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Polycystic ovary syndrome (PCOS) is the most common endocrine disorder in premeno-pausal women, with a wide spectrum of possible phenotypes, symptoms and sequelae according to the current clinical definition. However, there are women who do not fulfill at least two out of the three commonly used “Rotterdam criteria” and their risk of developing type 2 diabetes or obesity later in life is not defined. Therefore, we addressed this important gap by conducting a retrospective analysis based on 750 women with and without PCOS. We compared four different PCOS pheno-types according to the Rotterdam criteria with women who exhibit only one Rotterdam criterion and with healthy controls. Hormone and metabolic differences were assessed by analysis of vari-ance (ANOVA) as well as logistic regression analysis. We found that hyperandrogenic women have per se a higher risk of developing insulin resistance compared to phenotypes without hyperandro-genism and healthy controls. In addition, hyperandrogenemia is associated with developing insulin resistance also in women with no other Rotterdam criterion. Our study encourages further diagnostic and therapeutic approaches for PCOS phenotypes in order to account for varying risks of developing metabolic diseases. Finally, women with hyperandrogenism as the only symptom should also be screened for insulin resistance to avoid later metabolic risks.

Novel androgen receptor (AR) signaling inhibitors have improved the treatment of castration-resistant prostate cancer (CRPC). Nonetheless, the effect of these drugs is often time-limited and eventually most patients become resistant due to various AR alterations. Although liquid biopsy approaches are powerful tools for early detection of such therapy resistances, most assays investigate only a single resistance mechanism. In combination with the typically low abundance of circulating biomarkers, liquid biopsy assays are therefore informative only in a subset of patients. In this pilot study, we aimed to increase overall sensitivity for tumor-related information by combining three liquid biopsy approaches into a multi-analyte approach. In a cohort of 19 CRPC patients, we (1) enumerated and characterized circulating tumor cells (CTCs) by mRNA-based in situ padlock probe analysis, (2) used RT-qPCR to detect cancer-associated transcripts (e.g., AR and AR-splice variant 7) in lysed whole blood, and (3) conducted shallow whole-genome plasma sequencing to detect AR amplification. Although 44&ndash, 53% of patient samples were informative for each assay, a combination of all three approaches led to improved diagnostic sensitivity, providing tumor-related information in 89% of patients. Additionally, distinct resistance mechanisms co-occurred in two patients, further reinforcing the implementation of multi-analyte liquid biopsy approaches.

The 25-Hydroxyvitamin D (25[OH)D) serum concentration depends on vitamin D intake, endogenous vitamin D production and genetic factors. The latter have been demonstrated in large genome-wide association studies indicating that single nucleotide polymorphisms (SNPs) in genes related to the vitamin D metabolism are as important for serum 25(OH)D levels as the influence of season. The mechanism on how these SNPs influence serum 25(OH)D levels are still unclear. The aim of the present study was to investigate the genetic effects of ten selected SNPs related to vitamin D metabolism on 25-hydroxyvitamin D increase (∆25(OH)D) after vitamin D supplementation in three randomized controlled trials. Genotypes of SNPs related to vitamin D metabolism were determined in 411 participants with 25(OH)D concentrations &lt, 75 nmol/l receiving 20,000 IU cholecalciferol per week for 8 or 12 weeks after study inclusion. For the vitamin D receptor (VDR) rs10783219 polymorphism, the minor A-allele was associated with lower ∆25(OH)D values in the entire study population (p = 0.022), which was not consistent in all three cohorts when analysed separately. VDR rs10783219 might therefore be a genetic modulator of increasing 25-hydroxyvitamin D concentrations. Considering the wide-spread use of vitamin D supplementation, future large and well-designed randomized controlled trials (RCTs) should investigate the clinical impact of this polymorphism.

Background: Polycystic ovary syndrome (PCOS) affects 5–20% of women of reproductive age worldwide and is associated with disorders of glucose metabolism. Hormone and metabolic signaling may be influenced by phytoestrogens, such as isoflavones. Their endocrine effects may modify symptom penetrance in PCOS. Equol is one of the most active isoflavone metabolites, produced by intestinal bacteria, and acts as a selective estrogen receptor modulator. Method: In this interventional study of clinical and biochemical characterization, urine isoflavone levels were measured in PCOS and control women before and three days after a defined isoflavone intervention via soy milk. In this interventional study, bacterial equol production was evaluated using the log(equol: daidzein ratio) and microbiome, metabolic, and predicted metagenome analyses were performed. Results: After isoflavone intervention, predicted stool metagenomic pathways, microbial alpha diversity, and glucose homeostasis in PCOS improved resembling the profile of the control group at baseline. In the whole cohort, larger equol production was associated with lower androgen as well as fertility markers. Conclusion: The dynamics in our metabolic, microbiome, and predicted metagenomic profiles underline the importance of external phytohormones on PCOS characteristics and a potential therapeutic approach or prebiotic in the future.

