Skip to main page content
Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
, 58 (42), 4304-4316

Identification of Fatty Acid Binding Protein 5 Inhibitors Through Similarity-Based Screening

Affiliations

Identification of Fatty Acid Binding Protein 5 Inhibitors Through Similarity-Based Screening

Yuchen Zhou et al. Biochemistry.

Abstract

Fatty acid binding protein 5 (FABP5) is a promising target for development of inhibitors to help control pain and inflammation. In this work, computer-based docking (DOCK6 program) was employed to screen ∼2 M commercially available compounds to FABP5 based on an X-ray structure complexed with the small molecule inhibitor SBFI-26 previously identified by our group (also through virtual screening). The goal was discovery of additional chemotypes. The screen resulted in the purchase of 78 candidates, which led to the identification of a new inhibitor scaffold (STK-0) with micromolar affinity and apparent selectivity for FABP5 over FABP3. A second similarity-based screen resulted in three additional hits (STK-15, STK-21, STK-22) from which preliminary SAR could be derived. Notably, STK-15 showed comparable activity to the SBFI-26 reference under the same assay conditions (1.40 vs 0.86 μM). Additional molecular dynamics simulations, free energy calculations, and structural analysis (starting from DOCK-generated poses) revealed that R enantiomers (dihydropyrrole scaffold) of STK-15 and STK-22 have a more optimal composition of functional groups to facilitate additional H-bonds with Arg109 of FABP5. This observation suggests enantiomerically pure compounds could show enhanced activity. Overall, our study highlights the utility of using similarity-based screening methods to discover new inhibitor chemotypes, and the identified FABP5 hits provide a strong starting point for future efforts geared to improve activity.

Conflict of interest statement

The authors declare no competing financial interest.

Figures

Figure 1
Figure 1
Docking setup for virtual screening targeting FABP5 (PDB 5UR9), protein surface in tan, docking spheres represented by blue spheres, SBFI-26 reference ligand in cyan, and docking bounding box in gray.
Figure 2
Figure 2
DOCK6 Virtual Screening Protocol
Figure 3
Figure 3
Overlay of top-ranked clusterheads (200 compounds each) prioritized by different scoring functions with FABP5: (a) DCEVDW+ES, (b) FPSVDW+ES, (c) TotalScore, (d) HMS, (e) FMS, and (f) VOS. See text for scoring function descriptions.
Figure 4
Figure 4
Close-up view of FABP5 residue Arg129 with the 78 docked and purchased compounds (orange) highlighting a key conserved H-bond interaction (magenta lines).
Figure 5
Figure 5
Fluorescence displacement binding assay results for the first set of 78 compounds (5 μM) against (a) FABP5 and (b) FABP3. The results were arranged in the order of their mean % Fluorescence value against FABP5. Control compounds are arachidonic acid (red) and SBFI-26 (green).
Figure 6
Figure 6
Comparison of STK-0 and SBFI-26 showing (a) 2D structures, (b) activities with FABP5, and (c) binding site geometries (STK-0 orange, SBFI-26 green).
Figure 7
Figure 7
Comparison of DOCK predicted binding geometries for parents (cyan) vs analogues (magenta) derived from (a) 3D similarity searches of the originally docked library (N = 7) and (b) 2D similarity searches in ZINC based on parent compound STK-0 (N = 48).
Figure 8
Figure 8
Fluorescence displacement binding assay results for STK compounds (5 μM) against (a) FABP5 and (b) FABP3. The results were arranged in the order of their mean % Fluorescence value against FABP5. Control compounds are arachidonic acid (red) and SBFI-26 (green).
Figure 9
Figure 9
2-D structures and dose response curves for binding (blue) and HUVEC cell viability (red) for (a) STK-15, (b) STK-21, and (c) STK-22. Values represent the average of three independent experiments.
Figure 10
Figure 10
(a) Overlay of 3 of the 4 STK-15 enantiomers: S,R-STK-15 (blue), S,S-STK-15 (tan), and R,R-STK-15 (magenta). (b) Footprint (per-residue energy) comparison between S,R-STK-15 (blue) and R,R-STK-15 (magenta). Energies in kcal/mol.
Figure 11
Figure 11
Comparison of poses for (a) R,R-STK-15 and (b) S,R-STK-15 from representative snapshots from the largest MD clusters observed over 80 ns of simulation.
Figure 12
Figure 12
(a) Starting coordinates for R,R-STK-15 (meta O-phenyl, orange) and R,R-STK-0 (meta chlorine, gray) in the FABP5 X-ray crystal structure. (b) Comparison of starting coordinates (orange = R,R-STK-15, purple = Phe65) vs representative MD snapshot (green = R,R-STK-15, cyan = Phe65).
Figure 13
Figure 13
(a) Binding site comparison between S,R-STK-15 (orange ligand, tan protein) and R,R-STK-15 (green ligand, blue protein). (b) Overlay showing the S,R-STK-15 ligand (orange) in the R,R-STK-15 protein (blue) conformation through protein backbone alignment. (c) Overlay showing the R,R-STK-15 ligand (orange) in the S,R-STK-15 protein (blue) conformation.

Similar articles

See all similar articles

References

    1. Furuhashi M.; Hotamisligil G. S. (2008) Fatty acid-binding proteins: role in metabolic diseases and potential as drug targets. Nat. Rev. Drug Discovery 7, 489–503. 10.1038/nrd2589. - DOI - PMC - PubMed
    1. Zimmerman A. W.; Veerkamp J. H. (2002) New insights into the structure and function of fatty acid-binding proteins. Cell. Mol. Life Sci. 59, 1096–1116. 10.1007/s00018-002-8490-y. - DOI - PubMed
    1. Kaczocha M.; Glaser S. T.; Deutsch D. G. (2009) Identification of intracellular carriers for the endocannabinoid anandamide. Proc. Natl. Acad. Sci. U. S. A. 106, 6375–6380. 10.1073/pnas.0901515106. - DOI - PMC - PubMed
    1. LoVerme J.; Russo R.; La Rana G.; Fu J.; Farthing J.; Mattace-Raso G.; Meli R.; Hohmann A.; Calignano A.; Piomelli D. (2006) Rapid broad-spectrum analgesia through activation of peroxisome proliferator-activated receptor-alpha. J. Pharmacol. Exp. Ther. 319, 1051–1061. 10.1124/jpet.106.111385. - DOI - PubMed
    1. Lo Verme J.; Fu J.; Astarita G.; La Rana G.; Russo R.; Calignano A.; Piomelli D. (2005) The nuclear receptor peroxisome proliferator-activated receptor-alpha mediates the anti-inflammatory actions of palmitoylethanolamide. Mol. Pharmacol. 67, 15–19. 10.1124/mol.104.006353. - DOI - PubMed
Feedback