rIFN Gamma Research Articles

P004 Changes in thymic recent output function in patients with acute myeloid leukemia

The influence of recombinant human interferon alpha 2a (rIFN alpha), recombinant human interferon beta 1 (rIFN beta), and recombinant human interferon gamma (rIFN gamma) on human dermal microvascular endothelial cells (HDMEC) cultured in vitro was studied in various rIFN concentrations (0.1 lU/ml-104 IU/ml) over 2,3,4,6,8, and 10 d. Cell morphology and ultrastructure, cell proliferation, expression of class II alloantigens (HLA-DR and HLA-DQ), and intercellular adhesion molecule-1 (ICAM-1) were investigated using an in vitro technique established in our laboratory. All rIFN tested induced alterations of typical HDMEC morphology; the cells became spindle-shaped and fibroblastoid, although they maintained their endothelial cell marker expression. Also, all IFN dose- and time-dependently inhibited the proliferation of HDMEC in vitro (rIFN alpha> beta> gamma), whereby rIFN alpha exerted the strongest growth-inhibitory effect. Alkaline phosphatase anti-alkaline phosphatase (APAAP) immunocytochemistry of the cultured cells showed dose- and time-dependent stimulation of ICAM-1 and class II antigen expression only by rIFN gamma (HLA-DR> HLA-DQ), rIFN alpha and beta did not exert any immunomodulatory activity on HDMEC in vitro. These results indicate that HDMEC are an important target for the action of IFN. Besides growth inhibition, it seems that rIFN gamma in particular may be involved in the modulation of leucocyte adhesion and trafficking by altering the immunophenotype of the endothelial cell population.

Structural equation model testing and the quality of natural killer cell activity measurements

BackgroundBrowne et al. [Browne, MacCallum, Kim, Andersen, Glaser: When fit indices and residuals are incompatible. Psychol Methods 2002] employed a structural equation model of measurements of target cell lysing by natural killer cells as an example purportedly demonstrating that small but statistically significant ill model fit can be dismissed as "negligible from a practical point of view".MethodsReanalysis of the natural killer cell data reveals that the supposedly negligible ill fit obscured important, systematic, and substantial causal misspecifications.ResultsA clean-fitting structural equation model indicates that measurements employing higher natural-killer-cell to target-cell ratios are more strongly influenced by a progressively intrusive factor, whether or not the natural killer cell activity is activated by recombinant interferon γ (rIFN γ). The progressive influence may reflect independent rate limiting steps in cell recognition and attachment, spatial competition for cell attachment points, or the simultaneous lysings of single target cells by multiple natural killer cells.ConclusionsIf the progressively influential factor is ultimately identified as a mere procedural impediment, the substantive conclusion will be that measurements of natural killer cell activity made at lower effector to target ratios are more valid. Alternatively, if the individual variations in the progressively influential factor are modifiable, this may presage a new therapeutic route to enhancing natural killer cell activity. The methodological conclusion is that, when using structural equation models, researchers should attend to significant model ill fit even if the degree of covariance ill fit is small, because small covariance residuals do not imply that the underlying model misspecifications are correspondingly small or inconsequential.

Down-regulation of Th1-mediated pathology in experimental arthritis by stimulation of the Th2 arm of the immune response.

To assess whether systemic administration of recombinant interferon-gamma (rIFN gamma), a proinflammatory cytokine that influences the differentiation of naive T cells into Th1 cells, promotes the induction of arthritis in DBA/1 mice immunized with type II collagen (CII) in Freund's incomplete adjuvant (IFA) and to determine the antiarthritic effect of treatment with CII in IFA. DBA/1 mice were immunized with CII in IFA and injected intraperitoneally with rIFN gamma (8,000 units/mouse/day) or with recombinant interleukin-12 (rIL-12; 100 ng/mouse/day). In another experiment, mice were immunized with CII in Freund's complete adjuvant (CFA) and treated with a single dose of CII in IFA on the day of immunization or on the day of disease onset. Mice were monitored to assess the effect of the treatment on the severity of disease. Th1/Th2 cytokines and anti-CII antibodies were measured by enzyme-linked immunosorbent assay. The administration of rIL-12 or rIFN gamma to mice immunized with CII in IFA restored the Th1 response and resulted in the development of arthritis. We then determined whether immunization with CII in IFA had the capacity to prevent and/or ameliorate collagen-induced arthritis. A single intraperitoneal injection of CII in IFA prevented arthritis when given at the time of immunization with CII in CFA and reduced disease severity when given at the time of arthritis onset. The administration of CII in IFA resulted in a profound down-regulation of IFN gamma production and an up-regulation of IL-10 in cultures of draining lymph node cells. These findings demonstrate that it is possible to deflect an ongoing pathogenic Th1 response to an antigen by reimmunization of the same antigen with a Th2-polarizing adjuvant.

