Discovery and development of antiandrogens
Encyclopedia
In the 1960s, the first antiandrogen
, or androgen
antagonist, was discovered. Antiandrogens antagonise
the androgen receptor
(AR) and thereby block the biological effects of testosterone
and dihydrotestosterone
(DHT). Antiandrogens are important for men with hormonally responsive diseases like prostate cancer
, benign prostatic hyperplasia
(BHP), acne
, seborrhea
, hirsutism
and androgen alopecia
. Antiandrogens are mainly used for the treatment of prostate diseases. New research also suggests that ARs could be linked to the disease progression of triple-negative breast cancer and that antiandrogens can potentially be used to treat it.
Current antiandrogens are small molecule
antiandrogens and can be either steroid
al or nonsteroidal depending on ligand
chemistry. Steroidal antiandrogens share a similar steroid structure, while nonsteroidal antiandrogens may have structurally distinctive pharmacophores. Only a limited number of compounds are available for clinic
al use despite the fact that a very large variety of antiandrogen compounds have been discovered and researched.
, testes
and prostate gland
was established. Charles Brenton Huggins
established a method to measure the effect hormone
changes have on prostatic function
. He found out that castration
or estrogen
administration led to glandular atrophy
, which could be reversed by re-administration of androgen. In 1941 the beneficial effect of androgen ablation on metastatic prostate cancer was realised when Huggins and Clarence Hodges treated these patients by either castration or estrogen therapy. They monitored the prostate size and therapeutic efficacy by measuring serum acid-phosphatase
levels. They concluded that androgenic activity in the body influences prostate cancer at least with respect to serum phosphatase. Huggins was the first to use a systemic approach to treat prostate cancer. He was awarded the Nobel Prize in Physiology or Medicine
in 1966.
It became evident after many findings that androgen ablation, by means of castration or estrogen administration, was not enough to completely cure patients with advanced prostate cancer. In the late 1960s, the AR was discovered and characterised. Screening of chemical libraries
for AR blockers led to the discovery of cyproterone. An acetate
group was then added to cyproterone creating cyproterone acetate
. In the 1970s, flutamide
was discovered. The United States Food and Drug Administration
(FDA) approved it in 1989 for use in treating prostate cancer. In 1995, bicalutamide
was also approved and nilutamide
followed a year later.
subfamily of the nuclear receptor
superfamily. Its function is regulated by the binding of androgens, which initiates sequential conformation changes
of the receptor
that affects receptor-protein
and receptor-DNA interactions. Endogenous androgens are mainly testosterone and DHT.
The AR gene
is more than 90kb long and codes for a protein of 919 amino acid
s. Only one AR gene has been identified in humans which is located on chromosome X
. It comprises four main regions, see figure 1:
Two functions have been identified in AR that have critical roles in the regulation of target gene transactivation
, the N-terminal activation function 1 (AF1) and the C-terminal activation function 2 (AF2). AF1 is ligand-independent and plays the primary role in target gene transactivation. The AF2 is a ligand-dependent and only shows limited function.
, like a typical steroid receptor, and is associated with a complex of heat shock protein
s (HSP) through interactions with LBD. Androgens, either agonist
s or antagonists, position themselves in the ligand-binding pocket (LBP) of the cytosol
ic AR and bind to the LBD, see figure 2. The AR goes through a series of conformational
changes and HSP dissociate from AR. The transformed AR undergoes dimer
isation, phosphorylation
and translocates to the nucleus. The translocated receptor then binds to the androgen-response elements (ARE) on the promoter of the androgen responsive gene, a consensus sequence located either upstream or downstream of the transcription start site (TSS) of AR target genes. Recruitment of other transcription
co-factors (including co-activators and co-repressors) and general transcriptional machinery further ensures the transactivation of AR-regulated gene expression
. All these complicated processes are initiated by the ligand-induced conformational changes in the LBD. Ligand specific recruitment of coregulators
might be crucial for the agonist or antagonist activity of AR ligands. Binding of DNA is also required for AR-regulated gene expression, also known as classic genomic gene function of AR.
and pituitary. Therefore cyproterone blocks the negative feedback
of androgens at the hypothalamic-pituitary level leading to increased luteinizing hormone
(LH) serum levels. This rise in LH levels causes an increase in serum testosterone levels and ultimately diminishes the ability of cyproterone to compete for AR binding and to block androgenic stimulation
.
Cyproterone acetate was developed to overcome this problem. It is formed by adding an acetate group to cyproterone, see figure 3. Cyproterone acetate has a dual mode of action as it competes directly with DHT for binding to AR, but also inhibits gonadotropin
secretion. It thereby reduces androgen, estrogen and LH levels. Cyproterone acetate acts both directly as an antiandrogen in prostate cancer cells and also functions to indirectly decrease serum testosterone levels. The latter causes the limitations of cyproterone acetate, which are central effects on androgen secretion, with subsequent loss of libido
and sexual potency. Several reports also state that cyproterone acetate causes liver hyperplasia
. These side effect
s gave pharmaceutical companies the incentive to search for alternative non-steroidal "pure" antiandrogens that would not have these side effects. Pure antiandrogens block the androgen receptor without exerting any agonistic or any other hormonal activity.
Flutamide became the first non-steroidal antiandrogen to be tested clinically. Later the non-steroidal antiandrogens bicalutamide and nilutamide were developed. The alleged advantages of these compounds were that they did not affect libido or potency like the other centrally acting compounds under development, luteinizing-hormone-releasing hormone
(LHRH) agonists and cyproterone acetate. But this theory did not prove to be true. These non-steroidal antiandrogens eventually crossed the blood-brain barrier
, like cyproterone, leading to a subsequent increase in serum testosterone levels.
to its active form, 2-hydroxyflutamide, and hydrolysis
product, 3-trifluoromethyl-4-nitroaniline. Hydroxyflutamide is a more potent AR antagonist than flutamide in vivo
, with higher binding affinity for the AR. Hydroxyflutamide has an elimination half-life
of about 8 hours in humans. Hydrolysis of the amide bond
represents the major metabolic pathway
for this active metabolite
. By reversing the stimulatory effect of DHT on ventral prostate weight, flutamide is approximately 2-fold more potent
than cyproterone acetate. Hydroxyflutamide has relatively low binding affinity to AR and is therefore generally used at high doses in order to achieve complete AR blockage in therapy.
analog of flutamide, as seen in figure 5. Nilutamide is eliminated exclusively by metabolism, mainly by reduction of the aromatic
nitro group
. Although the hydrolysis of one of the carbonyl
functions of the imidazolinedione was identified, it is much less susceptible to hepatic metabolism than the amide bond in hydroxuflutamide. This results in a longer half-life of nilutamide in humans of 2 days. Nevertheless, the nitro anion-free radical
formed during nitro reduction could still be associated with hepatotoxicity
in humans, especially when using relatively high dosage employed for androgen blockage. Nilutamide causes side-effects which limit its usage, such as pneumonitis
and delayed adaption to darkness.
is an arylpropionamide analog, seen in figure 6. It has replaced flutamide and nilutamide as the first choice antiandrogen for prostate cancer treatment. Bicalutamide is not as hepatotoxic as flutamide and nilutamide and has a longer half-life, of 6 days in humans, that allows once a day administration at lower dosage. Bicalutamide shares the amide bond structure with flutamide. Even so, the amide bond hydrolysis was discovered in rats, not in humans, which could explain the prolonged half life of bicalutamide in humans.
Bicalutamide has a cyano group
at the para position instead of a nitro group like flutamide and nilutamide. This change in groups avoids the nitro reduction observed in nilutamide. Bicalutamide has a chiral carbon in its structure (labeled with an asterisk in figure 6), which is connected to the hydroxyl and methyl groups . It is therefore administered as a racemate. Post-approval investigation revealed that its antiandrogenic activity resides almost entirely in the (R)-enantiomer
. (R)-bicalutamide has an almost fourfold higher affinity for the prostate AR than hydroxyflutamide and has a better side-effect profile compared to other antiandrogens.
atome is replaced by a methyl group
the binding slightly decreases, whereas further removal of the C6 double bond modifies the binding kinetics, see figure 7.
gives weakly active compounds, which can be attributed to the lack of intramolecular hydrogen binding
or to poor hydrogen-bond donor capability. Various combinations of electron-withdrawing
substitutions in the aniline
ring of these drugs, have not shown higher binding to the AR receptor than compounds which have a chloro or trifluoromethyl group at the meta position (R1) and either a cyano or nitrogen group at the para position (R2).
