Drugs Affecting the Central Nervous System

1. Depressant Drugs

1.1. General Anesthetics

General anesthesia is a depression of the central nervous system carried out under controlled and reversible conditions so that loss of sensation and consciousness results.

Drugs given to induce or maintain general anesthesia are either given as:

- Gases or vapors (inhalational anesthetics)

- Injections (intravenous anesthetics)

Most commonly these two forms are combined, with an injection given to induce anesthesia and a gas used to maintain it, although it is possible to deliver anesthesia solely by inhalation or injection.

1.1.a. Inhalation

Inhalational anesthetic substances are either volatile liquids or gases and are usually delivered using an anesthesia machine. An anesthesia machine allows composing a mixture of oxygen, anesthetics and ambient air, delivering it to the patient and monitoring patient and machine parameters. Liquid anesthetics are vaporized in the machine.

Many compounds have been used for inhalation anesthesia, but only a few are still in widespread use. Desflurane, isoflurane and sevoflurane are the most widely used volatile anesthetics today. They are often combined with nitrous oxide.

Dr. Amged 1 Older, less popular, volatile anesthetics, include halothane, enflurane, and methoxyflurane.

1.1.b Injection

Injection anesthetics are used for induction and maintenance of a state of unconsciousness. Anesthetists prefer to use intravenous injections as they are faster, generally less painful and more reliable than intramuscular or subcutaneous injections. Among the most widely used drugs are:

1.2. Sedative & Hypnotics

A Sedative drug is a CNS depressant that decreases excitability but does not induce sleep. On the other hand, a hypnotic drug is a CNS depressant that produces sleep; used to induce sleep when natural sleep is impossible.

Sedative-hypnotics often are referred to as sleeping pills and are used to treat insomnia. This class of drugs causes drowsiness and facilitates the initiation maintenance of sleep. The observed pharmacological effects of most drugs in this class usually are dose related. Small doses cause sedation, larger doses cause hypnosis (sleep), and still larger doses may bring about surgical anesthesia.

Insomnia can be classified as primary (pathogenesis unknown) or secondary (from other causes). The latter is more common and can be the result of situational stress,

Dr. Amged 2 lifestyle habits, drugs, and psychiatric or medical disorders. The drugs currently used as hypnotics are effective, but there is ample need for newer and safer hypnotics. The ideal sedative-hypnotic should:

1) Cause transient decrease in the level of consciousness for the purpose of sleep without lingering effects (sleep induction and sleep maintenance).

2) Have no potential for decreasing or arresting respirations (even at relatively high doses).

3) Produce no abuse, addiction, tolerance or dependence.

1.2.1 Physiology of Sleep

1.2.1.a Sleep Cycle

Sleep is studied using related techniques that permit electronic monitoring of the head and neck muscle and eye movements. Form these and related studies, three states have been defined:

(1) Wakefulness.

(2) Nonrapid eye movement [NREM] sleep.

(3) Rapid eye movement [REM] sleep.

1.2.1.b Sleep Factors

The involvement of many autonomic, physiologic, and biochemical changes are associated with these wakefulness, NREM sleep, and REM sleep. The relationship of cause and effect in relation to these systems is still somewhat controversial and a rapidly changing area of research. Several brain regions that regulate sleep have now been identified; however, the specific contribution on any one region to sleep is still controversial. The roles of the major systems are important to be familiar with in relation to sleep. This not only helps one to understand the mechanism by which hypnotics work but also provides some understanding of why unrelated drugs, such as neuroleptics,

Dr. Amged 3 antihistamines, antidepressants, and antimanic drugs, occasionally are used as hypnotics to facilitate sleep.

Every neurotransmitter has, at one time or another, been implicated in sleep or wake fullness, for examples:

(1) Catecholamines.

(2) Serotonin.

(3) Histamine

(4) Acetylcholine.

(5) Adenosine.

(6) γ-Aminobutyric acid: it probably represents the most important inhibitory transmitter of the mammalian CNS. Both types of GABAergic inhibition (pre-

and postsynaptic) use the GABAA receptor subtype, which acts by regulation of the chloride channel of the neuronal membrane. A second GABAergic type,

GABAB, that is a G protein-coupled receptor is not considered to be important in

understanding the mechanism of hypnotics. Activation of a GABAA receptor by an agonist increases the inhibitory synaptic response of central neurons to GABA through hyperpolarization. Because many, if not all, central neurons receive some GABAergic input, this leads to a mechanism by which CNS activity can be depressed. For example, if the GABAergic interneurons are activated by an agonist that inhibits the monoaminergic structures of the brain stem, hypnotic activity will be observed.

Some hormones have been found to have a significant effect on sleep and circadian rhythmcity, for examples:

(1) Growth hormone.

(2) Prolactin.

(3) Melatonin

Dr. Amged 4 1.2.3 Classification of Hypnotics:

The hypnotic drugs are not characterized by common structural features. Instead, a wide variety of chemical compounds have been used in clinical therapy.

1) Chloral.

2) Barbiturates.

3) Benzodiazepines.

4) Nonbenzodiazepines

5) Melatonin receptor agonists.

6) Antihistamines.

7) Antidepressants.

1.2.3.a Chloral Hydrate

It was introduced as a sedative in 1869. During the 1950s and 1960s, chloral hydrate was widely promoted as a hypnotic. Today, it still finds use as a sedative in nonoperating room procedures for pediatric patients.

Chloral is a unique aldehyde because of the electron-withdrawing effect of CCl3 group. When chloral (an oily liquid) is treated with water or alcohol, a crystalline solid, chloral hydrate, is formed. Chloral hydrate is stable, but as indicated below, when it is dissolved in water, it is in equilibrium with the chloral form (equilibrium strongly favoring the chloral hydrate structure).

The CCl3 group is sufficiently electron-withdrawing that chloral hydrate is a weak acid (pKa = 10.04). This acidity makes it quite irritating to mucous membranes, such as in the stomach.

Dr. Amged 5 Chloral hydrate is readily absorbed from the gastrointestinal tract following oral or rectal doses and is quickly reduced to trichloroethanol, its active metabolite, by alcohol dehydrogenase in the liver and erythrocytes. Trichloroethanol is metabolized by alcohol dehydrogenase oxidation to chloral and then to the inactive metabolite trichloroacetic acid via ldehyde dehydrogenase. Trichloroacetic acid is excreted in the urine as a glucuronic acid conjugate.

Cl O alcohol Cl OH dehydrogenase Cl C Cl C H Cl H Cl H Chloral Trichloroethanol active metabolite aldehyde dehydrogenase

Cl O Cl C Conjugation Cl OH Trichloroacetic acid inactive metabolite

1.2.3.b Barbiturates

Older medications, such as the barbiturates, are used as sedative-hypnotics, but toxicity limits their widespread use. For example, they can cause significant central nervous system (CNS) depression, physical dependence, and tolerance. Additionally, they are potent inducers of liver enzymes, which can lead to clinically significant drug interactions when these medications are administered with other drugs extensively metabolized by the liver.

Barbiturates are cyclic ureides and are formed when a dicarboxylic acid reacts with urea. The acids used are generally in the form of ester and are condensed in the presence of sodium ethoxide (C2H5ONa).

Dr. Amged 6 Parabanic acid is a cyclic ureide containing five membered ring, which on hydrolysis by alkali may regenerate the corresponding acid and urea. The cyclic ureides are acidic owing to enolization and hence, they may form metallic salts by replacing the H atom of the –OH group as shown below:

Or

Many cyclic ureides are derived from malonic acid or malonic esters. They are collectively known as barbiturates because of their relationship of barbituric acid (malonyl urea).

Barbituric acid is prepared by the following two methods:

(a) By the interaction of urea and malonyl dichloride

Dr. Amged 7

(b) By the interaction of urea and diethyl malonate

Barbituric acid like parabanic acid exhibits keto-enol tautomerism as illustrated below:

In thiobarbiturate the oxygen of the carbonyl group of the urea residue is replaced by a sulfur atom (thiourea).

However, it is interesting to observe that the barbituric acid itself does not possess any hypnotic properties, but such characteristic is conferred only when the hydrogen atoms at C-5 are replaced by organic groups (alkyl or aryl).

Dr. Amged 8 The barbiturates are 5,5-disubstituted barbituric acids. The following scheme shows how the 5,5-dialkyl compounds are synthesized. Substitution of thiourea for urea produces the 2-thiobarbiturates, useful as induction anesthetics.

1.2.3.b.1 Clinical applications

A prescription for long-term use of barbiturates as hypnotics rarely is indicated. The currently available barbiturates are shown below:

Hypnotic Onset Duration

Barbiturate R5 R5 Dose (mg) (min) (hours)

Amobarbital C2H5– (CH3)2CHCH2CH2– 100–200 45–60 6–8

Aprobarbital CH2=CHCH2– (CH3)2CH– 40–160 45–60 6–8

Butabarbital C2H5– CH3CH2CH(CH3)– 50–100 45–60 6–8

Pentobarbital C2H5– CH3(CH2)2CH(CH3)– 100 10–15 3–4

Phenobarbital C2H5– C6H5– 100–320 30–60 10–16

Secobarbital CH2=CHCH2– CH3(CH2)2CH(CH3)– 100 10–15 3–4

Dr. Amged 9 1.2.3.b.2 Classification of Barbiturates

Barbiturates are classified, rather arbitrary, by the duration of their clinical effects.

A. Long Acting Barbiturates

The onset of action for long acting barbiturates is visible after an hour or so, and the duration of action last for 6-10 hours. They are largely excreted by the kidney. Examples:

Phenobarbital is prepared by treating phenyl malonic ester with ethyl bromide and sodium ethoxide. The lonely active hydrogen atom gets replaced with an ethyl group, through enol-form, thus forming phenyl malonic ester. Lastly, this on condensation with urea loses two molecules of ethanol and finally forms the desired compound phenobarbital.

Phenobarbital is used both as sedative and hypnotic. It is the drug of choice in the treatment of grandmal and petitmal epilepsy.

Dr. Amged 10 B. Intermediate Acting Barbiturates

The onset of action for intermediate acting barbiturates is 30 minutes and their hypnotic effect last for 2 to 6 hours. Most of them first degraded by the liver and the metabolized product subsequently excreted by the kidney. They are generally used in insomnia and also as a pre-operative sedative. They also find their use in the treatment of convulsions when administered intravenously. Examples:

C. Short Acting Barbiturates

The onset of action for short acting barbiturates falls within 15 minutes and their hypnotic action last for 1 to 2 hours. They are mostly metabolized in the liver. They are invariably used in the treatment of insomnia and pre-operative medication. Examples:

C. Ultra-Short Acting Barbiturates

These act almost instantaneously, i.e., within a few seconds after administration. Because of this peculiar characteristic they are usually employed to produce general anesthesia and to control convulsions. They may be used either alone or in conjugation with inhalation anesthesia. After administration, they are first deposited in adipose tissues but are eventually dependent on the liver and kidney for their ultimate metabolic degradation and elimination. Example:

Dr. Amged 11

1.2.3.b.3 Mechanism of Action of Barbiturates

The effect of the barbiturates is marked by a decrease in functional activities in the brain.

In vertebrates, γ-aminobutyric acid (GABA) acts at inhibitory synapses in the brain by binding to specific transmembrane receptors in the plasma membrane of both pre- and postsynaptic neuronal processes. This binding causes the opening of ion channels to allow the flow of either negatively charged chloride ions into the cell or positively charged potassium ions out of the cell. This action results in a negative change in the transmembrane potential, usually causing hyperpolarization. Two general classes of

GABA receptor are known: GABAA in which the receptor is part of a ligand-gated ion channel complex, and GABAB, which are G protein-coupled receptors that open or close ion channels via intermediaries (G proteins).

At therapeutic doses, the barbiturates enhance the GABAergic inhibitory response by influencing conductance at the chloride channel. At higher concentrations, the barbiturates can potentiate the GABAA-mediated chloride ion conductance and enhance both GABA and benzodiazepine binding.

1.2.3.b.4 Structure Activity Relationships

Hundreds of barbiturates have been synthesized on a trial-and-error basis. Although many structural features required for hypnotic activity have been recorded, no clear correlation between structure and activity has emerged. It was postulated that to possess good hypnotic activity, a barbituric acid must be a weak acid and must have a lipid/water partition coefficient between certain limits. Therefore, only the 5,5-disubstituted barbituric acids, the 5,5-disubstituted thiobarbituric acids, and the 1,5,5-trisubstituted barbituric acids possess acceptable hypnotic, anticonvulsant, or anesthetic activity. All

Dr. Amged 12 other substitution patterns, such as 5-monsubstituted barbituric acids, 1,3-disubstituted barbituric acids, or 1,3,5,5-tetrasubstituted barbituric acids, are inactive or produce convulsions.

1.2.3.b.4.1 5,5-Disubstitution

As the number of carbon atoms at the fifth carbon position increases, the lipophilic character of the substituted barbituric acids also increases. Branching, unsaturation, replacement of alicyclic or aromatic substituents for alkyl substituents, and introduction of halogen into the alkyl substituents all increase the lipid solubility of the barbituric acid derivatives. A limit is reached, however, because as the lipophilic character increases, the hydrophilic character decreases. Although lipophilic character determines the ability of compounds to cross the blood-brain barrier, hydrophilic character also is important, because it determines solubility in biological fluids and ensures that the compound reaches the blood-brain barrier. Introduction of polar groups into the alkyl substituent decreases lipid solubility below desirable levels. Modifications at this position by variation of the alkyl substituents were of primary importance in the development of barbiturates with short to intermediate duration of action.

Dr. Amged 13

1.2.3.b.4.2 Substitution on Nitrogen

Substitution of one imide hydrogen by alkyl groups increase lipid solubility. The result is a quicker onset and a shorter duration of activity. As the size of the N-alkyl substituent increases (methyl, ethyl, propyl), the lipid solubility increases and the hydrophilic character decreases beyond limits. Furthermore, attachment of large alkyl groups (starting with the ethyl group) to the nitrogen imparts convulsant properties to barbiturates. Attachment of alkyl substituents to both N1 and N3 renders the drug nonacidic, making them inactive. Modifications at this position are of primary importance in the barbiturates used as anticonvulsants and anesthetics.

1.2.3.b.4.3 Modification of Oxygen

Replacement of C2 oxygen by sulfur increases lipid solubility. Because maximal thiobarbiturate brain levels are quickly reached, onset of activity is rapid. As a result, these drugs (i.e., thiopental) are used as intravenous anesthetics.

Dr. Amged 14 1.2.3.b.5 Metabolism

Barbiturates lose their activities through metabolic transformation and redistribution. The metabolism of the barbiturates takes place primarily in the liver. After metabolism, the lipophilic character of barbiturates decreases and this is associated with a loss in depressant activity. Although not used as a hypnotic, the metabolic pathway for mephobarbital is representative of the metabolic pathway for the barbiturates.

O O O (1) (2) 5 NH HO 5 NH RO 5 NH

O N O O N O O N O

CH3 CH3 CH3 (R/S)-Mephobarbital R/S = 6.9 R = glucuronide and/or sulfate conjugate (3)

O O O (1) (2) 5 NH HO 5 NH RO 5 NH

O N O O N O O N O H H H phenobarbital O R = glucuronide and/or sulfate conjugate NH (2) 5 O N O

HO2C O OH H OH OH glucuronide conjugate S/R = 6.8

The major pathways by which the activity of barbiturates is terminated include the following:

(1) Oxidation of substituents at carbon 5. The initial products are alcohols or phenols that form glucuronide and sulfate conjugates.

(2) Conjugation of the heterocyclic nitrogen with glucuronides. This unusual conjugation pathway can be as important as oxidative metabolism in the

Dr. Amged 15 biotransformation of 5,5-disubstituted barbiturates (phenobarbital, amobarbital, pentobarbital).

