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Publication
Thioredoxin reductase inhibitors in anticancer therapy : thimerosal efficacy in Glioblastoma
Authors
Bramatti, Isabella
Abstract(s)
Glioblastoma multiforme (GBM) is the most common and aggressive primary malignant brain tumor with a low life expectancy. The standard treatment includes resection surgery, radiotherapy and chemotherapy with temozolomide (TMZ). TMZ is considered the first choice and standard treatment, but therapeutic success is limited due to neoangiogenesis, intratumor heterogeneity and therapy resistance, with overall patient survival increasing by just 14 months. This poor outcome of current therapies increases the demand for new, more effective therapeutic options, including drug repurposing.
Tumor hypoxia involves the development of non-functional blood vessels within neoplastic tissue, leading to tumor survival, invasion and metastasis. Cancer growth and resistance to therapy are frequently related to overexpression of antioxidant systems such as the thioredoxin system. This system includes the disulfide reductase, thioredoxin (Trx), the selenoenzyme thioredoxin reductase (TrxR) and NADPH. This system is fundamental for cell survival and proliferation with a key role in maintaining redox homeostasis, being able to provide electrons to a large range of enzymes critical for DNA synthesis and defense against oxidative stress, regulating hypoxia inducible factor-1alpha (HIF-1α) activity, which in turn controls vascular endothelial growth factor (VEGF), an indispensable factor for tumor invasiveness, and microenvironment tumor maintenance. HIF-1α can also be regulated by the signal transducer and activator of transcription 3 (STAT3), an oncogene stimulated by pro-inflammatory cytokines and growth factors.
Indeed, targeting of antioxidant systems arises as a potential strategy to tackle tumors since they are more sensitive to loss of redox homeostasis than normal cells and those systems are involved in more than one of the considered hallmarks of cancer. The thioredoxin system has several known inhibitors including mercury compounds such as thimerosal (TmHg), an ethylmercury-containing compound, that readily crosses the blood-brain barrier (BBB) and accumulates in the brain. Despite having a history of human use, epidemiological evidence of TmHg neurotoxicity was never obtained.
Therefore, since elevation of oxidative stress triggers different pathways that will lead to apoptosis of glioma cells and the overall survival for this cancer is still low even with emerging therapies, the goal of this thesis is to study the use of thioredoxin’ inhibitors, in this case thimerosal, as a possible candidate for hard repurposing towards GBM as well as to overcome therapy resistance to TMZ.
Results showed that TmHg significantly decreased the cell viability (> 50%) of two different glioblastoma cell lines (GL261 and U87 cell line) at much lower concentrations than TMZ, and strongly inhibited the Trx system. In fact, in GL261 cells besides an inhibition of cytosolic TrxR1/Trx1 there was a strong inhibition of mitochondrial TrxR2/Trx2 expression, predominantly with TmHg. In U87 cells, exposure to TmHg decreased cell migration, HIF-1α expression (35%) and in co-exposure with TMZ reduced HIF-1α, VEGF, and phosphorylation of STAT3, showing that tumour invasiveness related factors are significantly hampered. Furthermore, TmHg elicit N9 microglial inflammation markers - increased levels of IL1β, iNOS and TNFα and the autophagic process. Moreover, in co-cultures of microglia and GBM cells, the cytotoxicity of TmHg towards GL261 was higher.
The effectiveness of EtHg and TmHg in reducing GBM viability and affecting the Trx system could also be related with its interaction with TrxR. We demonstrated that EtHg, unlike other Hg compounds such as methylmercury, is a partial inhibitor of this selenoenzyme, forming a selenium-compromised thioredoxin reductase-derived apoptotic proteins (SecTRAPs), which enhances ROS-production and cytotoxicity.
Also, a mouse (C57BL/6) model was used in a distribution study following TmHg administration (i.v.; 20, 40 and 100 µM) for 14 days. We observed a concentration-dependent Hg accumulation in the brain, kidney and liver. Nevertheless, no neurobehavioral alterations were noticed as well as no histopathological damage in the organs, which led to the optimization and testing in a GBM model encompassing stereotactic injection of GBM cells.
Overall, thimerosal arose as a good candidate for hard repurposing towards GBM and the results obtained indicate its potential to be used alone as well as to overcome therapy resistance to TMZ.
Description
Keywords
glioblastoma stress oxidativo sistema da tiorredoxina hipóxia timerosal oxidative stress thioredoxin system hypoxia thimerosa