For hydroxyflutamide, a group of compounds that differed in the aromatic ring did not bind to the AR. This suggests that the bisubstitution in the hydroxyflutamide ring is essential for high AR binding affinity. It has also been demonstrated that hydroxyflutamide requires the strong hydrogen bond donor ability of the tertiary hydroxyl group and fixed conformers involved in intramolecular hydrogen binding, to bind effectively to AR.
For bicalutamide, the antiandrogenic activities of sulfide
and sulfone
substitions of the X-linkage were tested in vitro
. The sulfides showed in most cases at least 2-fold higher binding affinity than corresponding sulfones. However, this relationship was reversed when the R3 group was NHSO2CH3, where the binding affinity of sulfone was 3-fold higher than that of sulfide. These results indicate that substituents of the B-ring largely determine the effect of the X-linkage in AR binding. Researchers have proposed that the tertiary hydroxyl group is involved in direct interaction with AR because when an acetyl
group is introduced to that hydroxyl moiety, the receptor binding affinity greatly decreases.
Nilutamide has very low affinity for AR when tested on castrated rat prostate. Modifications such as replacing the N3 atom with oxygen has little effect on affinity of the compound for prostate AR. By replacing the oxygen
atom with a sulfur
atom at the C2 position of the imidazole
ring and adding butylalcohol
to the N3 atom, the receptor binding and biological activity of the compound increases 100 times that of non-steroidal antiandrogens. Also the compound does not bind to other steroid receptors. If a methyl group is changed for the butylalcohol group, the compound shows 3 and 10 times more antiandrogenic activity in vivo than bicalutamide and nilutamide, respectively.
growth. This phenomenon is called antiandrogen withdrawal syndrome (AWS) and is one of the major drawbacks of existing antiandrogens. AWS is defined as tumor regression or symptomatic relief observed upon discontinuation of the antiandrogen therapy. The mechanism for this is not fully understood but current theories include alterations of the AR gene, coregulator proteins and/or signal transduction pathways. This antiandrogen resistance may also be linked to the relative weakness of current antiandrogens as they have an affinity 50 times or more lower than that of DHT for the AR. This may also explain why compensatory AR overexpression is often observed.
s in the LBD that alter ligand specificity and/or functional activity exist and are thought to contribute to the conversion of some AR antagonists into agonists, which explains the paradoxical temporary improvement sometimes observed in patients when antiandrogen therapy is stopped. These mutations can have great affect on the antagonist activities of current small molecule antiandrogens and make them less efficient in blocking AR function via indirect modulation from inside of the LBP. Recent studies with circulating tumor cells, suggest that the mutation frequency is higher than previously assumed based on tumor biopsies
.
The T877A, W741L and W741C mutations are examples of known AR LBD mutations. The LNCaP
prostate cancer cell line
expresses AR with a T877A point mutation that causes proliferation in the presence of the antiandrogens hydroxyflutamide and cyproterone acetate. This mutation has also been discovered in patients with antiandrogen withdrawal syndrome being treated with these compounds. In another study, bicalutamide treatment of LNCaP cells resulted in two LBD mutations, W741L and W741C, causing bicalutamide to acquire agonist activity to both mutant ARs. The W741L mutation generates additional space such that the sulfonyl-linked phenyl
ring of bicalutamide is accommodated at the location of the missing indole
ring of W741. In non-mutant AR, the presence of the W741 side chain probably forces bicalutamide to protrude out thus precluding the active position of H12 on the AR receptor.
However, hydroxyflutamide worked as an antagonist for W741 mutant ARs. This concurs with the theory that flutamide and nilutamide antagonize AR through the mechanism of “passive antagonism”, as they are of a more modest size then bicalutamide. These drugs may therefore be effective as a second-line therapy for refractory prostate cancer previously treated with bicalutamide.
Steroid receptors have similarities in terms of gene sequences and protein structures, leading often to functional crosstalk among steroid receptors. One of the criteria for AR peptide antagonists is to achieve high degree of specificity for the AR. It is though important to realize that AR specificity does not necessarily translate in vivo, since peptide antagonists may also interact with protein targets other than AR.
. Therefore most of the research on peptide AR antagonists focuses on peptide
s that may directly block the AF2 in AR LBD from protein surface. Even in bound mutant AR, peptide antagonists would be able to block the AF2 function via direct surface interaction, regardless of the ligand bound.
Research on these peptide antagonists are usually carried out by affinity screening of phage display
libraries that express random peptides containing various signature motifs
. ARs seem to have a distinct preference for ‘FxxLF’ type of binding motifs (where F = phenylalanine
, L = leucine
, and X = any amino acid residue), whereas other nuclear receptors have a highly similar binding mechanism for ‘LxxLL’ type of binding motifs. This provides a unique opportunity for the development of AR-specific peptides.
Even though small molecule antagonists and peptide antagonist targeting AF2 surface differ in binding sites, they both inhibit AR function by disrupting AF2 function. Therefore mechanistically, these peptide antagonists may also be classified as ‘AF2 antagonists’.
and circular dichroism spectroscopy
analysis suggest that AR NTD is highly disordered under native conditions, making it a difficult target for drug discovery.
In 2008 there were reports of a chlorinated peptide, Sintokamide A, isolated from marine sponges that effectively inhibits AR N-terminal domain-activated reporter gene transcription, see figure 8. The evidence presented was not sufficient enough to support the conclusion that Sintokamide A directly inhibits the function of AR NTD, and the mechanism of action needs further investigation.
s (SARMs) has been proposed as a novel approach for the treatment of prostate cancer. These ligands should behave as antagonists in the prostate with either no activity or agonist activity in other target tissues, so as to have little or no effects in the anabolic tissues or central nervous system
(CNS). However discovering this new class of ligands might be challenging because the molecular mechanism of AR action is not well understood.
Several mechanisms have been proposed to achieve this tissue selectivity of AR ligands. The most definitive evidence exists for the role of 5-alpha reductase
. 5-alpha reductase is only expressed in specific tissues and could therefore be a unique contributor to tissue selectivity. Specific inhibition of the type 2 enzyme by finasteride
blocks the conversion of testosterone to DHT in the prostate.
Several approaches might make use of the potential tissue-specific conversion to develop SARMs, including:
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, see figures 9 and 10. In vitro assays have shown them both to be selective AR agonists and that they inhibit proliferation of several prostate cancer cell lines. Atraric acid also hinders extracellular matrix invasion and both compounds are able to prevent androgen-induced nuclear translocation of the AR. More potent derivatives are currently being synthesized in hope of improving the pharmacological profile of these two compounds.
Antiandrogen
Antiandrogens, or androgen antagonists, first discovered in the 1960s, prevent androgens from expressing their biological effects on responsive tissues. Antiandrogens alter the androgen pathway by blocking the appropriate receptors, competing for binding sites on the cell's surface, or affecting...
, or androgen
Androgen
Androgen, also called androgenic hormone or testoid, is the generic term for any natural or synthetic compound, usually a steroid hormone, that stimulates or controls the development and maintenance of male characteristics in vertebrates by binding to androgen receptors...
antagonist, was discovered. Antiandrogens antagonise
Receptor antagonist
A receptor antagonist is a type of receptor ligand or drug that does not provoke a biological response itself upon binding to a receptor, but blocks or dampens agonist-mediated responses...
the androgen receptor
Androgen receptor
The androgen receptor , also known as NR3C4 , is a type of nuclear receptor that is activated by binding of either of the androgenic hormones testosterone or dihydrotestosterone in the cytoplasm and then translocating into the nucleus...
(AR) and thereby block the biological effects of testosterone
Testosterone
Testosterone is a steroid hormone from the androgen group and is found in mammals, reptiles, birds, and other vertebrates. In mammals, testosterone is primarily secreted in the testes of males and the ovaries of females, although small amounts are also secreted by the adrenal glands...
and dihydrotestosterone
Dihydrotestosterone
Dihydrotestosterone is an androgen or male sex hormone. The enzyme 5α-reductase synthesises DHT in the prostate, testes, hair follicles, and adrenal glands...