(3) Oxidative N-dealkylation at the nitrogen. Introduction of an alkyl group on barbiturate nitrogen introduces a site of asymmetry at the 5 position. The S-isomer of these barbiturates primarily undergoes N-dealkylation, and the R-isomer primarily undergoes oxidation at the 5 position.

(4) Oxidative desulfurization of 2-thiobarbiturates takes place readily to yield the more hydrophilic barbiturates.

1.2.3.c Benzodiazepines

Benzodiazepines are compounds whose core chemical structure is the fusion of a benzene ring and diazepine ring.

Benzodiazepines and benzodiazepine-like drugs bind to a benzodiazepine recognition site or benzodiazepine receptor, one of several discrete allosteric sites that modulate the effect of the neurotransmitter γ-aminobutyric acid (GABA) when it binds to type A

GABA (GABAA) receptors in the brain. The GABAA receptor is a ligand-gated chloride ion channel. Thus, benzodiazepines enhance the effect of γ -aminobutyric acid, which results in sedative, hypnotic, anxiolytic, anticonvulsant, muscle relaxant, and amnesic action. Benzodiazepines are used clinically as daytime anxiolytic, sleep inducers, anesthetics, anticonvulsants, and muscle relaxants.

The benzodiazepines that are specifically promoted as sleep inducers are shown below, however, it is important to keep in mind that depending on the dose, any benzodiazepine may be used for its hypnotic effect.

Dr. Amged 16 NEt2 CF3 CH3 N N O S N N N N N N

Cl N Cl N Cl N Cl N F F Cl

Flurazepam Quazepam Triazolam Estazolam

CH H O 3 O N N OH

O2N N Cl N

Nitrazepam Temazepam

1.2.3.d Nonbenzodiazepine GABAA Agonists

A new group of sedative-hypnotic agents similar to the benzodiazepines –zaleplon, zolpidem, and zopiclone– have been developed with affinity for the GABA receptor complex. This produces a more efficacious clinical profile with fewer side effects than the benzodiazepines. Zaleplon, zolpidem, and zopiclone (the Z drugs) are structurally distinct, nonbenzodiazepine structures and are being used as short-acting sedative hypnotics in the United States and Europe. They act at the GABAA high affinity receptors comparable to the benzodiazepines but with different subunit specificity.

Dr. Amged 17 Some generalities associated with these Z compounds is that they are very lipophilic, facilitating their rapid absorption and the absence of active metabolites in the plasma and brain tissue.

1.2.3.d.1 Metabolism of zolpidem:

CH3 CH3 major N pathway N

N O O N O H2C C

OH N CH3 OH N CH3 H3C H3C

major pathway

CH3 CH3 N HO N

N O N O H3C minor H3C pathway N CH3 N CH3 H3C H3C Zolpidem

major pathway O CH OH C 2 OH N N

N O N O H3C major H3C pathway N CH3 N CH3 H3C H3C

Dr. Amged 18 1.2.3.d.2 Metabolism of zaleplon:

1.2.3.d.3 Metabolism of zoplicone:

1.2.3.e Melatonin Receptor Agonists

Melatonin, at times referred to as the hormone of darkness, is synthesized in the pineal gland and normally is secreted during the night.

Studies indicate that melatonin may have effects on circadian rhythm and sleep process. The presences of pharmacologically specific receptors for melatonin in which the molecular structures are known are referred to as MT1, MT2, and MT3 receptors. The

MT1 receptor appears to be primarily involved in initiating sleep. Melatonin is sold as a food supplement in the United States, but it has become popular for use as a hypnotic and

Dr. Amged 19 for alleviating jet lag (a flight across five or more time zones) and helping to resynchronize individuals who have difficulty adapting to night-shift work. However, as a neurohormone, melatonin is a poor drug because it is poor absorption, low oral bioavailability, rapid first-pass metabolism to 6-hydroxymelatonin (its primary metabolite), and ubiquitous effects. In the search for melatonin agonists as sedative, the melatonin molecule was reengineered by substituting the nitrogen of the indole ring with a carbon to give an indane ring bio-isostere of melatonin and by constraining the conformational flexibility of the 5-methoxy group into a furan ring to form either an angular indeno[5,4-b]furan or a linear indeno[5,6-b]furan heterocyclic ring systems.

CH3 CH3 HN O HN O

HN HN

CH O O 3

CH3 Conformer A Conformer B

CH3 CH3 HN O HN O

H H

O

O

Ramelteon (Indeno[5,6-b]furan) (Indeno[5,4-b]furan)

Subsequent MT1 receptor testing revealed that the indeno[5,6-b]furan has approximately 15000 fold weaker affinity for the MT1 receptor than indeno[5,4-b]furan. Furthermore, the S-enantiomer of indeno[5,4-b]furan showed approximately 500 fold greater affinity than the R-isomer for this receptor. Molecular modeling of the indeno[5,4-b]furan derivative with the MT1 receptor showed that by fixing the orientation of the 5-methoxyl group of melatonin into an angular furan ring plays a major factor for reinforcing the binding of the nonbonding pair oxygen electrons to a histidine residue in

Dr. Amged 20 the ligand binding pocket of the receptor. Thus, the methyl orientation of the 5-methoxy group of melatonin (conformer B) is critical for the optimal orientation of the oxygen lone pair for optimal ligand binding to the receptor. This approach culminated in the discovery and approval of Ramelteon as a very potent and very selective ligand for MT1 receptor, with superior in vivo activity and safety profile for use in the treatment of insomnia.

1.2.3.e.1 Synthesis of (S)-Ramelteon

1.2.3.f Antihistamines

Some of the histamine (H1)-receptor antagonists that can cross the blood-brain barrier are used for their hypnotic activity. The primary antihistamines used for their sedative effect are diphenhydramine and doxylamine, which belong to the ethanolamine class of antihistamines.

Dr. Amged 21 1.2.3.g Antidepressants

Antidepressants with sedation as a side effect are used to treat insomnia. This class of drugs will later on be discussed in details.

1.3. Antiseizure Drugs

The most important property of the nerve cell is its excitability. It responds to excitation by generating an action potential, which may, when excited, lead to repeated excessive activity.

An epileptic seizure is a transient symptom of excessive or synchronous neuronal activity in the brain. This excessive neuronal discharge may be brought about by a disturbance of physicochemical function and electrical activity of the brain. The cause of this abnormality, however, is not clearly understood. The epileptic seizures can manifest as an alteration in mental state, tonic or clonic movements, convulsions, and various other psychic symptoms. The outward effect can be as dramatic as a wild thrashing movement or as mild as a brief loss of awareness.

Two possible mechanisms for convulsive disorders have been suggested: a loss of the normal inhibitory control mechanism, and a chemical supersensitivity that increases excitability of neuronal elements.

1.3.1. Seizure Classification

Seizures are classified broadly as:

(1) Partial seizures, in which the abnormal firing initially occurs in a small number of neurons but may spread to adjacent areas.

(2) Generalized seizures, in which virtually the entire brain is affected simultaneously.

Dr. Amged 22

1.3.2. General Mechanism of Action of Antiseizure Drugs

In order to bring normal balance between excitatory and inhibitory postsynaptic potential, antiseizure drugs may use one or more of the following mechanisms:

(1) Enhancement of GABA-mediated inhibition: the drug may act directly on the GABA-receptor-chloride channel complex (e.g., benzodiazepines, barboturates), and inhibit the metabolism of GABA (e.g., vigabatrin, valproate) or increase the release of GABA (e.g., gabapentin). This mechanism provides protection against generalized and focal seizures.

Dr. Amged 23 (2) Suppression of rapid repetitive firing: this mechanism of action of antiseizure drugs (e.g., phenytoin, carbamazepine, valproate, lamotrigine) involves the prolongation and the closing of inactivation gate of Na+ channels, thus reducing the ability of neurons to fire at high frequencies. This mechansim provides protection against maximal electric shock in animals and focal seizures in humans.

(3) Reduction of current through T-type Ca++ channels: a low threshold Ca++ current (T-type) governs oscillatory responses in thalamic neurons. Reduction of this current by antiseizure drugs (e.g., ethosuximide, dimethadione, valproate) explains the mechanism of action against absence seizures.

(4) Reduction of excitatory glutaminergic neurotransmission: some antiseizure drugs (e.g., phenobarbital, topiramate) block the AMPA receptor and some (felbamate) block NMDA receptors.

1.3.3. Antiseizure drugs

The primary use of antiseizure drugs is in the prevention and control of epileptic seizures. Theoretically, the ideal antiseizure drug should, among other things, completely suppress seizures in doses that do not cause sedation or other undesired CNS toxicity. It should be well tolerated and highly effective against various types of seizures and be devoid of undesirable side effects on vital organs and functions. Its onset of action should be rapid after parenteral injection for control of status epilepticus, and it should have a long duration of effect after oral administration for prevention of recurrent seizures.

The first effective remedy, potassium bromide, was introduced in 1857. This drug was largely replaced by barbiturates (phenobarbital) in 1912.

The usefulness of both potassium bromide and phenobarbital in convulsive disorders was discovered by chance, but phenytoin (hydantoin derivative) was developed in 1937 as the result of a study of antiepileptic drugs (AEDs) in animals.

Many of the standard AEDs that contain the ureide structure, as shown, below, have been used clinically for more than 30 years without much change in their ureide

Dr. Amged 24 structures. Small change in X substituent of the ureide structure can cause significant changes in the type of seizures controlled.

As a result of rapid developments in molecular biological techniques for the study of the neurophysiology of epilepsy and in the interactions of AEDs with neurotransmitters at ion channels or brain receptors, a new generation of clinically available AEDs has emerged. These AEDs include felbamate, gabapentin, lamotrigine, levetiracetam, oxcarbazepine, tiagabine, topiramate, and zonisamide. Their mechanisms of action are targeted toward ion channels and brain receptors either by enhancing brain GABA activity or by inhibiting excitatory amino acids. This new generation of AEDs also exhibit limited drug interactions with fewer adverse effects.

Dr. Amged 25 A rational approach to the drug discovery process is necessary to develop new leads to novel effective therapy and to use structure-activity relationships to fine-tune the pharmacology of existing AEDs with the same or better efficacy and fewer adverse effects.

1.3.2.a. Hydantoins

The hydantoins have a 5-membered ring structure containing two nitrogens in an ureide configuration (imidazo-2,4-diones). They have comprised some of the most widely used drugs for treating severe motor and psychomotor epileptic seizures. The structures for the clinically available hydantoins are shown below:

Phenytoin is the prototype and most commonly prescribed member of the hydantoin family of drugs. Hydantoins usually have very poor water solubility. The apparent pKa of phenytoin is in the range of 8.06 to 8.33 and, thus, can form water soluble sodium salt (pH > 11). Aqueous solutions of phenytoin sodium (pH 11–12) gradually absorb carbon dioxide, neutralizing the alkalinity of the solution causing partial hydrolysis and crystallization of free phenytoin resulting in turbid solutions. When phenytoin sodium is administered intramuscularly (IM), its absorption may be erratic as a result of crystallization of insoluble phenytoin at the injection site because of the decrease in pH from 11.5.

Phenytoin is indicated for initial monotherapy or adjunct treatment of complex partial or tonic-clonic seizures, convulsive status epilepticus, and prophylaxis. It often is selected for initial monotherapy because of its high efficacy and relatively low incidence of side effects. Phenytoin is not used in the treatment of absence seizures, because it may increase their frequency of occurrence. Phenytoin binds to and stabilizes the inactivated

Dr. Amged 26 state of sodium channels, thus producing a use-dependent blockade of repetitive firing and inhibition of the spread of seizure activity to adjacent cortical areas.

Phenytoin is metabolized predominantly to its primary metabolite, 5-(4`- hydroxyphenyl)-5-phenylhydantoin (HPPH). Approximately 60 to 75% of an oral dose is excreted as HPPH glucuronide or sulfate metabolites.

Fosphenytoin sodium is a soluble pro-drug disodium phosphate ester of phenytoin that was developed as a replacement for parenteral phenytoin sodium to circumvent the pH and solubility problems associated with parenteral phenytoin sodium formulations. Unlike phenytoin, fosphenytoin is freely soluble in aqueous solutions and is rapidly absorbed by the IM route. It is rapidly metabolized (conversion half-life, 8–15 minutes) to phenytoin by in vivo phosphatases.

Dr. Amged 27 It is administered IV following benzodiazepines for control of status epilepticus or whenever there is a need to rapidly achieve therapeutic plasma concentrations. Severe bradycardiac adverse events to fosphenytoin, including some fatalities, have been reported. A dose reduction in patients who are elderly or have renal or hepatic impairment has been suggested.

The general reaction (see below) used to prepare this heterocyclic system involves the treatment of a carbonyl compound (2) with ammonium carbonate and KCN (one pot synthesis). The first step in the complex sequence can be visualized as the addition of the elements of the ammonia and hydrogen cyanide to give α-aminonitrile (3). Addition of ammonia to the cyano group would then lead to an amidine (4). Carbon dioxide (CO2) or -2 carbonate ion (CO3) present in the reaction mixture can then add to the quite basic amidine to afford a carbonic acid such as the intermediate (5); attack by adjacent amino group will then close the ring and afford the isolable amino derivative (6).

This is then hydrolyzed to a hydantoin (1) by treatment with aqueous acid.

HN O R H O R 1 NH 3 1 NH

R2 N R2 N H O H O 6 1

1.3.2.a.1. Synthesis of Phenytoin

Dr. Amged 28 1.3.2.a.2. Synthesis of Fosphenytoin

O

O O O O C6H5H2CO P OAg NH N OH N Cl OCH2C6H5 H H PCl3 N N N H O H O H O

Phenytoin

O O O OCH C H O OH P 2 6 5 P O H O N OCH2C6H5 2 N OH N (Hydrogenolysis) N H O H O

Fosphenytoin

1.3.2.a.3. Synthesis of Mephenytoin

1.3.2.b. Barbiturates

The barbiturates are substituted pyrimidine derivatives with an uredie configuration.

They are lipophilic weak acids (pKa 7–8) that are well distributed into brain. Although many barbiturates display sedative-hypnotic activity, only a few have antiseizure properties. Paradoxically, many barbiturates cause convulsions at larger doses. The barbiturates clinically useful as AEDs are:

O O O O O O

HN NH HN N HN NH CH3 O O Phenobarbital Mephobarbital Primidone pKa = 7.4 pKa = 7.7

Dr. Amged 29 The mechanism of antiseizure action for the barbiturates is unknown but is thought to involve blockade of sodium channels and enhancement of GABA-mediated inhibitory transmission.

1.3.2.c. Benzodiazepines

This class of drugs has been widely used as sedative-hypnotics and antianxiety drugs. The benzodiazepines diazepam, lorazepam, clonazeoam, clorazepate dipotassium, and midazolam are effective for seizure control.

They are thought to produce their antiseizure effects primarily by enhancing the effect of the inhibitory neurotransmitter GABA on the GABAA chloride channel. Additional evidence suggests that the benzodiazepines may diminish voltage-dependent sodium, potassium, and calcium currents in a manner independent of the GABAA/benzodiazepine receptor complex.

1.3.2.d. Oxazolidinediones

These compounds are some of the oldest AEDs in use, having been introduced into antiseizure therapy between 1946 and 1948. At that time, no effective drugs were available to control absence seizures (petit mal disorders). Therefore, the acceptance of

Dr. Amged 30 trimethadione in 1946 and paramethadione in 1948 for the control of absence seizures was rapid. At present, trimethadione is indicated only for control of absence seizures refractory to treatment with other AEDs. It is ineffective against other seizure types. Trimethadione is a pro-drug and is metabolized by N-demethylation to dimethadione, which is an effective antiepileptic.

