Predicting immune checkpoint therapy response in three independent metastatic melanoma cohorts.

While Immune checkpoint inhibition (ICI) therapy shows significant efficacy in metastatic melanoma, only about 50% respond, lacking reliable predictive methods. We introduce a panel of six proteins aimed at predicting response to ICI therapy. Evaluating previously reported proteins in two untreated melanoma cohorts, we used a published predictive model (EaSIeR score) to identify potential proteins distinguishing responders and non-responders. Six proteins initially identified in the ICI cohort correlated with predicted response in the untreated cohort. Additionally, three proteins correlated with patient survival, both at the protein, and at the transcript levels, in an independent immunotherapy treated cohort. Our study identifies predictive biomarkers across three melanoma cohorts, suggesting their use in therapeutic decision-making.

Identifying Ferroptosis Inducers, HDAC, and RTK Inhibitor Sensitivity in Melanoma Subtypes through Unbiased Drug Target Prediction.

The utilization of PD1 and CTLA4 inhibitors has revolutionized the treatment of malignant melanoma (MM). However, resistance to targeted and immune-checkpoint-based therapies still poses a significant problem. Here we mine large scale MM proteogenomic data integrating it with MM cell line dependency screen, and drug sensitivity data to identify druggable targets and forecast treatment efficacy and resistance. Leveraging protein profiles from established MM subtypes and molecular structures of 82 cancer treatment drugs, we identified nine candidate hub proteins, mTOR, FYN, PIK3CB, EGFR, MAPK3, MAP4K1, MAP2K1, SRC and AKT1, across five distinct MM subtypes. These proteins serve as potential drug targets applicable to one or multiple MM subtypes. By analyzing transcriptomic data from 48 publicly accessible melanoma cell lines sourced from Achilles and CRISPR dependency screens, we forecasted 162 potentially targetable genes. We also identified genetic resistance in 260 genes across at least one melanoma subtype. In addition, we employed publicly available compound sensitivity data (Cancer Therapeutics Response Portal, CTRPv2) on the cell lines to assess the correlation of compound effectiveness within each subtype. We have identified 20 compounds exhibiting potential drug impact in at least one melanoma subtype. Remarkably, employing this unbiased approach, we have uncovered compounds targeting ferroptosis, that demonstrate a striking 30x fold difference in sensitivity among different subtypes. This implies that the proteogenomic classification of melanoma has the potential to predict sensitivity to ferroptosis compounds. Our results suggest innovative and novel therapeutic strategies by stratifying melanoma samples through proteomic profiling, offering a spectrum of novel therapeutic interventions and prospects for combination therapy. Highlights: (1) Proteogenomic subtype classification can define the landscape of genetic dependencies in melanoma (2) Nine proteins from molecular subtypes were identified as potential drug targets for specified MM patients (3) 20 compounds identified that show potential effectiveness in at least one melanoma subtype (4) Proteogenomics can predict specific ferroptosis inducers, HDAC, and RTK Inhibitor sensitivity in melanoma subtypes.

An inflamed subtype of difficult-to-treat depression.

BACKGROUND: Chronic low-grade inflammation may play a role in the pathophysiology of depression, at least in a subset of patients. High-sensitivity C-reactive protein (hs-CRP) has been used to define an inflamed subgroup of depression with specific clinical characteristics and symptoms. In this study we investigated biochemical and clinical characteristics in patients with difficult-to-treat depression with and without chronic low-grade inflammation. METHOD: We assayed plasma levels of interferon-gamma, tumor necrosis factor-alpha, Interleukin (IL)-10, IL-6, IL-8, and vitamin D in a clinically well-characterized sample of patients with difficult-to-treat depression (n = 263) and healthy controls (n = 46). Serum hs-CRP levels were available in the patient group and were used to define "inflamed depression" (hs-CRP > 3 mg/L). Based on previous studies correlating specific depressive symptoms to inflammatory markers, we calculated a composite score of inflammatory depressive symptoms (Infl-Dep score). A principal component analysis (PCA) was performed to identify patterns of variance in cytokines and vitamin D among patients. RESULTS: Mean levels of IL-6 and IL-8 were significantly higher in depressed patients compared to controls, also after adjusting for sex, smoking, BMI, and age. None of the other inflammatory markers differed significantly between depressed patients and controls. Two components were extracted using PCA; one showed general cytokine elevations and one represented a pattern where IL-6 and IL-8 were inversely related to vitamin D (IL6-IL8-VitD component). The inflamed subgroup (hs-CRP > 3, n = 51) exhibited significantly higher BMI, higher Infl-Dep scores and higher IL6-IL8-VitD component scores than uninflamed patients (hs-CRP ≤ 3, n = 212). There were no significant differences in overall depression severity or suicidality between the inflamed and uninflamed groups. CONCLUSION: Our results support the hypothesis of an inflamed subgroup of depression as a meaningful construct. This subgroup may have certain biological and clinical characteristics and more studies are needed to determine potential clinical implications.

