Extensive metabolic consequences of human glycosyltransferase gene knockouts in prostate cancer.

BACKGROUND: Naturally occurring germline gene deletions (KO) represent a unique setting to interrogate gene functions. Complete deletions and differential expression of the human glycosyltransferase UGT2B17 and UGT2B28 genes are linked to prostate cancer (PCa) risk and progression, leukaemia, autoimmune and other diseases. METHODS: The systemic metabolic consequences of UGT deficiencies were examined using untargeted and targeted mass spectrometry-based metabolomics profiling of carefully matched, treatment-naive PCa cases. RESULTS: Each UGT KO differentially affected over 5% of the 1545 measured metabolites, with divergent metabolic perturbations influencing the same pathways. Several of the perturbed metabolites are known to promote PCa growth, invasion and metastasis, including steroids, ceramides and kynurenine. In UGT2B17 KO, reduced levels of inactive steroid-glucuronides were compensated by sulfated derivatives that constitute circulating steroid reservoirs. UGT2B28 KO presented remarkably lower levels of oxylipins paralleled by reduced inflammatory mediators, but higher ceramides unveiled as substrates of the enzyme in PCa cells. CONCLUSION: The distinctive and broad metabolic rewiring caused by UGT KO reinforces the need to examine their unique and divergent functions in PCa biology.

Preoperative Circulating 11-Oxygenated Androgens Are Associated With Metastasis-free Survival in Localized Prostate Cancer.

PURPOSE: Adrenal 11-oxygenated androgens may support cancer progression in men with prostate cancer owing to their abundance and androgenic potential. We hypothesized that preoperative circulating levels of 11-oxygenated androgens influence clinical outcomes in men with newly diagnosed localized prostate cancer. MATERIALS AND METHODS: We studied 1,793 treatment-naïve patients and 155 patients who received preoperative treatment with 5α-reductase inhibitors in the prospective PROCURE cohort, for which preoperative plasma samples were obtained prior to radical prostatectomy. Adrenal 11-oxygenated precursors, potent 11-oxygenated androgens and their metabolites (n=7), were quantified using liquid chromatography-tandem mass spectrometry. Circulating levels were evaluated in relation to prognostic factors, disease-free survival, and metastasis-free survival using multivariable Cox proportional hazards models. RESULTS: At a median follow-up of 93.8 months after surgery, 583 patients experienced biochemical recurrence, 104 developed metastatic disease, and 168 deceased. Higher levels of 11-hydroxytestosterone and 11-ketotestosterone were observed in men with PSA >20 ng/mL and positive nodal status (P < .05). In multivariable analyses, no significant association between 11-oxygenated androgens and disease-free survival was observed. Adrenal 11ß-hydroxyandrostenedione, the predominant androgenic 11-ketotestosterone, and its metabolite 11-ketoandrosterone, modeled as quartiles, were associated with metastasis-free survival (P = .06, P = .03, and P = .008, respectively). Significant accumulation of 11-oxygenated androgen precursors and bioactive androgens, but reduced metabolite levels, was observed in patients on 5α-reductase inhibitors (P < .001). CONCLUSIONS: Preoperative circulating 11-oxygenated androgen levels are associated with metastasis-free survival in men with localized prostate cancer undergoing radical prostatectomy and are affected by 5α-reductase inhibitor treatment.

Extensive Alteration of Androgen Precursor Levels After Castration in Prostate Cancer Patients and Their Association With Active Androgen Level.

