The effect of tricyclic antidepressants, selective serotonin reuptake inhibitors (SSRIs) and newer antidepressant drugs on the activity and level of rat CYP3A

Abstract

The aim of the present study was to investigate the influence of tricyclic antidepressants (TADs: imipramine, amitriptyline, clomipramine, and desipramine), selective serotonin reuptake inhibitors (SSRIs: fluoxetine and sertraline) and novel antidepressant drugs (mirtazapine and nefazodone) on the activity of CYP3A measured as a rate of testosterone 2β- and 6β-hydroxylation. The reaction was studied in control liver microsomes in the presence of the antidepressants, as well as in microsomes of rats treated intraperitoneally (i.p.) for 1 day or 2 weeks with pharmacological doses of the drugs (imipramine, amitriptyline, clomipramine, nefazodone 10 mg kg^− 1 i.p.; desipramine, fluoxetine, sertraline 5 mg kg^− 1 i.p.; mirtazapine 3 mg kg^− 1 i.p.), in the absence of the antidepressants in vitro. The investigated antidepressants added to control liver microsomes produced some inhibitory effects on CYP3A activity, which were very weak (most of TADs, K_i = 145–212 μM), modest (clomipramine and sertraline, K_i = 67.5 and 62 μM, respectively) or moderate (nefazodone and fluoxetine, K_i = 42 and 43 μM, respectively). Mirtazapine did not display this kind of properties. One-day exposure of rats to TADs substantially decreased the activity of CYP3A in liver microsomes, which was maintained during chronic treatment. The observed decreases in the enzyme activity were in contrast to the increased CYP3A protein level found after chronic treatment with TADs. On the other hand, sertraline increased the activity of the enzyme after its prolonged administration and its effect correlated positively with the observed elevation in CYP3A protein level. Fluoxetine, mirtazapine and nefazodone did not change the activity of CYP3A in liver microsomes after their administration to rats. Three different mechanisms of the antidepressants–CYP3A interaction are postulated: 1) a direct inhibition of CYP3A by nefazodone, SSRIs and clomipramine, shown in vitro, with the inhibitory effect of nefazodone being the strongest, but weaker than the effects of this drug on human CYP3A4; 2) in vivo inhibition of CYP3A produced by 1 day and maintained during chronic treatment with TADs, which suggests inactivation of the enzyme by reactive metabolites; 3) in vivo induction by sertraline of CYP3A produced only by chronic treatment with the antidepressant, which suggests its influence on the enzyme regulation.

Introduction

Among CYP3A isoforms, human CYP3A4 and rat CYP3A2 (male-dominant) belong to the main constitutive enzymes expressed in the liver. These isoforms are predominantly expressed in the liver (Komori and Oda, 1994), but they have also been found in the intestine, kidneys, lungs, brain and leukocytes (Watkins, 1992, Debri et al., 1995, Wang et al., 1996). The CYP3A subfamily is characterized by its ability to utilize a wide spectrum of substrates being both small molecules, such as paracetamol and large ones like cyclosporin. The CYP3A substrates are endogenous substances (e.g. steroids) or xenobiotics (e.g. drugs such as psychotropics, carbamazepine and calcium antagonists). 6β-Hydroxylation of cortisol in humans and 2β- and 6β-hydroxylation of testosterone in rats are used as marker reactions for studying CYP3A activities (Gibson et al., 2002).

The CYP3A expression is under physiological and environmental influence. Hormonal status is of great importance with a specific secretion of growth hormone and some steroids (corticosterone, hydrocortisone, testosterone) up-regulating it on the one hand and triiodothyronine and interleukins IL-1, IL-2 and IL-6 down-regulating it on the other (Waxman et al., 1995, Anderson et al., 1998, Liddle et al., 1998, Pascussi et al., 2000c, Sunman et al., 2004). It has been shown that glucocorticoid receptor (GR), pregnane X receptor (PXR), constitutive androstane receptor (CAR) and vitamin D receptor (VDR) play an important role in the regulation of CYP3A genes (Honkakoski and Negishi, 2000, Honkakoski et al., 2001, Quatrochi and Guzelian, 2001, Gibson et al., 2002, Burk and Wojnowski, 2004). Both PXR and CAR act as dimers with retinoid acid X receptor (RXR). GR may contribute to CYP3A induction via direct or indirect molecular mechanism, including functional cross talk between GR-, PXR- and CAR-signaling pathways (Zhang et al., 1999, Pascussi et al., 2000a, Pascussi et al., 2000b, Pascussi et al., 2003).

Administration of antidepressants to patients for months or years creates conditions for their interference with different kinds of proteins, such as receptors and enzymes. The literature data show that antidepressants are substrates of cytochrome P-450, mainly of CYP2D6 and CYP3A4. Some of them are also inhibitors of those enzymes, e.g. fluoxetine and paroxetine inhibit CYP2D6, fluvoxamine inhibits CYP1A2 (Crewe et al., 1992, Brøsen et al., 1993, Jeppesen et al., 1996), while nefazodone blocks CYP3A4 (von Moltke et al., 1999) in humans. Moreover, fluoxetine and fluvoxamine moderately inhibit CYP3A4 (Preskorn, 1997, Sproule et al., 1997).

However, there is only a few data on the effect of those drugs administered in clinical conditions (or simulating clinical conditions) on the activity of cytochrome P-450. It has been shown that chronic treatment with pharmacological doses of tricyclic antidepressants (TADs), selective serotonin reuptake inhibitors (SSRIs) and nefazodone decreased, while mirtazapine increased the activity of CYP2D in the rat liver (Daniel and Netter, 1992, Daniel et al., 2002). Their effect on other CYP isoforms is less known. Imipramine given at high daily doses (100 mg kg^− 1 p.o. once daily) for 5 days to rats increased the rates of CYP3A specific reactions, i.e. 2β- and 6β-hydroxylation of testosterone (Masubuchi et al., 1995), but the effect of chronic administration of pharmacological doses of imipramine or other antidepressants on that enzyme has not been investigated so far. Warrington (1992) suggested the induction of CYP by sertraline, expressed as an increase in antipyrine clearance by the antidepressant, the former drug being metabolized by many CYP isoforms (CYP3A4, CYP1A2, and CYP2C9).

On the other hand, the possibility of forming CYP-reactive metabolite complexes had been shown at high concentrations (100–200 μM) of some TADs and fluoxetine in vitro during their prolonged incubation in rat liver microsomes, indicating the involvement of CYP3A, CYP2C11 and CYP2A (Murray and Field, 1992, Bensoussan et al., 1995, McNeil and Murray, 1996, Murray and Murray, 2003). But it is not known whether the above mechanism can produce an inhibitory effect on CYP3A in vivo when animals are treated with pharmacological doses of antidepressants producing lower drug concentrations.

The aim of our study was to investigate the influence of 1-day and 2-week treatment with pharmacological doses of antidepressants (TADs, SSRIs, and the novel antidepressants nefazodone and mirtazapine) on the activity and protein level of CYP3A in liver microsomes (ex vivo study), which has not been done yet in either humans or rats. We also examined in our study the direct interaction of antidepressants with rat CYP3A in vitro (binding with cytochrome protein, which had been studied in humans, but not in rats) to show any possible effects of antidepressants on rat CYP3A, as well as differences in their direct effect on human and rat CYP3A. The obtained results indicate drug-dependent changes in the activity of rat CYP3A produced by antidepressants via different mechanisms.