Human Necrosis Factor Research Articles

Research progress of adalimumab in pediatric Crohn ′s disease

Adalimumab is an anti-TNF-α recombinant IgG1, a fully humanized monoclonal antibody binding soluble human tumor necrosis factor alpha with high affinity and specificity.It has lower immunogenicity and can be used clinically for induction and maintenance of remission in Crohn′s disease.The effectiveness, sustainability and safety of adalimumab in the treatment of pediatric Crohn′s disease has been gradually confirmed, which may pave the way for new therapy to treat children with refractory Crohn′s disease. Key words: Adalimumab; Crohn′s disease; Children

Effect of Tripterygium glycosides combined with Recombinant human tumor necrosis factor receptor II antibody fusion protein for injection on serum vascular endothelial growth factor, rheumatoid factor, receptor activator of nuclear factor κ B ligand levels in patients with rheumatoid arthritis

Objective To investigate the effect of tripterygium glycosides combined with recombinant human tumor necrosis factor receptor Ⅱ antibody fusion protein for injection on serum vascular endothelial growth factor(VEGF), rheumatoid factor(RF), receptor activator of nuclear factor κ B ligand(RANKL) levels in patients with rheumatoid arthritis.To provide reference for rational clinical application. Methods From December 2014 to January 2016, 132 patients with rheumatoid arthritis were divided into observation group and control group by random number table method, , with 66 cases in each group, The control group was treated with tripterygium glycosides alone (10 mg each time, 3 times daily, orally) for 3 months.The observation group was treated with a combination of tripterygium glycosides and recombinant human tumor necrosis factor receptor Ⅱ (0.4 mg/kg, once weekly, hypodermic injection)for 3 months.The clinical efficacy, and serum VEGF, RF and RANKL levels were compared between 2 groups. Results The effective rate of the observation group was 92.4% (61/66), which was significantly higher than that in the control group (80.3%, 53/66). There was a significant difference between the two groups (χ2=4.117, P 0.05). After treatment, the levels of serum VEGF, RF and RANKL in the observation group and the control group were (20.8±11.5) ng/L and (27.3±13.1)ng/L, (258.4±54.5)U/L and (298.1±49.5)U/L, (0.083±0.021) pmol/L and (0.197±0.064), respectively.There were significant differences between the two groups (t=3.029, 4.381, 13.750, all P<0.05). The incidence rate of adverse reactions in the control group was 10.5% (7/66), which was significantly higher than that in the observation group (1.5%, 1/66). There was a significant difference between the two groups(χ2=4.790, P<0.05). The one-year recurrence rate was 25.0% (13/52) in the control group and that was 6.7% (4/60) in the observation group, respectively, and there was a significant difference between the two groups(χ2=7.272, P<0.05). Conclusion Tripterygium glycosidescombined with recombinant human tumor necrosis factor receptor Ⅱ antibody fusion protein for injection is effective in the treatment of rheumatoid arthritis, which reduces the levels of serum VEGF, RF and RANKL, and has a low incidence of adverse reactions and recurrence. Key words: Tripterygium glycosides; Recombinant human tumor necrosis factor receptor Ⅱ antibody fusion protein for injection; Rheumatoid arthritis; Vascular endothelial growth factor; Rheumatoid factor; Receptor activator of nuclear factor κB ligand

Anti-rheumatic drugs combined with recombinant human tumor necrosis factor receptor antibody fusion protein lead to fatal lung infection and death: a case report

Anti-rheumatic drugs combined with recombinant human tumor necrosis factor receptor antibody fusion protein lead to fatal lung infection and death: a case report

Recombinant human tumor necrosis factor receptor type II: IgG Fc fusion protein combined with methotrexate for the treatment of psoriasis and their effects on levels of interleukin-17A and tumor necrosis factor-α