Changes in barrier function of a model intestinal epithelium by intraepithelial lymphocytes require new protein synthesis by epithelial cells.

Elements of the mucosal immune system may play an important part in regulating epithelial barrier function in the intestinal tract. Intraepithelial lymphocytes (IELs) represent a subtype of immunocyte which is strategically placed to regulate epithelial function at most mucosal sites. An IEL derived cell line (SC1) was used to examine its effects on the model epithelium T84--a tumour derived cell line which retains the phenotype of colonic crypt cells. Transepithelial electrical resistance (TER) was used as a marker of epithelial integrity. Coculture of T84 cells with SC1 produced a significant fall in TER as did exposure of T84 monolayers to IEL derived supernatant. Recombinant interferon-gamma (rIFN gamma) also reduced TER in T84 monolayers. Cycloheximide prevented the effects of IEL supernatant and of rIFN gamma on TER. The fall in TER in response to rIFN gamma was attenuated by blocking antibodies, which did not alter the fall in resistance induced by IEL supernatant. Fractions of IEL supernatant, separated on the basis of size, evoked temporally distinct changes in TER. Ultrastructural studies support the hypothesis that the slow onset but severe fall in TER indicates catastrophic effects on the monolayer. The more rapid onset fall in TER was not associated with gross changes in monolayer morphology. Reduction of TER by IEL supernatant was not influenced by inhibitors of tyrosine phosphatase or of protein kinase C. Although herbimycin did reduce the rapid onset change in TER, the tyrosine kinase inhibitor genistein did not alter responses to IEL supernatant. Mucosal T cells may influence barrier function by a process involving new protein synthesis by epithelial cells. This model may have relevance in some inflammatory conditions of the gastrointestinal tract.

Ginsenoside-Rh1 and Rh2 inhibit the induction of nitric oxide synthesis in murine peritoneal macrophages.

The effects of ginsenoside-Rh1 and Rh2 in the induction of nitric oxide (NO) synthesis by IFN-gamma plus LPS were investigated using murine peritoneal macrophages. The NO production from rIFN gamma plus LPS-treated macrophages was markedly reduced by ginsenoside-Rh1 or Rh2 in a dose dependent manner, but had no inhibitory effects by ginsenoside-Rb1, Rc or Re. In addition, treatment of the cells with ginsenoside-Rh2 6 hr before the stimulation with IFN-gamma plus LPS showed more inhibitory effect than the treatment with ginsenoside-Rh2 6 hr after or simultaneously with the stimulation with IFN-gamma plus LPS in the NO production. Ginsenoside-Rh2 also effectively inhibited IFN-gamma induced NO production when the cells were treated with IFN-gamma 6 hr after the treatment with ginsenoside-Rh2. Our findings suggest that this phenomenon might be caused by inhibition of priming signal such as IFN-gamma for the synergistic induction of NO synthesis.

Expression of ICAM-1 and TNF alpha in human alveolar macrophages from lung-transplant recipients.

Local activation of macrophages may play an important role in the immune process of pulmonary infections and in the inflammatory response of lung allograft rejection. To document macrophage activation within human lung allografts displaying various complications, we have investigated ICAM-1 expression in freshly isolated alveolar macrophages (AM) from lung-transplant recipients by immunocytofluorimetric analysis, and rIFN gamma induced in vitro by ELISA. A total of 21 bronchoalveolar lavage fluids (BAL) from 13 transplanted patients displaying no complication, acute rejection, bacterial/fungal infection, or CMV infection entered the study. ICAM-1 was expressed at a higher level in rejecting patients. Surprisingly, TNF alpha release from AM upon in vitro activation was significantly decreased during rejection. Furthermore, we have studied the effects of the glucocorticoid dexamethasone, the key drug for the treatment of allograft rejection, on the expression of ICAM-1 and TNF alpha induced in vitro in AM, at the levels of protein production and of transcription. Whereas dexamethasone did not influence ICAM-1 expression in AM, it downregulated TNF alpha production at least in part at the transcriptional level. Our results suggest strongly that the anti-inflammatory effects of corticosteroids are not related to ICAM-1 modulation on human AM but to the downregulation of the proinflammatory cytokine TNF alpha that is produced early in the inflammatory process. Moreover, our model of human AM activation induced in vitro by rIFN gamma appears a useful tool for in vitro investigation of the cellular and molecular targets of anti-inflammatory drugs for a more appropriate use.

