Proteomic profiling of peritoneal fluid-derived exosomes as a tool for the identification of prognostic markers in advanced ovarian carcinomas.

person Miguel Quiralte Clara Campal Comprehensive Cancer Center, Madrid, Madrid, Spain info_outline Miguel Quiralte, Arantzazu Barquin, Monica Yagüe, Paloma Navarro, Elena Sevillano, Juan Francisco Rodriguez-Moreno, Alejandra Balarezo, Héctor Peinado, Elena Izquierdo, Carlos Millan, Irene Lopez, Sergio Ruiz-Llorente, Jesús García-Donas

Authors person Miguel Quiralte Clara Campal Comprehensive Cancer Center, Madrid, Madrid, Spain info_outline Miguel Quiralte, Arantzazu Barquin, Monica Yagüe, Paloma Navarro, Elena Sevillano, Juan Francisco Rodriguez-Moreno, Alejandra Balarezo, Héctor Peinado, Elena Izquierdo, Carlos Millan, Irene Lopez, Sergio Ruiz-Llorente, Jesús García-Donas Organizations Clara Campal Comprehensive Cancer Center, Madrid, Madrid, Spain, Clara Campal Comprehensive Cancer Center, Madrid, Spain, Clara Campal Cancer Center, Madrid, Spain, HM Sanchinarro Centro Integral Oncologico Clara Campal (CIOCC), Madrid, Spain, HM Hospitales-Centro Integral Oncológico Clara Campal, Madrid, Spain, Centro Integral Oncológico Clara Campal, Madrid, Spain, CNIO, Madrid, Spain, Instituto de Medicina Molecular Aplicada, IMMA, Madrid, Madrid, Spain, Hospital Madrid Sanchinarro, Madrid, Spain Abstract Disclosures Research Funding No funding received None. Background: Exosomes have been involved in tumor progression and development of therapeutic resistances in several malignancies. This subtype of extracellular vesicles (EVs) promote angiogenesis, premetastatic niches formation and communication between the tumor and its microenvironment. Ovarian cancer (OC) is a highly lethal tumor that typically spreads within the abdominal cavity rather than through blood vessels. Based on this behavior, we aimed to study the presence of EVs in ascitic fluid of OC patients and explore their potential as predictors of tumor volume and chemoresistance. Methods: Observational prospective study in advanced OC patients who underwent a diagnostic laparoscopy or cytoreductive surgery. Abdominal washings of a cohort of non-cancer patients were collected as controls. All cases provided written consent. Clinical data and tumor genomic alterations (NGS, FoundationOne CDx) were recorded. EVs pellets were collected by ultracentrifugation and profiled by NTA and western blot (EVs markers: CD9, TSG101 and ALIX; non EVs related markers: ALB and APOB). EVs proteins were quantified by BCA assay and profiled by mass spectrometry. Results: We obtained 75 peritoneal fluids from 66 OC patients (median age at diagnosis [range] 62 [26-83] years) and 29 peritoneal washings from controls. Our cohort comprises various histological subtypes (high grade serous, n 55; low grade serous, n 2; endometrioid, n 6; clear cell, n 2 and mucinous, n 1) and tumor stages (I-II, n 7; III; n 39; IV, n 20). Samples were collected either from primary (n 36); interval, (n 25); or relapse surgery (n 14). Pathogenic BRCA events: 15 patients. Mean concentration of exosomes (MCE) was 3.0x10 6 vs. 1.6x10 6 particles/ml (p/ml) in cases vs. controls (p < 0.0017). No difference in MCE was observed regarding stages I-II vs. III-IV (3.9x10 6 vs. 3.3x10 6 p/ml; p = 0.38). A higher MCE was observed in those samples collected after relapse surgeries compared to those collected from primary (7.2x10 6 vs. 2.8x10 6 p/ml; p < 0.0001) and interval surgeries (7.2x10 6 vs. 3.4x10 6 p/ml; p < 0.0007). Median exosomal protein concentration (MEPC) in EVs of cancer patients vs. controls was 0.99µg/µl and 0.57µg/µl respectively (p = 0.02). Proteomic characterization by mass spectrometry of EVs cargo has identified 1827 proteins, of which 405 proteins were differentially expressed between cases and controls. We also found 38 differentially expressed proteins comparing platinum-sensitive vs. resistant patient samples. Conclusions: Our data show higher concentration of exosomes and exosomal proteins in OC cases vs controls. Differentially expressed proteins have also been found between samples from platinum-sensitive and platinum-resistant patients. These results suggest that exosomes may be involved in the progression and spread of OC and may provide insights into current treatment response.