Through the Looking-Glass: Reevaluating DHEA Metabolism Through HSD3B1 Genetics

Abstract

Dehydroepiandrosterone (DHEA) and DHEA-sulfate together are abundant adrenal steroids whose physiologic effects are mediated through their conversion to potent downstream androgens. 3β-hydroxysteroid dehydrogenase isotype 1 (3βHSD1) facilitates the rate-limiting step of DHEA metabolism and gates the flux of substrate into the distal portion of the androgen synthesis pathway. Notably, a germline, missense-encoding change, HSD3B1(1245C), results in expression of 3βHSD1 protein that is resistant to degradation, yielding greater potent androgen production in the periphery. In contrast, HSD3B1(1245A) encodes 3βHSD1 protein that is easily degraded, limiting peripheral androgen synthesis. These “adrenal-permissive” and “adrenal-restrictive” alleles have recently been associated with divergent outcomes in androgen-sensitive disease states, underscoring the need to reevaluate DHEA metabolism using HSD3B1 genetics.

Dehydroepiandrosterone Metabolism: A Genetically Determined Predictor of Clinical Phenotypes

DHEA is one of the most abundant steroids in human circulation [1]. It is a precursor of potent sex steroids (see Glossary) synthesized under the control of adrenocorticotropic hormone in the zona reticularis of the adrenal gland, along with its sulfated conjugate, DHEA-S (Box 1) [2]. DHEA production peaks at birth and crests again during the third decade of life before steadily declining with age [3]. Despite its potential significance in development, the physiologic effects of DHEA remain incompletely characterized.

Text Box 1. Significance of DHEA and DHEA-S.

DHEA and DHEA-S are together the most abundant steroids in human circulation. DHEA is synthesized from 17α-OH-pregnenolone via the action of CYP17A1 in the adrenal gland and, to a lesser extent, the gonads [20]. DHEA may be further metabolized to DHEA-S in the adrenal gland by sulfotransferases, which conjugate this adrenal steroid with a sulfate group [56]. Conjugation with the sulfate moiety strengthens DHEA-S’ affinity for albumin, extending its half-life, and increasing its serum level relative to DHEA and other steroids [57]. In fact, DHEA-S is nearly two orders of magnitude more abundant than DHEA and testosterone, and over three orders of magnitude more abundant than estradiol [58]. Given that the sulfate group may be removed from DHEA-S via steroid sulfatase (STS) in peripheral tissues, DHEA-S functionally serves as an inactive reservoir for DHEA and potent sex steroid derivatives [59]. Therefore, in this manuscript, we emphasize DHEA metabolism because this adrenal steroid serves as the primary substrate for 3βHSD1 and because DHEA is more proximal to downstream androgen and estrogens.

DHEA may be best understood as an androgenic precursor whose biologic actions are effectuated through the production of testosterone and dihydrotestosterone [4]. Accordingly, DHEA must be evaluated through the action and kinetics of the enzyme responsible for its metabolism in peripheral tissues, 3βHSD1 [5]. Through the rate-limiting conversion of DHEA to androstenedione, 3βHSD1 gates the flux of precursor steroids into the androgen and estrogen synthesis pathways and regulates peripheral exposure to sex steroids [6]. Interestingly, 3βHSD1 expression is distinguished by the inheritance of two functional HSD3B1 alleles: the adrenal-restrictive allele limits conversion of DHEA to downstream androgens through expression of 3βHSD1 protein that is easily degraded, whereas, the adrenal-permissive allele enhances DHEA metabolism by conferring resistance to ubiquitination-mediated degradation [7]. Inheritance of the adrenal-permissive allele consequently results in accumulation of 3βHSD1 protein, enhanced androgen production, and marked activation of the androgen receptor [8].

There has been significant progress in understanding how HSD3B1 genetics produce clinically distinct phenotypes. In patients with advanced prostate cancer, inheritance of the adrenal-permissive allele has been shown to confer resistance to androgen deprivation therapy and reduce progression-free survival [9–14]. These validated findings are the consequence of enriched DHEA metabolism in prostatic tissue, which drives this androgen-sensitive disease [15]. Conversely, inheritance of the adrenal-restrictive allele has recently been associated with worse pulmonary function and glucocorticoid resistance in severe asthma [16]. In this case, exogenous glucocorticoids suppress adrenal DHEA and DHEA-S, attenuate 3βHSD1-mediated DHEA metabolism, and dysregulate pulmonary immune responses [17]. These studies are part of a growing body of literature demonstrating how HSD3B1 genotype regulates DHEA metabolism in a fashion that dictates clinical outcomes in androgen-dependent conditions.