Augmented Procoagulant Activity in Cancer Patients, Treated with Recombinant Interferon-γ in Addition to Recombinant Tumor Necrosis Factor-α and Melphalan

Several investigators have reported that interferon-gamma (IFN gamma) can alter tumor necrosis factor alpha induced effects in vitro. We assessed in vivo effects of recombinant interferon-gamma (rIFN gamma) on recombinant tumor necrosis factor-alpha (rTNF alpha) induced activation of systemic blood coagulation in a non-randomized study in 20 consecutive cancer patients. Eight patients were treated with rIFN gamma prior to and during hyperthermic isolated limb perfusion with rTNF alpha and melphalan (IFN gamma group). They were compared with twelve patients who did not additionally receive rIFN gamma (non-IFN gamma group). Before start of perfusion, higher levels of TNF alpha, F1+2 and TAT levels were found in the IFN gamma group. Fibrinogen and ATIII levels tended to be lower in this group. High TNF alpha levels, due to leakage during perfusion, were associated with activation of coagulation in all patients, that became obvious after the end of perfusion, when heparin treatment had been antagonized. Activation, measured by increased F1+2 and TAT levels, was significantly stronger in the IFN gamma group. Monocytic TF remained low, possibly due to shedding of TF positive vesicles and/or sequestration of TF positive activated monocytes against the vessel wall. In both groups F1+2 and TAT levels declined 24 h after the perfusion, whereas monocytic TF increased to levels that were higher in the IFN gamma group. In conclusion, our data confirm a strong activation of coagulation induced by rTNF alpha in cancer patients. They suggest that rIFN gamma may lead to a slight activation of coagulation and augments TNF alpha induced procoagulant activity. These effects may be due to rIFN gamma induced sustained monocytic TF activity.

STREPTOCOCCAL PYROGENIC EXOTOXINS A (SpeA) AND C (SpeC) STIMULATE THE PRODUCTION OF INDUCIBLE NITRIC OXIDE SYNTHASE (iNOS) PROTEIN IN RAW 264.7 MACROPHAGES

Streptococcal pyrogenic exotoxins A (SpeA) and C (SpeC) are members of a family of superantigens produced by group A streptococci that appear to play a key role in the pathogenesis of streptococcal toxic shock syndrome. Since it is known that nitric oxide (NO) and tumor necrosis factor (TNF) are largely responsible for the shock and multiple organ dysfunction of Gram-negative sepsis, we hypothesized that SpeA and/or SpeC could trigger the production of inducible nitric oxide synthase (iNOS) and/or TNF by murine macrophages. We exposed RAW 264.7 macrophages to increasing concentrations of SpeA or SpeC alone and in combination with recombinant murine interferon-gamma (rIFN gamma) for 16-24 h. We found that both SpeA and SpeC triggered iNOS production in the presence of low concentrations of rIFN gamma, while neither provoked iNOS accumulation in the absence of rIFN gamma. Neither SpeA nor SpeC (with or without rIFN gamma) reproducibly induced TNF production by these murine macrophages. These data indicate that two streptococcal exotoxins up-regulate iNOS production by murine macrophages and suggest that nitric oxide production may play an important role in the pathogenesis of streptococcal toxic shock syndrome.

