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Immune Checkpoint Bioassays to Characterize Immune-Modulating Antibodi…

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Immune Checkpoint Bioassays to Characterize Immune-Modulating Antibodies

 




The human immune system is comprised of a complex network of co-inhibitory and co-stimulatory pathways that facilitate the elimination of cells expressing foreign antigens while maintaining tolerance to self antigen. Immune checkpoint pathways mediated by PD-1, CTLA-4, GITR, 4-IBB, OX4-O and TIGIT are promising immunotherapy targets for the treatment of cancer and autoimmunity. Immunotherapy agents that target combinations of these pathways (e.g., PD-1 and TIGIT) are particularly promising. Immune checkpoint reporter-based bioassays provide a simple, consistent and reliable cell-based assay to measure antibody function throughout the drug research and development pipeline.

 

 


  Biologically relevant measurement of Ab MOA
  • Blockade or engagement of specific immune checkpoint pathways induces luciferase activity
  • Well established and recognized reporter assay format
  • Predictive of true biology

  Consistent and reliable measure of Ab potency and stability
  • Demonstrated precision, accuracy, reproducibility and robustness
  • Thaw-and-use cell format, no cell culture required
  • Bioassay kits include all required reagents in standardized formats

  Easy To Implement
  • Convenient product formats, rapid and streamlined workflow
  • Amenable to high-throughput formats


Thaw-and-use cells provide significant time and labor savings over traditional methods, while reducing assay variability.

Measure Antibody efficacy to block immune checkpoint inhibitory signals. Disruption of the receptor-ligand interaction induces luciferase activity.



Schematic representation of the blockade bioassays for PD-1/PD-L1, TIGIT/CD155 and PD-1 + TIGIT. Each bioassay consists of a T effector cell (expressing PD-1, TIGIT or PD-1 + TIGIT) and an aAPC cell (expressing PD-L1, CD155 or PD-L1 + CD155), respectively. Co-culture of a T effector cell with its corresponding aAPC cell leads to three events as noted in the cell image.



  The PD-1/PD-L1 Blockade Bioassay is specific for PD-1 and PD-L1 Antibodies

PD-1 Effector Cells and PD-L1 aAPC Cells were incubated for 6 hr at 37°C with increasing concentrations of either an anti-PD-1, PD-L1, or CTLA-4 antibody. The anti-PD-1 and PD-L1 antibodies, but not the anti-CTLA-4 antibody, blocked the immune checkpoint inhibitory signal resulting in luciferase activity. Data were analyzed using a 4PL model and GraphPad® Prism software.


The performance characteristics of the PD-1/PD-L1 Blockade Bioassay are suitable for many applications of a bioassay used across antibody drug discovery research: It is specific, accurate over a relative potency range of 50-200%, and reproducible.



PD-1 Effector Cells and PD-L1 aAPC Cells were incubated with increasing concentrations anti-PD-L1 antibody preparations representing a 50-200% potency range. The PD-1/PD-L1 Blockade Bioassay demonstrated appropriate rank ordering of antibody potency.

PD-1 Effector Cells and PD-L1 aAPC Cells were incubated with increasing concentrations of anti-PD-L1 antibody preparations representing 100% potency. The PD-1/PD-L1 Blockade Bioassay was reproducible across four independent 96-well plates.


Bioassay Cell Components Catalog or CS#
PD-1/PD-L1 Blockade Bioassay Kit¹ PD-1 Effector Cells (Jurkat)
PD-L1 aAPC Cells (CHO-K1)²
J1250
PD-1/PD-L1 Blockade Bioassay Kit (5X) PD-1 Effector Cells (Jurkat)
PD-L1 aAPC Cells (CHO-K1)
J1255
PD-L1 Negative Cells³ PD-L1 Negative aAPC Cells (CHO-K1) J1191
PD-L1 Negative Cells (5X)³ PD-L1 Negative Cells (5X)³ J1195
  • ¹ Kits contain PD-1 Effector Cells, PD-L1 aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
  • ³ Negative control cells for the PD-1/PD-L1 Blockade Bioassay.
Bioassay Cell Components Catalog or CS#
PD-1/PD-L2 Blockade Bioassay Kit¹ PD-1 Effector Cells (Jurkat) CS187131-1
PD-1/PD-L2 Blockade Bioassay Kit¹, 5X PD-L2 Cells (CHO-K1)² CS187135-1
PD-1/PD-L2 Blockade Bioassay,
Propagation Model
NA CS187127
  • ¹ Kits contain PD-1 Effector Cells, PD-L1 aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
  • ³ Negative control cells for the PD-1/PD-L1 Blockade Bioassay.
Bioassay Cell Components Catalog or CS#
CTLA-4 Blockade Bioassay Kit¹ CTLA-4 Effector Cells (Jurkat)
aAPC Cells (Raji)²
CS186920
CTLA-4 Blockade Bioassay Kit (5X) CTLA-4 Effector Cells (Jurkat)
aAPC Cells (Raji)
CS186926
Control Ab, Anti-CTLA-4 NA CS186915
  • ¹ Kits contain CTLA-4 Effector Cells, aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
Bioassay Cell Components Catalog or CS#
TIGIT/CD155 Blockade Bioassays TIGIT Effector Cells (Jurkat)
CD155 aAPC Cells (CHO-K1)²
CS198807
TIGIT/CD155 Blockade Bioassay Kit¹ TIGIT Effector Cells (Jurkat)
CD155 aAPC Cells (CHO-K1)²
CS198811
  • ¹ Kits contain TIGIT Effector Cells, CD155 aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
Bioassay Cell Components Catalog or CS#
PD-1 + TIGIT Combination Bioassay Kit PD-1 + TIGIT Effector Cells (Jurkat)
PD-L1 + CD155 aAPC Cells (CHO-K1)²
CS198813
PD-1 + TIGIT Combination Bioassay Kit PD-1 + TIGIT Effector Cells (Jurkat)
PD-L1 + CD155 aAPC Cells (CHO-K1)²
CS198817
  • ¹ Kits contain PD-1 + TIGIT Effector Cells, PD-L1 + CD155 aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
Bioassay Cell Components Catalog or CS#
GITR Bioassay GITR Effector Cells (Jurkat) CS184003
CD40 Bioassay CD40 Effector Cells (Ramos)
PD-L1 + CD155 aAPC Cells (CHO-K1)²
CS193701
4-1BB Bioassay 4-1BB Effector Cells (Jurkat) CS196002
OX40 Bioassay OX40 Effector Cells (Jurkat) CS197701
  • ¹ Kits contain PD-1 Effector Cells, PD-L1 aAPC Cells, Cell Culture Medium, Assay Reagents.
  • ² aAPC (artificial antigen presenting cells) activate the TCR/CD3 complex on Jurkat Effector Cells.
  • ³ Negative control cells for the PD-1/PD-L1 Blockade Bioassay.
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