Abstract
Host defense caerin 1.1 and 1.9 peptides, isolated from the glandular secretion of Australian tree frogs, the genus Litoria, have been previously shown to have multiple biological activities, including the inhibition of human papillomavirus (HPV) 16 early protein E7 transformed murine as well as human cancerous cell proliferation both in vitro and in vivo. However, the mechanism underlying their anti-proliferative activities against HPV18+ cervical cancer HeLa cells remains unknown. This study comparatively investigated the anti-proliferation on HeLa cells by caerin 1.1, 1.9, and their mixture, followed by confocal microscopy examination to assess the cellular intake of the peptides. Tandem mass tag labeling proteomics was employed to reveal the proteins that were significantly regulated by the peptide treatment in cells and cell growth environment, to elucidate the signaling pathways that were modulated. Western blot was performed to confirm the modulation of the pathways. Both caerin 1.1 and 1.9 highly inhibited HeLa cell proliferation with a significant additive effect compared to untreated and control peptide. They entered the cells with different magnitudes. Intensive protein-protein interaction was detected among significantly upregulated proteins. Translation, folding and localization of proteins and RNA processing, apoptosis process was significantly enriched post the treatments. The apoptotic signaling was suggested as a result of tumor necrosis factor-α (TNF-α) pathway activation, indicated by the dose-dependent elevated levels of caspase 3 and caspase 9. The epidermal growth factor receptor and androgen receptor pathways appeared inhibited by the peptides. Moreover, the activation of T-cell receptor derived from the quantitation results further implies the likelihood of recruiting more T cells to the cell growth environment post the treatment and more sensitive to T cell mediated killing of HeLa cells. Our results indicate that caerin 1.1 and 1.9 mediate apoptotic signals of HeLa cells and may subsequently enhances adaptive T cell immune responses.
Highlights
Cervical cancer is the second most common cancer in woman worldwide, results from persistent infection of Human papillomavirus (HPV) infection, especially HPV subtype 16, 18 infection, which accounts for more than 70% of cervical cancer (Leo et al, 2017; McGregor et al, 2018)
We explored the presence of unconjugated caerin 1.1 and 1.9 in cytoplasm of HeLa cells post the treatment, to further confirm the entry of caerin peptides into the cells and exclude the effect of FITC labeling toward the cellular uptake
A previous study showed that caerin peptides were able to access HIV captured by dendritic cells either on the cell surface or intracellularly, effectively in inhibiting HIV infection by disrupting the viral membrane and preventing the entry the transfer of HIV to T cells (VanCompernolle et al, 2005)
Summary
Cervical cancer is the second most common cancer in woman worldwide, results from persistent infection of Human papillomavirus (HPV) infection, especially HPV subtype 16, 18 infection, which accounts for more than 70% of cervical cancer (Leo et al, 2017; McGregor et al, 2018). HPV prophylactic vaccine introduced a decade ago effectively prevents HPV infection and may eradicate cervical cancer in countries that the vaccines are well distributed (McGregor et al, 2018). The vaccine is ineffective for those already infected with HPV and developing countries that have most cervical cancer patients have limited resources to vaccinate those at risk (Ni et al, 2015). Development of effective cervical cancer therapies remains as an urgent task. An ideal therapeutic vaccine should be able to elicit high quality and enough numbers of effective T cells, which can migrate to the tumor site, and overcome the tumor immunosuppressive microenvironment and kill the tumor cells via the process such as apoptosis (Ni et al, 2015)
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