Lofgren James A, Dhandapani Sripriya, Pennucci Jason J, Abbott Christina M, Mytych Daniel T, Kaliyaperumal Arunan, Swanson Steven J, Mullenix Michael C
Department of Clinical Immunology, Amgen, Thousand Oaks, CA 91320, USA.
J Immunol. 2007 Jun 1;178(11):7467-72. doi: 10.4049/jimmunol.178.11.7467.
Evaluation of the immunogenicity of panitumumab, a fully human anti-epidermal growth factor receptor mAb approved for use in colorectal cancer patients, led to the development of two separate immunoassays for the detection of anti-panitumumab Abs. The first immunoassay used a bridging ELISA capable of detecting 10 ng/ml positive control anti-panitumumab Ab. The ELISA incorporated an acid dissociation step to reduce drug interference and tolerated the presence of approximately 100-fold molar excess of drug. During eight clinical trials, the ELISA detected developing Ab responses in 2 of 612 (0.3%) subjects. In one of the ELISA positive subjects, neutralizing Abs were detected using an epidermal growth factor receptor phosphorylation bioassay. The second immunoassay used a Biacore biosensor immunoassay format capable of detecting 1 mug/ml positive control Ab while tolerating the presence of equal molar amounts of drug. Although less sensitive and less tolerant to competing drug in the assay, the Biacore assay detected developing Ab responses in 25 of the 604 (4.1%) subjects. Additionally, the Biacore assay identified eight subjects who developed neutralizing Abs. Mouse mAbs with affinities ranging from 1.1 x 10(-6) to 8.4 x 10(-10) M were used to characterize both assay types. The ELISA was more sensitive for the detection of higher affinity mAbs and detected high-affinity mAbs in the presence of higher molar ratio of drug to mAb. The Biacore assay was more sensitive for detection of lower affinity mAbs and detected low affinity Abs in the presence of higher molar ratios of drug to mAb.
帕尼单抗是一种已被批准用于治疗结直肠癌患者的全人源抗表皮生长因子受体单克隆抗体，对其免疫原性的评估促使开发了两种用于检测抗帕尼单抗抗体的独立免疫测定法。第一种免疫测定法采用了一种桥联ELISA，能够检测浓度为10 ng/ml的阳性对照抗帕尼单抗抗体。该ELISA法包含一个酸解离步骤以减少药物干扰，并且能够耐受大约100倍摩尔过量的药物存在。在八项临床试验中，该ELISA法在612名受试者中的2名（0.3%）检测到了抗体反应的产生。在其中一名ELISA法检测为阳性的受试者中，使用表皮生长因子受体磷酸化生物测定法检测到了中和抗体。第二种免疫测定法采用了Biacore生物传感器免疫测定形式，能够检测浓度为1 μg/ml的阳性对照抗体，同时耐受等量摩尔药物的存在。尽管该Biacore测定法在检测中灵敏度较低且对竞争性药物的耐受性较差，但在604名受试者中的25名（4.1%）检测到了抗体反应的产生。此外，Biacore测定法还鉴定出了八名产生中和抗体的受试者。使用亲和力范围为1.1×10⁻⁶至8.4×10⁻¹⁰ M的小鼠单克隆抗体对两种测定类型进行了表征。ELISA法在检测高亲和力单克隆抗体时更为灵敏，并且在药物与单克隆抗体摩尔比更高的情况下能够检测到高亲和力单克隆抗体。Biacore测定法在检测低亲和力单克隆抗体时更为灵敏，并且在药物与单克隆抗体摩尔比更高的情况下能够检测到低亲和力抗体。
