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Keywords:

  • biomarker;
  • DPP IV;
  • LC15-0444;
  • pharmacodynamics;
  • pharmacokinetics

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

WHAT IS ALREADY KNOWN ABOUT THIS SUBJECT

• The importance of efficient drug development using biomarkers has been increasingly emphasized, from preclinical studies to clinical trials.

• However, as yet few ‘validated’ or ‘qualified’ biomarkers are used in early-stage drug development in terms of clinical pharmacology and disease pathophysiology.

WHAT THIS STUDY ADDS

• This first-time-in-human study provides evidence of the pharmacological activity of LC15-0444 in humans, by using dipeptidyl peptidase IV activity and active glucagon-like peptide-1 concentrations.

• LC15-0444 possesses pharmacokinetic and pharmacodynamic characteristics that support a once-daily dosing regimen.

AIMS LC15-0444 is a selective and competitive inhibitor of dipeptidyl peptidase (DPP) IV with potential for the treatment of Type 2 diabetes. The aim was to investigate the pharmacokinetic (PK) and pharmacodynamic (PD) profiles after multiple oral ascending doses of LC15-0444 in healthy male subjects.

METHODS A dose block-randomized, double-blind, placebo-controlled, parallel group study was performed in three groups with 10 subjects (eight for active drug; two for placebo) per group; each group received 200, 400 or 600 mg of LC15-0444 once daily for 10 days. Blood and urine samples were collected up to 24 h after the first dosing and up to 72 h after the last dosing.

RESULTS The LC15-0444 concentration–time profiles exhibited characteristics of multicompartment disposition. No dose- or time-dependent change in PK parameters was observed. Mean elimination half-life was in a range 16.6–20.1 h in the dose groups. Mean renal clearance and fraction of unchanged drug excreted in urine was 18.6–21.9 and 0.40–0.48 l h−1, respectively. In the steady state, mean accumulation ratios by dose groups were between 1.22 and 1.31. More than 80% inhibition of DPP IV activity from baseline was sustained for >24 h in all dose groups.

CONCLUSIONS This study provides evidence of the pharmacological activity of LC15-0444 in humans. LC15-0444 possesses PK and PD characteristics that support a once-daily dosing regimen.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

Dipeptidyl peptidase (DPP) IV inhibitors represent a new therapeutic approach to the treatment of Type 2 diabetes [1]. The therapeutic utility of these antihyperglycaemic agents rests on their ability to increase active levels of incretin peptides, including glucagon-like peptide-1 (GLP-1). Active GLP-1 is normally released from enteroendocrine L cells into the circulation after a meal to increase the release of insulin, but it is rapidly inactivated by DPP IV [2]. DPP IV inhibitors delay degradation of the active GLP-1, thereby promoting the release of insulin and glycaemic control.

LC15-0444 is a selective and competitive inhibitor of DPP IV, currently in clinical development for the treatment of Type 2 diabetes. Its mechanism of action is the direct inhibition of DPP IV, resulting in increased GLP-1 levels, increased insulin responses and improved glycaemic control. Preclinical studies indicate that LC15-0444 inhibits DPP IV activity, elevates active GLP-1 levels, and reduces glucose excursion after an oral glucose load (unpublished data). A metabolite of LC15-0444 was quantified as <8% of parent exposure in human. It was also found in rats and dogs in a similar ratio. The identification of other possible metabolites is on-going. The objectives of this study were to evaluate the pharmacokinetic (PK) and pharmacodynamic (PD) characteristics of LC15-0444 after multiple oral administrations in healthy male subjects.

Methods

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

Subjects and study design

Thirty healthy nonsmoking Korean male volunteers were enrolled in the study. The mean age was 26.0 years (range 19–38 years) and mean body weight was 69.2 kg (range 54.5–81.4 kg). There were no abnormalities in physical examination, vital signs, routine laboratory tests or 12-lead electrocardiogram, and none had any relevant medical disorders. All subjects denied any medication use within the 4-week period before the study, and this was confirmed by urine drug screening using REMEDi HS® (Bio-Rad Laboratories, Hercules, CA, USA). Written informed consent was obtained from all subjects after the study procedures had been fully explained. The study was approved by the Institutional Review Board of Seoul National University Hospital (SNUH).