PURPOSE: Dihydrotestosterone and testosterone are thought to be major contributors of prostate cancer progression and resistance. We studied the modulation of 15 circulating steroids by castration and their association with dihydrotestosterone and testosterone levels. MATERIALS METHODS: A total of 116 serum samples were collected from 99 prostate cancer patients and categorized as eugonadal, castration-sensitive prostate cancer, castration-resistant prostate cancer, or castration-resistant prostate cancer under abiraterone acetate. Serum levels of 15 steroids were measured using mass spectrometry and compared between groups using analysis of variance. Intrapatient association of steroid levels and the androgens testosterone and dihydrotestosterone were assessed using Pearson correlation and linear regression. RESULTS: Testosterone, dihydrotestosterone, androstenedione, dehydroepiandrosterone, dehydroepiandrosterone-sulfate, androsterone, androstenediol, estrone, estrone-sulfate, estradiol, and androsterone/3α-diol-3/3α-diol-17-glucuronide levels were significantly decreased in castration-sensitive prostate cancer (castrated) compared to eugonadal patients. Testosterone levels were strongly associated with multiple steroids under eugonadal conditions, whereas they were weakly affected by precursor steroids in castrated patients. By contrast, dihydrotestosterone levels under androgen deprivation therapy were associated with testosterone and the backdoor pathway metabolite androsterone. In castration-resistant prostate cancer patients, levels of androstenedione were significantly associated with testosterone level, while testosterone was the only steroid associated with dihydrotestosterone levels. CONCLUSIONS: Androgen deprivation therapy significantly reduces the levels of 13 circulating steroids. Upon androgen deprivation therapy initiation, the backdoor pathway metabolite androsterone are strongly associated with dihydrotestosterone levels. Under castration-resistant prostate cancer conditions, androstenedione was significantly associated with testosterone levels, suggesting the presence of tumor-related circulating androgens in these patients. These results provide further rationale to intensify treatments with androgen receptor axis signaling pathway inhibitors in patients with prostate cancer.

Urinary oestrogen steroidome as an indicator of the risk of localised prostate cancer progression.

BACKGROUND: Prostate cancer (PCa) is the most common cancer in North American men. Beyond the established contribution of androgens to disease progression, growing evidence suggest that oestrogen-related pathways might also be of clinical importance. The aim of this study was to explore the association of urinary oestrogen levels with clinical outcomes. METHODS: Urine samples from the prospective multi-institutional PROCURE cohort were collected before RP for discovery (n = 259) and validation (n = 253). Urinary total oestrogens (unconjugated + conjugated), including oestrone and oestradiol, their bioactive and inactive catechol and methyl derivatives (n = 15), were measured using mass spectrometry (MS). RESULTS: The median follow-up time for the discovery and replication cohorts was 7.6 and 6.5 years, respectively. Highly significant correlations between urinary oestrogens were observed; however, correlations with circulating oestrogens were modest. Our findings indicate that higher levels of urinary oestriol and 16-ketoestradiol were associated with lower risk of BCR. In contrast, higher levels of 2-methoxyestrone were associated with an increased risk of development of metastasis/deaths. CONCLUSIONS: Our data suggest that urinary levels of oestriol and 16-ketoestradiol metabolites are associated with a more favourable outcome, whereas those of 2-methoxyestrone are associated with an elevated risk of metastasis after RP. Further studies are required to better understand the impact of oestrogens on disease biology and as easily accessible urine-based risk-stratification markers.

Associations Between Prediagnostic Concentrations of Circulating Sex Steroid Hormones and Liver Cancer Among Postmenopausal Women.