Objective To evaluate the effect of recombinant human tumor necrosis factor receptor type Ⅱ: IgG Fc fusion protein (rhTNFR:Fc, trade name Etanercept) combined with methotrexate on levels of interleukin-17A (IL-17A) and tumor necrosis factor-α (TNF-α) in the serum and mononuclear cells of patients with moderate to severe plaque psoriasis. Methods A total of 30 patients with moderate to severe plaque psoriasis were enrolled from Department of Dermatology of Tenth People′s Hospital of Tongji University between August 2014 and February 2016, and then were randomly and equally divided into Etanercept group and Etanercept+ methotrexate group. The treatment lasted 24 weeks. Fifteen healthy blood donors served as healthy control group. Enzyme-linked immunosorbent assay (ELISA) and real-time quantitative PCR were performed to measure the serum levels and mRNA expression of IL-17A and TNF-α, respectively, in the patients of the above two groups before and after the treatment. Results Before the treatment, the serum levels of IL-17A and TNF-α, as well as the mRNA expression of IL-17A and TNF-α in peripheral blood mononuclear cells (PBMCs) , were all significantly higher in all the patients than in the healthy controls (all P < 0.05) . After the treatment, compared with the Etanercept group, the Etanercept+ methotrexate group showed significantly lower serum levels of IL-17A (142.67 ± 14.82 vs. 163.54 ± 23.18,P < 0.05) and TNF-α (70.07 ± 25.02 vs. 91.98 ± 14.62, P < 0.05) , as well as lower mRNA expression of IL-17A (1.12 ± 0.33 vs. 1.56 ± 0.77, P < 0.05) and TNF-α in PBMCs (2.50 ± 1.04 vs. 3.61 ± 2.14, P < 0.05) . Conclusion Etanercept combined with methotrexate is superior to Etanercept alone in the treatment of psoriasis, and can reduce treatment duration and improve therapeutic effect, likely by down-regulating the expression of IL-17A and TNF-α. Key words: Psoriasis; Receptors, tumor necrosis factor, type II; Recombinant fusion proteins; Methotrexate; Interleukin-17; Tumor necrosis factor-alpha

Recombinant human tumor necrosis factor receptor II: IgG Fc fusion protein for the treatment of erythrodermic psoriasis: a clinical observation

Objective To evaluate the efficacy of recombinant human tumor necrosis factor receptor Ⅱ: IgG Fc fusion protein (rhTNFR∶Fc) for the treatment of erythrodermic psoriasis. Methods A total of 23 patients with erythrodermic psoriasis were subcutaneously injected with rhTNFR∶Fc 25 mg (starting dose, 50 mg) twice a week for 12 weeks. Thereafter, they were followed up for 2 years. Outcome measures included the proportion of patients achieving 50%, 75% and 90% reduction in psoriasis area and severity index (PASI50, PASI75 and PASI90) , the serum level of tumor necrosis factor-α (TNF-α) , and incidence of adverse reactions. Statistical analysis was carried out by using nonparametric Friedman test and repeated measures ANOVA with the software SPSS 19.0. Results After 12-week treatment, the PASI score significantly decreased from 57.35 ± 3.45 at the baseline to 5.57 ± 3.60 in the patients with erythrodermic psoriasis (P < 0.01) , with the proportion of patients achieving PASI50, PASI75 and PASI90 being 100% (23 cases) , 95.65% (22 cases) , and 60.87% (14 cases) respectively. The serum level of TNF-α showed a significant decrease from 62.87 ± 15.23 ng/L before the treatment to 4.57 ± 2.99 ng/L (P < 0.01) after 12-week treatment. No adverse reactions were observed during the treatment. After 24-month follow-up, PASI scores experienced no significant changes in these patients compared with those at the end of 12-week treatment, and the number of patients achieving PASI50, PASI75 and PASI90 was 23, 20 and 15 respectively. The serum level of TNF-α was significantly lower at the end of 24-month follow-up than at that of 12-week treatment (3.37 ± 1.62 vs. 4.57 ± 2.99 ng/L, P < 0.05) . Conclusion rhTNFR∶Fc is effective for the control of acute inflammation in erythrodermic psoriasis. Key words: Psoriasis; Tumor necrosis factor-alpha; rhTNFR: Fc; TNF-alpha antagonists

Clinical observation and effect of stains collapse of traditional Chinese medicine in combination with tumor necrosis factor antagonist in treating ankylosing spondylitis

