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Histological and ultrastructural changes were investigated in lung, liver, and heart of mice given interleukin-2 (IL-2), either alone or in combination with other cytokines. IL-2 induced a vascular leak syndrome (VLS) of a moderate degree with infiltration of lymphoid cells, moderate endothelial damage, mild hepatic parenchymal damage, and minimal myocardial alterations. Interferon-alpha (IFN-alpha) produced infiltration mainly of monocytes/macrophages in liver and heart; endothelial cell damage was absent in lung and heart and minimal in liver. Interleukin-1 alpha (IL-1 alpha) caused an increased number of neutrophils in liver and lung; VLS and parenchymal cell and endothelial damage were not found. The VLS and the cellular damage caused by the combination of IL-2 and IFN were much more severe than those produced by IL-2 alone. In animals treated with IL-2, IFN-alpha, and IL-1 alpha, VLS was minimal and parenchymal and endothelial cell damage were less severe than after IL-2 alone or IL-2 plus IFN-alpha. Taken together, these observations show that IL-1 alpha reduces ultrastructural changes produced by IL-2 and IFN-alpha. This reduction may be clinically useful in the treatment of neoplasms.

A double-institution phase II study was performed in patients with metastatic renal cell carcinoma treated subcutaneously (s.c.) with interleukin 2 (IL-2) and alpha-interferon (INF-alpha). Thirty-eight patients were treated over a course of 7 weeks. Initially (day 1 + 2) patients received s.c. IL-2 at 18 x 10(6) IU m-2. During the following 6 weeks, patients received s.c. IL-2 at 3.6 x 10(6) IU m-2 for 5 days per week and s.c. INF-alpha at 5 x 10(6) for 3 days per week. Thirty-eight patients were evaluated for response. An objective response was seen in seven patients (18.4 +/- 12.3%), with one complete response and six partial responses. Median duration of response was 6.7 months. Toxicity could be evaluated in 38 patients and was limited. Mild to moderate toxicity included fever (97%), fatigue or malaise (76%), nausea or vomiting (50%), anorexia (32%), hypotension (26%), neurological disturbances (26%) and hypercreatininaemia (39%). In addition, four grade IV haematological toxicities were noted. No cardiac side-effects were seen. IL-2 and INF-alpha given by this schedule can be safely administered in an outpatient setting. The objective response rate was similar to our previous treatments with high-dose IL-2 given as a continuous infusion.

Interleukin (IL)-2 is known to induce vascular leak syndrome (VLS), which was suggested to be mediated by immune system-derived cytokines, including tumor necrosis factor (TNF). To characterize the role of TNF in IL-2 toxicity in C3H/HeN mice, we used two approaches to downregulate TNF production in vivo: treatment with dexamethasone (DEX) and induction of endotoxin (lipopolysaccharide) (LPS) tolerance by a 4-day pretreatment with LPS (35 micrograms/mouse/day). Mice were then treated with IL-2 for 5 days (1.8 x 10(5) IU/mouse, twice daily). Both DEX and LPS tolerance blocked development of hydrothorax in IL-2-treated mice and inhibited TNF induction. DEX and LPS tolerance also ameliorated IL-2 toxicity in terms of decrease in food intake and inhibited the increase of the acute-phase protein, serum amyloid A (SAA). The IL-2 activation of splenic natural killer (NK) cell activity was also diminished by DEX and, to a lesser extent, by LPS-tolerance. Treatment with IL-2 also caused induction of the superoxide-generating enzyme xanthine oxidase (XO) in tissues and serum and induced bacterial translocation in the mesenteric lymph nodes (MLN). These data suggest that multiple mechanisms, including NK cell activity, cytokines, and reactive oxygen intermediates, might be important in the vascular toxicity of IL-2.

The efficacy of ten daily injections of 500 or 500,000 U of recombinant interleukin 2 (IL-2) day-1 given 1.5 cm from the insertion of the sternocleidomastoid muscle on the mastoid was evaluated in 31 patients with recurrent head and neck squamous cell carcinoma. No toxic effects were noted. One complete response (CR) and three partial responses (PRs) were observed in the 16 patients who received 500 U of IL-2, whereas the higher dose was not effective. The CR was recorded in one of the seven patients with a oropharyngeal recurrence. Partial responses were obtained in 1/5 patients with hypopharyngeal recurrences, in 1/5 patients with oral cavity recurrences and 1/7 patients with laryngeal recurrences. The duration of the responses was 3-5 months and additional courses of ten injections of IL-2 had no further effect.

In a pilot study six patients received 4 days' treatment with interleukin 2 (IL-2) [cumulative dose (CD) 264 +/- 26 x 10(6) IU m-2] and C1 esterase inhibitor (C1-INH) (loading dose 2,000 U, followed by 500-1,000 U twice daily). Toxicity was compared with that in patients given 4 days' treatment with standard (CD 66 +/- 12 x 10(6) IU m-2) or escalating-dose (CD 99 +/- 8 x 10(6) IU m-2) IL-2. IL-2-induced hypotension was equivalent and complement activation was less after IL-2 + C1-INH (C3a = 10.5 +/- 3.2 nmol l-1) than following standard (14.1 +/- 8.4 nmol l-1) or escalating-dose (18.3 +/- 2.9 nmol l-1) IL-2. This study demonstrates that C1-INH administration during IL-2 treatment is safe and warrants further study to evaluate its ability to ameliorate IL-2-induced toxicity.

