Development of DANDYs, New 3,5-Diaryl-7-azaindoles Demonstrating Potent DYRK1A Kinase Inhibitory Activity

A series of 3,5-diaryl-1<i>H</i>-pyrrolo­[2,3-<i>b</i>]­pyridines were synthesized and evaluated for inhibition of DYRKIA kinase in vitro. Derivatives having hydroxy groups on the aryl moieties (<b>2c</b>, <b>2j</b>–<b>l</b>) demonstrated high inhibitory potencies with <i>K</i>_is in the low nanomolar range. Their methoxy analogues were up to 100 times less active. Docking studies at the ATP binding site suggested that these compounds bind tightly to this site via a network of multiple H-bonds with the peptide backbone. None of the active compounds were cytotoxic to KB cells at 10^–6 M. Kinase profiling revealed that compound <b>2j</b> showed 2-fold selectivity for DYRK1A with respect to DYRK2 and DYRK3

Theta-burst-induced LTP in Tg<i>hCBS</i>60.4 mice.

<p>Comparison of averaged LTP expressed as percent change in the slope of fEPSP <i>vs</i> time, induced by theta-burst stimulation (TBS, arrow) of glutamate afferents and recorded in slices from Tg<i>hCBS</i>60.4 (12 slices/9 animals) and control (10 slices/7 animals) mice. In the insert, representative traces of fEPSPs, recorded before and 60 min after TBS, are superimposed.</p

Involvement of CBS in sulfur-containing amino acid metabolism.

<p>From Stipanuk <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029056#pone.0029056-Stipanuk1" target="_blank">[54]</a>.</p

CBS expression and activity in brain regions of Tg<i>hCBS</i>60.4 mice.

<p>(A, top) Immunoblots for CBS and β-actin in cerebellum (left panel), hippocampus (middle panel) and forebrain (right panel). Total CBS proteins (human CBS plus mouse CBS) were detected using the anti-hCBS polyclonal antibody that recognises both the human and mouse proteins. (A, bottom) Quantification of CBS proteins normalized to β-actin and relative to control mice in the three brain regions (<i>n</i> = 3 Tg<i>hCBS</i>60.4 and <i>n</i> = 3 control) (B) CBS activity in cerebellum (left panel), hippocampus (middle panel) and forebrain (right panel). Units are nanomoles of cystathionine formed per milligram of protein extract per hour. For cerebellum and forebrain, data represent mean ± S.E.M. of activity measurements in each group (<i>n</i> = 9 Tg<i>hCBS</i>60.4 and <i>n</i> = 7 control; <i>n</i> = 8 Tg<i>hCBS</i>60.4 and <i>n</i> = 8 control, respectively). For hippocampus, data represent mean ± S.E.M. of three experimental assays performed on same pooled hippocampi (<i>n</i> = 7 Tg<i>hCBS</i>60.4 and <i>n</i> = 8 control). *for <i>p</i><0.05. ***for <i>p</i><0.001.</p

Basal synaptic transmission in Tg<i>hCBS</i>60.4 mice.

<p>(A) Superimposed sample traces of evoked AMPA-R-mediated fEPSPs induced in a control (left) and a Tg<i>hCBS</i>60.4 mouse (right) by increased intensities of electrical stimulation of glutamate afferents (arrow). Traces are averages of 3 consecutive responses. (B) Comparison of synaptic efficacy as determined by the fEPSP/PFV ratio calculated at a stimulus intensity from 500 to 900 µA in Tg<i>hCBS</i>60.4 (23 slices/5 animals) and control (22 slices/5 animals) mice.</p

Concentrations (nmoles/mg protein) of 30 amino acids and metabolite compounds in cerebellum extracts of Tg<i>hCBS</i>60.4 and control mice.

<p>Data correspond to mean ± S.E.M. BD: below detection. ND: not determined.</p

Behavioral assessments in Tg<i>hCBS</i>60.4 mice.

<p>The analysis was conducted on 8 Tg<i>hCBS</i>60.4 and 8 control mice. Data represent mean+S.E.M. for each group. (A) Rotarod test: latencies of first fall, second fall and third fall recorded in the 10 min accelerating periods of the two rotarod sessions (S1, S2) are presented. (B) Passive avoidance test: latency to enter into the dark compartment in acquisition session and in retention session was recorded.</p