Abstract
Lysophosphatidic acid (LPA) is a highly potent endogenous lipid mediator that protects and rescues cells from programmed cell death. Earlier work identified the LPA2 G protein-coupled receptor subtype as an important molecular target of LPA mediating antiapoptotic signaling. Here we describe the results of a virtual screen using single-reference similarity searching that yielded compounds 2-((9-oxo-9H-fluoren-2-yl)carbamoyl)benzoic acid (NSC12404), 2-((3-(1,3-dioxo-1H-benzo[de]isoquinolin-2(3H)-yl)propyl)thio)benzoic acid (GRI977143), 4,5-dichloro-2-((9-oxo-9H-fluoren-2-yl)carbamoyl)benzoic acid (H2L5547924), and 2-((9,10-dioxo-9,10-dihydroanthracen-2-yl)carbamoyl) benzoic acid (H2L5828102), novel nonlipid and drug-like compounds that are specific for the LPA2 receptor subtype. We characterized the antiapoptotic action of one of these compounds, GRI977143, which was effective in reducing activation of caspases 3, 7, 8, and 9 and inhibited poly(ADP-ribose)polymerase 1 cleavage and DNA fragmentation in different extrinsic and intrinsic models of apoptosis in vitro. Furthermore, GRI977143 promoted carcinoma cell invasion of human umbilical vein endothelial cell monolayers and fibroblast proliferation. The antiapoptotic cellular signaling responses were present selectively in mouse embryonic fibroblast cells derived from LPA1&2 double-knockout mice reconstituted with the LPA2 receptor and were absent in vector-transduced control cells. GRI977143 was an effective stimulator of extracellular signal-regulated kinase 1/2 activation and promoted the assembly of a macromolecular signaling complex consisting of LPA2, Na+-H+ exchange regulatory factor 2, and thyroid receptor interacting protein 6, which has been shown previously to be a required step in LPA-induced antiapoptotic signaling. The present findings indicate that nonlipid LPA2-specific agonists represent an excellent starting point for development of lead compounds with potential therapeutic utility for preventing the programmed cell death involved in many types of degenerative and inflammatory diseases.
Footnotes
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The online version of this article (available at http://molpharm.aspetjournals.org) contains supplemental material.
This research was supported by the National Institutes of Health National Institute of Allergy [Medical Countermeasures Radiological and Nuclear Threats Program AI80405].
G.T. is a founder of RxBio, Inc.
Article, publication date, and citation information can be found at http://molpharm.aspetjournals.org.
ABBREVIATIONS:
- LPA
- lysophosphatidic acid
- S1P
- sphingosine-1-phosphate
- OTP
- octadecenyl thiophosphate
- GPCR
- G protein-coupled receptor
- Ki16425
- 3-(4-[4-([1-(2-chlorophenyl)ethoxy]carbonyl amino)-3-methyl-5-isoxazolyl] benzylsulfanyl) propanoic acid
- AM152
- (R)-1-phenylethyl-5-(4-biphenyl-4-cyclopropanecarboxylic acid)-3-methylisoxazole-4-yl carbamate sodium salt
- AM095
- sodium, (4′-(3-methyl-4-((R)-1-phenylethoxycarbonylamino)-isoxazol-5-yl)-biphenyl-4-yl)-acetate
- GRI977143
- 2-((3-(1,3-dioxo-1H-benzo[de]isoquinolin-2(3H)-yl)propyl)thio)benzoic acid
- GRI
- Genome Research Institute
- Bax
- Bcl-2-associated X protein
- PARP-1
- poly(ADP-ribose)polymerase 1
- ERK1/2
- extracellular signal regulated kinases 1/2
- TRIP6
- thyroid receptor interacting protein 6
- NHERF2
- Na+-H+ exchange regulatory factor 2
- NSC12404
- 2-((9-oxo-9H-fluoren-2-yl)carbamoyl)benzoic acid
- H2L
- Hit2Lead
- H2L5547924
- 4,5-dichloro-2-((9-oxo-9H-fluoren-2-yl)carbamoyl)benzoic acid
- H2L5828102
- 2-((9,10-dioxo-9,10-dihydroanthracen-2-yl)carbamoyl)benzoic acid
- PBS
- phosphate-buffered saline
- MOE
- Molecular Operating Environment
- UC-DDC
- University of Cincinnati Drug Discovery Center
- TM
- transmembrane
- MEF
- mouse embryonic fibroblast
- DMEM
- Dulbecco's modified Eagle's medium
- FBS
- fetal bovine serum
- IEC-6
- intestinal epithelial cell line 6
- RH7777
- McArdle rat hepatoma cell line
- LPAR
- LPA receptor
- HUVEC
- human umbilical vein endothelial cell
- TNF-α
- tumor necrosis factor α
- CHX
- cycloheximide
- EGFP
- enhanced green fluorescent protein.
- Received April 29, 2012.
- Accepted September 10, 2012.
- Copyright © 2012 The American Society for Pharmacology and Experimental Therapeutics
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