Exploratory analysis of human urine by LC–ESI-TOF MS after high intake of olive oil: understanding the metabolism of polyphenols

Olive oil polyphenols have important biological properties which closely depend on their bioavailability, therefore it is essential to understand how polyphenols are absorbed, metabolized and eliminated from the body. An analytical methodology based on rapid resolution liquid chromatography (RRLC) coupled to mass spectrometry detection with a time of flight analyzer (RRLC-ESI-TOF MS) was developed for the determination of the main olive oil phenolic compounds and their metabolites in human urine. Urine samples from ten healthy volunteers were collected before and 2, 4 and 6 h after the intake of 50 mL of extra-virgin olive oil. The proposed method includes liquid-liquid extraction with ethyl acetate that provides extraction recoveries of the phenolic compounds studied between 35 and 75% from spiked urine samples. Good repeatability was obtained, since the relative standard deviations (RSDs) of peak areas in the intra- and inter-day studies were 4.3 and 6.5%, respectively. Statistical studies allowed us to discriminate between the urine samples before and after the intake, and facilitated to find out the m/z values responsible of this discrimination. Based on the very accurate mass information and the isotopic pattern provided by TOF-MS analyzer, together with other available information, ten of these biomarkers and more than 50 metabolites, obtained through phase I and phase II biotransformation reactions, were tentatively identified. Additionally, kinetic studies of the metabolites identified as possible biomarkers were developed, obtaining maximal values in the first two hours for most compounds.The authors are very grateful to Ministry of Education and Science (FPU, AP2005-4356 and Project AGL 2008- 05108-613 CO3-03/ALI), and Junta de Andalucía (P07-AGR-02629 and P09-FQM-5469)

Androgen receptor profiling predicts prostate cancer outcome

Prostate cancer is the second most prevalent malignancy in men. Biomarkers for outcome prediction are urgently needed, so that high-risk patients could be monitored more closely postoperatively. To identify prognostic markers and to determine causal players in prostate cancer progression, we assessed changes in chromatin state during tumor development and progression. Based on this, we assessed genomewide androgen receptor/chromatin binding and identified a distinct androgen receptor/chromatin binding profile between primary prostate cancers and tumors with an acquired resistance to therapy. These differential androgen receptor/chromatin interactions dictated expression of a distinct gene signature with strong prognostic potential. Further refinement of the signature provided us with a concise list of nine genes that hallmark prostate cancer outcome in multiple independent validation series. In this report, we identified a novel gene expression signature for prostate cancer outcome through generation of multilevel genomic data on chromatin accessibility and transcriptional regulation and integration with publically available transcriptomic and clinical datastreams. By combining existing technologies, we propose a novel pipeline for biomarker discovery that is easily implementable in other fields of oncology

APOBEC3B-Mediated Cytidine Deamination Is Required for Estrogen Receptor Action in Breast Cancer.

Estrogen receptor α (ERα) is the key transcriptional driver in a large proportion of breast cancers. We report that APOBEC3B (A3B) is required for regulation of gene expression by ER and acts by causing C-to-U deamination at ER binding regions. We show that these C-to-U changes lead to the generation of DNA strand breaks through activation of base excision repair (BER) and to repair by non-homologous end-joining (NHEJ) pathways. We provide evidence that transient cytidine deamination by A3B aids chromatin modification and remodelling at the regulatory regions of ER target genes that promotes their expression. A3B expression is associated with poor patient survival in ER+ breast cancer, reinforcing the physiological significance of A3B for ER action

From Omics to Multi-Omics Approaches for In-Depth Analysis of the Molecular Mechanisms of Prostate Cancer