(DHT). Antiandrogens are important for men with hormonally responsive diseases like prostate cancer
Prostate cancer
Prostate cancer is a form of cancer that develops in the prostate, a gland in the male reproductive system. Most prostate cancers are slow growing; however, there are cases of aggressive prostate cancers. The cancer cells may metastasize from the prostate to other parts of the body, particularly...
, benign prostatic hyperplasia
Benign prostatic hyperplasia
Benign prostatic hyperplasia also known as benign prostatic hypertrophy , benign enlargement of the prostate , and adenofibromyomatous hyperplasia, refers to the increase in size of the prostate....
(BHP), acne
Acne vulgaris
Acne vulgaris is a common human skin disease, characterized by areas of skin with seborrhea , comedones , papules , pustules , Nodules and possibly scarring...
, seborrhea
Seborrhoeic dermatitis
Seborrhoeic dermatitis is an inflammatory skin disorder affecting the scalp, face, and torso. Typically, seborrheic dermatitis presents with scaly, flaky, itchy, and red skin. It particularly affects the sebaceous-gland-rich areas of skin...
, hirsutism
Hirsutism
Hirsutism or frazonism is the excessive hairiness on women in those parts of the body where terminal hair does not normally occur or is minimal - for example, a beard or chest hair. It refers to a male pattern of body hair and it is therefore primarily of cosmetic and psychological concern...
and androgen alopecia
Alopecia areata
Alopecia areata is a medical condition in which hair is lost from some or all areas of the body, usually from the scalp. Because it causes bald spots on the scalp, especially in the first stages, it is sometimes called spot baldness. In 1–2% of cases, the condition can spread to the entire scalp ...
. Antiandrogens are mainly used for the treatment of prostate diseases. New research also suggests that ARs could be linked to the disease progression of triple-negative breast cancer and that antiandrogens can potentially be used to treat it.
Current antiandrogens are small molecule
Small molecule
In the fields of pharmacology and biochemistry, a small molecule is a low molecular weight organic compound which is by definition not a polymer...
antiandrogens and can be either steroid
Steroid
A steroid is a type of organic compound that contains a characteristic arrangement of four cycloalkane rings that are joined to each other. Examples of steroids include the dietary fat cholesterol, the sex hormones estradiol and testosterone, and the anti-inflammatory drug dexamethasone.The core...
al or nonsteroidal depending on ligand
Ligand
In coordination chemistry, a ligand is an ion or molecule that binds to a central metal atom to form a coordination complex. The bonding between metal and ligand generally involves formal donation of one or more of the ligand's electron pairs. The nature of metal-ligand bonding can range from...
chemistry. Steroidal antiandrogens share a similar steroid structure, while nonsteroidal antiandrogens may have structurally distinctive pharmacophores. Only a limited number of compounds are available for clinic
Clinic
A clinic is a health care facility that is primarily devoted to the care of outpatients...
al use despite the fact that a very large variety of antiandrogen compounds have been discovered and researched.
History
At the beginning of the twentieth century, a relationship between the pituitaryPituitary gland
In vertebrate anatomy the pituitary gland, or hypophysis, is an endocrine gland about the size of a pea and weighing 0.5 g , in humans. It is a protrusion off the bottom of the hypothalamus at the base of the brain, and rests in a small, bony cavity covered by a dural fold...
, testes
Testicle
The testicle is the male gonad in animals. Like the ovaries to which they are homologous, testes are components of both the reproductive system and the endocrine system...
and prostate gland
Prostate
The prostate is a compound tubuloalveolar exocrine gland of the male reproductive system in most mammals....
was established. Charles Brenton Huggins
Charles Brenton Huggins
Charles Brenton Huggins was a Canadian-born American physician and physiologist and cancer researcher at the University of Chicago specializing in prostate cancer. He and Peyton Rous were awarded the 1966 Nobel Prize for Physiology or Medicine for discovering that hormones could be used to control...
established a method to measure the effect hormone
Hormone
A hormone is a chemical released by a cell or a gland in one part of the body that sends out messages that affect cells in other parts of the organism. Only a small amount of hormone is required to alter cell metabolism. In essence, it is a chemical messenger that transports a signal from one...
changes have on prostatic function
Function (biology)
A function is part of an answer to a question about why some object or process occurred in a system that evolved through a process of selection. Thus, function refers forward from the object or process, along some chain of causation, to the goal or success...
. He found out that castration
Castration
Castration is any action, surgical, chemical, or otherwise, by which a male loses the functions of the testicles or a female loses the functions of the ovaries.-Humans:...
or estrogen
Estrogen
Estrogens , oestrogens , or œstrogens, are a group of compounds named for their importance in the estrous cycle of humans and other animals. They are the primary female sex hormones. Natural estrogens are steroid hormones, while some synthetic ones are non-steroidal...
administration led to glandular atrophy
Atrophy
Atrophy is the partial or complete wasting away of a part of the body. Causes of atrophy include mutations , poor nourishment, poor circulation, loss of hormonal support, loss of nerve supply to the target organ, disuse or lack of exercise or disease intrinsic to the tissue itself...
, which could be reversed by re-administration of androgen. In 1941 the beneficial effect of androgen ablation on metastatic prostate cancer was realised when Huggins and Clarence Hodges treated these patients by either castration or estrogen therapy. They monitored the prostate size and therapeutic efficacy by measuring serum acid-phosphatase
Prostatic acid phosphatase
Prostatic acid phosphatase , also prostatic specific acid phosphatase , is an enzyme produced by the prostate. It may be found in increased amounts in men who have prostate cancer or other diseases....
levels. They concluded that androgenic activity in the body influences prostate cancer at least with respect to serum phosphatase. Huggins was the first to use a systemic approach to treat prostate cancer. He was awarded the Nobel Prize in Physiology or Medicine
Nobel Prize in Physiology or Medicine
The Nobel Prize in Physiology or Medicine administered by the Nobel Foundation, is awarded once a year for outstanding discoveries in the field of life science and medicine. It is one of five Nobel Prizes established in 1895 by Swedish chemist Alfred Nobel, the inventor of dynamite, in his will...
in 1966.
It became evident after many findings that androgen ablation, by means of castration or estrogen administration, was not enough to completely cure patients with advanced prostate cancer. In the late 1960s, the AR was discovered and characterised. Screening of chemical libraries
Chemical library
A chemical library or compound library is a collection of stored chemicals usually used ultimately in high-throughput screening or industrial manufacture. The chemical library can consist in simple terms of a series of stored chemicals...
for AR blockers led to the discovery of cyproterone. An acetate
Acetate
An acetate is a derivative of acetic acid. This term includes salts and esters, as well as the anion found in solution. Most of the approximately 5 billion kilograms of acetic acid produced annually in industry are used in the production of acetates, which usually take the form of polymers. In...
group was then added to cyproterone creating cyproterone acetate
Cyproterone
Cyproterone acetate is an antiandrogen...
. In the 1970s, flutamide
Flutamide
Flutamide is an oral nonsteroidal antiandrogen drug primarily used to treat prostate cancer. It competes with testosterone and its powerful metabolite, dihydrotestosterone for binding to androgen receptors in the prostate gland. By doing so, it prevents them from stimulating the prostate cancer...
was discovered. The United States Food and Drug Administration
Food and Drug Administration
The Food and Drug Administration is an agency of the United States Department of Health and Human Services, one of the United States federal executive departments...
(FDA) approved it in 1989 for use in treating prostate cancer. In 1995, bicalutamide
Bicalutamide
Bicalutamide is an oral non-steroidal anti-androgen used in the treatment of prostate cancer and hirsutism...
was also approved and nilutamide
Nilutamide
Nilutamide is an antiandrogen medication used in the treatment of advanced stage prostate cancer. Nilutamide blocks the androgen receptor, preventing its interaction with testosterone. Because most prostate cancer cells rely on the stimulation of the androgen receptor for growth and survival,...
followed a year later.
