Pharmacokinetics (PK) and Toxicity Study in Non-human Primates

All non-human primate studies were conducted in the Good Laboratory Practice (GLP)-level laboratory of the Korea Institute of Toxicology (KIT, Jeollabuk-do, Korea) and approved by the KIT Institutional Animal Care and Use Committee. Cynomolgus monkeys (Macaca fascicularis) were obtained from the Nafovanny captive-breeding primate facility (Dong Nai Province, Vietnam) and housed individually in stainless cages (543 W × 715L × 818H in mm) during acclimation, pre-treatment, and dosing periods. All animals were at least 24 months old. The environment of the animal room was automatically controlled according to the Standard Operating Procedures of KIT (temperature: 20–29°C, relative humidity: 30–70%, light cycle: 12 h at 300–700 Lux, ventilation: 10–20 times/h, air pressure: negative). Males and females were assigned to treatment groups in a stratified manner using the Pristima System based on most recent body weight. Clinical signs, including mortality, moribundity, general appearance, and behavioral changes, were recorded with date, time, and duration. Body weight was measured prior to the first KDS2010 dose. For oral administration, KDS2010 was dissolved in water. In total, 42 monkeys were administered KDS2010 and ultimately sacrificed for autopsy. The four studies conducted on cynomolgus monkeys are summarized in Table ​Table11 and described in greater detail below.

Overview of non-human primates (cynomolgus monkey) studies

1 male/-

1 female/-

4 males/-

4 females/-

3 males/1 male

3 females/1 female

Once a day for 4 weeks

2 weeks for recovery

Repeated toxicity for 4 weeks

Recovery

for 2 weeks

Toxicokinetics

In the pharmacokinetic study (N118029), six male monkeys were assigned to 2 groups receiving either 10 or 30 mg/kg oral KDS2010. Approximately 0.5 mL of blood was collected from the cephalic vein pre-dose (time 0) and 0.5, 1, 2, 4, 6, 8, 10, and 24 h post-dose in tubes containing anti-coagulant (EDTA-2 K). Blood samples were mixed gently, stored in a wet-ice cryo-rack, and centrifuged (approximately 13,200 rpm, 5 min, 4°C) to obtain plasma. The separated plasma was aliquoted into polypropylene tubes and stored frozen until analysis.

In the single-dose escalation study (N118029), each male and female monkey was administered oral KDS2010 at increasing doses of 25, 50, and 100 mg/kg/day once a week. Mortality and general symptoms were observed during the trial period. Body weight was also measured. Blood was collected for hematology, coagulation, and clinical chemistry tests pre-dose (0) and 8, 15, and 22 days post-dose. After administration of 100 mg/kg/day on day 22, an autopsy was performed to observe individual macroscopic findings.

In the repeated 2-week oral administration toxicity study (G217013) for 4-week dose range finding (DRF), monkeys were assigned to 4 groups of one male and one female. Each group was orally administered KDS2010 at 0, 25, 50, or 100 mg/kg/day once a day for 2 weeks. During the test period, general symptoms were monitored, weight and feed intake measured, and ophthalmological tests performed. Clinical pathological tests (hematology, coagulation, blood biochemistry, and urine tests) were also conducted. All animals were autopsied, during which individual macroscopic findings were recorded, organ weights measured, and samples obtained for histopathology.

In the repeated 4-week oral administration toxicity with 2-week recovery and toxicokinetic test (G218027), monkeys were assigned to 4 groups of 3 males and 3 females. Each group was orally administered KDS2010 at 0, 10, 20, or 40 mg/kg/day once a day for 4 weeks. Toxicity tests were conducted as described for study G217013. For toxicokinetic measurements, blood samples were collected pre-dose (time 0) and at 1, 2, 4, 6, 8, 10, and 24 h after oral administration on the first day (day 1) and the last day (week 4; day 28) of dosing.

A non-compartmental analysis module in Phoenix® WinNonlin® (version 6.4) (Certara Inc., CA, USA) was used to calculate pharmacokinetic (PK) parameters. Systemic exposure to KDS2010 was calculated by applying the linear trapezoidal rule to the area under the plasma concentration–time curve from time zero to the last quantifiable time point (AUC_last), and the maximum observed peak plasma concentration (C_max) and the time to reach C_max (T_max) were determined based on the observed data. The apparent terminal elimination half-life (t_1/2) was calculated from the apparent terminal elimination rate constant using the formula t_1/2 = 0.693/kel.