Elevated tissue expression of hyaluronic acid and hyaluronidase validates the HA-HAase urine test for bladder cancer

Purpose: We examined the expression of 2 bladder tumor markers, hyaluronic acid (HA) and hyaluronidase (HAase), in bladder tissues and correlated tissue staining with the inferences of the HA-HAase urine test, which detects bladder cancer. Materials and Methods: A biotinylated HA binding protein and an antiHYAL1 antibody were used to localize HA and HYAL1 type HAase, respectively, in 83 bladder tissues. Immunoblot analysis was performed using an antiHYAL1 antibody to detect HYAL1. Results: A total of 12 normal bladder tissues showed no (66%) to 1+ (34%) HA staining and 0 (83%) to 1+ (17%) HYAL1 staining. The staining intensity of HA and HYAL1 increased in 71 bladder tumor specimens on chi-square analysis (p <0.001). Grade 1 tumors demonstrated 1+ (50%) to 2+ (50%) staining for HA and 1+ to 3+ staining for HYAL1 (37%, 37% and 26%, respectively). Grades 2 and 3 tumors showed 2+ to 3+ HA (94%) and HYAL1 (79%) staining. HA was expressed in tumor associated stroma and in tumor cells, whereas only tumor cells expressed HYAL1. In bladder tumor tissues HYAL1 expression was confirmed by immunoblot analysis. In 33 of the 34 patients (97%) with bladder cancer from whom urine and tumor tissue specimens were obtained at the same time 2+ to 3+ staining of HA and/or HYAL1 in 12 and 21, respectively, constituted a positive HA-HAase urine test (κ = 0.945). Conclusions: To our knowledge this is the first report of HA localization in bladder tissues and of HYAL1 in any normal or tumor tissue. A close correlation of elevated HA and HYAL1 levels in tumor tissues with a positive HA-HAase urine test indicates that in patients with bladder cancer tumor associated HA and HYAL1 are secreted in urine, causing the HA-HAase test to be positive.