Maternal serum vitamin D level in early pregnancy and risk for preeclampsia: A case-control study in Southern Sweden.

BACKGROUND: Preeclampsia is considered a major cause of maternal and fetal morbidity and mortality. The aim of the present case-control study in Sweden was to assess the hypothesized association between low serum vitamin D concentrations in early pregnancy and the risk of developing preeclampsia since vitamin D may play a role in early placental development. METHODS: The study included 296 women diagnosed with preeclampsia (cases) and 580 healthy pregnant women (controls). Serum samples were obtained from a biobank of samples collected in early pregnancy including almost all pregnancies in Southern Sweden. Concentrations of 25-hydroxyvitamin D3 (vitamin D) were analyzed using liquid chromatography-tandem-mass-spectrometry (LC/MS/MS). The cases were divided into two categories: i) infants were born before gestational week 34 (early onset) and/or born small-for-gestational age (SGA)(n = 51), ii) and others defined as late onset (n = 245). Vitamin D concentrations were analyzed both as a continuous and a categorized variable. RESULTS: When all preeclampsia cases were included in the analyses no consistent patterns were observed. However, the median serum concentrations of vitamin D were significantly lower among the cases who were early onset and/or were born SGA (median 39.2 nmol/L, range 1.2-93.6) as compared to the controls (49.0 nmol/L, 0.1-219; p = 0.01). In addition, high concentrations were statistically significantly associated with a decreased risk of preeclampsia (>66.9 vs ≤30.1 nmol/L; crude OR 0.39, 95% CI 0.16-0.96). When potential confounders were included in the models the associations were even more pronounced. CONCLUSIONS: Our results support the hypothesis that vitamin D deficiency is a risk factor for preeclampsia, but only in preeclampsia cases who were early-onset and/or were born SGA. Preeclampsia is not a homogenous condition and more studies are needed before vitamin D supplementation during pregnancy can be recommended.

Circulating vitamin D and breast cancer risk: an international pooling project of 17 cohorts.

Laboratory and animal research support a protective role for vitamin D in breast carcinogenesis, but epidemiologic studies have been inconclusive. To examine comprehensively the relationship of circulating 25-hydroxyvitamin D [25(OH)D] to subsequent breast cancer incidence, we harmonized and pooled participant-level data from 10 U.S. and 7 European prospective cohorts. Included were 10,484 invasive breast cancer cases and 12,953 matched controls. Median age (interdecile range) was 57 (42-68) years at blood collection and 63 (49-75) years at breast cancer diagnosis. Prediagnostic circulating 25(OH)D was either newly measured using a widely accepted immunoassay and laboratory or, if previously measured by the cohort, calibrated to this assay to permit using a common metric. Study-specific relative risks (RRs) for season-standardized 25(OH)D concentrations were estimated by conditional logistic regression and combined by random-effects models. Circulating 25(OH)D increased from a median of 22.6 nmol/L in consortium-wide decile 1 to 93.2 nmol/L in decile 10. Breast cancer risk in each decile was not statistically significantly different from risk in decile 5 in models adjusted for breast cancer risk factors, and no trend was apparent (P-trend = 0.64). Compared to women with sufficient 25(OH)D based on Institute of Medicine guidelines (50- < 62.5 nmol/L), RRs were not statistically significantly different at either low concentrations (< 20 nmol/L, 3% of controls) or high concentrations (100- < 125 nmol/L, 3% of controls; ≥ 125 nmol/L, 0.7% of controls). RR per 25 nmol/L increase in 25(OH)D was 0.99 [95% confidence intervaI (CI) 0.95-1.03]. Associations remained null across subgroups, including those defined by body mass index, physical activity, latitude, and season of blood collection. Although none of the associations by tumor characteristics reached statistical significance, suggestive inverse associations were seen for distant and triple negative tumors. Circulating 25(OH)D, comparably measured in 17 international cohorts and season-standardized, was not related to subsequent incidence of invasive breast cancer over a broad range in vitamin D status.

