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1.
Am J Transl Res ; 10(5): 1539-1551, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29887967

RESUMO

OBJECTIVE: This study aimed to: evaluate long-term toxicity and pharmacokinetic parameters; to identify the target organ of toxicity of a recombinant adenovirus vaccine expressing human papillomavirus 16 E6 and E7 proteins (HPV16 E6E7-Ad5 Vac) in primates; and to determine the specific immune response of this recombinant adenovirus vaccine. METHOD: HPV16 E6E7-Ad5 Vac (dose 4.68 × 109 IU/bottle) was administered to Macaca fascicularis (M. fascicularis) to evaluate its long-term toxicity. The Cynomolgus Monkeys were divided into a negative control group (sodium chloride injection group), a low-dose group (4.68 × 108 IU/macaque), and, a high-dose group (4.68 × 109 IU/macaque). The drugs were administered at intervals of once every three weeks (D1, D21, D42). The macaques were observed until the sixth week of the recovery period (D84) for safety and toxicological indicators and pharmacokinetic indicators. To study the specific immune response in Rhesus Macaque, empty viruses (rAd5-null) and buffer were inoculated as controls, respectively. Two doses of the vaccine were given at 1.0 × 108 IU/ml and 1.0 × 109 IU/ml and theHPV-16 E6-/HPV-16 E7-specific IFN-γ productions were measured. RESULTS: The macaques of both the high-dose group and the low-dose group did not exhibit any systemic toxic response. The administered safe dose of the vaccine was 4.68 × 109 IU per animal. Following vaccination, HPV16 E6/E7-specific antibodies were observed to be generated in both groups, indicating an immune response of the lymphocytes targeting HPV16 E6 and HPV16 E7 epitopes (specific NF-r) was elicited. The peak level of HPV-16 E6-/HPV-16 E7-specific IFN-γ production was observed in the ninth week.

2.
Exp Ther Med ; 9(4): 1161-1165, 2015 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-25780403

RESUMO

The aim of the present study was to understand the genetic stability of a master seed bank (MSB) and a working seed bank (WSB) of an adenovirus vector vaccine expressing the human papillomavirus (HPV) type 16 E6 and E7 fusion proteins (Ad-HPV16E6E7). Microscopic examination and viral infectious efficacy were used to measure the infectious titers of the Ad-HPV16E6E7 MSB and WSB. Polymerase chain reaction was used to analyze the stability of the Ad-HPV16E6E7 target gene insertion, while western blot analysis and immunofluorescence were used to assess the expression levels of the Ad-HPV16E6E7 target protein. A C57BL/6 mouse TC-1 tumor cell growth inhibition model was used to evaluate the biological effect of Ad-HPV16E6E7 administration. The infectious titers of the Ad-HPV16E6E7 MSB and WSB were 6.31×109 IU/ml and 3.0×109 IU/ml, respectively. In addition, the expression levels of the inserted target genes and target proteins were found to be stable. In the mouse TC-1 tumor inhibition analysis, when the virus titers of the Ad-HPV16E6E7 MSB and WSB were 109 IU/ml, the tumor inhibition rate was 100%, which was significantly different when compared with the control group (χ2MSB=20.00 and χ2WSB=20.00; P<0.01). Therefore, the Ad-HPV16E6E7 vaccine seed bank is genetically stable and meets the requirements for vaccine development.

3.
Hum Vaccin Immunother ; 9(7): 1430-7, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23571172

RESUMO

Enterovirus 71 (EV71) is one of the major causative agents for hand, foot and mouth disease (HFMD) in childhood. Nowadays, HFMD or EV71 infections have already become an important public health issue throughout the world. Vaccination may be the most effective measure to control the transmission of the virus. Therefore, to pave EV71 vaccine into human clinical trial, in the present study a comprehensive preclinical safety assessment of inactivated EV71 vaccine including single- and repeat-dose toxicity studies were conducted in rats and cynomolgus monkeys. No abnormal findings were observed in rats following single intramuscular administration with EV71 vaccine (640 U). The results also showed no obvious systemic toxicities from four repetitive intramuscular injections, with a 14-d interval, of two dosages of EV71 vaccine in the two animal species. Antinuclear antibody response was not detected after the repeated administrations. Histopathological examination demonstrated the minimal to severe inflammatory changes in muscle tissues of the injection sites in EV71 vaccine-injected animals and most of findings have been improved over time. Furthermore, test article could induce highly EV71-specfic neutralizing antibody response in both animal species. Taken together, these data suggested a favorable safety profile for inactivated EV71 vaccine and supported this product to enter human phase I clinical trial.


Assuntos
Enterovirus Humano A/imunologia , Doença de Mão, Pé e Boca/imunologia , Doença de Mão, Pé e Boca/prevenção & controle , Vacinas Virais/efeitos adversos , Vacinas Virais/uso terapêutico , Animais , Anticorpos Antinucleares/sangue , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Feminino , Macaca fascicularis , Masculino , Ratos , Ratos Wistar , Vacinas de Produtos Inativados/efeitos adversos , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/uso terapêutico , Vacinas Virais/imunologia
4.
World J Gastroenterol ; 10(22): 3284-8, 2004 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-15484301