BACKGROUND AND AIMS: In almost all countries, incidence rates of liver cancer (LC) are 100%-200% higher in males than in females. However, this difference is predominantly driven by hepatocellular carcinoma (HCC), which accounts for 75% of LC cases. Intrahepatic cholangiocarcinoma (ICC) accounts for 12% of cases and has rates only 30% higher in males. Hormones are hypothesized to underlie observed sex differences. We investigated whether prediagnostic circulating hormone and sex hormone binding globulin (SHBG) levels were associated with LC risk, overall and by histology, by leveraging resources from five prospective cohorts. APPROACH AND RESULTS: Seven sex steroid hormones and SHBG were quantitated using gas chromatography/tandem mass spectrometry and competitive electrochemiluminescence immunoassay, respectively, from baseline serum/plasma samples of 191 postmenopausal female LC cases (HCC, n = 83; ICC, n = 56) and 426 controls, matched on sex, cohort, age, race/ethnicity, and blood collection date. Odds ratios (ORs) and 95% confidence intervals (CIs) for associations between a one-unit increase in log2 hormone value (approximate doubling of circulating concentration) and LC were calculated using multivariable-adjusted conditional logistic regression. A doubling in the concentration of 4-androstenedione (4-dione) was associated with a 50% decreased LC risk (OR = 0.50; 95% CI = 0.30-0.82), whereas SHBG was associated with a 31% increased risk (OR = 1.31; 95% CI = 1.05-1.63). Examining histology, a doubling of estradiol was associated with a 40% increased risk of ICC (OR = 1.40; 95% CI = 1.05-1.89), but not HCC (OR = 1.12; 95% CI = 0.81-1.54). CONCLUSIONS: This study provides evidence that higher levels of 4-dione may be associated with lower, and SHBG with higher, LC risk in women. However, this study does not support the hypothesis that higher estrogen levels decrease LC risk. Indeed, estradiol may be associated with an increased ICC risk.

UGT2B17 modifies drug response in chronic lymphocytic leukaemia.

BACKGROUND: High UGT2B17 is associated with poor prognosis in untreated chronic lymphocytic leukaemia (CLL) patients and its expression is induced in non-responders to fludarabine-containing regimens. We examined whether UGT2B17, the predominant lymphoid glucuronosyltransferase, affects leukaemic drug response and is involved in the metabolic inactivation of anti-leukaemic agents. METHODS: Functional enzymatic assays and patients' plasma samples were analysed by mass-spectrometry to evaluate drug inactivation by UGT2B17. Cytotoxicity assays and RNA sequencing were used to assess drug response and transcriptome changes associated with high UGT2B17 levels. RESULTS: High UGT2B17 in B-cell models led to reduced sensitivity to fludarabine, ibrutinib and idelalisib. UGT2B17 expression in leukaemic cells involved a non-canonical promoter and was induced by short-term treatment with these anti-leukaemics. Glucuronides of both fludarabine and ibrutinib were detected in CLL patients on respective treatment, however UGT2B17 conjugated fludarabine but not ibrutinib. AMP-activated protein kinase emerges as a pathway associated with high UGT2B17 in fludarabine-treated patients and drug-treated cell models. The expression changes linked to UGT2B17 exposed nuclear factor kappa B as a key regulatory hub. CONCLUSIONS: Data imply that UGT2B17 represents a mechanism altering drug response in CLL through direct inactivation but would also involve additional mechanisms for drugs not inactivated by UGT2B17.

Alternative promoters control UGT2B17-dependent androgen catabolism in prostate cancer and its influence on progression.

BACKGROUND: Perturbation of the major UGT2B17-dependent androgen catabolism pathway has the potential to affect prostate cancer (PCa) progression. The objective was to evaluate UGT2B17 protein expression in primary tumours in relation to hormone levels, disease characteristics and cancer evolution. METHODS: We conducted an analysis of a high-density prostate tumour tissue microarray consisting of 239 localised PCa cases treated by radical prostatectomy (RP). Cox proportional hazard ratio analysis was used to evaluate biochemical recurrence (BCR), and a linear regression model evaluated variations in circulating hormone levels measured by mass spectrometry. The transcriptome of UGT2B17 in PCa was established by using RNA-sequencing data. RESULTS: UGT2B17 expression in primary tumours was associated with node-positive disease at RP and linked to circulating levels of 3α-diol-17 glucuronide, a major circulating DHT metabolite produced by the UGT2B17 pathway. UGT2B17 was an independent prognostic factor linked to BCR after RP, and its overexpression was associated with development of metastasis. Finally, we demonstrated that distinctive alternative promoters dictate UGT2B17-dependent androgen catabolism in localised and metastatic PCa. CONCLUSIONS: The androgen-inactivating gene UGT2B17 is controlled by overlooked regulatory regions in PCa. UGT2B17 expression in primary tumours influences the steroidome, and is associated with relevant clinical outcomes, such as BCR and metastasis.