Androgen receptor
The AR belongs to the steroid receptorSteroid hormone receptor
Steroid hormone receptors are found on the plasma membrane, in the cytosol and also in the nucleus of target cells. They are generally intracellular receptors and initiate signal transduction for steroid hormones which lead to changes in gene expression over a time period of hours to days...
subfamily of the nuclear receptor
Nuclear receptor
In the field of molecular biology, nuclear receptors are a class of proteins found within cells that are responsible for sensing steroid and thyroid hormones and certain other molecules...
superfamily. Its function is regulated by the binding of androgens, which initiates sequential conformation changes
Conformational change
A macromolecule is usually flexible and dynamic. It can change its shape in response to changes in its environment or other factors; each possible shape is called a conformation, and a transition between them is called a conformational change...
of the receptor
Receptor (biochemistry)
In biochemistry, a receptor is a molecule found on the surface of a cell, which receives specific chemical signals from neighbouring cells or the wider environment within an organism...
that affects receptor-protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...
and receptor-DNA interactions. Endogenous androgens are mainly testosterone and DHT.
The AR gene
Gene
A gene is a molecular unit of heredity of a living organism. It is a name given to some stretches of DNA and RNA that code for a type of protein or for an RNA chain that has a function in the organism. Living beings depend on genes, as they specify all proteins and functional RNA chains...
is more than 90kb long and codes for a protein of 919 amino acid
Amino acid
Amino acids are molecules containing an amine group, a carboxylic acid group and a side-chain that varies between different amino acids. The key elements of an amino acid are carbon, hydrogen, oxygen, and nitrogen...
s. Only one AR gene has been identified in humans which is located on chromosome X
X chromosome
The X chromosome is one of the two sex-determining chromosomes in many animal species, including mammals and is common in both males and females. It is a part of the XY sex-determination system and X0 sex-determination system...
. It comprises four main regions, see figure 1:
- N-terminal domain (NTD) which serves a modulatory function.
- DNA-binding domainDNA-binding domainA DNA-binding domain is an independently folded protein domain that contains at least one motif that recognizes double- or single-stranded DNA. A DBD can recognize a specific DNA sequence or have a general affinity to DNA...
(DBD) which recognises and binds to androgen response elements (ARE) in target gene sequence. - Ligand binding domain (LBD) which is responsible for ligand recognition and binding.
- A small hinge region between the DBD and LBD.
Two functions have been identified in AR that have critical roles in the regulation of target gene transactivation
Transactivation
In molecular biology and genetics, transactivation is an increased rate of gene expression triggered either by biological processes or by artificial means.- Natural transactivation :...
, the N-terminal activation function 1 (AF1) and the C-terminal activation function 2 (AF2). AF1 is ligand-independent and plays the primary role in target gene transactivation. The AF2 is a ligand-dependent and only shows limited function.
Mechanism of action
Unbound AR is mainly located in the cytoplasmCytoplasm
The cytoplasm is a small gel-like substance residing between the cell membrane holding all the cell's internal sub-structures , except for the nucleus. All the contents of the cells of prokaryote organisms are contained within the cytoplasm...
, like a typical steroid receptor, and is associated with a complex of heat shock protein
Heat shock protein
Heat shock proteins are a class of functionally related proteins involved in the folding and unfolding of other proteins. Their expression is increased when cells are exposed to elevated temperatures or other stress. This increase in expression is transcriptionally regulated...
s (HSP) through interactions with LBD. Androgens, either agonist
Agonist
An agonist is a chemical that binds to a receptor of a cell and triggers a response by that cell. Agonists often mimic the action of a naturally occurring substance...
s or antagonists, position themselves in the ligand-binding pocket (LBP) of the cytosol
Cytosol
The cytosol or intracellular fluid is the liquid found inside cells, that is separated into compartments by membranes. For example, the mitochondrial matrix separates the mitochondrion into compartments....
ic AR and bind to the LBD, see figure 2. The AR goes through a series of conformational
Conformational isomerism
In chemistry, conformational isomerism is a form of stereoisomerism in which the isomers can be interconverted exclusively by rotations about formally single bonds...
changes and HSP dissociate from AR. The transformed AR undergoes dimer
Dimer
A dimer is a chemical entity consisting of two structurally similar subunits called monomers joined by bonds that can be either strong or weak.- Organic chemistry :...
isation, phosphorylation
Phosphorylation
Phosphorylation is the addition of a phosphate group to a protein or other organic molecule. Phosphorylation activates or deactivates many protein enzymes....
and translocates to the nucleus. The translocated receptor then binds to the androgen-response elements (ARE) on the promoter of the androgen responsive gene, a consensus sequence located either upstream or downstream of the transcription start site (TSS) of AR target genes. Recruitment of other transcription
Transcription (genetics)
Transcription is the process of creating a complementary RNA copy of a sequence of DNA. Both RNA and DNA are nucleic acids, which use base pairs of nucleotides as a complementary language that can be converted back and forth from DNA to RNA by the action of the correct enzymes...
co-factors (including co-activators and co-repressors) and general transcriptional machinery further ensures the transactivation of AR-regulated gene expression
Gene expression
Gene expression is the process by which information from a gene is used in the synthesis of a functional gene product. These products are often proteins, but in non-protein coding genes such as ribosomal RNA , transfer RNA or small nuclear RNA genes, the product is a functional RNA...
. All these complicated processes are initiated by the ligand-induced conformational changes in the LBD. Ligand specific recruitment of coregulators
Transcription coregulator
In molecular biology and genetics, transcription coregulators are proteins that interact with transcription factors to either activate or repress the transcription of specific genes. Transcription coregulators that activate gene transcription are referred to as coactivators while those that...
might be crucial for the agonist or antagonist activity of AR ligands. Binding of DNA is also required for AR-regulated gene expression, also known as classic genomic gene function of AR.
Development of steroidal- and non-steroidal antiandrogens
Cyproterone is a steroidal antiandrogen that competitively inhibits the binding of testosterone or DHT to AR. Cyproterone binds to ARs that are expressed by prostate cancer cells as well as to the AR that are expressed in the hypothalamusHypothalamus
The Hypothalamus is a portion of the brain that contains a number of small nuclei with a variety of functions...
and pituitary. Therefore cyproterone blocks the negative feedback
Negative feedback
Negative feedback occurs when the output of a system acts to oppose changes to the input of the system, with the result that the changes are attenuated. If the overall feedback of the system is negative, then the system will tend to be stable.- Overview :...
of androgens at the hypothalamic-pituitary level leading to increased luteinizing hormone
Luteinizing hormone
Luteinizing hormone is a hormone produced by the anterior pituitary gland. In females, an acute rise of LH called the LH surge triggers ovulation and development of the corpus luteum. In males, where LH had also been called interstitial cell-stimulating hormone , it stimulates Leydig cell...
(LH) serum levels. This rise in LH levels causes an increase in serum testosterone levels and ultimately diminishes the ability of cyproterone to compete for AR binding and to block androgenic stimulation
Stimulation
Stimulation is the action of various agents on nerves, muscles, or a sensory end organ, by which activity is evoked; especially, the nervous impulse produced by various agents on nerves, or a sensory end organ, by which the part connected with the nerve is thrown into a state of activity.The word...
.
Cyproterone acetate was developed to overcome this problem. It is formed by adding an acetate group to cyproterone, see figure 3. Cyproterone acetate has a dual mode of action as it competes directly with DHT for binding to AR, but also inhibits gonadotropin
Gonadotropin
Gonadotropins are protein hormones secreted by gonadotrope cells of the pituitary gland of vertebrates. This is a family of proteins, which include the mammalian hormones follitropin , lutropin , placental chorionic gonadotropins hCG and eCG and chorionic gonadotropin , as well as at least two...
secretion. It thereby reduces androgen, estrogen and LH levels. Cyproterone acetate acts both directly as an antiandrogen in prostate cancer cells and also functions to indirectly decrease serum testosterone levels. The latter causes the limitations of cyproterone acetate, which are central effects on androgen secretion, with subsequent loss of libido
Libido
Libido refers to a person's sex drive or desire for sexual activity. The desire for sex is an aspect of a person's sexuality, but varies enormously from one person to another, and it also varies depending on circumstances at a particular time. A person who has extremely frequent or a suddenly...
and sexual potency. Several reports also state that cyproterone acetate causes liver hyperplasia
Hyperplasia
Hyperplasia means increase in number of cells/proliferation of cells. It may result in the gross enlargement of an organ and the term is sometimes mixed with benign neoplasia/ benign tumor....
