Tumor Biology

, Volume 36, Issue 10, pp 8107–8120 | Cite as

Cellular prion protein contributes to LS 174T colon cancer cell carcinogenesis by increasing invasiveness and resistance against doxorubicin-induced apoptosis

  • Cornelius Kwang-Lee Chieng
  • Yee-How SayEmail author
Research Article


As the cellular prion protein (PrPC) has been implicated in carcinogenesis, we aimed to investigate the effects of cancer cell-specific PrPC overexpression from the invasion, metastasis, and apoptosis aspects, by performing cell motility assays, cell proliferation assays under anchorage-dependent and anchorage-independent conditions, and apoptosis evasion when subjected to multiple anti-cancer drugs. Overexpression of PrPC in LS 174T was achieved by stable transfection. PrPC overexpression was shown to increase cell proliferation in anchorage-dependent and anchorage-independent manners, as shown by more viable cells in 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, more colonies formed in soft agar assay and increased resistance to anoikis in poly-2-hydroxyethyl methacrylate-coated surface. PrPC overexpression also increased cell motility and invasiveness of LS 174T. Cell adhesion to extracellular matrix using collagen- and fibronectin-coated surfaces revealed increased cell attachment in LS 174T cells overexpressing PrPC. Analysis of apoptotic and necrotic cells by propidium iodide/annexin V-fluorescein isothiocyanate microscopy and 7-amino-actinomycin D/annexin V-phycoerythrin flow cytometry revealed that PrPC overexpression attenuated doxorubicin-induced apoptosis. Human apoptosis antibody array with 35 apoptosis-related proteins revealed that three inhibitor of apoptosis proteins (IAPs)—survivin, X-linked inhibitor of apoptosis protein (XIAP), and cellular inhibitor of apoptosis protein-1 (cIAP-1)—were upregulated in LS 174T cells overexpressing PrPC in doxorubicin-induced apoptosis. In conclusion, the overexpression of PrPC could enhance the invasiveness and survival of LS 174T colorectal cancer cells, indicating that PrPC plays a role in colorectal cancer biology.


Cellular prion protein Colon cancer Invasion Metastasis Apoptosis 



This work was supported by grants from the UTAR Research Fund—IPSR/RMC/UTARRF/C210/S1 and IPSR/RMC/UTARRF/2012-C2/G05.

Conflicts of interest


Supplementary material

13277_2015_3530_Fig7_ESM.gif (45 kb)
Supplementary Fig. 1

PrPC overexpression had no effect on anchorage-dependent and anchorage-independent growth of HEK 293 cells. A. Stable expression of PrPC was determined by Western blot analysis using anti- PrPC, revealing the three glycosylation isoforms of PrPC namely, unglycosylated PrP (27 kDa), monoglycosylated PrP (30 kDa), and diglycosylated PrP (35 kDa); B. MTT cell viability assay for HEK 293 cells. Cell viability was calculated with the formula: (A570 of day n / A570 of day 0) × 100 %. Data of percentage of cells are expressed as mean ± SEM (error bars) obtained from three independent experiments. Mean values were compared using one-way ANOVA followed by LSD’s post hoc test for comparison of the means. * p < 0.05 as compared with HEK 293 and HEK 293–3.1. (GIF 44 kb)

13277_2015_3530_MOESM1_ESM.tif (7.9 mb)
High resolution image (TIFF 8078 kb)
13277_2015_3530_Fig8_ESM.gif (380 kb)
Supplementary Fig. 2

PrPC had no effect on the migration and invasion of HEK 293 cells. A. Scratch wound assay for HEK 293 cells. Wound closure for HEK 293 cells was up to 72 h. Images were acquired using NIS-Elements BR 3.0 software under Nikon Eclipse TS100 inverted microscope at 40× magnification; B. Remaining open wound area count for HEK 293 cells. Image analysis was performed using TScratch software version 1.0; HEK 293 cells were incubated with various concentrations of rat tail collagen I (C) for 15 min and fibronectin (D) for 30 min. Adherent cells were stained, lysed and quantified at OD570. Qualitative variations in adhesion were measured as changes in OD values; E. Cell scattering effect for HEK 293 cells was observed from 0 – 48 h. Cell morphology changed from compact to semiscattered and eventually to apoptotic cells. Inset images are enlargements of approximately 10× from original images. Images were acquired using NIS-Elements BR 3.0 software under Nikon Eclipse TS100 inverted microscope at 200× magnification. Data were expressed as mean ± SEM (error bars) obtained from at least two independent experiments. Mean values were compared using one-way ANOVA followed by LSD’s post hoc test for comparison of the means. * p < 0.05 as compared with HEK 293 and HEK 293–3.1. (GIF 380 kb)

13277_2015_3530_MOESM2_ESM.tif (20.8 mb)
High resolution image (TIFF 21341 kb)
13277_2015_3530_Fig9_ESM.gif (45 kb)
Supplementary Fig. 3

PrPC overexpression had no effect on protection against cell death of HEK 293 cells upon anti-cancer drugs treatment. Cells were exposed to DOX (A), etoposide (B) at concentrations of 2, 4, 8 and 16 μM, or vincristine sulphate - VIN (C) at concentrations of 0.025, 0.05, and 0.1 μM for 48 h before MTT assay was performed and absorbance was read at 550 nm; Data were expressed as mean ± SEM (error bars) obtained from two independent experiments. Mean values were compared using one-way ANOVA followed by LSD’s post hoc test for comparison of the means. * p < 0.05 as compared with HEK 293 and HEK 293–3.1. (GIF 45 kb)

13277_2015_3530_MOESM3_ESM.tif (12.1 mb)
High resolution image (TIFF 12341 kb)


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Copyright information

© International Society of Oncology and BioMarkers (ISOBM) 2015

Authors and Affiliations

  1. 1.Department of Biomedical Science, Faculty of ScienceUniversiti Tunku Abdul Rahman (UTAR) Perak CampusKamparMalaysia

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