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Cross-resistance to human interferon-gamma of human interferon-beta-resistant F-IFr cells.

F-IFr cells, which are more resistant to the anticellular and antiviral action of human alpha and beta interferons (IFN-alpha and IFN-beta) than the parental RSa cells, were also found to be more resistant to both the anticellular and antiviral effect of IFN-gamma. A high level of 2-5A synthetase was induced by treatment with IFN-alpha or -beta, but induction of 2-5A synthetase was not detected after IFN-gamma treatment of the cells. F-IFr cells had less than half the number of specific binding sites for IFN-alpha than the parental RSa cells.

2',5'-Oligoadenylate Synthetase↗

Biological similarities of rat fibroblast interferon to human and mouse alpha interferons.

High titres of rat interferon (IFN) can be produced by rat embryo fibroblast (CD) cells after treatment with Newcastle disease virus. This CD IFN was characterized by SDS-PAGE, and was found to contain three species at 30K to 33K, 25K to 27K and 17K to 22K mol. wt. Antibody affinity chromatography revealed that 95% of the CD IFN bound to an anti-human leukocyte IFN antibody column, 34% to an anti-mouse L cell IFN antibody column and none to an anti-human fibroblast IFN antibody column. The IFN that bound to the anti-human leukocyte IFN antibody column contained all three molecular weight species of IFN. However, the material bound to the anti-mouse L cell IFN antibody column only showed IFN activity of the 30K and 20K species. Examination of the heterologous antiviral activity of the unseparated CD IFN and of the IFN separated by antibody affinity chromatography revealed the same patterns of activity: 35 to 50% of homologous activity on guinea-pig cells, 10 to 20% on mouse cells, 2 to 4% on human cells and none on bovine cells. The data suggest that a major portion of this rat fibroblast IFN is related to alpha IFNs produced by other species.

Animals↗

Isolation of Daudi cells with reduced sensitivity to interferon. IV. Characterization of clones with altered binding of human interferon alpha subspecies.

Three clones of Daudi cells, DIF3, DIF8 and DIF9, have been isolated which possess specific interferon (IFN) receptors even though these cells are resistant to both the antiviral and antiproliferative actions of human IFN-alpha. Studies with 125I-labelled cloned human IFN-alpha subspecies showed that clone DIF3 bound IFN in a manner identical to that of the IFN-sensitive parental cells. The initial peak of binding observed 30 min after treatment of either Daudi or DIF3 cells at 37 degrees C with IFN-alpha 2 or IFN-alpha 8 was, however, either absent or much reduced when using DIF8 cells which are resistant to the antiproliferative action of IFN-alpha 2 and -alpha 8. Similarly, no initial peak of binding was observed after treatment of DIF9 cells with IFN-alpha 2 at 37 degrees C. In accord with its reduced biological activity, the binding of 125I-IFN-alpha 1 to either Daudi or DIF8 cells was considerably lower than that of either of the other two IFN-alpha subspecies and followed the form of a hyperbola without an initial transitory peak of binding. It is suggested that the phenotype of IFN resistance of DIF8 and DIF9 cells may be related to a defect in receptor activation whereas the same phenotype of DIF3 cells would appear to be unrelated to IFN binding.

Cell Division↗

HVJ (Sendai virus) stimulates release of interferon from leukocytes used once for interferon production.

Leukocytes, subjected once to interferon (IFN) induction by HVJ (Sendai virus), were studied for their capability to produce IFN after a second similar stimulus. Substantial amounts of IFN (about 30 000 IU/ml) were recovered. Experiments using cycloheximide or actinomycin D and kinetic studies showed that this IFN originated mainly in IFN which resided within the cell as a result of the first induction and was released after the second stimulation. Increasing amounts of HVJ used for the second stimulus resulted in proportionally increased yields of IFN, reaching a plateau at the same dose of HVJ (1000 HAU/ml) as that which gave optimal yields after the first stimulation. Evidence is presented that the capacity of HVJ to trigger the production of a second IFN harvest is closely associated with its infectivity.

Cells, Cultured↗

Tumour necrosis factor-alpha, interferon-gamma and interferon-beta exert antiviral activity in nervous tissue cells.

