Enzyme Inducers and Inhibitors

ENZYME INDUCTION
Some of the chemically dissimilar P450 substrate drugs, on
repeated administration, induceP450 expression by
enhancing the rate of its synthesis or reducing its rate of
degradation
Induction results in accelerated substrate metabolism and
usually in a decrease in the pharmacologic action of the
inducer and also of co-administered drugs.
o Mechanism of induction:
synthesis (through enhanced transcription and
translation and increased heme synthesis)
degradation (substrate stabilization)

o Effect of P450 Induction
ACCELERATED SUBSTRATE METABOLISM
pharmacologic action of inducer and co administered
drugs
However enzyme induction may also exacerbate metabolite
mediated toxicity if the drug is metabolically transformed to
reactive metabolites

ENZYME INDUCERS
CYP2B1 Phenobarbital
CYP1A1 Polycyclic Aromatic
Hydrocarbons (PAH)
Benzopyrene
CYP3A4/CYP3A Steroids
(glucocorticoids)/
Phenytoin (anti-
epilepsy)
CYP2E1 INH / Ethanol
CYP4A1 Clofibrate
CYP1A2/CYP2C9/
CYP3A4
Rifampin (anti-tb,
anti-leprosy,
antiviral)
Pollutants - Smoking, charcoal-broiled foods
With enzyme inducers, the action of the drug is
shortened and terminated
**CYP3 (most common)**, CYP2 (2nd), CYP1 (least
common)



INDUCER RECEPTOR INDUCED CYP
Polycyclic
Aromatic
Hydrocarbon
(PAH)
cruciferous
vegetables
Omeprazole
AhR CYP1A
Dexamethasone
Rifampin
Mifepristone
Phenobarbital
Atorvastatin
Hyperforin
(const. of
St. Johns wort)
Pregnane X
receptor
(PXR)
CYP3A
Phenobarbitals Constitutive
Androstane
receptor
(CAR)
CYP2B6
CYP2C9
CYP3A4
Peroxisome
Proliferator
receptor
(PPAR )
CYP4A
(metabolism
of FA -
arachidonic
acid &
derivatives)


ENZYME INHIBITION
Certain drug substrates inhibit cytochrome P450 enzyme
activity
Imidazole-containing drugs such as cimetidine and
ketoconazole bind tightly to the P450 heme iron and
effectively reduce the metabolism of endogenous substrates
(eg, testosterone) or other co-administered drugs through
competitive inhibition.
Some substrates irreversibly inhibit P450s via covalent
interaction of a metabolically generated reactive
intermediate that may react with the P450 apoprotein or
heme moiety or even cause the heme to fragment and
irreversibly modify the apoprotein.
The antibiotic chloramphenicol is metabolized by CYP2B1 to
a species that modifies the P450 protein and thus also
inactivates the enzyme.

o EFFECT OF P450 INHIBITION:
REDUCED SUBSTRATE METABOLISM pharmacologic
action of inhibitor and co administered drugs



.
Binding of inducer to receptor
.
translocation of inducer receptor complex into
nucleus
.
ligand-induced dimerization with nuclear protein
activation of CYP gene regulatory elements
ENZYME INHIBITORS
CYP3A4 Ketoconazole;
Cimetidine;
PRO-ADIPHEN HCl
(GSK-525-A)
Binds tightly to
heme iron
of CYP and
metabolism of
endogenous
substrates
(eg., testosterone)=
Competitive
inhibition
CYP2B1 Chloramphenicol;
Secobarbital
(shortacting
barbiturate)

CYP3A Erthromycin and
derivatives,
Troleandomycin,
Rendered inactive
by macrolide
antibiotics that
complex with the
CYP heme iron

SUICIDE INHIBITORS
Inactivators that attack the heme or the protein moiety
Irreversibly inhibit P450s via covalent interaction of a
metabolically reactive intermediate that may react with
P450 apoprotein or heme moiety or even cause the
heme to fragment and irreversibly modify the apoprotein.
includes :
-certain steroids (ethinyl estradiol, norethindrone, and
spironolactone);
-fluroxene; allobarbital;
-the analgesic sedatives allylisopropylacetyl- urea,
diethylpentenamide, and ethchlorvynol;
-carbon disulfide;
-grapefruit furanocoumarins;
-selegiline;
-phencyclidine;
-ticlopidine and clopidogrel;
-ritonavir; and
- propylthiouracil.
* With enzyme inhibitors, action of the drug is prolonged

