A 53-year-old female had clinical and laboratory findings suggestive of Cushing's syndrome. In contrast to the Cushing's syndrome caused by cortical adenoma, a high level of urinary <em>17</em>-<em>ketosteroids</em> (<em>17</em>-KS) was also noted. Imaging studies revealed a right adrenal tumor. Right adrenectomy was performed; the surgical specimen revealed a black adenoma consisting of compact cells with numerous pigments which seemed to be lipofuscin in nature. The present case indicates that black adenoma as well as adrenocortical carcinoma should be suspected, when patients with Cushing's syndrome show an increased level of urinary <em>17</em>-KS excretion.

Feminizing adrenal tumors in young boys are rare; such patients initially show bilateral gynecomastia, and may have signs of virilization. We present a patient with bilateral gynecomastia, left adrenal adenoma, and elevated estrogen levels but normal levels of <em>17</em>-<em>ketosteroids</em>.

<em>17</em> beta-Hydroxysteroid dehydrogenases (<em>17</em>HSDs) catalyze the interconversions between active <em>17</em> beta-hydroxysteroids and less-active <em>17</em>-<em>ketosteroids</em> thereby affecting the availability of biologically active estrogens and androgens in a variety of tissues. The enzymes have different enzymatic properties and characteristic cell-specific expression patterns, suggesting differential physiological functions for the enzymes. Epidemiological and endocrine evidence indicate that estrogens play a key role in the etiology of breast cancer while androgens are involved in mechanisms controlling the growth of prostatic cells, both normal and malignant. Recently, we have developed, using LNCaP prostate cancer cell lines, a cell model to study the progression of prostate cancer. In the model LNCaP cells are transformed in culture condition to more aggressive cells, able to grow in suspension cultures. Our results suggest that substantial changes in androgen and estrogen metabolism occur in the cells during the process. These changes lead to increased production of active estrogens during transformation of the cells. Data from studies of breast cell lines and tissues suggest that the oxidative <em>17</em>HSD type 2 may predominate in human non-malignant breast epithelial cells, while the reductive <em>17</em>HSD type 1 activity prevails in malignant cells. Deprivation of an estrogen response by using specific <em>17</em>HSD type 1 inhibitors is a tempting approach to treat estrogen-dependent breast cancer. Our recent studies demonstrate that in addition to sex hormone target tissues, estrogens may be important in the development of cancer in some other tissues previously not considered as estrogen target tissues such as colon. Our data show that the abundant expression of <em>17</em>HSD type 2 present in normal colonic mucosa is significantly decreased during colon cancer development.

The neuroendocrine effects of haloperidol therapy have been examined in 62 male chronic schizophrenic patients, aged 16-62 years. The duration of the disease varied between 2 and 29 years. The patients were divided into 48 hebephrenics with onset of the disease at puberty, or immediately after puberty, and 14 paranoids with onset of the disease in adulthood. They received 6 mg i.m.p.d. of haloperidol, for 30 days, up to a total dose of 180 mg. The following hormonal variables were examined before therapy and at 10-20 and 30 days of treatment: total urinary gonadotropins, serum FSH and LH, GH response to insulin stimulation, ACTH reserve (Metyrapone test), total urinary <em>17</em>-<em>ketosteroids</em> and <em>17</em>-hydroxycorticoids before and after an ACTH stimulation test, serum testosterone, insulin response to glucose load, plasma thyroxine before and after a TSH stimulation test. The basic hormonal values revealed decreased secretion of total gonadotropins, FSH, LH, ACTH and testosterone, and increased insulin secretion. The haloperidol therapy seemed to stimulate the secretion of FSH, LH, total gonadotropins, ACTH and testosterone, up to normal or low-normal levels. No modifications were observed in the other hormonal variables. The significance of these results is discussed.

