RESUMEN
Molecules involved in DNA damage response (DDR) are often overexpressed in cancer cells, resulting in poor responses to chemotherapy and radiotherapy. Although treatment efficacy can be improved with the concomitant use of DNA repair inhibitors, the accompanying side effects can compromise the quality of life of patients. Therefore, in this study, we identified a natural compound that could inhibit DDR, using the single-strand annealing yeast-cell analysis system, and explored its mechanisms of action and potential as a chemotherapy adjuvant in hepatocellular carcinoma (HCC) cell lines using comet assay, flow cytometry, Western blotting, immunofluorescence staining, and functional analyses. We developed a mouse model to verify the in vitro findings. We found that hydroxygenkwanin (HGK) inhibited the expression of RAD51 and progression of homologous recombination, thereby suppressing the ability of the HCC cell lines to repair DNA damage and enhancing their sensitivity to doxorubicin. HGK inhibited the phosphorylation of DNA damage checkpoint proteins, leading to apoptosis in the HCC cell lines. In the mouse xenograft model, HGK enhanced the sensitivity of liver cancer cells to doxorubicin without any physiological toxicity. Thus, HGK can inhibit DDR in liver cancer cells and mouse models, making it suitable for use as a chemotherapy adjuvant.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Daño del ADN/efectos de los fármacos , Flavonoides/farmacología , Animales , Apoptosis/efectos de los fármacos , Apoptosis/genética , Línea Celular Tumoral , Reparación del ADN/efectos de los fármacos , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Medicamentos Herbarios Chinos , Regulación de la Expresión Génica , Recombinación Homóloga/efectos de los fármacos , Humanos , Ratones , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Ensayos Antitumor por Modelo de Xenoinjerto , Levaduras/efectos de los fármacos , Levaduras/genética , Levaduras/metabolismoRESUMEN
Pazopanib-a multitargeted tyrosine kinase inhibitor with prominent antiangiogenic effects-has shown promise in the treatment of soft-tissue sarcomas. Hyperthermia has been also applied as an adjunctive treatment to chemotherapy for these malignancies. Here, we show that pazopanib and hyperthermia act synergistically in inhibiting uterine leiomyosarcoma (LMS) cell growth. Compared with either treatment alone, the combination of pazopanib and hyperthermia exerted the highest antitumor activity in a xenograft model. Mechanistically, we found that combined treatment with pazopanib and hyperthermia inhibited histone acetyltransferase 1 (HAT1) expression in LMS cells. The Clock element on the HAT1 promoter was critical for pazopanib- and hyperthermia-induced HAT1 downregulation. Inhibition of HAT1-either by pazopanib and hyperthermia or through HAT1 silencing-was mediated by suppression of Clock. Accordingly, Clock protein reconstitution rescued both HAT1 levels and HAT1-mediated histone acetylation. Immunohistochemistry revealed a higher expression of HAT1 in uterine LMS than in leiomyomas (p = 0.007), with high HAT1 expression levels being associated with poor clinical outcomes (p = 0.007). We conclude that pazopanib and hyperthermia exert synergistic effects against LMS growth by inhibiting HAT1. Further preclinical studies on HAT1 as a potential drug target in uterine LMS are warranted, especially in combination with hyperthermia. KEY MESSAGES: Pazopanib and hyperthermia inhibit the growth of leiomyosarcoma. Their combined use inhibits HAT1 expression in leiomyosarcoma cells. The promoter Clock element is required for HAT1 downregulation. HAT1 expression is higher in leiomyosarcoma than in leiomyomas. An increased HAT1 expression is associated with poor clinical outcomes.
Asunto(s)
Inhibidores de la Angiogénesis/farmacología , Antineoplásicos/farmacología , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Histona Acetiltransferasas/genética , Hipertermia Inducida , Indazoles/farmacología , Pirimidinas/farmacología , Sulfonamidas/farmacología , Biomarcadores , Proteínas CLOCK/genética , Proteínas CLOCK/metabolismo , Femenino , Histona Acetiltransferasas/metabolismo , Humanos , Hipertermia Inducida/métodos , Leiomiosarcoma/genética , Leiomiosarcoma/metabolismo , Leiomiosarcoma/patología , Neoplasias Uterinas/genética , Neoplasias Uterinas/metabolismo , Neoplasias Uterinas/patologíaRESUMEN
DNA repair systems are abnormally active in most hepatocellular carcinoma (HCC) cells due to accumulated mutations, resulting in elevated DNA repair capacity and resistance to chemotherapy and radiotherapy. Thus, targeting DNA repair mechanisms is a common treatment approach in HCC to sensitize cancer cells to DNA damage. In this study, we examined the anti-HCC effects of melatonin and elucidated the regulatory mechanisms. The results of functional assays showed that in addition to inhibiting the proliferation, migration, and invasion abilities of HCC cells, melatonin suppressed their DNA repair capacity, thereby promoting the cytotoxicity of chemotherapy and radiotherapy. Whole-transcriptome and gain- and loss-of-function analyses revealed that melatonin induces expression of the long noncoding RNA RAD51-AS1, which binds to RAD51 mRNA to inhibit its translation, effectively decreasing the DNA repair capacity of HCC cells and increasing their sensitivity to chemotherapy and radiotherapy. Animal models further demonstrated that a combination of melatonin and the chemotherapeutic agent etoposide (VP16) can significantly enhance tumor growth inhibition compared with monotherapy. Our results show that melatonin is a potential adjuvant treatment for chemotherapy and radiotherapy in HCC.