Excretion
1. Characteristics of Drugs to be excreted:

1. Water soluble
2. Ionized
3. Polar
4. Change in pH

Acid- gives sodium bicarbonate
Base- gives ammonium chloride

2. Ionization constant
electrostatic charge of an ionized molecule attracts
water dipoles and results in a polar, relatively
water-soluble and lipid-insoluble complex
Lipid diffusion depends on relatively high lipid
solubility, thus, ionization of drugs may markedly
reduce their ability to permeate membranes.
A very large percentage of the drugs in use are
weak acids or weak bases
Weak acid
- defined as a neutral molecule that can reversibly
dissociate into an anion (a negatively charged
molecule) and a proton
(a hydrogen ion)
- For example, aspirin dissociates as follows:


Weak base
- Defined as a neutral molecule that can form a cation (a
positively charged molecule) by combining with a proton.
- For example, pyrimethamine, an antimalarial drug,
undergoes the following association-dissociation process:


Note: All drugs are either weak bases or weak acids. So in
case of
overdose, one usually gives a weak base/acid to counteract
the drugs effects
WEAK ACIDS WEAK BASES
Acetaminophen
Ampicillin
Aspirin
C-Thiazide
Ciprofloxacin
Ibuprofen
L-Dopa
Methotrexate
(MTX)
Phenobarbital
Phenytoin
Thephylline
Albuterol
Allopurinol
Methamphetamine
Atropine
Chloroquine
Clonidine
Cocaine
Diazepam
Diphenydramine
Epinephrin
Lidocaine
Note: the protonated form of a weak acid is the neutral, more lipid-
soluble form, whereas the unprotonated form of a weak base is the
neutral form.
Henderson-Hasselbalch equation
relates the ratio of protonated to unprotonated weak acid
or weak base to the molecules pKa and the pH of the
medium
the lower the pH relative to the pKa, the greater will be the
fraction of drug in the protonated form
more of a weak acid will be in the lipid-soluble form at acid
pH, whereas more of a basic drug will be in the lipid-
soluble form at alkaline pH.

3. Organs of excretion
MAJOR ORGANS
1. Renal Excretion

Application of the principle of ionization is made in the
manipulation of drug excretion by the kidney. If a drug is in a lipid-
soluble form during its passage down the renal tubule, a
significant fraction will be reabsorbed by simple passive diffusion.
If the goal is to accelerate excretion of the drug (eg, in a case of
drug overdose), it is important to prevent its reabsorption from
the tubule. This can often be accomplished by adjusting urine pH
to make certain that most of the drug is in the ionized state. As a
result of this partitioning effect, the drug is trapped in the urine.
Acidification of urine increases reabsorption and
decreases excretion of weak acids and decreases
reabsorption of weak bases. Alkalinization of urine has
the opposite effect. Thus, weak acids are usually excreted
faster in alkaline urine; weak bases are usually excreted
faster in acidic urine
Sodium bicarbonate: alkalinizes the urine
Ammonium chloride: acidifies the urine.
Accounts for most drug excretion. About 15 of the plasma
reaching the glomerulus is filtered through pores in the glomerular
endothelium
nearly all water and most electrolytes are passively and actively
reabsorbed from the renal tubules back into the circulation
polar compounds, which include most drug metabolites, cannot
diffuse back into the circulation and are excreted unless a specific
transport mechanism exists for their reabsorption (eg, as for
glucose, ascorbic acid, and B vitamins)
With aging, renal drug excretion decreases
Drugs bound to plasma proteins remain in the circulation; only
unbound drug is contained in the glomerular filtrate. Un-ionized
forms of drugs and their metabolites tend to be reabsorbed readily
from tubular fluids.
Urine pH, which varies from 4.5 to 8.0, may markedly affect
drug reabsorption and excretion by determining whether a
weak acid or base is in an un-ionized or ionized form
the anion secretory system eliminates metabolites conjugated with
glycine, sulfate, or glucuronic acid. Anions compete with each other
for secretion. This competition can be used therapeutically;
eg, probenecid: blocks the normally rapid tubular secretion of
penicillin, resulting in higher plasma penicillin concentrations
for a longer time
In the cation transport system, cations or organic bases (eg,
pramipexole, dofetilide) are secreted by the renal tubules; this
process can be inhibited by cimetidine, trimethoprim,
prochlorperazin, megestrol , or ketoconazole