Cytokine Secretion by Macrophages in the Rat Testis1

The rat testis contains a large population of resident macrophages, the physiological roles of which are yet to be established. To investigate the functional capacity of these cells, we have analyzed the secretion of the cytokines interleukin (IL)-1, IL-6, tumor necrosis factor alpha (TNF alpha), and granulocyte macrophage-colony stimulating factor (GM-CSF) by isolated testicular macrophages (TMs) and, for comparison, by isolated rat peritoneal macrophages (PMs). Cells were cultured for 48 h in serum-free medium alone or with lipopolysaccharide (LPS, 10 micrograms/ml) and/or recombinant interferon-gamma (rIFN gamma, 200 U/ml). Specific bioassays were used to measure cytokines in the media collected from cultures. Basal production of IL-1, TNF alpha, and IL-6 by TMs and PMs were similar, but TMs produced 8-fold greater levels of GM-CSF than did PMs. LPS, alone or in combination with IFN gamma, significantly enhanced the secretion of all cytokines by PMs (340-840% increase). LPS alone had little effect on TM secretion except to reduce GM-CSF levels some 4-fold. The addition of LPS and IFN gamma increased IL-1, IL-6, and TNF alpha levels (200-750% increase) and reduced GM-CSF levels to 45% of basal levels. Treatment of cultures with indomethacin to minimize prostaglandin production enhanced the LPS-induced effects in both cell types. Expression of the mRNA for each cytokine in cultures of testicular and peritoneal macrophages, as well as in intact testis, was confirmed by reverse transcription polymerase chain reaction. These studies indicate that macrophages resident within the rat testis have a novel cytokine secretion profile and an altered responsiveness to inflammatory activators compared with macrophages from the peritoneal cavity. This may be important in physiological processes in the testis and may contribute to the dysfunctional afferent immune activity thought to underlie the immunologically privileged status of the testes.

High avidity IFN-neutralizing antibodies in pharmaceutically prepared human IgG.

This paper demonstrates and characterizes naturally occurring antibodies to interferon (IFN) in human IgG preparations. In vitro neutralization of the antiviral effect of IFN alpha and IFN beta, but not IFN gamma, was observed in 12 of 15 normal IgG preparations. The neutralizing capacity was higher against rIFN alpha 2A and rIFN alpha 2C than against lymphoblastoid IFN alpha and IFN beta. Frühsommer meningoencephalitis hyperimmune IgG and hepatitis-B hyperimmune IgG showed potent neutralization, whereas anti-rhesus D-, anti-rabies-, and anti-tetanus IgG showed weak neutralization. Saturable binding of 125I-rIFN alpha 2A was demonstrated only in those IgG preparations found to neutralize the antiviral effect of IFN. Significant correlation between IFN binding and neutralization capacity was observed. The antibodies bound with Fab to rIFN alpha 2A with an avidity of approximately 30 pM; the majority was of the IgG1 subclass. Maximum binding capacity was 490 pg rIFN alpha 2A/mg IgG. Cross-binding of rIFN alpha 2C, lyIFN alpha N1 and IFN beta occurred with 10 and 100-200 times lower activities than that of rIFN alpha 2A. There was no cross-binding with rIFN gamma or rIL-6. IgG preparations containing anti-IFN antibodies blocked the binding of 125I-rIFN alpha 2A to A549 cells. In conclusion, pharmaceutically prepared human IgG preparations contain variable but significant levels of high-avidity IFN alpha and IFN beta neutralizing antibodies.

Lysis of endothelial cells by autologous lymphokine-activated killer cells.

The mechanisms of lysis of endothelial cells derived from human umbilical vein (HUVEC) by autologous lymphokine-activated killer (LAK) cells, generated from cord blood lymphocytes of the same donor, were investigated. Freshly isolated HUVEC as well as HUVEC cultured for several passages were efficiently lysed by autologous LAK cells, and their susceptibility to the LAK cells was almost the some as that of allogenic HUVEC. Complement-depletion experiments revealed that the lysis was mainly dependent on CD16+ natural killer (NK) LAK cells. pretreatment of HUVEC with recombinant interferon gamma (rIFN gamma) for 24 h made them resistant to lysis by autologous LAK cells, while pretreatment with either rIL-1 beta. rTNF alpha, or acidic or basic fibroblast growth factor did not alter the lytic sensitivity of HUVEC. The resistance of rIFN gamma-treated HUVEC was specific to lysis by CD16+ NK LAK cells, and their lysis by CD3+ T-LAK cells was not significantly altered. Moreover, in comparison with control HUVEC or rIL-1 beta-treated HUVEC, rIFN gamma-treated HUVEC had a significantly less potent inhibitory effect on the lysis of untreated HUVEC, when used as an unlabeled target. This suggests that rIFN gamma treatment may down-regulate the recognition of some molecules on HUVEC by rIL-2-activated NK cells. These data suggest that damage of the endothelium during LAK therapy is mainly dependent on LAK cells with a NK phenotype that can specifically recognize a certain molecule on autologous endothelial cells.

