There is no effect of TAFI inhibition of the growth of an established AAA in the Angiotensin II model.

<p>AAA were induced in hyperlipidaemic mice by infusion of Ang II 750 ng/kg/min. After 1 week, mice were treated with a single injection of either MA-TCK26D6 or NaCl control. AAA progression in both groups was evaluated at 2 weeks post injection using Vevo2100 pre-clinical ultrasound scanning, and 3D reconstructions of the aortic segment at risk of AAA formation were created. Panel A Aortic diameter, Panel B Aortic volume, Panel C The process of creating the 3D aortic reconstruction. Panel D Example 3D reconstruction showing AAA progression over time (0, 1 and 3 weeks post Angiotensin II infusion). Data is shown as mean±standard deviation.</p

¹H NMR (500 MHz, methanol-D₄) (top) and ¹³C NMR (100 MHz, methanol-D₄) (bottom) spectra of 3-trifluoromethyl-4-(hydroxymethyl)phenol (18).

1H NMR (500 MHz, methanol-D4) (top) and 13C NMR (100 MHz, methanol-D4) (bottom) spectra of 3-trifluoromethyl-4-(hydroxymethyl)phenol (18).</p

Aortic distensibility decreases with AAA formation, but is not affected by TAFI-inhibition, in the Angiotensin II model of AAA.

<p>The aorta was imaged in longitudinal section using the Vevo2100 scanner, and the distensibility measured using ECG-gated images and VevoVasc software. Panel A shows aortic wall distensibility in all mice with AAA (MA-TCK26D6 treated and sham treated). One week following initiation of Ang II infusion, mice received either MA-TCK26D6 or control (NaCl 0.9%) as an IV injection via the femoral vein. Panel B and C There was no difference in the change of distensibility by week 3 in mice receiving treatment with MA-TCK26D6 at 1 week compared with controls (data shown is change in distensibility between week 1 and week 3). Data is shown as mean±standard deviation, *** p = 0.001 compared to baseline by student t-test.</p

¹H NMR (500 MHz, methanol-D₄) (top) and ¹³C NMR (125 MHz, methanol-D₄) (bottom) spectra of 2,6‐difluoro‐<i>N</i>‐{1‐[(2‐phenoxyphenyl)methyl]‐1<i>H</i>‐pyrazol‐3‐yl}benzamide (8).

1H NMR (500 MHz, methanol-D4) (top) and 13C NMR (125 MHz, methanol-D4) (bottom) spectra of 2,6‐difluoro‐N‐{1‐[(2‐phenoxyphenyl)methyl]‐1H‐pyrazol‐3‐yl}benzamide (8).</p

¹H NMR (400 MHz, DMSO-D₆) (top) and ¹³C NMR (100 MHz, DMSO-D₆) (bottom) spectra of 2,6-difluoro-<i>N</i>-(1<i>H</i>-pyrazol-3-yl)benzamide (21).

1H NMR (400 MHz, DMSO-D6) (top) and 13C NMR (100 MHz, DMSO-D6) (bottom) spectra of 2,6-difluoro-N-(1H-pyrazol-3-yl)benzamide (21).</p

Chemical structures of the small molecule SOCE inhibitors examined in this study and their common names.

Chemical structures of the small molecule SOCE inhibitors examined in this study and their common names.</p

The effect of TAFI inhibition on mortality in the Angiotensin II model of AAA.

<p>Panel A and B Blood pressure (BP) and heart rate (HR) measurements taken in all groups of mice in the second week of the experimental period using the CODA non-invasive BP device. Panel C and D Mortality of mice treated with Ang II, compared with NaCl 0.9% or Ang II plus MA-TCK26D6 or UK-396082. Data is shown as mean±standard deviation. Mortality in this model of AAA typically occurred early (Days 3–8 post initiation of Ang II infusion). * p<0.05 compared to Angiotensin II alone by Chi-Squared testing.</p

¹H NMR (400 MHz, CDCl₃) (top) and ¹³C NMR (100 MHz, CDCl₃) (bottom) spectra of 4,6-dimethoxypyridin-3-yl)boronic acid (30).

1H NMR (400 MHz, CDCl3) (top) and 13C NMR (100 MHz, CDCl3) (bottom) spectra of 4,6-dimethoxypyridin-3-yl)boronic acid (30).</p

S18 Fig -

Example fluorescence over time graph (A) and IC50 (B) for AnCoA4 following 30 minute preincubation and example fluorescence over time graph (C) and IC50 (D) for AnCoA4 following 90 minute preincubation. (TIF)</p

Convergent synthetic route to GSK7975A (9).

Calcium (Ca2+) is a key second messenger in eukaryotes, with store-operated Ca2+ entry (SOCE) being the main source of Ca2+ influx into non-excitable cells. ORAI1 is a highly Ca2+-selective plasma membrane channel that encodes SOCE. It is ubiquitously expressed in mammals and has been implicated in numerous diseases, including cardiovascular disease and cancer. A number of small molecules have been identified as inhibitors of SOCE with a variety of potential therapeutic uses proposed and validated in vitro and in vivo. These encompass both nonselective Ca2+ channel inhibitors and targeted selective inhibitors of SOCE. Inhibition of SOCE can be quantified both directly and indirectly with a variety of assay setups, making an accurate comparison of the activity of different SOCE inhibitors challenging. We have used a fluorescence based Ca2+ addback assay in native HEK293 cells to generate dose-response data for many published SOCE inhibitors. We were able to directly compare potency. Most compounds were validated with only minor and expected variations in potency, but some were not. This could be due to differences in assay setup relating to the mechanism of action of the inhibitors and highlights the value of a singular approach to compare these compounds, as well as the general need for biorthogonal validation of novel bioactive compounds. The compounds observed to be the most potent against SOCE in our study were: 7-azaindole 14d (12), JPIII (17), Synta-66 (6), Pyr 3 (5), GSK5503A (8), CM4620 (14) and RO2959 (7). These represent the most promising candidates for future development of SOCE inhibitors for therapeutic use.</div