Annette E. Maluish

A phase I trial of recombinant gamma interferon in patients with cancer

SummaryA total of 11 patients were treated on an escalating, single dose trial of recombinant gamma interferon (rIFN-γ), 6 patients by the i.m. and 5 patients by the i.v. route of administration. Dose ranges within each individual were from 0.05 mg/m2 of IFN (1 mg≥10×106 units of IFN) escalating to 10 mg/m2. All dosages were delivered twice weekly and the i.v. dose was infused over 5 min. The most common toxicities encountered included fever, chils, fatigue, anorexia, and granulocytopenia. The influenzalike symptoms were very similar to those encountered with IFN-α but were generally less severe. The granulocytopenia was dose-related and transient with recovery generally seen within 48–72 h following administration of rIFN-γ. Absolute granulocyte counts only rarely dropped below 1000 mm3. Hepatotoxicity was not observed. IFN levels were determined by both a bioassay and an enzyme-linked immunosorbent assay. By the i.v. route, the peak level of IFN activity could usually be seen at completion of the infusion with a serum half-life of 30 min. By the i.m. route, the peak level of serum activity was generally detected between 4–8 h with a serum half-life of 4.5 h after the initial elimination phase. Peak IFN levels appeared to correlate with maximum toxicity. One patient with melanoma had a 25% reduction in a cutaneous lesion, but there were no other minimal, partial, or complete responses.

Quantitation of anti-tumor cell-mediated immunity by a lymphokine-Dependent reaction using small volumes of blood

A two-stage modification of the leucocyte adherence inhibition (LAI) test is described, which quantitates cell-mediated immunity (CMI) in vitro with 0.5-ml samples of blood from mice exposed to methylcholanthrene-induced tumors. The total blood cells are washed and incubated for 30 min with tumor antigen, then centrifuged, and the supernatant assayed in the LAI test with normal peritoneal cells. The first stage of the reaction depends on the rapid formation of a soluble mediator which appears to be a lymphokine. By this method, the daily changes in specific CMI were followed in individual mice after syngeneic tumor transplantation.

Blocking and unblocking of cell-mediated anti-tumor immunity in mice, as detected by the leucocyte adherence inhibition test

Further observations have been made using the in vitro leucocyte adherence inhibition (LAI) test previously described. The test has been slightly modified and shown to give highly reproducible results in detecting cell-mediated immunity in peritoneal cells of mice exposed to tumors, and in demonstrating factors which modify this immunity. Two different methylcholanthrene-induced tumors stimulated specific reactivity in their hosts as shown by the leucocyte adherence inhibition test employing soluble tumor extracts as antigens. When serum from tumor-bearing mice was added to the mixture, reactivity was specifically blocked. Serum from mice after tumor removal lacked the ability to block the leucocyte adherence inhibition reaction, but specifically reversed the effect of blocking serum (unblocking). Thus three important phenomena of tumor immunity, previously discovered by tedious and complex techniques, are now detectable by a brief, simple procedure.

INTERFERON AND NK CELLS

Abstract: Interferon (IFN) has been shown to play a major role in the positive regulation of the activity of natural killer (NK) cells. All human IFN species tested to date of each of the three types, including homogeneous material and recombinant species, have had the ability to augment human NK activity. The mechanism for augmentation of this effector cell activity has been shown to be complex, with the potential for increasing the numbers of cells recognizing NK-susceptible targets, activating the lytic activity of target-binding cells, accelerating the rate of cytolysis of targets, and increasing the numbers of target cells killed by each effector cell. In addition, IFN has been shown to influence the growth and differentiation of NK cells from their progenitors: on the one hand, increasing the responsiveness of NK cells to interleukin-2 and thereby increasing cloning efficiency, and on the other hand, inhibiting generation of NK cells via a T cell-dependent mechanism. Some preliminary evidence has been obtained for the involvement of the 2′-5′A pathway in the mechanism of the augmentation of NK activity by IFN, since triphosphorylated 2′-5′A, after exposure of the cells to calcium chloride, caused an increase in NK activity. Nk cells appear to not only respond to IFN but also to produce IFN, in response to a wide range of stimuli. Overnight incubation of a highly purified population of human NK cells with viruses (herpes simplex or influenza), mycoplasma-contaminated tumor cell lines, BCG or C. parvum produced IFN-2, whereas under the same conditions, various mitogens (PHA, Con A, Staph enterotoxin) induced mainly IFN-α, and some IFN-β. Thus NK cells appear to be a major component of both the IFN-producing system and of the host effector mechanisms modulated by IFN. These observations have important implications in regard to clinical therapeutic trials with IFN or IFN inducers.