Recombinant human interleukin-2 (IL-2) was administered by the intravenous (i.v.) or intralymphatic (i.l.) route to 14 patients with advanced malignancy. IL-2 was given in doses of 600,000 IU/kg or 1,050,000 IU/kg daily x 5. Thoracic duct (TD) catheters were placed, and both TD lymphocytes (TDL) and peripheral blood lymphocytes (PBL) were studied. Five of eight patients at the 600,000 IU/kg dose experienced grade III toxicity as did five of six patients at the 1,050,000 IU/kg dose. Two episodes of grade IV toxicity were seen at the higher dose. The i.l. and i.v. routes had a similar toxicity profile excepting lymphangitis/pedal infection, seen only with i.l. administration. One partial response was seen in a patient with renal cell carcinoma. Lymphopenia was seen early in therapy, with lymphocytosis by day 6. Lymphoid yield of the TD catheter fell early in therapy, then increased over baseline by the end of treatment. Intralymphatic administration resulted in a prolonged serum t1/2 and lower serum levels than did i.v. administration, but resulted in higher TD levels. Antibodies against IL-2 were ubiquitous but had no clear effects. Lymphocyte trafficking studies suggested that IL-2 affected lymphocyte redistribution to liver, spleen, bone marrow, and lymph nodes. NK activity and phenotype and LAK activity increased in response to IL-2, with no advantage for TDL. Tumor necrosis factor-alpha and gamma-interferon levels increased sporadically with treatment. The i.l. route offered no advantage over the i.v. route, and TDL offered no advantage over PBL.

Thirty-six patients with unresectable squamous cell carcinoma of the head and neck were entered into a phase Ib trial evaluating the toxicity, maximally tolerated dose (MTD), and immunomodulating effects of locally administered interleukin-2 (IL-2). Patients received daily IL-2 injected perilesionally in divided doses in each of four quadrants and bilaterally into the superior jugular lymph nodes. The dose of IL-2 began at 200 U/day and was escalated to 4 x 10(6) U/day in groups of six patients. Overall, regionally administered IL-2 was well tolerated. The most frequently encountered toxicities were fever, hepatotoxicity, and hypotension. Dose-limiting toxicity was encountered at 4 x 10(6) U. Of the 36 patients treated, 2 partial responses were noted at 2,000 and 4 x 10(6) U. We conclude that regionally administered IL-2 is well tolerated in patients with head and neck cancer and that the MTD is 2 x 10(6) U/day, similar to what has been reported with systemically administered IL-2. Although the overall response rate was low, it may be improved with prolonged administration of IL-2 or by combining it with other biologic or cytotoxic agents.

The clinical application of interleukin-2 (IL-2) for the treatment of certain human malignancies has shown promise. However, the use of IL-2 in immunotherapeutic protocols has been limited due to its associated toxicities. The administration of therapeutic doses of IL-2 results in a vascular leak syndrome with associated multiple system organ edema, hypotension, and respiratory, renal, and hepatic dysfunction. Previous studies suggest that the mechanism of these toxicities involves the activation of both immune effector cells and the microvascular endothelium with resultant leukocyte-vessel wall interaction, endothelial cell injury, and subsequent invasion of normal tissues by activated leukocytes. Recently it has been demonstrated that interleukin-8 (IL-8) will inhibit leukocyte adherence to an activated endothelium. Thus, we hypothesized that IL-8 would ameliorate IL-2-evoked detrimental effects. We also investigated the influence of IL-8 on IL-2-induced antitumor efficacy. Four groups of nontumored, female, C57BL/6 mice and four groups of C57BL/6 mice with pulmonary metastases from a 3-methylcholanthrene-induced fibrosarcoma (MCA-105) were treated every 6 hr for 4 days by intraperitoneal injections of IL-2 alone, IL-2 and IL-8, IL-8 alone, or an equal volume of saline which served as our control. Upon completion of therapy, we found that IL-8 suppressed many of the IL-2-induced effects including multiple organ edema, hepatic dysfunction, leukopenia, and lymphocytic infiltration of normal organs. When the number of pulmonary metastases were counted 20 days after the cessation of therapy. IL-8 was also found to significantly ablate the IL-2-elicited antitumor efficacy.(ABSTRACT TRUNCATED AT 250 WORDS)

Conjugates of IL-2 with the ribosome-inactivating protein gelonin were prepared using heterobifunctional reagents to link the proteins via disulfide, acid-labile, and noncleavable linkers. In each case, one protein was modified using 2-iminothiolane. The sulfhydryl groups so introduced were then reacted either with 2-nitro-5-dithiobenzoate groups or with iodoacetamido groups which had been introduced into the second protein. In the case of the acid-labile linkage, a reagent which forms a labile bond upon reaction with amino groups, 4-(iodoacetamido)-1-cyclohexene-1,2-dicarboxylic acid anhydride (its synthesis is described in this paper) was used to modify the toxin. The conjugates were separated from nonconjugated proteins by gel filtration on Sephadex G100 (SF). Each was analyzed with respect to its ribosome-inactivating activity, its ability to bind to the IL-2 receptor, and its in vitro cytotoxicity. The ribosome-inactivating activity of gelonin was unaffected by modification with 2-iminothiolane and was retained in conjugates prepared using this reagent. Modification of the toxin with 4-(iodoacetamido)-1-cyclohexene-1,2-dicarboxylic acid anhydride to form the acid-labile link drastically reduced the activity of the toxin. However, the activity of the toxin was recovered following acid treatment to release the native protein. Conjugates containing each type of linkage exhibited both specific binding and selective cytotoxicity toward cells expressing the IL-2 receptor. The most potent of these toxins, that containing the disulfide linkage, exhibited a cytotoxicity which was 2 orders of magnitude greater than that of unconjugated gelonin.

