组 GM-CSF。使用该系统，在 5 名患者吸入 250μg 沙格司亭后，估计了血清药代动力学，在 Tmax 为 2±1.22h 时，Cmax 为 9.7±2.85pg/ml。
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