New Insight into the Structure–Activity Relationships of the Selective Excitatory Amino Acid Transporter Subtype 1 (EAAT1) Inhibitors UCPH‐101 and UCPH‐102

Abstract

In the present study, we made further investigations on the structure-activity requirements of the selective excitatory amino acid transporter 1 (EAAT1) inhibitor, 2-amino-4-(4-methoxyphenyl)-7-(naphthalen-1-yl)-5-oxo-5,6,7,8-tetrahydro-4H-chromene-3-carbonitrile (UCPH-101), by exploring 15 different substituents (R(1) ) at the 7-position in combination with eight different substituents (R(2) ) at the 4-position. Among the 63 new analogues synthesized, we identified a number of compounds that unexpectedly displayed inhibitory activities at EAAT1 in light of understanding the structure-activity relationship (SAR) of this inhibitor class extracted from previous studies. Moreover, the nature of the R(1) and R(2) substituents were observed to contribute to the functional properties of the various analogues in additive and non-additive ways. Finally, separation of the four stereoisomers of analogue 14 g (2-amino-4-([1,1'-biphenyl]-4-yl)-3-cyano-7-isopropyl-5-oxo-5,6,7,8-tetrahydro-4H-chromene) was carried out, and in agreement with a study of a related scaffold, the R configuration at C4 was found to be mandatory for inhibitory activity, while both the C7 diastereomers were found to be active as EAAT1 inhibitors. A study of the stereochemical stability of the four pure stereoisomers 14 g-A-D showed that epimerization takes places at C7 via a ring-opening, C-C bond rotation, ring-closing mechanism.

The Druggability of Solute Carriers

“…A series of allosteric EAAT1-selective inhibitors with novel scaffolds were discovered by screening a library of 3040 compounds in an [ 3 H]-aspartate cellular uptake assay. Optimization yielded UCPH-101 ( 26 ) with 0.66 μM IC 50 in uptake assays, which exhibited a noncompetitive and long-lasting EAAT1 inhibition through an allosteric binding mode confirmed later by structural studies . Although in vivo administration of UCPH-102 ( 27 ), an analogue with improved bioavailability, did not cause significant effects in a rodent locomotor model, presumably due to compensatory activities by EAAT2…”

Identification of a New Class of Selective Excitatory Amino Acid Transporter Subtype 1 (EAAT1) Inhibitors Followed by a Structure–Activity Relationship Study

“…However, it should be stressed that a definite conclusion as to whether 1a and the other inhibitors in this class are indeed allosteric inhibitors or actually target the substrate-binding pocket in EAAT1 will have to await further experimental data. Finally, it is noteworthy that even though 1a clearly is not nearly as potent and selective an EAAT1 inhibitor as 38 or 39 , the inhibitory potency and degree of selectivity exhibited by 1a at EAAT1 are nevertheless roughly comparable to the properties displayed by DHK at EAAT2 . In view of the extensive use of this prototypic EAAT2 inhibitor as a pharmacological tool in studies of EAAT2 function in native tissues over the years, we propose that 1a analogously could be a valuable tool in future explorations of the physiological roles mediated by and the therapeutic potential in EAAT1, be it as a supplement or alternative to 38 / 39 .…”

“…From a library screening, compound 1a was found to be a putative EAAT1-selective inhibitor . When tested at a concentration of ∼100 μM, 1a almost completely inhibited the EAAT1-mediated uptake of [ 3 H]- d -Asp uptake in EAAT1-HEK293 cells, whereas it exhibited no significant effects on [ 3 H]- d -Asp uptake measured in EAAT2- and EAAT3-HEK293 cells in concomitant screenings.…”

“…Compounds comprised in these library screenings are often structurally very different from α-amino acids, which leads to the assumption that hit compounds likely act through allosteric sites in the transporter. A successful outcome of this approach is discovery of the first class of EAAT1-selective inhibitors, represented by the analogues 2-amino-5,6,7,8-tetrahydro-4-(4-methoxyphenyl)-7-(naphthalen-1-yl)-5-oxo-4 H -chromene-3-carbonitrile ( 38 , UCPH-101) and 2-amino-5,6,7,8-tetrahydro-4-(4-methyl)-7-(naphthalen-1-yl)-5-oxo-4 H -chromene-3-carbonitrile ( 39 , UCPH-102) (Figure C). , This inhibitor class was developed based on a hit compound identified in a screening of a commercial 3040-compound library at EAAT1–3, and in subsequent studies the structure–activity-relationship (SAR) of this scaffold was explored in great detail. − In the present study, we report the discovery of a new class of selective EAAT1 inhibitors based on another hit compound from this screening and present an elaborate structure–activity relationship (SAR) study of this inhibitor class.…”

Bioavailability Studies and in vitro Profiling of the Selective Excitatory Amino Acid Transporter Subtype 1 (EAAT1) Inhibitor UCPH‐102

“…In the course of our previous work on the EAATs we discovered the first class of selectiveE AAT1 inhibitors [5,6] and subsequently conducted elaborate structure-activity relationship (SAR) studies to optimize the inhibitory potencies as well as the pharmacokinetic properties of this inhibitor class. [7][8][9][10] The analogue UCPH-101h as become as tandard tool compound in the EAATf ield ( Figure 1);h owever,a si td oes not penetrate the blood-brain barrier( BBB), it is not suitable for in vivo studies. [11][12][13][14] Herein we report that the close analogue UCPH-102, in contrastt oU CPH-101, is able to cross the BBB, and we present Although the selectivee xcitatory amino acid transporter subtype 1( EAAT1) inhibitor UCPH-101h as become as tandard pharmacological toolc ompound for in vitro and ex vivo studies in the EAATr esearch field, its inability to penetrate the blood-brain barrierm akes it unsuitable for in vivo studies.…”