The individual and synergistic antiviral effects of cytokines released by infiltrating immune cells or by cells of the nervous system may play an important role in inhibiting virus spread during infections of the central nervous system (CNS). We examined the antiviral activity against the neurotropic pseudorabies virus (PRV) of interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha), and combinations of these cytokines, as compared to that of IFN-beta, in rat nervous tissue cells. PRV replicated efficiently in all neural cell types tested, including neurons, astrocytes and oligodendrocytes. The inhibitory effects were determined by quantifying the inhibition of virus plaque formation, yields of infectious virus at various times after infection and synthesis of viral proteins. At a low m.o.i., IFN-gamma and IFN-beta inhibited viral plaque formation in all cell types; TNF-alpha was effective only in astrocytes but showed synergy with IFN-gamma. At a higher m.o.i., IFN-beta inhibited yields of infectious virus more effectively than IFN-gamma, whereas TNF-alpha had no effect on virus yields and was only marginally synergistic with the antiviral activity of IFN-gamma. The yield-reduction assays correlated well with cytokine-induced inhibition of viral protein synthesis. Our results show that both IFN-gamma and IFN-beta can induce a state of antiviral resistance in neural cells whereas TNF-alpha is effective only in astrocytes at low m.o.i.; they suggest an antiviral role of cytokines in the immune response to virus infections of the CNS.

Animals↗

Interferon-beta strong cytopathic effect on human papillomavirus type 16-immortalized HPK-IA cell line, unexpectedly not shared by interferon-alpha.

We report a novel, unusually severe cytopathic effect of interferon-beta (IFN-beta). Data concerning antibody neutralization, induction and recovery time course, CPE50 dose, impact on oxidative metabolic activity and 1D SDS-PAGE total cellular protein analysis are provided for preliminary characterization. This cytopathic effect appears to be linked to human papillomavirus type 16 (HPV-16) genome presence as it is markedly evident in the HPV-16-immortalized HPK-IA cell line, but is not induced in diploid keratinocytes. It is also not induced in highly malignant SiHa cells suggesting that it also requires a fairly conserved phenotype. This effect is unexpectedly not shared by IFN-alpha pointing to a discrimination between IFN-alpha and -beta signal despite the well-known sharing of a common receptor. It remains to be clarified whether this divergence, undetectable in other cellular systems, represents a direct effect of viral presence or a non-specific consequence of cellular homoeostatic disregulation induced by the papillomavirus genome.

Cell Line, Transformed↗

Role of the protein kinase PKR in the inhibition of varicella-zoster virus replication by beta interferon and gamma interferon.

Varicella-zoster virus (VZV) is sensitive to type I and type II interferons (IFNs), which mediate antiviral effects. In this study, it was demonstrated that IFN-beta and IFN-gamma inhibited the replication of VZV in vitro. Although IFN-beta was more effective than IFN-gamma, the level of inhibition of VZV replication achieved by the combination of both IFNs was more than additive and it was concluded that these two cytokines acted synergistically. Expression of the IFN-induced, double-stranded RNA-activated protein kinase PKR and its phosphorylation level were not modulated strongly during ongoing replication of VZV. However, in the presence of IFN-beta, but not IFN-gamma, PKR expression and its phosphorylation were increased, explaining in part the inhibition of virus replication by IFNs. The expression of herpes simplex virus Us11, a viral protein with several functions, including prevention of PKR activation, strongly increased the level of VZV replication.

Antiviral Agents↗

Sequential production of alpha and beta interferons and gamma interferon in the circulation of Listeria monocytogenes-infected mice after stimulation with bacterial lipopolysaccharide.