RESUMO

AIM: To isolate the subtypes of 3' region of cagA gene in Helicobacter pylori (H pylori) strains from Zhejiang Province in China and to investigate their relations to H pylori-associated gastroduodenal diseases. METHODS: One hundred and thirty-seven H pylori clinical strains were isolated from the gastric mucosa specimens of 74 patients with chronic gastritis, 61 with peptic ulceration, and 2 with gastric cancer. Bacterial genomic DNA was extracted and 3' region of cagA gene was amplified by polymerase chain reaction (PCR). Subtypes of 3' region of cagA gene were determined by the size of PCR amplified segments. The sequences of the subtypes were analyzed by PCR-based sequencing. RESULTS: Of the 137 H pylori isolates from Zhejiang Province, 132 (96.4%) yielded PCR products that could be classified into three groups of subtypes, named as subtypes I, II, and III according to their sizes. The sizes of subtypes I, II, and III were 648-650 bp, 705-707 bp, and 815 bp, respectively. Among the 132 cagA-positive H pylori strains, 123 (93.2%) belonged to the group of subtype I, 6 (4.5%) presented subtype II, 1 (0.8%) was subtype III, and 2 (1.5%) presented subtypes I and III both. The primary structure of subtype I was composed of 3 repeats of R1, 1 repeat of R2 and 1 repeat of R3. Subtype II possessing 4 repeats of R1, 2 repeats of R2 and 1 repeat of R3 was a newly found type of 3' region of cagA gene which had not been reported before. The primary structure of subtype III consisted of 4 repeats of R1, 1 repeat of R2 and 2 repeats of R3. Comparison of the sequences of subtype I strains with the corresponding sequences deposited in GenBank, showed a similarity of 95.0% (94.0-96.1%) for nucleotide sequences and 95.9% (94.9-97.4%) for deduced amino acid sequences. Comparison of the sequences of subtype III strains with the corresponding sequences deposited in GenBank, showed a similarity of 93.9% (90.8-96.9%) for nucleotide sequences and 93.2% (90.2-96.2%) for deduced amino acid sequences. Among subtype II strains, the nucleotide and deduced amino acid sequences showed a similarity of 95.2% (94.1-96.5%) and 96.4% (93.8-97.9%), respectively. There were no statistical differences in the distribution of subtypes of 3' region of cagA gene among different H pylori-associated gastroduodenal diseases (chi(2) = 11.544, P>0.05). CONCLUSION: There are three subtypes (I, II, and III) of 3' region of cagA gene in H pylori strains isolated from Zhejiang Province, and subtypeIis predominant. Subtype II is a newly found subtype of 3' region of cagA gene. The result of this study does not support the view that the subtypes of 3' region of cagA gene in H pylori isolated from Zhejiang Province are correlated with the clinical outcomes of H pylori infection.


Assuntos
Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Infecções por Helicobacter/microbiologia , Helicobacter pylori/genética , Adolescente , Adulto , Idoso , Sequência de Aminoácidos , Antígenos de Bactérias/química , Proteínas de Bactérias/química , China , Feminino , Gastrite/microbiologia , Helicobacter pylori/isolamento & purificação , Humanos , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Reação em Cadeia da Polimerase , Estrutura Terciária de Proteína
5.
World J Gastroenterol ; 10(14): 2060-2, 2004 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-15237434

RESUMO

AIM: To determine the prevalence of genotypes of cagII in Helicobacter pylori (H pylori)-infected patients in Zhejiang Province and investigate the relationship between these genotypes and the types of gastroduodenal diseases. METHODS: One hundred and seventy one clinical isolates were collected from 70 chronic superficial gastritis, 31 chronic atrophic gastritis, 41 gastric ulcer, 21 duodenal ulcer, 3 gastric and duodenal ulcer, and 5 gastric adenocarcinoma patients. Polymerase chain reaction assays were performed for analysis of cagT, ORF13 and ORF10 genes in the cagII region. RESULTS: Of 171 H pylori isolates from Zhejiang patients, 159(93.0%) were positive for all the three loci. One isolate (0.6%) was negative for all the three loci, and 11(6.4%) were partially deleted in cagII. The positive rates of cagT, ORF13 and ORF10 genes were 97.1%, 94.7% and 99.4%, respectively. In the strains isolated from the patients with diseases including chronic superficial gastritis, chronic atrophic gastritis, gastric ulcer and duodenal ulcer, the sitive rates of cagT were 95.7%, 100.0%, 95.1% and 100.0%, respectively. The positive rates of ORF13 were 94.3%, 93.5%, 95.1% and 100.0%, respectively. The sitive rates of ORF10 were 98.6%, 100.0%, 100.0% and 100.0%, respectively. The three genes were all positive in the three H pylori strains isolated from the patients with both gastric and duodenal ulcer. In the five strains isolated from the patients with gastric adenocarcinoma, only one isolate was negative for ORF13. There were no significant differences of the cagT, ORF13 and ORF10 genes among the different gastroduodenal diseases including chronic superficial gastritis, chronic atrophic gastritis, gastric ulcer, duodenal ulcer, both gastric and duodenal ulcer and gastric adenocarcinoma (chi(2)=3.098, P>0.05 for cagT; chi(2)=3.935, P>0.05 for ORF13 and chi(2)=6.328, P>0.05 for ORF10). CONCLUSION: The cagII is not a uniform and conserved entity. Although the genes in cagII are highly associated with the gastroduodenal diseases, the clinical outcome of H pylori infection is not reliably predicted by the three genes in cagII in patients from Zhejiang Province.


Assuntos
Ilhas Genômicas , Infecções por Helicobacter/genética , Helicobacter pylori , Adolescente , Adulto , Idoso , China , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade
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