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Kinase Inhibitor Screening In Self-assembled Human Protein Microarrays
Published on: October 23, 2019
Identification of a novel competitive inhibitor of p38alpha MAPK by a human PBMC screen
Yu-Chih Liu1, Chia-Chen Ko, Fong-Chi Cheng
1Graduate Institute of Biochemistry and Molecular Biology, College of Medicine, National Taiwan University, Taipei 100, Taiwan.
Abstract:
The pro-inflammatory cytokines TNF-alpha and IL-1beta are two of the important mediators involved in the several chronic inflammatory diseases. We used the release of TNF-alpha and IL-1beta from lipopolysaccharide-stimulated human PBMC as inflammatory indexes to discover the potential anti-inflammatory candidates. Among near 500 chemical compounds, MT4 had the suppressive action on the release of TNF-alpha and IL-1beta in PBMC with IC50 values of 22 and 44 nM, respectively. After verified the MT4 inhibitory mechanism, the results revealed that p38alpha and p38beta MAPK activity was inhibited by MT4 with an IC50 value of 0.13 and 0.55 microM, respectively. Further characterization of enzyme kinetics showed the binding mode of MT4 was competitive with the ATP substrate-binding site of p38alpha MAPK.
Insights
MT4 effectively suppresses the release of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) from human peripheral blood mononuclear cells (PBMC). This compound acts by competitively inhibiting p38alpha MAPK, indicating potential as an anti-inflammatory agent.
Area of Science:
- Immunology
- Pharmacology
- Molecular Biology
Background:
- Pro-inflammatory cytokines like tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) are key mediators in chronic inflammatory diseases.
- Human peripheral blood mononuclear cells (PBMC) stimulated with lipopolysaccharide (LPS) are utilized as an inflammatory model.
- Identifying novel anti-inflammatory compounds is crucial for managing inflammatory conditions.
Purpose of the Study:
- To discover novel anti-inflammatory candidates by screening chemical compounds.
- To evaluate the inhibitory effects of identified compounds on cytokine release from PBMC.
- To elucidate the molecular mechanism of action for promising anti-inflammatory agents.
Main Methods:
- Screening of approximately 500 chemical compounds for anti-inflammatory activity.
- Measurement of TNF-alpha and IL-1beta release from LPS-stimulated human PBMC.
- Determination of IC50 values for compound inhibition of cytokine release and MAPK activity.
- Enzyme kinetics studies to characterize the binding mode of the inhibitor.
Main Results:
- MT4 demonstrated suppressive activity on TNF-alpha and IL-1beta release from PBMC with IC50 values of 22 nM and 44 nM, respectively.
- MT4 inhibited p38alpha and p38beta MAPK activity with IC50 values of 0.13 microM and 0.55 microM, respectively.
- Enzyme kinetics revealed that MT4 competitively inhibits p38alpha MAPK at the ATP binding site.
Conclusions:
- MT4 is a potent inhibitor of pro-inflammatory cytokine release from human PBMC.
- MT4 exerts its anti-inflammatory effects through the inhibition of p38alpha and p38beta MAPK.
- The competitive inhibition of p38alpha MAPK by MT4 suggests a targeted therapeutic potential for inflammatory diseases.