Objective To study the clinical effect, safety of stains collapse of traditional Chinese medicine (TCM) combined with tumor necrosis factor-alpha (TNF-α) antagonist-a recombinant human tumor necrosis factor receptor:Fc fusion protein (rhTNF:Fc) and sulfasalazine (SSZ) in treatment of active ankylosing spondylitis (AS). Methods A total of 98 patients of AS of Gansu Provincial Hospital were divided into two groups in random, the western medicine group (rhTNF:Fc and SSZ, n=46) and the combining group (stains collapse of TCM, rhTNF:Fc and SSZ, n=52). The Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), the Bath Ankylosing Spondylitis Functional Index (BASFI), Patient Global Assessment (PGA), Visual Analogue Scale (VAS) of night pain, VAS of total pain, time of morning stiffness, TCM symptom score, distance between occipital and wall, distance between finger and ground, thoracic activity, Schober test, and erythrocyte sedimentation rate (ESR), C reactive protein (CRP), adverse reactions before and after treatment were observed and the clinical effect was evaluated in two groups. Results A total of 86 patients achieved the study finally, 45 patients in the combining group and 41 patients in the western medicine group. The effective rate of ASAS20 was better in the combining group than that in the western medicine group(χ2=4.814, P=0.028), the Bath ankylosing spondylitis disease activity index (BASDAI) 50 showed no significant difference between the two group (χ2=0.100, P=0.752). The total effective rate in the combining group than that in the western medicine group(χ2=3.873, P= 0.049). In comparison with the same group before treatment, the above indicators were improved after treatment (P 0.05). The adverse reactions showed no significant difference (P > 0.05). Conclusions Stains collapse of TCM combined with rhTNF:Fc was more effective than single method in treatment of AS, it can alleviated the symptoms, body signs and body function, decrease indexes of inflammation, which has dramatic effect and good safety in clinic. Key words: Spondylitis, ankylosing; Observation; Nursing care; Tumor necrosis factor-α; Stains collapse of traditional Chinese medicine

Clinical analysis of serum and pleural effusion inflammatory and specific indexes in children of mycoplasma pneumonia complicated with pleural effusion

Objective To investigate the changes of serum and pleural effusion inflammatory indexes in children with mycoplasma pneumoniae complicated with pleural effusion. Methods Enzyme linked immunosorbent assay (ELISA) double antibody sandwich method was used in the three of pleural effusion group, mycoplasma pneumonia group, control group and other groups to detect serum and pleural effusion human interleukin 6 (IL-6). human interleukin 2 (IL-2) and TNF-(human tumor necrosis factor alpha) etc.Inflammation index levels were detected and compared. Results The group of patients with mycoplasma pneumonia mycoplasma pneumonia patients was (9.93±1.05)pg/mL for the IL-6 level, IL-2 levels for (4.93±0.55)μg/mL, (1.45±0.17) ng/mL for the TNF alpha level, which were significantly higher than those of the control group, the IL-2 was significantly lower than that of the control group, with statistically significant difference (t=6.25, 5.88, 4.63, 5.95, 3.69, 5.95, all P<0.05). Comparing pleural effusion group and mycoplasma pneumonia group, we found serum IL-6, TNF alpha levels of pleural effusion group were significantly higher than that of mycoplasma pneumonia group, while IL-2 was lower than that in group mycoplasma pneumonia, with statistically significant difference (t=4.02, 3.06, 3.06 and 2.88, all P<0.05). The levels of serum IL-6, IL-2 and TNF-in the serum of patients with different effusion levels were significantly different from the other two groups (all P<0.05). Pleural effusion IL-6 and TNF level of pleural effusion group and mycoplasma pneumonia patients was significantly higher than those of the control group and IL-2 were significantly lower than that of the control group, the difference had statistical significance (t=5.69, 3.69, 602, 4.11, 2.94, 3.67, all P<0.05). Pleural effusion pleural effusion IL-6 and TNF levels were significantly higher than those in the mycoplasma pneumonia group and IL-2 were significantly lower than that in mycoplasma pneumonia group, the difference had statistical significance (t=3.58, 4.13, 3.21, all P<0.05). The inflammatory indexes of patients with different effusion volume could be found in the pleural effusion, in patients with a large number of pleural effusion, the IL-2, IL-6 and TNF-levels compared with the other two groups showed significantly different (t=6.11、5.44、4.82, all P<0.05). Conclusion IL-6, IL-2 and TNF alpha in the occurrence and development of mycoplasma pneumoniae pneumonia play an important role.Suffering from mycoplasma pneumonia in patients with pleural effusion, whoseTNF-and IL-6 inflammatory index and serum indexes are abnormal, is of certain clinical practical value for clinical diagnosis, treatment and prognosis, as the indicator of the development and prognosis of pneumonia and pleural effusion. Key words: Pneumonia, mycoplasma; Pleural effusion; Serum Inflammatory index