Plasma metabolome study reveals metabolic changes induced by pharmacological castration and testosterone supplementation in healthy young men.

Testosterone is a hormone that plays a key role in carbohydrate, fat, and protein metabolism. Testosterone deficiency is associated with multiple comorbidities, e.g., metabolic syndrome and type 2 diabetes. Despite its importance in many metabolic pathways, the mechanisms by which it controls metabolism are not fully understood. The present study investigated the short-term metabolic changes of pharmacologically induced castration and, subsequently, testosterone supplementation in healthy young males. Thirty subjects were submitted to testosterone depletion (TD) followed by testosterone supplementation (TS). Plasma samples were collected three times corresponding to basal, low, and restored testosterone levels. An untargeted metabolomics study was performed by liquid chromatography-high resolution mass spectrometry (UHPLC-HRMS) to monitor the metabolic changes induced by the altered hormone levels. Our results demonstrated that TD was associated with major metabolic changes partially restored by TS. Carnitine and amino acid metabolism were the metabolic pathways most impacted by variations in testosterone. Furthermore, our results also indicated that LH and FSH might strongly alter the plasma levels of indoles and lipids, especially glycerophospholipids and sphingolipids. Our results demonstrated major metabolic changes induced by low testosterone that may be important for understanding the mechanisms behind the association of testosterone deficiency and its comorbidities.

Lipid Metabolic Reprogramming Extends beyond Histologic Tumor Demarcations in Operable Human Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is among the deadliest malignancies and potentially curable only with radical surgical resection at early stages. The tumor microenvironment has been shown to be central to the development and progression of PDAC. A better understanding of how early human PDAC metabolically communicates with its environment and differs from healthy pancreas could help improve PDAC diagnosis and treatment. Here we performed deep proteomic analyses from diagnostic specimens of operable, treatment-naïve PDAC patients (n = 14), isolating four tissue compartments by laser-capture microdissection: PDAC lesions, tumor-adjacent but morphologically benign exocrine glands, and connective tissues neighboring each of these compartments. Protein and pathway levels were compared between compartments and with control pancreatic proteomes. Selected targets were studied immunohistochemically in the 14 patients and in additional tumor microarrays, and lipid deposition was assessed by nonlinear label-free imaging (n = 16). Widespread downregulation of pancreatic secretory functions was observed, which was paralleled by high cholesterol biosynthetic activity without prominent lipid storage in the neoplastic cells. Stromal compartments harbored ample blood apolipoproteins, indicating abundant microvasculature at the time of tumor removal. The features best differentiating the tumor-adjacent exocrine tissue from healthy control pancreas were defined by upregulation of proteins related to lipid transport. Importantly, histologically benign exocrine regions harbored the most significant prognostic pathways, with proteins involved in lipid transport and metabolism, such as neutral cholesteryl ester hydrolase 1, associating with shorter survival. In conclusion, this study reveals prognostic molecular changes in the exocrine tissue neighboring pancreatic cancer and identifies enhanced lipid transport and metabolism as its defining features. SIGNIFICANCE: In clinically operable pancreatic cancer, regions distant from malignant cells already display proteomic changes related to lipid transport and metabolism that affect prognosis and may be pharmacologically targeted.

Impact of add-back FSH on human and mouse prostate following gonadotropin ablation by GnRH antagonist treatment.