2. Biliary excretion

transported across the biliary epithelium against a concentration
gradient
active secretory transport is required
Drugs with a MW (molecular weight) > 300 g/mole and with both
polar and lipophilic groups are more likely to be excreted in bile;
smaller molecules are generally excreted only in negligible
amounts
Conjugation, particularly with glucuronic acid, facilitates biliary
excretion
MINOR ORGANS
1. Salivary glands
2. Sweat glands
3. Tear glands
4. Respiratory- exhalation
5. Breasts- suckling of milk
4. Clearance
The measure of the ability of the body to eliminate the
drug
factor that predicts the rate of elimination in relation to
the drug concentration:

Clearance is additive Rate of elimination at each organ of
elimination can be added to get the total systemic clearance



The two major sites of drug elimination are the kidneys and the
liver. Clearance of unchanged drug in the urine represents renal
clearance.
For most drugs, clearance is constant over the concentration
range ie. Elimination is not saturable and the rate of drug
elimination is directly proportional to concentration :


This is usually referred to as first-order elimination. When
clearance is first-order, it can be estimated by calculating
the area under the curve (AUC)of the time-concentration
profile after a dose. Clearance is calculated from the dose
divided by the AUC.

Two types of clearance:
I. Capacity-Limited
For drugs that exhibit capacity-limited elimination (eg,
phenytoin, ethanol), clearance varies depending on the
concentration of the drug that is achieved
Also known as mixed-order, saturable, dose- or
concentration-dependent, nonlinear, and Michaelis-
Menten elimination
Clearance has no real meaning for drugs with capacity
limited elimination - AUC cannot be used to describe the
elimination of such drugs

II. Flow-Dependent Elimination

In contrast to capacity-limited drug elimination, some
drugs are cleared very readily by the organ of elimination,
so that at any clinically realistic concentration of the drug,
most of the drug in the blood perfusing the organ is
eliminated on the first pass of the drug through it.
drugs under this category are called high extraction drugs
since they are completely extracted from the blood by the
organ
elimination is dependent on the rate of drug delivery to
the organ of elimination
thus, blood flow to the organ is the main determinant of
drug delivery
Clearance in Terms of Drug Concentration Measurement
Clearance is the single most important factor determining
drug concentrations
The interpretation of measurements of drug
concentrations depends on three factors that may
influence clearance:
a. Dose
b. organ blood flow
c. intrinsic function of the liver or kidneys
Development and Regulation of Drugs
a. Drug Discovery
6 approaches:
1. Identification of new drug target
2. Rational drug design of a new drug based on
understanding of biologic mechanisms, drug receptor
structure and drug structure.
3. Chemical modification of known molecule
4. Screening of biologic activity of large numbers of natural
products, previously discovered chemical entities, peptides,
nucleic acids and other organic molecules
5. Biotechnology and cloning using genes to produce peptides
and proteins.
6. Combination of known drugs to obtain additive or synergistic
effects or repositioning of a known drug to a new category.

b. Pre-clinical Safety-Toxicity Testing (In-vitro or
Animal Studies)
All drugs are toxic in some individuals at some dose.

These are done to identify potential human toxicities and to
design tests to further define the toxic mechanisms, and
predicting the most relevant toxicities to be monitored in
clinical trials.
the art of drug development consists of effective assessment and
management of risk versus benefit and not total risk avoidance.

Safety tests:
Acute toxicity
Usually two species, two routes
Goal is to determine the no-effect dose and the maximum
tolerated dose. In some cases, determine the acute dose that
is lethal in approximately 50% of animals.