Given these recent developments, it is time to reexamine DHEA physiology and metabolism through the lens of HSD3B1 genetics. In this paper, we assert this opinion by demonstrating how inheritance of the adrenal-permissive and adrenal-restrictive HSD3B1 alleles explain phenotypic variation in human disease, including metastatic prostate cancer and disorders of immune function. We also explore outcomes from DHEA supplementation studies and argue that HSD3B1 genotyping is necessary to elucidate the effects of this intervention.

HSD3B1 Genotype Alters DHEA Metabolism and Sex Steroid Biosynthesis

The importance of DHEA metabolism becomes clear when understood in terms of androgen biosynthesis (Figure 1). Though multiple synthetic pathways culminate in the production of potent androgens, all metabolic routes begin with the conversion of cholesterol to pregnenolone [18,19]. In adrenal pathways, pregnenolone is further metabolized to 17α-OH-pregnenolone and DHEA via 17α-hydroxylase/17,20-lyase (CYP17A1) through two sequential reactions [20]. DHEA is subsequently converted to androstenedione in peripheral tissues by 3βHSD1 in a rate-determining step (Box 2) [5,21]. Critically, 3βHSD1 gates the flux of DHEA into the distal portion of the androgen synthesis pathway, controlling the rate at which potent androgens may be synthesized [6,22]. DHEA metabolites (i.e., androstenedione and testosterone) may be further metabolized to estrone and estradiol in tissues expressing aromatase, underscoring the importance of 3βHSD1 in male and female sex steroid synthesis pathways [22].

Figure 1. 3βHSD1-Dependent Metabolism of DHEA Represents a Critical Juncture in Synthesis of Sex Steroids.

Dehydroepiandrosterone (DHEA) and dehydroepiandrosterone-sulfate (DHEA-S), highlighted in orange, together are the most abundant steroids in human circulation. DHEA-S may be converted to DHEA by the action of steroid sulfatase (STS), generating more of this essential sex steroid precursor. These adrenal steroids are derived from cholesterol, which is converted to pregnenolone by cholesterol side-chain cleavage enzyme (CYP11A1). Pregnenolone is subsequently metabolized to 17α-OH-pregnenolone and DHEA by 17α-hydroxylase/17,20-lyase (CYP17A1) in a stepwise reaction. 3β-hydroxysteroid dehydrogenase isotype 1 (3βHSD1) is necessary for the rate-limiting step in the peripheral conversion of DHEA to androstenedione, a reaction that gates the flux of metabolites downstream into sex steroid synthesis pathways. Potent androgens, including testosterone and dihydrotestosterone, may be generated via the action of 17β-hydroxysteroid dehydrogenase (17βHSD) and 5α-reductase (SRD5A). In tissues expressing aromatase, testosterone may be further metabolized to estradiol, a potent estrogen, as shown in pink. While this figure highlights the key enzymatic steps required for the generation of sex steroids, it is important to note that conversion of pregnenolone and 17-OH-pregnenolone to progesterone metabolites primarily occurs in the adrenal gland via the action of 3β-hydroxysteroid dehydrogenase isotype 2 (3βHSD2). The minor physiologic role 3βHSD1 plays in these reactions is indicated by the decreased thickness of the arrows reflecting this enzymatic activity. Not shown is conversion from androstenedione by aromatase to estrone.

Text Box 2. Distinguishing HSD3B1 and HSD3B2 Expression.

