The effect of biomarkers with analysis epigenetics expression profile in diagnosis and treatment of prostate cancer

Hamidreza Raeespour,1,* Nafise taromi,2 Sonia daraei,3 Mahdokht forouzan moheb,4 Elham zeinalifard,5 Asma marzanbakhsh,6

1. Gene Pajoohane Ebne Sina genetic research Laboratory
2. Gene Pajoohane Ebne Sina genetic research Laboratory
3. Gene Pajoohane Ebne Sina genetic research Laboratory
4. Gene Pajoohane Ebne Sina genetic research Laboratory
6. Gene Pajoohane Ebne Sina genetic research Laboratory

Abstract

Introduction

Once pca patients eventually relapse and develop castration-resistant pca (crpc), which is more aggressive and incurable, the mortality rate increases significantly. although psa screening has been widely applied for pca diagnosis, there are obvious limitations. psa may increase in bph, leading to a high rate of false-positive diagnosis. thus, there is a need to identify novel non-invasive pca-specific biomarkers in addition to psa detection to improve diagnostic accuracy. long noncoding rnas (lncrnas), which are known to participate in various biological events such as cell differentiation, proliferation and death, are rna transcripts of more than 200 bp without protein coding function. over the past decade, accumulating evidence has demonstrated that lncrnas modulate diverse processes in tumor suppression, metastasis, progression and clinical outcome in multiple types of cancers, including pca. for example, pcat-1 is a transcriptional repressor and, a target of prc2, so promoting cancer cell proliferation. pca3 is a prostate-tissue-specific lncrna that is selectively overexpressed in pca patients compared to healthy individuals. of the imprinting-associated lncrnas, schlap1 has been extensively studied in pca. schlap1 increases with progression of pca and predicts poor prognosis of patients with pca.

Methods

1- we performed a search in pubmed with the following mesh terms: biomarker, prostate cancer, microrna, biomarker prostate cancer, personal medicine 2- the search was narrowed to original articles published in english.

Results

We found 10 publications that analysed the clinical impact of isolation and characterization of exosomes from cell culture supernatant using dualanti body-functionalized immune affinity system exosomes were first isolated from conditioned cell culture medium using a commercial kit. tem pictures showed that the vesicle size of isolated exosomes ranged from 30 to 120 nm in diameter (fig. 1a). the presence of exosomal marker protein cd63 was then analyzed by western blotting (fig. 1b). exosomal marker protein cd63 was enriched in exosome fractions as compared to cell lysates (fig. 1b). to further enrich prostate tumor-derived exosomes, an immune affinity system was developed using magnetic beads conjugated with both anti-epcam and anti-psma antibodies based on our previous related researches (fig. 1c). to validate our isolation procedure, cd63 was probed on the samples from different fractions of the isolation process by western blotting. as shown in fig. 1d, the expression of cd63 for a certain number of total exosomes was pictured in the lane 1. then, the magnetic beads conjugated with anti-epcam and anti-psma antibody could effectively capture exosomes in equal amounts of total exosomes (lane 2 of fig. 1d), whereas there were significantly fewer exosomes in magnetic beads unbound fraction (lane 3 of fig. 1d). to test whether magnetic beads could nonspecifically bind to exosomes, unconjugated magnetic beads were incubated with exosomes, and it was found that magnetic beads alone had minimal capacity to bind to exosomes (lane 4 of fig. 1d), while most exosomes were present in the unbound fraction (lane 5 of fig. 1d). thus, the exosomes captured by conjugated magnetic beads were most likely to bind to dual-antibody as opposed to magnetic beads. based on recent literature and previous work in our laboratory (research in the molecular profiling of pooled circulating tumor cells from pca patients using dual-antibody-functionalized microfluidic device), it was decided to detect two lncrnas in tumor-derived exosomes and validate them in the tcga database (see online suppl. material, fig. s1).

Conclusion

In prostate cancer cells, androgen-regulated transcription factors, including fork head box a1, gata-binding protein 2 and octamer-binding protein 1, are recruited to ar chromosome binding sites. in coordination with ar, the ar-regulated signaling pathway is activated to modulate the overexpression of psa, transmembrane protease serine 2 (tmprss2) and other genes. tmprss2, a transmembrane serine protease, is expressed specifically in the prostate gland. ets transcription factors are important regulators of cell proliferation, differentiation and apoptosis. androgen-regulated ets gene fusions are the most commonly identified genetic alterations and are present in >50% of primary and metastatic prostate cancer cases. we can design a biomarker on a kit that lets us isolate the patient easily from a healthy person.

Keywords

Biomarker, prostate cancer, microrna