APE1 and SSRP1 is overexpressed in muscle invasive bladder cancer and associated with poor survival

Abstract

Background: Human apurinic/apyrimidinic (AP) endonuclease 1 (APE1) plays a critical role in DNA base excision repair (BER) pathway and has been reported to be overexpressed in multiple cancers. Previously, we have shown that histone chaperone FACT complex (Facilitates Chromatin Transcription, a heterodimer of SSRP1 and SPT16 proteins) facilitates the chromatin access and DNA repair function of APE1, and their expression levels are correlated with promoting drug resistance in cancer. FACT inhibitor has been introduced in phase I and II clinical trials for chemosensitization of advanced solid cancers. However, the expression profile and prognostic significance of APE1 and FACT complex in bladder cancer remains largely unknown.

Methods: Retrospectively, 69 bladder cancer samples were retrieved and submitted for immunohistochemical staining of APE1 and SSRP1. Expression profile including cytoplasmic and nuclear staining of APE1 and expression level of SSRP1 was examined and semi-quantified to render a H-score. The prognostic significance of APE1 and SSRP1 was evaluated by Kaplan-Meier survival analysis in our cohort and R2 database.

Results: APE1 expression is elevated in bladder cancer compared to normal adjacent tissues. Compared with low grade tumors, high grade tumors show a shift in the staining pattern including higher intensity and positive cytoplasmic staining. Carcinoma in situ has a similar staining pattern to high grade tumors. APE1 and SSRP1 staining intensity increases as tumor progresses with stage. There is a correlation between APE1 and SSRP1 staining in invasive bladder cancer (Spearman r = 0.5466, p < 0.0001). The increased expression of APE1 and SSRP1 is associated with poor survival in Kaplan-Meier analysis in our cohort and in R2-TCGA bladder cancer database.

Conclusions: The expression levels of APE1 and SSRP1 are significantly elevated in bladder cancer as compared to normal adjacent tissues. APE1 correlates with SSRP1 expression in high grade tumors. Overexpression of APE1 and SSRP1 is associated with poor survival in bladder cancer. This suggests the usage of FACT inhibitor curaxins in muscle invasive bladder cancer to target FACT complex and APE1 to improve chemosensitization after further validation.

Keywords: APE1; Bladder cancer; SSRP1.

Figure 1

Alteration of APE1 subcellular localization in high grade bladder cancer. A) Immunohistochemical staining of APE1 in primary bladder cancer tissues (original magnification x400). All samples were obtained from tissue bank in UNMC after IRB approval. Representative images of normal urothelium, low-grade tumor, carcinoma in situ and high-grade tumors were shown. B) Box chart of the APE1 nuclear and cytoplasmic immunohistochemical reactivity in normal bladder epithelium, low- and high-grade bladder cancers. Data report the median, 25th and 75th percentiles of APE1 at the nuclear and cytoplasmic levels. ∗p < 0.05.

Figure 2

APE1 correlates with SSRP1 in high grade tumors. A) Tumors at various T stages were stained with APE1 and SSRP1. Representative images were shown. B) & C) H-score of staining intensity was calculated based on percentage of positive staining in nucleus and cytoplasm. ∗∗∗p < 0.001; ∗∗∗∗p < 0.0001. D) H-score of APE1 and SSRP1 of each sample were entered in pairs in Graphpad Prism to perform correlation analysis. Spearman r = 0.5466, p < 0.0001.

Figure 3

Elevated APE1 and SSRP1 expression is associated with poor survival. A) & B) Patients with high grade tumors were divided into 2 groups based on H-score percentile: ≥50 vs < 50 percentile. Kaplan-Meier curves of APE1 and SSRP1 expression were plotted using Graphpad Prism 7.0.

Figure 4

Upregulation of APEX1 and SSRP1 was associated with poor survival in MIBC in R2 genomic database. A) & B) In R2 – TCGA database, survival analysis was performed using pT2, pT3 and pT4 patients regarding APEX1 and SSRP1. This captures only muscle invasive bladder cancer and has 371 (91%) patients in this cohort that has survival data for analysis. C) & D) In R2 – Hoglund database, survival analysis was performed using Ta and T1 patients regarding APEX1 and SSRP1. This captures only non-muscle invasive bladder cancer and has 213 (69%) patients in this cohort. Final analysis contains 173 (56%) patients with survival data available for analysis.