Señuelo dirigido a HIF-1 potencializa efectos citotóxicos de dos agentes quimioterapéuticos en MDA-MB-231
HIF decoy improve cytotoxic effects of two chemotherapeutic agents on MDA-MB-231
A decoy dirigida ao HIF-1 potencializa os efeitos citotóxicos de dois agentes quimioterápicos em MDA-MB-231
DOI:
https://doi.org/10.15446/rev.colomb.biote.v22n2.73114Palabras clave:
Cáncer, hipoxia, proliferación celular, apoptosis, sinergia (es)Procesos oncogénicos como proliferación incontrolable, resistencia apoptótica, aumento de mecanismos angiogénicos y evasión inmune son regulados generalmente por factores de transcripción, como HIF-1. Por tanto, se ha señalado a esta molécula como un blanco terapéutico prometedor. Para explorar esta posibilidad, un oligonucleótido tipo “señuelo” dirigido a HIF-1α (ODN) fue diseñado para evaluar su eficiencia en esquema tanto monoterapéutico, como en combinación con dos agentes quimioterapéuticos en un modelo in vitro de cáncer de seno. Después de comprobar, mediante citometría de flujo e inmunofluorescencia, la localización del blanco, el señuelo fue transfectado en la línea celular MDA-MB-231. Se estableció la IC-50 de HIF-1α ODN, Cisplatino y Taxol con el método de Resazurina. Mecanismos de muerte celular fueron evaluados con el método de TUNEL. Por último, se estableció el índice de combinación (IC) de cada uno de los quimio-agentes en combinación con el ODN. Se evidencio que HIF-1α ODN causa un efecto citotóxico en MDA-MB-231 de hasta un 90% hacia las 72h pos-tratamiento. Este efecto no se observa tanto en los controles del ensayo, como en el cultivo primario de células no tumorales (FIBRO), siendo este agente altamente selectivo hacia células tumorales, al activar mecanismos pro-apoptóticos. A su vez, HIF-1α ODN potencializa la actividad tumorogénica de Cisplatino y Taxol en la línea celular tumoral. Por tanto, HIF-1α ODN demostró tener actividad selectiva potencialmente antitumoral, al disminuir la proliferación celular e inducir apoptosis; optimizando de forma sinérgica, la eficacia de fármacos quimioterapéuticos de alto espectro, en tratamientos combinados.
Oncogenic processes like uncontrollable proliferation, resistance to apoptosis, increases in angiogenic mechanisms and immune evasion are regulated by transcription factors such HIF-1. Therefore, this molecule is regarded as a potential therapeutic target. To explore this possibility, a HIF-1α oligonucleotide decoy (ODN) was designed to evaluate its efficiency on both a mono-therapeutic scheme and a mixed treatment with two chemotherapeutic agents within an in vitro model of breast cancer. After confirming the target location with flow cytometry and immunofluorescence assays, that decoy was transfected over the MDA-MB-231 cell line. We established the HIF-1α ODN, Cisplatin and Taxol IC-50 using Resazurin tests. Cell death mechanisms was evaluated with TUNEL. Finally, we obtained the combination index (IC) of each chemical agents with the ODN. This study showed that HIF-1α ODN caused a cytotoxic effect (up to 90%) in MDA-MB-231 during 72 hours post treatment. That effect did not appear in either the assay controls nor in non-tumor cell cultures (FIBRO). This agent is highly selective towards tumor cells, activating pro-apoptotic mechanisms. Additionally, HIF-1α ODN increases the tumorigenic action of Cisplatin and Taxol on the cell line, due to an additive effect. For these reasons, HIF-1α ODN has potential antitumor selective activity, decreasing cell proliferation, inducing apoptosis, and optimizing, in a synergistic manner, the efficacy of wide spectrum chemotherapeutic compounds when it used in a combined treatment.
Processos oncogênicos, como proliferação incontrolável, resistência à apoptose, aumento de mecanismos angiogênicos e evasão imune, são geralmente regulados por fatores de transcrição, como o HIF-1. Portanto, esta molécula tem sido indicada como um alvo terapêutico promissor. Para explorar esta possibilidade, um tipo de oligonucleido "decoy" direccionamento HIF-1α (ODN) foi desenhado para avaliar a sua eficiência como esquema de monoterapêutica ou em combinação com dois agentes quimioterapêuticos, em um modelo in vitro de cancer da mama. Após verificação, por meio de citometria de fluxo e imunofluorescência, a localização do alvo, o chamariz foi transfectado na linhagem celular MDA-MB-231. IC-50 de ODN, Cisplatina e Taxol de HIF-l α foi estabelecido com o método Resazurina. Os mecanismos de morte celular foram avaliados pelo método TUNEL. Finalmente, o índice de combinação (CI) de cada um dos quimio-agentes em combinação com o ODN foi estabelecido. Foi evidenciado que o HIF-1α ODN causa um efeito citotóxico em MDA-MB-231 de até 90% para 72h após o tratamento. Este efeito não é observado tanto nos controles do ensaio, como na cultura primária de células não tumorais (FIBRO), sendo este agente altamente seletivo para células tumorais, ativando mecanismos pró-apoptóticos. Por sua vez, o ODN de HIF-1α potencia a actividade tumorigénica da Cisplatina e do Taxol na linha celular do tumor. Portanto, o HIF-1α ODN demonstrou atividade antitumoral seletiva, diminuindo a proliferação celular e induzindo a apoptose; otimizar sinergicamente, a eficácia de drogas quimioterápicas de alto espectro, em tratamentos combinados.
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