Date Published: March 6, 2019
Publisher: Public Library of Science
Author(s): Caio Raony Farina Silveira, Marcella Cipelli, Carolina Manzine, Silvia Helena Rabelo-Santos, Luiz Carlos Zeferino, Gretel Rodríguez Rodríguez, Josiane Betim de Assis, Suellen Hebster, Isabel Bernadinelli, Fabio Laginha, Enrique Boccardo, Luisa Lina Villa, Lara Termini, Ana Paula Lepique, Joao P.B. Viola.
Cervical cancer, caused by high oncogenic risk Human Papillomavirus (HPV) infection, continues to be a public health problem, mainly in developing countries. Using peptide phage display as a tool to identify potential molecular targets in HPV associated tumors, we identified α-mannosidase, among other enriched sequences. This enzyme is expressed in both tumor and inflammatory compartment of the tumor microenvironment. Several studies in experimental models have shown that its inhibition by swainsonine (SW) led to inhibition of tumor growth and metastasis directly and indirectly, through activation of macrophages and NK cells, promoting anti-tumor activity. Therefore, the aim of this work was to test if swainsonine treatment could modulate anti-tumor immune responses and therefore interfere in HPV associated tumor growth. Validation of our biopanning results showed that cervical tumors, both tumor cells and leukocytes, expressed α-mannosidase. Ex vivo experiments with tumor associated macrophages showed that SW could partially modulate macrophage phenotype, decreasing CCL2 secretion and impairing IL-10 and IL-6 upregulation, which prompted us to proceed to in vivo tests. However, in vivo, SW treatment increased tumor growth. Investigation of the mechanisms leading to this result showed that SW treatment significantly induced the accumulation of myeloid derived suppressor cells in the spleen of tumor bearing mice, which inhibited T cell activation. Our results suggested that SW contributes to cervical cancer progression by favoring proliferation and accumulation of myeloid cells in the spleen, thus exacerbating these tumors systemic effects on the immune system, therefore facilitating tumor growth.
Developing countries continue to be burdened by HPV associated cancer (WHO http://www.who.int/immunization/diseases/hpv/en/). In spite of the available prophylactic vaccines, poor countries still display high rates of cervical cancer and mortality due to this disease. In Brazil, for instance, the National Cancer Agency estimates 16.370 new cervical cancer cases in 2018. Once HPV infection is established, immunization against the virus cannot stop disease progression. Therefore, it is necessary to develop new strategies for clinical intervention, both preventively, in previously untreated patients, as well as patients after treatment with residual disease progression or new tumors. In the present study, we used peptide phage display to search for potential molecular targets in cervical cancer derived cell lines and tumors.
Using the peptide phage display method, we identified heptapeptide sequences with preferential affinity to cervical cancer cell lines and tumors. One of the sequences corresponded to α-mannosidase, which indicated that substrates of this enzyme should be abundant in these tumors. Interestingly, the overlay validation tests showed that the bacteriophage containing the α-mannosidase corresponding peptide displayed stronger binding to cervical tumor tissue than to normal cervical epithelium, again indicating that targets of this enzyme are more abundant in the tumor tissue. α-mannosidase aberrant activity, as well as the activity of other enzymes involved in protein glycosylation, has been associated with cancer progression due to the accumulation of N-glycans on the cell surface [32–34]. In line with this observation, the use of a plant derived α-mannosidase inhibitor, SW, has been shown to inhibit tumor growth and metastasis in experimental models  and induce tumor mass shrinkage in clinical trials .