Objective: During androgen ablation in prostate cancer by the standard gonadotropin-releasing hormone (GnRH) agonist treatment, only luteinizing hormone (LH) is permanently suppressed while circulating follicle-stimulating hormone (FSH) rebounds. We explored direct prostatic effects of add-back FSH, after androgen ablation with GnRH antagonist, permanently suppressing both gonadotropins. Methods: The effects of recombinant human (rFSH) were examined in mice treated with vehicle (controls), GnRH antagonist degarelix (dgx), dgx + rFSH, dgx + flutamide, or dgx + rFSH + flutamide for 4 weeks. Prostates and testes size and expression of prostate-specific and/or androgen-responsive genes were measured. Additionally, 33 young men underwent dgx-treatment. Seventeen were supplemented with rFSH (weeks 1-5), and all with testosterone (weeks 4-5). Testosterone, gondotropins, prostate-specific antigen (PSA), and inhibin B were measured. Results: In dgx and dgx + flutamide treated mice, prostate weight/body weight was 91% lower than in controls, but 41 and 11%, respectively, was regained by rFSH treatment (P = 0.02). The levels of seminal vesicle secretion 6, Pbsn, Nkx3.1, beta-microseminoprotein, and inhibin b were elevated in dgx + rFSH-treated animals compared with only dgx treated (all P < 0.05). In men, serum inhibin B rose after dgx treatment but was subsequently suppressed by testosterone. rFSH add-back had no effect on PSA levels. Conclusions: These data provide novel evidence for the direct effects of FSH on prostate size and gene expression in chemically castrated mice. However, in chemically castrated men, FSH had no effect on PSA production. Whether FSH effects on the prostate in humans also require suppression of the residual adrenal-derived androgens and/or a longer period of rFSH stimulation, remains to be explored.

Proteomic Alterations in Follicular Fluid of Human Small Antral Follicles Collected from Polycystic Ovaries-A Pilot Study.

Polycystic ovaries (PCO) contain antral follicles that arrest growing around 3-11 mm in diameter, perturbing the dominant follicle's selection and the subsequent ovulatory process. Proteomic alterations of PCO follicular fluid (FF) (i.e., microenvironment in which the oocyte develops until ovulation) have been studied from large follicles in connection with oocyte pickup during ovarian stimulation. The present study aimed to detect proteomic alterations in FF from unstimulated human small antral follicles (hSAF) obtained from PCO. After performing deep-sequencing label-free proteomics on 10 PCO and 10 non-PCO FF samples from unstimulated hSAF (4.6-9.8 mm), 1436 proteins were identified, of which 115 were dysregulated in PCO FF samples. Pathways and processes related to the immune system, inflammation, and oxidative stress appeared to be upregulated in PCO, while extracellular matrix receptors interactions, the collagens-containing extracellular matrix, and the regulation of signaling were downregulated. The secreted proteins SFRP1, THBS4, and C1QC significantly decreased their expression in PCO FF, and this downregulation was suggested to affect future oocyte competence. In conclusion, our study revealed, for the first time, evidence of proteomic alterations occurring in the FF of PCO hSAF that may be related to the dysfunction of follicular growth and subsequent oocyte competence.

Novel protein markers of androgen activity in humans: proteomic study of plasma from young chemically castrated men.

BACKGROUND: Reliable biomarkers of androgen activity in humans are lacking. The aim of this study was, therefore, to identify new protein markers of biological androgen activity and test their predictive value in relation to low vs normal testosterone values and some androgen deficiency linked pathologies. METHODS: Blood samples from 30 healthy GnRH antagonist treated males were collected at three time points: (1) before GnRH antagonist administration; (2) 3 weeks later, just before testosterone undecanoate injection, and (3) after additional 2 weeks. Subsequently, they were analyzed by mass spectrometry to identify potential protein biomarkers of testosterone activity. Levels of proteins most significantly associated with testosterone fluctuations were further tested in a cohort of 75 hypo- and eugonadal males suffering from infertility. Associations between levels of those markers and cardiometabolic parameters, bone mineral density as well as androgen receptor (AR) CAG repeat lengths, were explored. RESULTS: Using receiver operating characteristic analysis, 4-hydroxyphenylpyruvate dioxygenase (4HPPD), insulin-like growth factor-binding protein 6 (IGFBP6), and fructose-bisphosphate aldolase (ALDOB), as well as a Multi Marker Algorithm, based on levels of 4HPPD and IGFBP6, were shown to be best predictors of low (<8 nmol/l) vs normal (>12 nmol/l) testosterone. They were also more strongly associated with metabolic syndrome and diabetes than testosterone levels. Levels of ALDOB and 4HPPD also showed association with AR CAG repeat lengths. CONCLUSIONS: We identified potential new protein biomarkers of testosterone action. Further investigations to elucidate their clinical potential are warranted. FUNDING: The work was supported by ReproUnion2.0 (grant no. 20201846), which is funded by the Interreg V EU program.

