Accelerate Your PCSK9 Inhibitor Discovery with Cutting-Edge Assay Solutions

Background

PCSK9 (proprotein convertase kukulin/Kexin type 9) is a protein expressed primarily in the liver that regulates cholesterol metabolism. It binds low-density lipoprotein receptor (LDLR) through its catalytic domain and uses the C-terminal domain to induce LDLR degradation in lysosomes, reducing the amount of LDLR on the surface of hepatocytes and decreasing the efficiency of LDL-C clearance from plasma. High LDL-C is associated with stroke, complications of diabetes, chronic kidney disease and atherosclerosis. Therefore, researchers have designed small-molecule inhibitors to inhibit PCSK9, leveraging the advantages of high stability, high oral bioavailability, and low cost. Currently, some small-molecule PCSK9 inhibitors have entered preclinical and early clinical trial stages, demonstrating good safety and efficacy.

Biochemical Assays

ICE Bioscience has been providing high throughput screening services for PCSK9 enzyme activity and has developed several small-molecule PCSK9 inhibitor activity assays for in vitro studies, including the PCSK9-LDLR Homogeneous Time Resolved Fluorescence (HTRF) Assay and the PCSK9 Enzyme-linked Immunosorbent Assay (ELISA).

PCSK9-LDLR HTRF Assay

The PCSK9-LDLR HTRF Assay is a sensitive, high-throughput method ideal for screening small-molecule inhibitors. It uses the detection of fluorescence intensity to reflect the binding interaction between PCSK9 and LDLR. As shown by Pep2-8’s concentration-dependent inhibition and IC50 value, the inhibitor weakens the fluorescence signal, enabling quantitative inhibition assessment. HTRF technology features no-clean, rapid detection and time-resolved signal processing that reduces noise and improves accuracy, supporting cardiovascular drug discovery by facilitating efficient, high-throughput screening and improving data reliability.

PCSK9-ELISA

PCSK9-ELISA is a sensitive, specific assay for detecting PCSK9 binding to LDLR. It uses a primary antibody to capture PCSK9, a secondary HRP-labeled antibody for detection, and color intensity reflects binding strength. Adding inhibitors can reduce color intensity, allowing quantitative analysis of inhibition effects, demonstrated by the IC50 value of Alirocumab in Figure 4. Although ELISA involves more procedures, it eliminates label interference, providing high sensitivity and reliability, making it appropriate for developing small-molecule PCSK9 inhibitors in drug development.

Cell-Based Assay

PCSK9-HepG2?Cell-Based?Assay

The PCSK9-HepG2 Cell-based Assay is a novel platform designed for assessing small-molecule PCSK9 inhibitors in HepG2 cells. It evaluates PCSK9 secretion, LDL-R expression, and LDL uptake by ELISA and flow cytometry. In the reference test with PF-06446846 hydrochloride, PCSK9 secretion decreased, while LDL-R expression increased, confirming inhibitor efficacy. This assay provides multi-dimensional analysis and facilitates high-throughput screening (HTS), making it ideal for evaluating and optimizing PCSK9-related candidate compounds in drug discovery and development.

Summary

In summary, it is critical to select the appropriate assay in small-molecule PCSK9 inhibitor screening, which needs to be based on the mechanism of action of the inhibitor. ICE Bioscience has a comprehensive small-molecule PCSK9 inhibitor screening platform, including high-throughput HTRF assays, sensitive ELISA assays, and innovative cell-based screening platforms, which are able to meet the diverse needs of ?screening different small-molecule inhibitors. If you have any questions or needs about PCSK9 targets and related experiments, please feel free to contact us.