(1) Background: Prostate cancer (PCa) is the most frequently diagnosed cancer in men. Wide application of prostate specific antigen test has historically led to over-treatment, starting from excessive biopsies. Risk calculators based on molecular and clinical variables can be of value to determine the risk of PCa and as such, reduce unnecessary and invasive biopsies. Urinary molecular studies have been mostly focusing on sampling after initial intervention (digital rectal examination and/or prostate massage). (2) Methods: Building on previous proteomics studies, in this manuscript, we aimed at developing a biomarker model for PCa detection based on urine sampling without prior intervention. Capillary electrophoresis coupled to mass spectrometry was applied to acquire proteomics profiles from 970 patients from two different clinical centers. (3) Results: A case-control comparison was performed in a training set of 413 patients and 181 significant peptides were subsequently combined by a support vector machine algorithm. Independent validation was initially performed in 272 negative for PCa and 138 biopsy-confirmed PCa, resulting in an AUC of 0.81, outperforming current standards, while a second validation phase included 147 PCa patients. (4) Conclusions: This multi-dimensional biomarker model holds promise to improve the current diagnosis of PCa, by guiding invasive biopsies.

Prostate cancer (PCa) is the second most common cancer in men. Diagnosis and risk assessment are widely based on serum Prostate Specific Antigen (PSA) and biopsy, which might not represent the exact degree of PCa risk. Towards the discovery of biomarkers for better patient stratification, we performed proteomic analysis of Formalin Fixed Paraffin Embedded (FFPE) prostate tissue specimens using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Comparative analysis of 86 PCa samples among grade groups 1–5 identified 301 significantly altered proteins. Additional analysis based on biochemical recurrence (BCR; BCR+ n = 14, BCR- n = 51) revealed 197 significantly altered proteins that indicate disease persistence. Filtering the overlapping proteins of these analyses, seven proteins (NPM1, UQCRH, HSPA9, MRPL3, VCAN, SERBP1, HSPE1) had increased expression in advanced grades and in BCR+/BCR- and may play a critical role in PCa aggressiveness. Notably, all seven proteins were significantly associated with progression in Prostate Cancer Transcriptome Atles (PCTA) and NPM1NPM1, UQCRH, and VCAN were further validated in The Cancer Genome Atlas (TCGA), where they were upregulated in BCR+/BCR-. UQCRH levels were also associated with poorer 5-year survival. Our study provides valuable insights into the key regulators of PCa progression and aggressiveness. The seven selected proteins could be used for the development of risk assessment tools.

Background Empagliflozin (EMPA), Dapagliflozin (DAPA) and Ertugliflozin (ERTU) are selective sodium glucose co-transporter 2 inhibitors (SGLT2i) acting against type 2 diabetes mellitus. Purpose Due to differences in clinical trial outcomes, we aimed to 1) compare the cardioprotective effects of selective SGLT2i in terms of infarct size (IS) reduction and 2) reveal the mechanism of cardioprotection in non-diabetic mice. Methods C57BL/6 mice were randomized and orally received EMPA (10mg/kg/day), DAPA (9.0mg/kg/day), ERTU (9.7mg/kg/day) or vehicle for 7 days. IS was measured after 30' ischemia (I), and 120' reperfusion (R). EMPA, DAPA and ERTU were given at equivalent stoichiometrically doses (ESD). Body weight and fasting blood glucose (FBG) levels were determined at baseline and at the end of the treatment. On the 7th day, mice were housed in metabolic cages for 24 hours. Urine volume (UV), food and water uptake and 24h-glucose levels were determined to examine the extend of SGLT-2 inhibition by the drugs. In a second series, the ischemic myocardium was taken (10'R), shotgun proteomics were performed and several cardioprotective pathways were evaluated. In a third series, the dominant pathways were evaluated through molecular analyses and mitochondrial functionality. The causal relationships in the mechanism of protection, was established by inhibiting the concomitant cardioprotective pathways. Static, the specific STAT-3 inhibitor and wortmannin (a PI3K inhibitor) were administered and IS was measured upon 30'I/120' R. Results EMPA and DAPA but not ERTU reduced IS at this dose. Body weight and FBG levels were not affected by the treatments. EMPA, DAPA and ERTU lead to significant increase in UV and urinary glucose levels compared to the control group independently of the water and food intake. There was no significant difference in the parameters among the different SGLT-2i indicating that the chosen doses are sufficient to produce the same pharmacological SGLT-2 inhibition in mice. Proteomics revealed mitochondrial metabolism and NF-kB signaling as significant. Only EMPA preserved mitochondrial functionality in complex I & II linked oxidative phosphorylation. NF-kB, RISK and STAT-3 activation and the downstream reduction in apoptosis were evident in EMPA and DAPA groups coinciding with IS reduction. Static and wortmannin significantly attenuated IS reduction both in EMPA and DAPA groups indicating that STAT-3 and PI3K activation are the leading mechanisms of cardioprotection. Among several upstream mediators, fibroblast growth factor 2 (FGF-2) and caveolin-3 were increased in EMPA and DAPA groups. Conclusions Short term EMPA, DAPA and ERTU at the chosen ESD inhibit SGLT-2i in a similar extent but only EMPA and DAPA reduce IS. Our study reveals drug specific effects on cardioprotection against I/R injury. Cardioprotection afforded by EMPA and DAPA are STAT-3 and PI3K dependent and associated with increased FGF-2 and Cav-3 expression. Funding Acknowledgement Type of funding sources: None.

Defective Complement activation has been associated with various types of kidney disease. This led to the hypothesis that specific urine complement fragments may be associated with kidney disease etiologies, and disease progression may be reflected by changes in these complement fragments. Therefore, we investigated the occurrence of complement fragments in urine, their association with kidney function and disease etiology. Mass spectrometry based peptidomics data from the Human Urinary Proteome/Peptidome Database were extracted and the distribution of complement peptides in the different kidney disease etiologies and controls was investigated. All datasets where information on disease/health status was available and at least one complement peptide could be detected were included (n=16027). Twenty-three different urinary peptides derived from complement proteins could be identified, originating from the complement proteins C3, C4 and CFB. For most C3-derived peptides an inverse association with eGFR was observed, while the majority of peptides derived from CFB demonstrated positive association with eGFR. Highest levels of significant C3 excretion relative to controls were seen in minimal change disease (MCD), focal segmental glomerulosclerosis (FSGS), membranous glomerulonephritis (MGN), lupus nephritis (LN), diabetic kidney disease (DKD), IgAN, membranoproliferative glomerulonephritis (MPGN), and C3-glomerulonephritis. In conclusion, several peptides derived from the complement proteins C3, C4 and Factor B are significantly associated with specific kidney disease etiologies. These peptides may depict disease-specific complement activation, as well as damage to the glomerular basement membrane. Further investigation of these complement peptides may provide additional insight into disease pathophysiology and could possibly guide therapeutic decisions, especially when targeting complement factors.

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