Because of its potential fatal side effects, trimethadione rarely is used today. It causes malformations or fetal death in up to 87% of pregnancies. Paramethadione is no longer clinically available in the United States.

1.3.2.e. Succinimides

Because oxazolidinediones are toxic, an extensive search was undertaken to replace them with less toxic drugs. Substituting the ring O in the oxazolidinediones with a methylene group gave the antiseizure succinimides. The clinically used succinimides include ethosuximide, methsuximide, and phensuximide, which were introduced between 1951 and 1958 and widely accepted for the treatment of absence seizures.

Dr. Amged 31 1.3.2.f. Iminostilbenes

Carbamazepine (CBZ) was first marketed as a drug to treat trigeminal neuralgia in 1962. It has been used as an anticonvulsant since 1965, and it is presently indicated as entail or adjunct therapy for complex partial, tonic-clonic, and mixed-type seizures. It is one of the two safest and most effective older AEDs for these seizure types (phenytoin is the other) and is chosen for monotherapy because of its high effectiveness and relatively low incidence of side effects.

Carbamazepine can induce its own metabolism. It is metabolized in the liver to an epoxide and several other metabolites. A major metabolic pathway is oxidation by microsomal enzymes to form carbamazepine 10,11 epoxide. This is an active metabolite and is almost completely metabolized to an inactive metabolite, trans-10,11-dihydroxy- 10,11-dihydrocarbamazepine.

Oxacarbazepine is a structural derivative of CBZ, with a ketone in place of the –C=C– on the dibenzazepine ring. This difference helps reduce the impact on the liver of metabolizing the drug and also prevents the serious side effects generally associated with CBZ. Oxacarbazepine is a prodrug which is activated to eslicarbazepine (S-licarbazepine) in the liver.

Dr. Amged 32

Eslicarbazepine acetate (ESL) is a novel antiepileptic agent designed for improved efficacy and safety. It is a derivative of carbamazepine, oxacarbazepine, and a prodrug of the main active metabolite (S)-licarbazepine (one of the enantiomers of the monohydroxy derivative of oxacarbazepine). Eslicarbazepine acetate has the same mechanism of action of the structurally related carbamazepine and oxacarbazepine. It blocks voltage-gated sodium channels, making brain cells less excitable (less likely to generate action potentials).

In clinical trials, ESL demonstrated efficacy and was generally well tolerated. It also demonstrated low potential for drug-drug interaction. A new drug application for ESL for adjunctive therapy in adults with partial-onset seizures was accepted by FDA in June 2009.

1.3.2.g. Bis-Carbamates

Felbamate is a dicarbamate that was approved by the U.S. FDA for antiseizure use in 1993. Because of serious potential toxicity (aplastic anemia and severe hepatotoxicity), felbamate should be reserved for rare, compassionate use by physicians experienced in treating patients with epilepsy that is difficult to control.

Dr. Amged 33

These toxicity effects may be attributed to the formation of toxic metabolites.

Although the metabolism of felbamate has not been fully characterized, felbamate is esterase hydrolyzed to its monocarbamate metabolite, 2-phenyl-1,3-propanediol monocarbamate, which subsequently is oxidized via aldehyde dehydrogenase to its major human metabolite 3-carbamoyl-2-phenylpropionic acid. Other metabolites include the p- hydroxy and mercapturic acid metabolites of felbamate, which have been identified in human urine. An evidence has been provided for the formation of the reactive metabolite, 3-carbamoyl-2-phenylpropionaldehyde (CBMA), from alcohol oxidation of 2-phenyl-1,3- propanediol monocarbamate. CBMA then undergoes spontaneous elimination to another

Dr. Amged 34 reactive intermediate, 2-phenylpropenal (more commonly known as atropaldehyde), which is proposed to play a role in the development of toxicity during felbamate therapy. Evidence for in vivo atropaldehyde formation was confirmed with the identification of its mercapturic acid conjugates in human urine after felbamate administration. More recently, a fluorine analogue of felbamate was synthesized in which the benzylic C2 hydrogen of the propane chain was replaced with fluorine, preventing the formation of atropaldehyde and confirming that the acidic benzylic hydrogen plays a pivotal role in its formation. This analogue is presently undergoing drug development.

Although its mechanism of action is unknown, felbamate antagonizes the NMDA (N- methyl d-aspartate) receptor by binding to the glycine recognition site, preventing the usual glycine-induced increase in calcium channel opening frequency and lowering calcium currents. Flebamate has also been shown to have an effect on GABA receptors binding sites.

1.3.2.h. Gabapentin

It is a water-soluble amino acid originally designed to be a GABA-mimetic analogue capable of penetrating the CNS. Surprisingly, it has no direct GABA-mimetic activity, nor is it active on sodium channels. The mechanism of action remains unknown, although it has been suggested that gabapentin may alter the metabolism or release of GABA. In other words, gapapentin raises brain GABA levels in patients with epilepsy.

Dr. Amged 35 1.3.2.i. Valproic acid

O

OH

Valproic acid (dipropylacetic acid)

Valproate is available as valproic acid and valproate sodium for IV use. Its AED properties were discovered serendipitously when it was used a solvent for potential new

AEDs undergoing testing. Because the pKa of valproic acid is 4.7, the drug is completely ionized at physiologic pH; thus, the valproate ion is almost certainly the pharmacologically active species.

Although its mechanism of action is not clearly established, valproate appears to increase the inhibitory effect of GABA, possibly by activation of glutamic acid decarboxylase or inhibition of GABA-transaminase. The high drug concentrations required, however, cast doubt on the clinical relevance of this effect. Furthermore, valproate recently has been shown to decrease the uptake of GABA into cultured astrocytes; this action may contribute to the AED efficacy.

1.4. Anxiolytic Agents (Minor Tranquilizers)

1.4.1. Anxiety & Anxiety Disorders

Anxiety is a psychological and physiological state characterized by cognitive, somatic, emotional, and behavioral components. These components combine to create an unpleasant feeling that is typically associated with uneasiness, apprehension, fear, or worry.

Anxiety is considered to be a normal reaction to stress. It may help a person to deal with a difficult situation, for example at work or at school, by prompting one to cope with it. When anxiety becomes excessive, it may fall under the classification of an anxiety disorder.

Dr. Amged 36 1.4.2. Etiology of Anxiety Disorder

A variety of neurotransmitters, neuromodulators, (e.g., adenosine), and neuropeptides (e.g., corticotropin-releasing factor) are suggested to be involved in the pathophysiology of anxiety. Currently, abundant evidence exists to document the involvement of the neurotransmitters GABA, norepinephrine, and serotonin in anxiety.

1.4.3. Anxiolytics

1.4.3.a. Benzodiazepines

The benzodiazepines are the prototypic antianxiety agents. They target the GABAA receptor, and although other molecular targets (e.g., serotonin neuroreceptors) now are exploited for anxiolytic pharmacology, none of the alternative approaches has been shown to match either the efficacy or the rapid onset of the benzodiazepines.

1.4.3.a.1. Development of Benzodiazepine Anxiolytics

In the 1950s, the medicinal chemist, Sternbach noted that basic groups frequently impart biological activity, and in accordance with this observation, he synthesized a series of compounds by treating various chloromethylquinazoline N-oxide with amines to produce what he hope would be products with tranquilizer activity. Sternbach’s studies included the reaction of 6-chloro-2-chloromethyl-4-phenylquinazoline-3-oxide with methylamine, which yielded the unexpected rearrangement product 7-chloro-2-(N- methylamino)-5-phenyl-3H-1,4-benzodiazepin-4-oxide. This product was given the code name RO 50690 and screened for pharmacological activity in 1957. Fortunately enough, RO 50690 showed good sedative and hypnotic activities. Renamed chlordiazepoxide, RO 50690 was marketd in 1960 as Librium, a safe and effective anxiolytic agent.

Dr. Amged 37 N R N N R R Cl HN O R

N CH Cl 2 6-chloro-2-chloromethyl- 4-phenylquinazoline-3-oxide N Cl O NHCH N 3 CH3NH2 6-chloro-2-chloromethyl- 4-phenylquinazoline-3-oxide Cl N O

7-chloro-2-(N-methylamino)-5-phenyl-3H- 1,4-benzodiazepin-4-oxide (chlordiazepoxide)

Chlordiazepoxide turned out to have rather remarkable pharmacological properties and tremendous potential as a pharmacotherapeutic product, but it possessed a number of unacceptable physical chemical properties. In an effort to enhance its pharmaceutical elegance, structural modifications of chlordiazepoxide were undertaken that eventually led to the synthesis of diazepam in 1959. In contrast to the maxim that basic groups impart biological activity, diazepam contains no basic nitrogen moiety. Diazepam, however, was found to be 3- to 10-fold more potent than chlordiazepoxide and was marketed in 1963 as the still enormously popular anxiolytic drug Valium. Subsequently, thousands of benzodiazepines derivatives were synthesized, and more 20 benzodiazepines are in clinical use.

Dr. Amged 38 CH F C CH NHCH 3 O 3 2 S H O N 3 N N N

Cl N Cl N Cl N Cl N O O F O

chlordiazepoxide clobazam quazepam demoxepam

Generic name R1 R3 R8 X R1 O N Clonazepam H H NO2 Cl

R3 Clorazepate H COOK Cl H R8 N X Diazepam CH3 H Cl H

Flurazepam CH2CH2N(C2H5)2 H Cl F Class A Benzodiazepines

Halazepam CH2CF3 H Cl H

Lorazepam H OH Cl Cl

Oxazepam H OH Cl H

Temazepam CH3 OH Cl H

R N Generic name R X Y Y N Alprazolam CH3 H N

Cl N Estazolam H H N X Midazolam CH3 F CH

Triazolam CH Cl Class B Benzodiazepines 3 N

Dr. Amged 39 1.4.3.a.2. Structure-Activity Realationships

Ring A. In general, the minimum requirements for binding of 5-phenyl-1,4- benzodiazepin-2-one derivatives to benzodiazepines receptor (BZR) includes an aromatic or heteroaromatic ring (ring A). Substituents on ring A have varied effects on binding of benzodiazepine to the BZR, but such effects are not predictable on the basis of electronic or steric properties. It is generally true, however, that an electronegative group (e.g., halogen or nitro) substituted at the 7-position markedly increases functional anxiolytic activity, albeit effects on binding affinity in vitro are not as dramatic. On the other hand, substituents at position 6, 8, or 9 generally decrease anxiolytic activity. Other 1,4- diazepine derivatives in which ring A is replaced by a heterocycle generally show weak binding affinity in vitro and even less pharmacological activity in vivo when compared to phenyl-substituted analogues.

Ring B. A proton-accepting group is believed to be a structural requirement of benzodiazepine ligand binding to GABAA receptors. Optimal affinity occurs when the proton-accepting group in the 2-position of ring B (i.e., carbonyl moiety) is in a coplanar spatial orientation with the aromatic ring A. substitution of sulfur for oxygen at the 2- position may affect selectivity for binding to GABA BZR subpopulations, but anxiolytic activity is maintained.

Substitution of the methylene 3-position or the imine nitrogen is sterically unfavorable for antagonist activity but has no effect on agonist (i.e., anxiolytic) activity.

Dr. Amged 40 Derivative substituted with a 3-hydroxy moiety have comparable potency to nonhydroxylated analogues and are excreted faster. Esterification of a 3-hydroxy moiety also is possible without loss of potency. Neither the 1-position amide nitrogen nor its substituent is required for in vitro binding to the BZR, and many clinically used analogues are not N-alkylated. Although even relatively long N-alkyl side chains do not dramatically decrease BZR affinity, sterically bulky substituents like tert-butyl drastically reduce receptor affinity and in vivo activity. Neither the 4,5-double bond nor the 4- position nitrogen in ring B is required for in vivo anxiolytic activity, albeit in vitro BZR affinity is decreased if the C=N bond is reduced to C−N. It is proposed that in vivo activity of such derivatives results from oxidation back to C=N. it follows that the 4- oxide moiety of chlorodiazepoxide can be removed without loss of anxiolytic activity.

Ring C. The 5-phenyl ring C is not required for binding to the BZR in vitro. This accessory aromatic ring may contribute favorable hydrophobic or steric interactions to receptor binding, however, and its relationship to ring A planarity may be important. Substitution at the 4`-(para)-position of an appended 5-phenyl ring is unfavorable for agonist activity, but 2`-(ortho)-substituents are not detrimental to agonist activity, suggesting that limitations at the para position are steric, rather than electronic, in nature.

Annelating the 1,2-bond of ring B with an additional electron-rich (i.e., electron acceptor) ring, such as s-triazole or imidazole, also results in pharmacologically active benzodiazepine derivatives with high affinity for the BZR. For example, the s-triazolo- benzodiazepines triazolam, alprazolam, and estazolam and the imidazo-benzodiazepine midazolam are popularly prescribed, clinically effective anxiolytic agents.

Dr. Amged 41 1.4.3.a.3. Physiochemical and Pharmacokinetics Properties

In general, most benzodiazepines have relatively high lipid:water partition coefficients and are completely absorbed after oral administration and rapidly distributed to the brain and other highly perfused organs. A notable exception is clorazepate, which is rapidly decarboxylated, in the stomach (pH < 5), at the 3-position to the active metabolite N- desmethyldiazepam and, subsequently, quickly absorbed.

Hepatic microsomal oxidation, including N-dealkylation and aliphatic hydroxylation, accounts for the major metabolic disposition of most benzodiazepines. Subsequent conjugation of microsomal metabolites by glucuronyl transferase yields polar glucuronides that are excreted in urine. In general, the rate and product of benzodiazepine metabolism varies, depending on route of administration and the individual drug.

Dr. Amged 42 1.5. Antipsychotic Agents

An antipsychotic (or major tranquilizer or neuroleptic) is a medication primarily used to manage psychosis (e.g., delusion or hallucination), particularly in schizophrenia and bipolar disorder. The psychoses differ from the milder behavioral disorders, the neuroses, in that thinking tends to be illogical, bizarre, and loosely organized. Importantly, patients have difficulty understanding reality and their own conditions. There are often hallucinations (usually auditory) and delusions.

Psychoses can be organic or related to a specific toxic chemical, as in delirium produced by some central anticholinergic agents, or to a definite disease process, such as dementia, or they can be idiopathic. Idiopathic psychoses may be acute or chronic. Idiopathic acute psychotic reactions have been reported to follow extremely severe acute stress. Schizophrenia is a group of chronic idiopathic psychotic disorders.

Increased dopamine activity in the mesolimbic pathway of the brain is consistently found in schizophrenic individuals. Thus, suppression of the dopamine activity is the major mechanism by which the antipsychotics elicit their action. This may be achieved via direct interaction (antagonism) with D2 receptors. Antipsychotic drug clinical efficacy, however, is not solely accounted for by D2 receptor interactions; other CNS receptors systems (acetylcholine, histamine, norepinephrine, and serotonin) appear to be involved, especially for the atypical drugs.

Commonly used antipsychotic medications are listed below by group:

(1) Phenothiazine and thioxanthene derivatives.

(2) Butyrophenone derivatives.

(3) Benzamide derivatives.

(4) Benzazepine derivatives.

(5) Benzisoxazole and benzisothiazole derivatives.

