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BackgroundControversies exist as to whether the genetic polymorphisms of the enzymes responsible for the metabolism of tamoxifen can predict breast cancer outcome in patients using adjuvant tamoxifen. Direct measurement of concentrations of active tamoxifen metabolites in serum may be a more biological plausible and robust approach. We have investigated the association between CYP2D6 genotypes, serum concentrations of active tamoxifen metabolites, and long-term outcome in tamoxifen treated breast cancer patients.MethodsFrom an original observational study comprising 817 breast cancer patients, 99 women with operable breast cancer were retrospectively included in the present study. This cohort of patients were adjuvantly treated with tamoxifen, had provided serum samples suitable for measuring tamoxifen metabolites, and were relapse-free at 3 years after the primary treatment commenced. The median follow-up time from this entry point to breast cancer death was 13.9 years. Patients were CYP2D6 genotyped and grouped into four CYP2D6 phenotype groups (Ultra rapid, extensive, intermediate, and poor metabolizers). Tamoxifen and nine metabolites were quantified in serum (n = 86) and compared with CYP2D6 phenotype groups and outcome.ResultsBreast cancer patients with low concentrations of Z-4-hydroxy-tamoxifen (Z-4OHtam; ≤ 3.26 nM) had a breast cancer-specific survival (BCSS) of 60% compared to 84% in patients with Z-4OHtam concentrations > 3.26 nM (p = 0.020, log-rank hazard ratio (HR) = 3.56, 95% confidence interval (CI) = 1.14–11.07). For patients with Z-4-hydroxy-N-desmethyl-tamoxifen (Z-endoxifen) levels ≤ 9.00 nM BCSS was 57% compared to 84% for patients with concentrations > 9.00 nM (p = 0.029, HR = 3.73, 95% CI = 1.05–13.22). Low concentrations of Z-4OHtam and Z-endoxifen were associated with poorer survival also after adjusting for clinically relevant variables (HR = 4.27, 95% CI = 1.35–13.58, and HR = 3.70, 95% CI = 1.03–13.25, respectively). Overall survival analysis showed similar survival differences for both active metabolites. The Antiestrogen Activity Score showed comparable effects, but did not improve the prognostic information.ConclusionsPatients with Z-4OHtam and Z-endoxifen concentrations lower than 3.26 nM or 9.00 nM, respectively, showed an adverse outcome. Our results suggest that direct measurement of active tamoxifen metabolite concentrations could be of clinical value. Validation in larger study cohorts is warranted.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-017-0916-4) contains supplementary material, which is available to authorized users.

During the liquid chromatographic study of the phenolic fraction of monofloral honeys was detected in the asphodel honey ( Asphodelus microcarpus Salzm. et Viv.) chromatogram a distinctive peak not detected in other monofloral honeys such as Arbutus unedo L., Hedysarum coronarium , Eucalyptus spp., and Galactites tomentosa . After thin layer chromatography (TLC) purification and characterization by NMR and LC-MS/MS, the compound was identified as methyl syringate (MSYR) and confirmed against an original standard. Levels of MSYR were measured in honeys of 2005, 2006, and 2007 by HPLC-DAD analysis. Level determination of MSYR was repeated in 2008 for 2006 and 2007 honeys to evaluate chemical stability of this phenolic compound. Levels of MSYR measured 1 year after the sampling did not show significant statistical differences (p < 0.05). The stability of MSYR was also confirmed by 12 asphodel honey samples collected in 2005 that showed amounts of methyl syringate comparable with those found in fresh honey. For the evaluation of MSYR origin, samples of nectars were collected from flowers and the content of MSYR was measured. Levels of MSYR in honeys are originated from the nectar with an average contribution of the nectar to the honey of 80%. Melissopalinological analysis did not allow the attribution of the honey monofloral origin because levels of asphodel pollen were <6% for all analyzed samples. Previously reported levels of MSYR for robinia, rape, chestnut, clover, linden blossom, dandelion, sunflower, thyme, manuka, and fir honeys were <5 mg/kg. For this reason, a minimum level of 122.6 mg/kg for MSYR in asphodel honeys can be considered as a chemical marker and, unlike the melissopalynological analysis, can be used for the origin attribution and to evaluate the percent of asphodel nectar in the 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.

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