Sequential production of interferon (IFN)-alpha/beta and IFN-gamma in the circulation of mice which had been previously infected with viable Listeria monocytogenes was induced by injection of lipopolysaccharide (LPS) derived from Salmonella typhimurium. IFN-alpha/beta production occurred 2 hr after injection of LPS, thereafter IFN-gamma appeared and the maximum titer was demonstrated at 6 hr. At that time, almost all of the IFN was IFN-gamma. IFN-gamma production in response to LPS was observed from the 5th through the 11th day after infection with Listeria, but it was not demonstrated in either mice infected with lower doses of viable Listeria or mice immunized with heat-killed bacteria. IFN-alpha/beta production was not drastically affected by treatment with hydrocortisone, cyclophosphamide, carrageenan, antithymocyte serum, or anti-asialo GM1 antibody, whereas IFN-gamma production was suppressed by administration of all those agents. Noteworthily, IFN-alpha/beta, but not IFN-gamma, was produced even 6 hr after stimulation with LPS in cyclophosphamide- or antithymocyte serum-treated mice. IFN-gamma induction by LPS was markedly suppressed in mice in which IFN-alpha/beta produced by Listeria infection itself had been depleted by treatment with anti-mouse IFN-alpha/beta antibody, but it was not inhibited in mice when IFN-alpha/beta induced not by Listeria infection but by LPS had been depleted by treatment with anti-mouse IFN-alpha/beta antibody.

Animals↗

Human T-lymphoblastoid cell lines with high and low abilities to produce interferon-gamma constitutively and their susceptibilities to interferon.

A human T-lymphoblastoid cell line, TCL-Fuj, produces large amounts of interferon (IFN)-gamma constitutively. A variant cell line, 2M, was derived from it. Both cell lines express similar surface antigen markers, but differ in surface morphology. Compared with the parent TCL-Fuj cell line, 2M produced less IFN-gamma constitutively but more in response to IFN inducers. The IFNs produced constitutively and on stimulation with inducers were analyzed by SDS-polyacrylamide gel electrophoresis. In TCL-Fuj cells, the constitutive and induced IFNs consisted of the same molecular species (22K and 39K). In 2M cells, smaller IFNs were produced constitutively (18K and 32K) and induction resulted in a marked increase of 22K molecules. These two cell lines also differed in sensitivity to the antiviral activity of IFN. Other T-lymphoblastoid cell lines, HPB-ALL and TCL-Fuj 4 cells, which did not produce IFN-gamma were permissive for vesicular stomatitis virus (VSV) replication; its growth was markedly suppressed by IFN-gamma and -alpha. TCL-Fuj cells were also permissive for VSV, but were not susceptible to the antiviral effect of the IFNs. In contrast, in 2M cells the multiplication of VSV was restricted; the viral yield was further reduced by the IFNs and increased by treatment with anti-human IFN-gamma serum. Several clonal cell lines derived from TCL-Fuj and 2M cells had characteristics similar to the respective parent cell lines. The growth of both cell lines was not affected by IFN-gamma or by -alpha. The separation of antiviral and anti-proliferative susceptibilities was peculiar to 2M cells unlike other cell lines.

Cell Line↗

Characterization of anti-interferon-alpha antibodies appearing during recombinant interferon-alpha 2a treatment.

Patients with malignant midgut carcinoid tumours received recombinant interferon-alpha 2a (rIFN-alpha 2a) or rIFN-alpha 2a and chemotherapy (streptozocin and doxorubicin) for 6 months, and then rIFN-alpha 2a alone. Antibodies, mainly of IgG type, binding to rIFN-alpha 2a developed in nine of 22 patients (41%), as determined by immunoassay. In seven patients, antibodies also neutralized the biologic (anti-viral) activity of rIFN-alpha 2a. Anti-IFN-alpha 2a antibodies were equally frequent in both sexes and treatment groups, but were not observed in those patients (n = 8) that had previously received other types of IFN. Antibodies appeared after a median of 6 months of rIFN-alpha 2a treatment and had a median duration of 6 months. The anti-IFN-alpha 2a antibody titres declined with time with no obvious relation to change of therapy, also during continued IFN-alpha 2a treatment. High titres of neutralizing antibodies appeared to impair anti-tumoural effects in individual potential responders. Anti-IFN-alpha 2a antibodies further examined in six patients bound to native IFN-alpha subtypes present in both allogenic and autologous leucocyte IFN-alpha. Such autoantibodies neutralized the biologic activity of autologous IFN-alpha in two patients, and in a third were partially neutralizing.

Aged↗

A preliminary experience with GM-CSF plus interferon in patients with HBV and HCV resistant to interferon therapy.