Cancer arises following alterations at different cellular levels, including genetic and epigenetic modifications, transcription and translation dysregulation, as well as metabolic variations. High-throughput omics technologies that allow one to identify and quantify processes involved in these changes are now available and have been instrumental in generating a wealth of steadily increasing data from patient tumors, liquid biopsies, and from tumor models. Extensive investigation and integration of these data have led to new biological insights into the origin and development of multiple cancer types and helped to unravel the molecular networks underlying this complex pathology. The comprehensive and quantitative analysis of a molecule class in a biological sample is named omics and large-scale omics studies addressing different prostate cancer stages have been performed in recent years. Prostate tumors represent the second leading cancer type and a prevalent cause of cancer death in men worldwide. It is a very heterogenous disease so that evaluating inter- and intra-tumor differences will be essential for a precise insight into disease development and plasticity, but also for the development of personalized therapies. There is ample evidence for the key role of the androgen receptor, a steroid hormone-activated transcription factor, in driving early and late stages of the disease, and this led to the development and approval of drugs addressing diverse targets along this pathway. Early genomic and transcriptomic studies have allowed one to determine the genes involved in prostate cancer and regulated by androgen signaling or other tumor-relevant signaling pathways. More recently, they have been supplemented by epigenomic, cistromic, proteomic and metabolomic analyses, thus, increasing our knowledge on the intricate mechanisms involved, the various levels of regulation and their interplay. The comprehensive investigation of these omics approaches and their integration into multi-omics analyses have led to a much deeper understanding of the molecular pathways involved in prostate cancer progression, and in response and resistance to therapies. This brings the hope that novel vulnerabilities will be identified, that existing therapies will be more beneficial by targeting the patient population likely to respond best, and that bespoke treatments with increased efficacy will be available soon

Recent Advances in Prostate Cancer Treatment and Drug Discovery

Novel drugs, drug sequences and combinations have improved the outcome of prostate cancer in recent years. The latest approvals include abiraterone acetate, enzalutamide and apalutamide which target androgen receptor (AR) signaling, radium-223 dichloride for reduction of bone metastases, sipuleucel-T immunotherapy and taxane-based chemotherapy. Adding abiraterone acetate to androgen deprivation therapy (ADT) in order to achieve complete androgen blockade has proven highly beneficial for treatment of locally advanced prostate cancer and metastatic hormone-sensitive prostate cancer (mHSPC). Also, ADT together with docetaxel treatment showed significant benefit in mHSPC. Ongoing clinical trials for different subgroups of prostate cancer patients include the evaluation of the second-generation AR antagonists enzalutamide, apalutamide and darolutamide, of inhibitors of the phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) pathway, of inhibitors of DNA damage response, of targeted alpha therapy and of prostate-specific membrane antigen (PSMA) targeting approaches. Advanced clinical studies with immune checkpoint inhibitors have shown limited benefits in prostate cancer and more trials are needed to demonstrate efficacy. The identification of improved, personalized treatments will be much supported by the major progress recently made in the molecular characterization of early- and late-stage prostate cancer using “omics” technologies. This has already led to novel classifications of prostate tumors based on gene expression profiles and mutation status, and should greatly help in the choice of novel targeted therapies best tailored to the needs of patients

Comparative Proteomic and Transcriptomic Analysis of the Impact of Androgen Stimulation and Darolutamide Inhibition

Several inhibitors of androgen receptor (AR) function are approved for prostate cancer treatment, and their impact on gene transcription has been described. However, the ensuing effects at the protein level are far less well understood. We focused on the AR signaling inhibitor darolutamide and confirmed its strong AR binding and antagonistic activity using the high throughput cellular thermal shift assay (CETSA HT). Then, we generated comprehensive, quantitative proteomic data from the androgen-sensitive prostate cancer cell line VCaP and compared them to transcriptomic data. Following treatment with the synthetic androgen R1881 and darolutamide, global mass spectrometry-based proteomics and label-free quantification were performed. We found a generally good agreement between proteomic and transcriptomic data upon androgen stimulation and darolutamide inhibition. Similar effects were found both for the detected expressed genes and their protein products as well as for the corresponding biological programs. However, in a few instances there was a discrepancy in the magnitude of changes induced on gene expression levels compared to the corresponding protein levels, indicating post-transcriptional regulation of protein abundance. Chromatin immunoprecipitation DNA sequencing (ChIP-seq) and Hi-C chromatin immunoprecipitation (HiChIP) revealed the presence of androgen-activated AR-binding regions and long-distance AR-mediated loops at these genes