Discovery and characterization of potent small molecule inhibitors of the high affinity proline transporter.

The mammalian proline transporter (PROT) is a high affinity Na(+)/Cl(-)-dependent transporter expressed in specific regions of the brain. It is homologous to other neurotransmitter transporters such as glycine, norepinephrine, serotonin, and dopamine transporters. PROT is enriched in glutamatergic synaptic terminals and may play an important role in the regulation of excitatory neurotransmission. No non-peptide small molecule inhibitors have been described for this transporter. To study its physiological role in the central nervous system and evaluate its potential as a therapeutic target, we established cell lines that stably express recombinant hPROT and characterized its kinetic properties for proline uptake. We then screened for inhibitors and identified a series of compounds that inhibit hPROT-mediated proline uptake. A known compound, benztropine, was found to inhibit hPROT with an IC(50) of 0.75microM. A series of novel compounds were also found, one of which, LP-403812, showed an IC(50) of approximately 0.1microM on both recombinant human and mouse PROT without significant inhibition of glycine and dopamine transporters at concentrations up to 10microM. This compound also inhibited proline transporter activity of mouse brain synaptosomes with the same potency. These inhibitors provide important tools for the understanding of PROT functions in the brain and may lead to the development of therapeutic agents for certain neurological disorders.

Post-ischemic treatment with dipyruvyl-acetyl-glycerol decreases myocardial infarct size in the pig.

The beneficial effects of pyruvate in organ reperfusion injury have been documented, however the therapeutic use of pyruvate has been hindered by the lack of an appropriate delivery method. Pyruvic acid is unstable and high rates of sodium pyruvate infusion are toxic. Dipyruvyl-acetyl-glycerol (DPAG) ester was developed as a novel method for intravenous pyruvate delivery at a high rate without sodium overload. We tested the ability of DPAG to reduce myocardial infarct size when administered after severe myocardial ischemia in an anesthetized open-chest pig model of ischemia-reperfusion injury. Ischemia was induced by total occlusion of the distal 2/3 of the left anterior descending coronary artery for one hour, followed by two hours of reperfusion. Animals were either untreated (n = 7), or treated with intravenous DPAG (8.0 mg/kg(-1). min(-1), n = 8) during the two hours of reperfusion. Infarct size was measured on blinded samples using tetrazolium staining. The DPAG treated group had elevated pyruvate levels (0.82 +/- 0.07 mM) and reduced infarct size (20.1 +/- 4.2% of the volume at risk, compared to 30.8 +/- 4.6% in the untreated animals (p < 0.05)), with no difference in blood pressure or heart rate between groups. In conclusion, an intravenous infusion of DPAG safely increases arterial pyruvate concentration and reduces myocardial infarct size following myocardial ischemia.