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Idiotypes are distinct clonal markers for B-cell lymphomas. Previously we reported the use of anti-idiotype antibodies in the therapy of patients with B-cell malignancies. Because synergy was demonstrated with the addition of alpha interferon to anti-idiotype antibodies in a murine lymphoma model, we performed a clinical trial combining these two agents. Here we provide an update of the original trial of anti- idiotype antibodies alone and report the outcome of the new combination trial. In 16 treatment courses of anti-idiotype antibodies alone there were seven partial responses and one complete response. In 12 courses of combination anti-idiotype antibody and alpha interferon there were two complete responses and seven partial responses. Substantial tumor regressions occurred with minimal toxicity in both trials even in patients refractory to conventional chemotherapy. Tumor specimens obtained at the time of disease progression often contained a preponderance of idiotype-negative lymphoma cells, suggesting that anti- idiotype antibody treatment exerted a strong antitumor effect against antigen-positive cells. Anti-idiotype antibodies have reproducible objective antitumor activity in B-cell lymphoma. The addition of alpha interferon may improve the initial rate of response to this treatment. Strategies that deal effectively with idiotype-negative lymphoma cells should improve the extent and duration of these responses.

Follow-up data for 11 patients with non-Hodgkin's lymphoma treated with partially purified human leukocyte interferon is presented. The interferon preparation used was 0.1% pure and treatment consisted of 5 x 10(6) U given intramuscularly twice daily for 60 injections. One complete, three partial, and three minimal responses were observed in five of seven evaluable patients with nodular non-Hodgkin's lymphoma. Duration of response appears to be from 6 to 12 mo. One patient achieved a second partial response on retreatment with interferon in spite of having received chemotherapy in the interval between interferon treatments. No responses were seen in three patients with rapidly progressive diffuse histiocytic lymphoma. Dose-limiting toxicity is leukopenia, which necessitated modification or cessation of treatment in three patients. Nonhematologic toxicities consisted of fever, malaise, arthralgia, and loss of appetite. In conclusion, interferon has activity against non-Hodgkin's lymphoma, and prior treatment with chemotherapy does not preclude a response to interferon.

Mouse interferon preparations significantly suppress the in vivo antibody response to sheep red blood cells (SRBC), a thymus-dependent antigen, and to Salmonella typhimurium lipopolysaccharide (LPS), a thymus-independent antigen. It is also possible to effect the late responses of antigen sensitive "memory" cells observed during secondary immunization by administration of interferon prior to primary immunization. The immunosuppressive activity of interferon was time- and dose-dependent. Maximum suppression was produced when animals were given 1.5 times 10-5 units of interferon between 4 and 48 hr before antigenic stimulation. These findings suggested that interferon affects some early event(s) in the process of antibody synthesis which might be related to the general inhibitory effect of interferon on rapidly dividing cells and viral m-RNA translation. In addition, the use of nonadherent spleen cell cultures from interferon-treated mice, immunized in vitro with a thymus-independent antigen, indicated that in this situation the inhibitory effect of interferon was due to an action on B lymphocytes. A variety of soluble "suppressive" factors are secreted by T cells as a consequence of activation by mitogens or specific antigens in vitro. Since T cells are recognized as one of the sources of interferon, it is suggested that interferon should be investigated as a suppressor T cell-produced lymphokine which can regulate B cell expression.

Two murine models, C3H 38C13 B-cell lymphoma and AKR SL2 T-cell lymphoma were used to determine the efficacy of three different interferon preparations, recombinant human hybrid interferon-alpha A/D, recombinant murine interferon (rMIFN)-gamma, and natural MIFN-alpha/beta (greater than or equal to 85% beta), alone and in combination with tumor specific and nonspecific monoclonal antibody therapy. All three interferon preparations have direct in vitro antigrowth activity for 38C13 and SL2. All three interferons have direct antitumor activity in vivo for 38C13 lymphoma at high doses; however, none of these interferons has independent antitumor activity for SL2 in vivo. These data indicate that there is no relationship between in vitro growth cytostasis/cytolysis and in vivo antitumor response. All three interferon preparations will potentiate both tumor specific and nonspecific monoclonal antibody therapy. Natural MIFN-alpha/beta and recombinant human hybrid interferon-alpha A/D, which should share a common cell surface receptor, had similar antitumor activity in both models. Combining recombinant human hybrid interferon-alpha A/D and rMIFN-gamma therapy was not additive for 38C13 lymphoma and a three-way combination with antiidiotype was not significantly more effective than combination therapy with one interferon type. In general, rMIFN-gamma was more effective in in vivo combination therapy against the s.c. T-cell lymphoma than against the i.p. B-cell lymphoma and was more synergistic with anti-Thy1.1 than with antiidiotype.

