RESUMEN

BACKGROUND: Cellular immunotherapy has been widely used in the treatment of solid tumors. However, the clinical application of cord blood-derived dendritic cells and cytokine-induced killer cells (CB-DC-CIK) for the treatment of gastric cancer has not been frequently reported. In this study, the efficacy and safety of CB-DC-CIK for the treatment of gastric cancer were evaluated both in vitro and in vivo. METHODS: The phenotypes, cytokines, and cytotoxicity of CB-DC-CIK were detected in vitro. Patients with advanced gastric cancer were divided into the following two groups: the experimental group (CB-DC-CIK combined with chemotherapy) and the control group (chemotherapy alone). The curative effects and immune function were compared between the two groups. RESULTS: First, the results showed that combination therapy significantly increased the overall disease-free survival rate (P=0.0448) compared with chemotherapy alone. The overall survival rate (P=0.0646), overall response rate (P=0.410), and disease control rate (P=0.396) were improved in the experimental group, but these changes did not reach statistical significance. Second, the percentage of T-cell subsets (CD4(+), CD3(-)CD56(+), and CD3(+)CD56(+)) and the levels of IFN-γ, TNF-α, and IL-2, which reflect immune function, were significantly increased (P<0.05) after immunotherapy. Finally, no serious side effects appeared in patients with gastric cancer after the application of cellular immunotherapy based on CB-DC-CIK. CONCLUSION: CB-DC-CIK combined with chemotherapy is effective and safe for the treatment of patients with advanced gastric cancer.

RESUMEN

BACKGROUND: Currently, cytokine-induced killer cells (CIK)/dendritic cell (DC)-CIK-mediated immunotherapy is widely used to treat gastric cancer. However, limited information regarding clinical trials on CIK/DC-CIK therapy is available. Therefore, systemic evaluation of the efficacy and safety of the combination therapy is necessary. METHODS: A meta-analysis involving 1735 patients with gastric cancer was conducted. Before analysis, the study quality and heterogeneity were evaluated. The effects of chemotherapy combined with CIK/DC-CIK on gastric cancer were compared with the effects observed when chemotherapy alone was used. Pooled analysis was performed using RevMan version 5.2 from random or fixed-effect models. RESULTS: Seventeen trials were included. First, the analysis showed that the combination therapy significantly increased the overall survival rate and disease-free survival rate compared with those in patients treated using chemotherapy alone. The overall response rate (P = 0.002), disease control rate (P = 0.0007), and quality of life improved rate (P = 0.0008) were significantly improved in patients who received combined treatment than in patients who received chemotherapy alone. Second, the percentage of lymphocyte subsets (CD3(+), CD4(+) and CD3(-)CD56(+), CD3(+)CD56(+); P <0.01) and the levels of interleukin-12 and interferon-γ, which reflect immune function, were significantly increased (P <0.05) after the CIK/DC-CIK therapy. Further, carbohydrate antigen tumor markers were significantly reduced compared with the pre-therapy levels. Immunotherapy with CIK/DC-CIK obviously alleviated the adverse events caused by chemotherapy. CONCLUSION: The combination of CIK/DC-CIK therapy and chemotherapy was superior in prolonging the survival time, enhancing immune function and alleviating the adverse events caused by chemotherapy.

Asunto(s)

Antineoplásicos/uso terapéutico , Tratamiento Basado en Trasplante de Células y Tejidos/métodos , Células Asesinas Inducidas por Citocinas , Neoplasias Gástricas/terapia , China , Terapia Combinada , Células Asesinas Inducidas por Citocinas/inmunología , Células Dendríticas/inmunología , Supervivencia sin Enfermedad , Humanos , Inmunoterapia/métodos , Subgrupos Linfocitarios , Calidad de Vida , Neoplasias Gástricas/mortalidad , Tasa de Supervivencia , Resultado del Tratamiento

