Steroidogenic factor 1 (NR5A1) induces multiple transcriptional changes during differentiation of human gonadal-like cells

Peer reviewe

Association of CYP2D6 genotype and tamoxifen metabolites with breast cancer recurrence in a low-dose trial

Low-dose tamoxifen halves recurrence in non-invasive breast cancer without significant adverse events. Some adjuvant trials with tamoxifen 20 mg/day had shown an association between low endoxifen levels (9–16 nM) and recurrence, but no association with CYP2D6 was shown in the NSABP P1 and P2 prevention trials. We studied the association of CYP2D6 genotype and tamoxifen metabolites with tumor biomarkers and recurrence in a randomized phase III trial of low-dose tamoxifen. Median (IQR) endoxifen levels at year 1 were 8.4 (5.3–11.4) in patients who recurred vs 7.5 (5.1–10.2) in those who did not recur (p = 0.60). Tamoxifen and metabolites significantly decreased C-reactive protein (CRP, p < 0.05), and a CRP increase after 3 years was associated with higher risk of recurrence (HR = 4.37, 95% CI, 1.14–16.73, P = 0.03). In conclusion, endoxifen is below 9 nM in most subjects treated with 5 mg/day despite strong efficacy and there is no association with recurrence, suggesting that the reason for tamoxifen failure is not poor drug metabolism.publishedVersio

Low Z-4OHtam concentrations are associated with adverse clinical outcome among early stage premenopausal breast cancer patients treated with adjuvant tamoxifen

Low steady‐state levels of active tamoxifen metabolites have been associated with inferior treatment outcomes. In this retrospective analysis of 406 estrogen receptor‐positive breast cancer (BC) patients receiving adjuvant tamoxifen as initial treatment, we have associated our previously reported thresholds for the two active metabolites, Z‐endoxifen and Z‐4‐hydroxy‐tamoxifen (Z‐4OHtam), with treatment outcomes in an independent cohort of BC patients. Among all patients, metabolite levels did not affect survival. However, in the premenopausal subgroup receiving tamoxifen alone (n = 191) we confirmed an inferior BC ‐specific survival in patients with the previously described serum concentration threshold of Z‐4OHtam ≤ 3.26 nm (HR = 2.37, 95% CI = 1.02–5.48, P = 0.039). The ‘dose–response’ survival trend in patients categorized to ordinal concentration cut‐points of Z‐4OHtamoxifen (≤ 3.26, 3.27–8.13, > 8.13 nm) was also replicated (P‐trend log‐rank = 0.048). Z‐endoxifen was not associated with outcome. This is the first study to confirm the association between a published active tamoxifen metabolite threshold and BC outcome in an independent patient cohort. Premenopausal patients receiving 5‐year of tamoxifen alone may benefit from therapeutic drug monitoring to ensure tamoxifen effectiveness.publishedVersio

Steroidogenic factor 1 (NR5A1) induces multiple transcriptional changes during differentiation of human gonadal-like cells

Nuclear receptor subfamily 5 group A member 1 (NR5A1) encodes steroidogenic factor 1 (SF1), a key regulatory factor that determines gonadal development and coordinates endocrine functions. Here, we have established a stem cell-based model of human gonadal development and applied it to evaluate the effects of NR5A1 during the transition from bipotential gonad to testicular cells. We combined directed differentiation of human induced pluripotent stem cells (46,XY) with activation of endogenous NR5A1 expression by conditionally-inducible CRISPR activation. The resulting male gonadal-like cells expressed several Sertoli cell transcripts, secreted anti-Müllerian hormone and responded to follicle-stimulating hormone by producing sex steroid intermediates. These characteristics were not induced without NR5A1 activation. A total of 2691 differentially expressed genetic elements, including both coding and non-coding RNAs, were detected immediately following activation of NR5A1 expression. Of those, we identified novel gonad-related putative NR5A1 targets, such as SCARA5, which we validated also by immunocytochemistry. In addition, NR5A1 activation was associated with dynamic expression of multiple gonad- and infertility-related differentially expressed genes. In conclusion, by combining targeted differentiation and endogenous activation of NR5A1 we have for the first time, been able to examine in detail the effects of NR5A1 in early human gonadal cells. The model and results obtained provide a useful resource for future investigations exploring the causative reasons for gonadal dysgenesis and infertility in humans.</p

Antioxidant capacity and vasodilatory properties of Mediterranean food: the case of Cannonau wine, myrtle berries liqueur and strawberry-tree honey

The aim of this work was to use different assays to evaluate the antioxidant and vasodilatory properties of three typical food products from the Mediterranean area and to correlate these activities with their phenolic content. For this purpose, red wines Cannonau, liqueurs obtained by cold maceration of myrtle (Myrtus communis L.) berries and bitter honeys obtained from strawberry-tree flowers (Arbutus unedo L.) were analysed. The total phenolic (TP) content was measured spectrophotometrically with a modified Folin–Ciocalteau method and phenolic compounds were identified and dosed by HPLC-DAD and LCMS/MS. Antioxidant activities were evaluated with DPPH, FRAP and ABTS assays and the in vitro vasodilatory effects were assessed using norepinephrine precontracted rat aortic rings. Cannonau wines and myrtle liqueurs showed high levels of TP (1978 ± 279 and 1741 ± 150 mg GAE/L, respectively), linearly correlated to the results of FRAP, ABTS, and DPPH assays. Their maximal vasodilatory activity was 61.7 ± 4.1% and 53.0 ± 3.0%, respectively. Although strawberry-tree honey contained relatively high levels of phenolic compounds (922 ± 38 mg GAE/kg), it did not induce vasodilation, even at the highest dose tested (0.206 g/L). These results indicate that foods with high levels of phenolic compounds should be studied using several different biological assays before being recommended to the general public as functional foods

Floral markers of strawberry tree (<i>Arbutus unedo</i> L.) honey

Strawberry tree honey, due to its characteristic bitter taste, is one of the most typical Mediterranean honeys, with Sardinia being one of the largest producers. According to specific chemical studies, homogentisic acid was identified as a possible marker of this honey. This work, based on HPLC-DAD-MS/MS analysis of strawberry tree (Arbutus unedo L.) honeys, previously selected by sensory evaluation and melissopalynological analysis, showed that, in addition to the above-mentioned acid, there were other high levels of substances useful for the botanical classification of this unifloral honey. Two of these compounds were isolated and identified as (±)-2-cis,4-trans-abscisic acid (c,t-ABA) and (±)-2-trans,4-trans-abscisic acid (t,t-ABA). A third compound, a new natural product named unedone, was characterized as an epoxidic derivative of the above-mentioned acids. Structures of c,t-ABA, t,t-ABA, and unedone were elucidated on the basis of extensive 1D and 2D NMR experiments, as well as HPLC-MS/MS and Q-TOF analysis. In selected honeys the average amounts of c,t-ABA, t,t-ABA, and unedone were 176.2 ± 25.4, 162.3 ± 21.1, and 32.9 ± 7.1 mg/kg, respectively. Analysis of the A. unedo nectar confirmed the floral origin of these compounds found in the honey. Abscisic acids were found in other unifloral honeys but not in such high amount and with a constant ratio of about 1:1. For this reason, besides homogentisic acid, these compounds could be used as complementary markers of strawberry tree honey