Extragonadal Steroids Contribute Significantly to Androgen Receptor Activity and Development of Castration Resistance in Recurrent Prostate Cancer after Primary Therapy.

PURPOSE: Beyond testosterone, several steroids contribute to the activation of the androgen receptor pathway, but their relative contributions to the activation of the androgen receptor signaling axis in patients with castrated prostate cancer remain unknown. MATERIALS AND METHODS: Serum levels of 9 steroids were measured by mass spectrometry from continuously castrated patients of the PR.7 study (219) and from the PCA24 cohort (116). For each steroid standard curves for dose dependent prostate specific antigen promoter activation were built in castration sensitive (LAPC4) and resistant (VCaP) prostate cancer models. Standard curves were used to determine the androgen receptor activation potency for each steroid measurement from patients in these trials. RESULTS: In LAPC4 and VCaP cells testosterone, dihydrotestosterone and androstenedione induced androgen receptor transcriptional activity, while dehydroepiandrosterone, 5alpha-androstan-3beta,17beta-diol, androstenediol and androsterone stimulated androgen receptor only in VCaP cells. Extragonadal steroids were responsible for 34% (LAPC4) and 88% (VCaP) of the serum total androgen receptor transcriptional activity found in castrated cases. The total androgen receptor transcriptional activity secondary to testosterone, dihydrotestosterone and androstenedione was associated with time to castration resistance in patients from the PR.7 study (HR 2.17, 95% CI 1.12-4.23, p=0.02) in multivariate analysis using the castration sensitive model (LAPC4). Androgen receptor transcriptional activity of extragonadal androstenedione was the only steroid statistically associated with time to castration resistance in univariate analysis (HR 1.89, 95% CI 1.04-3.44, p=0.036). CONCLUSIONS: Extragonadal steroids contribute significantly to the androgen receptor axis activation at castration levels of testosterone in recurrent nonmetastatic prostate cancer and these sustain the development of castration resistance after primary local treatment.

Glucuronidation of Abiraterone and Its Pharmacologically Active Metabolites by UGT1A4, Influence of Polymorphic Variants and Their Potential as Inhibitors of Steroid Glucuronidation.

Abiraterone (Abi) acetate (AA) is a prodrug of Abi, a CYP17A1 inhibitor used to treat patients with advanced prostate cancer. Abi is a selective steroidal inhibitor that blocks the biosynthesis of androgens. It undergoes extensive biotransformation by steroid pathways, leading to the formation of pharmacologically active Δ4-Abi (D4A) and 5α-Abi. This study aimed to characterize the glucuronidation pathway of Abi and its two active metabolites. We show that Abi, its metabolites, and another steroidal inhibitor galeterone (Gal) undergo secondary metabolism to form glucuronides (G) in human liver microsomes with minor formation by intestine and kidney microsomal preparations. The potential clinical relevance of this pathway is supported by the detection by liquid chromatography-tandem mass spectrometry of Abi-G, D4A-G, and 5α-Abi-G in patients under AA therapy. A screening of UGT enzymes reveals that UGT1A4 is the main enzyme involved. This is supported by inhibition experiments using a selective UGT1A4 inhibitor hecogenin. A number of common and rare nonsynonymous variants significantly abrogate the UGT1A4-mediated formation of Abi-G, D4A-G, and 5α-Abi-G in vitro. We also identify Gal, Abi, and its metabolites as highly potent inhibitors of steroid inactivation by the UGT pathway with submicromolar inhibitor constant values. They reduce the glucuronidation of both the adrenal precursors and potent androgens in human liver, prostate cancer cells, and by recombinant UGTs involved in their inactivation. In conclusion, tested CYP17A1 inhibitors are metabolized through UGT1A4, and germline variations affecting this metabolic pathway may also influence drug metabolism. SIGNIFICANCE STATEMENT: The antiandrogen abiraterone (Abi) is a selective steroidal inhibitor of the cytochrome P450 17α-hydroxy/17,20-lyase, an enzyme involved in the biosynthesis of androgens. Abi is metabolized to pharmacologically active metabolites by steroidogenic enzymes. We demonstrate that Abi and its metabolites are glucuronidated in the liver and that their glucuronide derivatives are detected at variable levels in circulation of treated prostate cancer patients. UDP-glucuronosyltransferase (UGT)1A4 is the primary enzyme involved, and nonsynonymous germline variations affect this metabolic pathway in vitro, suggesting a potential influence of drug metabolism and action in patients. Their inhibitory effect on drug and steroid glucuronidation raises the possibility that these pharmacological compounds might affect the UGT-associated drug-metabolizing system and pre-receptor control of androgen metabolism in patients.

