Title:HIF1A is Overexpressed in Medulloblastoma and its Inhibition Reduces Proliferation and Increases EPAS1 and ATG16L1 Methylation
Volume: 18
Issue: 3
Author(s): Gustavo Alencastro Veiga Cruzeiro, Maristella Bergamo dos Reis, Vanessa Silva Silveira, Regia Caroline Peixoto Lira, Carlos Gilberto Carlotti Jr, Luciano Neder, Ricardo Santos Oliveira, Jose Andres Yunes, Silvia Regina Brandalise, Simone Aguiar, Agda Karina Eterovic, Luiz Gonzaga Tone, Carlos Alberto Scrideli*Elvis Terci Valera
Affiliation:
- Department of Pediatrics, Ribeirao Preto Medicine School, University of Sao Paulo, Avenida dos Bandeirantes 3900, 14049-900, Ribeirao Preto, SP,Brazil
Keywords:
HIF1A, EPAS1, medulloblastoma, epigenetics, ATG16L1, hypoxia.
Abstract: Background: Genetic and epigenetic modifications are closely related to tumor initiation
and progression and can provide guidance for understanding tumor functioning, potentially leading
to the discovery of new therapies. Studies have associated hypoxia-related genes to tumor progression
and chemo/radioresistance in brain tumors. Information on the expression profile of hypoxiarelated
genes in pediatric medulloblastoma, although scarce, may reveal relevant information that
could support treatment decisions.
Objective: Our study focused on evaluation the of CA9, CA12, HIF1A, EPAS1, SCL2A1 and VEGF
genes in 41 pediatric fresh-frozen medulloblastoma sample. Additionally, we analyzed the effect of
hypoxia and normoxia in the pediatric medulloblastoma cell-line UW402. Furthermore, we assessed
the effects of HIF1A knockdown in cell-proliferation and methylation levels of genes related to hypoxia,
apoptosis and autophagy.
Method: qPCR was performed to evaluate mRNA levels, and Western blot to confirm HIF1A silencing
in both patient samples and cell line. Pyrosequencing was performed to asses the methylation
levels after HIF1A knockdown in the UW402 cell line.
Results: A higher HIF1A mRNA level was observed in MB patients when compared to the cerebellum
(non-tumor match). In UW402 MB cell-line, chemically induced hypoxic resulted in an increase
of mRNA levels of HIF1A, VEGF, SCL2A1 and CA9 genes. Additionally, HIF1A knockdown induced
a decrease in the expression of hypoxia related genes and a decrease of 30% in cell proliferation
was also observed. Also, a significant increase in the methylation of ATG16L1 promoter and
decrease in the methylation of EPAS1 promoter were observed after HIF1A knockdown.
Conclusion: HIF1A knockdown in medulloblastoma cells lead to decreased cellular proliferation,
suggesting that HIF1A can be a potential therapeutic target to be explored in the medulloblastoma.
However, the mechanisms behind HIF1A protein stabilization and function are very complex and
more data need to be generated to potentially use HIF1A as a therapeutical target.