BACKGROUND Liquid biopsies can be used in castration-resistant prostate cancer (CRPC) to detect androgen receptor splice variant 7 (AR-V7), a splicing product of the androgen receptor. Patients with AR-V7-positive circulating tumor cells (CTCs) have greater benefit of taxane chemotherapy compared with novel hormonal therapies, indicating a treatment-selection biomarker. Likewise, in those with pancreatic cancer (PaCa), KRAS mutations act as prognostic biomarkers. Thus, there is an urgent need for technology investigating the expression and mutation status of CTCs. Here, we report an approach that adds AR-V7 or KRAS status to CTC enumeration, compatible with multiple CTC-isolation platforms. METHODS We studied 3 independent CTC-isolation devices (CellCollector, Parsortix, CellSearch) for the evaluation of AR-V7 or KRAS status of CTCs with in situ padlock probe technology. Padlock probes allow highly specific detection and visualization of transcripts on a cellular level. We applied padlock probes for detecting AR-V7, androgen receptor full length (AR-FL), and prostate-specific antigen (PSA) in CRPC and KRAS wild-type (wt) and mutant (mut) transcripts in PaCa in CTCs from 46 patients. RESULTS In situ analysis showed that 71% (22 of 31) of CRPC patients had detectable AR-V7 expression ranging from low to high expression [1–76 rolling circle products (RCPs)/CTC]. In PaCa patients, 40% (6 of 15) had KRAS mut expressing CTCs with 1 to 8 RCPs/CTC. In situ padlock probe analysis revealed CTCs with no detectable cytokeratin expression but positivity for AR-V7 or KRAS mut transcripts. CONCLUSIONS Padlock probe technology enables quantification of AR-V7, AR-FL, PSA, and KRAS mut/wt transcripts in CTCs. The technology is easily applicable in routine laboratories and compatible with multiple CTC-isolation devices.

Liquid biopsies can be used in castration-resistant prostate cancer (CRPC) to detect androgen receptor splice variant 7 (AR-V7), a splicing product of the androgen receptor. Patients with AR-V7-positive circulating tumor cells (CTCs) have greater benefit of taxane chemotherapy compared with novel hormonal therapies, indicating a treatment-selection biomarker. Likewise, in those with pancreatic cancer (PaCa),We studied 3 independent CTC-isolation devices (CellCollector, Parsortix, CellSearch) for the evaluation of AR-V7 or KRAS status of CTCs with in situ padlock probe technology. Padlock probes allow highly specific detection and visualization of transcripts on a cellular level. We applied padlock probes for detecting AR-V7, androgen receptor full length (AR-FL), and prostate-specific antigen (PSA) in CRPC andIn situ analysis showed that 71% (22 of 31) of CRPC patients had detectable AR-V7 expression ranging from low to high expression [1-76 rolling circle products (RCPs)/CTC]. In PaCa patients, 40% (6 of 15) hadPadlock probe technology enables quantification of AR-V7, AR-FL, PSA, and

Although tumor genotyping is still the most currently used method for categorizing tumors for clinical decisions, tumor tissues provide only a snapshot or are often difficult to obtain. To overcome these issues, methods are needed for a rapid, cost-effective, and noninvasive identification of biomarkers at various time points during the course of disease. The analysis of circulating tumor cells (CTCs), cell-free circulating tumor DNA (ctDNA), circulating RNAs, and exosomes, frequently referred to as liquid biopsy, has recently gained enormous momentum. Due to technological advances, novel circulating tumor biomarkers were shown to have a great potential to improve patient treatment in terms of estimation of prognosis, monitoring treatment response, early detection of resistance mechanisms, identification of actionable targets, and detection of minimal residual disease. However, despite all efforts, liquid biopsies are not yet routinely used mainly due to technological hurdles, lack of analytical and pre-analytical standards and conclusive evidence that patients indeed benefit from such analyses. In this chapter, the different entities with respect to state-of-the-art technologies, potential clinical applications, and their limitations are discussed.

Osteoporosis and diabetes mellitus type 2 (T2DM) are frequently coinciding in elderly patients, increasing with age. The disease is associated with a higher fracture risk despite normal to high planar bone mineral density (BMD) as measured by dual energy bone densitometry (DXA) in these individuals. As BMD measurements do not identify T2DM patients at risk for bone fractures, this coincidence has not been recognized for a long time, though higher mortality and the risk of a delayed wound healing have been found in T2DM patients with major fractures. In patients with diabetes mellitus type 1 (T1DM) fracture risk is even higher, with an up to 12-fold risk as compared to patients with normal glucose metabolism. These patients might be affected from early childhood on, with worse fracture outcomes depending on their glucose control. Pathophysiological conditions that might contribute to the increased fracture risk in both T1DM and T2DM include decreased skeletal quality and properties, aggravated by comorbidities such as macro- and microvascular complications, and chronic inflammation, but there are many open questions to resolve. As specific diagnostic approaches are urgently required, several attempts have been made by new imaging technologies, namely trabecular bone score (TBS), high resolution peripheral quantitative computed tomography (HRpQCT), bone biopsies, histomorphometry and microindentation. New biomarkers of diabetoporosity have been addressed. The complex interaction of these new biomarkers is illustrated by several articles within this issue. We report on the current knowledge in pathophysiology, diagnostics and management of bone in diabetes mellitus. Given the increasing numbers of T2DM and osteoporosis patients worldwide—with a current number of about 415 million T2DM patients, a growing community of T1DM patients and about 9 million osteoporotic fractures annually in 200 million women and probably 120 million men—this is in fact an important problem for future health care.