Comparison of effects of fentanyl, sufentanil and remifentanil on immune function of dendritic cells in human umbilical cord blood

Objective To compare the effects of fentanyl,sufentanil and remifentanil on the immune function of dendritic cells in human umbilical cord blood.Methods Human umbilical cord blood mononuclear cells were obtained by density gradient centrifugation and seeded in 24-well plates with a density of 1 × 106/ml (2ml/hole).The cells were randomly divided into 7 groups (n =15 each):control group (group C),fentanyl 1.0 ng/ml group (group F1),fentanyl 5.0 ng/ml group (group F5),sufentanil 0.1 ng/ml group (group S1),sufentanil 0.5 ng/ml group (group S5),remifentanil 1.0 ng/ml group (group R1),and remifentanil 5.0 ng/ml group (group R5).The cells were incubated for 10 days in serum-free culture medium containing 50 ng/ml recombinant human granulocyte colony stimulating factor,10 ng/ml recombinant human interleukin-4 or the corresponding concentration of fentanyl,sufentanil or remifentanil,and then 50 ng/ml recombinant human tumor necrosis factor alpha was added to the culture medium and the cells were incubated for another 4 days in the seven groups.Three holes in each group were chosen and the cell morphology was examined with inverted microscope.Six holes in each group were chosen for determination of the concentration of IL-12 in the supernatant and expression of CD80/CD86.Six holes in each group were chosen for measurement of the cell viability.Results Compared with group C,the concentration of IL-12 and cell viability were significantly decreased and the expression of CD80/CD86 was down-regulated in groups F5,S1,S5,R1 and R5 (P ＜ 0.05).The concentration of IL-12,cell viability and expression of CD80/CD86 were significantly lower in groups S1 and R1 than in group F1 (P ＜ 0.05).Compared with group F5,the concentration of IL-12 was significantly decreased in group S5,and the concentration of IL-12 and cell viability were significantly decreased and the expression of CD80/CD86 was down-regulated in group R5 (P ＜ 0.05).The concentration of IL-12 and cell viability were significantly lower in group R1 than in group S1 (P ＜ 0.05).The concentration of IL-12,cell viability and expression of CD80/CD86 were significantly lower in group R5 than in group S5 (P ＜ 0.05).Conclusion Remifentanil has stronger inhibitory effect on the immunological function of dendritic cells in human umbilical cord blood than sufentanil,and the inhibitory effect of sufentanil is stronger than that of fentanyl. Key words: Fentanyl; Sufentanil; Piperidines; Immunity, cellular; Dendritic cells

Effects of recombinant human TNFR: Fc fusion protein on rat acute liver injury induced by lipopolysaccharide