The enzymes of the <em>17</em> beta-hydroxysteroid dehydrogenase (<em>17</em> beta-HSD) gene family are responsible for a key step in the formation and degradation of androgens and estrogens: catalyzing the interconversion of <em>17</em>-<em>ketosteroids</em> and their active <em>17</em> beta-hydroxysteroid counterparts. The structure of human type II <em>17</em> beta-HSD cDNA was recently reported. This enzyme catalyzes the interconversion of delta 4-androstenedione and testosterone, androstanedione and dihydrotestosterone, and estrone and <em>17</em> beta-estradiol, whereas type I <em>17</em> beta-HSD catalyzes exclusively the interconversion of estrogens. To locate the HSD<em>17</em>B2 gene, the novel dinucleotide CA repeat sequence found 571 bp downstream from the end of exon 1 was genotyped into eight CEPH reference families by PCR. Two-point linkage analysis was performed between the latter polymorphism and the 2066 microsatellite markers of Généthon. The maximal pairwise lod score (Zmax = 33.3) with a maximal recombination fraction (theta max) of 0.008 was obtained with the marker D16S422 located on 16q24.1-q24.2. To define further the localization of the HSD<em>17</em>B2 gene, we constructed a high-resolution genetic map of the region flanking the polymorphic HSD<em>17</em>B2 gene including eight Généthon markers. The order of the HSD<em>17</em>B2 gene and markers is qter-D16S516-D16S504-D16S507-D16S505-D16S511+ ++-[HSD<em>17</em>B2-D16S422]-D16S520- D16S413-tel.

In addition to reproductive tissue, sex hormones induce transcriptional events in many connective tissue cells, including osteoblasts. Some sex hormone receptor modulators with bone sparing effects selectively target estrogen or androgen receptors, whereas others appear more promiscuous, in part through enzymatic metabolism. Rat osteoblasts express significant oxidative 3alpha-hydroxysteroid dehydrogenase activity, which can convert precursor substrates to potent androgen receptor agonists. Here we show that they also express 3-<em>ketosteroid</em> reductase activity, exemplified by 7-methyl-<em>17</em>-ethynyl-19-norandrostan-5 (10)en-3-one (tibolone) conversion to potent estrogen receptor alpha agonists. Conversion was rapid and quantitative, with 3alpha-hydroxytibolone as the primary metabolite. Consistently, tibolone induced estrogen receptor alpha-dependent gene promoter activity through cis-acting estrogen response elements, increased the stimulatory effect of TGF-beta on Smad-dependent gene promoter activity, and enhanced prostaglandin E2-induced activity of transcription factor Runx2. Rat osteoblasts express the 3-<em>ketosteroid</em> reductase AKR1C9, an aldo-keto reductase gene family member. Exposure to prostaglandin E2 increased AKR1C9 gene promoter activity and mRNA expression. AKR1C9 promoter activity was also enhanced by overexpression of protein kinase A catalytic subunit or transcription factor C/EBPdelta, and the effect of PGE2 was reduced by dominant negative C/EBPdelta competition or C/EBPdelta antisense expression. Moreover, prostaglandin E2 increased the amount of functional endogenous nuclear C/EBPdelta that could bind specifically to a distinct domain approximately 1.8-kb upstream from the start site of AKR1C9 transcription. In summary, in addition to 3alpha-hydroxysteroid dehydrogenase, rat osteoblasts express significant and regulatable 3-<em>ketosteroid</em> reductase activity. Through these enzymes, they may selectively metabolize precursor compounds into potent steroid receptor agonists locally within bone.

