Label-free cervicovaginal fluid proteome profiling reflects the cervix neoplastic transformation

Cervicovaginal fluid (CVF) is a valuable source of clinical information about the female reproductive tract in both nonpregnant and pregnant women. The aim of this study is to specify the CVF proteome at different stages of cervix neoplastic transformation by label-free quantitation approach based on liquid chromatography tandem mass spectrometry (LC-MS/MS) method. The proteome composition of CVF from 40 women of reproductive age with human papillomavirus (HPV)-associated cervix neoplastic transformation (low-grade squamous intraepithelial lesion [LSIL], high-grade squamous intraepithelial lesion [HSIL], and CANCER) was investigated. Hierarchical clustering and principal component analysis (PCA) of the proteomic data obtained by a label-free quantitation approach show the distribution of the sample set between four major clusters (no intraepithelial lesion or malignancy [NILM], LSIL, HSIL and CANCER) depending on the form of cervical lesion. Multisample ANOVA with subsequent Welch's t test resulted in 117 that changed significantly across the four clinical stages, including 27 proteins significantly changed in cervical cancer. Some of them were indicated as promising biomarkers previously (ACTN4, VTN, ANXA1, CAP1, ANXA2, and MUC5B). CVF proteomic data from the discovery stage were analyzed by the partial least squares-discriminant analysis (PLS-DA) method to build a statistical model, allowing to differentiate severe dysplasia (HSIL and CANCER) from the mild/normal stage (NILM and LSIL), and receiver operating characteristic (ROC) area under the curve (AUC) were obtained on an independent set of 33 samples. The sensitivity of the model was 77%, and the specificity was 94%; AUC was equal to 0.87. CVF proteome proved to be reflect the stage of cervical epithelium neoplastic process.