. These side effect
Side effect
In medicine, a side effect is an effect, whether therapeutic or adverse, that is secondary to the one intended; although the term is predominantly employed to describe adverse effects, it can also apply to beneficial, but unintended, consequences of the use of a drug.Occasionally, drugs are...
s gave pharmaceutical companies the incentive to search for alternative non-steroidal "pure" antiandrogens that would not have these side effects. Pure antiandrogens block the androgen receptor without exerting any agonistic or any other hormonal activity.
Flutamide became the first non-steroidal antiandrogen to be tested clinically. Later the non-steroidal antiandrogens bicalutamide and nilutamide were developed. The alleged advantages of these compounds were that they did not affect libido or potency like the other centrally acting compounds under development, luteinizing-hormone-releasing hormone
Gonadotropin-releasing hormone
Gonadotropin-releasing hormone , also known as Luteinizing-hormone-releasing hormone and luliberin, is a tropic peptide hormone responsible for the release of follicle-stimulating hormone and luteinizing hormone from the anterior pituitary. GnRH is synthesized and released from neurons within...
(LHRH) agonists and cyproterone acetate. But this theory did not prove to be true. These non-steroidal antiandrogens eventually crossed the blood-brain barrier
Blood-brain barrier
The blood–brain barrier is a separation of circulating blood and the brain extracellular fluid in the central nervous system . It occurs along all capillaries and consists of tight junctions around the capillaries that do not exist in normal circulation. Endothelial cells restrict the diffusion...
, like cyproterone, leading to a subsequent increase in serum testosterone levels.
Flutamide
Flutamide is an arylpropionamide analog with pure antiandrogenic properties, seen in figure 4. It is completely absorbed from the gastrointestinal tract after oral administration and undergoes extensive first pass metabolismFirst pass effect
The first-pass effect is a phenomenon of drug metabolism whereby the concentration of a drug is greatly reduced before it reaches the systemic circulation. It is the fraction of lost drug during the process of absorption which is generally related to the liver and gut wall...
to its active form, 2-hydroxyflutamide, and hydrolysis
Hydrolysis
Hydrolysis is a chemical reaction during which molecules of water are split into hydrogen cations and hydroxide anions in the process of a chemical mechanism. It is the type of reaction that is used to break down certain polymers, especially those made by condensation polymerization...
product, 3-trifluoromethyl-4-nitroaniline. Hydroxyflutamide is a more potent AR antagonist than flutamide in vivo
In vivo
In vivo is experimentation using a whole, living organism as opposed to a partial or dead organism, or an in vitro controlled environment. Animal testing and clinical trials are two forms of in vivo research...
, with higher binding affinity for the AR. Hydroxyflutamide has an elimination half-life
Half-life
Half-life, abbreviated t½, is the period of time it takes for the amount of a substance undergoing decay to decrease by half. The name was originally used to describe a characteristic of unstable atoms , but it may apply to any quantity which follows a set-rate decay.The original term, dating to...
of about 8 hours in humans. Hydrolysis of the amide bond
Peptide bond
This article is about the peptide link found within biological molecules, such as proteins. A similar article for synthetic molecules is being created...
represents the major metabolic pathway
Metabolic pathway
In biochemistry, metabolic pathways are series of chemical reactions occurring within a cell. In each pathway, a principal chemical is modified by a series of chemical reactions. Enzymes catalyze these reactions, and often require dietary minerals, vitamins, and other cofactors in order to function...
for this active metabolite
Metabolite
Metabolites are the intermediates and products of metabolism. The term metabolite is usually restricted to small molecules. A primary metabolite is directly involved in normal growth, development, and reproduction. Alcohol is an example of a primary metabolite produced in large-scale by industrial...
. By reversing the stimulatory effect of DHT on ventral prostate weight, flutamide is approximately 2-fold more potent
Potency (pharmacology)
In the field of pharmacology, potency is a measure of drug activity expressed in terms of the amount required to produce an effect of given intensity. A highly potent drug evokes a larger response at low concentrations, while a drug of lower potency evokes a small response at low concentrations...
than cyproterone acetate. Hydroxyflutamide has relatively low binding affinity to AR and is therefore generally used at high doses in order to achieve complete AR blockage in therapy.
Nilutamide
Nilutamide is a nitroaromatic hydantoinHydantoin
Hydantoin, which is also known as glycolylurea, is a heterocyclic organic compound that can be thought of as a cyclic "double-condensation reaction" product of glycolic acid and urea...
analog of flutamide, as seen in figure 5. Nilutamide is eliminated exclusively by metabolism, mainly by reduction of the aromatic
Aromaticity
In organic chemistry, Aromaticity is a chemical property in which a conjugated ring of unsaturated bonds, lone pairs, or empty orbitals exhibit a stabilization stronger than would be expected by the stabilization of conjugation alone. The earliest use of the term was in an article by August...
nitro group
Nitro compound
Nitro compounds are organic compounds that contain one or more nitro functional groups . They are often highly explosive, especially when the compound contains more than one nitro group and is impure. The nitro group is one of the most common explosophores used globally...
. Although the hydrolysis of one of the carbonyl
Carbonyl
In organic chemistry, a carbonyl group is a functional group composed of a carbon atom double-bonded to an oxygen atom: C=O. It is common to several classes of organic compounds, as part of many larger functional groups....
functions of the imidazolinedione was identified, it is much less susceptible to hepatic metabolism than the amide bond in hydroxuflutamide. This results in a longer half-life of nilutamide in humans of 2 days. Nevertheless, the nitro anion-free radical
Radical (chemistry)
Radicals are atoms, molecules, or ions with unpaired electrons on an open shell configuration. Free radicals may have positive, negative, or zero charge...
formed during nitro reduction could still be associated with hepatotoxicity
Hepatotoxicity
Hepatotoxicity implies chemical-driven liver damage.The liver plays a central role in transforming and clearing chemicals and is susceptible to the toxicity from these agents. Certain medicinal agents, when taken in overdoses and sometimes even when introduced within therapeutic ranges, may injure...
in humans, especially when using relatively high dosage employed for androgen blockage. Nilutamide causes side-effects which limit its usage, such as pneumonitis
Pneumonitis
Pneumonitis or pulmonitis is a general term that refers to inflammation of lung tissue.Pneumonia is pneumonitis combined with consolidation and exudation...
and delayed adaption to darkness.
Bicalutamide
BicalutamideBicalutamide
Bicalutamide is an oral non-steroidal anti-androgen used in the treatment of prostate cancer and hirsutism...
is an arylpropionamide analog, seen in figure 6. It has replaced flutamide and nilutamide as the first choice antiandrogen for prostate cancer treatment. Bicalutamide is not as hepatotoxic as flutamide and nilutamide and has a longer half-life, of 6 days in humans, that allows once a day administration at lower dosage. Bicalutamide shares the amide bond structure with flutamide. Even so, the amide bond hydrolysis was discovered in rats, not in humans, which could explain the prolonged half life of bicalutamide in humans.
Bicalutamide has a cyano group
Cyanide
A cyanide is a chemical compound that contains the cyano group, -C≡N, which consists of a carbon atom triple-bonded to a nitrogen atom. Cyanides most commonly refer to salts of the anion CN−. Most cyanides are highly toxic....
at the para position instead of a nitro group like flutamide and nilutamide. This change in groups avoids the nitro reduction observed in nilutamide. Bicalutamide has a chiral carbon in its structure (labeled with an asterisk in figure 6), which is connected to the hydroxyl and methyl groups . It is therefore administered as a racemate. Post-approval investigation revealed that its antiandrogenic activity resides almost entirely in the (R)-enantiomer
Enantiomer
In chemistry, an enantiomer is one of two stereoisomers that are mirror images of each other that are non-superposable , much as one's left and right hands are the same except for opposite orientation. It can be clearly understood if you try to place your hands one over the other without...
. (R)-bicalutamide has an almost fourfold higher affinity for the prostate AR than hydroxyflutamide and has a better side-effect profile compared to other antiandrogens.
