Date Published: December 25, 2012
Publisher: Hindawi Publishing Corporation
Author(s): Faris Azzouni, Alejandro Godoy, Yun Li, James Mohler.
Despite the discovery of 5 alpha-reduction as an enzymatic step in steroid metabolism in 1951, and the discovery that dihydrotestosterone is more potent than testosterone in 1968, the significance of 5 alpha-reduced steroids in human diseases was not appreciated until the discovery of 5 alpha-reductase type 2 deficiency in 1974. Affected males are born with ambiguous external genitalia, despite normal internal genitalia. The prostate is hypoplastic, nonpalpable on rectal examination and approximately 1/10th the size of age-matched normal glands. Benign prostate hyperplasia or prostate cancer does not develop in these patients. At puberty, the external genitalia virilize partially, however, secondary sexual hair remains sparse and male pattern baldness and acne develop rarely. Several compounds have been developed to inhibit the 5 alpha-reductase isozymes and they play an important role in the prevention and treatment of many common diseases. This review describes the basic biochemical properties, functions, tissue distribution, chromosomal location, and clinical significance of the 5 alpha-reductase isozyme family.
Testosterone (T) is the most abundant androgen in serum. Approximately 97% of T is bound to albumen and sex-hormone binding globulin and the remaining 3% is free and biologically active. T is synthesized by the Leydig cells of the testes under the control of the hypothalamus and anterior pituitary gland. In male fetuses, T stimulates the differentiation of the Wolffian duct into male internal genitalia (epididymis, vas deferens, and seminal vesicles) and development of libido, enlargement of the vocal cords, skeletal muscles, penis, and scrotum and the initiation of spermatogenesis at puberty [1, 2]. T is taken from circulation to cells through processes that remain poorly understood. Intracellular T is converted to dihydrotestosterone (DHT), the preferred ligand for androgen receptor (AR) transactivation, by the enzyme 5 alpha-reductase (5α-R). Upon ligand binding and transactivation, the DHT-AR complex translocates from cytoplasm to nucleus and activates the transcription of certain genes (the androgen receptor-regulated genes, ARRG).
Steroids are a special type of lipid. The backbone of steroids is the compound “gonane”, a 17-carbon molecule composed of 4 rings. The three cyclohexane rings are labeled A, B, and C. These 3 rings together are called phenanthrene. Ring D is a cyclopentane ring. The carbon atoms are numbered from 1 to 17. Typically, steroids have a methyl group (–CH3) at carbons C-10 and C-13 and an alkyl side chain (R) at C-17 (Table 1). Alkanes are saturated hydrocarbons composed of carbon and hydrogen atoms linked by single bonds. The simplest alkyl group is a methyl group. Steroids vary by the configuration of the alkyl side chain, the number of additional methyl groups, and the functional groups attached to the steroid nucleus. Carbons number 18 and 19 are attached to carbons number 13 and 10, respectively. Additional carbon atoms are usually a part of the R side chain or attached elsewhere to the steroid backbone . Androgens are derivatives of androstane and contain 19 carbons and either a keto group (e.g., dehydroepiandrosterone (DHEA) and androstenedione (ASD)) or a hydroxy group (e.g., T and DHT) at position 17 of the steroid nucleus (Figure 1).
Steroid-5-reductases (5α-R and 5β-R) were first discovered, purified, and characterized in rat liver homogenates . These early experiments demonstrated that these enzymes were capable of irreversibly reducing the delta 4, 5 bond (double bond between carbons 4 and 5; Δ4,5) of C-19 and C-21 steroids to 5α- and 5β-stereoisomers.
The 5α-R family is composed of 3 subfamilies and 5 members (isozymes) in total. Isozymes are different proteins that perform the same function:
5α-R1 and 2 isozymes are NADPH-dependent, membrane-associated (microsomal) enzymes, composed of 259 and 254 amino acids, and have molecular weights of 29.5 and 28.4 kilodaltons, respectively. They contain a high content of hydrophobic amino acids distributed throughout their sequences, which suggests that they are intrinsic membrane proteins deeply embedded in the lipid bilayer.
When examined in lysates of transfected cells, 5α-R1 exhibits a broad pH optimum, which ranges between 6.0 and 8.5, while 5α-R2 shows a narrow acidic pH optimum (pH 5–5.5). However, there is evidence to suggest that inside intact human cells, 5α-R2 isozyme functions optimally at a more neutral pH range (6.0–7.0). 5α-R1 has a larger turnover number, as indicated by its Kcat value and a lower substrate affinity for T, Km = 1–5 μM. 5α-R2 has a lower turnover number (Kcat) and a higher substrate affinity, as indicated by Km = 0.004–1 μM for T. Under optimal conditions, 5α-R2 has a higher 5α-reducing activity than 5α-R1, as indicated by its high Vmax/km ratio. Both isozymes contain an NH2-terminal steroid (ligand) binding domain and a COOH-terminal NADPH binding domain. The apparent dissociation constant for NADPH cofactor is similar for both isozymes (3–10 μM). No such comparisons exist for 5α-R3 except that it appears to be efficient at pH 6.5–6.9 (unpublished work from our group).
Goal of development of 5α-reductase inhibitors (5α-RI) was to bind to 5α-R with little or no affinity for the androgen or other steroid receptors. The first inhibitors were steroids that mimicked T and, in many cases, were substrates themselves (i.e., not true inhibitors). The inhibitors can be broadly classified into two categories: steroidal and nonsteroidal. The steroidal class has more inhibitors thus far.
Numerous reports exist in the literature on the expression pattern of 5α-R1 and 5α-R2 in human tissue at various stages of development. The results vary due to differences in antibody sensitivity and specificity, mRNA analysis (in situ hybridization versus northern blotting versus reverse transcriptase-polymerase chain reaction), protein analysis (immunohistochemistry versus western blotting), tissue preparation, nature of tissue, evaluation of results, tissue fixation protocols, and control tissue. In addition, normal, benign, and malignant human tissue specimens are heterogeneous with variable expression of proteins among specimens from different individuals and within the same specimen, that is, inter- and intraindividual variability. Therefore, a summary of many studies that discussed the tissue distribution of 5α-R1-3 in different human tissues was tabulated to demonstrate differences in results (Table 3).
Alterations in the conversion of T into DHT by the enzyme 5α-R are associated with a number of human disorders:
The 5α-R system is an important player in human physiology and pathology. More work is needed to identify the biochemical characteristics and role of 5α-R3 in several human conditions such as CaP and androgen-stimulated skin diseases. Clinical data are inconclusive regarding the benefit of 5α-RI for CaP prevention. Clinical trials are ongoing to define the role of dutasteride for treatment of CaP, such as REDEEM (dutasteride in low-risk CaP patients on active surveillance), ARTS (biochemical failure after local treatment with curative intent), AVO 108943 (bicalutamide and dutasteride versus bicalutamide and placebo in CR-CaP), and ARI40006 (2-year follow-up study of REDUCE participants who received dutasteride or placebo) . Future trials should focus on blocking multiple steroidogenic enzymes at once, such as in men with biochemical failure after local therapy or men with CR-CaP. Blocking several different steps in steroidogenesis simultaneously may not allow CaP cells time to adjust to loss of androgen stimulation.