Effects of nIFN β and rifn γ on growth and morphology of two human melanoma cell lines: Comparison between two‐and three‐dimensional culture

We compared the anti-proliferative effects of natural interferon beta (nIFN beta) and recombinant interferon gamma (rIFN gamma) on 2 human melanoma cell lines, IGRI and SK-Mel28, grown in 2-dimensional monolayer and in 3-dimensional spheroid culture. In monolayer culture, growth of both lines was inhibited in a dose-dependent manner by 5-day treatments with IFN in concentrations ranging between 1 and 5,000 IU/ml. Incubations with 120 IU/ml nIFN beta or 25 IU/ml rIFN gamma led to a 50% growth inhibition of IGRI cells. A 50% growth inhibition of SK-Me128 cells was obtained with 60 IU/ml nIFN beta, whereas even 5,000 IU/ml rIFN gamma inhibited the growth of this line by only 30%. Growing these melanoma cell lines in 3-dimensional spheroid culture for 5 days reduced their sensitivity to interferon. Growth inhibition values of 50% were achieved with 3,000 IU/ml rIFN gamma or 9,000 IU/ml nIFN beta for IGRI spheroids and 10,000 IU/ml nIFN beta for SK-Me128 spheroids, while 10,000 IU/ml rIFN gamma reduced the growth of SK-Me128 spheroids by only 25%. Outgrowth tests showed that the proliferative capacity after 5-day incubations with IFN was only reduced in IGRI spheroids treated with high doses of nIFN beta. The macroscopically observed increased density of interferon-treated spheroids could be confirmed by light microscopy as corresponding to reduced intercellular space in these spheroids. Scanning electron microscopy furthermore showed variations on the surface of IFN-treated spheroids as well as in cellular organization and structures between cells, hinting at a possible involvement of extracellular matrix substances in the reaction to interferons.(ABSTRACT TRUNCATED AT 250 WORDS)

Poor induction of interleukin-2 receptor expression on CD8bright+ cells in whole blood cell cultures with CD3 mAb. Implications for immunotherapy with CD3 mAb.

To induce better stimulation of T cells during recombinant interleukin-2 (rIL-2) therapy of renal cell carcinoma patients, pretreatment with low-dose CD3 monoclonal antibody (mAb) has been proposed. However, in our clinic, such a treatment did not induce additional activation of T cells. To investigate this we performed whole blood cell cultures with rIL-2 or CD3 mAb as a stimulant. Cultures using isolated blood mononuclear cells were used as a control. When stimulated by the addition of rIL-2, the lymphocyte composition and activation of whole blood cultures did not differ from those of mononuclear cell (MNC) cultures. However, when stimulation was performed with CD3 mAb, CD8bright+ cells in whole blood cultures were not or only minimally induced to express CD25 or IL-2 receptor beta (IL-2R beta). This is in contrast to the situation found in MNC cultures where all CD8bright+ cells expressed CD25 or IL-2R beta to a high extent at the end of culture. When rIL-2 or recombinant interferon gamma (rIFN gamma) was added to whole blood cultures together with CD3 mAb, significantly more CD8bright+ cells were induced to express CD25 or IL-2R beta. These results suggest that whole blood cultures represent the in vivo situation better than MNC cultures. In addition, the results suggest that, also in vivo, administration of low-dose CD3 mAb alone might not be sufficient to induce IL-2R expression on CD8bright+ cells, and would therefore not induce additional specific T cell activation in rIL-2-based immunotherapy. The presented results suggest that in vivo simultaneous administration of rIFN gamma or rIL-2 with low-dose CD3 mAb might induce better stimulation of CD8+ T cells than CD3 mAb only.

In vitro immunomodulating effect of protein-bound polysaccharide, PSK on peripheral blood, regional nodes, and spleen lymphocytes in patients with gastric cancer.