Factors Affecting Interindividual Variability of Hepatic UGT2B17 Protein Expression Examined Using a Novel Specific Monoclonal Antibody.

Accurate quantification of the metabolic enzyme uridine diphospho-glucuronosyltransferase (UGT) UGT2B17 has been hampered by the high sequence identity with other UGT2B enzymes (as high as 94%) and by the lack of a specific antibody. Knowing the significance of the UGT2B17 pathway in drug and hormone metabolism and cancer, we developed a specific monoclonal antibody (EL-2B17mAb), initially validated by the lack of detection in liver microsomes of an individual carrying no UGT2B17 gene copy and in supersomes expressing UGT2B enzymes. Immunohistochemical detection in livers revealed strong labeling of bile ducts and variable labeling of hepatocytes. Expression levels assessed by immunoblotting were highly correlated to mass spectrometry-based quantification (r = 0.93), and three major expression patterns (absent, low, or high) were evidenced. Livers with very low expression were carriers of the functional rs59678213 G variant, located in the binding site for the transcription factor forkhead box A1 (FOXA1) of the UGT2B17 promoter. The highest level of expression was observed for individuals carrying at least one rs59678213 A allele. Multiple regression analysis indicated that the number of gene copies explained only 8% of UGT2B17 protein expression, 49% when adding rs59678213, reaching 54% when including sex. The novel EL-2B17mAb antibody allowed specific UGT2B17 quantification and exposed different patterns of hepatic expression. It further suggests that FOXA1 is a key driver of UGT2B17 expression in the liver. The availability of this molecular tool will help characterize the UGT2B17 level in various disease states and establish more precisely the contribution of the UGT2B17 enzyme to drug and hormone metabolism.

Sex-dependent association of circulating sex steroids and pituitary hormones with treatment-free survival in chronic lymphocytic leukemia patients.

Chronic lymphocytic leukemia (CLL) is not considered a hormone-regulated cancer although sex is a recognized risk factor with men more frequently diagnosed and developing progressive disease. We hypothesized that variable hormonal exposure may have a sexually dimorphic influence on treatment-free survival (TFS). In 156 CLL cases, we quantitatively profiled 29 circulating steroids (progesterone, adrenal precursors, androgens, estrogens, and catechol estrogens) as well as luteinizing hormone (LH) and follicle-stimulating hormone. Median TFS was shorter for men than that for women (80.7 vs. 135.0 months, P = 0.033). Circulating hormone profiles in CLL patients were significantly different from those of healthy donors. In male CLL cases, higher LH levels were associated with shorter TFS (adjusted hazard ratio (HRadj) 2.11; P = 0.004). In female CLL cases, high levels of the potent androgens testosterone and dihydrotestosterone and the sum of methoxy estrogens were associated with an improved TFS with HRadj values of 0.24 (P = 0.007), 0.54 (P = 0.023), and 0.31 (P = 0.034), respectively. Reduced TFS was observed for women with CLL exhibiting high expression of the steroid-inactivating UGT2B17 enzyme. This study is the first to establish a link between the outcome of CLL patients, sex steroids, and pituitary hormones, revealing a sex-specific hormonal imbalance associated with disease progression.

Food systems for sustainable development: proposals for a profound four-part transformation.

