Pharmacokinetics And Drug Drug Interactions Biology Essay
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Drug development is a complex multidisciplinary process during which a preclinical drug candidate is evaluated to assess its potential efficacy, safety, pharmacokinetic and metabolism. The data produced in vitro and in vivo should be predictive of what will be observed in humans during clinical trials. A preclinical drug metabolism and pharmacokinetic group (DMPK) should investigate several properties of a drug candidate that are related to its ability to be absorbed through the gastrointestinal tract, to bind to plasma proteins, to partition between water and hydrophobic compartments, to be actively expelled by cells, to be metabolized, to enter in specific compartments. Initial measurements of solubility and stability in water at different PH and in simulated gastric and intestinal fluids, provides useful data to understand if the drug could be administered orally. The biopharmaceutical classification system (BCS) categorizes drugs into four classes using solubility and permeability (usually measured using Caco2 cells, a continuous line of heterogeneous human epithelial colorectal adenocarcinoma cells). Class I contains molecules with high permeability and solubility, class II contains molecules with high permeability and low solubility, class III is characterized by low permeability and high solubility, class IV by low permeability and solubility. The difficulty in achieving oral absorption is increasing with the classes and only in the latest years it has been possible to obtain oral formulations with good bioavailability for compounds belonging to class IV. Lipophylicity is determined measuring LogP and is an important parameter to determine drug distribution among lipophilic and hydrophilic compartments. For ionized compounds it is preferable to use LogD which take into consideration the partitioning of all species, with and without charges.
Understanding whether a compound is likely to enter the brain and central nervous system (CNS) via the BBB may also be important for any drug, not only psychoactive medicines. If the compound in question is a potential CNS active drug, crossing the BBB is a prerequisite. Conversely, BBB penetration might cause unwanted off - target side effects for those drugs not intended for the CNS. The BBB penetration can be estimated using cell monolayers or artificial membranes (PAMPA), while the actual measurement of blood /brain ratio will be determined in a suitable pharmacokinetic experiment.
Both influx transporters, which facilitate the entry of drugs into cells, and efflux transporters, which limit the entry of drugs or enhance the removal of drugs from the cells, are crucial in determining concentrations of drug in the body and within various cells and tissues. There are many transporters that have been identified, but P-glycoprotein (Pgp) is the one that has been studied in more detail and the majority of the clinical interactions reported are mediated by this glycoprotein. A widely used in vitro method to investigate Pgp uses Caco2 cells and the MDCKII cell line. These cells can be grown as monolayers which are polarized allowing investigating the bidirectional transport of the drug. Preclinical studies should investigate the inhibition the Pgp as the effects on co-administered drugs could be quite dramatic and result in toxic effects.
Another important parameter is serum protein binding that can affect the elimination of the compound from blood and also its activity, as this latter is determined by the unbound fraction. The protein binding is different in each animal species and when is greater than 95% it could significantly affect the plasma half-life as the bound fraction is not excreted through glomerular filtration. Protein binding should be determined in man and also all animal species that are used in pharmacology and toxicology. A compound with a high plasma protein binding could also affect the protein binding of other drugs that are co-administered, with a potential effect on their pharmacokinetic profile and therefore with the risk of modifying the efficacy or the toxicity.
The pharmacokinetic studies have two main scopes: to determine the drug concentration at different doses that are related to the pharmacological effect, and to determine the drug concentration achieved during toxicological studies. In this latter case it is generally defined as toxicokinetics. The studies should be conducted using escalating doses and, where appropriate different regimens. The formulation can dramatically change the adsorption and in some cases also the metabolism of the drug under investigation, therefore this phenomenon should be carefully evaluated when performing pharmacokinetic, pharmacological and toxicological studies. Ideally, it would be preferable to use the same formulation that should be used in humans, however this is rarely possible as the dosage form for clinical trials is generally developed in parallel to pharmacokinetic studies or even later. For orally administered drugs, both intravenous and oral pharmacokinetic parameters should be determined and relative bioavailability should be estimated (area under the curve per os / area under the curve i.v.). The pharmacokinetic study could investigate drug concentration in several biological fluids and tissues, like cerebro spinal fluid (CSF), urines, kidneys, lungs, liver. In addition, if the drug is metabolized into active compounds, the study may also include these substances as they could significantly affect the overall biological activity of the drug candidate. The pharmacokinetic studies are usually performed, according to ICH guidelines and regulations in at least two animal species: one rodent and one non rodent and preferably in the species in which the efficacy studies have been conducted, unless there are valid justifications not to do so. The main parameters that are determined in pharmacokinetic studies include plasma or whole blood half-life, Cmax (maximal concentration reached by the administered drug), tmax (time at which Cmax is achieved), distribution volume, area under the curve.
Another important aspect that must be evaluated is the metabolism of the drug candidate. This is evaluated using several systems that range from enzyme preparations like CYP450s, to cellular fractions like S9 liver microsomes in which only phase I metabolism occurs, to hepatocytes, which express the full metabolic capabilities.
Subcellular fractions such as liver microsomes S9 are in vitro models of hepatic clearance (also called intrinsic clearance), that are often used in drug discovery and early stages of preclinical development.
