Rapid, sensitive and cost-effective determination of immune checkpoint inhibitor activity using a magnetic bead-based
Qinglai Meng1, Yujia Fu1, Shenzhi Li1
1Institute of Biomedical Sciences, The Key Laboratory of Chemical Biology and Molecular Engineering of National Ministry of Education, Shanxi Provincial Key Laboratory of Medical Molecular Cell Biology, Shanxi University, Taiyuan, Shanxi Province 030006, China.
Insights
New magnetic bead assays offer a rapid, sensitive, and cost-effective method for evaluating immune checkpoint inhibitor (ICI) activity. These assays accelerate ICI development and research by overcoming limitations of traditional cell-based methods and ELISA.
Area of Science:
- Immunology
- Biotechnology
- Cancer Therapeutics
Background:
- Immune checkpoint inhibitors (ICIs) are crucial cancer immunotherapeutics.
- Accurate and rapid assessment of ICI blocking activity is vital for development and research.
- Existing methods like cell-based assays and ELISA have limitations including time, cost, and complex preparation.
Purpose of the Study:
- To develop novel, rapid, and sensitive magnetic bead-based binding assays for evaluating ICI activity.
- To compare the performance of these new assays against traditional methods.
- To establish cost-effective alternatives for ICI characterization.
Main Methods:
- Developed two magnetic bead-based assays: soluble ligand/bead immobilized receptor (sL/bR) and soluble receptor/bead immobilized ligand (sR/bL).
- Assessed ICI efficacy by measuring the blockage of ligand-receptor binding.
- Calculated half maximal inhibitory concentration (IC50) values to quantify ICI activity.
Main Results:
- The sL/bR assay accurately determined the activity of TIGIT blocking antibodies, showing results comparable to cell-based assays.
- The sR/bL assay demonstrated significantly enhanced sensitivity for PD-1 blocking antibodies compared to the sL/bR assay.
- Both magnetic bead assays required over 10 times less recombinant protein and had shorter turnaround times than functional ELISA.
Conclusions:
- The developed magnetic bead-based assays are sensitive, rapid, and cost-effective for determining ICI blocking activity.
- These assays offer a significant improvement over traditional methods for ICI evaluation.
- The findings support the utility of these assays in accelerating ICI development and immunological research.
Abstract:
Immune checkpoint Inhibitors (ICIs) are effective immunno-therapeutic agents for cancer. Rapid and sensitive determination of the blocking activity of ICIs is important for ICIs development and immunological research. Among various immune checkpoint (IC) binding assays, cell-based binding assays are widely regarded, and the functional ELISA is a convenient alternative. However, these methodologies are limited by time-consuming preparation of cell lines stably expressing IC molecules, or long turnaround time with high cost. In this study, two magnetic bead based binding assays were developed to evaluate activity of ICIs, which was determined by a soluble ligand/bead immobilized receptor based binding assay (sL/bR binding assay) that assessed efficacy to block binding of one soluble IC ligand on its cognate receptor immobilized beads, or by a soluble receptor/bead immobilized ligand based binding assay (sR/bL binding assay) that assessed efficacy to block binding of soluble IC receptor on its cognate ligand immobilized beads. Half maximal inhibitory concentration (IC50) values of ICIs were calculated to determine ICIs activity. The sL/bR binding assay accurately determined the activity of two TIGIT blocking antibodies, since the relative blocking activity of two TIGIT antibodies determined by the sL/bR binding assay established in this study and that by the cell based binding assay were almost identical. In contrast, the sR/bL binding assay showed significantly improved sensitivity to determine activity of two PD-1 blocking antibodies than the sL/bR binding assay that was tested in this study and previous reports. Moreover, both amount of the used recombinant protein of ICI receptor/ligand and turnaround time of the two binding assays were more than 10 times less than those of the functional ELISA. These data indicate that the two magnetic bead based binding assays are sensitive, rapid and cost-effective methods to determine blocking activity of ICIs.
