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Updated: Jul 2, 2025
Achieving Efficient Fragment Screening at XChem Facility at Diamond Light Source
Published on: May 29, 2021
Novel Fragment Inhibitors of PYCR1 from Docking-Guided X-ray Crystallography
Kaylen R Meeks1, Juan Ji1, Mykola V Protopopov2
1Department of Biochemistry, University of Missouri, Columbia, Missouri 65211, United States.
Abstract:
The proline biosynthetic enzyme Δ1-pyrroline-5-carboxylate (P5C) reductase 1 (PYCR1) is one of the most consistently upregulated enzymes across multiple cancer types and central to the metabolic rewiring of cancer cells. Herein, we describe a fragment-based, structure-first approach to the discovery of PYCR1 inhibitors. Thirty-seven fragment-like carboxylic acids in the molecular weight range of 143-289 Da were selected from docking and then screened using X-ray crystallography as the primary assay. Strong electron density was observed for eight compounds, corresponding to a crystallographic hit rate of 22%. The fragments are novel compared to existing proline analog inhibitors in that they block both the P5C substrate pocket and the NAD(P)H binding site. Four hits showed inhibition of PYCR1 in kinetic assays, and one has lower apparent IC50 than the current best proline analog inhibitor. These results show proof-of-concept for our inhibitor discovery approach and provide a basis for fragment-to-lead optimization.
Insights
Researchers discovered novel PYCR1 inhibitors using a structure-first approach. These inhibitors target cancer cell metabolism by blocking key enzyme binding sites, offering a new avenue for cancer drug development.
Area of Science:
- Biochemistry
- Structural Biology
- Cancer Metabolism
Background:
- Proline biosynthesis enzyme PYCR1 is upregulated in many cancers.
- PYCR1 is crucial for cancer cell metabolic reprogramming.
- Targeting PYCR1 offers a potential strategy for cancer therapy.
Purpose of the Study:
- To discover novel inhibitors of PYCR1 using a fragment-based, structure-first approach.
- To identify compounds that block both P5C substrate and NAD(P)H binding sites.
- To provide a foundation for fragment-to-lead optimization for PYCR1 inhibitors.
Main Methods:
- Fragment-based drug discovery.
- Structure-first approach utilizing docking and X-ray crystallography.
- Screening of 37 fragment-like carboxylic acids.
- Kinetic assays to determine enzyme inhibition.
Main Results:
- Identified 8 crystallographic hits from 37 screened fragments (22% hit rate).
- Discovered novel fragments that bind to both P5C substrate and NAD(P)H sites.
- Four fragments inhibited PYCR1 activity in kinetic assays.
- One fragment exhibited a lower IC50 than existing proline analog inhibitors.
Conclusions:
- The fragment-based, structure-first approach is effective for PYCR1 inhibitor discovery.
- Novel PYCR1 inhibitors targeting distinct binding sites were identified.
- These findings provide a strong basis for developing new cancer therapeutics targeting PYCR1.