Given its critical role in the metabolism of adrenal precursor steroids, 3βHSD isoenzymes must be differentiated. 3βHSD isoenzyme 1 (3βHSD1) is encoded by HSD3B1 and is expressed predominantly in peripheral tissues such as breast, placenta, prostate, and skin. In contrast, 3βHSD isoenzyme 2 (3βHSD2) is encoded by HSD3B2 and is expressed in the adrenal gland, ovary, and testis [6]. Though there is a high-degree of structural homology between 3βHSD1 and 3βHSD2, their tissue-specific expression confers disparate phenotypic effects. This is classically demonstrated in 3βHSD2-associated congenital adrenal hyperplasia, in which homozygous inactivating mutations in HSD3B2 disrupt adrenal steroidogenesis in the setting of preserved HSD3B1 expression [60]. In this condition, severely reduced 3βHSD2 function prevents conversion of Δ⁵-3β-OH-steroids (including pregnenolone, 17α-OH-pregnenolone, DHEA, and androstenediol) to Δ⁴-3-keto-steroids (including progesterone, 17α-OH-progesterone, androstenedione, and testosterone), resulting in mineralocorticoid deficiency and DHEA excess [61]. Overabundance of adrenal sex steroid precursors drives production of potent androgens by peripheral 3βHSD1 in female infants, leading to mild virilization of the genitalia [62]. In male infants, peripheral DHEA metabolism alone does not produce adequate levels of androgens required for normal sexual differentiation, often resulting in ambiguous genitalia and hypospadias [60]. This example highlights the divergent functional roles of these isoenzymes: 3βHSD2 expression in the adrenal gland generates a circulating pool of Δ⁴-3-keto-steroids, while peripheral expression of 3βHSD1 facilitates sex steroid synthesis in non-endocrine tissues. Thus, the ensuing discussion of 3βHSD1 and HSD3B1 genetics pertains to physiologic and pathologic effects that are localized to specific peripheral tissues.

HSD3B1 genotype regulates the rate of DHEA metabolism by encoding 3βHSD1 protein with divergent susceptibilities for degradation. A common missense-encoding polymorphism in HSD3B1, rs1047303 (c.1245C>A, p.T367N), is responsible for these varying phenotypes [7]. Though HSD3B1(1245C) may develop somatically [8] and has been associated with transcriptional changes associated with cell cycle dysregulation, the discussion that follows will focus on the effects of germline HSD3B1 polymorphisms [23]. The HSD3B1(1245A) allele encodes 3βHSD1 protein that readily associates with autocrine mobility factor receptor, an ubiquitin E3 ligase that facilitates proteolysis through the endoplasmic-reticulum-associated degradation pathway [24]. Conversely, HSD3B1(1245C) encodes 3βHSD1 protein that exhibits resistance to ubiquitination and accrues intracellularly [25]. Accumulation of the protein responsible for gating the flux of adrenal steroids to potent sex steroids would be expected to generate an androgen-enriched milieu in the tissues expressing 3βHSD1 (Figure 2, Key Figure). This expected finding has been demonstrated consistently in vitro: experiments using LAPC4 cells transfected with the HSD3B1(1245C) allele revealed enhanced androgen synthesis, augmented prostate-specific antigen production, and accelerated xenograft growth when compared to cells transfected with the HSD3B1(1245A) allele [8]. Additionally, prostatic cell lines expressing the HSD3B1(1245C) allele, including LNCaP and VCaP, demonstrate more robust DHEA metabolism than those expressing the HSD3B1(1245A) allele, such as LAPC4 [24,26].

Figure 2. Inheritance of the Adrenal-Permissive Allele Enhances Peripheral Androgen Production.

HSD3B1 encodes 3β-hydroxysteroid dehydrogenase isotype 1 (3βHSD1) in peripheral tissues including the breast, placenta, prostate, and skin. 3βHSD1 is required for the conversion of dehydroepiandrosterone (DHEA), an adrenal steroid, to androstenedione in a rate-limiting step. Androstenedione may subsequently be converted to downstream androgens including testosterone and dihydrotestosterone in the androgen synthesis pathway. HSD3B1(1245A) encodes 3βHSD1 protein that readily associates with autocrine motility factor receptor (AMFR), an ubiquitin ligase that targets protein for endoplasmic-reticulum-associated degradation (ERAD). This results in a low concentration of intracellular 3βHSD1 protein and limits the conversion of DHEA precursor steroids to downstream androgens. Given this effect, we refer to HSD3B1(1245A) as the adrenal-restrictive allele because its inheritance limits the production of potent sex steroids from adrenal precursors. In contrast, HSD3B1(1245C) encodes a missense version of the 3βHSD1 protein that weakly interacts with AMFR, a property that confers resistance to ERAD. As a consequence, 3βHSD1 accumulates intracellularly and facilitates a greater flux of DHEA metabolites into the distal portion of the androgen biosynthesis pathway. We therefore refer to HSD3B1(1245C) as the adrenal-permissive allele because its inheritance enhances the metabolism of adrenal precursors to potent sex steroids, enriching production of androgens and estrogens in the periphery.