Dr. Amged 43 1.5.1. Phenothiazines and Thioxanthenes

1.5.1.a Development of Phenothiazines and Related Neuroleptics

O CH3 O CH3 O CH3 CHOCH2CH2N CH2N CH2N CH3 I CH3 II CH3 III benzodioxanes ethanolamines diphenhydramine (antihistaminic) (antihistaminic) (antihistaminic)

C2H5 CH3 CH3 S NCH2CH2N NCH2CH2N NCH2CH2N C2H5 CH3CH2 CH3 CH3 VI IV V diethazine ethylenediamines tripelennamine (anti-Parkinson) (antihistaminic) (antihistaminic)

Cl

CH3 CH3 S NCH2 CH N S NCH2 CH2 CH2 N CH3 CH3 CH3 VII VIII promethazine chlorpromethazine (antihistaminic) (antipsychotic)

Although the phenothiazine nucleus was synthesized in 1883, and although it was used as an anthelmintic for many years, it has no antipsychotic activity.

Dr. Amged 44 The basic structural type from which the phenothiazine antipsychotic drugs trace their origins in the antihistamines of the benzodioxane type I. In 1937, Bovet hypothesized that specific substances antagonizing histamine ought to exist, tried various compounds known to act on the autonomic nervous system, and was the first to recognize antihistamine activity. With the benzodioxanes as a starting point, many molecular modifications were carried out in various laboratories in a search for other types of antihistamines. The benzodioxanes led to ethers of ethanolamine of type II, which after further modifications led to the benzhydryl ethers (type III), which are characterized by the clinically useful antihistamine diphenhydramine, or to ethylenediamine (type IV), which led to antihistamine drugs, such as tripelennamine (type V). Further modification of the ethylenediamine type of antihistamine resulted in the incorporation of one of the nitrogen atoms into a phenothiazine ring system, which produced phenothiazine (type VI), a compound that was found to have antihistaminic properties and, similar to many other antihistaminic drugs, a strong sedative effect. Diethazine (type VI) is more useful in the treatment of Parkinson’s disease (because of its potent antimuscarinic action) than in allergies, whereas promethazine (type VII) is clinically used as an antihistaminic. After the ability of promethazine to prolong barbiturate-induced sleep in rodents was discovered, the drug was introduced into clinical anesthesia as a potentiating agent.

To enhance the sedative effects of such phenothiazines, Charpentier and Courvoisier synthesized and evaluated many modifications of promethazine. This research effort eventually led to the synthesis of chlorpromazine (type VIII) in 1950. Soon thereafter, the French surgeon Laborit and his coworkers described the ability of this compound to potentiate anesthetics and produce artificial hibernation. The first attempts to treat mental illness with chlorpromazine alone were made in Paris in 1951 and early 1952 by Paraire and Sigwald. Thus, what initially involved minor molecular modifications of an antihistamine that produced sedative side effects resulted in the development of a major class of drugs that initiated a new era in the drug therapy for the mentally illness.

Today, there are many phenothiazine and thioxanthene derivatives in clinical use, for examples:

Dr. Amged 45

1.5.1.b. Structure-Activity Relationships of Phenothiazines and Thioxanthenes

It is presumed that phenothiazine and thioxanthene neuroleptics mediate their pharmacological effects mainly through interaction at D2-type dopamine receptors. Examination of the x-ray structures of dopamine (in the preferred trans α-rotamer conformation) and chlorpromazine (in the preferred conformer) shows that these two structures can be partly superimposed. On the other hand, the active structure of dopamine does not superimpose with the trans-like conformer of chlorpromazine that would be predicted to be inactive. In the preferred conformation of chlorpromazine, its side chain tilts away from the midline toward the chlorine-substituted ring.

The electronegative chlorine atom on ring A is responsible for imparting asymmetry to this molecule, and the attraction of the amine side chain (protonated at physiologic pH)

Dr. Amged 46 toward the ring containing the chlorine atom indicates an important structural feature of such molecules.

Phenothiazines and related compounds lacking a chlorine atom in this position are, in most cases, inactive as neuroleptic drugs. In addition to ring A substituent, another major requirement for therapeutic efficacy of phenothiazines is that the side-chain amine contain three carbons separating the two nitrogen atoms. Phenothiazines with two carbon atoms separating the two nitrogen atoms lack antipsychotic efficacy. Compounds such as promethazine are primarily antihistaminic and are less likely to assume the preferred conformation.

When thioxanthene derivatives that contain an olefinic double bond between the tricyclic ring and the side chain are examined, it can be seen that such structures can exist in either the cis or trans isomeric configuration. The cis isomer of the neuroleptic thiothixene is several-fold more active than both the trans isomer and the compound obtained from saturation of the double bond.

The duration of action of many of the neuroleptics with a free hydroxy (OH) moiety can be considerably prolonged by the preparation of long-chain fatty acid esters. Thus, fluphenazine decanoate and fluphenazine enanthate were the first of these esters to appear

Dr. Amged 47 in clinical use and are longer acting, with fewer side effects, than the unesterified precursor.

1.5.1.c. Metabolism of Phenothiazines and Thioxanthenes

Chlorpromazine can be demethylated, sulfoxidized, hydroxylated, and glucuronidated to yield 7-O-glu-nor1-CPZ-SO.

The combination of such processes leads to more than 100 identified metabolites. Evidence indicates that the 7-hydroxylated derivatives and, possibly, other hydroxylated derivatives as well as the mono- and didemethylated products (nor1-CPZ, nor2-CPZ) are active in vivo and at dopamine D2 receptors, whereas, the sulfoxide (CPZ-SO) is inactive. Although the thioxanthenes are closely related to the phenothiazines in their pharmacological effects, there seems to be at least one major difference in metabolism: Most of thioxanthenes do not form ring hydroxylated derivatives.

Dr. Amged 48 1.5.1.d Synthesis of Chlorpromazine

1.5.2. Butyrophenones

1.5.2.a. Development of Butyrophenone Neuroleptics

In the late 1950s, Janssen and coworkers synthesized the propiophenone and butyrophenone analogues of meperidine (an analgesic) in an effort to increase its analgesic potency. The propiophenone analogue had 200-fold the analgesic potency of meperidine, but the butyrophenone analogue also displayed activity resembling that of chlorpromazine. Janssen and coworkers found that it was possible to eliminate the morphine type of analgesic activity and, simultaneously, to accentuate the chlorpromazine type of neuroleptic activity in butyrophenone series, provided that certain structural changes are made.

Dr. Amged 49 1.5.2.b. Structure-Activity Relationships of Butyrophenones

All butyrophenone derivatives displaying high neuroleptic potency have the following general structure:

The attachment of a tertiary amino group to the fourth carbon of the butyrophenone skeleton is essential for neuroleptic activity; lengthening, shortening, or branching of the three-carbon propyl chain decreases neuroleptic potency. Replacement of the keto moiety (e.g., with the thioketone group, with olefinic or phenoxy groups, or reduction of the carbonyl group) decreases neuroleptic potency. In addition, most potent butyrophenone compounds have a fluorine substituent in the para position of the benzene ring. Variations are possible in the tertiary amino group without loss of neuroleptic potency; for example, the basic nitrogen usually is incorporated into a 6-membered ring (piperidine, tetrahydropyridine, or piperazine) that is substituted in the para position.

Haloperidol (the prototype) was introduced for the treatment of psychoses in Europe in 1958 and in the United States in 1967. It is an effective alternative to more familiar antipsychotic phenothiazine drugs and also is used for manic phase of bipolar (manic- depressive) disorder. Haloperidol decanoate has been introduced as depot maintenance therapy (taken every 4 to 6 weeks).

Other currently available butyrophenones include the followings:

Dr. Amged 50

Modification of the haloperidol butyrophenone side chain by replacement of the keto function with a di-4-flurophenylmethane moiety results in diphenylbutylpiperidine neuroleptics, such as pimozide, penfluridol, and fluspirilene. The diphenylbutyl piperidines neuroleptics have a longer duration of action than the butyrophenone analogues.

F F

N O N OH N NH

F F CF3 pimozide penfluridol Cl F

N N

NH O F fluspirilene

1.5.2.c. Metabolism of Butyrophenones

The oxidative metabolic pathway of butyrophenones, exemplified by haloperidol, is shown below:

Dr. Amged 51

1.5.2.d. Synthesis of Haloperidol

1.5.3. Benzamide Derivatives

Certain benzamide derivatives have both local anesthetic and antiemetic properties. The benzamide metoclopramide has limited local anesthetic activity but is an efficacious antiemetic drug that modifies the gastric motility. Similar to the phenothiazine antiemtics

(e.g., promethazine), metoclopramide was found to antagonize dopamine D2-type receptors in the chemoreceptor trigger zone of the brain stem and, subsequently, was shown to be neuroleptic. Metoclopramide has relatively low affinity and selectivity for several receptors in addition to D2/D3 antagonism. It blocks muscarinic M3 and 5-HT1A as well as 5-HT3 ligand-operated ion channel. Moreover, numerous studies have documented its anticholinesterase activity.

Dr. Amged 52

The weak affinity and lack of selectivity of metoclopramide likely is explained by the large number of permissible conformers arising from the fexible 2-(diethylamino)ethyl moiety.

Several analogues of metoclopramide in which the side chain is incorporated into a pyrrolidine ring include S-(–)-sulpiride and S-(–)-remoxipride. Both drugs display neuroleptic properties.

Dr. Amged 53

1.5.4. Benzazepine Derivatives

Clozapine, olanzapine, loxapine, and quetiapine are benzazepine-type derivatives with antipsychotic activity and a typically low risk of extrapyramidal side effects. Currently, it generally is agreed that their mechanism of action involves occupancy of both D2 and 5-

HT2A receptors. The high receptor 5-HT2A affinity of atypical antipsychotic agents led to the proposal that 5-HT2A antagonism accounts for the lower propensity of these drugs to cause extrapyramidal side effects.

1.5.5. Benzisoxazole and Benzisothiazole Derivatives

Combination of the chemical features present in the potent benzamide D2 antagonists

(e.g., remoxipride) with those of the benzothiazolyl piperazine 5-HT2A antagonists (e.g., tiospirone) led to the development of the 3-(4-piperidinyl)-1,2-benzisoxazole nucleus

Dr. Amged 54 present in the 5-HT2A/D2 antagonist risperidone and ziprasidone, which also have relatively high affinity at histamine H1 and adrenergic α1/α2 receptors.

N S N O

O N O F N N N N

O N Cl tiospirone risperidone N S

N N O N Cl H ziprasidone

2. Antidepressant Drugs

Depression is an illness that involves the body, mood, and thoughts and affects the way one eats and sleeps, the way one feels about oneself, and the way one thinks about things. It often interferes with normal functioning, causing pain and suffering not only to themselves but also to those around them.

Depression affects approximately 10 to 12% of the Western population, with females outnumbering males 2:1: One in four women and 1 in 10 men can expect to develop depression during their lifetime. Thus, depression ranks among the top 5 diseases in the Western countries. Depression affects at least 1 in 50 children under the age 12 years and 1 in 20 teenagers, mostly girls. The increase in the rate of depression among adolescent girls is related more to physical changes that occur during puberty, suggestive of hormonal changes. Premenstrual syndrome and postpartum depression are additional conditions involving depression that specifically affect women and are suggestive of hormonal involvement in the pathogenesis of depression.

Dr. Amged 55 2.1. Types of Depressive Disorders

2.1.a. Major depression

Major depression (also called unipolar depression) is the most serious type of depression; is manifested by a combination of symptoms that interfere with the ability to work, study, sleep, eat and enjoy once-pleasurable activities; and may reoccur several times during a lifetime. Major depression seems to run in families, suggesting that depressive illness can be inherited. The treatment for major depression are medication, psychotherapy, and in extreme cases, electroconvulsive therapy.

2.1.b. Dysthymia

This is a mild, chronic depression that lasts for 2 years or longer and is characterized by chronic symptoms that do not disable but that keep one from functioning well or from feeling good about oneself. Many of those with dysthmia also experience major depressive episodes at some point in their lives. Most people may not realize that they are depressed and continue to function at work or school, but often with the feeling that they are just going through the motions. Antidepressants and psychotherapy can help.

2.1.c. Bipolar Disorder (Manic-Depressive Illness)

Bipolar disorders can be divided into bipolar I (manic-depressive episodes), bipolar II (hypomanic-depressive episodes). Bipolar disorders also appear to run in families and affect men and women equally. Not nearly as prevalent as the other forms of epressive disorders, bipolar disorders are characterized by cyclical periods of depression (lows) with periods of abnormal behavior (highs) known as hypomanic or mania. Sometimes, the mood switches are dramatic and rapid, but most often, they are gradual. When in the depressed cycle, an individual can exhibit the symptoms of depressive disorder. When in the hypomanic manic cycle, the individual may be overactive, overtalkative, and have a great deal of energy. The hypomanic/manic cycle often affects thinking, judgment, and social behavior in ways that cause serious problems and embarrassment. Lithium, carbamazepine, topiramide, and valproic acid are effective mood-stabilizing treatments for bipolar disorders. Also, the typical antipsychotic, olanzepine is used as a mood

Dr. Amged 56 stabilizer for acute bipolar mania or in conjunction with antidepressants (e.g., Symbyax, a combination of olanzepine and fluoxetine).

2.1.d. Other Types of Depressive Disorders

Other, less common types of depression include seasonal effective disorder. This disorder involves symptoms of depression that occur during the fall and winter seasons, when the days are shorter and there is less exposure to natural sunlight. When the spring and summer seasons begin and there is greater exposure to longer hours of daylight, the symptoms of depression disappear. Adjustment disorder with depressed mood is a type of depression that results when a person has something bad happen that depresses him (e.g., loss of one’s job can cause this type of depression). It generally fades as time passes and the person gets over whatever it was that happened. Additional factors involved in its onset include stresses at home, work, or school, and symptoms may persist for as long as six months.

2.2. Biological basis of depression

Current theories regarding the cause of depression support the role of the neurotransmitters serotonin (5-HT) and norepinephrine (NE) in depression and their interrelationships with each other and with dopamine. Although the precise nature of the depression is not fully understood at the level of the chemistry in the brain, several theories explain the role of NE and 5-HT in the cause of depression.

2.2.a. Monoamine Hypothesis

The monoamine hypothesis proposes that depression results from a deficiency in 5- HT and/ or NE and that antidepressant therapy aims to correct these deficiencies. The role of dopamine in depression remains unclear.

NH2 OH HO NH 2 HO

HO N norepinephrine H serotonin (5-HT)

Dr. Amged 57 storage vesicle

neurotransmitter synapse

postsynaptic receptors

The neurotransmitter precursor is taken up into the neuron and converted to the neurotransmitter. The neurotransmitter is stored in synaptic vesicles. Under the appropriate conditions, the synaptic vesicles migrate to and fuse with the cell membrane, releasing their store of the neurotransmitter. Released neurotransmitter interacts withand activate postsynaptic receptors. The action of the neurotransmitter is terminated either by diffusion of the neurotransmitter away from the synapse, with subsequent metabolism, or by the neurotransmitter being taken back up by a transporter into the presynaptic neuron (i.e., reuptake), where it can be stored or metabolized.

2.2.b. Receptor Sensitivity Hypothesis

The receptor sensitivity hypothesis proposes that it is not simply the level of NE or 5- HT in the synapse that matters but, rather, the sensitivity of the postsynaptic receptors to these neurotransmitters. Those with depression, it is speculated, posses postsynaptic receptors that have grown hypersensitivity to NE and 5-HT because of their depletion in the synaptic cleft.

2.2.c. Permissive Hypothesis

The permissive hypothesis emphasizes the importance of the balance between 5-HT and NE in regulating mood, not the absolute levels of these neurotransmitters or their receptors. If 5-HT levels are too low, the balanced control of the NE system is lost, permitting abnormal levels of NE to cause mania, as seen in bipolar disorders. If the NE

Dr. Amged 58 levels fall, the balanced control of the 5-HT system is lost, allowing abnormal levels of 5- HT to cause the person to exhibit the symptoms of depression.