Evidence shows the importance of food systems for sustainable development: they are at the nexus that links food security, nutrition, and human health, the viability of ecosystems, climate change, and social justice. However, agricultural policies tend to focus on food supply, and sometimes, on mechanisms to address negative externalities. We propose an alternative. Our starting point is that agriculture and food systems' policies should be aligned to the 2030 Agenda for Sustainable Development. This calls for deep changes in comparison with the paradigms that prevailed when steering the agricultural change in the XXth century. We identify the comprehensive food systems transformation that is needed. It has four parts: first, food systems should enable all people to benefit from nutritious and healthy food. Second, they should reflect sustainable agricultural production and food value chains. Third, they should mitigate climate change and build resilience. Fourth, they should encourage a renaissance of rural territories. The implementation of the transformation relies on (i) suitable metrics to aid decision-making, (ii) synergy of policies through convergence of local and global priorities, and (iii) enhancement of development approaches that focus on territories. We build on the work of the "Milano Group," an informal group of experts convened by the UN Secretary General in Milan in 2015. Backed by a literature review, what emerges is a strategic narrative linking climate, agriculture and food, and calling for a deep transformation of food systems at scale. This is critical for achieving the Sustainable Development Goals and the Paris Agreement. The narrative highlights the needed consistency between global actions for sustainable development and numerous local-level innovations. It emphasizes the challenge of designing differentiated paths for food systems transformation responding to local and national expectations. Scientific and operational challenges are associated with the alignment and arbitration of local action within the context of global priorities.

Evidences of Biological Functions of Biliverdin Reductase A in the Bovine Epididymis.

Epididymal sperm binding protein 1 (ELSPBP1) is secreted by the epididymal epithelium via epididymosomes and is specifically transferred to dead spermatozoa during epididymal transit. We identified biliverdin reductase A (BLVRA) as a partner of ELSPBP1 by immunoprecipitation followed by tandem mass spectrometry. Pull down assays showed that these two proteins interact in the presence of zinc ions. The BLVRA enzyme is known to convert biliverdin to bilirubin, both of which possess antioxidant activity. Assessment by real-time RT-PCR showed that BLVRA is highly expressed in the caput and the corpus epididymis, but is expressed at lower levels in the testis and the cauda epididymis. It is primarily found in the soluble fraction of the caput epididymal fluid, is barely detectable in the cauda fluid, and is detectable to a lesser extent in the epididymosome fraction of both caput and cauda fluids. Immunocytometry on epididymal sperm showed that BLVRA is found on all sperm recovered from the caput region, whereas it is undetectable on cauda sperm. Biliverdin and bilirubin are found in higher concentrations in the caput epididymal fluid, as measured by mass spectrometry. Lipid peroxidation was limited by 1 µM of biliverdin, but not bilirubin when caput spermatozoa were challenged with 500 µM H2O2. Since immature spermatozoa are a source of reactive oxygen species, BLVRA may be involved in the protection of maturing spermatozoa. It is also plausible that BLVRA is implicated in haemic protein catabolism in the epididymal luminal environment.

Association between circulating levels of sex steroid hormones and Barrett's esophagus in men: a case-control analysis.

BACKGROUND & AIMS: Esophageal adenocarcinoma is believed to result from the progression of gastroesophageal reflux disease to erosive esophagitis and re-epithelialization of the esophagus with a columnar cell population termed Barrett's esophagus (BE). Men develop BE and esophageal adenocarcinoma more frequently than women, yet little is known about the mechanisms of this difference. We assessed whether sex steroid hormones were associated with BE in a male population. METHODS: We analyzed data from the Barrett's Esophagus Early Detection Case Control Study, based at the Walter Reed National Military Medical Center. Blood samples were collected from 174 men with BE and 213 men without BE (controls, based on endoscopic analysis); 13 sex steroid hormones were measured by mass spectrometry and sex hormone binding globulin was measured by enzyme-linked immunosorbent assay. We also calculated free estradiol, free testosterone, and free dihydrotestosterone (DHT). We used multivariable logistic regression to estimate odds ratios (ORs) and 95% confidence intervals (CIs) adjusted for age, race, smoking status, alcohol consumption, body mass index, heartburn, regurgitation, and gastroesophageal symptom score (excluding heartburn and regurgitation). RESULTS: Levels of free testosterone and free DHT were associated positively with BE risk; patients in the highest quartile for these hormones were most likely to have BE (free testosterone: OR, 5.36; 95% CI, 2.21-13.03; P = .0002; free DHT: OR, 4.25; 95% CI, 1.87-9.66; P = .001). Level of estrone sulfate was associated inversely with BE risk (P for trend = .02). No other hormone was associated with BE risk. Relationships were not modified by age or BMI. CONCLUSIONS: In an analysis of men, levels of free testosterone and free DHT were significantly associated with BE.