Though HSD3B1(1245C) has often been referred to as the “variant” allele because of its lower prevalence, it serves as the reference sequence in humans. These naming conventions are confusing, therefore, we will refer to the HSD3B1(1245C) as the “adrenal-permissive” allele, given that its inheritance amplifies the physiologic effects of adrenal steroid metabolism by encoding 3βHSD1 protein resistant to degradation. In contrast, HSD3B1(1245A) will be referred to as the “adrenal-restrictive” allele because its inheritance blunts conversion of DHEA to downstream metabolites. The adrenal-permissive and adrenal-restrictive terminology also describes the physiological basis by which inheritance of each allele contributes to disease phenotypes, giving this nomenclature clinical significance.

HSD3B1 Alleles Serve as Biomarkers for Prostate Cancer Clinical Outcomes

Prostate cancer is the most common non-dermatological cancer in men and is the second most common cause of oncologic death [27]. Localized disease is definitively treated with radiation or radical prostatectomy, though active surveillance may be offered to select patients [28]. Metastatic prostate cancer is often initially treated with androgen deprivation therapy (ADT), which consists of medical or surgical castration that ablates gonadal production of androgens [29]. Although adding other therapies to ADT have been found to be effective in recent years, these intensifying strategies will not be covered here. ADT encompasses a range of therapeutic options, including luteinizing hormone releasing hormone (LHRH) agonists, LHRH antagonists, and surgical management with bilateral orchiectomy [30]. Given that prostate cancer growth is androgen-driven, ADT is a highly effective index treatment for hormone-sensitive disease [31]. However, over time, metastatic prostate cancer develops a castration-resistant phenotype as intratumoral conversion of DHEA to downstream androgens drives tumor growth and disease progression [32,33].

Given the mechanistic relationship between HSD3B1 genetics and cellular phenotype, it was hypothesized that inheritance of the adrenal-permissive allele would permit a greater flux of DHEA metabolites into the androgen synthesis pathway and confer resistance to ADT. This hypothesis was evaluated by Hearn et al. in a multicohort study that assessed longitudinal outcomes in cohorts treated with ADT for post-prostatectomy biochemical failure (Table 1) [9]. Analysis of the primary cohort demonstrated that inheritance of the adrenal-permissive (AP) HSD3B1 allele worsened progression-free survival for patients on ADT, with hazard increasing in a dose-dependent fashion (HR: 1.7 for 1 AP allele, p=0.041; HR: 2.4 for 2 AP alleles, p=0.029). Distant metastasis-free survival (HR: 1.7 for 1 AP allele, p=0.074; HR: 2.7 for 2 AP alleles, p=0.022) and overall survival (HR: 2.0 for 1 AP allele, p=0.036; HR=3.3 for 2 AP alleles, p=0.013) followed a similar trend, worsening with the inherence of each additional allele. Clinical outcomes associated with HSD3B1 genotype were affirmed in two validation cohorts, providing strong evidence that inheritance of the AP allele predicts poor responsiveness to ADT [9].

Table 1.

Inheritance of the AP HSD3B1Allele Worsens Clinical Outcomes with ADT for Castration, Sensitive Prostate Cancer^a,b

		1 AP vs. 0 AP Alleles		2 AP vs. 0 AP Alleles
Cohort	Outcome	HR (95% CI)	P-Value	HR (95% CI)	P-Value	Reference
118 men treated with ADT for post-prostatectomy BCR	PFS	1.7 (1.0 – 2.9)	0.041	2.4 (1.1 – 5.3)	0.029	[9]
	Distant MFS	1.7 (1.0 – 2.8)	0.074	2.7 (1.2 – 6.2)	0.022
	OS	2.0 (1.1 – 3.7)	0.036	3.3 (1.3 – 8.3)	0.013
	PCaSpS	1.7 (0.8 – 3.3)	0.17	3.1 (1.1 – 8.7)	0.027
137 men treated with ADT for post-prostatectomy BCR	PFS	1.0 (0.7 – 1.7)	0.85	3.4 (1.6 – 7.0)	0.0013
188 men diagnosed with mCSPC	PFS	1.1 (0.8 – 1.5)	0.38	2.0 (1.1 – 3.8)	0.027
	OS	1.5 (1.0 – 2.1)	0.036	2.5 (1.2 – 5.0)	0.013
102 men treated with ADT for mCSPC	PFS	1.04 (0.64 – 1.07)	0.86	2.16 (1.01 – 4.58)	0.046	[10]
213 men treated with ADT for BCR after primary RT	TTM	1.19 (0.74 – 1.92)	0.48	2.01 (1.02–3.97)	0.045	[14]
		1 or 2 AP vs. 0 AP Alleles
Cohort	Outcome	HR (95% CI)			P Value	Reference
104 Japanese men treated with ADT for mCSPC	PFS	2.34 (1.08 – 4.49)			0.03	[11]
	OS	1.36 (0.52 – 2.92)			0.50
44 Spanish men treated with ADT for prostate cancer	PFS	2.4 (NR)c			.038	[12]
174 Caucasian men treated with ADT or ADT and docetaxel for LV mCSPC	2-year FF CRPC	1.89 (1.13 – 3.14)			0.02	[13]
	5-year OS	1.74 (1.01 – 3.00)			0.045
301 Caucasian men treated with ADT or ADT and docetaxel for HV mCSPC	2-year FF CRPC	1.10 (0.82 – 1.47)			0.52
	5-year OS	0.89 (0.65 – 1.22)			0.48