2.2.d. Hormonal Hypothesis

The hormonal hypothesis suggests that changes in the hypothalamus-pituitary-adrenal axis can influence the levels of 5-HT and NE released by nerve cells in the brain and, subsequently, their function. In the event of stress, the hypothalamus produces a hormone locally in the brain called corticotrophin-releasing factor, which in turn stimulates the pituitary gland to secret adrenocotricotropic hormone into the blood, where it stimulates the adrenal glands to release hydrocortisone (cortisol), which prepares the body for dealing with stress. Stress also directly stimulates the adrenal gland to secrete epinephrine and NE. Hydrocortisone can cause depression, especially when released in high-than- usual amounts.

2. 3.a. Biosynthesis of Norepinephrine

2.3.b. Metabolism of Norepinephrine

Dr. Amged 59 2.4.a. Biosynthesis of Serotonin

2.4.b. Metabolism of Serotonin

2.5. General Approaches to Treatment of Depression

Before 1950, there were no antidepressants―at least not as we know them today. The two treatments for depressive illness were either amphetamine stimulants, which often were ineffective and had the general affect of increasing energy and activity, or electroconvulsive therapy, which was effective but had the disadvantage of terrifying and often endangering the patient. Not until the late 1950s were the first generation of antidepressants discovered (the TCAs and MAOIs), not by design but by chance. While searching for chlorpromazine-like compounds to treat schizophrenia, imipramine was recognized by Kuhn for its antidepressant properties, thus becoming the forerunner of the tricyclic class of monoamine reuptake inhibitor antidepressants (i.e., the TCAs).

Dr. Amged 60 The second compound to be discovered was the antitubercular drug isoniazid, which proved to have powerful mood enhancing properties, becoming the forerunner of the MAOIs. With the introduction of imipramine and isoniazid, the theory and treatment of depression changed.

Between 1960 and 1980, TCAs were the major pharmacological treatment for depression. The TCAs, however, have many other actions in addition to blocking monoamine reuptake, including anticholinergic, antihistaminergic, and cardiotoxic side effects that are related to their affinity for muscarinic, histamine, and α1-adrenergic receptors as well as their action on cardiac and central nervous system (CNS) sodium channels in membranes. The improved safety, tolerability, and reuptake selectivity of the newer antidepressants (i.e., selective serotonin reuptake inhibitors [SSRIs], selective NE reuptake inhibitors [SNRIs], and nonselective NE and 5-HT reuptake inhibitors [NSRIs]) have resulted in displacement of the TCAs as the first choice for prophylactic treatment of major depression. The TCAs occupy a narrower, but still important, role in the psychopharmacological therapy.

The early MAOIs irreversibly inhibited the oxidative deamination of the neurotransmitter monoamines, the proposed mechanism of their antidepressant activity. The biggest liability for these MAOIs was their potential to cause life-threatening hypertensive reactions, resulting from the nonselective irreversible inhibition of MAO-A and MAO-B, which decreases the intestinal and hepatic degradation of dietary sources of tyramine. This inhibition of MAO allows excessive amounts of dietary tyramine, a weak sympathomimetic vasoconstrictor, to be absorbed from the food, resulting in increased blood pressure. Inhibition of monoamine oxidases also can alter the pharmacokinetics of monoamine drugs, allowing them to accumulate in the blood and, thus, increasing their potential for causing adverse drug effects and drug―drug interactions. Minimizing drug and food interactions of these early MAOIs inspired the development of a new generation of MAOIs that are both reversible and selective for MAO-A.

Dr. Amged 61 2.6. Antidepressants

An antidepressant is a psychiatric medication used to alleviate mood disorders, such as major depression and dysthymia. Drugs including the monoamine oxidase inhibitors (MAOIs), tricyclic antidepressants (TCAs), selective serotonin reuptake inhibitors (SSRIs), and serotonin-norepinephrine reuptake inhibitors (SNRIs) are most commonly associated with the term.

Traditionally, antidepressants have been classified according to their structure (i.e., secondary or tertiary amine TCAs) or their principle mechanism of action (i.e., MAOIs and SSRIs).

2.6.1. Selective Norepinephrine Reuptake Inhibitors (SNRIs)

2.6.1.a. Tricyclic Secondary Amines Antidepressants

The TCAs contain a 6-7-6 ring arrangement in which the central 7-membered ring is either carbocyclic or heterocyclic, saturated or unsaturated, which is fused to two phenyl rings. The side chain may be attached to any one of the atoms in the central 7-membered ring, but it must be three carbon atoms, either saturated (propyl) or unsaturated (propylidine), and have a terminal amine group (secondary or tertiary). The TCAs differ structurally from the antipsychotic phenothiazines in that the two phenyl (aromatic) rings are connected by a 2-carbon link to form a central 7-membred ring instead of a sulfur bridge.

2.6.1.b. Nontricyclic Secondary Amines Antidepressants

Reboxetine is a potent nontricyclic SNRI in which the propylamine side chain of the TCAs is constrained into a morpholine ring. It is a chiral compound that is marketed as a

Dr. Amged 62 racemic mixture of R,R- and S,S-reboxetine. The antidepressant activity for reboxetine appears to reside with the S,S-(+)-enantiomer, which has approximately 2-fold the inhibition potency of the R,R-enantiomer.

CH CH 3 H 3 H N N O O O (R) O (S) (R) O (S) O

R,R-reboxetine S,S-reboxetine

Each reboxetine enantiomer is metabolized to one primary metabolite, O- desethylreboxetine, and three minor metabolites, two arising via oxidation of the ethoxy aromatic ring and a third yet-unidentified metabolite.

H H N N OH OH O (R) O (S) (R) O (S) O

major metabolite major metabolite of of R,R-reboxetine S,S-reboxetine

Nisoxetine was the initial phenoxyphenylpropylamine synthesized in Lilly research laboratories during the early 1970s from the rearrangement of an oxygen atom in diphenhydramine, a diphenmethoxyethylamine, to a phenoxyphenylpropylamine. Nisoxetine was discovered to be a potent very selective norepinephrine reuptake inhibitor (SNRI), with little affinity for other receptors. It underwent clinical studies as an alternative to Lilly’s best-selling antidepressant, nortriptyline, but without the adverse effects associated with the tricyclic secondary amines. Nisoxetine is now widely used in scientific research as a standard selective noradrenaline reuptake inhibitor. It was never marketed, however, because of a greater interest in developing its 4- trifluoromethylanalogue, fluoxetine, an SSRI.

Dr. Amged 63

The type and position of the ring substitution plays a critical role in the mechanism of action for the nisoxetine analogues (phenoxyphenylpropylamine derivatives). The unsubstituted molecule is a weak SSRI. However, 2-substitutions into the phenoxy ring

(except for the 2-CF3) yields compounds with high potency and selectivity for blocking norepinephrine reuptake (SNRIs), whereas the 4-substitution results in compounds having potent SSRI activity, with the 4-trifuormethyl group (fluoxetine) being the most potent and selective serotonin reuptake transporter (SERT). The substantial changes in transporter selectivity for norepinephrine reuptake transporter (NET) and SERT and the differences in affinity is more likely attributed to the bulky 2-(ortho)-substituted groups, which restricts the flexibility of the aromatic rings, thereby enhancing alignment of the hydrogen-bond acceptor group (the methoxy) with a donor group on the binding site on the NET for NE that is not available for 5-HT binding site. The R-isomer of nisoxetine has 20 times greater affinity than its S-isomer for NET.

Dr. Amged 64

2.6.2. Selective 5-HT Reuptake Inhibitors (SSRIs)

5-HT is a major player in depressive illness, and serotonergic pathways are closely related to mood disorders, especially depression. Thus, drugs affecting the 5-HT levels in the neural synapse and serotonergic pathways may lead to effective therapy of depression.

Although the TCAs, as a group, are effective antidepressants, their adverse-event profile and high potential for toxicity have limited their use. The early antidepressants indicated that 5-HT might play a significant role in depression. Therefore, medicinal chemists set out in search of the ideal SSRI with the goal for developing drugs with:

(1) High affinity and selectivity for the 5-HT uptake transporter (SERT).

(2) Ability to slow or inhibit the transporter when bound to it.

(3) Low affinity for the multiple neuroreceptors known to be responsible for many of the adverse effects of the TCAs (e.g., acetylcholine, histamine, and adrenergic receptors).

(4) No inhibition of the fast sodium channels which cause the cardiotoxicity problems associated with TCAs.

The discovery that certain antihistaminic agents without the condensed aromatic ring systems are selective inhibitors of 5-HT reuptake with little affinity for other neuroreceptors and almost devoid of cardiotoxicity questioned the need for the 10.11-

Dr. Amged 65 ethylene bridge for the TCAs. Thus, the search for inhibitors that selectively blocked 5- HT reuptake without the 7-membered central ring of the TCAs resulted in the synthesis of the diarylpropylamine analogues of the TCAs. Thus, during the late 1960s and early 1970s, antihistamine molecules were structurally manipulated in the search for compounds that selectively inhibited 5-HT reuptake with greater potency. The initial breakthrough came with the synthesis of Z-zimeldine, the first SSRI that selectively inhibited the presynaptic reuptake of 5-HT without the adverse events associated with the multireceptor activities of the TCAs. Zimeldine was synthesized from the manipulation of the antihistamine pheniramine into a diaryl allylamine, the Z-isomer (rigid analogue) of the propylamine group.

Other structural changes that enhanced its potency and selectivity for blocking 5-HT reuptake was moving the regional position of the 2-pyridyl ring of pheniramine to the 3- pyridyl position and substitution of a halogen into the 4-position of the phenyl ring (2- substitutions selectively block NE reuptake). The secondary amine and primary metabolite, norzimeldine, was 15 times more potent than zimeldine for blocking 5-HT reuptake. On the other hand, E-zimeldine is a nonselective inhibitor of 5-HT and NE reuptake, whereas its corresponding secondary amine is a potent and selective inhibitor of NE reuptake.

Dr. Amged 66

It is not unusual for geometric isomers to differ markedly from each other with regard to their receptor or transporter selectivity, affinity, and pharmacodynamic properties. Thus, zimeldine became the first SSRI to be marketed as an antidepressant, but unfortunately, several cases of Guillain-Barre syndrome (an autoimmune disorder attacking the peripheral nervous system) were associated with the use of this drug and led to its withdrawal from the market in 1983.

The success of zimeldine as an SSRI, however, led to the discovery and marketing of several nontricyclic SSRIs from multiple pharmaceutical companies worldwide, for example fluoxetine (patented 1982, Lilly).

Fluoxetine is a 3-phenoxy-3-phenylpropylamine that exhibits selectivity and high affinity for human SERT and low affinity for NET. It is marketed as a racemic mixture of R- and S-fluoxetine. Its selectivity for SERT inhibition depends on the position of the substituent in the phenoxy ring. Monosubstitution in the 4-(para) position of the phenoxy group (with an electron-withdrawing group, e.g., trifluoromethyl group, as in fluoxetine) results in selective inhibition of 5-HT reuptake.

Fluoxetine is metabolized primarily by N-demethylation to its active metabolite norfluoxetine and, to a lesser extent, O-dealkylation to form the inactive metabolite p- trifluoromethylphenol.

Dr. Amged 67

Both R- and S-norfluoxetine are less potent than the corresponding enantiomers of fluoxetine as inhibitors of NE uptake.

2.6.2.a Synthesis of (±)-Fluoxetine

In trying to create a new antidepressant to inhibit NE reuptake, Lundbeck chemists accidently synthesized two new compounds (talopram and tasulopram) having the phenylspiro-isobenzofuran nucleus. These compounds were potent SNRIs, but considering that a number of suicide attempts were reported during clinical studies with these compounds, Lundbeck discontinued the studies. Undeterred, the chemists subsequently converted talopram into citalopram by a single 6-cyano substitution and other modifications (1989). Citalopram was marketed in the United States in 1996 as the

Dr. Amged 68 most selective SSRI and, therefore, as the least likely to cause the adverse effects observed with most of the other antidepressants.

Citalopram is metabolized via hepatic N-demethylation to its major metabolite, N- desmethylcitalopram which exhibits approximately 50% of the potency of citalopram as an inhibitor of 5-HT reuptake.

The patent expired in 2003, allowing other companies to legally produce generic versions. Lundbeck has recently released an updated formulation called escitalopram (also known as Cipralex or Lexapro), which is the S-enantiomer of the racemic citalopram, and acquired a new patent for it.

Escitalopram, the S-enantiomer of citalopram, binds with high affinity and selectivity to the human SERT equivalent to (±)-citalopram. It has been reported that nearly all the activity resides in the S-(+)-isomer and that R-citalopram actually counteracts the action of the S-enantiomer. Studies show that escitalopram exhibits twice the activity of citalopram and is at least 27 times more potent than the R-enantiomer. The R-enantiomer inhibits the S-enantiomer at the transporter.

Dr. Amged 69 2.6.2.b. Synthesis or (±)-Citalopram

OH OH NC NC NC F MgBr O OH NaBH O 4

O

F F H C NC NC 3 CH3 O N CH Cl N O 3 + H H base CH3

reflux

F F ( )-citalopram

2.6.3. Norepinephrine and Serotonin Reuptake Inhibitors (NSRIs)

The NSRI antidepressant drugs in this class block both the NET and SERT (i.e., they combine the mechanism of action of both the SSRIs and SNRIs), exhibiting dual affinity for NET and SERT. Historically, the tertiary amine TCAs displayed dual inhibition of 5- HT and NE presynaptic reuptake, but they also bind to other types of neuroreceptors, which is responsible for their narrow therapeutic window and adverse effects.

2.6.3.a. Tricyclic Tertiary Amine NSRI Antidepressants

The discovery of the antipsychotic activity of chlorpromazine opened the modern era of psychopharmacology. An intense effort ensued in many laboratories to investigate the structure-activity relationships of related compounds. This included the preparation of what may be regarded as a carbocyclic bioisostere of a phenothiazine, with the sulfur bridge of the prototype being replaced by an ethylene chain of approximately the same size, the nitrogen bridge by a trigonal sp2 carbon, and the chlorine substituent by hydrogen. The resulting compound, amitriptyline, unexpectedly showed antidepressant rather than antipsychotic activity.

Dr. Amged 70 S

N Cl

H H C H C 3 N 3 N

CH3 CH3 amitriptyline chlorpromazine

The TCAs in this class belong to the tertiary amine TCAs. The nucleus for the prototype TCA imipramine consists of a dibenzazepine system. The activity is retained when the nucleus is replaced by a dibenzoxepine and/ or the propylamino side chain is attached to a trigonal sp2 carbon rather than trigonal nitrogen. The relatively low bioavailability for the tertiary amine TCAs suggests first-pass metabolism (N- demethylation) to their secondary amine active metabolites (nor or desmethyl metabolites) and aromatic ring hydroxylation.

Dr. Amged 71 2.6.3.a.1. Synthesis of Amitriptyline

2.6.3.a.2. Synthesis of Imipramine

Cl Cl NaNH2

NO NO NO2 2 2 NO2 NO2

H2 heat NaNH2 N NH2 NH2 H

CH3 Cl N CH3

N CH3 N

CH3 imipramine

2.6.3.b. Nontricyclic NSRI Antidepressants

Clinical studies suggest that compounds which increase the synaptic availability of both NE and 5-HT have greater efficacy than single-acting drugs in the treatment of major depression. Thus began the efforts to design a drug that combined the properties of SSRIs and SNRIs that only blocked SERTs and NETs and without the unwanted adverse effects of TCAs. Currently, the nontricyclic dual inhibitors of 5-HT and NE uptake are venlafaxine, milnacipran, and duloxetine.

