Exosomal ephrinA2 derived from serum as a potential biomarker for prostate cancer

Abstract

Up-regulation of serum ephrinA2 is common in various malignancies and has been suggested as a potential biomarker for the diagnosis and prognosis of prostate cancer (PCa). However, the type of serum ephrinA2 expressed in PCa patients remains elusive. Furthermore, the level of exosomal ephrinA2 derived from serum is increased in patients with osteoporosis, a common complication of PCa patients undergoing androgen deprivation therapy. It is unknown whether exosomes derived from PCa patient serum contains ephrinA2. In this study, we explored the ephrinA2 expression in whole serum and tissues and identified the circulating exosomal ephrinA2 as a potential biomarker for PCa. Exosomes were isolated from patient sera by differential centrifugation and the presence of ephrinA2 was confirmed via electron microscopy and western blotting. The type of ephrinA2 in serum was evaluated by western blotting. The expression of serum ephrinA2 including secreted and cleaved ephrinA2 and exosomal ephrinA2 were detected by ELISA and western blotting. Compared with benign prostatic hyperplasia (BPH) and controls, the levels of whole serum ephrinA2 and exosomal ephrinA2 were significantly higher in PCa patients. Moreover, exosomal ephrinA2 expression was positively correlated with TNM staging and Gleason score of PCa patients. The diagnostic efficiency of exosomal ephrinA2 was superior to that of whole serum ephrinA2 and serum PSA in distinguishing PCa patients from those from BPH patents. Our study indicates that exosomal ephrinA2 has high potential as a biomarker for the presence of PCa and offers a new therapeutic target for this disease.

Keywords: EphrinA2; Exosomal ephrinA2; Gleason score; Prostate cancer; TNM staging..

Fig 1

Expression of ephrinA2 in tissues and serum. A. Expression of ephrin A2 protein in 9 pairs of PCa tissues by western blotting (N, adjacent tissue; T, tumor tissue). The result shows the representative western blots in four pairs. B. Expression of ephrinA2 protein in 20 healthy controls, 21 BPH patients and 50 PCa patients measured via ELISA. C. The correlation between exosomal ephrinA2 and TNM staging or Gleason score respectively.

Fig 2

Confirmation of the ultracentrifugation pellets containing exosomes. A. TEM micrographs of exosome isolation by ultracentrifugation. Bar represents 100nm. B. Western blotting for exosomes using exosomal marker CD63.

Fig 3

Expression of ephrinA2 in exosomes and exosome-depleted supernatant respectively. E, exosomes. Sn, exosome-depleted supernatant.

Fig 4

Exosomal ephrinA2 expression in clinical samples. A. Expression of ephrinA2 protein in 20 healthy controls, 21 BPH patients and 50 PCa patients was measured via ELISA. B. The correlation between exosomal ephrinA2 and TNM staging or Gleason score respectively. C. Expression of exosomal ephrinA2 in patients evaluated by western blotting.

Fig 5

ROC curve for ephrinA2 and serum PSA. A. ROC curve analysis of serum ephrinA2 expression in BPH patients versus PCa patients with a specificity of 80.39%, and a sensitivity of 80.59%. B. ROC curve analysis of exosomal ephrinA2 expression in BPH patients versus PCa patients with a specificity of 88% and a sensitivity of 80.59%. C. ROC curve analysis of serum PSA expression in BPH patients versus PCa patients with a specificity of 42.8%, and a sensitivity of 82%.