^aClinical studies reporting hazard ratios for prostate cancer progression, metastasis, or survival in cohorts that underwent HSD3B1 genotyping

^bAbbreviations: ADT, androgen deprivation therapy; AP, adrenal-permissive; BCR, biochemical recurrence; CRPC, castration-resistant prostate cancer; FF, freedom from; HV, high-volume; LV, low-volume; mCSPC, metastatic castration-sensitive prostate cancer; MFS, metastasis-free survival; NR, not reported; OS, overall survival; PCaSpS, prostate-cancer-specific survival; PFS, progression-free survival; RT, radiotherapy; TTM, time to metastasis

^cMedian PFS ratio calculated

The differential effect of HSD3B1 genotype on ADT responsiveness has been independently validated in multiple, international cohorts. Agarwal and colleagues independently demonstrated the finding that inheritance of the adrenal-permissive HSD3B1 allele decreases progression-free survival by evaluating an American cohort of 102 men initially treated with ADT for metastatic, castration-sensitive prostate cancer (HR: 1.04 for 1 AP allele, p=0.86; HR: 2.16, for 2 AP alleles, p=0.046) [10]. Similarly, Shiota and colleagues replicated these results in a cohort of 104 Japanese men treated with ADT for metastatic, castration-sensitive prostate cancer [11]. These investigators found that men inheriting the AP allele experienced worse progression-free survival (HR: 2.34 for 1 or 2 AP alleles, p=0.03) and similar overall survival (HR: 1.36 for 1 or 2 AP alleles, p=0.50) when compared to those homozygous for the adrenal-restrictive HSD3B1 allele. However, the comparisons were likely underpowered in this study given that AP allele frequency in the cohort was only 8.7%. Poorer progression-free survival was also exhibited in a Spanish cohort of 44 men receiving ADT with inheritance of the AP allele (24 months vs. 57 months, p=0.038), providing further evidence that HSD3B1 genotype serves as a biomarker for responsiveness to ADT [12].

Because metastatic prostate is characterized by a high degree of tumor heterogeneity in advanced disease states, the effect of HSD3B1 genotype was evaluated in patients with low-volume and high-volume disease. This was achieved by analyzing 475 patients in the CHAARTED clinical trial, which included Caucasian men with metastatic prostate cancer who were randomized to ADT with or without docetaxel [13]. Inheritance of 1 or 2 AP alleles was associated with reduced two-year freedom from castration-resistant disease 51.0% vs. 70.5%, p=0.01), worse five-year overall survival (57.5% vs. 70.8%, p=0.03), and a greater risk of death (HR: 1.74, p=0.045) in patients with low-volume metastatic cancer. Though these findings were not observed in the high-volume disease group, this study provides further support that HSD3B1 genotype serves as a biomarker for early resistance to castration therapy and overall survival in patients with low-volume, metastatic disease.

