Human Papillomavirus 16 E6 increases the radiosensitivity of p53-mutated cervical cancer cells, associated with up-regulation of aurora A

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Abstract

Purpose: To examine the effect of the human papillomavirus (HPV) type 16-E6 (HPV `early' gene) oncoprotein on in vitro radiosensitivity of HPV-negative/p53 mutant C33a cervical cancer cells. Methods and materials: The human cervical cancer cell line C33a was stably transfected with either the HPV16 E6 cDNA cloned into the vector pcDNA3.0 (C33aE6) or the empty-vector control (C33aV). Radiosensitivity, DNA damage, and cell cycle measurements were made using standard clonogenic assays, immunofluorescent assessment of nuclear histone H2AX phosphorylated on serine-139 (g-H2AX) foci, and flow cytometry. Western immunoblotting and fluorescence confocal microscopy were used to analyse the changes in cellular proteins. Real-time polymerase chain reaction (PCR) was used to compare levels of aurora A mRNA. Results: Compared to C33aV cells, C33aE6 cells showed enhanced radiation cell killing. This was associated with a large amount of polyploidy which was followed by late cell death in C33aE6 cells. Aurora A was highly expressed in C33aE6 cells at pre- and post-irradiation times compared to C33aV cells. Silencing aurora A resulted in a reduced amount of residual g-H2AX foci in C33aE6 cells, and diminished the difference in radiosensitivity between the C33aE6 and C33aV cells. Conclusion: Our in vitro results indicate that genetic instability could be augmented in the HPV-infected cancer cells by up-regulation of aurora A, especially against a background of dysfunctional p53. Further studies are needed to examine whether aurora A could be a viable therapeutic target in HPV-related tumours.

Keyword(s)

ASSAY; CANCER; CANCER CELLS; CELL-CYCLE; CELLS; Cell Death; Cell Line; DAMAGE; DEATH; DNA damage; DNA-DAMAGE; Dna; Flow Cytometry; GENE; Human; IN-VITRO; IONIZING-RADIATION; P53; PROTEIN; RADIATION; SENSITIVITY; cell cycle; cervical cancer; ionizing radiation; methods; proteins; radiosensitivity

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