PSK, a protein-bound polysaccharide, has been widely used for cancer immunotherapy in Japan. However, the mechanism of its immunomodulatory effect has not been fully clarified. In the present study the in vitro effect of PSK on the lymphocytes of patients with gastric cancer was studied. Culturing lymphocytes with PSK at 5-100 micrograms/ml increased the level of DNA synthesis, and augmented the cytotoxicities against K562 and KATO-3. Flow-cytometric analysis also showed an increase in the proportion of interleukin-2 (IL-2)-receptor-positive cells after the lymphocytes were cultured with PSK. However the cytotoxicity of cells cultured with PSK was not augmented by the addition of recombinant interferon gamma (rIFN gamma) and rIL-2. Further experiments using fractionated PSK showed that its biological action is present mainly in fractions having molecular masses greater than 10(5) Da. However, these immunomodulations were not seen in all patients. These results suggest that the susceptibility of lymphocytes to PSK may be different in each patients, and that the immunomodulation by PSK may be mediated by mechanisms independent of IFN and IL-2.

The effect of treatment with recombinant γ-interferon on adjuvant-induced arthritis in rats

We have investigated the effects of recombinant rat gamma-interferon (rIFN gamma) on adjuvant-induced arthritis (AA). Lewis rats, inoculated in the left hind-paw with adjuvant (day 0), were given 10(5) U/rat of rIFN gamma daily (days 0 to 20), subcutaneously and intramuscularly on alternate days. rIFN gamma suppressed the secondary phase of swelling of both hind-paw on and after day 18 without influencing the earlier phases, both primary and secondary, of swelling. rIFN gamma also reduced the hind-paw bone lesions, the degree of splenomegaly, and the increase in erythrocyte sedimentation rate and plasma fibrinogen. These results indicate a new aspect of the regulatory role of IFN gamma in chronic inflammation.

A randomised controlled trial of recombinant interferon‐γ in chinese patients with chronic hepatitis B virus infection

Nineteen Chinese patients with chronic hepatitis B virus (HBV) infection, seropositive for HBV e antigen (HBeAg) and HBV DNA on at least three occasions in 6 months, were randomised to receive either recombinant human interferon-gamma (rIFN gamma) 0.1 mg/m2 intramuscularly thrice weekly for 16 weeks (n = 11) or no anti-viral therapy (controls, n = 8). Five patients in the treatment group and four patients in the control group had persistently elevated serum alanine aminotransferases (ALT) of over two times the upper limit of normal before entering into the trial. rIFN gamma had no or minimal inhibitory effect on serum HBV DNA during treatment and no patient developed e-seroconversion or sustained loss of serum HBV DNA. Hepatitic flare, which occurred in a proportion of patients responding successfully to interferon-alpha (IFN alpha) therapy, was not observed with rIFN gamma treatment. Side-effects included pyrexia and mild headache that showed tachyphylaxis and were well tolerated by all patients. In the control group, one patient with elevated pre-entry serum ALT lost serum HBV DNA and seroconverted to anti-HBe. Another patient with elevated ALT lost serum HBV DNA transiently during therapy. In the dose given, rIFN gamma was safe but had no apparent anti-viral effects in Chinese patients with chronic HBV infection.

Effects of lobenzarit disodium on human endothelial cells. Lobenzarit disodium inhibits proliferative response, HLA–DR antigen expression, and T cell adherence toward endothelial cells

The therapeutic action of lobenzarit disodium (CCA) on the function of endothelial cells (EC) isolated from human umbilical cord veins was investigated. CCA suppressed 3H-thymidine incorporation into EC in a dose-dependent manner. Significant inhibition was detected at a concentration of 50 micrograms/ml. The expression of HLA-DR antigen on the surface of EC was increased when EC were cultured with recombinant interferon-gamma (rIFN gamma). Treatment of EC with either IFN gamma or interleukin-1 enhanced the adhesion of T cells to EC. The kinetics of HLA-DR antigen expression by EC cultured with IFN gamma was different from the kinetics of T cell-EC adhesion, however. Neither anti-HLA-DR nor anti-HLA-ABC monoclonal antibody inhibited T cell binding to EC monolayers. CCA suppressed the expression of HLA-DR antigen by EC cultured with rIFN gamma. In an EC monolayer adhesion assay, CCA also inhibited T cell adhesion to EC in the presence of either IFN gamma or interleukin-1. Significant inhibition was observed at a CCA concentration of 10 micrograms/ml, a level that is easily attainable in serum. These results suggest that CCA may suppress rheumatoid synovitis by reducing the angiogenesis and emigration of chronic inflammatory cells from the blood into the synovium.