Hematopoietic recovery in 115 patients with metastatic breast cancer or metastatic melanoma, enrolled in phase-I studies of recombinant growth factors while undergoing treatment with high-dose chemotherapy with autologous bone marrow support, was examined with assays of bone marrow progenitor cells and peripheral blood progenitor cells, and by evaluation of peripheral blood counts. Groups of patients receiving hematopoietic cytokine support [with interleukin-1 (IL-1), interleukin-2 (IL-2), granulocyte colony-stimulating factor (G-CSF), granulocyte-macrophage CSF (GM-CSF), or monocyte CSF (M-CSF)] post marrow infusion were compared with contemporaneous control patients not receiving growth factor support. Patients receiving GM-CSF demonstrated statistically significant increases in the growth of granulocyte/macrophage colony-forming units (CFU-GM) in the bone marrow and peripheral blood compared with control patients. The effect of GM-CSF was dose dependent in the early period post marrow infusion (day +6) with bone marrow CFU-GM colonies at doses 8-16 micrograms/kg/day 34 times those measured in controls. Significant increases in bone marrow multipotential progenitor cells (CFU-GEMM) were seen in patients receiving GM-CSF day +21 post marrow infusion. Patients receiving IL-1 demonstrated significant increases in bone marrow CFU-GM at day +21, maximal at dosages of 24-32 ng/kg/day. There were no significant increases in burst forming unit-erythroid (BFU-E) among any study group. Patients receiving G-CSF had significantly increased absolute neutrophil counts (ANC) and total white blood cell counts (WBC) by day +11 post transplant compared with control patients. Patients receiving GM-CSF demonstrated significantly increased WBC (greater than 2000/mm3) at day +11 and ANC greater than 500/mm3 at day +16. Optimal dose of G-CSF and GM-CSF to stimulate neutrophil recovery post transplant was 4-8 micrograms/kg/day and 8-16 micrograms/kg/day, respectively. Platelet recovery did not differ among the six study groups. These data demonstrate accelerated myeloid recovery after high-dose chemotherapy and autologous bone marrow support in patients receiving either G-CSF or GM-CSF. Moreover, GM-CSF and IL-1 stimulate myelopoiesis at the level of bone marrow CFU-GM, while G-CSF causes earlier neutrophil recovery peripherally.

DNA fusions encoding chimeric proteins in which human interleukin 2 (IL2) was fused to the A subunit of the plant cytotoxin ricin (RA) have been expressed in Xenopus oocytes. The constructs contained N-terminal IL2 and C-terminal RA, or N-terminal RA and C-terminal IL2. In the expressed chimeric proteins, the IL2 and RA moieties were joined by a peptide sequence containing a proteolytic cleavage site. Two proteolytically-sensitive peptide sequences were utilized; a peptide that forms the trypsin-sensitive disulfide-bonded loop in diphtheria toxin (DT) or a synthetic peptide containing the factor Xa recognition site in a sequence flanked by two cysteine residues. In an in vitro cell free system the RA component was biologically active in all chimeric proteins produced since it specifically depurinated 28S ribosomal RNA. Proteolytic cleavage of the chimeras with either trypsin or factor Xa as appropriate separated the IL2 and RA moieties, but they did not remain covalently linked by a disulfide bond. Because of this, the cytotoxicity of protease-treated chimeras could not be assessed. Chimeras not pretreated with factor Xa but which contained the factor Xa target sequence were not cytotoxic to CTLL-2 cells. Rather, these molecules had a stimulatory effect that was ascribed to the IL2 moiety. In contrast, recombinant chimeric toxins containing the DT loop sequence were cytotoxic to CTLL-2 cells. Taken together the data suggest that RA-containing chimeras require intracellular proteolytic cleavage to release the RA moiety to render them cytotoxic to target cells.

The primary, known physiologic effect of interleukin-2 (IL-2) is to act as a T lymphocyte growth factor. We investigated the potential contribution of IL-2 to intraocular inflammation by studying the inflammation resulting from the intravitreal injection of recombinant, human IL-2 in New Zealand white rabbits. Serial slit lamp observations indicated that 40 microgram of intravitreally injected IL-2 induced an anterior uveitis which was maximal 5 days after the injection. Inflammation was less marked but still significant with amounts of IL-2 as low as 400 ng. Direct examination of aqueous humor confirmed elevations of protein, prostaglandin E2, and mononuclear cells which correlated with the clinical observations. The kinetics of the response to intravitreal IL-2 distinguished it from the responses to other intravitreally injected cytokines such as interleukins 1, 6, or 8 as well as tumor necrosis factor. Intramuscular injection of cyclosporine A significantly reduced the protein extravasation associated with IL-2 injection, but cyclosporine had no effect on inflammation secondary to an intravitreal injection of interleukin-1. These observations implicate IL-2 as a potential contributor to uveitis. In addition, the studies with cyclosporine indicate the heterogeneity of inflammation such that pharmacologic agents which affect one cause of uveitis are not necessarily efficacious in another model.