This was a dose block-randomized, double-blind, placebo-controlled study. This multiple ascending dose, parallel-group study was performed in three groups with 10 male subjects (including two placebo subjects) per group: each group was administered 200, 400 or 600 mg of the study drug once a day for 10 days. Dose escalation to the next dose proceeded only after the safety of the previous dose had been confirmed. After overnight fasting, each dose was administered at approximately 09.00 h. Blood and urine samples for evaluations of PK and PD were collected up to 24 h after the first dosing and up to 72 h after the last dosing of the study drug.

Assay of LC15-0444 levels in plasma and urine

Following deproteinated extraction using acetonitrile with 2% formic acid, plasma concentrations of LC15-0444 were measured by tandem mass spectrometry (MS; API 4000; Applied Biosystems/MDS Sciex, Toronto, Canada) coupled with high-performance liquid chromatography (HPLC; Agilent 1100 series; Agilent Technologies, Wilmington, DE, USA). Chromatographic separation was achieved under gradient conditions on a Gemini C18 110A column (50 × 3.0 mm, 3 µm; Phenomenex, Torrance, CA, USA) with a mobile phase A (10 mM ammonium acetate : acetonitrile = 90:10, v/v) with 0.1% formic acid and a mobile phase B (10 mM ammonium acetate : methanol : acetonitrile = 10:45:45, v/v) with 0.1% formic acid. The MS/MS system was operated using an electrospray in positive ionization mode. For LC15-0444 and the internal standard, LC15-0510, the precursor-to-product ion reactions monitored were m/z 490.243[RIGHTWARDS ARROW]338.071 and 488.160[RIGHTWARDS ARROW]336.079, respectively. The lower limit of quantification (LLOQ) was 0.5 ng ml−1. The day-to-day coefficients of variation (CV) were 6.5% at 1 ng ml−1, 3.6% at 10 ng ml−1 and 4.3% at 1600 ng ml−1.

Urine concentrations of LC15-0444 were measured by HPLC (SymbiosisPharma, Spark, Plainsboro, NJ, USA) with an MS/MS (API 4000QTRAP; Applied Biosystems/MDS Sciex) following deproteinated extraction using acetonitrile with 2% formic acid. Chromatographic separation was achieved under gradient conditions on a Gemini C18 110A column (50 × 3.0 mm, 5 µm; Phenomenex) with the same mobile phase as plasma, as described above. The MS/MS system was operated using an electrospray in positive ionization mode. For LC15-0444 and LC15-0510, the precursor-to-product ion reactions monitored were the same as those of plasma described above. The LLOQ was 4 ng ml−1. The day-to-day CVs were 3.5% at 8 ng ml−1, 2.0% at 200 ng ml−1 and 2.0% at 1600 ng ml−1.

Pharmacokinetic assessments

Blood sampling for plasma LC15-0444 was performed just before dosing (0 h) and at 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 12 and 16 h after dosing (day 1), and predose on days 2–9, and just before dosing (0 h) and 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 12, 16, 24, 36, 48 and 72 h after dosing (day 10). Blood (6 ml) was collected in ethylenediamine tetraacetic acid (EDTA) tubes, shaken gently, and kept in an ice box temporarily for 10 min. Afterwards, samples were centrifuged for 10 min at 1600 g; two aliquots of 1.0 ml were transferred to cryotubes prefilled with 1.0 ml 5% formic acid (88%) in water, frozen at −20°C for 24 h, and finally stored at −70°C until analysis. Urine samples were collected after dosing during intervals of 0–6, 6–12 and 12–24 h (day 1), and 0–6, 6–12, 12–24, 24–48 and 48–72 h (day 10), for determination of the LC15-0444 concentrations. Urine was collected and stored at −4°C immediately; during each interval two aliquots of 1.0 ml were transferred to cryotubes prefilled with 1.0 ml 5% formic acid (88%) in water, and frozen at −20°C for 24 h, and finally stored at −70°C until analysis.