Objective To investigate the protective effects and the undedying mechanism of recombinant human tumor necrosis factor receptor: Fc fusion protein (Yisaipu, rhu TNFR: Fc) on the lipopolysaccharide (LPS) induced acute liver injury of rats. Method Totally48 SD rats were randondy divided into four groups , in-cluding control gronp (n = 12), Yisaipu group(n = 12), LPS gronp(n = 12) and Yisaipu + IPS group(n = 12). The models of acute liver injury were produced by injection of LPS intravenously. Being fasted for 12 h, the rats were anaesthetized (60 mg/kg pentobarbital sodium, i.p.) and cannulated into carotid arteries. The cannula was connected with the multi-channel creature signal analysis system. The rata in control group and LPS group were injected with normal saline or LPS in dose of 5 mg/kg through rats' sublingual vein respectively. While the rats in Yisaipu group and Yisaipu + LPS group was pretreated with Yisaipu in dose of 0.4 mg/kg subcutaneously 24 h be-fore normal saline or LPS infusion. Six rats of each goup were randomly selected and mean arterial pressure (MAP) were monitored for 6 h via multi-channel creature signal analysis system, and rats' survival rate was calcu-lated. The rats whose MAP less than 10 mmHg were considered to die and the alive rats during period of observa-tion sacrificed by exsanguination. The liver tissue at the same site was removed, fixing in 10% formalin or stored at -80 ℃. To detect serum TNF-α, alanine aminotransferase (ALT) and aspartate aminotransferase (AST) level, 0.2 mL blood samples were collected from the carotid artery 2 and 3 h after the injection of saline or LPS. The serum was collected from centrifuged blood samples and stored at -80 ℃. Enzyme-linked immunosorbent assay (ELISA) and flow cytometry were used to assess serum TNF-α level and bioactivity respectively. We also measured the serum ALT and AST levels, the myeloperoxiase (MPO) and superexide dismutase (SOD) activity, and malon-dialdehyde (MDA) content in liver tissue The pathology of hepatic tissue was evaluated by HE staining. Statistical-ly,the data of TNF-α level and bioactivity, ALT and AST release, and MDA content were analyzed by ANOVA, and rat survival rate were analyzed by Chi-square Tests. Results The rats in control group and Yisaipu group were all survived. Rat survival rate was significantly higher in Yisaipu + LPS group (67%) than in LPS group (17%) (P < 0.05). Serum TNF-α bioactivity was significantly lower in Yisaipu + LPS group than in LPS group [(7.3±2.8)% vs.(51.3±6.4)%, P <0.05]. Compared with IPS group, Yisaipu pretreatment decreased MDA content [(1.40±0.10)vs. (2.81±0.11) nmol/mgprot, P <0.05]and MPO acticity [(0.38±0.04) vs. (0.54±0.02) U/g, P <0.05]in hepatic tissue, while SOD activity [(188.4±20.2) vs. (142.5 ± 18.3) U/mgprot, P <0.05]was increased. The serum AST level, ALT level and the pathology in the liver were also ameliorated correspondingly. Conclusions These data suggest that Yisaipu could protect rats from LPsinduced a-cute liver injury by inhibiting TNF-α bioactivity and by enhancing anti-oxidation. Key words: Yisaipu; Tumor necrosis factor-α; Lipopolysaccharide; Acute liver injury

Effects of ovarian carcinoma cells on the maturation and function of human dendritic cells

To investigate the effects of ovarian carcinoma cells on the differentiation, maturation, and function of the dendritic cells (DC). Human epithelial ovarian carcinoma cells were isolated from specimens of ovarian carcinoma from 12 patients obtained during operation, and cultured. Peripheral blood samples were collected from these patients. DC were isolated and co-cultured with recombinant human tumor necrosis factor (TNF)-alpha to promote their maturation. Immunostaining and flow cytometry (FC) were used to detect the DC specific marker CD1a, maturation marker CD83, co-stimulating factors CD86 and human leucocyte antigen (HLA)-DR. FITC-labeled glucan was added and FC was used to detect the phagocytic function. T cells were co-cultured with immature and mature DC of different concentrations respectively for 96 h, 18 h before the end of culture 3H-TdR was added, liquid scintillation counter was used to measure the level of count per minute (cpm) Ovarian carcinoma cells and peripheral mononuclear cells were put into Transwell to contact each other directly or indirectly, TNF-alpha was add to promote the maturation, and then FC was used to detect the CD1a, CD83, CD86, and HLA-DR levels, and the glucan uptake level. ELISA was used to detect the levels of IL-12, IL-10, and interferon (IFN)-gamma, and Western blotting was used to detect the p-38 protein and phosphorylated p-38 protein in the supernatant. After stimulation of TNF-alpha the expression; levels of CD1a, CD86, CD83, and HLA-DR of the DCs were remarkably increased. Immature DCs showed a strong ability at phagocytosis of glucan, and this ability was decreased by the stimulation of TNF-alpha. The proliferative function of mature DCs on allogenic T lymphocytes was significantly increased dose-dependently (P<0.05). Co-culture with ovarian carcinoma cells decreased the expression levels of CD1a, CD86, and HLA-DR, increased the expression level of CD83, and decreased the ability in up taking glucan by 43.05%. The proliferative function of the DC cultured in direct contact with ovarian carcinoma cells was decreased by 56.35%, and such inhibitory function of the DCs cultured not in direct contact with ovarian carcinoma cells was weaker, and the levels of CD1a, CD86, CD83, and HLA-DR expressed by the DC cultured not in direct contact with ovarian carcinoma cells was between those of the DC cultured in direct contact with ovarian carcinoma cells and those of the DC undergoing routine culture. The levels of IL-12 and phosphorylated p-38 protein were decreased in the supernatant of the DC co-cultured with ovarian carcinoma cells. Ovarian carcinoma cells suppress the differentiation of monocyte into DC, negatively regulates the phagocytotic and antaean managing functions of DC, thus the DC can not effectively stimulate the proliferation of T lymphocytes and inhibit the activity of cytotoxic T lymphocytes. That may be one of the underlying mechanisms of immune escape of tumor and immune tolerance of T lymphocytes.

