Background and Methods: PCOS is common and associated with significant comorbidity. However, its pathogenesis is complex and poorly understood. We have developed new methods for deep phenotyping discovery proteomic profiling of urine, aiming to identify disease mechanisms, drug targets and novel non-invasive biomarkers for PCOS in adolescents. Here, we present the baseline proteomic data from our prospective, longitudinal study (n=40). We undertook proteomic analysis (nano 2D-LC-QTOF MSe) on a subset of 15 samples. We compared the urinary proteome of adolescents with PCOS (n=6), controls (n=6), and insulin resistance (IR) (n=3), to identify markers of PCOS as distinct from those of IR.
Results: We identified 3,793 proteins, of which, 314 were significantly and differentially expressed (P<0.05) in the PCOS cohort in comparison to healthy controls, and 397 in the PCOS vs. IR analysis. Of these, 66 proteins overlapped and were differentially expressed in the PCOS cohort in comparison to both controls and IR, and as such are potential novel biomarkers for PCOS. These proteins had a median fold-change of -2.2 (range -16.6 – +1775.5) and eight were upregulated in the PCOS cohort. Gene ontology analysis revealed these proteins were mediators of complement, coagulation and apoptosis cascades, glucose homeostasis, pro-inflammatory cytokines, and cytoskeletal components, the latter important in folliculogenesis. Bioinformatic Ingenuity Pathway Analysis (IPA) of all significant proteins identified differential activation of 23 significant biological pathways in the PCOS cohort. The top five canonical pathways were the coagulation system, the intrinsic prothrombin activation pathway, 14-3-3-mediated signalling, remodelling of epithelial adherens junctions, and acute phase response signalling. Almost half of all significant pathways (10/23; 43%) related to inflammatory/immunological responses and the thrombotic/fibrinolytic systems. Nearly a quarter (n=5/53; 22%) were associated with glucose homeostasis/insulin resistance, hyperandrogenism, and neuroendocrine pathways. Whilst, one third (8/23; 35%) were involved in folliculogenesis, cytoskeleton reorganisation, remodelling, apoptosis and autophagy. IPA diseases and function analysis revealed the inflammatory response as the most significant biological process associated with PCOS in our study (P<0.001).
Conclusions: To the best of our knowledge, this is the first study to utilise non-invasive matrices to map the proteome of PCOS in adolescents. We have identified 66 potential novel biomarkers, provided promising insight into the molecular pathways of PCOS, and demonstrated that inflammation could be a major contributory factor in the pathophysiology of this common condition. To confirm these findings, we are performing targeted multiplexed assays comprising 70 inflammatory proteins on our entire longitudinal cohort.
22 Sep 2021 - 26 Sep 2021