Hepatocytes have advantages over subcellular fractions in that they are whole cell systems that are energetically competent with active transporters. Recent improvements in cryopreservation techniques have solved several practical issues associated with isolation of the hepatocytes. Recently, the use of cryopreserved hepatocytes for the determination of intrinsic clearance has increased. Hepatocytes tend to be the system of choice for applications such as metabolite profiling and induction studies. Ideally these studies should be conducted using human and animal derived hepatocytes to facilitate the interpretation of differences in pharmacokinetic and pharmacodynamic parameters, as well as support allometric scaling calculations. These determinations become of particular importance if the compound under investigation is converted in active or potentially toxic metabolites.
Moreover, I would like to highlight the attention to the fact that although liver is the main site of metabolism, there are several other tissues that are capable of metabolizing drugs like kidneys and lungs. In particular for drugs that are administered as inhalatory formulation, the lungs are the first organ that the drug encounters with significant capability to metabolize it.
CYPs (cytochrome P450) are a class of enzymes which all have similar core structures and mechanism of action. They are involved in more than 75% of all drug biotransformations. It is believed that 9 out of 10 drugs in use today are metabolized by only five of these isoforms; CYPs 1A2, 2C9, 2C19, 2D6 and 3A4/5.
Drugs that are metabolized by the CYPs can also alter the metabolism of other drugs and act as inducer or inhibitor of their metabolism. This aspect is particularly important as it may alter the circulating concentration of the drugs increasing it to a toxic level or reducing it to a level that is no more effective. This type of studies is generally addressed during phase II clinical development as they may be quite long and expensive.
There are several broad groups of drugs capable of inducing hepatic metabolism: these include: anticonvulsants, steroids, antibiotics, recreational agents; nicotine, alcohol, herbal remedies, protease inhibitors.
A new drug is generally regarded as an inducer if it produces a change in drug clearance which is equal to or greater than 40 per cent of an established potent inducer, usually taken as rifampicin.
Drug candidates should also be tested for their CYP450 inhibitory activity as this phenomenon could reduce the metabolism of other co-administered drugs and result in toxic effects.
To summarize, there are various attributes of an investigational new drug that need to be determined, or at least estimated, prior to submitting an application for starting clinical trials in man. In particular with respect to metabolism the following aspects should be evaluated:
• Metabolic rate: drugs which are very rapidly, or very slowly, cleared can present problems in accurate control of plasma levels and with very long half - life agents, the risk of toxicity can be considerable.
• Multiple CYP metabolism: ideally a drug candidate should be metabolized by several cytochromes in order to allow its clearance even in the presence of co-administered drugs which inhibit a specific enzyme.
• Likelihood of DDIs, inhibition: if a drug is likely to inhibit a major CYP isoform, or its metabolism is blocked by a known series of inhibitors, then the likelihood of DDIs or drug - drug interactions will be high.
• Likelihood of DDIs, induction: a powerful inducer will be problematic in complex regimens and will accelerate the clearance of other drugs.
• Polymorphisms: a drug that is subject to a single polymorphic CYP clearance may well be restricted in its usage or would require close medical supervision, as the risks of toxicity and drug failure would be considerable.
• Linearity of metabolism: most drugs are cleared at a rate proportional to their intake (linear metabolism). If the metabolism of the drug is non - linear, then it will be difficult to predict plasma levels with ascending dose, which will also make the drug difficult to use and subject to toxicity and potential failure.
A novel candidate drug, FBXL5 is under development as a H2 receptor antagonist. You are presented with the following information and need to make a decision about possible in vivo drug inhibition caused by FBXL5.
In table 1 FBXL5 was tested at 1.5mM in human microsomes in the presence of various probes CYP substrates (phenacetin de-ethylation for CYP1A2, tolbutamide hydroxylation for CYP2C9, debrisoquine hydroxylation for CYP2D6, chlorzoxazone hydroxylation for CYP2E1, Nifedipine aromatisation for CYP3A4). The data show a marked inhibition of debrisoquine hydroxylation (about 20% of residual activity) and a less pronounced inhibition of chlorzoxazone hydroxylation and nifedipide aromatisation (about 60% of residual activity). These data suggest that FBXL5 is an ihibitor of CYP2D6 and to a lesser extent of CYP2E1 and CYP3A4.
In table 2 the data show that FBXL5 has an IC50 of 210M and 480M respectively in inhibiting CYP2D6 and CYP3A4 when dextromethorphan or dextrorphan are used as substrate.
Table 3 reports the predicted therapeutic dose that is 1-4M, a concentration that is about 50 times lower than the IC50 for CYP2D6, is about 100 times lower than IC50 for CYP3A4 and is much lower in all other cases.
The overall data show a potential for drug-drug interaction (DDI), particularly for those drugs that are metabolized by CYP2D6 and to a lesser extent for CYP3A. The route of administration and the formulation adopted will be an important additional parameter to take into consideration as the Cmax could reach values that are closer or even higher than the IC50 for CYP2D6.
In particular in the case of an intravenous bolus administration the risk of inhibiting the CYP2D6 and pheraps the CYP3A4 is quite high, while in the case of an oral administration using an extended release formulation would minimize this risk.
More in depth studies to determine the mechanism of inhibition, its reversibility and its effect on different CYP2D6 and CYP3A4 polymorphs should be conducted to define the relative risk and potential limitations in the use of the investigational drug FBXL5.
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