In vitro gene imaging by luciferase to detect the expression and effect of human tumor necrosis factor related apoptosis-inducing ligand in lung cancer A549 cells

Objective To detect the expression and effect of human tumor necrosis facctor related apoptosis-inducing ligand(hTRAIL)in vitro by using a novel double expressing adenoviral vector encoding hTRAIL and firefly lueiferase (luc) gene (Ad-hTRAIL-luc),in which luc wag used, as reporter gene.Methods A549 cells were transduced with the adenoviral vector encoding enhanced green fluorescent protein (EGFP) gene(Ad-EGFP)at variable multiplicity of infection(MOI).Adenoviral transducfion efficiency wag determined 48 h later.A549 cells were transduced with Ad-hTRAIL-luc at variable MOI.and the following tests were performed 48h later,respectively:the expressive ratio of hTRAIL and the apeptotic ratio of A549 cells were meagnred by flow eytometer;counts per minute(cpm)of luminescence were measurde by scintiUation counters. A549 cells were transduced with Ad-luc at variable MOI, and cpm of luminescence was measured by scintillation counters 48 h later. After A549 cells were transduced with AdhTRAIL-luc,the expressive ratio of hTRAIL,the apoptotic ratio of A549 cells and cpm of luminescence were analyzed by one-way ANOVA.The positive ratio of EGFP and cpm of luminescence (Ad-luc) were analyzed bv nonparametric ANOVA.Results After A549 cells were transfected with Ad-hTRAIL-luc,the expressive ratio of hTRAIL on the cell membrane of the groups were(2.37±0.04)/,(3.16±0.03)/,(3.64±0.03)/,(3.96±0.02)/,(4.24±0.02)/,(4.34±0.02)/ respectively,which showed significant difference between each other (P＜0.01);and the apoptotic ratio of A549 cells were (1.52±0.04)/,(2.93±0.02)/,(3.39±0.02)/,(3.64±0.02)/,(3.86±0.02)/,(4.08±0.02)/,(4.20±0.02)/,respectirely,and it showed significant difference between each other (P＜0.01);cpm of luminescence were 465 561±26 801,1 038 576±29 417,937 655±23 197,786 432±20 028,524 288±16 338,401 566±15 961,respectively,and it also showed significant difference between each other(P＜0.01).There was a positive relationship between the expressive ratio of hTRAIL and cell apoptotic ratio of A549 cells (r=0.984,P＜0.01).Conclusion The double expressing adenoviral vector Ad-hTRAIL-luc can transfer luc and hTRAIL gene to A549 cells efficiently,and the activity of luc may reflect the effect of hTRAIL as well as the expression of hTRAIL. Key words: Gene expression regulation; Receptors,yulnor necrosis factor; Gene therapy; Diagnostic imaging

Clinical evaluation and radiological observation of recombinant human tumor necrosis factor receptor-Fc fusion protein in treatment of active ankylosing spondylitis

To evaluate the efficacy of recombinant human tumor necrosis factor receptor-Fc fusion protein (rh TNFR:Fc) in the treatment of active ankylosing spondylitis (AS). 68 patients with active AS underwent subcutaneous injection of rh TNFR: Fc 25 mg twice a week for 24 weeks. The following indexes were observed: improvement of at least 20% of reported symptoms, based on the multicomponent Assessments in AS (ASAS) response criteria (ASAS 20) at weeks 2, 6, 12 and 24, ASAS 50 and ASAS 70 responses, and improved scores on individual components of ASAS, including Bath AS disease activity index (BASDAI), acute phase reactants, and Bath AS metrology index (BASMI). And the X-ray images of patient's pelvis at weeks 0, 12, and 24 were graded based on BASRI. After treatment of rh TNFR:Fc the primary and secondary efficacy end points of the 68 patients were all improved compared with the baseline values. During the process of treatment, the number of patients with the efficacy achieving ASAS20 was gradually rising: 44 cases (64.7%) at week 12 and 58 (85.0%) at week 24. The efficacy of 41 cases (60.3% ) reached ASAS50 and that of 34 cases reached ASAS70 at week 24. Other efficacy end points also reflected the similar effect and were improved significantly compared with the baseline values at different time points ( all P < 0.01). BASRI showed that the radiological improvement of sacroiliac joint was not significant (P > 0.05) and that the radiological improvement of hip was not obvious at week 12 (P > 0.05) but significant at week 24 (P < 0.05). rh TNFR:Fc rapidly improves the signs and symptoms of active AS.

