Mechanism of Action

Apremilast is a novel oral small-molecule inhibitor of PDE4 (IC₅₀ = 74 nM), a cAMP-specific PDE as a dominant PDE in inflammatory cells.

Apremilast inhibits PDE4 and elevates intracellular cAMP levels, which in turn downregulates the inflammatory response by modulating the expression of pro- and anti-inflammatory cytokines, such as tumor necrosis factor alpha (TNF-α), interleukin (IL)-23, IL-17, and anti-inflammatory cytokines.

The metabolites of apremilast had no significant inhibition of PDE4, but the apremilast S-isomer metabolite M17 significantly inhibited PDE4 (IC₅₀ = 94 nM) and TNF-α (IC₅₀ = 21 nM).

Apremilast (10 µM) had no significant activity in an in vitro screening assay for 68 cell surface receptors or 17 enzymes, except for L-calcium channel receptor.  It did not significantly inhibit any of the 255 kinases in Invitrogen’s Select Screen profile either.^[8,9]

In Vitro Efficacy

Effects on cytokines and anti-inflammatory of apremilast in cell lines:

●    In LPS-stimulated PBMCs:

v    Decreased TNF-α, MIP-1α, IL-12: IC₅₀ = 110-440 nM.

v    Increased IL-10: EC₅₀ = 80 nM.

●    In LPS-stimulated human cutaneous lupus PBMCs:

v    TNF-α: IC₅₀ = 77 nM.

v    IL-12: IC₅₀ = 140 nM.

v    IL-10: EC₅₀ = 2300 nM.

●    In TLR9 agonist CpG-A stimulated human cutaneous lupus PBMCs:

v    Significantly inhibited gene expression of CXCL9, CXCL10, and CXCL11.

●    IL-1β-stimulated human cutaneous lupus PBMC model:

v    TNF-α: IC₅₀ = 83 nM.

●    In anti-CD3 mAb stimulated T-cell line:

v    Decreased TNF-α, IL-5, IL-10, IL-13, IL-17: IC₅₀ = 30-930 nM.

●    In CIA synovial membrane tissue:

v    TNF-α: IC₅₀ = 100 nM.

●    In pDC/HEKa co-culture cells: Inhibited the production of IFN-α and TNF-α protein, and expression of HEKa intracellular MxA protein.

In Vivo Efficacy

In rat models:

●    Plasma TNF-α in LPS-stimulated CD rats: ED₅₀ = 0.018 mg/kg.

●    Air pouch TNF-α in carrageenan-stimulated CD rats: 82% inhibition at 10 mg/kg.

●    Lung neutrophilia in LPS-stimulated CD rat lung models: ED₅₀ = 0.25 mg/kg.

In mouse models:

●    Serum TNF-α in LPS-stimulated Balb/c mice: ED₅₀ = 0.05 mg/kg.

●    Type II CIA: 49% reduction of mean severity score on day 14.

●    Mab/LPS-induced: Reduced paw thickness and hind-paw arthritic reactions in dose-dependent manner at 5 and 25 mg/kg/day.

Absorption of Apremilast

Exhibited a non-linear pharmacokinetics in humans following oral dosing.  The increases in C_max and AUC appeared to be less than dose-proportional in the dose range of 10 to 100 mg apremilast.

Had high oral bioavailability in humans (73.2%), monkeys (78%) and female rats (63%), but moderate in mice (27%) and low in male rats (11.5%). ^[8,9,12]

Was absorbed moderately with the T_max occurring 1-3 h in humans.

Showed a half-life ranging between of 4.5-7.6 h in humans, much longer than that in mice (1.7 h for male and 2.3 h for female). ^[8,9,12]

Had low clearance in humans (169 mL/min) and rabbits (2039 mL/h/kg), in contrast to liver blood flow, after intravenous administration. ^[8,9,12]

Exhibited an extensive tissue distribution in humans but moderate in rabbits, with apparent volumes of distribution at 87 L and 1843 mL/kg, after intravenous administration. ^[8,9,12]

Showed a low permeability, with a Papp_(A→B) of 21 × 10^-6 cm/s in LLC-PK1 cell monolayer model. ^[9]

Distribution of Apremilast

Exhibited moderate plasma protein binding in humans (68.3%), mice (88.6%), rabbits (80.9%) and monkeys (84.3%), but high in rats (90.6%).

Albino mice following a single oral administration: ^[8,9]

●    Absorbed radioactivity was rapidly distributed into the tissues.

●    Relatively higher concentration levels were observed in these tissues generally associated with the principal of biotransformation and excretion (i.e the liver and kidneys), the pancreas, and gall bladder (biliary elimination).

●    Concentrations measured in the central nervous system were consistently low, indicating that penetration of the blood-brain barrier was limited.

●    Males and females had similar patterns of tissue distribution.

●    Radioactivity was not associated with melanin-containing tissues (i.e uveal tract and pigmented skin) in pigmented male mice.