Background: One of the side effects of interleukin 2 (IL-2) cancer immunotherapy in humans is the vascular leak syndrome, which is frequently associated with depression of myocardial function, myocarditis, and myocardial necrosis., Methods and Results: To investigate this cardiotoxicity, IL-2 (three doses of 5 x 10(5) Cetus units/day i.p.) was given to rats for 2, 3, or 5 days. Heart, lung, liver, spleen, and kidney tissues were studied by light and electron microscopy and with immunoperoxidase techniques. Cardiac changes consisted of focal lymphocytic and eosinophilic infiltration, myocyte vacuolization, myofibrillar loss, and necrosis. Ultrastructural alterations included swelling of endothelial cells, with dissociation of intercellular junctions, migration of lymphocytes into the interstitium, and interstitial hemorrhage and edema. Close contact between infiltrating lymphocytes, particularly large granular lymphocytes, and cardiac myocytes was often observed in areas of tissue damage. All lesions were more severe on day 5 than on days 2 and 3. Immunoperoxidase stains demonstrated asialo GM1 ganglioside antibody-positive, granular lymphocytes to be much more frequent in myocardium of IL-2-treated rats than in that of control rats., Conclusions: Although we cannot exclude the possibility of a direct toxic effect of IL-2 on myocytes, our observations suggest that the myocardial damage produced by this agent is triggered by IL-2-activated lymphocytes that exert cytolytic effects, first on endothelial cells and then on cardiac myocytes, thus producing lesions that involve both the cardiac microcirculation and the muscle cells.

It should be pointed out that although repeated systemic administration of exogenous rIL-2 in nonphysiological doses to mice, rats, and monkeys produced various toxicity-related exaggerated pharmacodynamic effects in many experimental parameters, pathological findings were substantially similar among all species. Furthermore, because toxicities observed in our studies were identical to those reported in humans dosed with rIL-2, preclinical toxicity using animal models was fully predictive of the pathogenesis of rIL-2 in human. With continuous infusion, which maintained high blood concentrations of the product during the course of the study, rIL-2 constantly stimulated proliferation of lymphoblastoid cells in peripheral blood, subsequently causing vascular leakage, which consists of edema and extravasation of activated lymphoblastoid cells into perivascular areas. The migrating lymphoblastoid cells, identified as cytotoxic T lymphocytes and NK-LAK cells, caused perivascular and hepatocytic necrosis by means of enzymatic secretions from the cells in association with infiltration into parenchymal tissues. In addition to the stimulation of lymphoblastoid cells in peripheral blood, prolonged daily bolus administration of rIL-2 induced extravasation and infiltration of activated macrophages and B cells in tissues of all organs. Eosinophilia, fibrosis in the subcutis, or pseudolobular formations in the liver could be mediated by systemic release of many other cytokines from activated T cells, i.e., CTLs or LAK cells, B cells, and macrophages.

We examined the direct effects of human recombinant interleukin 2 (hrIL-2) in vitro on the phenotypic and the functional changes of rat Kupffer cells (KCs) isolated by collagenase perfusion and differential centrifugation. When KCs were cultured with high concentrations of hrIL-2, marked changes were dose-dependently observed in phenotypic expression, phagocytic activity, and accessory cell function compared to untreated KCs. In the phenotypic changes, approximately 70% of KCs expressed class 2 antigen after 48-hr culture. Moreover, rapid expression of iC3b receptor on KCs was observed after 6 hr culture. KC phagocytic activity of both 0.81- and 2-microns latex particles was increased by incubating with hrIL-2 (10(4) U/ml) as compared to those of untreated KCs. Complement-mediated phagocytosis was more dominant than that of Fc receptor-mediated phagocytosis without hrIL-2 and augmented by increasing concentrations of hrIL-2. Accessory cell function of KCs was greatly augmented by preincubation with hrIL-2 (10(4) U/ml) for 24 hr. In the presence of 10(-6) M of indomethacin or prostaglandin E2, accessory cell function was at a lower level than without. This suppressive effect of indomethacin or prostaglandin E2 was reduced at lower concentrations. These results suggest that the potential ability of hrIL-2-activated KCs is closely associated with the ongoing pathophysiological process in the liver during IL-2 immunotherapy.

Previous studies on continuous hepatic artery infusions of recombinant interleukin-2 (IL-2) have shown that in a nontumor-bearing animal a continuous infusion given in a circadian "day cycled" pattern was much less toxic and could be given with 10 times higher doses of IL-2 than if the constant pattern of infusion was used. In the present study, circadian-patterned continuous hepatic artery infusions of IL-2 were used in hepatoma-bearing rats. Doses of 10 mg/m2/day could be tolerated when IL-2 was given in a "day cycle" rhythm. Control animals were given 1 mg/m2/day of constant infusion IL-2, which was the highest hepatic artery infusion dose tolerated at a constant rate without mortality in nontumor-bearing animals. Animals treated with the constant infusions of IL-2 had a 37.5% mortality rate and a 25% objective response rate in measurable tumor size. Animals receiving the "day cycle" had no mortality and a 100% objective response rate. The conclusion was that "day cycled" circadian-patterned continuous hepatic artery infusions of IL-2 could be given with much lower toxicity and much improved tumor response rates than constant continuous infusions.