Dr. Amged 72

2.6.4. Monoamine Oxidase Inhibitors

MAOIs act by inhibiting the activity of monoamine oxidase, thus preventing the breakdown of monoamine neurotransmitters and thereby increasing their availability. At least two isoforms of monoamine oxidase exist, MAO-A and MAO-B, with differences in substrate preference, inhibitor specificity, and tissue distribution. The MAO-A substrates include 5-HT, and the MAO-B substrates include phenylethyamine. Tyramine, epinephrine, NE, and dopamine are substrates for both MAO-A and MAO-B.

The discovery of MAOIs resulted from a search for derivatives of isoniazid (isonicotinic acid hydrazide) with antitubercular activity. During clinical trials with this hydrazide derivative, a rather consistent beneficial effect of mood elevation was noted in depressed patients with tuberculosis. Although no longer used clinically, iproniazid, the first derivative to be synthesized, was found to be hepatotoxic at dosage levels required for antitubercular and antidepressant activity. The antidepressant activity of iproniazid, however, promoted a search for other MAOIs, which resulted in the synthesis of hydrazine and nonhydrazine MAOIs that were relatively less toxic than iproniazid.

Dr. Amged 73

The MAOIs can be classified as hydrazines (e.g., phenelzine) and nonhydrazines (e.g., tranylcypromine), which can block the oxidative deamination of naturally occurring monoamines. MAOIs can also be classified according to their ability to selectively or nonselectively inhibit MAO. The currently available MAOI antidepressants (phenelzine and tranylcypromine) are considered to be irreversible nonselective inhibitors of MAO.

The mechanism of antidepressant action of the MAOIs suggests that an increase in free 5-HT and NE and/ or alterations in other amine concentrations within the CNS is mainly responsible for their antidepressant effect.

The major goal for developing new reversible MAO-A inhibitors is to avoid the severe, life-threatening hypertensive reactions that can occur with irreversible inhibitors. Irreversible inhibition of intestinal and hepatic MAO-A can lead to inhibition of tyramine degradation, thus allowing excessive amounts of naturally occurring tyramine to be absorbed from the food. Because these reversible compounds form unstable complexes with the MAO-A subtype, they can be easily displaced from MAO-A by tyramine. Thus, it becomes possible for ingested tyramine to be metabolized, diminishing the need for the dietary restrictions that plague the use of older irreversible nonselective MAOIs. This new class of selective and reversible inhibitors of MAO-A includes moclobemide.

Dr. Amged 74 Moclobemide is a benzamide derivative containing a morpholine ring with a pKa of 6.2 and a partition coefficient (octanol/pH 7.4 buffer solution) of 40. Moclobemide undergoes a complex metabolism, initially involving morpholine carbon and nitrogen oxidation, deamination, and aromatic hydroxylation.

2.6.4.a. Synthesis of Moclobemide

3. Opioid Analgesics

3.1. Introduction

Agents that decrease pain are referred to as analgesics, or analgetics. Although analgetic is grammatically correct, common use has made analgesic preferable to analgetic for the description of the pain-killing drugs. Pain relieving agents also are called antinociceptives.

A number of classes of drugs are used to relieve pain. The nonsteroidal anti- inflammatory agents have primarily a peripheral site of action, are useful for mild to moderate pain, and often have an anti-inflammatory effect associated with their pain- killing action. Local anesthetics inhibit pain transmission by inhibition of voltage- regulated sodium channels. These agents often are highly toxic when used in concentrations sufficient to relieve chronic or acute pain in ambulatory patients. Dissociative anesthetics (ketamine), and other compounds that act as inhibitors of N- methyl-D-aspartate (NMDA)-activated glutamate receptors in the brain, are effective antinociceptive agents when used alone or in combination with opioids. Compounds,

Dr. Amged 75 such as the antiseizure drug pregabulin, which inhibits voltage regulated calcium ion channels, are useful in treating neuropathic pain. Most central nervous system depressants (e.g., ethanol, barbiturates, and antipsychotics) will cause a decrease in pain perception. Inhibitors of serotonin and norepinephrine reuptake (i.e., antidepressant drugs) are useful either alone or in combination with opioids in treating certain cases of chronic pain. Current research into the antinociceptive effects of centrally acting α- adrenergic-, cannabinoid-, and nicotinic-receptor agonists may yield clinically useful analgesics working by nonopioid mechanisms. Research in one or more of the above areas may lead to new drugs, but at present, severe acute or chronic pain generally is treated most effectively with opioid agents.

Historically, opioid analgesics have been called narcotic analgesics. Narcotic analgesics literally means that the agent cause sleep or loss of consciousness (narcosis) in conjunction with its analgesic effect. The term narcotic has become associated with the addictive properties of opioids and other CNS depressants. The term narcotic analgesic is misleading because the great therapeutic value of the opioids is their ability to induce analgesia without causing narcosis, and because not all opioids are addicting.

3.1.a. Historical Background of Opioid Analgesics

The juice (opium in Greek) or latex from the unripe seed pods of the poppy Papaver somniferum is among the oldest recorded medication used by humans. The writings of Theophrastus around 200 BC describe the use of opium in medicine; however, evidence suggests that opium was used in the Sumerian culture as early as 3500 BC. The initial use of opium was as a tonic, or it was smoked. The pharmacist Surturner first isolated an alkaloid from opium in 1803. He named the alkaloid morphine, after Morpheus, the Greek god of dreams. Codeine, thebaine, and papaverine are other medically important alkaloids that were later isolated from the latex of opium poppies.

Dr. Amged 76

Morphine was among the first compounds to undergo structure modification. Ethylmorphine (the 3-ethyl ether of morphine) was introduced as a medicine in 1898. Diacetylmorphine (heroin), which may be considered to be the first synthetic pro-drug, was synthesized in 1874 and marketed as a nonaddicting analgesic, antidiarrheal, and antitussive agent in 1898.

3.1.b. Opiate/Opioid

The use of the terms opiate and opioid requires clarification. Until the 1980s, the term opiate was used extensively to describe any natural or synthetic agent that was derived from morphine. One could say an opiate was any compound that was structurally related to morphine. In the mid-1970s, the discovery of peptides in the brain with pharmacological actions similar to morphine promoted a change in nomenclature. The peptides were not easily related to morphine structurally, yet their actions were like those produced by morphine. At this time, the term opioid, meaning opium- or morphine-like in terms of pharmacological action, was introduced. The broad group of opium alkaloids and the many naturally occurring and synthetic peptides with morphine-like pharmacological effects are called opioids. In addition to having pharmacological effects similar to morphine, a compound must be antagonized by an opioid antagonist, such as

Dr. Amged 77 naloxone, to be classed as an opioid. The neuronal-located proteins to which opioid agents bind and initiate biological responses are called opioid receptors.

3.2. Endorphins

Scientists had postulated for some time, based on structure-activity relationships (SARs), that opioids bind to specific receptor sites to cause their actions. It was also reasoned that morphine and the synthetic opioid derivatives are not the natural ligands for the opioid receptors and that some analgesic substance must exist within the brain. Techniques to prove these two points were not developed until the mid-1970s. Hughes and co-workers used the electrically stimulated contractions of guinea pig ileum and the mouse vas deferens, which are very sensitive to inhibition by opioids, as bioassays to follow the purification of compounds with morphine-like activity from mammalian brain tissue. These researchers were able isolate and determine the structures of two pentapeptides, Tyr-Gly-Gly-Phe-Met (Met-enkephalin) and Tyr-Gly-Gly-Phe-Leu (Leu- enkephalin), that caused the opioid activity. The compounds were named enkephalins after the Greek word Kaphale, which translates as “from the head”.

Discovery of the enkephalins was soon followed by the identification of other endogenous opioid peptides, including β-endorphin, the dynorphins, and the endomorphins.

The opioid peptides isolated from mammalian tissue are known collectively as endorphins, a word that is derived from a combination of endogenous and morphine. The opioid alkaloids and all of the synthetic opioid derivatives are exogenous opioids. Interestingly, the isolation of morphine and codeine in small amounts has been reported from mammalian brain. The functional significance of endogenous morphine remains unknown.

3.3. Opioid Receptors

There are three major types of opioid receptors: μ, κ, and δ. All three of the receptor types have been well characterized and cloned. A nomenclature adopted by the International Union of Pharmacology (IUPHAR) in 1996 classifies the three opioid

Dr. Amged 78 receptors by the order in which they were cloned. By this classification, δ opioid receptors are OP1 receptors, κ opioid receptors are OP2 receptors, and μ opioid receptors are OP3 receptors. The IUPHAR approved a new nomenclature in 2000, naming the receptors as MOP-μ, DOP-δ, and KOP-κ. In current literature, however, the opioid receptors often are referred to DOP (δ), KOP (κ), and MOP (μ).

3.3.a. The Mu Opioid Receptors (μ)

A number of therapeutically useful compounds have been found that are selective for μ opioid receptors. All of the opioid alkaloids and most of their synthetic derivatives are μ-selective agonists. Morphine, normorphine, and dihydromorphinone have 10- to 20- fold μ receptor selectivity.

Studies have confirmed that all the major pharmacological actions observed on injection of morphine (e.g., analgesia, respiratory depression, tolerance, withdrawal symptoms, decrease gastric motility, and emesis) occur by interactions with μ receptors. We can now see why it is so difficult to remove the side effects of morphine and its analogues, since the receptor with which they bind most strongly is also inherently involved with these side effects.

Naloxone and naltrexone are antagonists that have weak (5- to 10 times) selectivity for μ receptors.

3.3.b. The Kappa Opioid Receptors (κ)

In general, κ agonists (including morphine) produce analgesia and other effects including diuresis, sedation, and dysphoria. Compared to μ agonists, κ agonists lack respiratory depressant, constipation, and strong addictive (euphoria and physical dependence) properties. It was hoped that κ agonists would become useful strong analgesics that lacked addictive properties; however, clinical trails with several highly selective and potent κ agonists were aborted because of the occurrence of unacceptable sedative and dysphoric side effects.

Nalorphine acts as an antagonist at the μ receptor, thus blocking morphine from acting there. However, it acts as a weak agonist at the κ receptor (as does morphine) and so the

Dr. Amged 79 slight analgesia observed with nalorphine is due to the partial activation of the κ receptor. Unfortunately, nalorphine has hallucinogenic side effects. This is causes by nalorphine also binding to completely different, non-analgesic receptor in the brain called the sigma receptor (σ) where it acts as an agonist.

Pentazocine interacts with the μ and κ receptors in the same way, but is able to switch on the κ receptor more strongly. It too suffers the drawback that it switches on the σ receptor. Buprenorphine is slightly different. It binds strongly to all three analgesic receptors and acts as an antagonist at the δ and κ receptors, but act as a partial agonist at the μ receptor to produce its analgesic effect. This might suggest that buprenorphine should suffer the same side effects as morphine. The fact that it does not is related in some way to the rate at which buprenorphine interacts with the receptor. It is slow to bind but, once it has bound, it is slow to leave.

3.3.c. The Delta Opioid Receptors (δ)

Enkephalins, the natural ligands at δ (DOP) receptors, are only slightly selective for δ over μ receptors. Changes in the amino acid composition of the enkephalins can give compounds with high potency and selectivity for δ receptors. Morphine can also bind quite strongly to this receptor.

Naltrindol and naltriben are highly selective non-peptide antagonists for δ receptors. Naltrindol penetrates the CNS and displays antagonist activity that is selective for δ receptors in in vitro and in vivo systems. The δ opioid receptor antagonists have shown clinical potentials as immunosuppressants and in treatment of cocaine abuse.

The following table shows the relative activities of morphine, nalorphine, pentazocine, enkephalins, pethidine, and naloxone. A plus sign indicates that the compound is acting as an agonist. A minus sign means that it acts as an antagonist. A zero sign means that there is no activity or minor activity.

Dr. Amged 80

3.4. Morphine

3.4.a. Isolation

Opium contains a complex mixture of almost 25 alkaloids. The principle alkaloid in the mixture, and the one responsible for analgesic activity, is morphine. Although pure morphine was isolated 1n 1803, it was not until 1833 that chemists at Macfarlane & Co. (now Macfarlane-Smith) in Edinburgh were able to isolate and purify it on a commercial scale. However, since morphine was poorly absorbed orally, it was little used in medicine until the hypodermic syringe was invented in 1853, allowing doctors to inject morphine directly into the blood supply.

Morphine was then found to be a particularly good analgesic and sedative, and was far more effective than crude opium. But there was also a price to be paid. Morphine was used during the American Civil war (1861-1865) and the Franco-Prussian war. However, there was poor understanding about safe dose levels, the effects of long term use, and the increased risks of addiction, tolerance, and respiratory depression. As a result, many casualties were either killed by overdoses or became addicted to the drug.

At this stage, it is worth pointing out that all drugs have side-effects of one sort or another. This is usually due to the drug not being specific enough in its action, and interacting with receptors other than the one of interest. One reason for drug development is to try and eliminate the side effects without losing the useful activity. Therefore, the medicinal chemist has to try and modify the structure of the original drug molecule to make it more specific for the target receptor. Admittedly, this has often been a case of

Dr. Amged 81 trial and error in the past, but there are various strategies (refer to the drug design course) which can be employed. The development of narcotic analgesics is a good example of the traditional approach to medicinal chemistry and provides good examples of various strategies which can be employed in drug development. We can identify several stages:

Stage 1: Recognition that a natural plant or herb (opium from the poppy) has a pharmacological action.

Stage 2: Extraction and identification of the active principle (morphine).

Stage 3: Synthetic studies (full and partial synthesis).

Stage 4: Structure-activity relationships – the synthesis of analogues to see which parts of the molecule are important to biological activity (pharmacophore).

Stage 5: Drug development – the synthesis of analogues to try and improve activity or reduce side effects.

Stage 6: Theories on the analgesic receptors. Synthesis of analogues to test theories.

Stages 5 and 6 are the most challenging and rewarding parts of the procedure as far as the medicinal chemist is concerned, since the possibility exists of improving on what nature has provided. In this way, the chemist hopes to gain a better understanding of the biological process involved, which in turn suggests further possibilities for new drug.

3.4.b. Structure and Properties

By 19th century standards, morphine was an extremely complex molecule and provided a huge challenge to chemists. By 1881, the functional groups on morphine had been identified, but it took many more years to establish the full structure. In those days the only way to find the structure of a complicated molecule was to break it down into simpler fragments which were already known and could be identified. So, for example, the degradation of morphine with strong basic solutions to produce methylamine gas established that there was an N-CH3 fragment in the molecule.

Dr. Amged 82 From these fragments, chemists would propose a structure. Once a structure had been proposed, chemists would then attempt to synthesize the structure. If the properties of the synthesized compound were the same as the natural compound, then the structure would be established. This was a long drawn out affair, made all the more difficult since there were fewer of the synthetic reagents or procedures which are available today. As a result, it was not until 1925 that Sir Robert Robinson proposed the correct structure. A full synthesis of morphine was achieved in 1952 and the structure proposed by Robinson was finally established when it was studied by X-ray crystallography in 1968 (165 years after the original isolation).

Morphine is the active principle of opium and is still one of the most effective painkillers available to medicine. It is especially good for treating dull, constant pain rather than sharp, periodic pain. It acts in the brain’s awareness of pain. Unfortunately, it has a large number of side effects which include the following:

 Depression of the respiratory centre.

 Constipation.

 Excitation.

 Euphoria.

 Nausea.

 Pupil constriction.

 Tolerance.

 Dependence.

Dr. Amged 83

Some side effects are not particularly serious. Some, in fact, can be advantageous. Euphoria, for example, is a useful side effect when treating pain in terminally ill patients. Other side effects, such as constipation, are uncomfortable but can give clues to other possible uses for opiate-like structures. For example, opiate structures are widely used in cough medicines and the treatment of diarrhea.