Individual PK parameters were calculated by noncompartmental methods using WinNonlin® (version 5.1; Pharsight Corp., Mountain View, CA, USA). Terminal elimination half-life was calculated from linear regression of log-transformed plasma concentration and time. AUC (AUC0,τ, AUCss,τ and AUC0–∞,1d) was calculated using the linear-up and log-down trapezoidal method in plasma concentration–time curves. AUC0–∞,1d was calculated as AUC0,τ+ C24z, where C24 is the plasma concentration just before the second dose, and λz is the terminal elimination rate constant. The accumulation ratio was calculated as AUCss,τ divided by AUC0,τ. Apparent clearance (CL/F) after the first dose was calculated as the administered dose over AUC0–∞,1d, and CL/F in the steady state was calculated as the administered dose over AUCss,τ. Renal clearance on day 1 was calculated as the cumulative amount of unchanged drug excreted in urine (Ae0,τ) divided by AUC0,τ, and that of day 10 was calculated as Aess,τ over AUCss,τ. Ae was calculated by first multiplying urinary concentrations by the corresponding urine volumes during each collection interval, and then summing the amount from time 0 to 24 after the first dose (Ae0,τ) or the last dose (Aess,τ). The fraction of unchanged drug excreted in urine of day 10 was calculated as Aess,τ over the dose.

Pharmacodynamic assessments

Blood samples for the measurement of various biomarkers related to the pharmacology of this drug were obtained at selected time points. For the DPP IV activity assay, blood samples were collected just before dosing (0 h) and at 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 12 and 16 h after dosing (day 1), and predose on days 2–9, and just before dosing (0 h) and 0.5, 1, 1.5, 2, 3, 4, 5, 6, 8, 12, 16, 24, 36, 48 and 72 h after dosing (day 10). Blood for DPP IV activity was collected in EDTA tubes, shaken gently and kept in an ice box temporarily for 10 min. Afterwards, samples were centrifuged for 10 min at 1600 g; two aliquots of 100 µl were transferred to cryotubes, frozen at −20°C for 24 h and finally stored at −70°C until analysis. For active GLP-1, insulin and glucose assay, standardized regular meals were served at 4, 10 and 24 h, and blood sampling was performed at 4 (premeal), 4.25, 4.5, 5, 6, 10 (pre-meal), 10.25, 10.5, 11, 12, 24 (pre-meal), 24.25, 24.5, 25 and 26 h after first dosing (day 1) and last dosing (day 10). Blood (5 ml) for active GLP-1 was collected in EDTA tubes (pretreated with protease inhibitor) immediately, shaken gently and kept in an ice box temporarily for up to 1 h. Samples were then centrifuged for 10 min at 1100 g; two aliquots of 250 µl for active GLP-1 were transferred to cryotubes, frozen at −20°C for 24 h and finally stored at −70°C until analysis. Blood (3 ml) for serum glucose was collected in serum separator tubes and kept at room temperature for up to 1 h. Afterwards, samples were analysed by the Department of Laboratory Medicine, SNUH. Blood (3 ml) for whole-blood insulin was collected in EDTA tubes, shaken gently and kept in an ice box temporarily for up to 1 h. Samples were then analysed by the Department of Nuclear Medicine, SNUH.

For DPP IV, the percent inhibition from baseline activity was calculated. For active GLP-1, glucose and insulin, time-weighted average concentrations for 2 h after the start of each meal (AUC over 2 h) were calculated.