Subacute or subchronic toxicity
Three doses, two species
2 weeks to 3 months of testing may be necessary before
clinical trial
the longer the duration of expected clinical use, the
longer the subacute test
goal is to determine biochemical, physiologic effects.
Chronic toxicity
Rodent and at least one nonrodent species for 6 months
required when drug is intended to be used in humans for
prolonged periods
usually run concurrently with clinical trials
goal is to determine same end points as subacute toxicity
tests.
Effect on Reproductive performance
Two species, usually one rodent and rabbits.
Test effects on animal mating behaviour, reproduction,
parturition, progeny, birth defects, postnatal
development.
Carcinogenic potential
Two years, two species
Required when drug is intended to be used in humans for
prolonged periods.
goal is to determine gross and histologic pathology
Mutagenic potential
test effects on genetic stability and mutations in
bacteria (Ames test) or mammalian cells in culture;
dominant lethal test and clastogenicity in mice
It is important to recognize the limitations of preclinical testing.
These include the following:
1. Toxicity testing is time-consuming and expensive.
2. Large numbers of animals may be needed to obtain valid
preclinical data.
3. Extrapolations of therapeutic index and toxicity data from animals
to humans are reasonably predictive for many but not for all
toxicities.
4. For statistical reasons, rare adverse effects are unlikely to be
detected in preclinical testing.

c. Clinical Drug Trial
Less than 1/3 of drugs tested in clincal trials reach
the market place
The need for careful desing and execution is based on
3 major confounding factors inherent in the study of any
drug in humans:
1. The variable Natural History of most diseases
2. The presence of other diseases and risk factor
3. Subject and Observer Bias and other risk factor

Phase I
20-100 healthy human volunteers
Done in research centers
Non-blind or open technique
Determine: Safety & pharmacokinetics

Phase II
100-200 volunteer patients with the target disease
Done in special clinical centers e.g. university hospitals
Single-blind with placebo technique
Determine: Efficacy, doses to be used on follow on trials &
toxicity

Phase III
1000-6000 (usually thousands, did not mention any
specific number-katzung 12
th
ed) volunteer patients with
target disease
Done in settings similar to those anticipated for the
ultimate use of drug e.g. clinical hospitals
Double-blind and cross-over techniques
Expensive
Investigators are usually specialists in the disease
Determine: safety-efficacy
Toxic effects caused by immunologic process will be
apparent in this phase, if drug passed phase 3
submission of New Drug Application (NDA) to FDA.

Phase IV
Monitoring the safety of the new drug under actual
conditions of use in large number of patients.
post-marketing surveillance
filing of a patent application to approval for
marketing of a new drug may be 5 years or
considerably longer
lifetime of a patent is 20 years in the USA, the owner
of the patent (usually a pharmaceutical company) has
exclusive rights for marketing the product for only a
limited time after approval of the new drug
application
Orphan drug - drug for rare diseases
d. Government Agencies
Government Agencies
Food and Drug Administration (FDA)
Administrative body that oversees the drug evaluation
process in the USA (same with the Philippines- which is
previously called BFAD) and grants approval for
marketing of new drug products.
In addition to the approval of the sponsoring
organization and the FDA, an interdisciplinary
institutional review board (IRB) at the facility where
the clinical drug trial will be conducted must review
and approve the scientific and ethical plans for testing in
humans
For procurement, advertisement, marketing and
information (PDIA/ Phil. Drug Information Agency)
Also for control and regulation of drugs (PDEA/ Phil. Drug
Enforcement Agency)

e. Discovery of Anti-angiogenesis
1961
Dr. J. Folkman had a hunch that cancerous
tumors, in order to expand, needed to trigger the
growth of new blood vessels to feed themselves
1971
Biologic hypothesis - tumors are angiogenic
dependent. If a tumor could be stopped from
growing its own blood supply, it would wither and
die
1984
chemical hypothesis (angiogenic factors for
formation of capillary blood vessels necessary for
perfusion, healing/repair)
1997
Discovery of angiostatin/endostatin (a globular
protein that serves as anti angiogenic agent)
1989
Dr. N. Ferrera discovered vascular endothelial
growth factor (VEGF) capable of forming ne blood
vessels.
1993
block VEGF
Genentech team (monoclonal antibody mab)
2004
Bevacizumab (1st anti-angiogenesis to prevent
metastasis)
Approved by FDA
R. Damato thalidomide, a drug used for leprosy
(2nd anti-angiogenesis)