An open label trial of GM-CSF plus high-dose interferon (IFN) alpha 2b was performed in eight patients with chronic hepatitis B infection and 16 patients with chronic hepatitis C, who either failed to clear virus with 6 months of daily high-dose IFN (5 MU daily) therapy (n = 22) or were considered untreatable because of advanced disease and leukopenia (n = 2). The dose of GM-CSF used was 500 micrograms subcutaneously twice weekly. The dose of IFN used was 5 MU daily. Both agents were administered for 4 months. Five of the eight hepatitis B patients and five of the 16 hepatitis C virus patients responded to combined therapy having previously failed IFN therapy alone. The hepatitis B virus responders had low entry ALT, AST, and gamma GPT levels as compared to the non-responders. No such differences for responders and non-responders were seen with the hepatitis C virus patients. These data suggest that the combination of GM-CSF and IFN may be more effective at achieving viral clearance than IFN alone.

Adult↗

Comparison of long-term effects of lymphoblastoid interferon alpha and recombinant interferon alpha-2a therapy in patients with chronic hepatitis B.

To compare the long-term effect of natural lymphoblastoid interferon-alpha (IFN-alpha nl) and recombinant IFN-alpha 2a therapy in patients with chronic hepatitis B, 210 patients in two trials were followed-up for 1.1-15.5 years following the end of therapy. They included 34 patients who received placebo (control), 67 treated with IFN-alpha nl (36 after prednisolone priming) and 109 treated with IFN-alpha 2a (56 after prednisolone priming). The cumulative sustained response was higher in patients who had been treated with IFN-alpha nl after prednisolone priming than was exhibited using IFN-alpha nl alone, IFN-alpha 2a alone or the placebo (P < 0.05), or IFN-alpha 2a following prednisolone priming (P = 0.052) at the end of 11 years. Hepatocellular carcinoma (HCC) was detected in 1.5% of the IFN-alpha nl group, 3.7% of the IFN-alpha 2a group and 14.7% of the control group (control vs IFN-alpha nl or IFN-alpha 2a, P < 0.05). The cumulative HCC development was higher in the control group than in the IFN-alpha nl group (P < 0.002) and the IFN-alpha 2a group (P = 0.06). The cumulative survival rate was lower in the control group than in the IFN-alpha nl group (P < 0.01) and the IFN-alpha 2a group (P = 0.02). Multivariate analysis revealed that IFN-alpha nl therapy and female gender are significant predictors of sustained response; preexisting cirrhosis, age at entry and IFN therapy are significant factors in both HCC development and survival. In conclusion, IFN-alpha nl treatment may have a better long-term effect on hepatitis B virus (HBV) clearance than IFN-alpha 2a and placebo, and IFN therapy may provide better long-term beneficial effects than placebo in terms of HBV clearance, reduction of HCC and prolonged survival.

Adolescent↗

A randomized trial of prolonged high dose of interferon plus ribavirin for hepatitis C patients nonresponders to interferon alone.

Retreatment of chronic hepatitis C patients nonresponders to interferon (IFN) alone with the standard dose of IFN [3 million units (MU) thrice weekly (TIW)] plus ribavirin for 24 weeks has yielded low sustained virological response (SVR), averaging 8%. The aim of the present, open-labelled, randomized study was to evaluate the efficacy of IFN induction therapy followed by prolonged high dose of IFN plus ribavirin in nonresponders. One hundred and fifty-one patients were randomized to receive 5 MU daily of IFN alfa-2b (group 1, n = 73) or 5 MU TIW of IFN alfa 2b (group 2, n = 78) for 4 weeks followed by IFN (5 MU TIW) plus ribavirin (1000/1200 mg/daily) for 48 weeks in both groups. In an intention-to-treat analysis, the sustained virological response (SVR) at 24-week follow-up was 33 and 23% for group 1 and 2, respectively (P = 0.17). The overall SVR was 52 and 18% in patients with genotype 2/3 and 1/4, respectively. Among genotype 1/4 patients the SVR was 29 and 11% for age younger or older than 40 years. Compared with genotype 2/3 patients, the risk (95% confidence interval) of nonresponse to retreatment was 3.0-fold (1.17-8.0) in younger genotype 1/4 patients and 8.4-fold (3.0-23.29) in older genotype 1/4 patients. In conclusion these results suggest that retreatment with a reinforced regimen should be focused in nonresponder genotype 2/3 patients and younger genotype 1/4 patients, who are most likely to benefit. Induction therapy does not improve SVR.

