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J Cancer Metastasis Treat 2020;6:5 I http://dx.doi.org/10.20517/2394-4722.2020.13 Page 27 of 38

and poor prognosis. The development of cancer is often associated with dysregulation of epigenetic
mechanisms, including DNA methylation. The aim of our study was to evaluate MGMT (O6-methylguanine
DNA methyltransferase), BRCA1 (breast cancer 1), and MEG3 (maternally expressed 3) methylation in
TNBC.

Experimental procedure: In this study, 44 TNBC patients were included. The methylation status of the
MGMT, BRCA1, and MEG3 promoter regions were analyzed by methylation-specific PCR.

Results: MGMT, BRCA1, and MEG3 promoter methylation was found in 70.4%, 61.3%, and 61.3% of TNBC
patients, respectively. Moreover, we showed that the frequency of MGMT and BRCA1 methylation is higher
in older patients compared to younger patients (P-value for MGMT is P = 0.0194 and for BRCA1 is P =
0.0188). Additionally, in one of TNBC patient with glandular and squamous histopathological components,
it was shown that the promoter status of all analyzed genes changed from methylated to unmethylated after
chemotherapy of this patient.

Conclusion: The high frequency of MGMT, BRCA1, and MEG3 methylation indicates that epigenetic
changes are important mechanisms in breast cancer. Moreover, our results indicate that MGMT and BRCA1
methylation may have greater impact in the development of breast cancer in older patients compared to
younger patients.

38. Sphingosine kinase 2 in oral squamous cell carcinoma

Lais Brigliadori Fugio, Andréia Machado Leopoldino

Department of Clinical Analysis, Toxicology and Food Sciences, School of Pharmaceutical Sciences of Riberião
Preto, University of São Paulo, Ribeirão Preto, SP 13010, Brazil.

Background and aim: Sphingosine kinase 2 (SK2) is one of the enzymes responsible for producing
[1]
sphingosine-1-phosphate (S1P) . Recently, SK2 has been associated with protective autophagy and
[2,3]
survival, and regulation of p21 in breast and colon cancer cells . However, the role of SK2 in oral
squamous cell carcinoma (OSCC) is still unclear. Thus, our study aimed to investigate the involvement of
SK2 in autophagy and proliferation in OSCC cells.

Experimental procedure: HN13 and HN12 (OSCC) cell lines were transduced with short hairpin RNA
interference against SK2 and a lentiviral vector containing cDNA for SK2, respectively. Cell cycle
analyses were performed by propidium iodide staining and flow cytometry. Western blotting and
immunofluorescence assays were adopted to analyze protein levels and cellular distribution.

Results: HN13 cells with SK2 knockdown showed a decrease of pAkt, c-MYC, and LC3 levels (an
autophagy marker) while p21 was increased. SK2 knockdown in HN13 cells caused cell arrest in S phase
with reduction of the cells in G2/M. SK2 overexpression in HN12 cells led to an increase of pAkt, c-Myc,
and LC3 levels.

Conclusion: Our work is the first to demonstrate the role of SK2 in proliferation and autophagy in OSCC
cells. Other studies are in progress to understand the molecular mechanism underlying the role of SK2 and
its potential as a target.

Financial support: FAPESP (grant: 2016/19103-2; scholarship: 2018/14225-8, CAPES, CNPq- Brazil).