Determinação dos efeitos da hipertermia em modelo in vitro de HIPEC sobre as linhagens de células de câncer de ovário: implicações da expressão dos genes das proteínas do choque térmico e resistência à platina
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Data
2021-09-24
Autores
Andrade, Warne Pedro de
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Universidade Estadual Paulista (Unesp)
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INTRODUÇÃO: O câncer epitelial de ovário (EOC) é a neoplasia maligna ginecológica mais letal, com a presença de quimiorresistência contribuindo para o pior prognóstico. Aproximadamente 80 por cento dos casos são diagnosticados no estágio III C e são tratados com cirurgia de citorredução seguida de quimioterapia adjuvante. No entanto, 70 por cento desses pacientes têm recorrências pélvicas e peritoneais. As Proteínas de Choque Térmico são produzidas em resposta ao estresse fisiopatológico e participam de diversos estágios da carcinogênese, atuando principalmente como agentes antiapoptóticos. Elas também estão implicadas na resistência à quimioterapia em vários tipos de tumores. Na tentativa de melhorar os resultados oncológicos, novas abordagens terapêuticas, como quimioterapia intraperitoneal e HIPEC, foram propostas em estudos recentes com ganhos na sobrevida global (SG). No entanto, algumas questões ainda não foram respondidas. MÉTODO: No estudo, culturas de células de câncer de ovário TOV-21G (carcinoma de células claras), SKOV-3 (carcinoma seroso resistente à platina) e OV-90 (seroso de alto grau) foram utilizadas. O ensaio de citotoxicidade celular (MTT) foi realizado. As linhagens de células de câncer de ovário foram tratadas com cisplatina em normotermia (37ºC) e cisplatina em hipertermia (41ºC) e um grupo controle tratado com solução salina PBS de 37 a 41ºC) por 24 horas, seguido por nova suplementação e uma nova incubação de três horas. Um ensaio clonogênico foi realizado. Em seguida, foram submetidos à extração de RNA e transcrição reversa. O qRT-PCR foi realizado para comparar a expressão de TRAP1, HSPB1, HSPD1, HSPA1A e HSPA1L em diferentes tratamentos. RESULTADOS: Não houve diferença significativa quanto à citotoxicidade no tratamento com cisplatina aquecida em relação ao tratamento com normotermia. Não foi possível avaliar a expressão gênica das proteínas de choque térmico na linhagem SKOV-3. Os genes HSPB1, HSPD1 e TRAP1 foram regulados positivamente em OV-90 submetido à hipertermia em relação à normotermia. Não houve alterações significativas na expressão gênica na linhagem TOV-21G. CONCLUSÃO: A linhagem de células serosas de câncer de ovário OV-90, após tratamento com cisplatina aquecida, teve os genes de choque térmico HSPA1A, TRAP1 e HSPB1 regulados positivamente. O gene HSPB1 apresentou a expressão de valor mais significativa. Nosso resultado mostrou que os genes do choque térmico podem estar relacionados à fenotipagem de resistência à quimioterapia encontrada neste tumor. O uso de everolimus tem o potencial de sensibilizar a citotoxicidade em um modelo usando cisplatina aquecida e melhorar resultados. Assim, é necessário avaliar esses genes em um estudo clínico de HIPEC.