Adult↗

Comparison of two PEG-interferon alpha-2b doses (1.0 or 1.5 microg/kg) combined with ribavirin in interferon-naïve patients with chronic hepatitis C and up to moderate fibrosis.

Health regulatory approval of the 1.5 microg/kg body weight dose of pegylated interferon (PEG-I) alpha-2b in combination with ribavirin for the treatment of chronic hepatitis C was based on a study using PEG-I alpha-2b at doses of only 0.5 and 1.5 microg/kg body weight (BW), in spite of the previously shown flat dose-response curve at doses of > or =1.0 microg/kg. Our aim was to compare PEG-I alpha-2b 1.0 microg/kg with 1.5 microg/kg, both in combination with ribavirin. Open-label, randomized study in 227 patients with biopsy-proven chronic hepatitis C (Metavir < or =F2), receiving oral ribavirin (400 mg, twice daily) in combination with subcutaneous PEG-I alpha-2b (1.0 or 1.5 microg/kg, once weekly) for 24 weeks (genotype 2 or 3), or 48 weeks (other genotypes), followed by a 24-week drug-free period. Virologic response rates did not differ between the two doses of PEG-I alpha-2b: in patients infected with hepatitis C virus (HCV) genotype 1 or 4 treated with PEG-I 1.0 microg/kg BW, 38% (22/58) had a sustained virologic response compared with 39% (27/70) in the PEG-I 1.5 microg/kg BW dose group (P = ns). The corresponding values in patients infected with HCV genotype 2 or 3 were 71% (39/55) and 81% (29/36) respectively (P = ns). Adverse events led to transient or permanent dose reductions in fewer patients in the 1.0 microg/kg BW dose group (48/113 patients; 42%) than in the 1.5 microg/kg BW dose group (63/106 patients; 59%, P = 0.015). Furthermore, 89% of patients treated for 24 weeks but only 58% of patients treated for 48 weeks (P < 0.001) tolerated the treatment without relevant dose reduction or premature termination. In combination with ribavirin, PEG-I alpha-2b 1.0 microg/kg was as effective as 1.5 microg/kg but was better tolerated in patients with chronic hepatitis C and up to moderate fibrosis.

Adolescent↗

Effects of interferon-alpha/beta and interferon-gamma preparations on phagocytosis by mouse peritoneal macrophages.

The influence of murine alpha/beta-interferon (Mu IFN-alpha/beta) and murine gamma-interferon (Mu IFN-gamma) preparations on the attachment and ingestion phase of phagocytosis by mouse peritoneal macrophages (MPM) was studied. A non-opsonized strain of Escherichia coli, IgG-opsonized E. coli, and sheep erythrocytes opsonized with IgG (E-IgG) and IgM plus complement factor C3b (E-IgMC) were used as test particles. Pretreatment of MPM with 10(2)-10(3) U/ml of Mu IFN-alpha/beta for 24 h enhanced both attachment and ingestion of bacteria or erythrocytes mediated by the non-specific receptor, the Fc receptor, or the C3b receptor. Higher concentrations had no such effects. In contrast, treatment of MPM with 10(1)-10(2)U/ml of Mu IFN-gamma suppressed attachment and ingestion of non-opsonized and IgG-opsonized E. coli and of E-IgG by 10-40%. Mu IFN-gamma did not influence attachment and ingestion of E-IgMC. The effects were neutralized by specific anti-IFN antiserum. The data indicate that the IFN effect on phagocytic activity is, at least to a large extent, due to modifications of the surface receptors.

Animals↗

Interferon-alpha production and tissue localization of interferon-alpha/beta producing cells after intradermal administration of Aujeszky's disease virus-infected cells in pigs.