Between 1910 and 1940, 146 young females, aged 15-30 years, underwent bilateral salpingo-oophorectomy as part of a radical operation for salpingo-oophoritis. These women or their records were reviewed in 1971. 42 women had died in the meantime. More than half (22) of them had died from cardiovascular diseases, 5 from carcinoma of the uterus and 4 had committed suicide. None had died from carcinoma of the breast. Of 68 who were still alive, information by questionnaire was obtained and 32 were admitted to hospital for extensive examination. 32 age-matched women to be operated on for prolapse but with no other known disease of the reproductive tract served as controls. A further control group was added as 11 of the 68 were found to have menstruated again after the operation which had evidently not completely removed the gonads. Complete oophorectomy was found to have been followed by: (a) an increased incidence of cardiac symptoms and nervous diseases as well as an increased use of drugs; (b) a significant increase in the frequency of coronary vascular diseases in ages up to 70 years; (c) an increase in the serum cholesterol and triglycerides, most significantly in the ages below 60-65 years. Women with symptomatic coronary disease had a higher serum cholesterol level than women without and women with signs of peripheral vascular diseases had a significantly higher concentration of serum triglycerides; (d) an increased frequency of fractures (radius and femoral neck), increased osteoporosis and thinner cortical bone. The brittleness of the skeleton was correlated with low excretion of oestrogens in the urine. No vertebral compression or abnormal decrease in height was observed. (e) an increased adrenocortical activity with significantly increased excretion of <em>17</em>-<em>ketosteroids</em>, <em>17</em>-OH-<em>ketosteroids</em> and low polar total oestrogens. This activity abated in women above 65 years. (f) a traumatic psychological experience of the accompanying sterility while sexuality seemed to be largely unaffected in many of them. Almost half of the women examined by the psychiatrist were unusually mentally active and agile and they had a lower excretion of estriol than the rest.

The conventional treatment of CAH with hydrocortisone (16-19 mg/m2 per day) and 9 alpha-F-cortisol (just enough to normalise renin concentrations, started at 07:00 h) was ineffective in suppressing the early morning rise of <em>17</em>-OH-progesterone and in turn androgens in about 20% of our patients. The present work explored the effect of a modified dosage regimen of the drug in five patients. The schedule was: 03:00 h F 33% + 9 alpha-F-F 33%; 07:00 h F 30%; 12:00 h F 22% + 9 alpha-F-F 33%; <em>17</em>:30 h F 15% + 9 alpha-F-F 33%. Monitored levels of circulating <em>17</em>-OH-progesterone, testosterone, and individual urinary <em>17</em>-<em>ketosteroids</em> showed significant improvement, which was not achieved by giving higher or later evening doses. Menarche was induced in two girls (bone age 15 years). The modified dosage schedule offers on the one hand the possibility of better management of CAH, and on the other, cuts down the risk of enhanced Cushing-like effects, which in animal models have been related frequently to dosage schedules not corresponding to the circadian rhythm. The difficulty of administering the drugs at 03:00 h should be overcome by the development of a late-releasing preparation.

Acne is known to be one of the features of hyperandrogenism. The aim of the present work was to study women with persistent acne and without other evidence of hyperandrogenism, such as hirsutism, alopecia, or irregular menses. Among 87 female patients with acne and/or hirsutism, we defined three groups: group 1 (n = 29), patients having treatment-resistant acne without menstrual disturbance, alopecia, or hirsutism; group 2 (n = 27), patients with acne and hirsutism; and group 3 (n = 31), patients with hirsutism alone. Clinical chemistry criteria for hyperandrogenism were based on elevated values of one or more of the following parameters: plasma testosterone, delta-4-androstenedione, dehydroepiandrosterone, urinary 5 alpha-androstane 3 alpha-<em>17</em> beta-diol, and <em>17</em>-<em>ketosteroids</em> (with chromatography). Plasma and urine samples were drawn between the 18th and 25th days of the cycle. Among group 1 patients, we found 25 subjects (86%) with hyperandrogenism, according to these laboratory criteria. The etiologies were: polycystic ovary syndrome (36%), adrenal hypersecretion (40%, of which 12% showed secondary polycystic ovaries), isolated increase in 5 alpha-androstane 3 alpha-<em>17</em> beta-diol (20%), and hyperandrogenism without diagnosis (4%). The parameters were found to be more elevated in these patients than in a control group of 30 normal volunteer women. In groups 2 and 3, the findings were essentially the same as in group 1, except for increased levels of testosterone and the testosterone/SHBG ratio. Furthermore, it was evident that persistent acne may be an isolated sign of hyperandrogenism.