DPP IV enzyme assay  DPP IV activity in human plasma was determined using a continuous spectrophotometric assay with the substrate Gly-Pro-pNA (Bachem, Bubendorf, Switzerland). Enzyme activity was determined by measuring the increase in absorbance at 390 nm resulting from the cleavage of the substrate Gly-Pro-pNA by DPP IV. A typical reaction solution consisted of 40% human plasma, 400 µM Gly-Pro-pNA and 50 mM N-2-hydroxyethylpiperazine-N′-2-ethanesulphonic acid buffer (pH 7.4) in a total volume of 100 µl. The release of pNA was measured continuously for 15 min using a 96-well plate spectrophotometer (SpectraMax 340; Molecular Devices Corp., Sunnyvale, CA, USA) at 30°C. Enzyme activity was defined as the slope (in mOD min−1) from 4 to 14 min. Within-run precisions of the sample with the highest and lowest DPP IV activity were 0.96 and 1.25%, respectively. Between-run precisions of the respective samples were 2.06 and 2.42%. The range satisfying precision and linearity criteria was 0.637–64.3 mOD min−1.

Active GLP-1, glucose and insulin assays  Plasma active GLP-1 was assayed with an enzyme-linked immunosorbent assay kit according to the manufacturer's specifications (Linco Research Inc., St Charles, MO, USA). The range between the lower and upper concentrations satisfying precision, accuracy and linearity criteria was 2.0–100 pmol l−1. Serum glucose levels were measured with an oxidase enzymatic assay on an automated analyser (TBA 200FR-1; Toshiba Co. Ltd., Tokyo, Japan). This assay was linear in the range of 0–600 mg dl−1 and the interassay CV was <2.0%. Whole blood insulin levels were measured using an immunoradiometric assay (IRMA) based on coated-tube separation, using a BioSource INS-IRMA Kit (BioSource, Nivelles, Belgium). This assay had a working range of 1.0–416 µIU ml−1 and an interassay CV of 6.5%.

Safety and tolerability

Adverse events (AEs) were monitored throughout the study. In addition, various evaluations were performed before the study and at various time points after study drug administration, as follows: physical examinations, vital sign measurements, 12-lead electrocardiography, clinical laboratory tests including haematology, blood chemistry, coagulation and urinalysis, and computerized impedance cardiography.

Statistical analysis

For PK data, summary statistics are presented as the mean ± standard deviation (SD). For PD data, summary statistics are presented as the mean ± standard error (SE). As for AUCτ,ss and Cmax,ss, dose linearity was tested using linear regression by log-transformed values (power model) [3]. If 95% confidence intervals (CI) of the slope of linear regression included unity, it was considered to have linear PK characteristics. A linear anova model was applied to compare PD parameters among the treatment groups, and Dunnett's test was used for multiple comparisons. The placebo group consisted of six subjects, originating from the two placebo subjects from each dosing group pooled together. SPSS® version 12.0 (SPSS Inc., Chicago, IL, USA) was used for statistical analysis, and the level of significance was 0.05.

Results

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

Pharmacokinetics

LC15-0444 was readily absorbed during multiple oral dosing, reaching maximum plasma levels (Tmax,ss) at 1.0–5.0 h after dosing and apparently achieving a steady state after the second dose (Figure 1). The terminal elimination half-life (t1/2) measured after the last dose was 16.6–20.1 h across the dose groups (Table 1). The difference in t1/2 between day 1 and day 10 was attributed to different sampling schemes (up to 24 h on day 1 vs. up to 72 h on day 10). This caused underestimation of the area under the plasma LC15-0444 concentration–time curve (AUC) extrapolated to infinity (AUC0–∞) after first dose, compared with the AUC during the dosing interval of the steady state (AUCss,τ), resulting in relatively larger apparent clearance (CL/F) on day 1 compared with that of day 10 (Table 1). The mean CL/F by dose groups ranged from 43.0 to 52.7 l h−1, under steady-state conditions. The AUC during the dosing interval (AUC0,τ and AUCss,τ) generally increased in proportion to the dose over the dose range studied. The mean accumulation ratios were between 1.22 and 1.31 among the dose groups.