Although it is best known for its role in developing male sex organs and maintaining sexual function, the hormone testosterone is important for many parts of the human body. A deficiency can cause an increased risk of serious conditions such as diabetes, cancer and osteoporosis. Testosterone deficiency can develop due to disease or age-related changes, and men affected by this can be given supplements of this hormone to restore normal levels. The most common way to test for testosterone deficiency is by measuring the concentration of the hormone in the blood. However, this does not accurately reflect the activity of the hormone in the body. This may lead to men who need more testosterone not receiving enough, and to others being unnecessarily treated. Several factors may lead to discrepancy between testosterone concentration in blood and its physiological activity. One of the most common is obesity. Additionally, certain genetic factors, which cannot be controlled for yet, regulate sensitivity to this hormone: some people do well at low levels, while others need high concentrations to be healthy. Therefore, to improve the diagnosis of testosterone deficiency it is necessary to identify biological markers whose levels act as a proxy for testosterone activity. Giwercman, Sahlin et al. studied the levels of a large number of proteins in the blood of 30 young men before and after blocking testosterone production. The analysis found three proteins whose concentrations changed significantly after testosterone deprivation. Giwercman, Sahlin et al. then validated these markers for testosterone deficiency by checking the levels of the three proteins in a separate group of 75 men with fertility problems. The results also showed that the three protein markers were better at predicting diabetes and metabolic syndrome than testosterone levels alone. These newly discovered markers could be used to create a test for measuring testosterone activity. This could help to identify deficiencies and finetune the amount of supplementary hormone given to men as treatment. However, further research is needed to understand the clinical value of such a test in men, as well as women and children.

An Observational Study on the Molecular Profiling of Primary Melanomas Reveals a Progression Dependence on Mitochondrial Activation.

Melanoma in advanced stages is one of the most aggressive tumors and the deadliest of skin cancers. To date, the histopathological staging focuses on tumor thickness, and clinical staging is a major estimate of the clinical behavior of primary melanoma. Here we report on an observational study with in-depth molecular profiling at the protein level including post-translational modifications (PTMs) on eleven primary tumors from melanoma patients. Global proteomics, phosphoproteomics, and acetylomics were performed on each sample. We observed an up-regulation of key mitochondrial functions, including the mitochondrial translation machinery and the down-regulation of structural proteins involved in cell adhesion, the cytoskeleton organization, and epidermis development, which dictates the progression of the disease. Additionally, the PTM level pathways related to RNA processing and transport, as well as chromatin organization, were dysregulated in relation to the progression of melanoma. Most of the pathways dysregulated in this cohort were enriched in genes differentially expressed at the transcript level when similar groups are compared or metastasis to primary melanomas. At the genome level, we found significant differences in the mutation profiles between metastatic and primary melanomas. Our findings also highlighted sex-related differences in the molecular profiles. Remarkably, primary melanomas in women showed higher levels of antigen processing and presentation, and activation of the immune system response. Our results provide novel insights, relevant for developing personalized precision treatments for melanoma patients.

Deep Proteomic Analysis on Biobanked Paraffine-Archived Melanoma with Prognostic/Predictive Biomarker Read-Out.

The discovery of novel protein biomarkers in melanoma is crucial. Our introduction of formalin-fixed paraffin-embedded (FFPE) tumor protocol provides new opportunities to understand the progression of melanoma and open the possibility to screen thousands of FFPE samples deposited in tumor biobanks and available at hospital pathology departments. In our retrospective biobank pilot study, 90 FFPE samples from 77 patients were processed. Protein quantitation was performed by high-resolution mass spectrometry and validated by histopathologic analysis. The global protein expression formed six sample clusters. Proteins such as TRAF6 and ARMC10 were upregulated in clusters with enrichment for shorter survival, and proteins such as AIFI1 were upregulated in clusters with enrichment for longer survival. The cohort's heterogeneity was addressed by comparing primary and metastasis samples, as well comparing clinical stages. Within immunotherapy and targeted therapy subgroups, the upregulation of the VEGFA-VEGFR2 pathway, RNA splicing, increased activity of immune cells, extracellular matrix, and metabolic pathways were positively associated with patient outcome. To summarize, we were able to (i) link global protein expression profiles to survival, and they proved to be an independent prognostic indicator, as well as (ii) identify proteins that are potential predictors of a patient's response to immunotherapy and targeted therapy, suggesting new opportunities for precision medicine developments.

