A Preliminary Proteomic Investigation of Circulating Exosomes and Discovery of Biomarkers Associated with the Progression of Osteosarcoma in a Clinical Model of Spontaneous Disease

Abstract

Circulating cancer exosomes are microvesicles which originate from malignant cells and other organs influenced by the disease and can be found in blood. The exosomal proteomic cargo can often be traced to the cells from which they originated, reflecting the physiological status of these cells. The similarities between cancer exosomes and the tumor cells they originate from exhibit the potential of these vesicles as an invaluable target for liquid biopsies. Exosomes were isolated from the serum of eight osteosarcoma-bearing dogs, five healthy dogs, and five dogs with traumatic fractures. We also characterized exosomes which were collected longitudinally from patients with osteosarcoma prior and 2 weeks after amputation, and eventually upon detection of lung metastasis. Exosomal proteins fraction were analyzed by label-free mass spectrometry proteomics and were validated with immunoblots of selected proteins. Ten exosomal proteins were found that collectively discriminate serum of osteosarcoma patients from serum healthy or fractured dogs with an accuracy of 85%. Additionally, serum from different disease stages could be distinguished with an accuracy of 77% based on exosomal proteomic composition. The most discriminating protein changes for both sample group comparisons were related to complement regulation, suggesting an immune evasion mechanism in early stages of osteosarcoma as well as in advanced disease.

The Emerging Roles of Extracellular Vesicles in Osteosarcoma

“…As far as EV-associated proteins are concerned, circulating levels of EV-associated TGF-β have been found to be increased in osteosarcoma patients, when compared to healthy control subjects (27). A proteomic investigation of circulating EVs in canine serum samples identified EV related proteins useful to distinguish serum of osteosarcoma from serum of healthy or fractured dogs (72). Furthermore, Brady et al identified two proteins associated with EVs (serpinD1 and MHC class III-complement C6) which allow to discriminate serum derived from different disease stages of OS (72).…”

Exosomal communication by metastatic osteosarcoma cells modulates alveolar macrophages to an M2 tumor-promoting phenotype and inhibits tumoricidal functions

“…22 In osteosarcoma, exosomes have been shown to be related to migration, invasion, and tumor progression, but the complex interplay between osteosarcoma cells and macrophages has yet to be investigated. [23][24][25] Because a tumor-promoting microenvironment contains M2 macrophages, and M2 macrophages are linked to poor outcomes in patients with osteosarcoma lung metastases, we compared the extracellular communications between metastatic and non-metastatic osteosarcoma cells and alveolar macrophages. It was our hypothesis that metastatic osteosarcoma cells help to create a tumor-permissive environment by exosomal communication.…”

Evaluation of nucleosome concentrations in healthy dogs and dogs with cancer

“…Though there are no published studies specifically describing cancer detection using nucleosomes in dogs, several publications have described the utility of cfDNA (13)(14)(15)(16)(17).…”