Project description:Current protocols for the screening of prostate cancer cannot accurately discriminate clinically indolent tumors from more aggressive ones. One reliable indicator of outcome has been the determination of organ-confined versus nonorgan-confined disease but even this determination is often only made following prostatectomy. This underscores the need to explore alternate avenues to enhance outcome prediction of prostate cancer patients. Fluids that are proximal to the prostate, such as expressed prostatic secretions (EPS), are attractive sources of potential prostate cancer biomarkers as these fluids likely bathe the tumor. Direct-EPS samples from 16 individuals with extracapsular (n = 8) or organ-confined (n = 8) prostate cancer were used as a discovery cohort, and were analyzed in duplicate by a nine-step MudPIT on a LTQ-Orbitrap XL mass spectrometer. A total of 624 unique proteins were identified by at least two unique peptides with a 0.2% false discovery rate. A semiquantitative spectral counting algorithm identified 133 significantly differentially expressed proteins in the discovery cohort. Integrative data mining prioritized 14 candidates, including two known prostate cancer biomarkers: prostate-specific antigen and prostatic acid phosphatase, which were significantly elevated in the direct-EPS from the organ-confined cancer group. These and five other candidates (SFN, MME, PARK7, TIMP1, and TGM4) were verified by Western blotting in an independent set of direct-EPS from patients with biochemically recurrent disease (n = 5) versus patients with no evidence of recurrence upon follow-up (n = 10). Lastly, we performed proof-of-concept SRM-MS-based relative quantification of the five candidates using unpurified heavy isotope-labeled synthetic peptides spiked into pools of EPS-urines from men with extracapsular and organ-confined prostate tumors. This study represents the first efforts to define the direct-EPS proteome from two major subclasses of prostate cancer using shotgun proteomics and verification in EPS-urine by SRM-MS.
Project description:Label-free quantitative proteomics was employed to compare the protein content of extracellular vesicles isolated by various differential centrifugation-based approaches from expressed prostatic secretions in urine (EPS-urine) from men with prostate cancer. The developed optimized approach improved EV purity by depleting the high-abundance urine protein Tamm-Horsfall protein (THP) and other common contaminants and achieved relative enrichment of prostate cancer-associated EV-resident proteins.
Project description:Prostate cancer is one of the major cancers that seriously affect men's health. The low specificity of prostate-specific antigen (PSA) for prostate cancer has resulted in the overdiagnosis and subsequent overtreatment of clinically indolent tumors. There is an urgent need for noninvasive and easy diagnostic assays to help evaluate whether a prostate biopsy is warranted. Many non-coding RNAs (eg, microRNAs, long non-coding RNAs, circular RNAs) have been reported to play key roles in prostate cancer progression, showing great potential to impact cancer diagnostics and therapies. Remarkably, exosomes secreted by cells into body fluids contain molecules that reflect the disease information, and urinary exosomes could be used to detect prostate cancer as a new type of liquid biopsies. Non-coding RNAs are enriched and stable in exosomes. We performed high-throughput sequencing on urine-derived exosomes of 11 patients with high-grade (Gleason score 7 or greater) prostate cancer and 11 patients with benign prostatic hyperplasia to screen differentially expressed non-coding RNAs.
Project description:To identify which lncRNAs are differentially expressed in prostate cancer, total RNA from prostate cancer cell line, PC3, and normal epithelial prostatic cells were screened using NCode (Life Technologies) . The NCode microarray is designed to interrogate 12,784 lncRNAs and 25,409 mRNA target protein-coding genes.
Project description:Although an increased level of the prostate-specific antigen can be an indication for prostate cancer, other reasons often lead to a high rate of false positive results. Therefore, an additional serological screening of autoantibodies in patients’ sera could improve the detection of prostate cancer. We performed protein macroarray screening with sera from 49 prostate cancer patients, 70 patients with benign prostatic hyperplasia and 28 healthy controls and compared the autoimmune response in those groups. We were able to distinguish prostate cancer patients from normal controls with an accuracy of 83.2%, patients with benign prostatic hyperplasia from normal controls with an accuracy of 86.0% and prostate cancer patients from patients with benign prostatic hyperplasia with an accuracy of 70.3%. Combining seroreactivity pattern with a PSA level of higher than 4.0 ng/ml this classification could be improved to an accuracy of 84.1%. For selected proteins we were able to confirm the differential expression by using Lluminex on 84 samples. We provide a minimally invasive serological method to reduce false positive results in detection of prostate cancer and according to PSA screening to distinguish men with prostate cancer from men with benign prostatic hyperplasia.
Project description:Research conducted using the novel approach of Next Generation Sequencing to determine the differentially expressed microRNAs in whole blood samples from prostate cancer patients.
Project description:To identify which lncRNAs are differentially expressed in prostate cancer, the total RNA from prostate cancer cell lines (PC3, LNCaP), normal epithelial prostatic cells and the pool of 10 prostate tumor tissues and 10 adjacent normal prostate tissues were screened using SurePrint G3 human lncRNA microarrays (Agilent). The SurePrint G3 Human Gene array contains 16,472 lincRNAs and 34,127 mRNA genes.
Project description:To identify which lncRNAs are differentially expressed in prostate cancer, total RNA from prostate cancer cell line, PC3, and normal epithelial prostatic cells were screened using NCode (Life Technologies) . The NCode microarray is designed to interrogate 12,784 lncRNAs and 25,409 mRNA target protein-coding genes. Gene expression in prostate epithelial cells and PC3 was measured. Three independent experiments (biological replicates) were performed.