The effect of the AP allele on ADT responsiveness was also evaluated in patients experiencing biochemical recurrence following radiation treatment for localized disease [14]. Cohort analysis demonstrated that inheritance of the HSD3B1 genotype was associated with a copy-number-dependent decrease in time to metastasis (HR: 1.19 for 1 AP allele, p=0.48; HR: 2.01, for 2 AP alleles, p=0.045), though significant differences in composite progression-free survival (0 AP Alleles: 2.3 years, 1 AP Allele: 2.3 years, 2 AP Alleles: 1.4 years; p=0.68) and overall survival (0 AP Alleles: 7.7 years, 1 AP Allele: 6.9 years, 2 AP Alleles: 7.2 years; p=0.31) were not observed. This finding underscores that inheritance of the AP allele accelerates metastasis, as shown in prior studies. However, the absence of significant survival findings may explained by significant portions of the cohorts receiving prior treatment with ADT or concurrent treatment with androgen receptor blockers, which likely diluted the signal associated with HSD3B1 genotype.

HSD3B1 Genotype Is Associated with Glucocorticoid Responsiveness in Severe Asthma

There is strong evidence that higher levels of androgens improve outcomes in patients with asthma, as demonstrated by well-characterized gender disparities in disease prevalence [34]. In childhood, males exhibit an increased prevalence of asthma and are more likely to require hospitalization due to a severe exacerbation. This trend reverses in adolescence and adulthood, when the prevalence of asthma in women and need for hospitalization greatly exceeds those of men [35]. This epidemiologic shift has been attributed to the increased levels of androgens generated during male puberty, underscoring the role male sex steroids play in reactive airway disease. Androgen receptor signaling is thought to decrease IL-4, IL-5, IL-9, and IL-13 production from CD4⁺ T_H2 cells and group 2 innate lymphoid cells, attenuating IgE-mediated hypersensitivity responses and eosinophil activation [36,37]. Androgens are also thought to suppress IL-17 and IFNγ production, reducing neutrophil and natural killer cell recruitment to pulmonary tissues [38]. Though these mechanisms are still being investigated, it appears androgen receptor activation decelerates the immune responses associated with pathologic lung inflammation and airway hyperreactivity.

Intriguingly, increased circulating levels of DHEA and gonadal androgens have been associated with improved pulmonary function in asthmatics [39]. Given that 3βHSD1 facilitates androgen production from adrenal precursors in peripheral tissues, Zein and colleagues evaluated how HSD3B1 genotype affects respiratory function, as measured by the percent of predicted forced expiratory volume in one second (FEV₁PP), in a cohort of patients diagnosed with severe asthma [16]. Patients homozygous for the adrenal-restrictive allele on glucocorticoid therapy were found to have worse FEV₁PP than those not treated with glucocorticoids (54.3% vs. 75.1%, p<0.001), which was not observed in patients homozygous for the adrenal-permissive HSD3B1 allele (73.4% vs. 78.9%, p=0.39). Notably, pulmonary function worsened in a stepwise fashion with inheritance of each additional adrenal-restrictive allele in severe asthmatics on glucocorticoid therapy. Results were validated in a second asthma cohort. Exogenous glucocorticoids suppress DHEA production, limiting the 3βHSD1 substrate, which specifically affects individuals with the adrenal-restrictive genotype and effectively weaker enzyme, thus curbing peripheral androgen production in a fashion that exacerbates inflammatory immune responses in the airway. Expression of the adrenal-restrictive allele, therefore, blunts the benefit of glucocorticoid therapy and may serve as predictor of treatment resistance.

HSD3B1 Genetics May Clarify the Effects of DHEA Supplementation in Other Inflammatory Diseases

Given that multiple, validated cohort studies have demonstrated differential, genotype-specific effects in prostate cancer and glucocorticoid-resistant asthma, it is time to consider how other disease states may benefit from analysis through the lens of HSD3B1 genetics. One of these conditions is Systemic Lupus Erythematosus (SLE), a chronic, relapsing-remitting autoimmune disease characterized by multiorgan involvement with symptoms that include arthritis, nephritis, and mucocutaneous lesions [40]. Epidemiologic observations suggest that low androgens may contribute to the pathogenesis of SLE: the condition is nine times more prevalent in women, women diagnosed with the condition have lower circulating levels of DHEA and testosterone when compared to controls, and hypogonadal men are at greater risk of developing SLE than eugonadal counterparts [41–43]. Given their immunosuppressive effects, androgenic compounds have been assessed as potential treatments for SLE. For example, Danazol has been evaluated in small clinical trials that have demonstrated symptomatic improvement, increased serum complement and platelet levels, and decreased anti-double-stranded DNA immunoglobulin in a subset of patients [44]. DHEA supplementation studies have also been performed, with randomized control trials demonstrating reduced flare frequency, reduced need for prednisone treatment, and stabilization of symptom severity without loss of bone mineral density [45,46]. While not every DHEA supplementation trial has demonstrated benefit, the heterogeneity in clinical outcomes supports the hypothesis that some SLE patients may experience a relative androgen deficiency that is corrected with adrenal steroid supplementation, a phenotype consistent with inheritance of the adrenal-permissive HSD3B1 allele [37]. This signal, if present, may be obfuscated by including patients with sufficient peripheral androgen production conferred by expression of the adrenal-permissive allele. Future DHEA supplementation studies must stratify by HSD3B1 genotype in order to evaluate whether differential metabolism of adrenal steroids is associated with therapeutic benefit in SLE patients.