A major and a minor form of dihydrodiol dehydrogenase were co-purified with <em>17</em> beta-hydroxysteroid dehydrogenase and aldehyde reductase, respectively, to apparent homogeneity from liver cytosol of male ddY mice. The activities of dihydrodiol dehydrogenase and testosterone dehydrogenase or aldehyde reductase of the two enzyme forms comigrated electrophoretically. The major form of the enzyme oxidized <em>17</em> beta-hydroxysteroids and nonsteroidal alicyclic alcohols and reduced <em>17</em>-<em>ketosteroids</em> and various synthetic carbonyl compounds, showing higher affinity for steroids than for xenobiotics. The activity of this enzyme form toward benzene dihydrodiol and testosterone exhibited identical thermostability and susceptibility to inhibition by quercitrin, SH-reagents, nonsteroidal estrogens and anti-inflammatory agents. On the other hand, the minor form of the enzyme, which oxidized benzene dihydrodiol but not <em>17</em> beta-hydroxysteroids, also reduced various aldehydes well and was specifically inhibited by barbiturates and sorbinil. These results indicate that the major form of dihydrodiol dehydrogenase is identical to <em>17</em> beta-hydroxysteroid dehydrogenase and the minor enzyme form to aldehyde reductase.

An adrenal cortical tissue tumor developed in a patient with poorly controlled salt-losing congenital adrenal hyperplasia. A 16-year-old girl became progressively virilized from 13 to 16 years of age. Base line serum progesterone, <em>17</em>-hydroxyprogesterone, and testosterone levels were high and there was a diurnal pattern of the hormones. Initially elevated urinary <em>17</em>-<em>ketosteroid</em> and serum steroid levels were decreased by high dose dexamethasone therapy, and at laparotomy an adenoma was found in the cortex of the hyperplastic left adrenal gland. It is inferred that persistent adrenocorticotrophic hormone stimulation may result in neoplastic transformation of hyperplastic adrenal cortical tissue in patients with congenital adrenal hyperplasia.

Sexual development was evaluated in 9 female and 2 male subjects with Prader-Willi syndrome. The process of sexual development and degree of genital development attained were found to be variable but abnormal in all subjects. Hypothalamic-pituitary-gonadal functions were evaluated by measurement of serum Luteinizing Hormone and plasma testosterone responses to stimulation by clomiphene citrate and plasma testosterone responses to stimulation by human chorionic gonadotrophin. The degree of vaginal estrogenization was variable. The testicular biopsies showed abnormalities mainly in the germinal epithelium. In agreement with previous studies, it was concluded that the abnormalities of sexual development in this syndrome are mainly due to a defect in the hypothalamic pituitary axis. Adrenal function was not found to be grossly abnormal. The <em>17</em> <em>ketosteroid</em> excretion values were low, probably explaining the rather sparse pubic and axillary hair observed in these patients. The urinary <em>17</em>-hydroxycorticosteroid creatinine ratios were found to be elevated, probably due to decreased creatinine excretion, reflecting the muscular abnormalities of these subjects.

Serum beta-glucuronidase activity is shown to differ quantitatively in the following strains of mice, listed in order of increasing activity: C3H, C57BL/6 less than BALB/c, DBA/2, ICR less than SENCAR, A/He. The level of the enzyme in the murine strains is shown to correlate with the urinary excretion of <em>17</em>-<em>ketosteroids</em>, which in turn reflects the endogenous level of androgens. Dietary calcium D-glucarate, an in vivo beta-glucuronidase inhibitor, reduced the steady state level of both beta-glucuronidase and <em>17</em>-<em>ketosteroid</em> excretion in the highly susceptible A/He and SENCAR strains to that of strains known to be resistant to chemical carcinogenesis. Sensitivity of the A/He strain is significantly reduced by dietary calcium glucarate, which is shown to inhibit DNA binding and the induction of pulmonary adenomas by benzo[a]pyrene.

Gonadotropin levels and secretory patterns were studied in 28 oligomenorrheic patients with various types of polycystic ovary disease (PCO). On the basis of ovarian morphology and histology, the patients PCOuld be separated into two distinct categories arbitarily designated "typical" (type I) and "atypical" (type II) PCO. Although no differences were noted in symptomatology or <em>17</em>-<em>ketosteroid</em>, testosterone, or follicle-stimulating hormone levels, the 12 type I patients demonstrated widely fluctuating, but markedly elevated, luteinizing hormone (LH) levels, while the 16 type II patients demonstrated lower and less fluctuating LH levels which were comparable to those found during the normal follicular phase. It is likely that type I PCO is a distinct entity similar to that described by Stein and Leventhal, while type II co represents a heterogenous spectrum of disorders, many of which remain obscure.