In vitro growth inhibition of cisplatin-resistant human lung cancer cell lines by recombinant human tumor necrosis factor and/or recombinant human interferon-gamma by virtue of collateral sensitivity.

The colony-inhibitory effects of recombinant human tumor necrosis factor (rH-TNF) and recombinant human interferon-gamma (rH-IFN-gamma) were evaluated in four human lung cancer cell lines and their cisplatin-resistant sublines. The cell lines tested were PC-7 and PC-9 (adenocarcinoma), H69 and N231 (small cell lung cancer) and four cisplatin-resistant sublines, PC-7/1.0, PC-9/0.5, H69/0.2 and N231/0.2, which were 20.0, 7.1, 4.8 and 8.4 fold resistant to cisplatin, respectively, compared to the respective parental cell line in terms of IC50 in a soft agar colony assay. All parental cell lines were resistant to rH-TNF and rH-IFN-gamma, alone or in combination. However, two resistant sublines showed sensitivity to rH-TNF and rH-IFN-gamma. Colony formation by PC-9/0.5 was significantly inhibited, in the absence or presence of cisplatin, by 10(2) U/ml of rH-TNF (less than 50% of control) and the inhibition was synergistic with that produced by 10(3) or 10(4) U/ml of rH-IFN-gamma. RH-IFN-gamma inhibited the colony formation of H69/0.2 only at the highest concentration tested (10(4) U/ml) (less than 50% of control) and the combined effect with rH-TNF was additive. These results suggest that rH-TNF and rH-IFN-gamma may have some potential in overcoming cisplatin resistance by virtue of collateral sensitivity.

A multicenter randomized phase III trial of domestic product of rmhTNF in the treatment of non-small cell lung cancer

To evaluate and compare the effects and toxicity of the domestic product of recombinant mutant human tumor necrosis factor (rmhTNF) combined with chemotherapy and chemotherapy alone in the treatment of patients with non-small cell lung cancer (NSCLC). Two hundred patients with NSCLC in multicenter were randomly devided into trial group (150 cases) and control group (50 cases). Chemotherapy with CAP regimen was given to the patients. Meanwhile, rmhTNF injection of 4×10⁶U/m² was also given from the 1st to 7th days, the 11th to 17th days on the chemotherapy cycle in the trial group. The control patients received chemotherapy alone. Twenty-one days were as a cycle, 2 cycles were given to each patient. The chemotherapeutic effects and toxicity were observed and compared between the two groups after the therapy. of the 200 patients, 5 cases in the trial group and 3 cases in the control group were out of the trial because of economy. The other 192 cases (145 cases in the trial group and 47 cases in the control group) could be analyzed and evaluated the clinical effects and toxicity. The response rate of chemotherapy was 46.90% (68/145) in the trial group and 17.02% (8/47) in the control group respectively ( P =0.001). The KPS scores was 86.02±9.74 in the trial group, and 80.14±9.10 in the control group ( P =0.025). No significant difference of degree III+IV toxicity was observed between the two groups ( P > 0.05). The side effects related to rmhTNF included slight fever, cold-like symptoms, pain and red and swelling in the injection site. All of them were mild and didn't need any treatment and disappeared after the therapy. There were no severe abnormality of liver and kidney function and ECG in both groups. The results demonstrate that the effects of domestic rmhTNF combined with chemotherapy are remarkably higher than that of chemotherapy alone in the treatment of NSCLC. rmhTNF can increase the sensitivity to chemotherapy and improve the quality of life of the patients with slight toxicity. Hence rmhTNF is worth expanding clinical use.

Regulation of telomerase activity by recombinant human tumor necrosis factor in vitro

Regulation of telomerase activity by recombinant human tumor necrosis factor in vitro

Expression of a partial synthetic human TNF cDNA in E. coli.