Interleukin 2 (IL-2) mediates the regression of metastatic cancer but clinical use has been limited due to associated toxicities. Tumor necrosis factor (TNF) is an important mediator of IL-2 toxicity and may have a limited role in IL-2 antitumor efficacy. Because pentoxifylline (PTXF) inhibits TNF production, we hypothesized that PTXF would ameliorate IL-2 toxicity without compromising antitumor efficacy. Four groups of female C57BL/6 mice with pulmonary metastases from a 3-methylcholanthrene-induced fibrosarcoma (MCA-105) and four groups of nontumored mice were treated every 6 h for 4 d by intraperitoneal injections of either IL-2 alone, IL-2 and PTXF, PTXF alone, or equal volumes of saline. Upon completion of therapy, we found that PTXF suppressed many of the IL-2-induced effects including TNF production, lymphocytic infiltration of multiple organs, multiple organ edema, hepatic dysfunction, leukopenia, and thrombocytopenia. Tumor response was determined 21 d after cessation of therapy by quantitating the number and surface area of pulmonary metastases. PTXF preserved antitumor efficacy while reducing the morbidity and mortality caused by IL-2 treatment. These data strongly support the use of PTXF in extending the therapeutic index of IL-2 in the treatment of cancer.

We have performed a phase IB study of polyinosinic-polycytidylic acid complexed with poly-L-lysine and carboxymethylcellulose (poly-ICLC) in combination with interleukin 2 (IL-2) in 25 patients with a variety of cancers. Patients received weekly or biweekly poly-ICLC by i.m. injection, at doses ranging from 0.01 to 1.0 mg/m2, for 1 month. This was followed by 2 months of outpatient therapy with biweekly i.m. poly-ICLC in combination with IL-2 (3 x 10(6) units/m2) given i.v. by 24-h continuous infusion twice weekly, using a portable infusion pump. No objective tumor responses were observed. Toxicity was moderate at all poly-ICLC doses tested and increased only slightly following the addition of IL-2. No increases in peripheral blood natural killer (NK) activity were observed after treatment with poly-ICLC alone. However, high dose poly-ICLC (greater than or equal to 0.3 mg/m2) in combination with IL-2 resulted in NK activity greater than that seen using the same dose of IL-2 in combination with lower poly-ICLC doses. Increases in the number and percentage of CD56+ cells were evident only after initiation of IL-2 therapy and were unaffected by the poly-ICLC dose. In the majority of patients, these increases were preferentially associated with the subset of CD56+ cells coexpressing CD8, while the CD56+/CD16+ population was elevated to a lesser extent. Moderate increases in serum neopterin levels and 2',5'-oligoadenylate synthetase activity in peripheral blood mononuclear cells were noted at 72 h following initial treatment with 1.0 mg/m2 poly-ICLC. No induction of alpha or gamma interferon was detected. This study shows that the addition of poly-ICLC to a well tolerated IL-2 regimen can significantly enhance NK activity. Poly-ICLC can be used to enhance IL-2-induced NK lytic activity without increases in the dose and, therefore, the toxicity of IL-2 treatment.

DAB486IL-2 is a recombinant fusion toxin in which the native receptor binding domain of diphtheria toxin has been replaced with human interleukin-2 (IL-2). It selectively binds and intoxicates only cells that bear the high-affinity receptor for IL-2. In the first clinical trial of a genetically engineered ligand fusion-toxin, we have treated 18 patients with chemotherapy-resistant IL-2 receptor expressing hematologic malignancies with escalating doses of DAB486IL-2. The maximal tolerated dose of a daily intravenous bolus of DAB486IL-2 was 0.1 mg/kg per day for 10 doses, established by asymptomatic, reversible elevations of hepatic transaminases without changes in other tests of liver function. Other mild reversible side effects noted were rash, nausea, elevated creatinine, chest tightness, and fever. Pharmacokinetic analysis showed a monophasic clearance of 5.8 +/- 0.7 minutes with peak levels of 3,549 +/- 1,041 mg/mL at the 0.1 mg/kg dose. Approximately 50% of patients developed an antibody response to diphtheria toxin or DAB486IL-2. The presence of such antibodies did not preclude patients from experiencing an antitumor response as four of the six patients with antitumor effect had detectable antibody titers. Although this was a phase I trial designed to define the safety of DAB486IL-2, remissions were observed in three patients lasting from 5 to over 18 months. The ability to achieve significant tumor reductions in this group of heavily treated patients is encouraging and suggests additional trials are warranted in hematologic malignancies.