HSD3B1 Genotype May Clarify the Effects of DHEA Supplementation in Menopause

Female sexual dysfunction is a disorder with psychological and hormonal inputs that increases in prevalence in post-menopausal women [47]. Common symptoms—including vaginal atrophy, dyspareunia, reduced arousal, low libido, and increased difficulty achieving orgasm—occur as ovarian estrogen production and adrenal DHEA synthesis naturally decline [47,48]. Given that DHEA is an essential estrogen and androgen precursor, multiple investigators have investigated the effect of supplementation on sexual function in post-menopausal women. A recent review article summarizing these studies reported that DHEA supplementation typically improved sexual outcomes—including arousal, desire lubrication, and pain—in post-menopausal women who received 12 weeks or more of therapy [49]. Interestingly, these findings were also demonstrated in younger patients with adrenal insufficiency in a randomized control trial coordinated by Arlt and colleagues: four months of DHEA supplementation elevated levels of androgens, increased the frequency of sexual thoughts, enhanced sexual interest, and improved overall sexual satisfaction [50]. However, not all studies have demonstrated benefit from DHEA supplementation, particularly those including women with serum DHEA levels within reference ranges [48,49]. The variation in these findings underscores how age-related declines in circulating adrenal steroids contribute to sexual dysfunction by diminishing the availability of sex steroids in the periphery [51]. Given the importance of DHEA metabolism in correcting this relative deficiency, HSD3B1 genotype should be evaluated in future supplementation trials to assess how expression of adrenal-permissive and adrenal-restrictive alleles associates with sexual function outcomes in women. Inheriting the adrenal-restrictive allele, for example, may limit downstream sex steroid production in women already experiencing significant declines in adrenal steroid synthesis, making these individuals more symptomatic and more amenable to the potential benefits of supplementation.

Concluding Comments and Future Perspectives

We describe how HSD3B1 genetics serves as a lens that clarifies the physiologic and pathophysiologic significance of DHEA metabolism. By encoding a version of the 3βHSD1 protein that accumulates intracellularly, the adrenal-permissive allele confers a phenotype in which peripheral tissues are relatively enriched with potent sex steroids. Conversely, inheritance of the adrenal-restrictive allele results in 3βHSD1 protein that is more easily degraded and confers a phenotype in which peripheral tissues are relatively depleted with DHEA metabolites. This mechanistic relationship explains how HSD3B1 genetics highlights the need to reexamine disease states in which adrenal androgens have been implicated (see Outstanding Questions).

In particular, androgen-sensitive conditions may be better understood using HSD3B1 genetics. Given that HSD3B1 genotype has repeatedly been associated with prostate cancer progression, it is reasonable to hypothesize that inheritance of the adrenal-permissive allele would accelerate androgen-sensitive conditions such as benign prostatic hypertrophy and androgenic alopecia. Additionally, androgens may be converted to estrogens in tissues expressing aromatase, making the adrenal-permissive allele a potential prognostic factor in estrogen-sensitive conditions, like breast cancer [52–54]. 3βHSD1 inhibitors, such as trilostane and 17β-N,N-diethylcarbamoyl-4-methyl-4-aza-5-α-androstan-3-one (4-MA), have been investigated as potential therapeutic agents for androgen-sensitive disease states [55]. However, they each have their limitations, including, for example, partial androgen receptor agonist activity for trilostane and hepatotoxicity for 4-MA. Future developments in drug discovery and further characterization of the effects of HSD3B1 genotype in these disease states are certainly warranted.

