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Updated: Apr 28, 2026
Identifying PD-1/PD-L1 Inhibitors with Surface Plasmon Resonance Technology
Published on: May 2, 2025
Insights into PARP Inhibitors' Selectivity Using Fluorescence Polarization and Surface Plasmon Resonance Binding
Gianluca Papeo1, Nilla Avanzi1, Serena Bettoni1
1Nerviano Medical Sciences S.r.l., Nerviano, Italy.
Abstract:
PARP inhibitors are an exciting new class of antineoplastic drugs that have been proven to be efficacious as single agents in cancer settings with inherent DNA repair defects, as well as in combination with DNA-damaging chemotherapeutics. Currently, they are designed to target the catalytic domain of PARP-1, the most studied member of the family, with a key role in the DNA-damage repair process. Because PARP inhibitors are substrate (NAD(+)) competitors, there is a need for a deeper understanding of their cross-reactivity. This is particularly relevant for PARP-2, the PARP-1 closest homologue, for which an embryonic lethal phenotype has been observed in double knockout mice. In this study, we describe the development and validation of binding assays based on fluorescence polarization (FP) and surface plasmon resonance (SPR) techniques. PARP-1, PARP-2, PARP-3, and TNKS-1 FP displacement assays are set up by employing ad hoc synthesized probes. These assays are suitable for high-throughput screening (HTS) and selectivity profiling, thus allowing the identification of NAD(+)binding site selective inhibitors. The PARP-1 and PARP-2 complementary SPR binding assays confirm displacement data and the in-depth inhibitor characterization. Moreover, these formats have the potential to be broadly applicable to other members of the PARP family.
Insights
New assays were developed to understand PARP inhibitor cross-reactivity, crucial for targeting DNA repair defects in cancer. These methods enable high-throughput screening for selective inhibitors, advancing cancer drug development.
Area of Science:
- Biochemistry
- Molecular Biology
- Pharmacology
Background:
- Poly(ADP-ribose) polymerase (PARP) inhibitors are effective cancer drugs, particularly in tumors with DNA repair deficiencies.
- Current PARP inhibitors target the catalytic domain of PARP-1, a key enzyme in DNA damage repair.
- Understanding cross-reactivity, especially with PARP-2, is vital due to its homology with PARP-1 and observed embryonic lethality in knockout models.
Purpose of the Study:
- To develop and validate binding assays for assessing PARP inhibitor cross-reactivity.
- To enable high-throughput screening (HTS) for identifying selective inhibitors targeting the NAD(+) binding site.
- To provide tools for in-depth characterization of PARP inhibitors.
Main Methods:
- Development of fluorescence polarization (FP) displacement assays for PARP-1, PARP-2, PARP-3, and TNKS-1 using synthesized probes.
- Utilization of surface plasmon resonance (SPR) binding assays for complementary validation and inhibitor characterization.
- Application of these assays for high-throughput screening and selectivity profiling.
Main Results:
- Successfully established FP displacement assays for multiple PARP family members.
- Validated FP assay findings with complementary SPR binding assays for PARP-1 and PARP-2.
- Demonstrated the suitability of the developed assays for HTS and selectivity profiling of NAD(+) binding site inhibitors.
Conclusions:
- The developed FP and SPR assays are robust tools for characterizing PARP inhibitor cross-reactivity.
- These assays facilitate the identification of selective inhibitors targeting specific PARP family members.
- The assay formats show broad applicability for studying other members of the PARP enzyme family.