Concentrations of testosterone, luteinizing hormone (LH), and follicle-stimulating hormone (FSH) in serum and <em>17</em>-<em>ketosteroids</em> (<em>17</em>-KS) in urine of 10 paraplegic and 10 quadriplegic subjects were measured from onset of injury and followed once a week for 4 months. Compared with age-matched normal controls, paraplegic subjects showed significantly lower serum levels of LH and FSH for 2 weeks and of testosterone for 6 weeks after spinal cord trauma, following which periods of time these hormones attained normal levels. By contrast, in quadriplegic subjects, serum testosterone concentrations remained significantly lower than those of the controls during the entire 4-month testing period. Furthermore, in another group of 10 chronic (1 to 6 years after onset of injury) paraplegic and 10 chronic quadriplegic subjects, serum testosterone and FSH concentrations were comparable to those of the normal controls. Serum LH concentrations were at control levels in chronic paraplegic but significantly depressed in chronic quadriplegic subjects. The concentrations of urinary <em>17</em>-KS exhibited sharp fluctuations over the 4-month period and were below control levels in paraplegic but within control limits in quadriplegic subjects. The results indicate that the function of the hypothalamic-pituitary-gonadal axis is disturbed for at least 4 months in quadriplegic subjects.

Dihydrodiol dehydrogenase activity was detected in the cytosol of various mouse tissues, among which kidney exhibited high specific activity comparable to the value for liver. The enzyme activity in the kidney cytosol was resolved into one major and three minor peaks by Q-Sepharose chromatography: one minor form cross-reacted immunologically with hepatic 3 alpha-hydroxysteroid dehydrogenase and another with aldehyde reductase. The other minor form was partially purified and the major form was purified to homogeneity. These two forms, although different in their charges, were monomeric proteins with the same molecular weight of 39,000 and had similar catalytic properties. They oxidized cis-benzene dihydrodiol and alicyclic alcohols as well as trans-dihydrodiols of benzene and naphthalene in the presence of NADP+ or NAD+, and reduced several xenobiotic aldehydes and ketones with NAD(P)H as a cofactor. The enzymes also catalyzed the oxidation of 3 alpha-hydroxysteroids and epitestosterone, and the reduction of 3- and <em>17</em>-<em>ketosteroids</em>, showing much lower Km values (10(-7)-10(-6) M) for the steroids than for the xenobiotic alcohols. The results of mixed substrate experiments, heat stability, and activity staining on polyacrylamide gel electrophoresis suggested that, in the two enzymes, both dihydrodiol dehydrogenase and 3(<em>17</em>)alpha-hydroxysteroid dehydrogenase activities reside on a single enzyme protein. Thus, dihydrodiol dehydrogenase existed in four forms in mouse kidney cytosol, and the two forms distinct from the hepatic enzymes may be identical to 3(<em>17</em>)alpha-hydroxysteroid dehydrogenases.

Rodent <em>17</em>beta-hydroxysteroid dehydrogenase/<em>17</em>-<em>ketosteroid</em> reductase type 7 (<em>17</em>HSD/KSR7) catalyzes the conversion of estrone (E1) to estradiol (E2) and is abundantly expressed in the ovaries of pregnant animals in particular. In the present work we demonstrate cell-specific expression of <em>17</em>HSD/KSR7 in the ovaries, uteri, and placentas of pregnant and nonpregnant mice using in situ hybridization. The results show that mouse <em>17</em>HSD/KSR7 (m<em>17</em>HSD/KSR7) messenger RNA is distinctly and exclusively expressed in a proportion of corpora lutea (CLs). During pregnancy, expression of m<em>17</em>HSD/KSR7 is most abundant around embryonic day 14.5 (E14.5), when the ovaries are filled with CLs expressing <em>17</em>HSD/KSR7. In the uterus, m<em>17</em>HSD/KSR7 is first detected on E5.5, when expression surrounds the implantation site on the antimesometrial side. As gestation progresses, m<em>17</em>HSD/KSR7 is expressed in the decidua capsularis on E8 and E9.5, disappearing thereafter from the antimesometrial decidua. On E9 onward, m<em>17</em>HSD/KSR7 messenger RNA expression takes place at the junctional zone of the developing placenta. On E12.5 and E14.5, m<em>17</em>HSD/KSR7 is abundantly expressed in the spongiotrophoblasts, where expression gradually declines toward parturition. In conclusion, m<em>17</em>HSD/KSR7 expression in the CL is related to the life span of the CL. Moreover, spatial and temporal expression of m<em>17</em>HSD/KSR7 in the uterus suggests that locally produced E2 plays a role in implantation and/or decidualization. Finally, the results indicate that mouse placenta is capable of converting E1 to E2 in situ, and that the synthesized E2 may be effective in a paracrine, autocrine, and/or intracrine manner and be involved in placentation.

