Sialylation transmogrifies human breast and pancreatic cancer cells into 3D multicellular tumor spheroids using cyclic RGD-peptide induced self-assembly
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Roman Akasov1, Sabah Haq2, Fiona Haxho2, Vanessa Samuel2, Sergey V. Burov3, Elena Markvicheva1, Ronald J. Neufeld4, Myron R. Szewczuk2
1Shemyakin-Ovchinnikov Institute of Bioorganic Chemistry, Russian Academy of Sciences, 117997 Moscow, Russia
2Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, Ontario, K7L 3N6 Canada
3Institute of Macromolecular Compounds, Russian Academy of Sciences, Petersburg, 119004 Russia
4Department of Chemical Engineering, Queen’s University, Kingston, Ontario, K7L 3N6 Canada
Elena Markvicheva, email: email@example.com
Ronald J. Neufeld, email: firstname.lastname@example.org
Myron R. Szewczuk, email: email@example.com
Keywords: sialylation, spheroids, oseltamivir phosphate, pancreatic cancer, breast cancer
Received: June 21, 2016 Accepted: August 25, 2016 Published: September 06, 2016
Multicellular tumor spheroids (MTS) have been at the forefront of cancer research, designed to mimic tumor-like developmental patterns in vitro. Tumor growth in vivo is highly influenced by aberrant cell surface-specific sialoglycan structures on glycoproteins. Aberrant sialoglycan patterns that facilitate MTS formation are not well defined. Matrix-free spheroids from breast MCF-7 and pancreatic PANC1 cancer cell lines and their respective tamoxifen (TMX) and gemcitabine (Gem) resistant variants were generated using the RGD platform of cyclic Arg-Gly-Asp-D-Phe-Lys peptide modified with 4-carboxybutyl-triphenylphosphonium bromide (cyclo-RGDfK (TPP)). MCF-7 and MCF-7 TMX cells formed tight spheroids both in the classical agarose-and RGD-based platforms while all PANC1 cells formed loose aggregates. Using lectin histochemistry staining, sialidase assay, neuraminidase (Vibrio cholerae) and oseltamivir phosphate (OP) neuraminidase inhibitor treatments, MCF-7 and PANC1 cells and their drug-resistant variants expressed different sialic acid (SA) content on their cell surfaces. α-2,3- and α-2,6-sialic acid surface residues facilitated spheroid formation under cyclo-RGDfK(TPP)-induced self-assembly. Pretreatment with α-2,3- SA specific Maackia amurensis (MAL-II) lectin, α-2,6-SA specific Sambucus nigra (SNA) lectin, and exogenous α-2,6-SA specific neuraminidase (Vibrio cholerae) dose-dependently reduced spheroid volume. OP enhanced cell aggregation and compaction forming spheroids. PANC1 and MDA-MB231 xenograft tumors from untreated and OP-treated RAGxCγ double mutant mice expressed significantly higher levels of α-2,3- SA over α-2,6-SA. MCF-7 spheroids also expressed a high α-2,3-SA to α-2,6-SA ratio. These results suggest that the relative levels of specific sialoglycan structures on the cell surface correlate with the ability of cancer cells to form avascular multicellular tumor spheroids and in vivo xenograft tumors.
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