HSD3B1 genetics must also be used to investigate conditions associated with reduced DHEA biosynthesis, such as autoimmune disease and age-related changes in mental and sexual function. While DHEA supplementation studies are key to evaluating how restoration of adrenal steroids affects symptom burden, no studies to date have stratified participants by HSD3B1 genotype. Given that inheritance of the adrenal-restrictive allele suppresses peripheral androgen synthesis, genotyping must be performed to fully appreciate how DHEA supplementation affects those with differing metabolic profiles.

Though much has yet to be elucidated about DHEA physiology, what remains certain is the tremendous role 3βHSD1 plays in mediating the potent downstream effects of DHEA metabolism. As the Wonderland of DHEA physiology is further characterized, the looking-glass of HSD3B1 genetics cannot be ignored.

Outstanding Questions.

• What is the physiological and phenotypic significance of DHEA as a sex steroid precursor?

• How does DHEA-S contribute to the circulating pool of DHEA? Are circulating levels of DHEA or DHEA-S correlated with disease development or severity? How does HSD3B1 genotype affect the conversion between DHEA and DHEA-S?

• How does HSD3B1 genotype affect age-related changes in human physiology, including immune dysregulation, sexual dysfunction, and unfavorable changes in body composition?

• Does inheritance of the adrenal-permissive allele worsen outcomes in other androgen-driven disease states like benign prostatic hypertrophy, androgenic alopecia, and polycystic ovarian syndrome?

• How does the relative suppression of peripheral androgen production associated with inheritance of the adrenal-restrictive allele affect immune functioning? In particular, how does inheritance of this allele affect outcomes autoimmune disorders such as systemic lupus erythematosus, rheumatoid arthritis, and multiple sclerosis?

• How does supplementation affect patients with lower serum levels of DHEA? How does HSD3B1 genotype magnify or attenuate these effects? Does DHEA supplementation improve outcomes in disease states associated with inheritance of the adrenal-restrictive allele?

Highlights.

• In vitro studies using prostatic cell lines demonstrated that the adrenal-permissive HSD3B1 allele encodes 3βHSD1 protein resistant to degradation, resulting in enhanced DHEA metabolism and increased androgen biosynthesis.

• Multiple clinical cohort studies have demonstrated that inheritance of the adrenal-permissive allele increases hazard for progression, metastasis, and death in patients with castration-sensitive prostate cancer treated with androgen deprivation therapy.

• Respiratory function worsens in a dose-dependent fashion with inheritance of the adrenal-restrictive allele in patients with severe asthma on glucocorticoid therapy, suggesting that suppressed DHEA metabolism in peripheral tissue dysregulates the immune response.

• HSD3B1 genotype may explain phenotypic variation in autoimmune disease, menopausal changes, and clinical responses to DHEA supplementation.

Glossary

Androgens

A group of 19-carbon steroids that facilitate the development and maintenance of male sex characteristics by activating the androgen receptor. Testosterone and dihydrotestosterone are potent androgens that activate the androgen receptor, whereas DHEA, androstenedione, and androstenediol primarily serve as precursor steroids.

Androgen deprivation therapy

Medical therapy or surgical interventions that ablate gonadal androgen production, typically utilized in the setting of metastatic, hormone-sensitive prostate cancer. Medical castration is achieved by using luteinizing hormone releasing hormone (LHRH) antagonists and LHRH agonists. Surgical castration typically involves bilateral orchiectomy.

Adrenal-permissive

Refers to the HSD3B1(1245C) allele and the phenotype resulting from its expression. The adrenal-permissive allele encodes 3βHSD1 protein that is resistant to degradation, expression of which results in enhanced peripheral DHEA metabolism and escalated production of potent downstream androgens.

Adrenal-restrictive

Refers to the HSD3B1(1245A) allele and the phenotype resulting from its expression. The adrenal-restrictive allele encodes 3βHSD1 protein that is readily degraded, expression of which results in limited peripheral DHEA metabolism and suppressed production of potent downstream androgens.

Endoplasmic-reticulum-associated degradation pathway

A proteolytic pathway in which misfolded, mutated, or aged proteins are transported to the cytoplasm, polyubiquitinated, and degraded by the proteasome.

Estrogens

A group of 18-carbon steroids that facilitate the development and maintenance of female sex characteristics by activating estrogen receptors. Physiologically relevant estrogens include estradiol, estriol, and estrone.

Sex steroids

A group of steroids produced in bulk from male and female gonads encompassing androgens and estrogens. Sex hormone production may occur in peripheral tissues expressing enzymes that metabolize precursor steroids.