The effect of interferon on mouse and human in vitro erythropoiesis was investigated using the mouse marrow CFU-E and the human blood BFU-E systems, respectively. Homologous interferons were found to have a marked inhibitory effect on mouse CFU-E and human BFU-E proliferation. This inhibitory effect was dose related and independent of the erythropoietin concentration used. The effective concentration of human interferon was within the range observed in humans following viral infections. It is suggested that interferon may play a role in the mechanism of the acute erythroblastopenic crisis observed at times in patients with chronic anemia following viral infections.

Human peripheral blood lymphocytes, highly enriched for T cells, were obtained by passing gravity-sedimented leukocytes through nylon wool columns. The eluted cells were cultured with autologous macrophages and the mixture was studied for its capacity to produce interferon in vitro in response to stimulation with herpes simplex virus antigen. The interferon produced by the combined macrophage-lymphocyte cultures was shown to depend upon the presence of T cells; elimination of these cells by treatment with an anti-T cell serum plus complement greatly diminished the amount of interferon produced. The memory for the immune-specific release of interferon also appeared to be carried by the T lymphocytes rather than the glass-adherent macrophages. Furthermore, the results suggest that under our conditions of culture immune-specific interferon originates from T cells.

Several chemicals and interferons have been utilized in the prophylaxis and the treatment of a number of human herpesvirus infections. For example, adenine arabinoside as well as acyclovir and human leukocyte interferon appear active against acute varicella zoster and herpes simplex infections in controlled trials. Some of these agents have transient in vivo actions against CMV and EBV as well. Further open trial work with these 2 viruses appears necessary to improve dose regimens or to employ combination therapy or study new derivatives or agents before controlled trials are warranted. Recombinant DNA methodology is currently providing much greater yields of pure interferon for such uses. Recent successes in eradicating chronic hepatitis B virus infection suggest that it is even possible in the future that persistent herpesviruses in the CNS or lymphoreticular systems may be amenable to long-term therapy. As hyperimmune globulin appears to diminish the severity of CMV disease, it is also possible that human monoclonal antibodies may provide a useful approach for immunoprophylaxis as therapy.

At different times of exposure, interferon (IFN) enhanced and suppressed pokeweed mitogen- (PWM) induced IgG synthesis by human peripheral blood lymphocytes (PBL). Pretreatment of PBL and IFN frequently increased antibody production by more than 100% when compared with that by untreated PBL. Results of experiments in which PBL were separated into T and B subpopulations indicated that IFN preparations acted directly on B cells. Thus, mixtures of IFN-treated B cells and untreated T cells from 5 of 7 persons tested produced 81% to 500% more IgG than untreated, matched control cells. However, IFN-treated monocytes mixed with untreated B and T cells or IFN-treated T cells mixed with untreated B cells failed to enhance IgG production significantly in similar assays. In contrast to the pretreatment protocol, when IFN was present in the incubation mixture throughout the PWM assay, IgG production decreased. Sephadex chromatography of the IFN and tests of the resulting fractions indicated that the IgG production-enhancing activity was located in the fraction carrying the antiviral activity.