Quantitative profiling of human renal UDP-glucuronosyltransferases and glucuronidation activity: a comparison of normal and tumoral kidney tissues.

Renal metabolism by UDP-glucuronosyltransferase (UGT) enzymes is central to the clearance of many drugs. However, significant discrepancies about the relative abundance and activity of individual UGT enzymes in the normal kidney prevail among reports, whereas glucuronidation in tumoral kidney has not been examined. In this study, we performed an extensive profiling of glucuronidation metabolism in normal (n = 12) and tumor (n = 14) kidneys using targeted mass spectrometry quantification of human UGTs. We then correlated UGT protein concentrations with mRNA levels assessed by quantitative polymerase chain reaction and with conjugation activity for the major renal UGTs. Beyond the wide interindividual variability in expression levels observed among kidney samples, UGT1A9, UGT2B7, and UGT1A6 are the most abundant renal UGTs in both normal and tumoral tissues based on protein quantification. In normal kidney tissues, only UGT1A9 protein levels correlated with mRNA levels, whereas UGT1A6, UGT1A9, and UGT2B7 quantification correlated significantly with their mRNA levels in tumor kidneys. Data support that posttranscriptional regulation of UGT2B7 and UGT1A6 expression is modulating glucuronidation in the kidney. Importantly, our study reveals a significant decreased glucuronidation capacity of neoplastic kidneys versus normal kidneys that is paralleled by drastically reduced UGT1A9 and UGT2B7 mRNA and protein expression. UGT2B7 activity is the most repressed in tumors relative to normal tissues, with a 96-fold decrease in zidovudine metabolism, whereas propofol and sorafenib glucuronidation is decreased by 7.6- and 5.2-fold, respectively. Findings demonstrate that renal drug metabolism is predominantly mediated by UGT1A9 and UGT2B7 and is greatly reduced in kidney tumors.

Multiplexed Targeted Quantitative Proteomics Predicts Hepatic Glucuronidation Potential.

Phase II metabolism is prominently governed by UDP-glucuronosyltransferases (UGTs) in humans. These enzymes regulate the bioactivity of many drugs and endogenous small molecules in many organs, including the liver, a major site of regulation by the glucuronidation pathway. This study determined the expression of hepatic UGTs by targeted proteomics in 48 liver samples and by measuring the glucuronidation activity using probe substrates. It demonstrates the sensitivity and accuracy of nano-ultra-performance liquid chromatography with tandem mass spectrometry to establish the complex expression profiles of 14 hepatic UGTs in a single analysis. UGT2B7 is the most abundant UGT in our collection of livers, expressed at 69 pmol/mg microsomal proteins, whereas UGT1A1, UGT1A4, UGT2B4, and UGT2B15 are similarly abundant, averaging 30-34 pmol/mg proteins. The average relative abundance of these five UGTs represents 81% of the measured hepatic UGTs. Our data further highlight the strong relationships in the expression of several UGTs. Most notably, UGT1A4 correlates with most measured UGTs, and the expression levels of UGT2B4/UGT2B7 displayed the strongest correlation. However, significant interindividual variability is observed for all UGTs, both at the level of enzyme concentrations and activity (coefficient of variation: 45%-184%). The reliability of targeted proteomics quantification is supported by the high correlation between UGT concentration and activity. Collectively, these findings expand our understanding of hepatic UGT profiles by establishing absolute hepatic concentrations of 14 UGTs and further suggest coregulated expression between most abundant hepatic UGTs. Data support the value of multiplexed targeted quantitative proteomics to accurately assess specific UGT concentrations in liver samples and hepatic glucuronidation potential.