A 72-year-old postmenopausal woman with high plasma testosterone levels and virilization, as demonstrated by hirsutism and balding, is presented. Urinary <em>17</em>-<em>ketosteroids</em> and <em>17</em>-hydroxycorticosteroids, as well as computed axial tomography scan of the adrenal glands were normal. Although no pelvic mass was detected by ultrasonography or pelvis examination, the patient was found to have small bilateral hilus cell tumors of the ovary. Following bilateral salpingo-oophorectomy the plasma testosterone dropped to normal level. This is the third case of bilateral hilus cell tumors of the ovary to be reported.

Natural and synthetic steroid hormones excreted into the environment are potentially threatening the population dynamics of all kinds of animals and public health. We have previously isolated a steroid degrading bacterial strain (H5) from the Baltic Sea, at Kiel, Germany. 16S-rRNA analysis showed that bacterial strain H5 belongs to the genus Vibrio, family Vibrionaceae and class Gamma-Proteobacteria. Bacterial strain H5 can degrade steroids such as testosterone and estrogens, which was shown in this study by determining the (3)H labeled steroid retaining in the bacterial H5 culture medium at incubation times of 5 h and 20 h. Since 3α-hydroxysteroid dehydrogenase/carbonyl reductase (3α-HSD/CR) is a key enzyme in adaptive steroid degradation in Comamonas testosteroni (C. testosteroni), in previous investigations, a meta-genomic system with the 3α-HSD/CR gene as a positive control was established. By this meta-genomic system, two estradiol inducible genes coding 3-<em>ketosteroid</em>-delta-1-dehydrogenase and carboxylesterase, respectively, which are involved in steroid degradation, were found in marine strain H5. In the present work, the 3-<em>ketosteroid</em>-delta-1-dehydrogenase and carboxylesterase genes were subcloned into plasmids pET38-12 and pET24-<em>17</em>, respectively. Overexpression in Escherichia coli (E. coli) strain BL21(DE3)pLysS cells resulted in corresponding proteins with an N-terminal His-tag sequence. After induction with isopropyl-β-D-thiogalactoside, 3-<em>ketosteroid</em>-delta-1-dehydrogenase and carboxylesterase were purified in one step using nickel-chelate chromatography. After protein determination, 3-<em>ketosteroid</em>-delta-1-dehydrogenase (0.48 mg/ml) and carboxylesterase (1.28 mg/ml) were used to prepare antibodies to determine steroid binding specificity in future research. In summary, we have shown that the marine strain H5 could metabolize steroids; have isolated two estradiol inducible genes from strain H5 chromosomal DNA, and purified the corresponding proteins for further research. The exact characterization and systematic classification of the marine steroid degrading bacterial strain H5 is envisaged. The strain might be used for the bioremediation of steroid contaminations in seawater.

