Office of Biotechnology Products, Office of Pharmaceutical Quality, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, Maryland, United States of America.
Office of Clinical Pharmacology, Office of Translational Sciences, Center for Drug Evaluation and Research, U.S. Food and Drug Administration, Silver Spring, Maryland, United States of America.
Sci Rep. 2020 Feb 12;10(1):2476. doi: 10.1038/s41598-020-59346-z.
PEGylated recombinant human granulocyte colony stimulating factor (pegfilgrastim) is used clinically to accelerate immune reconstitution following chemotherapy and is being pursued for biosimilar development. One challenge to overcome in pegfilgrastim biosimilar development is establishing pharmacokinetic (PK) similarity, which is partly due to the degree of PK variability. We herein report that commercially available G-CSF and PEG ELISA detection kits have different capacities to detect pegfilgrastim aggregates that rapidly form in vitro in physiological conditions. These aggregates can be observed using SDS-PAGE, size-exclusion chromatography, dynamic light scattering, and real-time NMR analysis and are associated with decreased bioactivity as reflected by reduced drug-induced cellular proliferation and STAT3 phosphorylation. Furthermore, individual variability in the stability and detectability of pegfilgrastim in human sera is also observed. Pegfilgrastim levels display marked subject variability in sera from healthy donors incubated at 37 °C. The stability patterns of pegfilgrastim closely match the stability patterns of filgrastim, consistent with a key role for pegfilgrastim's G-CSF moiety in driving formation of inactive aggregates. Taken together, our results indicate that individual variability and ELISA specificity for inactive aggregates are key factors to consider when designing and interpreting studies involving the measurement of serum pegfilgrastim concentrations.
聚乙二醇化重组人粒细胞集落刺激因子（PEG 重组人粒细胞集落刺激因子）临床上用于加速化疗后的免疫重建，目前正在开发其生物类似药。在 PEG 重组人粒细胞集落刺激因子生物类似药开发中需要克服的一个挑战是建立药代动力学（PK）相似性，这部分是由于 PK 变异性的程度。本文报道了市售的 G-CSF 和 PEG ELISA 检测试剂盒在检测体外生理条件下迅速形成的 PEG 重组人粒细胞集落刺激因子聚集体的能力存在差异。这些聚集体可以通过 SDS-PAGE、尺寸排阻色谱、动态光散射和实时 NMR 分析观察到，并且与生物活性降低有关，表现为药物诱导的细胞增殖和 STAT3 磷酸化减少。此外，还观察到 PEG 重组人粒细胞集落在人血清中的稳定性和可检测性存在个体差异。在 37°C 孵育的健康供体血清中，PEG 重组人粒细胞集落刺激因子水平显示出明显的个体变异性。PEG 重组人粒细胞集落刺激因子的稳定性模式与粒细胞集落刺激因子的稳定性模式密切相关，这与 PEG 重组人粒细胞集落刺激因子的 G-CSF 部分在驱动无活性聚集体形成中的关键作用一致。总之，我们的结果表明，个体变异性和针对无活性聚集体的 ELISA 特异性是在设计和解释涉及测量血清 PEG 重组人粒细胞集落刺激因子浓度的研究时需要考虑的关键因素。