A factor with antiviral activity has been produced in vitro by combined macrophage-lymphocyte cultures from patients with recent herpes labialis in response to HSV antigen stimulation. It has been designated "immune interferon" and characterized in comparison to several other human interferons. It was shown to be relatively unstable at pH 2 and at 56 degrees C. Rabbit anti-human leukocyte interferon serum was shown to be less active against immune interferon than against diploid cell interferon or against vesicle fluid interferon. The possibility of immune interferon being a totally different anti-viral protein or a protein with certain shared antigen determinants or structures with classical viral interferon is discussed. A simplified method for the assay of anti-interferon sera with microtiter plates is also described.

Seventeen patients with disseminated cancer were treated with a human leukocyte interferon preparation in doses ranging from 3 X 10(6) to 50 X 10(6) IU daily for 30 days. Doses above 18 X 10(6) IU were considered intolerable in this schedule of administration due to severe fatigue and weight loss. Serum concentrations of interferon were lower than those achieved with either partially pure native or recombinant leukocyte interferon. Three of 17 patients in this study showed minimal evidence of tumor regression. Two patients treated at doses of 18 X 10(6), 36 X 10(6), and 50 X 10(6) IU also received a 5 X 10(6)-IU dose of a second human leukocyte interferon preparation. The latter resulted in less toxicity but similar serum levels. These results suggest that human leukocyte interferons prepared in the same manner may differ significantly in their in vivo biologic properties.

Our studies of in vitro cellular immune responses to herpesviruses have shown that, before treatment, lymphoma patients have decreased transformation to varicella-zoster virus antigen compared to herpes simplex virus and cytomegalovirus antigens. Interferon production to varicella-zoster virus and cytomegalovirus antigens is also decreased. During the first six months of treatment, decreased lymphocyte transformation and interferon production to all herpesviral antigens was observed. Among patients in long-term remission, recovery from the suppression associated with treatment was noted except that interferon production to varicella-zoster virus antigen remained decreased.

The clinical and immunologic effects of a biosynthetic human leukocyte interferon, recombinant leukocyte A interferon (IFL-rA), are reported in eight patients with advanced cancer. Single escalating doses from 3 X 10(6) units to 198 X 10(6) units were given by intramuscular injection in a phase I study. Major toxic effects included pyrexia, fatigue, myalgia, and headache. Data on the effects of IFL-rA on lymphocyte subpopulations and peripheral blood mononuclear-cell surface beta 2-microglobulin are presented. Four of eight patients had objective tumor regression, indicating that further investigation of this biologically active material is warranted.

Natural killing by mouse spleen cells can be stimulated in vivo by interferon or by agents that stimulate interferon, such as poly I.C. Natural killing can be suppressed in vivo by the sustained administration of 17 beta-estradiol. In BALB/c mice that had been treated with 17 beta-estradiol for 10 weeks, natural killing did not respond to intravenous poly I.C, although stimulation of circulating interferon was equal to controls. Estradiol, then, does not block interferon production but does suppress the response of natural killer cells to interferon. It is suggested that estrogens either block the maturation of natural killer cells or reduce the number of natural killer cell precursors.

Interferons are proteins elaborated by infected cells that protect noninfected cells from viral infection. These proteins produce a temporary "antiviral state" by altering nucleotide metabolism and cytoplasmic enzyme induction. Interferons appear early after viral infection locally and systematically to limit spread of viral infection; they also affect cell differentiation, growth, surface, antigen expression, morphologic findings, and immunoregulation. Several human disorders have diminished interferon production. Newborns have normal interferon alpha but deficient interferon gamma production. Infants with congenital infections may also have defects in interferon production. Immunosuppressed patients receiving transplants (marrow, heart, of kidney) have diminished interferon production, particularly immediately after transplant. Deficiencies of interferon have also been noted in Down's syndrome, cellular immunodeficiencies, uremia, malnutrition, and hematopoietic malignancy. Leukocyte interferon has been of therapeutic value in herpes zoster infections, in patients with cancer, and in patients with hepatitis B infection. Interferon has not been proved to help children with congenital cytomegalovirus or rubella. Interferon can shrink lymphoid tumors, particularly non-Hodgkin's lymphoma.

