Next Article in Journal
Editorial of Special Issue “Non-Coding RNAs in Pathogen-Host Interaction”
Previous Article in Journal
Comprehensive Identification of Deleterious TP53 Missense VUS Variants Based on Their Impact on TP53 Structural Stability
Article

Simultaneous Inhibition of PD-1 and Stimulation of CD40 Signaling Pathways by Anti-PD-L1/CD40L Bispecific Fusion Protein Synergistically Activate Target and Effector Cells

1
Bioassay Development, Analytical Sciences, Biopharmaceutical Development, R&D, AstraZeneca, Gaithersburg, MD 20878, USA
2
Physicochemical-Analytical Sciences, Biopharmaceutical Development, AstraZeneca, R&D, Gaithersburg, MD 20878, USA
*
Author to whom correspondence should be addressed.
Academic Editor: Maria Grazia Cifone
Int. J. Mol. Sci. 2021, 22(21), 11302; https://doi.org/10.3390/ijms222111302
Received: 3 September 2021 / Revised: 22 September 2021 / Accepted: 15 October 2021 / Published: 21 October 2021
(This article belongs to the Section Molecular Pathology, Diagnostics, and Therapeutics)
Bispecific antibodies (BsAbs) or fusion proteins (BsAbFPs) present a promising strategy for cancer immunotherapy. Numerous BsAbs targeting coinhibitory and costimulatory pathways have been developed for retargeting T cells and antigen presenting cells (APCs). It is challenging to assess the potency of BsAb that engages two different signaling pathways simultaneously in a single assay format, especially when the two antigen targets are expressed on different cells. To explore the potency of anti-PD-L1/CD40L BsAbFP, a fusion protein that binds to human CD40 and PD-L1, we engineered CHO cells as surrogate APCs that express T cell receptor activator and PD-L1, Jurkat cells with PD-1 and NFAT-luciferase reporter as effector T cells, and Raji cell with NFkB-luciferase that endogenously expresses CD40 as accessory B cells. A novel reporter gene bioassay was developed using these cell lines that allows anti-PD-L1/CD40L BsAbFP to engages both PD-1/PD-L1 and CD40/CD40L signaling pathways in one assay. As both reporters use firefly luciferase, the effects of activating both signaling pathways is observed as an increase in luminescence, either as a higher upper asymptote, a lower EC50, or both. This dual target reporter gene bioassay system reflects potential mechanism of action and demonstrated the ability of anti-PD-L1/CD40L BsAbFP to synergistically induce biological response compared to the combination of anti-PD-L1 monovalent monoclonal antibody and agonist CD40L fusion protein, or either treatment alone. The results also showed a strong correlation between the drug dose and biological response within the tested potency range with good linearity, accuracy, precision, specificity and stability indicating properties, suggesting that this “three-cell-in-one” dual target reporter gene bioassay is suitable for assessing potency, structure-function and critical quality attributes of anti-PD-L1/CD40L BsAbFP. This approach could be used for developing dual target bioassays for other BsAbs and antibodies used for combination therapy. View Full-Text
Keywords: bioassay; bispecific antibody; potency; mechanism of action; qualification bioassay; bispecific antibody; potency; mechanism of action; qualification
Show Figures

Figure 1

MDPI and ACS Style

Pandey, M.S.; Wang, C.; Umlauf, S.; Lin, S. Simultaneous Inhibition of PD-1 and Stimulation of CD40 Signaling Pathways by Anti-PD-L1/CD40L Bispecific Fusion Protein Synergistically Activate Target and Effector Cells. Int. J. Mol. Sci. 2021, 22, 11302. https://doi.org/10.3390/ijms222111302

AMA Style

Pandey MS, Wang C, Umlauf S, Lin S. Simultaneous Inhibition of PD-1 and Stimulation of CD40 Signaling Pathways by Anti-PD-L1/CD40L Bispecific Fusion Protein Synergistically Activate Target and Effector Cells. International Journal of Molecular Sciences. 2021; 22(21):11302. https://doi.org/10.3390/ijms222111302

Chicago/Turabian Style

Pandey, Madhu S., Chunlei Wang, Scott Umlauf, and Shihua Lin. 2021. "Simultaneous Inhibition of PD-1 and Stimulation of CD40 Signaling Pathways by Anti-PD-L1/CD40L Bispecific Fusion Protein Synergistically Activate Target and Effector Cells" International Journal of Molecular Sciences 22, no. 21: 11302. https://doi.org/10.3390/ijms222111302

Find Other Styles
Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Article Access Map by Country/Region

1
Back to TopTop