Mapping the Melanoma Plasma Proteome (MPP) Using Single-Shot Proteomics Interfaced with the WiMT Database.

Plasma analysis by mass spectrometry-based proteomics remains a challenge due to its large dynamic range of 10 orders in magnitude. We created a methodology for protein identification known as Wise MS Transfer (WiMT). Melanoma plasma samples from biobank archives were directly analyzed using simple sample preparation. WiMT is based on MS1 features between several MS runs together with custom protein databases for ID generation. This entails a multi-level dynamic protein database with different immunodepletion strategies by applying single-shot proteomics. The highest number of melanoma plasma proteins from undepleted and unfractionated plasma was reported, mapping >1200 proteins from >10,000 protein sequences with confirmed significance scoring. Of these, more than 660 proteins were annotated by WiMT from the resulting ~5800 protein sequences. We could verify 4000 proteins by MS1t analysis from HeLA extracts. The WiMT platform provided an output in which 12 previously well-known candidate markers were identified. We also identified low-abundant proteins with functions related to (i) cell signaling, (ii) immune system regulators, and (iii) proteins regulating folding, sorting, and degradation, as well as (iv) vesicular transport proteins. WiMT holds the potential for use in large-scale screening studies with simple sample preparation, and can lead to the discovery of novel proteins with key melanoma disease functions.

Short-Term Effect of Induced Alterations in Testosterone Levels on Fasting Plasma Amino Acid Levels in Healthy Young Men.

Long term effect of testosterone (T) deficiency impairs metabolism and is associated with muscle degradation and metabolic disease. The association seems to have a bidirectional nature and is not well understood. The present study aims to investigate the early and unidirectional metabolic effect of induced T changes by measuring fasting amino acid (AA) levels in a human model, in which short-term T alterations were induced. We designed a human model of 30 healthy young males with pharmacologically induced T changes, which resulted in three time points for blood collection: (A) baseline, (B) low T (3 weeks post administration of gonadotropin releasing hormone antagonist) and (C) restored T (2 weeks after injection of T undecanoate). The influence of T on AAs was analyzed by spectrophotometry on plasma samples. Levels of 9 out of 23 AAs, of which 7 were essential AAs, were significantly increased at low T and are restored upon T supplementation. Levels of tyrosine and phenylalanine were most strongly associated to T changes. Short-term effect of T changes suggests an increased protein breakdown that is restored upon T supplementation. Fasting AA levels are able to monitor the early metabolic changes induced by the T fluctuations.

The Human Melanoma Proteome Atlas-Complementing the melanoma transcriptome.

The MM500 meta-study aims to establish a knowledge basis of the tumor proteome to serve as a complement to genome and transcriptome studies. Somatic mutations and their effect on the transcriptome have been extensively characterized in melanoma. However, the effects of these genetic changes on the proteomic landscape and the impact on cellular processes in melanoma remain poorly understood. In this study, the quantitative mass-spectrometry-based proteomic analysis is interfaced with pathological tumor characterization, and associated with clinical data. The melanoma proteome landscape, obtained by the analysis of 505 well-annotated melanoma tumor samples, is defined based on almost 16 000 proteins, including mutated proteoforms of driver genes. More than 50 million MS/MS spectra were analyzed, resulting in approximately 13,6 million peptide spectrum matches (PSMs). Altogether 13 176 protein-coding genes, represented by 366 172 peptides, in addition to 52 000 phosphorylation sites, and 4 400 acetylation sites were successfully annotated. This data covers 65% and 74% of the predicted and identified human proteome, respectively. A high degree of correlation (Pearson, up to 0.54) with the melanoma transcriptome of the TCGA repository, with an overlap of 12 751 gene products, was found. Mapping of the expressed proteins with quantitation, spatiotemporal localization, mutations, splice isoforms, and PTM variants was proven not to be predicted by genome sequencing alone. The melanoma tumor molecular map was complemented by analysis of blood protein expression, including data on proteins regulated after immunotherapy. By adding these key proteomic pillars, the MM500 study expands the knowledge on melanoma disease.

RUBIC (ReproUnion Biobank and Infertility Cohort): A binational clinical foundation to study risk factors, life course, and treatment of infertility and infertility-related morbidity.