Sex Hormones, the Stool Microbiome, and Subclinical Atherosclerosis in Women With and Without HIV.

CONTEXT: Cardioprotective roles of endogenous estrogens may be particularly important in women with HIV, who have reduced estrogen exposure and elevated cardiovascular disease risk. The gut microbiome metabolically interacts with sex hormones, but little is known regarding possible impact on cardiovascular risk. OBJECTIVE: To analyze potential interplay of sex hormones and gut microbiome in cardiovascular risk. METHODS: Among 197 postmenopausal women in the Women's Interagency HIV Study, we measured 15 sex hormones in serum and assessed the gut microbiome in stool. Presence of carotid artery plaque was determined (B-mode ultrasound) in a subset (n = 134). We examined associations of (i) sex hormones and stool microbiome, (ii) sex hormones and plaque, and (iii) sex hormone-related stool microbiota and plaque, adjusting for potential confounders. RESULTS: Participant median age was 58 years and the majority were living with HIV (81%). Sex hormones (estrogens, androgens, and adrenal precursors) were associated with stool microbiome diversity and specific species, similarly in women with and without HIV. Estrogens were associated with higher diversity, higher abundance of species from Alistipes, Collinsella, Erysipelotrichia, and Clostridia, and higher abundance of microbial ß-glucuronidase and aryl-sulfatase orthologs, which are involved in hormone metabolism. Several hormones were associated with lower odds of carotid artery plaque, including dihydrotestosterone, 3α-diol-17G, estradiol, and estrone. Exploratory mediation analysis suggested that estrone-related species, particularly from Collinsella, may mediate the protective association of estrone with plaque. CONCLUSION: Serum sex hormones are significant predictors of stool microbiome diversity and composition. The gut microbiome may play a role in estrogen-related cardiovascular protection.

Menopause and Estrogen Associations With Gut Barrier, Microbial Translocation, and Immune Activation Biomarkers in Women With and Without HIV.

OBJECTIVES: Estrogens may protect the gut barrier and reduce microbial translocation and immune activation, which are prevalent in HIV infection. We investigated relationships of the menopausal transition and estrogens with gut barrier, microbial translocation, and immune activation biomarkers in women with and without HIV. DESIGN: Longitudinal and cross-sectional studies nested in the Women's Interagency HIV Study. METHODS: Intestinal fatty acid binding protein, lipopolysaccharide binding protein, and soluble CD14 (sCD14) levels were measured in serum from 77 women (43 with HIV) before, during, and after the menopausal transition (∼6 measures per woman over ∼13 years). A separate cross-sectional analysis was conducted among 72 postmenopausal women with HIV with these biomarkers and serum estrogens. RESULTS: Women in the longitudinal analysis were a median age of 43 years at baseline. In piecewise, linear, mixed-effects models with cutpoints 2 years before and after the final menstrual period to delineate the menopausal transition, sCD14 levels increased over time during the menopausal transition (Beta [95% CI]: 38 [12 to 64] ng/mL/yr, P = 0.004), followed by a decrease posttransition (-46 [-75 to -18], P = 0.001), with the piecewise model providing a better fit than a linear model (P = 0.0006). In stratified analyses, these results were only apparent in women with HIV. In cross-sectional analyses, among women with HIV, free estradiol inversely correlated with sCD14 levels (r = -0.26, P = 0.03). Lipopolysaccharide binding protein and intestinal fatty acid binding protein levels did not appear related to the menopausal transition and estrogen levels. CONCLUSIONS: Women with HIV may experience heightened innate immune activation during menopause, possibly related to the depletion of estrogens.