RESUMEN

AIM: To investigate whether the combination of fluvastatin and losartan synergistically relieve atherosclerosis and plaque inflammation induced by a high-cholesterol diet in rabbits. METHODS: Atherosclerosis was induced with a high-cholesterol diet for 3 months in 36 New Zealand white rabbits. The animals were randomly divided into model group, fluvastatin (10 mg·kg(-1)·d(-1)) group, losartan (25 mg·kg(-1)·d(-1)) group, and fluvastatin plus losartan group. After the 16-week treatments, the blood samples the animals were collected, and the thoracic aortas were examined immunohistochemically. The mRNA and protein expression levels of monocyte chemotactic protein-1 (MCP-1) were measured using RT-PCR and Western blot. RESULTS: Compared to the treatment with losartan or fluvastatin alone, the combined treatment did not produce higher efficacy in reduction of blood cholesterol level. However, the combination did synergistically decrease the intimal and media thickness of thoracic aortas with significantly reduced macrophage infiltration and MCP-1 expression in the plaques. CONCLUSION: The combined treatment with losartan and fluvastatin significantly inhibited atherosclerotic progress and reduced inflammation associated with atherosclerotic plaques.

Asunto(s)

Aterosclerosis/tratamiento farmacológico , Ácidos Grasos Monoinsaturados/uso terapéutico , Indoles/uso terapéutico , Losartán/uso terapéutico , Macrófagos/efectos de los fármacos , Placa Aterosclerótica/tratamiento farmacológico , Animales , Aterosclerosis/inmunología , Aterosclerosis/patología , Quimiocina CCL2/genética , Colesterol/sangre , Sinergismo Farmacológico , Ácidos Grasos Monoinsaturados/farmacología , Fluvastatina , Regulación de la Expresión Génica/efectos de los fármacos , Indoles/farmacología , Inflamación/tratamiento farmacológico , Inflamación/inmunología , Inflamación/patología , Losartán/farmacología , Macrófagos/patología , Placa Aterosclerótica/inmunología , Placa Aterosclerótica/patología , Conejos , Proteínas Quinasas p38 Activadas por Mitógenos/genética

RESUMEN

Ornithine decarboxylase (ODC), the first rate-limiting enzyme of polyamine biosynthesis, was found to be associated with cell growth, proliferation and transformation. ODC gene expression in gastric cancer was increased and its level was positively correlated with the degree of malignity of gastric mucosa and development of gastric lesions. In order to evaluate the therapeutic effects of antisense RNA of ODC on gastric cancer, an antisense RNA of ODC expressing plasmid pcDNA-ODCr which delivered a 120 bp fragment complementary to the initiation codon of ODC gene was constructed and transfected to gastric cancer cells SGC7901 and MGC803. Expression of ODC in gastric cancer cells was determined by western blot. Cell proliferation was assessed by MTS assay. Cell cycle was analyzed by flow cytometry and Matrigel assay was performed to assess the ability of gastric cancer cell invasiveness. The results showed that the ODC gene expression in gastric cancer cells transfected with the pcDNA-ODCr was downregulated efficiently. Tumor cell proliferation was suppressed significantly, and cell cycle was arrested at G1 phase. Gastric cancer cells had reduced invasiveness after gene transfer. Our study suggested that antisense RNA of ODC expressing plasmid pcDNA-ODCr had antitumor activity by inhibiting the expression of ODC, and downregulation of ODC expression using a gene therapy approach might be a novel therapeutic strategy for gastric cancer.

