Effect of spironolactone dosage (combined 2 mg/kg/day and 4 mg/kg/day) on RAS-Fingerprint^™ analytes in 10 healthy purpose-bred Beagle dogs at baseline (combined D0 and D21) and post-treatment (combined D7 and D28) using a cross-over study design at 07:00 (T1) prior to morning dosing.

Data are presented as median (IQR) in pM/L for AngI, AngII, aldosterone, Ang1-7, Ang1-5, AngIII, AngIV, and PRA-S and as a ratio for ACE-S, AA2, and ALT-S. P value compares overall treatment to baseline.</p

Effect of spironolactone dosage (combined 2 mg/kg/day and 4 mg/kg/day) in 10 healthy purpose-bred Beagle dogs using a cross-over study design on RAS-Fingerprint^™ analytes at each sampling period at 12:00 (T2) 5-hours after oral dosing of spironolactone.

The sampling periods include two baseline sampling periods at Day 0 (D0) and Day 21 (D21) as well as following two seven-day treatment periods at Day 7 (D7) and Day 28 (D28). Data are presented as median (IQR) in pM/L for AngI, AngII, aldosterone, Ang1-7, Ang1-5, AngIII, AngIV, and PRA-S and as a ratio for ACE-S, AA2, and ALT-S. P value compares overall treatment to baseline at each specific timepoint (D0 vs. D7 and D21 vs. D28).</p

Effect of spironolactone treatment (combined 2 mg/kg/day and 4 mg/kg/day) in 10 healthy purpose-bred Beagle dogs using a cross-over study design on select serum biochemistry variables and SAP between baseline (combined D0 and D21) and treatment (combined D7 and D28) at 07:00 (T1; prior to feeding or morning dosing).

Effect of spironolactone treatment (combined 2 mg/kg/day and 4 mg/kg/day) in 10 healthy purpose-bred Beagle dogs using a cross-over study design on select serum biochemistry variables and SAP between baseline (combined D0 and D21) and treatment (combined D7 and D28) at 07:00 (T1; prior to feeding or morning dosing).</p

Mean (one standard deviation) plasma canrenone and 7-α-thiomethyl spironolactone (TMS) concentration following treatment with spironolactone at either 2 mg/kg/day or 4mg/kg/day for seven days at T1 (immediately prior to spironolactone dosing) and T2 (5 hours post-spironolactone dose) in 10 healthy purpose-bred Beagle dogs using a cross-over study design.

The range of plasma canrenone and TMS at each timepoint following spironolactone treatment is also reported.</p

RAS-Fingerprint^™ analytes at baseline (combined D0 and D21) and post-treatment (D7 and D28 combined) with spironolactone at 2 mg/kg/day and 4 mg/kg/day combined at 07:00 (T1) in 10 healthy purpose-bred Beagle dogs.

There was a significant increase in aldosterone (P Table 3). Spheres show relative concentrations of angiotensin peptides. Blue spheres generally signify that the angiotensin peptide’s action is inert, red indicates predominantly vasoconstrictive and pro-fibrotic effects, and green indicates vasodilatory and anti-fibrotic actions. Enzymes are shown as blue connecting lines between peptides.</p

Effect of spironolactone dosage (combined 2 mg/kg/day and 4 mg/kg/day) on RAS-Fingerprint^™ analytes in 10 healthy purpose-bred Beagle dogs using a cross-over study design at each sampling period at 07:00 (T1) immediately prior to oral dosing of spironolactone.

The sampling periods include two baseline sampling periods at Day 0 (D0) and Day 21 (D21) as well as following two seven-day treatment periods at Day 7 (D7) and Day 28 (D28). Data are presented as median (IQR) in pM/L for AngI, AngII, aldosterone, Ang1-7, Ang1-5, AngIII, AngIV, and PRA-S and as a ratio for ACE-S, AA2, and ALT-S. P value compares overall treatment to baseline at each specific timepoint (D0 vs. D7 and D21 vs. D28).</p

Depiction of study design using a complete cross-over (AB/BA) two-arm design.

Study dogs were randomly allocated to two dosing groups (spironolactone at 2 mg/kg or 4 mg/kg, by mouth every 24 hours). Each treatment was given for seven consecutive days (D1 –D7 and D22 –D28) with a 14-day washout period between treatment periods. Samples were obtained at baseline (D0 and D21) prior to each treatment period and upon the completion of each treatment period (D7 and D28) at two timepoints: T1, immediately prior to daily spironolactone administration (07:00) and T2, peak plasma concentration (12:00).</p

Scatterplot graphs showing the weak association (R < 0.4) between plasma canrenone and plasma TMS concentration and serum aldosterone in 10 healthy purpose-bred Beagle dogs receiving spironolactone at 2mg/kg/day and 4mg/kg/day combined in a cross-over study.

Scatterplot graphs showing the weak association (R < 0.4) between plasma canrenone and plasma TMS concentration and serum aldosterone in 10 healthy purpose-bred Beagle dogs receiving spironolactone at 2mg/kg/day and 4mg/kg/day combined in a cross-over study.</p

Excel file containing raw study data for all dogs at all timepoints (D0, D7, D21, D28).

Data includes dog identification number, spironolactone dose, average blood pressure, complete blood count and serum biochemical profile values, and RAS-Fingerprint™ analytes. Footnotes: aSpironolactone, 25mg tablets, NorthStar Healthcare ULC, bRAS-Fingerprint™, Attoquant Diagnostics, Vienna, Austria. (XLSX)</p

RAS-Fingerprint^™ analytes on Day 21 of the study (baseline) vs. Day 28 of the study (post-treatment with spironolactone at 2 mg/kg/day and 4 mg/kg/day combined) at 07:00 (T1) in 10 healthy purpose-bred Beagle dogs.

There were significant increases in AngII, AngI, Ang1-5, and PRA-S (P Table 4). Spheres show relative concentrations of angiotensin peptides. Blue spheres generally signify that the angiotensin peptide’s action is inert, red indicates predominantly vasoconstrictive and pro-fibrotic effects, and green indicates vasodilatory and anti-fibrotic actions. Enzymes are shown as blue connecting lines between peptides.</p