BACKGROUND: Infertility affects 15%-25% of all couples during their reproductive life span. It is a significant societal and public health problem with potential psychological, social, and economic consequences. Furthermore, infertility has been linked to adverse long-term health outcomes. Despite the advanced diagnostic and therapeutic techniques available, approximately 30% of infertile couples do not obtain a live birth after fertility treatment. For these couples, there are no further options to increase their chances of a successful pregnancy and live birth. OBJECTIVES: Three overall questions will be studied: (1) What are the risk factors and natural life courses of infertility, early embryonic loss, and adverse pregnancy outcomes? (2) Can we develop new diagnostic and prognostic biomarkers for fecundity and treatment success? And (3) what are the health characteristics of women and men in infertile couples at the time of fertility treatment and during long-term follow-up? MATERIAL AND METHODS: ReproUnion Biobank and Infertility Cohort (RUBIC) is established as an add-on to the routine fertility management at Copenhagen University Hospital Departments in the Capital Region of Denmark and Reproductive Medicine Centre at Skåne University Hospital in Sweden. The aim is to include a total of 5000 couples equally distributed between Denmark and Sweden. The first patients were enrolled in June 2020. All eligible infertile couples are prospectively asked to participate in the project. Participants complete an extensive questionnaire and undergo a physical examination and collection of biospecimens (blood, urine, hair, saliva, rectal swabs, feces, semen, endometrial biopsies, and vaginal swabs). After the cohort is established, the couples will be linked to the Danish and Swedish national registers to obtain information on parental, perinatal, childhood, and adult life histories, including disease and medication history. This will enable us to understand the causes of infertility and identify novel therapeutic options for this important societal problem.

Apelin promotes blood and lymph vessel formation and the growth of melanoma lung metastasis.

Apelin, a ligand of the APJ receptor, is overexpressed in several human cancers and plays an important role in tumor angiogenesis and growth in various experimental systems. We investigated the role of apelin signaling in the malignant behavior of cutaneous melanoma. Murine B16 and human A375 melanoma cell lines were stably transfected with apelin encoding or control vectors. Apelin overexpression significantly increased melanoma cell migration and invasion in vitro, but it had no impact on its proliferation. In our in vivo experiments, apelin significantly increased the number and size of lung metastases of murine melanoma cells. Melanoma cell proliferation rates and lymph and blood microvessel densities were significantly higher in the apelin-overexpressing pulmonary metastases. APJ inhibition by the competitive APJ antagonist MM54 significantly attenuated the in vivo pro-tumorigenic effects of apelin. Additionally, we detected significantly elevated circulating apelin and VEGF levels in patients with melanoma compared to healthy controls. Our results show that apelin promotes blood and lymphatic vascularization and the growth of pulmonary metastases of skin melanoma. Further studies are warranted to validate apelin signaling as a new potential therapeutic target in this malignancy.

Serum amyloid P component: a new biomarker for low sperm concentration?

Serum amyloid P component (SAP) is present in seminal plasma, on spermatozoa, and in different tissues of the male reproductive tract, but its function is not known. The aims of this study were to determine if the concentration of SAP in seminal plasma is associated with commonly assessed semen parameters and to investigate if SAP could be a new, indirect biomarker for these parameters. In a cross-sectional study of 203 young volunteers, the concentration of SAP in seminal plasma was measured with a in-house developed enzyme-linked immunosorbent assay. Scatter plots, Pearson's correlation coefficients (r), and linear regression models were produced, and SAP showed a statistically significant correlation with sperm concentration (r = 0.75), sperm number (r = 0.68), semen volume (r = -0.19), progressive sperm motility (r = 0.24), and sperm immotility (r = -0.20). When the study group was dichotomized, SAP could be used to discriminate samples with a sperm concentration < or ≥5 × 106 ml-1, 15 × 106 ml-1, or 40 × 106 ml-1, and in receiver operating characteristic curves, the corresponding areas under the curves were 0.97, 0.93, and 0.82, respectively, with P < 0.001 for all three cutoff values studied. The concentration of SAP in seminal plasma showed a strong, positive correlation with the concentration of spermatozoa in semen. SAP may be used as a new indirect potential biomarker for sperm concentration in fresh and in frozen, stored samples. In addition, it is envisaged that the assay could be developed into a home fertility test to differentiate between a low and a normal sperm concentration.