High-temperature-required protein A2 as a predictive marker for response to chemotherapy and prognosis in patients with high-grade serous ovarian cancers

Abstract

Background: High-temperature-required protein A2 (HtrA2), a protein relating with apoptosis in a caspases-dependent and non-dependent manner, has been reported to be associated with chemosensitivity in several human cancers.

Methods: Tissue microarrays made from 142 patients with high-grade serous ovarian adenocarcinoma were evaluated to assess whether HtrA2 expression was related with several clinical parameters.

Results: Negative HtrA2 expression was observed in 36 cases (25%) of the patients, and related with significantly lower response rates of primary chemotherapy than those with positive HtrA2 expression (56% vs 83%, P<0.01). In addition, negative HtrA2 expression was identified as an independent worse prognostic factor for progression-free survival and overall survival by multivariate analyses. Furthermore, HtrA2 downregulation modulated sensitivity to platinum in serous ovarian cancer cells in vitro.

Conclusions: HtrA2 expression was a predictor for sensitivity to chemotherapy, and could be a candidate of molecular target in the treatment of high-grade serous ovarian cancers.

Figure 1

Immunochemical stains of HtrA2 in tissue microarray-based samples of ovarian serous cell carcinomas ( × 20). Immunohistochemical stains by a rabbit monoclonal antibody for HtrA2 (EPR22; dilution 1 : 100, Abcam) was evaluated. Immunoreactivity was scored according to the staining intensity as follows: weak (0), moderate (1+), and strong (2+). (A) Negative control (NC). (B) Score 0. (C) Score 1+. (D) Score 2+. Scale bar, 100 μm.

Figure 2

Progression-frees survival (PFS) and overall survival (OS) curves of the 142 patients with high-grade serous adenocarcinomas according to expression levels of HtrA2. (A) PFS of the patients with negative HtrA2 expression was significantly worse than that with positive HtrA2 expression (P=0.042). (B) OS of the patients with negative HtrA2 expression was significantly worse than that with positive HtrA2 expression (P<0.01).

Figure 3

HtrA2 protein expression and cisplatin sensitivity. (A) HtrA2 protein concentrations of KF28, KFr13, and MH were analysed by western blotting. (B) HtrA2 protein expression level that was densitometrically normalised against β-actin expression is shown. Blots were scanned and quantified by densitometry. HtrA2 protein concentration was significantly lower in KFr13 and MH compared with that of KF28. MH had significantly lower HtrA2 protein expression in comparison with KFr13. *P<0.01. (C) Cell viability of KF28, KFr13, and MH at the fifth day from cisplatin treatment was evaluated using the MTT assay. IC₅₀ of KFr13 was higher than KF28 (P<0.01), and IC₅₀ of MH was higher than that of KFr13 (P=0.01). The data represented the mean±s.d. of at least three experiments. *P<0.01.

Figure 4

Downregulation of HtrA2 by a specific siRNA decreased the sensitivity to cisplatin in cisplatin-sensitive serous ovarian cancer cells. (A) Downregulation of HtrA2 by a specific siRNA was observed in KF28 cells by western blotting analysis. (B) HtrA2 protein expression level of KF28 cells transfected with non-specific control siRNA (KF28-C), and HtrA2-specific siRNA (KF28-H) were analysed. Blots were scanned and quantified by densitometry. HtrA2 expression level of KF28-H was significantly lower than that of KF28-C. *P<0.01 (C) Cell viability was evaluated by the MTT assay. Five days after transfection with siRNA, sensitivity to cisplatin was measured by 24-h cisplatin treatment, and KF28-H showed resistance to cisplatin treatment. The data represented the mean±s.d. of at least three experiments. *P<0.01.