Androgens delay lung maturation through their action on lung fibroblasts. Knowing that testosterone is secreted by the lung epithelial-like cell line A-549, we have studied the metabolism of androgens by several human lung diploid fibroblasts cell lines. No <em>17</em>-<em>ketosteroid</em> reductase activity was detected. In contrast, testosterone was transformed mainly into androstenedione and androstanedione with no 5 alpha-dihydrotestosterone formed, indicating the presence of <em>17</em> beta- hydroxysteroid dehydrogenase (HSD) type 2 and 5 alpha-reductase activities. The eight cell lines analyzed had either a low or high <em>17</em> beta-HSD type 2 activity level. No correlation between these levels and the sex or age stage of cells was established, but Northern blot analysis of human lung RNA samples of five adult subjects revealed very similar variations between subjects in the level of <em>17</em> beta-HSD type 2 mRNA. The 5 alpha-reductase activity had a marked substrate preference for androstenedione, the product of <em>17</em> beta-HSD type 2. When tritiated testosterone was used as substrate, only barely detectable levels of 5 alpha-dihydrotestosterone were observed by HPLC in the presence of the <em>17</em> beta-HSD type 2 inhibitor EM-919. The use of unlabeled testosterone or of the antiandrogen hydroxyflutamide demonstrated that the tritiated testosterone substrate itself had no effect on levels of 5 alpha-reduction. In fact, in these cells, 5 alpha-reductase has no significant activity on testosterone, but it further converts the product of <em>17</em> beta-HSD type 2, thus playing a role with <em>17</em> beta-HSD type 2 in androgen inactivation. Because androgens delay lung maturation and surfactant synthesis by their action on lung fibroblasts, it is of particular interest to find that the steroid metabolism of these lung fibroblast cells is oriented toward androgen inactivation. Because lung fibroblasts of subjects with low <em>17</em> beta-HSD type 2 expression levels are likely to be exposed to higher levels of androgens, an allelic variation of the <em>17</em> beta-HSD-2 gene is suspected, which would result in familial incidence of respiratory distress. This is in line with reported cases of familial incidence of respiratory distress.

The preparation of steroidal[<em>17</em>,16-d][1,2,4]triazolo[1,5-a]pyrimidines and their biological evaluation as potential anticancer agents are herein reported. These novel heterosteroids (2, 4) were prepared through the condensation reaction of 3-amino-1,2,4-triazole with 16-arylidene-<em>17</em>-<em>ketosteroids</em> (1, 3). All the synthesized compounds were evaluated for their anticancer activity in vitro against PC-3 (human prostatic carcinoma), MCF-7 (human breast carcinoma) and EC9706 (human esophageal carcinoma) cell lines. Among the screened compounds, 2i, 2n and 4f showed significant inhibitory activity against all the three human cell lines.

Mycobacterium neoaurum ST-095 and its mutant M. neoaurum JC-12, capable of transforming phytosterol to androst-1,4-diene-3,17-dione (ADD) and androst-4-ene-3,17-dione (AD), produce very different molar ratios of ADD/AD. The distinct differences were related to the enzyme activity of 3-ketosteroid-Δ(1)-dehydrogenase (KSDD), which catalyzes the C1,2 dehydrogenation of AD to ADD specifically. In this study, by analyzing the primary structure of KSDDI (from M. neoaurum ST-095) and KSDDII (from M. neoaurum JC-12), we found the only difference between KSDDI and KSDDII was the mutation of Val(366) to Ser(366). This mutation directly affected KSDD enzyme activity, and this result was confirmed by heterologous expression of these two enzymes in Bacillus subtilis. Assay of the purified recombinant enzymes showed that KSDDII has a higher C1,2 dehydrogenation activity than KSDDI. The functional difference between KSDDI and KSDDII in phytosterol biotransformation was revealed by gene disruption and complementation. Phytosterol transformation results demonstrated that ksdd I and ksdd II gene disrupted strains showed similar ADD/AD molar ratios, while the ADD/AD molar ratios of the ksdd I and ksdd II complemented strains were restored to their original levels. These results proved that the different ADD/AD molar ratios of these two M. neoaurum strains were due to the differences in KSDD. Finally, KSDD structure analysis revealed that the Val(366)Ser mutation could possibly play an important role in stabilizing the active center and enhancing the interaction of AD and KSDD. This study provides a reliable theoretical basis for understanding the structure and catalytic mechanism of the Mycobacteria KSDD enzyme.