Steroidal antiandrogens
Cyproterone acetate is a 6-chloro-1,2-methylene derivative of 17 α-acetoxyprogesterone. It shows major antiandrogenic activity together with androgenic activities. Cyproterone acetate displays high affinity for AR in rats which increases when the 1,2-methylene group is removed from the compound. If the chlorineChlorine
Chlorine is the chemical element with atomic number 17 and symbol Cl. It is the second lightest halogen, found in the periodic table in group 17. The element forms diatomic molecules under standard conditions, called dichlorine...
atome is replaced by a methyl group
Methyl group
Methyl group is a functional group derived from methane, containing one carbon atom bonded to three hydrogen atoms —CH3. The group is often abbreviated Me. Such hydrocarbon groups occur in many organic compounds. The methyl group can be found in three forms: anion, cation and radical. The anion...
the binding slightly decreases, whereas further removal of the C6 double bond modifies the binding kinetics, see figure 7.
Non-steroidal antiandrogens
Hydroxyflutamide and it‘s analogs, bicalutamide and nilutamide, share an anilide ring structure. The structures can be seen in figure 7, where the anilide ring is coloured red. These three compounds require an electron-deficient aromatic ring for efficient AR binding. Replacing the anilide with an alkeneAlkene
In organic chemistry, an alkene, olefin, or olefine is an unsaturated chemical compound containing at least one carbon-to-carbon double bond...
gives weakly active compounds, which can be attributed to the lack of intramolecular hydrogen binding
Hydrogen bond
A hydrogen bond is the attractive interaction of a hydrogen atom with an electronegative atom, such as nitrogen, oxygen or fluorine, that comes from another molecule or chemical group. The hydrogen must be covalently bonded to another electronegative atom to create the bond...
or to poor hydrogen-bond donor capability. Various combinations of electron-withdrawing
Polar effect
The Polar effect or electronic effect in chemistry is the effect exerted by a substituent on modifying electrostatic forces operating on a nearby reaction center...
substitutions in the aniline
Aniline
Aniline, phenylamine or aminobenzene is an organic compound with the formula C6H5NH2. Consisting of a phenyl group attached to an amino group, aniline is the prototypical aromatic amine. Being a precursor to many industrial chemicals, its main use is in the manufacture of precursors to polyurethane...
ring of these drugs, have not shown higher binding to the AR receptor than compounds which have a chloro or trifluoromethyl group at the meta position (R1) and either a cyano or nitrogen group at the para position (R2).
For hydroxyflutamide, a group of compounds that differed in the aromatic ring did not bind to the AR. This suggests that the bisubstitution in the hydroxyflutamide ring is essential for high AR binding affinity. It has also been demonstrated that hydroxyflutamide requires the strong hydrogen bond donor ability of the tertiary hydroxyl group and fixed conformers involved in intramolecular hydrogen binding, to bind effectively to AR.
For bicalutamide, the antiandrogenic activities of sulfide
Sulfide
A sulfide is an anion of sulfur in its lowest oxidation state of 2-. Sulfide is also a slightly archaic term for thioethers, a common type of organosulfur compound that are well known for their bad odors.- Properties :...
and sulfone
Sulfone
A sulfone is a chemical compound containing a sulfonyl functional group attached to two carbon atoms. The central hexavalent sulfur atom is double bonded to each of two oxygen atoms and has a single bond to each of two carbon atoms, usually in two separate hydrocarbon substituents.-IUPAC name and...
substitions of the X-linkage were tested in vitro
In vitro
In vitro refers to studies in experimental biology that are conducted using components of an organism that have been isolated from their usual biological context in order to permit a more detailed or more convenient analysis than can be done with whole organisms. Colloquially, these experiments...
. The sulfides showed in most cases at least 2-fold higher binding affinity than corresponding sulfones. However, this relationship was reversed when the R3 group was NHSO2CH3, where the binding affinity of sulfone was 3-fold higher than that of sulfide. These results indicate that substituents of the B-ring largely determine the effect of the X-linkage in AR binding. Researchers have proposed that the tertiary hydroxyl group is involved in direct interaction with AR because when an acetyl
Acetyl
In organic chemistry, acetyl is a functional group, the acyl with chemical formula COCH3. It is sometimes represented by the symbol Ac . The acetyl group contains a methyl group single-bonded to a carbonyl...
group is introduced to that hydroxyl moiety, the receptor binding affinity greatly decreases.
Nilutamide has very low affinity for AR when tested on castrated rat prostate. Modifications such as replacing the N3 atom with oxygen has little effect on affinity of the compound for prostate AR. By replacing the oxygen
Oxygen
Oxygen is the element with atomic number 8 and represented by the symbol O. Its name derives from the Greek roots ὀξύς and -γενής , because at the time of naming, it was mistakenly thought that all acids required oxygen in their composition...
atom with a sulfur
Sulfur
Sulfur or sulphur is the chemical element with atomic number 16. In the periodic table it is represented by the symbol S. It is an abundant, multivalent non-metal. Under normal conditions, sulfur atoms form cyclic octatomic molecules with chemical formula S8. Elemental sulfur is a bright yellow...
atom at the C2 position of the imidazole
Imidazole
Imidazole is an organic compound with the formula C3H4N2. This aromatic heterocyclic is a diazole and is classified as an alkaloid. Imidazole refers to the parent compound, whereas imidazoles are a class of heterocycles with similar ring structure, but varying substituents...
ring and adding butylalcohol
Butanol
Butanol or butyl alcohol can refer to any of the four isomeric alcohols of formula C4H9OH:*n-Butanol, butan-1-ol, 1-butanol, n-butyl alcohol;*Isobutanol, 2-methylpropan-1-ol, isobutyl alcohol;...
to the N3 atom, the receptor binding and biological activity of the compound increases 100 times that of non-steroidal antiandrogens. Also the compound does not bind to other steroid receptors. If a methyl group is changed for the butylalcohol group, the compound shows 3 and 10 times more antiandrogenic activity in vivo than bicalutamide and nilutamide, respectively.
Antiandrogen withdrawal syndrome
Antiandrogens that are currently on the market are particularly useful for the treatment of prostate cancer during the early stages. However, prostate cancer often progresses to a hormone-refractory state in which the cancer progresses in the presence of continued androgen ablation or antiandrogen therapy. This suggests that long term use of these antiandrogens during prostate cancer can lead to the development of androgen-independent prostate cancer cells or the ability of adrenal androgens to support tumorTumor
A tumor or tumour is commonly used as a synonym for a neoplasm that appears enlarged in size. Tumor is not synonymous with cancer...
growth. This phenomenon is called antiandrogen withdrawal syndrome (AWS) and is one of the major drawbacks of existing antiandrogens. AWS is defined as tumor regression or symptomatic relief observed upon discontinuation of the antiandrogen therapy. The mechanism for this is not fully understood but current theories include alterations of the AR gene, coregulator proteins and/or signal transduction pathways. This antiandrogen resistance may also be linked to the relative weakness of current antiandrogens as they have an affinity 50 times or more lower than that of DHT for the AR. This may also explain why compensatory AR overexpression is often observed.
Androgen receptor gene mutations
AR gene mutationMutation
In molecular biology and genetics, mutations are changes in a genomic sequence: the DNA sequence of a cell's genome or the DNA or RNA sequence of a virus. They can be defined as sudden and spontaneous changes in the cell. Mutations are caused by radiation, viruses, transposons and mutagenic...
s in the LBD that alter ligand specificity and/or functional activity exist and are thought to contribute to the conversion of some AR antagonists into agonists, which explains the paradoxical temporary improvement sometimes observed in patients when antiandrogen therapy is stopped. These mutations can have great affect on the antagonist activities of current small molecule antiandrogens and make them less efficient in blocking AR function via indirect modulation from inside of the LBP. Recent studies with circulating tumor cells, suggest that the mutation frequency is higher than previously assumed based on tumor biopsies
Biopsy
A biopsy is a medical test involving sampling of cells or tissues for examination. It is the medical removal of tissue from a living subject to determine the presence or extent of a disease. The tissue is generally examined under a microscope by a pathologist, and can also be analyzed chemically...