MODULATION OF NK AND MONOCYTE ACTIVITY IN ADVANCED CANCER PATIENTS RECEIVING INTERFERON

ABSTRACT : 134 patients with a variety of malignancies were treated in two Phase 1 clinical trials of recombinant leukocyte A interferon (IFN) produced by recombinant DNA methodology by Hoffmann-La Roche Inc. in collaboration with Genentech Inc. IFN was given either twice daily or three times weekly for 28 days. Extensive monitoring of immune function was done on these patients, in an attempt to define the range of doses of interferon and timing of administration that would optimally alter immune functions which might have beneficial anti-tumor effects. These two protocols failed to result in an appreciable increase in NK activity but rather in the majority of patients there was no significant change in NK activity from the pretreatment baseline levels. A particularly unexpected finding was the depresssion in NK activity seen in 30% of the patients. In contrast, the IFN preparation did induce significant augmentation of monocyte function, as measured in a monocyte-mediated cytostatic assay, with increases observed in 80% of the patients. There is some evidence for the presence of a suppressor factor in the sera of these patients which interfered with in vitro boosting of NK activity by IFN and therefore might have been responsible for the paradoxical NK results. These data have also been analyzed in terms of their possible relationship to doses, and protocol of IFN administration, and to tumor response.

A Phase I Trial of Immune Interferon

A variety of interferons have now been tested in clinical trials. Most of these trials have utilized leukocyte (α) interferon preparations. Early trials were conducted with partially purified material derived from the supernatants of virus-stimulated leukocytes that were of low purity and inconsistent pharmaceutical quality. More recently, trials have been conducted with a lymphoblastoid cell line interferon (α) of high purity and good reproducibility (Knost et al., 1983). Several recent studies have utilized recombinant α-interferon derived by cloning a gene for α-interferon in an E. coli expression system (Sherwin et al., 1982, 1983). Despite the major differences between these α-interferon preparations, many of the toxicities, immunological modulating effects, and therapeutic effects have been similar. Fever, chills, headache, fatigue, and anorexia have been rather constant side effects of these interferon preparations. At higher doses, mild hematological depression and transient hepatic enzyme abnormalities have been seen. Occasional cardiac effects including arrhythmias and ischemic effects have been observed in the context of these trials. Some central nervous toxicity including confusion, decreased ability to concentrate, and rarely seizures at very high doses have been seen in these studies. It is unclear whether all these effects are due to the direct action of the interferon preparation since the induction of fever, tachycardia, and fatigue may have secondary effects (Oldham, 1983a). The phase I trials for the α-interferons are virtually complete.

Immunological reactions in medullary carcinoma of the thyroid

Three patients with medullary carcinoma of the thyroid (MCT) were tested for antitumor cell-mediated immunity and serum blocking factors by the leukocyte adherence inhibition (LAI) test. Leukocyte reactivity with both autochthonous and allogeneic MCT extracts was detected after surgical removal of tumor. Blocking factors were present in patients with progressing MCT but disappeared after tumor removal; in one patient the serum then became unblocking. The leukocytes of seven control subjects did not react in the LAI test with MCT extracts. The value of the LAI test in MCT is discussed as an adjunct to the serum calcitonin assay.