Genetic associations at 53 loci highlight cell types and biological pathways relevant for kidney function.

Reduced glomerular filtration rate defines chronic kidney disease and is associated with cardiovascular and all-cause mortality. We conducted a meta-analysis of genome-wide association studies for estimated glomerular filtration rate (eGFR), combining data across 133,413 individuals with replication in up to 42,166 individuals. We identify 24 new and confirm 29 previously identified loci. Of these 53 loci, 19 associate with eGFR among individuals with diabetes. Using bioinformatics, we show that identified genes at eGFR loci are enriched for expression in kidney tissues and in pathways relevant for kidney development and transmembrane transporter activity, kidney structure, and regulation of glucose metabolism. Chromatin state mapping and DNase I hypersensitivity analyses across adult tissues demonstrate preferential mapping of associated variants to regulatory regions in kidney but not extra-renal tissues. These findings suggest that genetic determinants of eGFR are mediated largely through direct effects within the kidney and highlight important cell types and biological pathways

Molecular structures of cdc2-like kinases in complex with a new inhibitor chemotype

Cdc2-like kinases (CLKs) represent a family of serine-threonine kinases involved in the regulation of splicing by phosphorylation of SR-proteins and other splicing factors. Although compounds acting against CLKs have been described, only a few show selectivity against dual-specificity tyrosine phosphorylation regulated-kinases (DYRKs). We here report a novel CLK inhibitor family based on a 6,7-dihydropyrrolo[3,4-g]indol-8(1H)-one core scaffold. Within the series, 3-(3-chlorophenyl)-6,7-dihydropyrrolo[3,4-g]indol-8(1H)-one (KuWal151) was identified as inhibitor of CLK1, CLK2 and CLK4 with a high selectivity margin towards DYRK kinases. The compound displayed a potent antiproliferative activity in an array of cultured cancer cell lines. The X-ray structure analyses of three members of the new compound class co-crystallized with CLK proteins corroborated a molecular binding mode predicted by docking studies

Results of a docking experiment with 8a in CLK1 (PDB-ID: 1Z57).

<p>A: front view; B: top view; dashed lines: H-bonds and edge to face interaction.</p

In vitro growth inhibition of cancer cell lines by KuWal151 (8c)^a.

<p>In vitro growth inhibition of cancer cell lines by KuWal151 (8c)<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196761#t002fn001" target="_blank">^a</a>.</p

Structures of and CMGC kinase inhibition by 3-phenyl-6,7-dihydropyrrolo[3,4-<i>g</i>]indol-8(1<i>H</i>)-ones (IC₅₀-values [μM])^a.

<p>Structures of and CMGC kinase inhibition by 3-phenyl-6,7-dihydropyrrolo[3,4-<i>g</i>]indol-8(1<i>H</i>)-ones (IC₅₀-values [μM])<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196761#t001fn001" target="_blank">^a</a>.</p

Data collection and refinement statistics of CLK/ligand complexes.

<p>Data collection and refinement statistics of CLK/ligand complexes.</p

CLK1 inhibitors described in the literature.

<p>TG003 (<b>1</b>); NCGC00185963 (<b>2</b>), KH-CB19 (<b>3</b>); benzo[<i>b</i>]thiophen-2-carboxamide <b>4</b>; T3 (<b>5</b>); TG693 (<b>6</b>); [1,2,3]triazolo[4,5-<i>c</i>]quinoline <b>7</b>.</p

Comparison of co-crystal structures of 8g (PDB-ID: 6FT8; upper row) and 16 (PDB-ID: 6FT9; lower row) in complex with CLK1, respectively; red spheres: water molecules; black dashed lines: hydrogen bonds.

<p>A: Top view of <b>8g.</b> B: Side view of <b>8g</b>. C: Top view of <b>16</b>. D: Side view of <b>16</b>. A, C: While the lactam motives of the ligands perform the canonical hydrogen bonds to the hinge region, the indole nitrogen atoms are connected to a conserved water molecule. B, D: An area near the Lys191 and Glu206 side chains, unoccupied by both <b>7g</b> and <b>16</b>, is filled by three water molecules. The opening to the entrance of the binding pocket is delimited by Asp250. Compared to the shape of the pocket with bound <b>8g</b> (B), the entrance of the binding pocket is widened to accommodate the 2-bromo substituent of <b>16</b> (D).</p

3-Aryl-6,7-dihydropyrrolo[3,4-<i>g</i>]indol-8(1<i>H</i>)-ones listed in Table 1.

<p>3-Aryl-6,7-dihydropyrrolo[3,4-<i>g</i>]indol-8(1<i>H</i>)-ones listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196761#pone.0196761.t001" target="_blank">Table 1</a>.</p

Synthesis procedures for 3-aryl-6,7-dihydropyrrolo[3,4-<i>g</i>]indol-8(1<i>H</i>)-ones.

<p>For residues R^<b>1</b>-R^<b>5</b> refer to <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0196761#pone.0196761.t001" target="_blank">Table 1</a>. Reagents and conditions: (reagents and conditions a, for synthesis of <b>10b</b>) NBS, CH_<b>2</b>Cl_<b>2</b>, - 8°C, 1 h, 78%; (reagents and conditions b, for synthesis of <b>10c</b>) NCS, acetonitrile, 60°C → reflux, 2 h, 81%; (reagents and conditions c, for synthesis of <b>10d</b>) CH_<b>3</b>I, KO<i>t</i>Bu, THF, RT, N_<b>2</b>, 24 h, 54%; (d) 1. 37% HCl, NaNO_<b>2</b>, < 0°C; 2. 37% HCl, SnCl_<b>2</b> x 2 H_<b>2</b>O, 30 min; (e) aldehyde or ketone or acetal, acetic acid, 95°C, 3.5 h, 10%-31%; (f) PdCl_<b>2</b>(MeCN)_<b>2</b>, <i>p</i>-benzoquinone, <i>tert</i>-butanol, water, 80°C; (g) <b>11a</b>, ethanol, H_<b>2</b>SO_<b>4</b>, H_<b>2</b>O, 50°C, 2.5 h, 23%; (h) BBr_<b>3</b>, CH_<b>2</b>Cl_<b>2</b>, RT, N_<b>2</b>, 1 h, 33%; (i) acetic anhydride, pyridine, 4-DMAP, RT, 3 h, 29%; (j) alkyl halide, KO<i>t</i>Bu, acetone, RT, N_<b>2</b>, 24 h, 15%-48%; (k) NBS, CH_<b>2</b>Cl_<b>2</b>/acetic acid, < 10°C, N_<b>2</b>, 1.5 h, 29%; (l) appropriate arylboronic acid, Cs_<b>2</b>CO_<b>3</b>, toluene/ethanol, mircowaves, 150°C, 20 min, 5.4%-31%.</p