.
The T877A, W741L and W741C mutations are examples of known AR LBD mutations. The LNCaP
Lncap
LNCaP cells are a cell line of human cells commonly used in the field of oncology. LNCaP cells are androgen-sensitive human prostate adenocarcinoma cells derived from the left supraclavicular lymph node metastasis from a 50-year-old caucasian male in 1977...
prostate cancer cell line
Cell culture
Cell culture is the complex process by which cells are grown under controlled conditions. In practice, the term "cell culture" has come to refer to the culturing of cells derived from singlecellular eukaryotes, especially animal cells. However, there are also cultures of plants, fungi and microbes,...
expresses AR with a T877A point mutation that causes proliferation in the presence of the antiandrogens hydroxyflutamide and cyproterone acetate. This mutation has also been discovered in patients with antiandrogen withdrawal syndrome being treated with these compounds. In another study, bicalutamide treatment of LNCaP cells resulted in two LBD mutations, W741L and W741C, causing bicalutamide to acquire agonist activity to both mutant ARs. The W741L mutation generates additional space such that the sulfonyl-linked phenyl
Phenyl group
In organic chemistry, the phenyl group or phenyl ring is a cyclic group of atoms with the formula C6H5. Phenyl groups are closely related to benzene. Phenyl groups have six carbon atoms bonded together in a hexagonal planar ring, five of which are bonded to individual hydrogen atoms, with the...
ring of bicalutamide is accommodated at the location of the missing indole
Indole
Indole is an aromatic heterocyclic organic compound. It has a bicyclic structure, consisting of a six-membered benzene ring fused to a five-membered nitrogen-containing pyrrole ring. Indole is a popular component of fragrances and the precursor to many pharmaceuticals. Compounds that contain an...
ring of W741. In non-mutant AR, the presence of the W741 side chain probably forces bicalutamide to protrude out thus precluding the active position of H12 on the AR receptor.
However, hydroxyflutamide worked as an antagonist for W741 mutant ARs. This concurs with the theory that flutamide and nilutamide antagonize AR through the mechanism of “passive antagonism”, as they are of a more modest size then bicalutamide. These drugs may therefore be effective as a second-line therapy for refractory prostate cancer previously treated with bicalutamide.
Peptide antiandrogens
Peptide antiandrogens have been proposed to overcome the limitations of current antiandrogens regarding mutant ARs, by directly blocking AR function from protein surface, outside of the LBP. This direct blockade is thought to provide a more efficient strategy to avoid or overcome abnormal AR action during AWS, as well as allowing for more flexibility in structural modification without the space limitations of the rigid LBP.Steroid receptors have similarities in terms of gene sequences and protein structures, leading often to functional crosstalk among steroid receptors. One of the criteria for AR peptide antagonists is to achieve high degree of specificity for the AR. It is though important to realize that AR specificity does not necessarily translate in vivo, since peptide antagonists may also interact with protein targets other than AR.
Ligand binding domain as target site
AR activation requires the formation of a functional activation function 2 (AF2) region in AR LBD that mediates the interactions between AR and various transcription cofactorsCofactor (biochemistry)
A cofactor is a non-protein chemical compound that is bound to a protein and is required for the protein's biological activity. These proteins are commonly enzymes, and cofactors can be considered "helper molecules" that assist in biochemical transformations....
. Therefore most of the research on peptide AR antagonists focuses on peptide
Peptide
Peptides are short polymers of amino acid monomers linked by peptide bonds. They are distinguished from proteins on the basis of size, typically containing less than 50 monomer units. The shortest peptides are dipeptides, consisting of two amino acids joined by a single peptide bond...
s that may directly block the AF2 in AR LBD from protein surface. Even in bound mutant AR, peptide antagonists would be able to block the AF2 function via direct surface interaction, regardless of the ligand bound.
Research on these peptide antagonists are usually carried out by affinity screening of phage display
Phage display
Phage display is a method for the study of protein–protein, protein–peptide, and protein–DNA interactions that uses bacteriophages to connect proteins with the genetic information that encodes them. Phage Display was originally invented by George P...
libraries that express random peptides containing various signature motifs
Sequence motif
In genetics, a sequence motif is a nucleotide or amino-acid sequence pattern that is widespread and has, or is conjectured to have, a biological significance...
. ARs seem to have a distinct preference for ‘FxxLF’ type of binding motifs (where F = phenylalanine
Phenylalanine
Phenylalanine is an α-amino acid with the formula C6H5CH2CHCOOH. This essential amino acid is classified as nonpolar because of the hydrophobic nature of the benzyl side chain. L-Phenylalanine is an electrically neutral amino acid, one of the twenty common amino acids used to biochemically form...
, L = leucine
Leucine
Leucine is a branched-chain α-amino acid with the chemical formula HO2CCHCH2CH2. Leucine is classified as a hydrophobic amino acid due to its aliphatic isobutyl side chain. It is encoded by six codons and is a major component of the subunits in ferritin, astacin and other 'buffer' proteins...
, and X = any amino acid residue), whereas other nuclear receptors have a highly similar binding mechanism for ‘LxxLL’ type of binding motifs. This provides a unique opportunity for the development of AR-specific peptides.
Even though small molecule antagonists and peptide antagonist targeting AF2 surface differ in binding sites, they both inhibit AR function by disrupting AF2 function. Therefore mechanistically, these peptide antagonists may also be classified as ‘AF2 antagonists’.
N-terminal domain as target site
Functionally, AR N-terminal domain (NTD) plays the primary role in regulating target gene transcription activation and mediating various receptor-protein and intra-receptor N-terminal and C-terminal interactions. Therefore modulation of NTD function is considered an efficient strategy to target AR action. Among various functional domains in different nuclear receptors, NTD is the least conserved and so could maybe become the best target site for peptide antagonists to achieve AR specificity. However the structural features of the NTD are undetermined due to a high degree of flexibility in its conformation. Both biochemicalBiochemistry
Biochemistry, sometimes called biological chemistry, is the study of chemical processes in living organisms, including, but not limited to, living matter. Biochemistry governs all living organisms and living processes...
and circular dichroism spectroscopy
Circular dichroism
Circular dichroism refers to the differential absorption of left and right circularly polarized light. This phenomenon was discovered by Jean-Baptiste Biot, Augustin Fresnel, and Aimé Cotton in the first half of the 19th century. It is exhibited in the absorption bands of optically active chiral...
analysis suggest that AR NTD is highly disordered under native conditions, making it a difficult target for drug discovery.
In 2008 there were reports of a chlorinated peptide, Sintokamide A, isolated from marine sponges that effectively inhibits AR N-terminal domain-activated reporter gene transcription, see figure 8. The evidence presented was not sufficient enough to support the conclusion that Sintokamide A directly inhibits the function of AR NTD, and the mechanism of action needs further investigation.
Selective androgen receptor modulators
Small molecule antiandrogens that are available today have undesirable side effects caused by complete, non-selective inhibition of AR action. To minimize these side effects, a new class of tissue selective androgen receptor modulatorSelective androgen receptor modulator
Selective androgen receptor modulators or SARMs are a novel class of androgen receptor ligands. They are intended to have the same kind of effects as androgenic drugs like anabolic steroids but be much more selective in...
s (SARMs) has been proposed as a novel approach for the treatment of prostate cancer. These ligands should behave as antagonists in the prostate with either no activity or agonist activity in other target tissues, so as to have little or no effects in the anabolic tissues or central nervous system
Central nervous system
The central nervous system is the part of the nervous system that integrates the information that it receives from, and coordinates the activity of, all parts of the bodies of bilaterian animals—that is, all multicellular animals except sponges and radially symmetric animals such as jellyfish...
(CNS). However discovering this new class of ligands might be challenging because the molecular mechanism of AR action is not well understood.
Several mechanisms have been proposed to achieve this tissue selectivity of AR ligands. The most definitive evidence exists for the role of 5-alpha reductase
5-alpha reductase
5α-reductases, also known as 3-oxo-5α-steroid 4-dehydrogenases, are enzymes involved in steroid metabolism. They participate in 3 metabolic pathways: bile acid biosynthesis, androgen and estrogen metabolism, and prostate cancer....
