Flow Cytometric Analysis of Nucleoside Transporters Activity in Chemoresistant Prostate Cancer Model

Klin Onkol. 2018 Spring;31(Supplementum1):140-144.

Abstract

Background: Nucleoside analogues represent a relevant class of antimetabolites used for therapy of various types of cancer. However, their effectivity is limited by drug resistance. The nucleoside transport capability of tumour cells is considered to be a determinant of the clinical outcome of treatment regimens using antimetabolites. Due to hydrophilic properties of antimetabolites, their transport across the plasma membrane is mediated by two families of transmembrane proteins, the SLC28 family of cation-linked concentrative nucleoside transporters (hCNTs) and SLC29 family of energy-independent equilibrative nucleoside transporters (hENTs). Loss of functional nucleoside transporters has been associated with reduced efficacy of antimetabolites and their derivatives and treatment failure in diverse malignancies including solid tumours, such as pancreatic adenocarcinoma.

Material and methods: The effectivity and kinetics of antimetabolite uptake were analysed using control and docetaxel-resistant PC3 cells. For this purpose, fluorescent nucleoside analogue probe uridine-furane and inhibitor of nucleoside transporters, S-(4-nitrobenzyl) -6-thioinosine were exploited. Combination of flow cytometry, confocal microscopy and real-time quantitative polymerase chain reaction methodology were used for the analysis.

Results: Here we utilized flow cytometric assay for analysis of nucleoside transporters activity employing fluorescent nucleoside analogue, uridine-furane. We have determined the long-time kinetics of uridine-furane incorporation and quantified its levels in the parental prostate cancer cell line PC3 and its chemoresistant derivative. Finally, we have shown an association between the activity and mRNA expression of nucleoside transporters and sensitivity to various nucleoside analogues.

Conclusion: Fluorescent techniques can serve as an effective tool for the detection of nucleoside transporter activity which has the potential for application in clinical oncology.Key words: nucleoside transporter proteins - drug resistance - prostatic neoplasm - chemotherapy.

MeSH terms

  • Affinity Labels / pharmacology
  • Antineoplastic Agents / pharmacology
  • Docetaxel / pharmacology
  • Drug Resistance, Neoplasm / genetics*
  • Flow Cytometry / methods*
  • Fluorescent Dyes / pharmacology
  • Gene Expression Regulation, Neoplastic*
  • Humans
  • Male
  • Nucleoside Transport Proteins / genetics*
  • PC-3 Cells
  • Prostatic Neoplasms / drug therapy
  • Prostatic Neoplasms / genetics*
  • Thioinosine / analogs & derivatives
  • Thioinosine / pharmacology

Substances

  • Affinity Labels
  • Antineoplastic Agents
  • Fluorescent Dyes
  • Nucleoside Transport Proteins
  • Docetaxel
  • Thioinosine
  • 4-nitrobenzylthioinosine