Interleukin-2 was recently shown to cause acute lung injury characterized by microvascular permeability defect, interstitial edema, and leukosequestration. Similar responses can also be produced by platelet activating factor (PAF). Thus, the present study aimed to examine whether PAF plays a key role in the development of IL-2-induced lung injury in the anesthetized rat. Intravenous infusion (60 min) of recombinant human IL-2 at 10(5)-10(6) U/rat (n = 7-9) dose-dependently elevated lung water content (27 +/- 1%, P less than 0.01), myeloperoxidase activity (+84 +/- 23%, P less than 0.05), and serum thromboxane B2 (990 +/- 70%, P less than 0.01), but failed to alter blood pressure, hematocrit, serum tumor necrosis factor-alpha, and circulating leukocytes and platelets. Pretreatment (-30 min) with a potent and specific PAF antagonist, BN 50739 (10 mg/kg, intraperitoneally, n = 6) prevented the pulmonary edema (P less than 0.05) and thromboxane B2 production (P less than 0.01), and attenuated the elevation of lung myeloperoxidase activity (+18 +/- 16%, P less than 0.05) induced by IL-2. These data suggest that PAF is involved in the pathophysiological processes leading to IL-2-induced lung injury, and point to the potential therapeutic capacity of PAF antagonists in preventing pulmonary edema during IL-2 therapy.

A preclinical pilot study was done to evaluate the effects of a continuous regional hepatic arterial infusion of human recombinant interleukin-2 (IL-2) in dogs with an infusion pump. Preliminary studies demonstrated the ability to culture canine lymphokine-activated killer (LAK) cells in vitro and a canine LAK cell 15Cr assay was developed with a canine tumor cell line (CTAC) with appropriate controls. An in vitro study of the stability of IL-2 in the pump was done with a bioassay and enzyme-linked immunosorbent assay for IL-2 that demonstrated the stability of IL-2 during a 14-day period at 37 degrees C. Infusions of 300, 600, and 1200 units/kg/hr IL-2 were tested in vivo in dogs. LAK cell and natural killer cell activity, blood counts, and hepatic and renal function were monitored for 1 month. No significant natural killer or LAK response or toxicity was found at the 300 unit/kg/hr level. Infusion of 600 units/kg/hr was associated with a significant increase of the cytotoxic activity of peripheral blood lymphocytes after 3 weeks of treatment. At the 1200 unit/kg/hr level, increased activity occurred at 1 week and thereafter. The only significant toxicity was a 15% increase in body weight occurring during the infusion of 1200 units/kg/hr. Results of renal and hepatic function studies remained normal except for a slight elevation of transaminase levels after 4 weeks of 1200 units/kg/hr. A significant rise in eosinophil count was noted at each dosage level. Results of autopsies were unremarkable. These data demonstrate that continuous hepatic arterial regional infusion with relatively low doses of IL-2 is able to stimulate a sustained in vivo peripheral blood LAK cell effect in dogs with the absence of major side effects. These findings suggest that these methods may have both research application in large animals and clinical application in patients with tumors that are responsive to LAK cell lysis.

Methods: Seventy-three patients with advanced malignancy were treated with the recombinant lymphokine interleukin-4 either as the sole immunotherapeutic reagent or in combination with recombinant interleukin-2., Results: Twelve of 84 courses of therapy were complicated by gastroduodenal erosion or ulceration. Three of these courses were associated with significant bleeding, which required multiple red blood cell transfusions and endoscopic therapy. No treatment-related deaths occurred. Eleven of 57 courses administered with concomitant indomethacin and 11 of 62 courses administered with ranitidine resulted in gastroduodenal mucosal injury. No acute change in gastric acid output occurred after one dose of interleukin-4 in patients prospectively evaluated with an indwelling nasogastric tube. An intravenous ranitidine infusion appropriately reduced acid output in these patients. In contrast, we have treated over 650 patients with interleukin-2 and indomethacin without interleukin-4, none of whom developed signs or symptoms of gastroduodenal ulceration., Conclusions: These observations suggest that systemically administered cytokines may exert an effect on the integrity of the gastroduodenal mucosa.

Antibody to tumour necrosis factor (TNF Ab) markedly decreases the toxicity of systemic interleukin-2 (IL-2) in mice but does not completely abrogate the anti-tumour response in terms of number of pulmonary metastases. Experiments were performed with a murine model of pulmonary metastases treated with high-dose IL-2 and concomitant TNF Ab or control antibody (CON Ab) to determine the effects of TNF Ab on survival. Mice were given either equal doses of IL-2 and TNF Ab or CON Ab or equitoxic doses of IL-2. In four consecutive experiments mice given TNF Ab tolerated 5 to 6 additional IL-2 doses (a 40-60% increase in total doses) in the equitoxic IL-2 dose group compared to the maximally tolerated dose with CON Ab. In all four experiments TNF Ab-treated mice had decreased survival compared to the CON Ab group given equal doses of IL-2 and in two of four experiments this difference was statistically significant (P2 < 0.01). Mice given 40-60% additional doses of IL-2 with TNF Ab had no improvement in survival compared with equitoxic doses of IL-2 with CON Ab in three of four experiments (P2 = 0.32, P2 = 0.67, P2 = 0.69). The TNF Ab preparation had no direct inhibition of IL-2 activity in an in vitro IL-2 proliferation bioassay. TNF Ab consistently blocks IL-2 toxicity and it also abrogates IL-2 therapeutic efficacy such that survival parallels treatment toxicity in this experimental model.

