RESUMEN
The present study aimed to investigate the effects of photothermal therapy with gold nanorods (AuNRs) or epidermal growth factor receptor monoclonal antibodyconjugated AuNRs (EGFRmAbAuNRs) on hypopharyngeal carcinoma (HC) in nude mice. In addition, the associated signaling pathways were explored. Briefly, a subcutaneous transplantable hypopharyngeal tumor model was established in nude mice injected with FaDu human HC cells. A total of 70 nude mice were randomly divided into seven groups, each of which received a different treatment. Mice were treated with AuNRs, locally or through intravenous injection, whereas EGFRmAb or EGFRmAbAuNRs were only administered locally. Near infrared spectroscopy (NIR) was also applied for plasmonic photothermal therapy (PPTT). The growth curve and the inhibitory rate for tumor growth were used to evaluate the effects of each treatment. Flow cytometry and the terminaldeoxynucleotidyl transferase dUTP nick end labeling assay were adopted to detect apoptosis of cells in the transplanted tumors. Reverse transcriptionquantitative polymerase chain reaction and western blotting were used to determine the mRNA and protein expression levels of target genes, respectively. Local treatment with AuNRs + NIR or EGFRmAb significantly inhibited tumor growth, and EGFRmAb conjugation further increased the inhibitory effects. Furthermore, there was a significant increase in apoptosis of tumor cells in the AuNRs + NIR, EGFRmAb and EGFRmAbAuNRs + NIR groups; treatment with EGFRmAbAuNRs + NIR induced the highest apoptotic effect. Mechanistic studies indicated that EGFRmAbAuNRs + NIR may inhibit tumors through the AKT serine/threonine kinase (Akt) and DNA damage signaling pathways. In the AKT pathway, the mRNA and protein expression levels of phosphatase and tensin homolog were increased, whereas the expression levels of Akt and glycogen synthase kinase 3ß were decreased. In the DNA damage signaling pathway, the mRNA and protein expression levels of ATR serine/threonine kinase, checkpoint kinase 1 and p53 were enhanced, whereas phosphorylatedp53 protein expression was reduced. The present findings indicated that AuNRs + NIR inhibited HC tumor growth, and conjugating EGFRmAb to AuNRs further enhanced the inhibitory effects. EGFRmAb conjugation may increase the antitumor effects of AuNRsinduced PPTT by downregulating the phosphatidylinositol3kinase/Akt pathway and upregulating the DNA damage pathway. These findings may provide novel insights into tumortargeting PPTT in vivo.
Asunto(s)
Antineoplásicos Inmunológicos/administración & dosificación , Oro/administración & dosificación , Hipertermia Inducida/métodos , Neoplasias Hipofaríngeas/terapia , Administración Intravenosa , Animales , Antineoplásicos Inmunológicos/química , Antineoplásicos Inmunológicos/farmacología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Receptores ErbB/antagonistas & inhibidores , Oro/química , Oro/farmacología , Humanos , Neoplasias Hipofaríngeas/genética , Neoplasias Hipofaríngeas/metabolismo , Ratones , Ratones Desnudos , Nanotubos/química , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , Distribución Aleatoria , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Hypopharyngeal carcinoma (HC) is one of the most malignant tumors in the upper aerodigestive tract. Currently, there are no effective treatments for HC. Gold nanoparticles (AuNPs) are a promising tool that can be used for plasmonic photothermal therapy (PPTT), which refers to the use of electromagnetic radiation, most often in near infrared (NIR) region, for the treatment of various medical conditions including cancer. AuNPs have been proved to be a promising tool for NIR spectroscopy-mediated photothermal therapies. In this study, we chemically conjugated AuNPs with a monoclonal antibody (mAb) targeting the epidermal growth factor receptor (EGFR), a cell-surface receptor that is overexpressed in many cancers. We then assessed the effect of NIR photothermal treatment with the EGFRmAb-AuNPs in FaDu HC cells. Our data showed that nanoparticle conjugation with the EGFRmAb improved the specific targeting towards FaDu cells and reduced cytotoxicity towards normal (293 T) cells which do not overexpress the EGFR. A significant amount of our EGFRmAb-conjugated AuNPs could enter the nucleus. Moreover, the expression levels of double strand DNA break repair proteins, including p-ATR, p-CHK1, and p-CHK2 were increased following AuNPs treatment, indicating the presence of DNA damage. These findings suggest that the AuNPs can potentially disrupt genome integrity and induce apoptosis. In addition, EGFRmAb-AuNPs+NIR could induce FaDu cell apoptosis, accompanied by the inhibition of the PI3K/AKT/mTOR pathway and stimulation of DNA damage response. Based on these data, PPTT using the EGFRmAb-AuNPs could be a new promising treatment for HC.
Asunto(s)
Apoptosis/efectos de los fármacos , Daño del ADN , Inmunoconjugados/farmacología , Fosfotransferasas/metabolismo , Transducción de Señal/efectos de los fármacos , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/inmunología , Apoptosis/efectos de la radiación , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/patología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Supervivencia Celular/efectos de la radiación , Receptores ErbB/inmunología , Oro/química , Células HEK293 , Humanos , Neoplasias Hipofaríngeas/genética , Neoplasias Hipofaríngeas/metabolismo , Neoplasias Hipofaríngeas/patología , Inmunoconjugados/química , Inmunoconjugados/inmunología , Rayos Infrarrojos , Nanopartículas del Metal/química , Fosfatidilinositol 3-Quinasas/metabolismo , Fototerapia/métodos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Transducción de Señal/genética , Transducción de Señal/efectos de la radiación , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Gold nanorods (AuNRs) have been used in plasmonic photothermal therapy (PPTT), which is thought to be more efficient and selective than conventional photothermal therapy. The efficiency and safety of PPTT can be improved by functionally modifying the gold nanorods with proteins or biomolecules. In this study, AuNRs were modified with anti-epidermal growth factor receptor (EGFR) monoclonal antibody (mAb), and the apoptotic potential of EGFRmAb-AuNR was assessed in Hep-2 cells in vitro and in vivo. The EGFRmAb modification had no obvious influence on the original optical property of the AuNRs, but it significantly increased the entry of AuNRs into Hep-2 cells. EGFRmAb-AuNRs, with appropriate laser irradiation, resulted in higher Hep-2 cells apoptosis than AuNRs did alone, in vitro, and was accompanied by alteration of reactive oxygen species (ROS) production, Ca(2+) release, change in mitochondrial membrane potential (ΔΨm), cytochrome c (Cyt-c) release, active caspase-3 expression, and level of B-cell lymphoma 2 (Bcl-2) and B-cell lymphoma 2 protein-associated X protein (Bax). EGFRmAb-AuNR-mediated apoptosis in Hep-2 cells was also observed in vivo and had an inhibitive effect on growth of Hep-2 tumor xenografts. Our data suggest that the EGFRmAb modification improves AuNR-mediated apoptosis and may have the potential to be used clinically.