RESUMEN
Minicircle DNA (mcDNA) has been suggested as a vanguard technology for gene therapy, consisting of a nonviral DNA vector devoid of prokaryotic sequences. Unlike conventional plasmid DNA (pDNA), this small vector is able to sustain high expression rates throughout time. Thus, this work describes the construction, production, and purification of mcDNA-p53 and its precursor parental plasmid (PP)-p53 for a comparative study of both DNA vectors in the growth suppression of human papillomavirus (HPV)-18-infected cervical cancer cells. First, live cell imaging and fluorescence microscopy studies allowed to understand that mcDNA-p53 vector was able to enter cell nuclei more rapidly than PP-p53 vector, leading to a transfection efficiency of 68% against 34%, respectively. Then, p53 transcripts and protein expression assessment revealed that both vectors were able to induce transcription and the target protein expression. However, the mcDNA-p53 vector performance stood out, by demonstrating higher p53 expression levels (91.65 ± 2.82 U/mL vs. 74.75 ± 4.44 U/mL). After assuring the safety of both vectors by viability studies, such potential was confirmed by proliferation and apoptosis assays. These studies confirmed the mcDNA-p53 vector function toward cell cycle arrest and apoptosis in HPV-18-infected cervical cancer cells. Altogether, these results suggest that the mcDNA vector has a more promising and efficient role as a DNA vector than conventional pDNA, opening new investigation lines for cervical cancer treatment in the future.
Asunto(s)
Papillomavirus Humano 18/genética , Infecciones por Papillomavirus/terapia , Proteína p53 Supresora de Tumor/genética , Neoplasias del Cuello Uterino/terapia , Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , ADN de Cinetoplasto/genética , ADN de Cinetoplasto/farmacología , Femenino , Técnicas de Transferencia de Gen , Terapia Genética/tendencias , Papillomavirus Humano 18/patogenicidad , Humanos , Infecciones por Papillomavirus/genética , Infecciones por Papillomavirus/patología , Infecciones por Papillomavirus/virología , Plásmidos/genética , Proteína p53 Supresora de Tumor/farmacología , Neoplasias del Cuello Uterino/genética , Neoplasias del Cuello Uterino/patología , Neoplasias del Cuello Uterino/virologíaRESUMEN
Six transmembrane epithelial antigen of the prostate 1 (STEAP1) is overexpressed in numerous types of tumors, especially in prostate cancer. STEAP1 is located in the plasma membrane of epithelial cells and may play an important role in inter- and intracellular communication. Several studies suggest STEAP1 as a potential biomarker and an immunotherapeutic target for prostate cancer. However, the role of STEAP1 in cell proliferation and apoptosis remains unclear. Therefore, the role of STEAP1 in prostate cancer cells proliferation and apoptosis was determined by inducing STEAP1 gene knockdown in LNCaP cells. In addition, the effect of DHT on the proliferation of LNCaP cells knocked down for STEAP1 gene was evaluated. Our results demonstrated that silencing the STEAP1 gene reduces LNCaP cell viability and proliferation, while inducing apoptosis. In addition, we showed that the cellular and molecular effects of STEAP1 gene knockdown may be independent of DHT treatment, and blocking STEAP1 may reveal to be an appropriate strategy to activate apoptosis in cancer cells, as well as to prevent the proliferative and anti-apoptotic effects of DHT in prostate cancer.
Asunto(s)
Andrógenos/farmacología , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Oxidorreductasas/antagonistas & inhibidores , Neoplasias de la Próstata/patología , ARN Interferente Pequeño/genética , Antígenos de Neoplasias/genética , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Humanos , Masculino , Oxidorreductasas/genética , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/metabolismoRESUMEN
Liver, the largest intern organ of the human body, is responsible for several vital tasks such as digestive and excretory functions, as well as for nutrients storage and metabolic functions, synthesis of new molecules and purification of toxic chemicals. Cirrhosis, fibrosis and hepatocellular carcinoma are the most prevalent liver diseases. Despite all the studies performed so far, treatment options for these diseases are very limited. For this reason, it is urgent to find effective therapies for these pathologies. Several studies have been performed during the last decade about the possible application of human amniotic membrane in hepatic diseases therapy. Promising results about human amniotic membrane or its derived cells, in vitro and in vivo, applications in fibrosis, cirrhosis and hepatocellular carcinoma were already published. Since it is an attractive study area, it is becoming a dynamic scientific subject. However, the action mechanisms of human amniotic membrane and its derived cells in hepatic diseases therapy must be precisely known in order that this promising therapy could be clinically used.
