(A) Summary of the shotgun proteomics workflow applied to FFPE tumors from ovarian cancer patients. Following tissue lysis and homogenization, purified proteins were digested and analyzed in single-run high-performance liquid chromotography (HPLC)-MS using a quadrupole Orbitrap mass spectrometer. Data were analyzed and quantified in MaxQuant (Cox et al., 2014; Cox and Mann, 2008).
(B) Number of quantified proteins from chemotherapy-resistant (n = 11) and -sensitive (n = 14) tumors. Error bars show standard deviation for each group.
(C) Grouping of chemotherapy-resistant and -sensitive tumor proteomes by PCA. Component 1 accounts for 12.8% of total data variability and component 2 accounts for 9.7%.
(D) Volcano plot of chemotherapy-resistant versus -sensitive tumor proteomes. Expression fold changes are plotted against the t test p value (−log10). Dashed lines indicate the significance threshold (FDR < 0.05, s0 = 2). CT45 is highlighted in green. Chemoresistant patients were defined as those with less than 6 months disease-free survival from the date of last treatment to recurrence, and sensitive patients were defined as those with more than 12 months disease-free survival from the date of last treatment to recurrence.
(E) Immunohistochemistry for CT45 and corresponding H&E staining in serial tumor sections from three representative patients.
(F) Kaplan-Meier survival analysis (log-rank test) of overall survival based on CT45 protein levels in the proteomics dataset. The patients with the highest CT45 expression (top 25%, n = 6, green, median overall survival = 95.4 months) are compared to the lower 75% (n = 19, purple, median overall survival = 35.4 months).
(G) Kaplan-Meier analysis of disease-free survival based on CT45 staining of HGSOC TMAs. Advanced-stage HGSOC patients with a staining score of 0 (n = 82) versus 1+ (n = 42) are compared.
(H) Kaplan-Meier survival analysis based on CT45 RNA levels in 284 HGSOC patients from TCGA (p = 0.0094, log-rank test for trend, IlluminaHiseq BC dataset) (Cancer Genome Atlas Research Network, 2011).
See also Figure S1 and Tables S1, S2, S3, and S4.