Asunto(s)

Regulación Neoplásica de la Expresión Génica , Ornitina Descarboxilasa/biosíntesis , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/terapia , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Colágeno/química , Combinación de Medicamentos , Citometría de Flujo/métodos , Perfilación de la Expresión Génica , Técnicas de Transferencia de Gen , Terapia Genética/métodos , Humanos , Laminina/química , Oligonucleótidos Antisentido/genética , Plásmidos/metabolismo , Proteoglicanos/química , ARN/metabolismo

RESUMEN

BACKGROUND: Tumstatin is a novel endogenous angiogenesis inhibitor which is widely studied using purified protein. The current study evaluates the antiangiogenic effects of tumstatin-overexpression plasmid in vitro, reveals the mechanism underlying the vascular endothelial cell growth inhibition and searches for a novel method administering tumstatin persistently. METHODS: The eukaryotic expression plasmid pcDNA-tumstatin encoding tumstatin gene was constructed and transfected to human umbilical vein endothelial cell ECV304 and human renal carcinoma cell ACHN. Expression of tumstatin in the two cell lines was determined by RT-PCR and Western blotting. Vascular endothelial cell proliferation was assessed by CCK-8 assay and cell cycle was analyzed by flow cytometry. To investigate the mechanism by which pcDNA-tumstatin inhibited vascular endothelial cell proliferation in vitro, cyclin D1 protein was detected by Western blotting. RESULTS: DNA sequence confirmed that pcDNA-tumstatin was successfully constructed. RT-PCR and Western blotting indicated that tumstatin could express in the two cell lines effectively. After tumstatin gene transfer, ECV304 cell growth was significantly inhibited and the cell cycle was arrested in G1 phase. And Western blotting showed that pcDNA-tumstatin decreased the level of cyclin D1 protein. CONCLUSIONS: Overexpression of tumstatin mediated by pcDNA 3.1 (+) specially inhibited vascular endothelial cells by arresting vascular endothelial cell in G1 phase resulting from downregulation of cyclin D1 and administration of tumstatin using a gene therapy might be a novel strategy for cancer therapy.

Asunto(s)

Autoantígenos/genética , Autoantígenos/metabolismo , Colágeno Tipo IV/genética , Colágeno Tipo IV/metabolismo , Células Endoteliales/citología , Células Endoteliales/metabolismo , Western Blotting , Ciclo Celular/genética , Ciclo Celular/fisiología , Línea Celular , Línea Celular Tumoral , Proliferación Celular , Citometría de Flujo , Humanos , Plásmidos/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa

RESUMEN

OBJECTIVE: Angiotensin converting enzyme 2 (ACE2) efficiently hydrolyses the potent vasoconstrictor angiotensin II to vasodilative angiotensin (1-7). We hypothesized that ACE2 overexpression may inhibit inflammation response in atherosclerotic plaque by degrading Ang II into Ang-(1-7). METHODS: Atherosclerosis (AS) plaques were induced in the abdominal aorta of 38 rabbits by endothelial injury and atherogenic diet for 3 months. Rabbits were then underwent injection of a recombinant adenovirus (2.5 x 10(9) pfu/ml) carrying a murine ACE2 gene (Ad-ACE2) through a catheter into the abdominal aortic segments rich in plaques (n = 19) or injection of a control vector Ad-EGFP (n = 19). One month later, all rabbits were sacrificed and plaques from aortic segments were analyzed. RESULTS: ACE2 expression in aortic tissues of the Ad-ACE2 group were confirmed by immunohistochemistry. Macrophage infiltration (13.6% +/- 4.2% vs. 23.6% +/- 6.9%, P < 0.01) and MCP-1 expression (13.2% +/- 0.4% vs. 25.0% +/- 7.4%, P < 0.01) were significantly reduced in Ad-ACE2 group compared to Ad-EGFP group. CONCLUSIONS: Overexpression of ACE2 inhibited atherosclerotic plaque inflammation response in hypercholesterolemic rabbits.