The dangerous side effects of morphine are those of tolerance and dependence, allied with the effects morphine can have on breathing. In fact, the most common cause of death from a morphine overdose is by suffocation. Tolerance and dependence in the one drug are particularly dangerous and lead to severe withdrawal symptoms when the drug is no longer taken.

Withdrawal symptoms associated with morphine include anorexia, weight loss, pupil dilatation, chills, excessive sweating, abdominal cramps, muscle spasm, hyperirritability, lacrimation, tremor, increased heart rate, and increased blood pressure. No wonder addicts find it hard to kick the habit!

The isolation and structural identification of morphine mark the first two stages of out story and have already been described. The molecule contains five rings, labeled A, B, C, D, and E, and has a pronounced T shape. It is basic because of the tertiary amino group, but it also contains a phenolic group, an alcohol group, an aromatic ring, an ether bridge, and a double bond. The next stage in the procedure is to find out which of these functional groups is essential to the analgesic activity.

Dr. Amged 84 3.4.c. Structure-Activity Relationships

The story of how morphine’s secrets were uncovered is presented here in a logical step-by-step fashion. However, in reality this was not how the problem was tackled at the time. Different compounds were made in a random fashion depending on the ease of synthesis, and the logical pattern followed on from the results obtained. By presenting the development of morphine in the following manner, we are distorting history but we do get a better idea of the general strategies and the logical approach to drug development as a whole.

The first and easiest morphine analogues which can be made are those involving peripheral modifications of the molecule (that is, changes which do not affect the basic skeleton of the molecule). In this approach, we are looking at the different functional groups and discovering whether they are needed or not.

We now look at each of these functional groups in turn.

3.4.c.1. The Phenolic OH

Codeine is the methyl ether of morphine and is also present in opium. It is used for treating moderate pain, coughs, and diarrhea.

MeO EtO H3C O O

O O O

N N N H H H H H H CH3 CH3 CH3

HO HO HO codeine 3-ethylmorphine 3-acetylmorphine

analgesic activity decreases

By methylating the phenolic OH, the analgesic activity drops drastically and codeine is only 0.1% as active as morphine. This drop in activity is observed in other analogues containing a masked phenolic group. Clearly, a free phenolic group is crucial for analgesic activity.

Dr. Amged 85 However, the above result refers to isolated receptors in laboratory experiments. If codeine is administered to patients, its analgesic effect is 20% that of morphine – much better than expected. Why is this so?

The answer lies in the fact that codeine can be metabolized in the liver to give morphine. The methyl ether is removed to give the free phenolic group. Thus, codeine can be viewed as a prodrug for morphine. Further evidence supporting this is provided by the fact that codeine has no analgesic effect at all if it is injected directly into the brain. By doing this, codeine is injected directly into the CNS and does not pass through the liver. As a result, demethylation does not take place.

In all the following examples, the test procedures were carried out on animals or humans and so it must be remembered that there are several possible ways in which a change in activity could have resulted.

3.4.c.2. The 6-Alcohol

It has been shown that masking or complete loss of the alcohol group does not decrease analgesic activity and, in fact, often has the opposite effect. Again, it has to be emphasized that the testing of analgesics has generally been done in vivo and that there are many ways in which improved activity can be achieved.

HO HO HO

O O O

N N N H H H H H H CH3 CH3 O CH3

MeO EtO H3C O heterocodeine 6-ethylmorphine 6-acetylmorphine more active than morphine more active than morphine more active than morphine HO HO HO

O O O

N N N H H H H H H CH3 CH3 CH3

H H O OH active H active active

Dr. Amged 86 In these examples, the improvement in activity is due to the pharmacokinetic properties of these drugs rather than their affinity for the analgesic receptor. In other words, it reflects how much of the drug can reach the receptor rather than how well it binds to it.

There are a number of factors which can be responsible for affecting how much of a drug reaches its target. For example, the active compound might be metabolized to an inactive compound before it reaches the receptor. Alternatively, it might be distributed more efficiently to one part of the body than another.

In this case, the morphine analogues shown are able to reach the analgesic receptor far more efficiently than morphine itself. This is because the analgesic receptors are located in the brain and, to reach the brain, the drugs have to cross a barrier called the blood- brain barrier. The capillaries which supply the brain are lined by a series of fatty membranes which overlap more closely than in any other part of the body. To enter the brain, drugs have to negotiate this barrier. Since the barrier is fatty, highly polar compounds are prevented from crossing. Thus, the more polar groups a molecule has, the more difficulty it has in reaching the brain. Morphine has three polar groups (phenol, alcohol, and an amine), whereas the analogues above have either lost the polar alcohol group or have it masked by an alkyl or acyl group. They therefore enter the brain more easily and accumulate at the receptor sites in greater concentrations; hence, the better analgesic activity.

It is interesting to compare the activities of morphine, 6-acetylmorphine, and diamorphine (heroin). The most active (and the most dangerous) compound of the three is 6-acetylmorphine, which is four times more active than morphine. Heroin is also more active than morphine by a factor of two, but is less active than 6-acetylmorphine. How do we explain this?

Dr. Amged 87

6-Acetylmorphine, as we have seen already, is less polar than morphine and will enter the brain more quickly and in greater concentrations. The phenolic group is free and therefore it will interact immediately with the analgesic receptors.

Heroin has two polar groups which are masked and is therefore the most efficient compound of the three to cross the blood-brain barrier. However, before it can act at the receptor, the acetyl group on the phenolic group has to be removed by esterases in the brain. Therefore, it is more powerful than morphine because it enters the brain more easily, but it is less powerful than 6-acetylmorphine because the 3-acetyl group has to be removed before it can act.

Heroin and 6-acetylmorphine are both more potent analgesics than morphine. Unfortunately, they also have greater side effects and have severe tolerance and dependence characteristics. Heroin is still used to treat terminally ill patients, such as those dying of cancer, but 6-acetylmorphine is so dangerous that its synthesis is banned in many countries.

To conclude, the 6-hydroxyl group is not required for analgesic activity and its removal can be beneficial to analgesic activity.

3.4.c.3. The Double Bond at 7–8

Several analogues, including dihydromorphine have shown that the double bond is not necessary for analgesic activity.

Dr. Amged 88

3.4.c.4. The N-Methyl Group

The N-oxide and N-methyl quaternary salts of morphine are both inactive, which might suggest that the introduction of charge destroys analgesic activity. However, we have to remember that those experiments were performed on animals and it is hardly surprising that no analgesia is observed, since a charged molecule has very little chance of crossing the blood-brain barrier. If these same compounds are injected directly into the brain, a totally different result is obtained and both these compounds are found to have similar analgesic activity to morphine. This fact, allied with the fact that neither compound can lose its charge, shows that nitrogen atom of morphine is ionized when it binds to the receptor.

The replacement of the N-CH3 group with N-H reduces activity but does not eliminate it. The secondary N-H group is more polar than the tertiary N-CH3 group and therefore finds it more difficult to cross the blood-brain barrier, leading to a drop in activity. The fact that significant activity is retained shows that the methyl substituent is not essential to activity.

Dr. Amged 89 However, the nitrogen itself is crucial. If it is removed completely, all analgesic activity is lost. To conclude, the nitrogen atom is essential to analgesic activity and interacts with the analgesic receptor in the ionized form.

3.4.c.5. The Aromatic Ring

The aromatic ring is essential. Compounds lacking it show no analgesic activity.

3.4.c.6. The Ether Bridge

As we shall see later, the ether bridge is not required for analgesic activty.

3.4.c.7. Stereochemistry

At this stage, it is worth making some observations on stereochemistry. Morphine is an asymmetric molecule containing several asymmetric centres, and exists naturally as a single enantiomer. When morphine was synthesized, it was made as a racemic mixture of the naturally occurring enantiomer plus its mirror image. These were separated and the unnatural mirror image was tested for analgesic activity. It was turned out to have no activity whatsoever.

This is not particularly surprising if we consider the interactions which must take place between morphine and its receptor. We have identified that there are at least three important interactions involving the phenol, the aromatic ring, and the amine on morphine. Let us consider a diagrammatic representation of morphine as T-shaped block with three groups marked as shown below. The receptor has complementary binding groups placed in such a way that they can interact with all three groups. If we now

Dr. Amged 90 consider the mirror image of morphine, then we can see that it can interact with only one binding region at any one time.

Epimerization of a single asymmetric centre such as the 14-position is not beneficial either, since changing the stereochemistry at even one asymmetric centre can result in a drastic change of shape, making it difficult for the molecule to bind to the analgesic receptors.

8 2 7 HO 3 HO 1 H3C N 14 9 4 11 6

12 15 10 16 15 13 5 OH O O 16 10 9 12 13 14 O 5 N N 11 H H 4 H H CH3 CH3 8 6 HO 1 3 7 HO OH morphine less aactive than morphine (10% )

To summarize, the important functional groups for analgesic activity in morphine are shown below:

O H

van der Waals Binding H-Bonding O Groups Ionic N H H H CH3

HO morphine

Dr. Amged 91 3.4.d. Development of Morphine Analogues

We now move on to consider the development of morphine analogues. As mentioned before (see the drug design course), there are several strategies used in drug development. The following have been particularly useful in the development of morphine analogues:

 Variation of substituents.

 Drug extension.

 Simplification.

 Rigidification.

3.4.d.1. Variation of Substituents

A series of alkyl chains on the phenolic group give compounds which are inactive or poorly active. We have already identified that the phenol group must be free for analgesic activity.

The removal of the N-methyl group to give normorphine allows a series of alkyl chains to be added to the basic centre. These results are discussed under drug extension since the results obtained are more relevant under that heading.

3.4.d.2. Drug Extension

Drug extension is a strategy by which the molecule is extended by the addition of extra binding groups. The aim here is to probe for further binding regions which might be available in the receptor’s binding site and which might improve the interaction between the drug and the receptor.

MORPHINE MORPHINE Extra Extra binding binding region group

Drug Extension

RECEPTOR RECEPTOR

Dr. Amged 92 However, are such extra binding regions likely? Perhaps morphine is already an ideal fit for the binding site and has already found all the possible binding interactions? Perhaps morphine is produced naturally in the body and there is a receptor specifically for it?

All these scenarios are certainly possible, but rather unlikely. Alkaloids such as morphine are secondary metabolites which are not essential to the growth of the plant and are only produced once the plant is mature. They could be looked upon as luxury items which are not widespread in the chemistry of life and which have evolved in individual species. It seems highly unlikely then, that the ability to synthesize morphine was evolved separately in poppies and humans. It also seems highly unlikely that morphine evolved in the poppy to act as an analgesic, and so we have to conclude that it is a happy coincidence that morphine is able to interact with a painkilling receptor in the body. For this reason, it is quite possible that a search for further binding regions would be productive. For example, it is perfectly possible that there are four important binding regions in the binding site and that morphine only uses three of them. Therefore, why not add binding groups to the morphine skeleton to search for that fourth binding interaction?

Many analogues of morphine have been made with extra functional groups attached. These have rarely shown any improvement. However, there are two exceptions. The introduction of a hydroxyl group at position 14 has been particularly useful. This might be taken to suggest that there is a possible hydrogen bond interaction taking place between the 14-OH group and a suitable amino acid residue on the receptor. However, an alternative explanation is provided in Section 2.3.5.

Dr. Amged 93

The easiest position at which to add substituents (and the most advantageous) has been the nitrogen atom. The synthesis is easily achieved by removing the N-methyl group from morphine to give normorphine, then alkylating the amino group with an alkyl halide. Removal of the N-methyl group was originally achieved by von Braun degradation with cyanogen bromide, but is now more conveniently carried out using a chloroformate reagent such as vinyloxycarbonyl chloride (VOC). The final alkylation step can sometimes be profitably replaced by a two-step process involving an acylation to give an amide, followed by reduction.

The results obtained from the alkylation studies are quite dramatic. As the alkyl group is increased in size from a methyl to butyl group, the activity drops to zero. However, with a larger group such as a pentyl or a hexyl group, activity recovers slightly. None of this is particularly exciting, but when a phenethyl group is attached, the activity increases

Dr. Amged 94 14-fold – a strong indication that a hydrophobic binding region has been located which interacts favorably with the new aromatic ring.

To conclude, the size and nature of the group on the nitrogen is important to activity spectrum. Drug extension can lead to better binding by making use of additional binding interactions.

Before leaving this subject, it is worth describing series of important results arising from varying susbstituents on the nitrogen atom. Spectacular results were obtained when an allyl group or a cyclopropylmethylene group were attached.

No increase in analgesic activity was observed and, in fact, the results were quite the opposite. Naloxone and naltrexone, for example, have no analgesic activity at all, whilst nalorphine retains only weak analgesic activity. However, the important feature about these molecules is that they act as antagonists to morphine. They do this by binding to the

Dr. Amged 95 analgesic receptors without switching them on. Once they have bound to the receptors, they block morphine from binding. As a result, morphine can no longer act as an analgesic. One might be hard pushed to see an advantage in this and with good reason. If we are just considering analgesia, there is none. However, the fact that morphine is blocked from all its receptors means that none of its side-effects are produced either, and it is the blocking of these effects which make antagonists extremely useful.

In particular, accident victims have sometimes been given an overdose of morphine. If this is not treated, then the casualty may die of suffocation. By administering nalorphine, the antagonist displaces morphine from the receptor and binds more strongly, thus preventing morphine from continuing its action.

Naltrexone is eight times more active than naloxone as an antagonist and is given to drug addicts who have been weaned off morphine or heroin. Since naltrexone blocks the opiate receptors, it blocks the effects which the addicts might seek by restarting their habit and makes it less likely they will do so.

There is, however, another interesting observation arising from the biological results of these antagonists. For many years, chemists had been trying to find a morphine analogue with analgesic properties, but without the depressant effects on breathing, or the withdrawal symptoms. There had been so little success that many workers believed that the two properties were directly related, perhaps through the same receptor. The fact that the antagonist naloxone blocked both the analgesia and side effects of morphine did nothing to change that view.

However, the properties of nalorphine offered a glimmer of hope. Nalorphine is a strong antagonist and blocks morphine from its receptors. Therefore, no analgesic activity should be observed. However, a very weak analgesic activity is observed and, what is more, this analgesia appears to be free of the undesired side effects. This was the first sign that a non-addictive, safe analgesic might be possible.

But how can this be? How can a compound be an antagonist of morphine but also act as an agonist and produce analgesia? If it is acting as an agonist, why is the activity so weak and why is it free of the side-effects?

Dr. Amged 96 As has been mentioned before, there is not one single type of analgesic receptor, but several. There are at least three types of analgesic receptors. The differences between them are slight, such that morphine can not distinguish between them and activates them all, but in theory it should be possible to find compounds which would be selective for one type of analgesic receptor over another. However, this is not the way that nalorphine works.

Nalorphine binds to all three types of analgesic receptor and therefore blocks morphine from all three. Nalorphine itself is unable to switch on two of the receptors and is therefore a true antagonist at these receptors. However, at the third type of receptor, nalorphine acts as a weak or partial agonist. In other words, it has activated the receptor, but only weakly. We could imagine how this might occur if the third receptor is controlling something like an ion channel.

Morphine is a strong agonist and interacts strongly with this receptor, leading to a change in receptor conformation which fully opens the ion channel. Ions flow in or out of the cell, resulting in the activation or deactivation of enzymes. Naloxone is a pure antagonist. It binds strongly, but does not produce the correct change in the receptor conformation. Therefore, the ion channel remains closed. Nalorphine binds to the third receptor and changes the tertiary structure of the receptor very slightly, leading to a slight opening of the ion channel. It is therefore a weak agonist at this receptor, but it is also an antagonist since it blocks morphine from full switching on the receptor.