The purpose of this study was to evaluate the efficacy of treatment with recombinant interleukin-2 (rIL-2) and lymphokine-activated killer cells in patients with advanced bladder cancer and to study the induced changes in the distribution of leukocyte subsets in blood and tumor. Nine patients with metastatic transitional cell cancer of the bladder were treated with a continuous infusion of rIL-2 combined with lymphocytes stimulated in vitro with rIL-2. None of the patients responded to the therapy despite substantial changes observed in the immunological cells, both in tumor and blood. The rIL-2 infusion induced migration of leukocytes to the tumors, which was related to increased expression of the adhesion molecule VLA-1 on both peripheral blood mononuclear cells and the endothelial cells of small tumor vessels. Only T-cells, predominately expressing IL-2 receptors, and macrophages infiltrated the tumors. Natural killer cells remained few or absent in the tumors, even though the natural killer cells in peripheral blood were activated by the treatment. This study shows that the present technique of rIL-2 and lymphokine-activated killer cell therapy is able to induce substantial changes in the immune system of patients with metastatic bladder cancer. However, this treatment did not induce tumor regression, which may be due to the advanced stage of disease.

In this Phase I trial, the feasibility of sequential administration of continuous intravenous recombinant interleukin-2 (rIL-2) at 18 x 10(6) IU/m2/day for 6 days, followed by three daily bolus intravenous recombinant tumor necrosis factor (rTNF) infusions at doses escalating between 10 and 120 micrograms/m2/day, was investigated in 31 patients with metastatic malignancies. Prophylactic use of indomethacin prior to and during rTNF administration was found to significantly reduce toxicity. However, despite prophylactic indomethacin, a maximum tolerated dose of rTNF of 120 micrograms/m2 was reached. The limiting toxicity was hypotension. Predictable flu-like toxicities (i.e., fever/chills, hypotension, gastrointestinal toxicity, edema, malaise) were seen in most patients. These started during the rIL-2 infusion and continued during rTNF administration, particularly in the absence of indomethacin. Hematological, renal, and hepatic toxicities were not dose limiting. These toxicities were all reversible after treatment interruption. Pulmonary toxicity [i.e., anaphylactic-like reactions, bronchospasms, and adult respiratory distress syndrome (ARDS)] was seen in several patients immediately after rTNF infusions, irrespective of the rTNF dose or treatment cycle, and mainly in patients with extensive pulmonary metastases. The combined effect of treatment-related ARDS, lung metastases, and a Guillain-Barré syndrome led to the death of one patient. Two partial responses were seen in this study (i.e., breast and renal cancer). Based on these results, a Phase II trial of rIL-2 followed by rTNF has been initiated in metastatic breast cancer patients.

Interleukin 2 (IL-2), a T lymphocyte product released upon antigen stimulation, has been used for cancer therapy in high doses for more than five years. More recently, its potential as a stimulant of cell-mediated immunity in infectious diseases, particularly those caused by intracellular microbes, has become appreciated. Drawing on the extensive information available as to the structure, cellular and molecular effects of IL-2, this review focuses on its use in patients with lepromatous leprosy and AIDS in low, physiologic doses. The data indicate that IL-2 is effective in stimulating cell-mediated immunity without systemic toxicity.

Using an enzyme-linked immunosorbent assay (ELISA), we have measured serum levels of a soluble form of the p55 subunit of the interleukin-2 receptor complex, soluble CD25 (sCD25), at regular intervals in the sera of 51 pediatric and adult cancer patients receiving recombinant human interleukin-2 (IL-2). The IL-2 was administered in repetitive weekly cycles alone or in combination with lymphokine-activated killer (LAK) cells. Levels of CD25 correlated with clinical toxicities reflected by nadir blood pressures, percentages of weight gained, and minimum Karnofsky performances during IL-2 therapy. Coadministration of autologous in vitro activated LAK cells together with IL-2 did not significantly affect the pattern of sCD25 release relative to administration of IL-2 alone. Examination of sCD25 release in response to different doses of IL-2 revealed a statistically significant dose effect of IL-2 on the sCD25 levels in patient sera. In addition, the level of sCD25 in patient sera also correlated strongly with expression of CD25 on the surface of peripheral blood lymphocytes (PBL) obtained from patients following IL-2 therapy. These studies demonstrate the utility of the sCD25 ELISA as a clinical tool for monitoring patients on treatment regimens that include IL-2.

Recombinant bovine interleukin-2 (rBoIL-2) was administered as a single intramuscular bolus to healthy calves to determine the minimal dose capable of exerting a biological response. Doses ranging from 2.5 to 0.05 micrograms rBoIL-2/kg did not induce pyrexia, diarrhea, or depression, nor did they alter any blood chemistry or hematological parameters commonly associated with IL-2 toxicity. Moreover, the only significant immunological change observed was a reduction in the number of peripheral blood lymphocytes identified with the monoclonal antibodies B7A, BAQ4A (WC1+ cells), CACTB6A (WC2+ cells) and DH59B (monocytes). The decrease in cells associated with these markers did not influence non-MHC restricted cytotoxicity or in vitro lymphocyte proliferative responses to mitogens and IL-2. The treatments had no effect on delayed type hypersensitivity responses to phytohemagglutinin. These results indicate that IL-2 may be involved in the regulation of trafficking patterns of a unique subpopulation of lymphocytes in cattle.