Asunto(s)

Aterosclerosis/genética , Peptidil-Dipeptidasa A/genética , Transfección , Enzima Convertidora de Angiotensina 2 , Animales , Aterosclerosis/metabolismo , Células Cultivadas , Dieta Aterogénica , Vectores Genéticos , Conejos

RESUMEN

Spermidine/spermine N(1)-acetyltransferase (SSAT) is the rate-limiting step in polyamine catabolism. In a previous study, we constructed a recombinant adenovirus, Ad-SSAT, which can express human SSAT. In the present study, we investigated the effect of upregulated and downregulated SSAT on gastric cancer cells. We found that upregulated SSAT could inhibit the growth of MGC803 and SGC7901 cells, whereas adverse results were found with downregulated SSAT. We further analyzed cell cycle profiles and the expression levels of the major cell cycle regulatory proteins of S phase. The results showed that the growth inhibition was caused by S phase arrest. Ad-SSAT suppressed the expression of cyclin A and nuclear factor E2F1 in MGC803 and SGC7901 cells. We observed the E2F promoter activity caused by Ad-SSAT using a reporter gene assay. We also investigated the antitumorigenicity of upregulated SSAT by Ad-SSAT using a SGC7901 xenograft model in nude mice. Our results suggest that the upregulation of SSAT by Ad-SSAT infection inhibited the growth of gastric cancer in vitro and in vivo. Ad-SSAT arrested gastric cancer cells in S phase, which was mediated through downregulation of the cyclin A-E2F signaling pathway.

Asunto(s)

Acetiltransferasas/fisiología , Adenoviridae/genética , Neoplasias Gástricas/prevención & control , Acetiltransferasas/genética , Animales , Poliaminas Biogénicas/análisis , Línea Celular Tumoral , Ciclina A/antagonistas & inhibidores , Factor de Transcripción E2F1/antagonistas & inhibidores , Vectores Genéticos , Humanos , Ratones , Ratones Endogámicos BALB C , Fase S , Neoplasias Gástricas/química , Neoplasias Gástricas/enzimología , Neoplasias Gástricas/patología , Regulación hacia Arriba

RESUMEN

Spermidine/spermine N1-acetyltransferase (SSAT) is a key enzyme of polyamine catabolism. In a previous study, we constructed a recombinant adenovirus, Ad-SSAT, which can express human SSAT. In the present study, we investigated the effect of Ad-SSAT on the growth and cell cycle of colorectal cancer cells. We found that Ad-SSAT increased the expression of SSAT and inhibited the growth of HT-29 and Lovo cells. The growth inhibition was caused by cell cycle arrest in the S phase. Furthermore, Ad-SSAT was shown to suppress the expression of cyclin A and nuclear factor E2F-1 in HT-29 and Lovo cells. The inhibitory effect of Ad-SSAT on cyclin A promoter activity was also observed in a reporter gene assay. Our results suggest that the expression of SSAT mediated by Ad-SSAT infection inhibits the growth of colorectal cancer cells and induces cell cycle arrest at the S phase, through a mechanism involving the suppression of cyclin A and E2F-1 expression.

Asunto(s)

Acetiltransferasas/genética , Ciclo Celular/fisiología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Espermidina/fisiología , Acetiltransferasas/metabolismo , Adenoviridae/genética , Western Blotting , Línea Celular Tumoral , Ciclina A/metabolismo , Factor de Transcripción E2F1/metabolismo , Expresión Génica , Vectores Genéticos , Humanos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transfección

RESUMEN

OBJECTIVE: The purpose of this study was to test the hypothesis that ACE2 overexpression may enhance atherosclerotic plaque stability by antagonizing ACE activity and converting angiotensin II to angiotensin 1-7. METHODS AND RESULTS: Atherosclerotic plaques were induced in the abdominal aorta of 114 rabbits by endothelial injury and atherogenic diet. Gene therapy was performed in group A at week 4 and in group B at week 12, respectively. Each group of rabbits were randomly divided into 3 subgroups which received, respectively, a recombinant ACE2 expressing vector (AdACE2), a control vector AdEGFP and AdACE2+A779, an antagonist of angiotensin 1-7 receptor. Local ACE2 overexpression attenuated the progression of lesions from week 4 to week 8, but not progression of plaque size from week 12 to week 16. In group B rabbits, local ACE2 overexpression resulted in stable plaque compositions, ie, fewer macrophages, less lipid deposition and more collagen contents, higher plaque stability scores, decreased angiotensin II levels, and increased angiotensin 1-7 levels in plaque tissues in the AdACE2 subgroup compared with those in the AdEGFP subgroup. CONCLUSIONS: Overexpression of ACE2 results in stabilized atherosclerotic plaques and the mechanism is probably the conversion of vasoconstrictive angiotensin II to vessel protective angiotensin 1-7.