image

Figure 1. Mean plasma concentration–time profiles of LC15-0444. Data are presented as mean ± SD. 200 mg (N = 8) (—●—); 400 mg (N = 8) (—▵—); 600 mg (N = 8) (inline image)

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Table 1.  Summary of mean pharmacokinetic parameters of LC15-0444 in healthy male subjects
 200 mg (n= 8)400 mg (n= 8)600 mg (n= 8)
Day 1Day 10Day 1Day 10Day 1Day 10
  1. Tmax, time to maximum concentration observed in plasma; t1/2, terminal elimination half-life; Cmax, maximum concentration observed in plasma; AUCτ, area under the concentration–time curve during the dosing interval; AUC0–∞, area under the concentration–time curve from time 0 to infinity; R, accumulation ratio; CL/F, apparent clearance; CLR, renal clearance; Ae0,τ, cumulative amount of unchanged drug excreted in urine from time 0 to 24; fe, fraction of unchanged drug excreted in urine. All values represent arithmetic means (CV %), except for Tmax, median [range].

Tmax (h)1.82.52.01.83.01.8
(1.0–5.0)(1.0–4.0)(1.0–4.0)(1.0–3.0)(1.0–5.0)(1.0–4.0)
t1/2 (h)9.320.17.416.68.417.4
(14.3%)(8.5%)(16.3%)(8.2%)(18.1%)(8.1%)
Cmax (µg l−1)511.2626.71078.71118.31 739.82 230.5
(19.2%)(22%)(23.1%)(26.1%)(20.2%)(20.5%)
AUCτ (µg*h l−1)3444.94532.36477.97856.511 422.014 122.0
(14.1%)(20.9%)(19.3%)(19.5%)(10.3%)(11.8%)
AUC0–∞ (µg*h l−1)3980.57184.412 747.2
(16.7%)(21.8%)(12.1%)
R1.311.221.24
(10.4%)(9.7%)(7.7%)
CL/F (l h−1)51.445.858.652.747.743.0
(15.2%)(20.3%)(13.1%)(20.3%)(11.9%)(10.7%)
CLR (l h−1)16.321.921.620.915.518.6
(24.8%)(34.1%)(35.2%)(16.8%)(17.4%)(12.9%)
Ae0,τ (mg)1.475.676.6112.27.9119.1
(32.1%)(25.5%)(37.0%)(40.6%)(19.5%)(26.9%)
fe0.480.400.44
(30.5%)(14.9%)(15.1%)

The mean fraction of unchanged drug excreted in urine by dose group was independent of dose and in the range 0.40–0.48. Renal clearance (CLR) of LC15-0444 ranged from 18.6 to 21.9 l h−1 or 310 to 365 ml min−1 at steady state (Table 1).

Regarding dose linearity in the PK parameters, the 95% CIs of the slope of linear regression for log-transformed Cmax,ss and AUCss,τ were 0.89, 1.36 and 0.84, 1.20, respectively (power model, Figure 2), so it was considered to possess linear PK properties. No dose- or time-dependent change in CL/F or CLR of LC15-0444 was observed.

image

Figure 2. Linear regressions of LC15-0444 pharmacokinetic parameters after multiple oral administrations of 200, 400 or 600 mg to subjects. Left, Cmax,ssvs. dose; right, AUCss,τvs. dose. Regression line (——); 95% Confidence interval (········)

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Pharmacodynamics

LC15-0444 produced a dose-related inhibition of plasma DPP IV activity over the dose range studied (Figure 3). Plasma DPP IV activity was significantly inhibited at all doses compared with placebo (P < 0.001 at every measured time point after dosing). Inhibition of DPP IV activity by >80% was maintained for the whole duration of the dosing interval after the first dose, for at least 36 h after the last dose, and for 48 h in the 600-mg group.