INTRODUCTION: Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy, with the presence of chemoresistance contributing to the poor prognosis. Approximately 80% of cases are diagnosed in stage III C and are treated with cytoreduction surgery followed by adjuvant chemotherapy. However, 70 percent of these patients have pelvic and peritoneal recurrences. Heat Shock Proteins are produced in response to pathophysiological stress and take part in several stages of carcinogenesis, acting primarily as anti-apoptotic agents. They are also implicated in resistance to chemotherapy in several types of tumors. In an attempt to improve oncological results, new therapeutic approaches such as intraperitoneal chemotherapy and HIPEC have been proposed in recent studies with gains in overall survival (OS). However, some questions have not yet been answered. METHODS: In the study, cultures of ovarian cancer cells TOV-21G (clear cell carcinoma), SK-OV3 (platinumresistant serous carcinoma), and OV-90 (high-grade serous). Cell cytotoxicity (MTT) assay was performed. The ovarian cancer cells lines were treated with cisplatin in normothermia (37 degrees Celsius) and cisplatin in hyperthermia (41 degrees Celsius) and a control group treated with PBS saline solution at (37 degrees Celsius and 41 degrees Celsius) for 24 hours, followed by new supplementation and a new three hours incubation. A clonogenic assay was performed. Then they were submitted to RNA extraction and reverse transcription. qRT-PCR was performed to compare the expression of TRAP1, HSPB1, HSPD1, HSPA1A and HSPA1L in different treatments. RESULTS: There was no statistical difference concerning cytotoxicity between treatment with heated cisplatin and treatment with normothermia. It was not possible to evaluate the expression of the heat shock genes in the SKOV-3 lineage. The HSPB1, HSPD1 and TRAP1 genes were positively regulated in OV-90 submitted to hyperthermia in relation to normothermia, and there were no significant changes in expression in the TOV-21G. CONCLUSION: In conclusion, we observed that OV-90 serous ovarian cancer cell line, after heated cisplatin treated had the HSPA1A, TRAP1, and HSPB1 heat shock genes upregulated. The HSPB1 genes had the most significant value expression. Our result showed that the heat shock genes can be related at chemotherapy resistance phenotyping founded this tumor-like. The use of everolumus has the potential to sensitize cytotoxicity in a model using heated cisplatin and improved results Thus, it is necessary to evaluate these genes in a clinical study of HIPEC.
INTRODUCTION: Epithelial ovarian cancer (EOC) is the most lethal gynecological malignancy, with the presence of chemoresistance contributing to the poor prognosis. Approximately 80% of cases are diagnosed in stage III C and are treated with cytoreduction surgery followed by adjuvant chemotherapy. However, 70 percent of these patients have pelvic and peritoneal recurrences. Heat Shock Proteins are produced in response to pathophysiological stress and take part in several stages of carcinogenesis, acting primarily as anti-apoptotic agents. They are also implicated in resistance to chemotherapy in several types of tumors. In an attempt to improve oncological results, new therapeutic approaches such as intraperitoneal chemotherapy and HIPEC have been proposed in recent studies with gains in overall survival (OS). However, some questions have not yet been answered. METHODS: In the study, cultures of ovarian cancer cells TOV-21G (clear cell carcinoma), SK-OV3 (platinumresistant serous carcinoma), and OV-90 (high-grade serous). Cell cytotoxicity (MTT) assay was performed. The ovarian cancer cells lines were treated with cisplatin in normothermia (37 degrees Celsius) and cisplatin in hyperthermia (41 degrees Celsius) and a control group treated with PBS saline solution at (37 degrees Celsius and 41 degrees Celsius) for 24 hours, followed by new supplementation and a new three hours incubation. A clonogenic assay was performed. Then they were submitted to RNA extraction and reverse transcription. qRT-PCR was performed to compare the expression of TRAP1, HSPB1, HSPD1, HSPA1A and HSPA1L in different treatments. RESULTS: There was no statistical difference concerning cytotoxicity between treatment with heated cisplatin and treatment with normothermia. It was not possible to evaluate the expression of the heat shock genes in the SKOV-3 lineage. The HSPB1, HSPD1 and TRAP1 genes were positively regulated in OV-90 submitted to hyperthermia in relation to normothermia, and there were no significant changes in expression in the TOV-21G. CONCLUSION: In conclusion, we observed that OV-90 serous ovarian cancer cell line, after heated cisplatin treated had the HSPA1A, TRAP1, and HSPB1 heat shock genes upregulated. The HSPB1 genes had the most significant value expression. Our result showed that the heat shock genes can be related at chemotherapy resistance phenotyping founded this tumor-like. The use of everolumus has the potential to sensitize cytotoxicity in a model using heated cisplatin and improved results Thus, it is necessary to evaluate these genes in a clinical study of HIPEC.
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Palavras-chave
Câncer de ovário, Resistência à platina, HIPEC, Proteínas de choque térmico, Ovarian cancer, , Resistance to chemotherapy, Heat shock proteins