Asunto(s)

Angiotensina II/metabolismo , Aorta Abdominal/enzimología , Aterosclerosis/enzimología , Peptidil-Dipeptidasa A/metabolismo , Adenoviridae/genética , Angioplastia de Balón/efectos adversos , Angiotensina I/metabolismo , Angiotensina II/análogos & derivados , Angiotensina II/farmacología , Enzima Convertidora de Angiotensina 2 , Animales , Aorta Abdominal/efectos de los fármacos , Aorta Abdominal/patología , Aterosclerosis/etiología , Aterosclerosis/patología , Línea Celular , Células Cultivadas , Colágeno/metabolismo , Dieta Aterogénica , Grasas de la Dieta/administración & dosificación , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Vectores Genéticos , Humanos , Ratones , Fragmentos de Péptidos/metabolismo , Fragmentos de Péptidos/farmacología , Peptidil-Dipeptidasa A/genética , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/metabolismo , Conejos , Receptores Acoplados a Proteínas G/antagonistas & inhibidores , Receptores Acoplados a Proteínas G/metabolismo , Factores de Tiempo , Transducción Genética , Regulación hacia Arriba

RESUMEN

AIM: To construct a recombinant adenovirus that can express human spermidine/ spermine N1-acetyltransferase (SSAT) and detect its inhibitory effect on colorectal cancer cell growth in vitro. METHODS: A 516 bp cDNA of SSAT was amplified and cloned into a pGL3-hTERT plasmid. The pGL3-hTERT-SSAT recombinant was digested, and the small fragment was cloned into the shuttle vector pAdTrack. The pAdTrack-hTERT-SSAT plasmids were recombined with pAdEasy-1 vectors in AdEasy-1 cells. Positive clones were selected and transfected into the HEK293 packaging cells (transformed human embryonic kidney cells) after they were linearized by PacI. The process of adenovirus packaging and amplification was monitored by green fluorescent protein (GFP) expression. The SSAT protein levels were determined by Western blotting, and the intracellular polyamine content was detected by reverse-phase high performance liquid chromatography. The MTS (3-(4, 5-dimethylthiaol-2-yl)-5-(3-carboxy-methoxyphenyl)-2-(-4- sulfophenyl)-2H-tetrazolium, inner salt) and colony-forming assays were used to analyze the gene transduction efficiency and effect on the growth of HT-29 and LoVo cells. A viable cell count was used to determine the cell growth with or without exogenous polyamines. RESULTS: The GFP expression in 293 cells during virus packing and amplification was observed by fluorescence microscopy. Western blotting results demonstrated that Ad-hTERT-SSAT could increase the expression of SSAT, and consequently, spermidine and spermine were reduced to low levels. The MTS and colony-forming assay results showed that HT-29 and LoVo cell growth were significantly inhibited, and the inhibitory effect could be partially reversed by exogenous spermidine and spermine. CONCLUSION: The successfully constructed recombinant adenovirus Ad-hTERT-SSAT could accelerate polyamine catabolism and inhibit the colorectal cell growth in vitro. It also has therapeutic potential in the treatment of colorectal cancer.

Asunto(s)

Acetiltransferasas/genética , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Espermidina/metabolismo , Espermina/metabolismo , Acetiltransferasas/metabolismo , Adenoviridae/genética , Secuencia de Bases , Línea Celular , ADN Complementario/química , Expresión Génica , Vectores Genéticos , Proteínas Fluorescentes Verdes/metabolismo , Células HT29 , Humanos , Riñón/citología , Datos de Secuencia Molecular , Plásmidos , Transfección