Development of Thiophenic Analogues of Benzothiadiazine Dioxides as New Powerful Potentiators of 2‑Amino-3-(3-hydroxy-5-methylisoxazol-4-yl)propionic Acid (AMPA) Receptors

On the basis of the results obtained in previous series of AMPA potentiators belonging to 3,4-dihydro-2<i>H</i>-benzo- and 3,4-dihydro-2<i>H</i>-pyrido-1,2,4-thiadiazine 1,1-dioxides, the present work focuses on the design of original isosteric 3,4-dihydro-2<i>H</i>-thieno-1,2,4-thiadiazine 1,1-dioxides. Owing to the sulfur position, three series of compounds were developed and their activity as AMPA potentiators was characterized. In each of the developed series, potent compounds were discovered. After screening the selected active compounds on a safety in vivo test, 6-chloro-4-ethyl-3,4-dihydro-2<i>H</i>-thieno­[2,3-<i>e</i>]-1,2,4-thiadiazine 1,1-dioxide (<b>24</b>) appeared as the most promising compound and was further evaluated. Its effects on long-term potentiation in vivo and on AMPA-mediated noradrenaline release were measured to predict its potential cognitive enhancing properties. Finally, an object recognition test performed in mice revealed that <b>24</b> was able to significantly enhance cognition, after oral administration, at doses as low as 0.3 mg/kg. This study validates the interest of the isosteric replacement of the benzene or pyridine nuclei by the thiophene nucleus in the ring-fused thiadiazine dioxides class of AMPA potentiators

GluA1, GluA2 and GluA4 subunits and splice variants selectivity of S 47445 at human AMPA receptors.

<p>(A) Typical examples of glutamate-evoked currents in absence and presence of 100 μM S 47445 obtained on the same oocyte expressing either GluA1 flip (GluA1i, n = 5) or flop (GluA1o, n = 3) variants. (B) Concentration response curves for S 47445 obtained in oocytes injected with GluA1i and GluA1o receptors. (C) Maximal fold potentiation (E_max) induced by S 47445 on homomeric GluA1 flip (GluA1i), GluA1 flop (GluA1o), GluA4 flip (GluA4i), GluA4 flop (GluA4o) and heteromeric GluA1/GluA2 or GluA4/GluA2 receptors. E_max is given as the amplitude of current evoked in the presence of S 47445 normalized to unity versus the control current evoked by glutamate alone on the same oocytes. n is indicated in brackets. ** p≤0.01 GluA1o versus GluA1i, unpaired T-test. # p≤0.05 GluA1 flop/ GluA2 flip (GluA1o/2i) versus GluA1 flip/ GluA2 flip (GluA1i/2i), Tukey’s test following significant one-way ANOVA performed on GluA1/GluA2 heterodimers. In (A), (B) and (C), cells were held at -80 mV. Values are mean ± SEM.</p

Detailed data of the article

in vitro and in vivo dat

Effect of S 47445 on spontaneous alternation in T-maze in mice (n = 12).

<p>Mice were treated i.p. with either S 47445 (0.1, 0.3 and 1 mg/kg) or vehicle, 30 min prior to the test. Histograms illustrate the mean ± SEM of the percentage of spontaneous alternation of mice. ⁺ p = 0.07 and * p≤0.05 versus vehicle group, one way ANOVA followed by a Dunnett’s test.</p

GluA1, GluA2 and GluA4 subunits and splice variants selectivity of S 47445 at human AMPA receptors.

<p>(A) Typical examples of glutamate-evoked currents in absence and presence of 100 μM S 47445 obtained on the same oocyte expressing either GluA1 flip (GluA1i, n = 5) or flop (GluA1o, n = 3) variants. (B) Concentration response curves for S 47445 obtained in oocytes injected with GluA1i and GluA1o receptors. (C) Maximal fold potentiation (E_max) induced by S 47445 on homomeric GluA1 flip (GluA1i), GluA1 flop (GluA1o), GluA4 flip (GluA4i), GluA4 flop (GluA4o) and heteromeric GluA1/GluA2 or GluA4/GluA2 receptors. E_max is given as the amplitude of current evoked in the presence of S 47445 normalized to unity versus the control current evoked by glutamate alone on the same oocytes. n is indicated in brackets. ** p≤0.01 GluA1o versus GluA1i, unpaired T-test. # p≤0.05 GluA1 flop/ GluA2 flip (GluA1o/2i) versus GluA1 flip/ GluA2 flip (GluA1i/2i), Tukey’s test following significant one-way ANOVA performed on GluA1/GluA2 heterodimers. In (A), (B) and (C), cells were held at -80 mV. Values are mean ± SEM.</p

Chemical structure of S 47445.

<p>Chemical name of S 47455 is 8-cyclopropyl-3-[2-(3-fluorophenyl)ethyl]-7,8-dihydro-3H-[1,3]oxazino[6,5-g][1,2,3]benzotriazine-4,9-dione.</p

Effects of GYKI52466 attenuates the potentiation caused by S 47445 but not the sensitivity.

<p>Determination of the potentiation caused by a series of S 47445 on the response evoked by 10 μM glutamate conducted in absence or presence of GYKI52466. Typical currents evoked by a 10 μM glutamate test pulse recorded in the same cell are illustrated in the inset. Currents were normalized to unity for the maximal value recorded in presence of 30 μM glutamate alone on the same oocytes (n = 3). Note that exposure to GYKI52466 caused a significant reduction of the magnitude of the potentiation but does not alter the sensitivity to S 47445.</p

EC₅₀, EC_2X and maximal potentiation values obtained with S 47445 and other positive allosteric modulators at rat or human AMPA receptors on oocytes injected with poly(A+) mRNA.

<p>EC₅₀, EC_2X and maximal potentiation values obtained with S 47445 and other positive allosteric modulators at rat or human AMPA receptors on oocytes injected with poly(A+) mRNA.</p

EC₅₀ values obtained on different GluA1, GluA2 and GluA4 subunits and flip (i) and flop (o) splice variants at human AMPA receptors.

<p>EC₅₀ values obtained on different GluA1, GluA2 and GluA4 subunits and flip (i) and flop (o) splice variants at human AMPA receptors.</p

Effects of S 47445 on native rat and human AMPA, NMDA or kainate receptors.

<p>(A) Typical currents evoked by AMPA, kainate or NMDA on oocytes injected with rat cortex or human hippocampal poly(A+) mRNA in absence or presence of 100 μM S 47445 recorded in the same cell. (B) Concentration-activity curves obtained with S 47445 on AMPA-, kainate- or NMDA-evoked current on oocytes injected with rat cortex poly(A+) mRNA (n = 3, 3 and 4, respectively). (C) Comparative effects of S 47445 on AMPA-evoked current on oocytes injected with either human hippocampal or rat cortex poly(A+) mRNA (n = 5 and 3, respectively). In (A), (B) and (C), cells were held at -60 mV. The amplitude of current evoked in the presence of S 47445 was normalized to unity versus the control current evoked by the agonist alone on the same oocytes (either 10 μM AMPA, 1 mM kainate or 300 μM NMDA/30 μM glycine). Results are expressed as mean ± SEM.</p