Understandably, there is interest in further defining the nature of 'non-specific' defenses. This, in fact, would clarify the specific nature of their appearance and action, as well as their role in host defense. Therefore, 'non-specific' defenses in my discussion will be taken to mean non-immune defenses in virus infections. Tissue or respiratory tract organ cultures are convenient means of studying host defenses in the absence of immune responses. Factors to be controlled in making observations with such systems will be discussed which help 'in vitro' results to parallel 'in vivo' findings. 'In vitro' and 'in vivo' methods and results probing importance of mucous barrier, ciliary activity, temperature and pH will be reviewed. Critical points in separating cellular and humoral immune functions from non-specific mediators will also be touched on. This distinction is complicated by the fact that both limbs of immunity work in close concert with macrophages, PMN's complement and other inflammatory responses. Interferon and macrophage mobilization are examples of 'non-specific' responses which have more recently become complicated by their recognition as part of the effector limb of the cell-mediated immune responses. Finally, methods will also be presented for distinguishing human interferon of the immune-specific variety associated with cell-mediated immunity from that produced by viral infection of non-immune cells.

Antigen-specific type II interferon was produced in vitro by harvesting supernatants of spleen cell cultures from Swiss-Webster mice sensitized with Mycobacterium bovis strain BCG and challenged with old tuberculin. Treatment of C3H mouse spleen cell cultures with appropriate anti-Ia, anti-IgG, anti-Thy-1 or anti-Ly-2,3 sera resulted in a significant decrease in production of type II interferon. Removal of nylon wool adherent cells or cells with histamine receptors by column chromatography similarly caused reduced production of type II interferon. Recombination of spleen cell cultures treated with anti-Ia and anti-Thy-1 sera or of cells treated with anti-IgG and anti-Thy-1 resulted in restored production of type II interferon. Interferon production was also restored by combination of cells passed through histamine columns with anti-Ia treated cells, or those passed through nylon wool columns with anti-Thy-1 treated cells. Anti-Ly-1 serum treatment had no effect on interferon production. Removal of plastic-adherent cells or cells that had phagocytosed carbonyl iron also decreased interferon production, suggesting that macrophages were also involved in type II interferon production. Recombination of non-adherent spleen cells with anti-Ia and anti-Thy-1 sera treated spleen cells, however, did not restore interferon production, suggesting that other cells in addition to macrophages are depleted by the adherence procedure. These findings indicate that type II interferon is produced by suppressor or cytotoxic (Ly-2,3+) T lymphocytes in co-operation with one or two additional cell types: (i) B lymphocytes, and (ii) macrophages.

Ten young adult patients with chronic hepatitis B virus infection and positive hepatitis B e antigen and DNA polymerase (DNAP) levels were treated with alternating courses of seven to 28 days of 5 to 7.5 mg/kg of vidarabine monophosphate (adenine arabinoside monophosphate) and 28 days of human leukocyte interferon (IFN-alpha); three different regimens were given on an outpatient basis. All patients with a fall in their DNAP level, and the DNAP remained undetectable six months after treatment was stopped in one patient. The major side effect, which most often occurred in those patients receiving 7.5 mg/kg of vidarabine monophosphate, was severe muscular pains. This study demonstrated the feasibility of administering vidarabine monophosphate and interferon to outpatients. Based on data from this and other studies, it is now possible to use a relatively nontoxic regimen that includes 28 days of 5 mg/kg of vidarabine monophosphate in a larger controlled study to answer the question of efficacy.

SummaryIncreasing the in vitro age of chick cell monolayers was found to alter the dose-response to interferon so that aged cells responded over a narrower concentration range than young cells. This effect was not observed on aged L-cells. No consistent effect on the PDD50 of chick and mouse interferons was observed under the conditions of aging used in these experiments.The percent of plaques appearing on interferon-treated chick cell monolayers was found to be independent of the amount of vesicular stomatitis virus used for challenge over an eightfold range. The percent of plaques appearing on interferon-treated L-cells, however, showed a progressive decrease with increasing amounts of challenge virus.I am grateful to Dr. Walter D. Foster for helpful criticism of the manuscript and to Tony Verga for technical assistance.