Immunologic Changes in Renal Cell Carcinoma Patients Receiving Gamma Interferon

Immunologic changes were monitored in 5 patients with metastatic renal cell carcinoma participating in a phase II trial of recombinant interferon gamma (rIFN gamma). Treatment consisted of a 4-week period of intravenous infusion of either 0.1 mg/m2 given over 4 h on alternate days (3 times/week) or 2 mg/m2 given over 1 h for 5 days every other week. One minor response was seen at the higher dose level and toxicity was minimal (grades I-II). Absolute leukocyte counts and lymphocyte subpopulations did not change significantly. Both natural killer cytolytic activity and spontaneous monocyte-mediated tumoricidal activity increased. Prostaglandin E2 synthesis of patient leukocytes was abnormally high and pretreatment levels dropped during treatment. Data suggest that rIFN gamma may be potentially useful for enhancing immunologic function in renal cell carcinoma.

Effects of rIFN Alpha, Beta, and Gamma on the Morphology, Proliferation, and Cell Surface Antigen Expression of Human Dermal Microvascular Endothelial Cells In Vitro

The influence of recombinant human interferon alpha 2a (rIFN alpha), recombinant human interferon beta 1 (rIFN beta), and recombinant human interferon gamma (rIFN gamma) on human dermal microvascular endothelial cells (HDMEC) cultured in vitro was studied in various rIFN concentrations (0.1 IU/ml-10(4) IU/ml) over 2, 3, 4, 6, 8, and 10 d. Cell morphology and ultrastructure, cell proliferation, expression of class II alloantigens (HLA-DR and HLA-DQ), and intercellular adhesion molecule-1 (ICAM-1) were investigated using an in vitro technique established in our laboratory. All rIFN tested induced alterations of typical HDMEC morphology; the cells became spindle-shaped and fibroblastoid, although they maintained their endothelial cell marker expression. Also, all IFN dose- and time-dependently inhibited the proliferation of HDMEC in vitro (rIFN alpha greater than beta greater than gamma), whereby rIFN alpha exerted the strongest growth-inhibitory effect. Alkaline phosphatase anti-alkaline phosphatase (APAAP) immunocytochemistry of the cultured cells showed dose- and time-dependent stimulation of ICAM-1 and class II antigen expression only by rIFN gamma (HLA-DR greater than HLA-DQ), rIFN alpha and beta did not exert any immunomodulatory activity on HDMEC in vitro. These results indicate that HDMEC are an important target for the action of IFN. Besides growth inhibition, it seems that rIFN gamma in particular may be involved in the modulation of leucocyte adhesion and trafficking by altering the immunophenotype of the endothelial cell population.

A phase IA trial of sequential administration recombinant DNA-produced interferons: combination recombinant interferon gamma and recombinant interferon alfa in patients with metastatic renal cell carcinoma.

This study investigated the effects of sequentially administered recombinant interferon gamma (rIFN gamma) and recombinant interferon alfa (rIFN alpha) in 36 patients with metastatic renal cell carcinoma (RCC). rIFN alpha was subcutaneously administered daily for 70 days at dosages that varied (2.5, 5, 10, and 20 x 10(6) U/m2) across four cohorts of patients. Within each cohort of patients receiving a given dose of rIFN alpha, three subsets of patients received either 30, 300, or 1,000 micrograms/m2 rIFN gamma. rIFN gamma was administered intravenously for 5 days every third week, 6 hours prior to administration of rIFN alpha. Dose-limiting toxicity (DLT) included constitutional symptoms, leukopenia, nephrotic syndrome with acute renal failure, hypotension associated with death, and congestive heart failure. DLT was related more often to the rIFN alpha dose level than to rIFN gamma dose level. Maximum-tolerated dose (MTD) was 10 x 10(6) U/m2 rIFN alpha and 1,000 micrograms/m2 rIFN gamma. Six patients failed to complete a minimum of 21 days of therapy due to toxicity or rapid progression of disease. Clinical responses were seen in eight of 30 assessable patients. Two patients experienced complete remission and have remained in complete remission 20+ and 22+ months. An additional six patients have shown partial responses for 4 to 18+ months. One patient in partial remission continues to show slow regression of pulmonary and liver lesions off therapy with rIFNs. Clinical responses have remained durable for patients with complete remissions and patients with partial remissions. The results of this study suggest that toxicities associated with combination rIFN therapy can be reduced by administering these agents sequentially as opposed to simultaneously.