The antitumor effects of recombinant interleukin-2 (rIL-2) and mismatched double-stranded RNA (dsRNA) were assessed in tissue culture and in a nude mouse model. Mismatched dsRNA did not show a direct antiproliferative effect against the human malignant melanoma cell line, BRO, in tissue culture. However, treatment of the BRO cells with up to 1000 units/ml rIL-2 in culture showed a slight increase in growth rate. Combined rIL-2/mismatched dsRNA treatment also demonstrated a similar slight enhancement of growth. Nude mice bearing subcutaneous tumors were treated by intraperitoneal injection of low doses (5000-20,000 units) of rIL-2 and mismatched dsRNA (500 micrograms). The in vivo tumor growth was significantly inhibited by the combined treatments (P less than 0.05) and survival was significantly increased (P less than 0.05). Measurement of cytotoxicity using splenocytes from treated animals showed significant augmentation of lytic activity against natural killer (NK)-sensitive YAC-1 cells in all rIL-2/mismatched dsRNA treatment groups, compared to the individual treatments or controls (P less than 0.05). Cytotoxicity of the splenocytes against the NK-resistant BRO cells was also augmented in animals treated with mismatched dsRNA and the highest rIL-2 dose utilized here (P less than 0.01). Renal, liver, and hematological toxicity was evaluated by measurement of blood urea nitrogen, creatinine, serum asparrtate aminotransferase, and a complete blood count with differential. There were no significant differences in these parameters in any of the treatment groups. Similarly, no differences in weight of the animals was seen in any treatment group. These results indicate that the combination of low-dose rIL-2 and mismatched dsRNA can potentiate host-mediated antitumor effects, yielding increased survival, without significant toxicity.

Between April 1988 and December 1990, 37 patients with progressive histologically proven metastatic melanoma were treated with interleukin-2 according to three multicentric successive protocols. Eighteen males and 19 females entered the trial. Mean age was 44 years (range 20-66); none of the patients had severe visceral disease or brain metastasis. Superficial and visceral metastatic sites were equally distributed. Interleukin-2 was administered as a 3 to 5 day continuous intravenous infusion, at a dose varying from 16 to 24 million international units/m2/day, as previously described by West. The second course was given after a 9 to 16 day free interval and one to seven courses were administered (mean three courses). A total of 132 courses has been given to the 37 patients. All are evaluable for toxicity and efficacy. Toxicity was tolerable and not different from that presented in recent reports. Only four patients had to definitively stop therapy for toxicity, one of them for cardiotoxicity; a dose modification or a transient suspension of therapy occurred in 18% of treatment cycles. One hypothyroidism with anti-thyroglobulin and anti-microsome antibodies was observed. We observed eight major responses (21.6%), usually of short duration (2-6 months). Most responses occurred in superficial lesions. One patient remains in complete remission, as therapy is stopped for 40 months. Immunological parameters, although demonstrating induced immunostimulation, did not correlate with clinical outcome. With an overall response rate of 21.6%, we confirm the activity of interleukin-2 in melanoma, as previously reported by others.

Nine patients with a recurrent malignant glioma were treated with repeated intracavitary or intracerebroventricular injections of human recombinant interleukin-2 (rIL-2) alone or in combination with systemic interferon-alpha (IFN-alpha). Five patients received only rIL-2 and four were treated with rIL-2 plus subcutaneous injections of IFN-alpha. Therapy was administered on a Monday, Wednesday, Friday schedule for up to 10 weeks, beginning with a dose of 10,000 IU rIL-2/injection. Doses were escalated every two weeks until some toxicity was apparent. The maximum amount of rIL-2 any one patient in this group received was 580,000 IU. Patients on combination immunotherapy were held at an rIL-2 dosage of 10,000 IU while IFN-alpha, which began at 3 million IU, was escalated every other week up to 18 million IU/dose. They were then held at that IFN-alpha dosage and rIL-2 was increased to 50,000 IU. The total amount of rIL-2 and IFN-alpha any one in this group received was 510,000 IU and 417 million IU, respectively. Repeated injections of 10,000 IU rIL-2 were well-tolerated by all nine patients and no change in their functional status was seen. At doses at 50,000 IU rIL-2, increased edema around the tumor cavity was observed by MRI/CT scand in 3/5 patients and clinical side-effects in the form of somnolence and headache along with some morbidity specifically associated with tumor location were also seen. Patients receiving rIL-2+ IFN-alpha showed progressive fatigue, muscle weakness, and occasionally nausea. Two of these patients showed increased peritumoral edema on MRI/CT scan.(ABSTRACT TRUNCATED AT 250 WORDS)

A phase I trial was performed to assess the immunomodulatory activities, maximum tolerated doses, and the toxicity of recombinant interleukin-2 (rIL-2) administered in combination with doxorubicin to patients with refractory malignancies. Therapy was administered to successive cohorts of four to six patients who were treated at three different dose levels (1A, 1B, 2A). Levels 1-2 refer to doxorubicin (40 or 60 mg/m2) given as an intravenous (i.v.) bolus on day 1, and levels A-B refer to rIL-2 (1.0 or 3.0 x 10(6) U/m2) given as a continuous i.v. infusion on days 2-5, 9-12, and 16-19. Cycles were repeated every 28 days. Seventeen patients were entered in the trial. Dose limiting toxicity consisted of neutropenia, and the maximum tolerated dose (MTD) of the combination was doxorubicin 40 mg/m2 and rIL-2 3.0 x 10(6) U/m2. No objective responses were observed. