Estudi dels potencials mecanismes involucrats en la carcinogènesi de l'arsènic
- Autores: Jana Peremartí Brosel
- Directores de la Tesis: Ricardo Marcos Dauder (dir. tes.), Alba Hernández Bonilla (dir. tes.)
- Lectura: En la Universitat Autònoma de Barcelona ( España ) en 2017
- Idioma: catalán
- Tribunal Calificador de la Tesis: Amadeu Creus Capdevila (presid.), Elisabet Teixidó Condomines (secret.), Joan Maria Llobet Mallafré (voc.)
- Programa de doctorado: Programa Oficial de Doctorado en Genética
- Materias:
- Enlaces
- Resumen
Inorganic arsenic is a widely spread environmental contaminant known to be cytotoxic, genotoxic and carcinogenic to humans. Epidemiological data clearly show an increase in the incidence of cancer in human populations living in arsenic-rich areas, mainly on skin, lung, bladder, liver, prostate and kidney cancers. However, the associated carcinogenic mechanisms remain incompletely characterized. Hence, the aim of this Thesis is to deeper in the mechanisms of arsenic-related effects, trying to disclose new insights.
Several studies suggest that reactive oxygen species (ROS) generated during arsenic biotransformation play an important role in arsenic-related carcinogenesis, but the lack of studies with environmentally relevant type of exposures makes this point uncertain. One of the goals of this Thesis is to assess the behavior of oxidative and genotoxic DNA damage along the acquisition of the in vitro cancer phenotype, and to determine the involvement of AS3MT and MTH1 enzymes in the process. Thus, mouse embryonic fibroblast (MEF) cells previously transformed by 30 weeks of chronic arsenite exposure were monitored for oxidative DNA damage (ODD) and chromosome damage, by using the comet and the micronucleus assay, at different time-of-exposure intervals for the complete duration of the exposure. Expression changes of As3mt and Mth1 genes, both related to oxidative damage, were also evaluated by real time RT-PCR, as well as, the epigenetic consequences. Results indicate that the oxidative and genotoxic damage of chronically exposed MEF cells increases up to the point of transformation, but drop drastically afterwards. Interestingly, As3mt expression follows the same pattern than DNA damage, while Mth1 increases its expression after the point of transformation, rather being the responsible of the observed DNA damage decrease. In addition, a global DNA hymomethylation was observed during the complete duration of the exposure.
On the other hand, chronic arsenic exposure interferences with NER and BER pathways are well documented, presumably due to the arsenic ability of interacting and inhibiting proteins, but interactions with other DNA repair pathways are poorly explored. So, another point of interest of this Thesis is to scope whether arsenic disrupts the normal function of FA/BRCA pathway. For this, cells were exposed for two weeks to subtoxic concentrations of two trivalent arsenic compounds, ATO and MMAIII and, after that, the cellular response to different inducers of the studied pathway was evaluated and compared to one FA/BRCA-deficient cell line. Our results show that preexposure to the trivalent arsenic compounds sensitizes cells in front of crosslinking agents, increases accumulation of cells in G2/M phase, reduces the amount of activated FANCD2 protein and increases the micronuclei frequency. Altogether, these findings indicate that environmental relevant arsenic exposures disrupt the FA/BRCA function, displaying a novel mechanism of arsenic co- and carcinogenesis.
Our data are certainly important for the potential use of arsenicals in cancer therapy. As they sensitize cells to crosslinking agents, we suggest that combination chemotherapy of ATO and a crosslinking agent would increase the response and improve outcomes by lowering the associated toxicity side-effects. To prove this hypothesis, three different bladder cancer cell lines were treated with a combination of low concentrations of ATO and cisplatin, a well-known chemotherapeutic drug. After proving the altered FA/BRCA repair capacity of the bladder cancer cells treated with ATO and cisplatin, several endpoints like toxicity and extrinsic hallmarks of cancer phenotype were assessed. Our results show that combined treatments significantly decrease the viability of exposed cells in all cases. Also, we find that the ATO/cisplatin cotreatment decreases the number of colonies able to grow in soft-agar, as well as cell migration rate, MMP secretion, and cell invasion capacity.
Overall, our results demonstrate the implication of Mth1 and As3mt in the accumulation and subsequent drop of the oxidative DNA damage during arsenic-induced transformation process, and set Mth1 as a useful biomarker for arsenic carcinogenesis. On the other hand, our data demonstrate for the first time that environmentally relevant arsenic exposures disrupt the normal function of the FA/BRCA pathway, supporting a novel source of arsenic-related carcinogenic and co-carcinogenic effects that in turn can be used as a novel cancer therapy strategy, where a combination of ATO and cisplatin would synergistically improve the anticancer outcomes requiring lower cisplatin doses, eventually preventing from its toxic side effects and common cancer recurrence.