image

Figure 3. Inhibition from baseline plasma dipeptidyl peptidase IV activity–time profiles during multiple oral administrations of LC15-0444. Data are presented as mean ± SE. Placebo (N = 6) (—⋆—); 200 mg (N = 8) (inline image); 400 mg (N = 8) (—▵—); 600 mg (N = 8) (inline image)

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The weighted average plasma concentrations of active GLP-1 were remarkably higher with all doses of LC15-0444 compared with placebo (Figure 4). Multiple doses of LC15-0444 produced 1.8–2.8-fold increases in weighted active GLP-1 levels after meals, compared with placebo, at 4, 10 and 24 h after administration (Table 2).

image

Figure 4. Weighted average plasma concentrations of active glucagon-like peptide-1 by dose groups after single and multiple oral administrations of LC15-0444. Data are presented as mean ± SE. *P < 0.05, †P < 0.01, ‡P < 0.001 compared with placebo, using Dunnett's test. Placebo (N = 6) (inline image); 200 mg (N = 8) (inline image); 400 mg (N = 8) (inline image); 600 mg (N = 8) (inline image)

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Table 2.  Weighted average plasma concentrations (pmol l-1) of active glucagon-like peptide 1 (GLP-1) over 2 h after standardized meals following once-daily administration of LC15-0444 or placebo for 10 days in healthy male subjects
TreatmentDay 1Day 10
4 h after dosing10 h after dosing24 h after dosing4 h after dosing10 h after dosing24 h after dosing
Mean*GMR95% CIMean*GMR95% CIMean*GMR95% CIMean*GMR95% CIMean*GMR95% CIMean*GMR95% CI
  • *

    Geometric least squares means.

  • †GMR and 95% CI were for the comparisons between the active doses and placebo, and were obtained by Dunnett's test. CI, confidence interval; GMR, geometric mean ratio based on least squares means.

Placebo (n= 6)4.12  4.29  5.28  4.07  3.79  4.38  
200 mg (n= 8)8.792.131.46, 3.138.802.051.40, 4.008.811.671.14, 2.447.841.931.18, 3.148.352.201.43, 3.397.791.781.08, 2.93
400 mg (n= 8)7.171.741.19, 2.557.601.771.21, 2.5911.342.151.47, 3.148.522.091.28, 3.417.752.041.33, 3.159.162.091.27, 3.44
600 mg (n= 8)13.683.322.27, 4.8611.882.771.89, 4.0512.792.421.66, 3.559.802.411.48, 3.9210.692.821.83, 4.3411.152.551.55, 4.19

Multiple doses of LC15-0444 generally decreased the postprandial glucose levels, and there was a statistically significant decrease in glucose at 10 h after dosing on day 10, but had no remarkable effect on insulin levels. Results for the 10-h period post dose are shown in Table 3.

Table 3.  Weighted average plasma concentrations of glucose and insulin over 2 h after a standardized meal ingested at 10 h following once-daily administration of LC15-0444 or placebo for 10 days in healthy male subjects
TreatmentGlucose (mg dl−1)Insulin (µIU ml−1)
Mean*GMR95% CIMean*GMR95% CI
  • *

    Geometric least squares means.

  • †GMR and 95% CI were for the comparisons between the active doses and placebo, and were obtained by Dunnett's test. CI, confidence interval; GMR, geometric mean ratio based on least squares means.

Placebo (n= 6)142.6  53.6  
200 mg (n= 8)121.90.850.75, 0.9750.70.950.55, 1.64
400 mg (n= 8)126.80.890.78, 1.0145.80.850.49, 1.48
600 mg (n= 8)112.30.790.69, 0.9048.90.910.53, 1.58

Safety and tolerability

LC15-0444 was generally well tolerated. None of the subjects developed any serious clinical or laboratory adverse experiences, or discontinued the study due to an AE. All AEs were mild or moderate, and no dose-related trends were observed. Thirteen of the 30 subjects (43.3%) reported a total of 26 AEs (Table 4). Of these AEs, a total of 14 (11 in the LC15-0444 group and three in the placebo group) were considered to be possibly related to the study drug. AEs considered to be related to the study drug were headache and dizziness (three cases each), somnolence, dyspepsia, aphthous stomatitis, rash morbilliform, hyperthermia, pyrexia, palpitations and increased heart rate (a single case each). All AEs resolved spontaneously without any concomitant medication. There were no reports of hypoglycaemic symptoms or signs, or of any significant abnormalities on clinical laboratory tests, 12-lead electrocardiogram, or impedance cardiography.