The results observed with nalorphine show that activation of this third type of analgesic receptor leads to analgesia without the undesirable side effects associated with the other two analgesic receptors.

Unfortunately, nalorphine has hallucinogenic side effects resulting from the activation of a non-analgesic receptor, and is therefore unsuitable as an analgesic, but for the first time a certain amount of analgesia had been obtained without the side effects of respiratory depression and tolerance.

Dr. Amged 97 3.4.d.3. Simplification or Drug Dissection

We turn now to more drastic alterations of the morphine structure and ask whether the complete carbon skeleton is really necessary. After all, if we could simplify the molecule, it would be easier to make it in the laboratory. This in turn would allow the chemist to make analogues more easily, more efficiently, and more cheaply.

There are five rings present in the structure of morphine and analogues were made to see which rings could be removed.

3.4.d.3.1. Removing Ring E

Removing ring E leads to a complete loss of activity. This result emphasizes the importance of the basic nitrogen to analgesic activity.

3.4.d.3.2. Removing Ring D

Removing the oxygen bridge gives a series of compounds called the morphinans which have useful analgesic activity. This demonstrates that the oxygen bridge is not essential. Examples are shown below:

Dr. Amged 98

N-Methyl morphinan was the first such compound tested and is only 20% as active as morphine but, since the phenolic group is missing, this not surprising. The more relevant levorphanol structure is five times more active than morphine and, although side effects are also increased, levorphanol has a massive advantage over morphine in that it can be taken orally and lasts much longer in the body. This is because levorphanol is not metabolized in the liver to the same extent as morphine. As might be expected, the mirror image of levorphanol (dextrorphan) has insignificant analgesic activity.

The same strategy of drug extension already described for the morphine structures was also tried on the morphinans with similar results. For example, adding an allyl substituent on the nitrogen gives antagonists. Adding a phenethyl group to the nitrogen greatly increases potency. Adding a 14-OH group also increases activity.

To conclude:

 Morphinans are more potent and longer acting than their morphine counterparts, but they also have higher toxicity and comparable dependence characteristics.

 The modifications carried out on morphine, when carried out on the morphinans, lead to the same biological results. This implies that both types of molecule are binding to the same receptors in the same way.

 The morphinans are easier to synthesize since they are simpler molecules.

Dr. Amged 99 3.4.d.3.3. Removing Rings C and D

Opening both rings C and D gives an interesting group of compounds called the benzomorphans which are found to retain analgesic activity. One of the simplest of these structures is metazocine, which has the same analgesic activity as morphine. Notice that the two methyl groups in metazocine are cis with respect to each other and represent the stumps of the ring C.

HO HO

Me N Me N H H H H CH3 CH2 CH2 Me Me metazocine phenazocine (same potency as morphine) (4X more potent than morphine)

If the same types of chemical modifications are carried out on the benzomorphans as were described for the morphinans and morphine, then the same biological effects are observed. This suggests that the benzomorphans interact similarly with the analgesic receptors as the morphinans and morphine analogues. For example, replacing the N- methyl group of metazocine with a phenethyl group gives phenazocine, which is four times more active than morphine and the first compound to have a useful level of analgesia without dependence properties.

Further developments led to pentazocine, which has proved to be a useful long term analgesic with a very low risk of addiction. A newer compound (bremazocine) has a longer duration, has 200 times the activity of morphine, appears to have no addictive properties, and does not depress breathing.

Dr. Amged 100 These compounds appear to be similar in their action to nalorphine in that they act as antagonists at two of the three types of analgesic receptors, but acts as an agonist at the third. The big difference between nalorphine and compounds like pentazocine is that the latter are far stronger agonists, resulting in a more useful level of analgesia.

Unfortunately, many of these compounds have hallucinogenic side effects because of interactions with a non-analgesic receptor.

We shall come back to the interaction of benzomorphans with analgesic receptors later. For the moment, we can make the following conclusions about benzomorphans:

 Rings C and D are not essential to analgesic activity.

 Analgesia and addiction are not necessarily co-existent.

 6,7-Benzomorphans are clinically useful compounds with reasonable analgesic activity, less addictive liability, and less tolerance.

 Benzomorphans are simpler to synthesize.

3.4.d.3.4. Removing Rings B, C, and D

Removing rings B, C, and D gives a series of compounds known as 4- phenylpiperidines. The analgesic activity of these compounds was discovered by chance in the 1940s when chemists were studding analogues of cocaine for antispasmodic properties. Their structural relationship to morphine was only identified when they were found to be analgesics, and is evident if the structure is drawn as shown below.

Activity can be increased 6-fold by introducing the phenolic group and altering the ester to a ketone to give ketobemidone.

Dr. Amged 101

Meperidine (pethidine) is not as strong an analgesic as morphine and also shares the same undesirable side effects. However, it has a rapid onset and a shorter duration, and as a result has been used as an analgesic for difficult childbirths. The rapid onset and short duration of action mean that there is less chance of the drug depressing the baby’s breathing.

The piperidines are more easily synthesized than any of the previous groups and a large number of analogues have been studied. There is some doubt as to whether they act in the same way as morphine at analgesic receptors, since some of the chemical adaptations we have already described do not lead to comparable biological results. For example, adding allyl or cyclopropyl groups does not give antagonists.

The replacement of the methyl group of meperidine with a cinnamic acid residue increases the activity by 30 times, whereas putting the same group on morphine eliminates activity.

Dr. Amged 102 HO

O EtO N N H H O R R 30X more potent than pethidine HO zero activity

H

R =

O H

These results might have something to do with the fact that the piperidines are far more flexible molecules than the previous structures and are thus more likely to bind with receptors in different ways.

One of the most successful piperidine derivatives is fentanyl, which is up to 100 times more active than morphine. The drug lacks a phenolic group, but is very lipophilic. As a result, it can cross the blood-brain barrier efficiently.

CO2Et

N N CH2 CH2

fentanyl

HO

O O N EtO N H H N R

fentanyl R HO 100X more potent than morphine 14X more potent than morphine

R = CH2 CH 2

Dr. Amged 103 To conclude:

 Rings C, D, and E are not essential for analgesic activity.

 Piperidines retain side effects such as addiction and depression of the respiratory centre.

 Piperidine analgesics are faster acting and have shorter duration.

 The quaternary centre present in piperidines is usually necessary (fentanyl is an exception).

 The aromatic ring and basic nitrogen are essential to activity, but the phenol group is not.

 Piperidine analgesics appear to bind with analgesic receptors in a different manner to previous groups.

3.4.d.3.5. Removing Rings B, C, D, and E

The analgesic methadone (a diphenylheptanone analogue) was discovered in Germany during the Second World War and has proved to be a useful agent, comparable in activity to morphine. Unfortunately, methadone retains morphine-like side effects. However, it is orally active and has less severe emetic and constipation effects. Side effects such as sedation, euphoria, and withdrawal are also less severe and therefore the compound has been given to drug addicts as a substitute for morphine or heroin in order to wean them off these drugs. This is not a complete cure since it merely swaps an addiction to heroin/morphine for an addiction to methadone. However, this is considered less dangerous.

Dr. Amged 104 The molecule has a single asymmetric centre and, when the molecule is drawn in the same manner as morphine, we could expect the R-enantiomer to be more active. This proves to be the case, with the R-enantiomer being twice as powerful as morphine, whereas the S-enantiomer is inactive. This is quite a dramatic difference. Since R- and S- enantiomers have identical physical properties and lipid solubility, they should both reach the receptor site to the same extent, and so the difference in activity is most probably due to receptor-ligand interactions.

Many analogues of methadone have been synthesized, but with little improvement over the parent drug.

3.4.d.4. Rigidification

Up until now, we have considered minor adjustments of functional groups on the periphery of the morphine skeleton or drastic simplification of the morphine skeleton.

A completely different strategy is to make the molecule more complicated or more rigid. This strategy is usually employed in an attempt to remove the side effects of a drug or to increase activity.

It is usually assumed that the side effects of a drug are due to interactions with additional receptors other than the one in which we are interested. These interactions are probably because of the molecule taking up different conformations or shapes. If we make the molecule more rigid so that it takes up fewer conformations, we might eliminate the conformations which are recognized by undesirable receptors, and thus restrict the molecule to the specific conformation which fits the desired receptor. In this way, we would hope to eliminate such side effects as dependence and respiratory depression. We might also expect increased activity since the molecule is more likely to be in the correct conformation to interact with the receptor.

The best example of this tactic in the analgesic field is provided by a group of compounds known as the oripavines. These structures often show remarkably high activity.

Dr. Amged 105 The oripavines are made from an alkaloid which is called thebaine. Thebaine can be extracted from opium along with codeine and morphine, and is very similar in structure to both these compounds. However, unlike morphine and codeine, thebaine has no analgesic activity. There is a diene group present in ring C and, when thebaine reacts with methyl vinyl ketone, a Diels Alder reaction takes place to give an extra ring and increased rigidity to the structure.

A comparison with morphine shows that the extra ring sticks out from what used to be the crossbar of the T-shaped structure.

Dr. Amged 106 Since a ketone group has been introduced, it is now possible to try the strategy of drug extension, this time by adding various groups to the ketone via a Grignard reaction.

By varying the groups added by the Grignard reaction, some remarkably powerful compounds have been obtained. Etorphine, for example, is 10000 times more potent than morphine. This is a combination of the fact that it is a very hydrophobic molecule and can cross the blood-brain barrier 300 times more easily than morphine, as well as the fact that it has 20 times more affinity for the analgesic receptor site because of better binding interactions.

At slightly higher doses than those required for analgesia, it can act as a knock-out drug or sedative. The compound has a considerable margin of safety and is used to immobilize large animals such as elephants. Since the compound is so active, only very small doses are required and these can be dissolved in such small volumes (1 ml) that they can be placed in crossbow darts and fired into the hide of the animal.

Dr. Amged 107 The addition of lipophilic groups is found to improve activity dramatically, indicating the presence of a hydrophobic binding region close by on the receptor. The group best able to interact with this region is a phenethyl substituent, and the product containing this group is even more active than etorphine.

Because of their rigid structures, these compounds are highly selective agents for the analgesic receptors. Unfortunately, the increased analgesic activity is also accompanied by unacceptable side effects. It was therefore decided to see whether N-substituents such as an allyl or cyclopropyl group, would give antagonists as found in the morphine, morphinan, and benzomorphan series of compounds. If so, it might be possible to obtain an oripavine equivalent of pentazocine or nalorphine – an antagonist with some agonist activity and with reduced side effects.

Adding a cyclopropyl group gives a very powerful antagonist called diprenorphine, which is 100 times more potent than nalorphine and can be used to reverse the immobilizing effects of etorphine. Diprenorphine has no analgesic activity.

Dr. Amged 108

Replacing the methyl group derived from the Grignard reagent with a tert-butyl group gives buprenorphine, which has similar properties to drugs like nalorphine and pentazocine in that it has analgesic activity with a very low risk of addiction. This feature appears to be related to the slow onset and removal of buprenorphine from the analgesic receptors. Since these effects are so gradual, the receptor system is not subjected to sudden changes in transmitter levels.

Buprenorphine is the most lipophilic compound in the oripavine series of compounds and therefore enters the brain very easily. Usually, such a drug would react quickly with its receptor. The fact that it does not is therefore a feature of its interaction with the receptor rather than the ease with which it can reach the receptor. It is 100 times more active than morphine as an agonist and four times more active than nalorphine as an antagonist. It is a particularly safe drug since it has very little effect on respiration and what little effect it does have actually decreases at high doses. Therefore, the risks of suffocation from a drug overdose are much smaller than with morphine. Buprenorphine has been used in hospitals to treat patients suffering from cancer and also following

Dr. Amged 109 surgery. Its drawbacks include side effects such as nausea and vomiting, as well as the fact it can not be taken orally. A further use for buprenorphine is as an alternative means to methadone for weaning addicts off heroin.

Buprenorphine binds slowly to analgesic receptors but, once it does bind, it binds very strongly. As a result, less buprenorphine is required to interact with a certain percentage of analgesic receptors than morphine. On the other and, buprenorphine is only a partial agonist. In other words, it is not very efficient at switching the analgesic receptor on. This means that it is unable to reach the maximum level of analgesia which can be acquired by morphine. Overall, buprenorphine’s stronger affinity for analgesic receptors outweighs its relatively weak action, such that buprenorphine can produce analgesia at lower does than morphine. However, if pain levels are high, buprenorphine is unable to reach the levels of analgesia required and morphine has to be used.

Nevertheless, buprenorphine provides another example of an opiate analogue where analgesia has been separated form dangerous side effects.

3.5. Agonists and Antagonists

We return now to look at a particularly interesting problem regarding the agonist/antagonist properties of morphine analogues. Why should such a small changes as replacing an N-methyl group with an allyl group result in such a dramatic change in biological activity such that an agonist becomes an antagonist? Why should a molecule such as nalorphine act as an agonist at one analgesic receptor and an antagonist at another? How can different receptors distinguish between such subtle changes in a molecule?

We shall consider one theory which attempts to explain how these distinctions might take place, but it is important to realize that there are alternative theories. In this particular theory, it is suggested that there are two necessary hydrophobic binding regions present in an analgesic receptor. It is then proposed that a structure will act as an agonist or as an antagonist depending on which of these extra binding region is used. In other words, one of the hydrophobic binding regions is an agonist binding region, whereas the other is an antagonist binding region.

Dr. Amged 110 The model was proposed by Synder and co-workers and is shown below. In the model, the agonist binding region is further away from the nitrogen and is positioned axially with respect to it. The antagonist region is closer and positioned equatorially.

Let us now consider the morphine analogue containing a phenethyl substituent on the nitrogen. It is proposed that this structure binds as already described, such that the phenol, aromatic ring, and basic centre are interacting with their respective binding regions. If the phenethyl group is in the axial position, the aromatic ring is in the correct position to interact with the agonist binding region. However, if the phenethyl group is in the equatorial position, the aromatic ring is placed beyond the antagonist binding region and can not bind. The overall result is increased activity as an agonist.

Let us now consider what happens if the phenethyl group is replaced with an allyl group. In the equatorial position, the allyl group is able to bind strongly to the antagonist binding region, whereas the axial position is barely reaches the agonist binding region, resulting in a weak interaction.

Dr. Amged 111 In this theory, it is proposed that a molecule such as phenazocine (with a phenethyl group) acts as an agonist since it can only bind to the agonist binding region. A molecule such as nalorphine (with an allyl group) can bind to both agonist and antagonist regions and therefore acts as an agonist at one receptor and an antagonist at another. The ration of these effects would depend on the relative equilibrium ratio of the axial and equatorial substituted isomers.

A compound which is a pure antagonist would be forced to have a suitable substituent in the equatorial position. It is believed that the presence of a 14-OH group sterically hinders the isomer with the axial substituent, and forces the substituent to remain equatorial.

3.6. Ideal Opioid Analgesic

Thousands of derivatives of morphine and other μ agonists have been prepared and tested. The objective of most of the synthetic efforts has been to find an analgesic with improved pharmacological properties over known μ agonists. Specifically, one would like to have an orally active drug that retains the strong analgesic properties of morphine yet lacks its ability to cause tolerance, physical dependence, respiratory depression, emesis, and constipation. The success of this search has been limited. Many compounds that are more potent than morphine have been discovered. Also, compounds with pharmacodynamic properties different from those of morphine have been discovered, and some of these compounds are preferred to morphine for selected medical uses. The ideal analgesic drug, however, is yet to be discovered. Research to find new centrally acting analgesics has turned away from μ agonists and now is focused on agents that act through

Dr. Amged 112 other types of subtypes of opioid receptors or through nonopioid neurotransmitter systems.

3.6. Metabolism of Morphine & Codeine

Dr. Amged 113