Intradermal administration of glutaraldehyde-fixed Aujeszky's disease virus (ADV) infected autologous or allogeneic cells resulted in the induction of an interferon (IFN)-alpha response in pigs. Using a sensitive dissociation-enhanced lanthanide fluoroimmunoassay (DELFIA), IFN-alpha was detected in blood at 8 and 24 h after injection of ADV-infected cells. In parallel, by means of in situ hybridization, IFN-alpha/beta mRNA containing cells were demonstrated in regional lymph nodes. Occasional IFN-alpha/beta mRNA positive cells were also seen in injected dermal areas, but not in contralateral lymph nodes, spleen, bone marrow, blood or liver. The ability of leucocytes in whole blood cultures to produce IFN-alpha upon stimulation by ADV was markedly diminished 3-7 days after intradermal injection of ADV-infected cells. In contrast, cultures of purified peripheral blood mononuclear cells (PBMC) had intact IFN-alpha responses. Further, serum from ADV-injected pigs inhibited the in vitro ADV-induced IFN-alpha responses in PBMC from control pigs, most likely due to the demonstrated presence of anti-ADV antibodies. We suggest that the IFN-alpha/beta producing cells in lymph nodes may participate in the development of antiviral immunity and could be equivalent to Natural IFN-alpha/beta producing (NIP) cells.

Animals↗

Quantitative analysis of interferon alpha/beta receptor mRNA in the liver of patients with chronic hepatitis C: correlation with serum hepatitis C virus-RNA levels and response to treatment with interferon.

We analysed the expression of interferon (IFN) alpha/beta receptor mRNA in the liver of patients with chronic hepatitis C and examined the relationship between the expression of this receptor gene and the level of hepatitis C virus (HCV)-RNA as well as the response to 16 weeks of 6 x 10(6) units IFN. The mean level of IFN alpha/beta receptor mRNA in patients with chronic HCV infection (expressed as delta cycle; 10.8 +/- 1.9 (mean +/- SD); n = 39) was significantly higher than that of control subjects (9.4 +/- 0.5; n = 6; P < 0.01). There was a significant negative correlation between the level of IFN alpha/beta receptor mRNA and serum HCV-RNA in 39 patients with chronic hepatitis C (R = -0.546; P < 0.01). The mean level of IFN alpha/beta receptor mRNA in six patients who showed a complete response to IFN therapy (12.3 +/- 1.6) was higher than that of 15 patients who failed to respond to therapy (10.1 +/- 1.5; P < 0.01). Our results are consistent with the suggestion that the anti-viral activity of IFN depends on the level of the IFN alpha/beta receptor on hepatocytes in patients with chronic hepatitis C.

Adult↗

Efficacy of hepatitis B vaccination and interferon-alpha-2b combination therapy versus interferon-alpha-2b monotherapy in children with chronic hepatitis B.

BACKGROUND: Although interferon (IFN) has been approved in the treatment of chronic hepatitis B in children, it is effective only in 30-40% of patients. In some studies it has been suggested that therapeutic use of anti-hepatitis B virus (HBV) vaccine may be beneficial in patients with chronic hepatitis B. The aim of the present study was to compare the efficacy of hepatitis B vaccination and IFN-alpha-2b in combination and IFN-alpha-2b monotherapy in children with chronic hepatitis B. METHODS: Fifty treatment-naive children with chronic hepatitis B infection were randomly assigned to receive either 5 million units/m(2) recombinant IFN-alpha-2b subcutaneously three times per week for 9 months, and pre-S2/S vaccine at the beginning and 4 and 24 weeks after initiation of IFN therapy (n = 25) or recombinant IFN-alpha-2b (5 million units/m(2) subcutaneously thrice weekly) alone for 9 months (n = 25). Children were followed for at least 6 months after the end of therapy. RESULTS: There was no statistically significant difference in the mean alanine aminotransferase levels, histologic activity index and fibrosis scores between combination and IFN monotherapy groups at the end of the therapy and end of the follow-up period. When combination and monotherapy groups were compared, the mean HBV-DNA values were significantly reduced in combination group at the end of the therapy (P = 0.004), but no statistically significant difference was found at the end of the follow up. Sustained HBeAg seroconversion with clearance of HBV-DNA was obtained in 13 of 25 children (52%) treated with combination therapy, and in eight of 25 patients (32%) treated with IFN monotherapy (P = 0.251). CONCLUSION: Although the difference was statistically insignificant, the sustained response rates were better in the combination therapy group than in the monotherapy group. The potential benefit of combining IFN and hepatitis B vaccine should be investigated in further studies with different regimens of combination therapy.

Adolescent↗