Genomic Network-Based Analysis Reveals Pancreatic Adenocarcinoma Up-Regulating Factor-Related Prognostic Markers in Cervical Carcinoma

Abstract

We previously showed that PAUF is involved in tumor development and metastases in cervical cancer. This study was conducted to discover novel molecular markers linked with PAUF in cervical cancer using genomic network analysis and to assess their prognostic value in cervical cancer. Three PAUF-related genes were identified using in-silico network-based analysis of the open genome datasets. To assess the expression of these genes and their relationship to the outcome of cervical cancer, immunohistochemical analysis was performed using cervical cancer TMA. The associations of the identified proteins with clinicopathologic characteristics and prognosis were examined. AGR2, BRD7, and POM121 were identified as interconnected with PAUF through in-silico network-based analysis. AGR2 (r = 0.213, p < 0.001) and POM121 (r = 0.135, p = 0.013) protein expression were positively correlated with PAUF. BRD7^High and AGR2^Low were significantly associated with favorable disease-free survival (DFS) (p = 0.009 and p < 0.001, respectively), and in combination with PAUF^High, even more significantly favorable DFS observed (p < 0.001 for both). In multivariate analysis, AGR2^High (HR = 3.16, p = 0.01) and BRD7^High (HR = 0.5, p = 0.025) showed independent prognostic value for DFS. In a random survival forest (RSF) model, the combined clinical and molecular variable model predicted DFS with significantly improved power compared with that of the clinical variable model (C-index of 0.79 vs. 0.75, p < 0.001). In conclusion, AGR2 and BRD7 expression have prognostic significance in cervical cancer and provide opportunities for improved treatment options. Genomic network-based approaches using the cBioPortal may facilitate the discovery of additional biomarkers for the prognosis of cervical cancer and may provide new insights into the biology of cervical carcinogenesis.

Keywords: AGR2; BRD7; PAUF; POM121; prognosis; uterine cervical neoplasms.

Figure 1

(A) Flow chart identifying the candidate PAUF-associated prognostic markers. (B) Network view of 22 genes using interactive analysis with cBioPortal network analysis (http://cbioportal.org). Among candidate genes, high expression of AGR2 protein and low expression of BRD7 protein are significantly associated with favorable disease-free survival while the low expression of POM121 protein shows a trend of worse prognosis in patients with cervical cancer.

Figure 2

Expression of the PAUF-associated proteins AGR2, BRD7, and POM121 in patients with cervical cancer. (A) Representative examples of positive and negative expression of the AGR2, BRD7, and POM121 proteins. The scale bar represents 50 μm. (B) Correlations between expression of PAUF and PAUF-related proteins in cervical cancer tissues. AGR2 protein expression was positively correlated with POM121 expression (r = 0.33, p < 0.001).

Figure 3

Hierarchical clustering analysis for immunohistochemical expression of PAUF-associated proteins in cervical cancer patients. (A) Three cluster groups were identified in the present study. Category 1 (n = 118, blue arrow) consists of relatively high AGR2 and POM121 expression. Category 2 (n = 194, red arrow) consists of relatively low AGR2 and PAUF expression. Category 3 (n = 24, green arrow) consists of relatively high PAUF and low POM121 expression. (B) Kaplan–Meier plot for overall survival according to group by hierarchical clustering. A tendency toward decreased overall survival was observed in Category 3 than in the other groups (log-rank test, p = 0.053).

Figure 4

Kaplan–Meier plot of disease-free survival according to expression of PAUF and PAUF-associated proteins. (A,B) Patients with high AGR2 expression (AGR2^High) and low BRD7 expression (BRD7^Low) showed worse disease-free survival (log-rank test, p < 0.001 and p = 0.009, respectively) than patients with low AGR2 expression (AGR2^Low) and high BRD7 expression (BRD7^High). (C) Patients with low POM121 expression (POM121^Low) showed a trend of worse disease-free survival. (D–F) Patients with combined PAUF^High/AGR2^High, PAUF^High/BRD7^Low, or PAUF^High/POM121^Low expression showed shorter disease-free survival (log-rank test, p < 0.001, p < 0.001, and p < 0.001, respectively) than patients with the opposite combined expression profile. (G–J) Patients with combined AGR2^High/BRD7^Low, BRD7^Low/POM121^Low, or AGR2^High/BRD7^Low/POM121^Low expression showed shorter disease-free survival (log-rank test, p < 0.001, p = 0.009, and p = 0.044, respectively) than patients with combined AGR2^Low/BRD7^High, BRD7^High/POM121^High, or AGR2^Low/BRD7^High/POM121^High expression.

Figure 5

Comparison of survival predictive power between the clinical variables and the combined clinical and molecular data. The plots indicate the distribution of C-indexes from 100 rounds of cross-validation. The yellow box highlights the model built from the clinical variables, and the green box highlights the models integrating the molecular variables of PAUF-related proteins and the clinical variables. (A) Using the combined clinical and molecular markers (median C-index 0.79) to predict recurrence yielded improved predictive power compared with the clinical variable model (median C-index 0.74; p < 0.001). (B) The combined clinical/molecular-variable model showed similar performance compared with the clinical variable model in predicting death (p = 0.397). The dashed line indicates the C-index equivalent to a random guess (C-index = 0.5). A C-index of 1 indicates perfect prediction.