●    By the 72 h sampling time, levels were below the lower limit of quantification of 0.71 μg·eq/g, except for the liver for males only, the kidneys (cortex and medulla), skin, uveal tract, nasal mucosa and gastrointestinal mucosa.

●    Radioactivity was not detected in any tissues at 168 h after dosing or even later.

Metabolism of Apremilast

Could be metabolized in human and animal liver microsomes and hepatocytes with different extents. ^[8,9]

In vitro, oxidative metabolism of apremilast was mediated by CYP3A4 to a large extent, with minor contributions from CYP1A2 and CYP2A6.

Overall, the parent drug represented the most abundant component, with M12 (glucuronide conjugate of O-demethylated apremilast) as the major metabolites in human plasma.  The major component in mice was M1/2.

The major circulating M12 was not pharmacologically active.

Excretion of Apremilast

Was predominantly excreted in urine, with M12 as the major component in human urine.

Was predominantly eliminated in feces in rats and monkeys, and the major component in feces was M3.

About 59% in male and 54% in female of apremilast was recovered via biliary excretion in bile duct-cannulated (BDC) female mice.

Drug-Drug Interaction

Apremilast did not inhibit CYP1A2, 2A6, 2B6, 2C9, 2C19, 2D6, 2E1 or 3A4, but had a weak inhibition for CYP2C8 (IC₅₀ = 56.1 μM). ^[9]

Apremilast had no effects on CYP3A4, 1A2 or 2C9 at 1 μM.  A 3.7-fold induction of CYP3A4 was observed at 100 μM.  Treatment at higher concentrations caused 35% (10 μM) and up to 70% (100 μM) decreases in CYP1A2 and CYP2C9 activities. ^[9]

Apremilast was a substrate of P-gp, but not of BCRP.

Apremilast had no inhibitions for P-gp or BCRP, but had weak inhibitions for MRP1 and MRP2 (IC₅₀ >10 μM).

Apremilast was not a substrate of OATP1B1, OATP1B3, OAT1, OAT3 or OCT2, and had weak inhibitions for them (IC₅₀ >10 μM).

Single-Dose Toxicity

Acute toxicity by the i.v. and p.o. route in two rodent species, and the observed MNLDs/MLDs were concluded:

●    For mice: The oral MNLD was >2000 mg/kg, and the intravenous MLD was ≥120 mg/kg.

●    For rats: The oral MLD was 2000 mg/kg in males and >300 mg/kg in females, and the intravenous MLD was >60 mg/kg.

Repeated-Dose Toxicity

Sub- and chronic toxicity by the oral route in mice (up to 6 months), and monkeys (up to 12 months):

●    In mice:

v    The major apremilast-related finding was arteritis within the thoracic organs, particularly prominent at the junction of the aortic root and the heart, but non-prevalent in the thymus.  Perivascular inflammation of the lung was also observed.

v    Other findings included centrilobular hepatocyte hypertrophy and gastrointestinal effects at high doses such as histopathological changes in the stomach (distension, thickening, irregular surface and raised foci) and associated reduction in food consumption and weight loss.

●    In monkeys:

v    Arteritis was observed in a shorter duration and higher doses studies.  Arteritis was generally found in the myocardium and other locations that included connective tissues of the sciatic nerve and kidney.

v    Other findings at higher doses included emesis, retching or excessive salivation, and thin body condition.

Safety Pharmacology

Both in vitro and in vivo safety pharmacology studies were conducted to assess the effects of apremilast on neurological, cardiovascular, respiratory system, and gastrointestinal effects.

●    Neurological effects: Apremilast produced sympathetic activation responses, and one death at HD the day after neurological testing.

●    Cardiovascular effects: The IC₅₀ of 184 μM for I_Kr would be >100-fold the human C_max at 30 mg BID dose, suggesting rarely potential for QT prolongation, which was confirmed by an in vivo dog study.

●    No findings of initial concern in studies of respiratory function or gastrointestinal transport.

Genotoxicity

Apremilast was considered neither mutagenic nor clastogenic, based on the whole battery of genotoxilogical assays employed.

Reproductive and Developmental Toxicity

Fertility and early embryonic development in mice:

●    Prolonged the estrus cycle and increased the interval to mating in female mice.

Embryo-fetal developmental in mice and monkeys:

●    Dystocia reduced viability, fetal weight and litter size, and increased abortion and embyo-fetal death.

●    Not teratogenic in mice or monkeys.

Pre- and postnatal development in mice:

●    Premature delivery, dystocia, reduced viability and birth weights at doses ≥10 mg/kg.

Repeated-dose toxicity in juvenile mice:

●    Apremilast was considered to be well tolerated in male and female juvenile mice.

Carcinogenicity

Two 2-year oral carcinogenicity studies with apremilast assays were conducted in mice and rats: Not carcinogenic in mice at doses up to 1000 mg/kg/day or in rats at doses up to 20 mg/kg/day in males and 3 mg/kg/day in females.