Table 4.  Adverse events (AEs) per system organ class and treatments (n= 30)
System organ class/AETreatment group/dose
PlaceboGroup 1Group 2Group 3
 200 mg400 mg600 mg
n= 6n= 8n= 8n= 8
  1. Data are no. of events, no. (%) of subjects.

Cardiac disorders1, 1 (12.5)
 Palpitations1, 1 (12.5)
Gastrointestinal disorders1, 1 (16.7)1, 1 (12.5)
 Aphthous stomatitis1, 1 (16.7)
 Dyspepsia1, 1 (12.5)
General disorders and administration site conditions1, 1 (16.7)2, 1 (12.5)
 Feeling hot1, 1 (12.5)
 Pyrexia1, 1 (12.5)
 Vessel puncture site bruise1, 1 (16.7)
Investigations1, 1 (12.5)
 Heart rate increased1, 1 (12.5)
Musculoskeletal and connective tissue disorders1, 1 (12.5)
 Myalgia1, 1 (12.5)
Nervous system disorders3, 2 (33.3)3, 2 (25.0)1, 1 (12.5)5, 4 (50.0)
 Dizziness2, 1 (16.7)1, 1 (12.5)1, 1 (12.5)
 Dizziness postural1, 1 (12.5)
 Headache2, 2 (25.0)2, 2 (25.0)
 Paraesthesia1, 1 (16.7)1, 1 (12.5)
 Somnolence1, 1 (12.5)
Respiratory, thoracic and mediastinal disorders4, 2 (33.3)1, 1 (12.5)
 Cough1, 1 (16.7)
 Epistaxis3, 1 (16.7)1, 1 (12.5)
Skin and subcutaneous tissue disorders1, 1 (12.5)
 Rash mobilliform 1, 1 (12.5)
Total9, 4 (66.7)4, 2 (25.0)6, 2 (25.0)7, 5 (62.5)

Discussion

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

After multiple oral daily doses of LC15-0444, the AUC and Cmax increased in general proportion to the dose. The accumulation of LC15-0444 was modest, with mean AUC accumulation ratios ranging from 1.22 to 1.31 across the dose range of 200–600 mg administered once a day. This is not unexpected in the accumulation of a drug with a terminal elimination half-life of 16.6–20.1 h, administered every 24 h.

Approximately 40–48% of the oral dose was excreted unchanged in the urine, and the mean CLR was estimated to be 310–365 ml min−1. CLR and the fraction of dose excreted unchanged in the urine were not different between the doses. LC15-0444 is likely to be excreted through an active renal secretion process, because the CLR of LC15-0444 exceeds the typical glomerular filtration rate of 125 ml min−1 in subjects of similar age and normal renal function [4].

Multiple dosing of LC15-0444 produced dose-related, but ultimately almost saturated inhibition of plasma DPP IV activity. The pattern of DPP IV inhibition in the steady state was similar to that observed on day 1. All doses were associated with an inhibition of DPP IV activity of >80% over 24 h after a single dose and in the steady state. This level of inhibition is equivalent to the target level of DPP IV inhibition required for near-maximal acute glucose lowering in rodent models [5]. These findings, together with the observation that the apparent terminal t1/2 ranges from 16.6 to 20.1 h, support a once-daily dosing regimen for LC15-0444 in diabetic patients.

