Sialylation facilitates self-assembly of 3D multicellular prostaspheres by using cyclo-RGDfK(TPP) peptide
Received 31 January 2017
Accepted for publication 31 March 2017
Published 4 May 2017 Volume 2017:10 Pages 2427—2447
Checked for plagiarism Yes
Review by Single-blind
Peer reviewers approved by Dr Lucy Goodman
Peer reviewer comments 2
Editor who approved publication: Dr Jianmin Xu
Sabah Haq,1 Vanessa Samuel,1 Fiona Haxho,1 Roman Akasov,2,3 Maria Leko,4 Sergey V Burov,4 Elena Markvicheva,2 Myron R Szewczuk1
1Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON, Canada; 2Polymers for Biology Laboratory, Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 3Sechenov First Moscow State Medical University, Institute for Regenerative Medicine, Moscow, 4Synthesis of Peptides and Polymer Microspheres Laboratory, Institute of Macromolecular Compounds, Russian Academy of Sciences, St Petersburg, Russia
Background: Prostaspheres-based three dimensional (3D) culture models have provided insight into prostate cancer (PCa) biology, highlighting the importance of cell–cell interactions and the extracellular matrix (EMC) in the tumor microenvironment. Although these 3D classical spheroid platforms provide a significant advance over 2D models mimicking in vivo tumors, the limitations involve no control of assembly and structure with only limited spatial or glandular organization. Here, matrix-free prostaspheres from human metastatic prostate carcinoma PC3 and DU145 cell lines and their respective gemcitabine resistant (GemR) variants were generated by using cyclic Arg-Gly-Asp-D-Phe-Lys peptide modified with 4-carboxybutyl-triphenylphosphonium bromide (cyclo-RGDfK(TPP)).
Materials and methods: Microscopic imaging, immunocytochemistry (ICC), flow cytometry, sialidase, and WST-1 cell viability assays were used to evaluate the formation of multicellular tumor spheroid (MCTS), cell survival, morphologic changes, and expression levels of α2,6 and α2,3 sialic acid (SA) and E- and N-cadherin in DU145, PC3, and their GemR variants.
Results: By using the cyclo-RGDfK(TPP) peptide platform in a dose- and time-dependent manner, both DU145 and DU145GemR cells formed small MCTS. In contrast, PC3 and PC3GemR cells formed irregular multicellular aggregates at all concentrations of cyclo-RGDfK(TPP) peptide, even after 6 days of incubation. ICC and flow cytometry results revealed that DU145 cells expressed higher amounts of E-cadherin but lower N-cadherin compared with PC3 cells. By using Maackia amurensis (α2,3-SA-specific MAL-II) and Sambucus nigra (α2,6-SA specific SNA) lectin-based cytochemistry staining and flow cytometry, it was found that DU145 and DU145GemR cells expressed 5 times more α2,6-SA than α2,3-SA on the cell surface. PC3 cells expressed 4 times more α2,3-SA than α2,6-SA, and the PC3GemR cells showed 1.4 times higher α2,6-SA than α2,3-SA. MCTS volume was dose-dependently reduced following pretreatment with α2,6-SA-specific neuraminidase (Vibrio cholerae). Oseltamivir phosphate enhanced cell aggregation and compaction of 3D MCTS formed with PC3 cells.
Conclusion: The relative levels of specific sialoglycan structures on the cell surface correlate with the ability of PCa cells to form avascular multicellular prostaspheres.
Keywords: PC3 and DU145 cell lines, cadherin, oseltamivir phosphate, gemcitabine, chemoresistance, neuraminidase, EMT, spheroid
This work is published and licensed by Dove Medical Press Limited. The full terms of this license are available at https://www.dovepress.com/terms.php and incorporate the Creative Commons Attribution - Non Commercial (unported, v3.0) License. By accessing the work you hereby accept the Terms. Non-commercial uses of the work are permitted without any further permission from Dove Medical Press Limited, provided the work is properly attributed. For permission for commercial use of this work, please see paragraphs 4.2 and 5 of our Terms.Download Article [PDF] View Full Text [HTML][Machine readable]