. 5-alpha reductase is only expressed in specific tissues and could therefore be a unique contributor to tissue selectivity. Specific inhibition of the type 2 enzyme by finasteride
Finasteride
Finasteride is a synthetic antiandrogen that inhibits type II 5-alpha reductase, the enzyme that converts testosterone to dihydrotestosterone...
blocks the conversion of testosterone to DHT in the prostate.
Several approaches might make use of the potential tissue-specific conversion to develop SARMs, including:
- Inactive parent compounds that are activated by type 2 5-alpha reductase in the prostate to form antiandrogens.
- AR agonists that are inactivated by type 2 5-alpha reductase in the prostate.
- AR agonists that are converted to antiandrogens only by type 2 5-alpha reductase in the prostate.
Other small molecule antiandrogens
The development status of other small molecule antiandrogens undergoing research in 2011 can be seen in table 1.| Name of compound | Structure | Company | Stage of development | Other information | |
|---|---|---|---|---|---|
| RU58642 | Roussel-Uclaf SA Roussel Uclaf Roussel Uclaf S.A. was the second largest French pharmaceutical company before it was acquired by Hoechst AG of Frankfurt, Germany in 1997, with pharmaceutical operations combined into the Hoechst Marion Roussel division... |
Preclinical Pre-clinical development In drug development, pre-clinical development is a stage of research that begins before clinical trials can begin, and during which important feasibility, iterative testing and drug safety data is collected.... – no further developments since 1998 |
Orally active and more potent than current small molecular antiandrogen Antiandrogen Antiandrogens, or androgen antagonists, first discovered in the 1960s, prevent androgens from expressing their biological effects on responsive tissues. Antiandrogens alter the androgen pathway by blocking the appropriate receptors, competing for binding sites on the cell's surface, or affecting... s. |
||
| LG120907 | Ligand Pharmaceuticals | Preclinical | Orally active, strong antagonistic Receptor antagonist A receptor antagonist is a type of receptor ligand or drug that does not provoke a biological response itself upon binding to a receptor, but blocks or dampens agonist-mediated responses... activity in the prostate Prostate The prostate is a compound tubuloalveolar exocrine gland of the male reproductive system in most mammals.... without raising plasma levels of LH Luteinizing hormone Luteinizing hormone is a hormone produced by the anterior pituitary gland. In females, an acute rise of LH called the LH surge triggers ovulation and development of the corpus luteum. In males, where LH had also been called interstitial cell-stimulating hormone , it stimulates Leydig cell... and testosterone Testosterone Testosterone is a steroid hormone from the androgen group and is found in mammals, reptiles, birds, and other vertebrates. In mammals, testosterone is primarily secreted in the testes of males and the ovaries of females, although small amounts are also secreted by the adrenal glands... . |
||
| LG105 | Ligand Pharmaceuticals | Preclinical | Orally available, strong antagonistic activity in the prostate without raising plasma levels of LH and testosterone. Seems to be more potent than LG120907. | ||
| RD162 | Medivation | Preclinical | High binding affinity to AR. Unlike bicalutamide Bicalutamide Bicalutamide is an oral non-steroidal anti-androgen used in the treatment of prostate cancer and hirsutism... , it does not promote nuclear translocation and impairs both DNA binding to androgen Androgen Androgen, also called androgenic hormone or testoid, is the generic term for any natural or synthetic compound, usually a steroid hormone, that stimulates or controls the development and maintenance of male characteristics in vertebrates by binding to androgen receptors... response elements and recruitment of coactivators Coactivator (genetics) A coactivator is a protein that increases gene expression by binding to an activator which contains a DNA binding domain. The coactivator is unable to bind DNA by itself.... . |
||
| MDV3100 MDV3100 MDV3100 is an experimental androgen receptor antagonist drug developed by the pharmaceutical company Medivation for the treatment of castration-resistant prostate cancer currently in Phase 3 clinical trials. Results so far have been encouraging; Medivation has reported up to an 89% decrease in... |
Medivation | Phase III clinical | High binding affinity to AR. Unlike bicalutamide, it does not promote nuclear translocation and impairs both DNA binding to androgen response elements and recruitment of coactivators. Induces tumor cell apoptosis Apoptosis Apoptosis is the process of programmed cell death that may occur in multicellular organisms. Biochemical events lead to characteristic cell changes and death. These changes include blebbing, cell shrinkage, nuclear fragmentation, chromatin condensation, and chromosomal DNA fragmentation... and has not agonist Agonist An agonist is a chemical that binds to a receptor of a cell and triggers a response by that cell. Agonists often mimic the action of a naturally occurring substance... activity. |
||
| BMS-641988 | Bristol-Myers Squibb Bristol-Myers Squibb Bristol-Myers Squibb , often referred to as BMS, is a pharmaceutical company, headquartered in New York City. The company was formed in 1989, following the merger of its predecessors Bristol-Myers and the Squibb Corporation... |
Phase I clinical – trial terminated | Showed increased potency compared to bicalutamide. Phase I trial was discontinued because of an epileptic seizure in a patient. Led to the findings that several antiandrogens produce off-target antagonist binding to GABA-A receptors. | ||
| CH5137291 | Chugai Pharmaceutical Co. Ltd. Chugai Pharmaceutical Co. Chugai Pharmaceutical Co., Ltd. is a subsidiary drug manufacturer operating in Japan controlled by Hoffmann–La Roche, which owns 52% of the company... |
Preclinical | Completely inhibits AR-mediated transactivation and proliferation of the CRPC xenograft model LNCaP-BC2, which is bicalutamide-resistant. |
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Natural antiandrogens
Atraric acid and N-butylbenzenesulfonamide are natural compounds with antiandrogen properties which have been purified from the bark of the African tree Pygeum africanumPrunus africana
Prunus africana, or Red Stinkwood , is an evergreen tree native to the montane regions of Sub-Saharan Africa and the Islands of Madagascar, Sao Tome, Fernando Po and Grande Comore at about 900–3400 m. of altitude. The mature tree is 10–25 m. high, open-branched and often pendulous in forest,...
, see figures 9 and 10. In vitro assays have shown them both to be selective AR agonists and that they inhibit proliferation of several prostate cancer cell lines. Atraric acid also hinders extracellular matrix invasion and both compounds are able to prevent androgen-induced nuclear translocation of the AR. More potent derivatives are currently being synthesized in hope of improving the pharmacological profile of these two compounds.
See also
- AntiandrogenAntiandrogenAntiandrogens, or androgen antagonists, first discovered in the 1960s, prevent androgens from expressing their biological effects on responsive tissues. Antiandrogens alter the androgen pathway by blocking the appropriate receptors, competing for binding sites on the cell's surface, or affecting...
- Androgen receptorAndrogen receptorThe androgen receptor , also known as NR3C4 , is a type of nuclear receptor that is activated by binding of either of the androgenic hormones testosterone or dihydrotestosterone in the cytoplasm and then translocating into the nucleus...
- Prostate cancerProstate cancerProstate cancer is a form of cancer that develops in the prostate, a gland in the male reproductive system. Most prostate cancers are slow growing; however, there are cases of aggressive prostate cancers. The cancer cells may metastasize from the prostate to other parts of the body, particularly...
- CyproteroneCyproteroneCyproterone acetate is an antiandrogen...
- FlutamideFlutamideFlutamide is an oral nonsteroidal antiandrogen drug primarily used to treat prostate cancer. It competes with testosterone and its powerful metabolite, dihydrotestosterone for binding to androgen receptors in the prostate gland. By doing so, it prevents them from stimulating the prostate cancer...
- BicalutamideBicalutamideBicalutamide is an oral non-steroidal anti-androgen used in the treatment of prostate cancer and hirsutism...
- NilutamideNilutamideNilutamide is an antiandrogen medication used in the treatment of advanced stage prostate cancer. Nilutamide blocks the androgen receptor, preventing its interaction with testosterone. Because most prostate cancer cells rely on the stimulation of the androgen receptor for growth and survival,...
- Selective androgen receptor modulatorSelective androgen receptor modulatorSelective androgen receptor modulators or SARMs are a novel class of androgen receptor ligands. They are intended to have the same kind of effects as androgenic drugs like anabolic steroids but be much more selective in...