A recombinant human tumour necrosis factor (rhTNF) cDNA was constructed. The TNF gene was isolated from a human genomic gene library. There are four exons in the TNF gene. The fourth exon codes for 140 amino acids of the TNF matured protein which is composed of 157 amino acids. A major portion of the fourth exon was isolated and then ligated to a synthesized DNA fragment coding for the remaining amino acids. The partial synthetic hTNF (rhTNF) cDNA thus generated was subcloned into a vector and successfully expressed in E. coli. 5-1 fermentator was used to produce rhTNF. About 20 g (wet weight) of bacterial pellet per liter medium and 10(6)-10(7) units of cytotoxicity to L929 cells per milliliter medium were obtained. rhTNF was purified by HPLC and dried with a freeze dryer. rhTNF with a purity of about 95% in the form of white powder was obtained. The sequence of ten amino acids at the amino terminus of the rhTNF was determined. The result showed that it was identical with that of the natural human TNF.

Isolation, characterization and expression of the complementary DNA for human tumor necrosis factor (TNF-alpha).

A cDNA for human TNF-alpha (615bp) was isolated by means of polymerase chain reaction (PCR) using first strand cDNA from PMA-induced HL-60 cells as template. The result from sequencing the 615 bp cDNA fragment indicated that it corresponded to the entire sequence of mature human TNF coding region. Direct expression of mature human TNF was achieved using a plasmid pHT-1 constructed by ligation of the cDNA and a synthetic DNA. The IPTG-induced bacterial product (hTNF) showed cytotoxicity to mouse L-929 cells. The TNF activity was further identified by neutralization of a specific monoclonal antibody against human TNF-alpha. Approximately 80,000 units of activity were detected per ml of culture at A600 = 2.

In vitro synergistic effects of natural human tumor necrosis factor and natural human interferon-alpha.

Tumor necrosis factor and interferons are multifunctional cytokines. The present studies were undertaken to investigate the biologic interactions of highly purified natural human tumor necrosis factor and highly purified natural human interferon-alpha, which were derived from a B cell acute lymphatic leukemia line (BALL-1 cells) sensitized with hemagglutinating virus of Japan (HVJ). Combined treatment with natural human tumor necrosis factor and natural human interferon-alpha synergistically inhibited the in vitro proliferation of P4788 cells derived from a human colon cancer. Flow cytometric analysis of the cell cycle of asynchronous cells indicated that target cells treated with natural human tumor necrosis factor alone accumulate in the S phase. This accumulation in the S phase of the cell cycle was augmented by combined treatment with natural human tumor necrosis factor and natural human interferon-alpha. The growth inhibitions appeared to be a result of arrest in the S phase of the cell cycle. Combined treatment with these cytokines had potent cytostatic and cytotoxic effects on most of the tested malignant cell lines of human epithelial origin.

Potential limitation of growth-inhibitory action of recombinant human tumor necrosis factor (PAC-4D) due to easy induction of resistance: evidence in vitro.

The growth-inhibitory activity of recombinant human tumor necrosis factor (rH-TNF: PAC-4D) against 32 human cultured cell lines derived from leukemias and lymphomas (7 T cell, 11 B cell, 5 non-T,non-B cell and 9 myeloid cell lines) was measured quantitatively by in vitro regrowth assay. The growth of two non-T,non-B-cell lines (Reh, P30/OHK) and six myeloid cell lines (ML-1, U937, THP-1-0, P31/FUJ, P39/TSU, HEL) was found to be significantly inhibited by a 72 hr treatment with PAC-4D. Although the levels of sensitivity of these cell lines to PAC-4D were different, it was common to all these cell lines that increasing the dose of PAC-4D did not augment the growth-inhibitory action above a certain level. Neither dose-dependent nor time-dependent growth-inhibitory action was observed, namely, exposure to 100 U/ml of PAC-4D for 48 hr was sufficient to exhibit maximum growth-inhibitory action. Furthermore U937 cells were found to become completely resistant to PAC-4D during a continuous 12-day exposure to it. This resistance, however, was lost on culture of the cells with PAC-4D-free growth medium for 15 days. These results suggested that some non-T,non-B acute lymphoblastic leukemias and acute myelogenous leukemias might show an initial response to PAC-4D therapy, but prolonged administration might induce resistance to PAC-4D rather than increase the anti-tumor effect.