Further evidence of the pharmacological activity of LC15-0444 in the steady state was obtained by measuring active GLP-1 levels. Consistent with the marked inhibition of DPP IV activity, LC15-0444 resulted in increases in the postprandial rise in the active GLP-1 levels, an incretin hormone that is degraded by plasma DPP IV [6]. LC15-0444 produced postprandial increases in active GLP-1 that were 1.8–2.8-fold higher than corresponding values for placebo. This increase is also sufficient for near-maximal short-term glucose-lowering efficacy, based on previous reports [5, 7, 8]. The sustained inhibition of DPP IV and augmentation of active GLP-1 levels provide pharmacological proof of concept for LC15-0444 in humans.

There was no clinically meaningful effect of LC15-0444 on fasting and postprandial insulin levels. However, multiple doses of LC15-0444 decreased fasting and postprandial glucose levels at 4, 10 and 24 h after multiple dosing. A previous study has show no meaningful changes in insulin and glucose after multiple oral administrations of sitagliptin in healthy normoglycaemic subjects [8]. This can be explained by the fact that although active GLP-1 levels were enhanced by sitagliptin, these levels were within physiological range, so significant changes in insulin and glucose were not observed in nondiabetic subjects [8–10]. The significant decrease in weighted average glucose levels after LC15-0444 dosing might be explained by the fact that LC15-0444 produced more prominent postprandial increases in active GLP-1, especially at 10 h after a meal (2.0–2.8-fold) than sitagliptin (1.9–2.3-fold in the once-a-day 200–600-mg dose groups) [8].

Administration of multiple doses of LC15-0444 over 10 days did not result in any serious AEs or discontinuation of the medication. Clinical AEs associated with LC15-0444 were qualitatively similar to those seen with placebo or drugs of the same class and were generally transient, self-limiting, and mild in severity. In particular, there were no episodes of hypoglycaemic symptoms or signs. LC15-0444 was not associated with any clinically significant changes as assessed by vital signs, clinical laboratory tests, or electrocardiographic parameters such as QT- or QTc-interval prolongation.

Since the US Food and Drug Administration released the ‘Critical Path Initiative’ report, the importance of efficient drug development using biomarkers has been increasingly emphasized, from preclinical studies to clinical trials [11]. However, as yet few ‘validated’ or ‘qualified’ biomarkers are used in early-stage drug development in terms of clinical pharmacology and disease pathophysiology [12].

The development of a DPP IV inhibitor is one of the most successful cases to date using proximal or target engagement biomarkers, such as DPP IV and active GLP-1, and also distal or disease biomarkers, such as glucose and insulin, in early-phase clinical trials [7, 8, 13]. The present PK/PD and tolerability study is the first assessment of multiple doses of LC15-0444 in human subjects; PK/PD and tolerability profiles of LC15-0444 after single dose in healthy subjects have previously been reported [14].

In summary, multiple daily doses of LC15-0444 exhibited little accumulation, inhibited plasma DPP IV activity by >80% over a 24-h dosing interval, and significantly increased active GLP-1 levels after a standard meal. Furthermore, LC15-0444 decreased postprandial glucose levels and was generally well tolerated in healthy male subjects. This study provides pharmacological proof of concept for LC15-0444 in humans, and supports a once-daily dosing regimen for further clinical development in Type 2 diabetic patients.

Competing interests

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES

D-K.K., H.J.Y., S-H.L. and S-H.K. are employees of LG Life Sciences Ltd. None of the other authors have any competing interests to declare.

Presented in part as a poster at the 109th annual meeting of the American Society for Clinical Pharmacology and Therapeutics, Orlando, FL, 2–5 April 2008. An abstract has been published in Clin. Pharmacol. Ther. 2008; 83 (Suppl. 1): S95. This study was supported by LG Life Sciences Ltd., Seoul, Republic of Korea (H-0608-028-180). It was also partially supported by a grant from the Ministry of Knowledge Economy (Bio-Star, 10024075).

REFERENCES

  1. Top of page
  2. Abstract
  3. Introduction
  4. Methods
  5. Results
  6. Discussion
  7. Competing interests
  8. REFERENCES
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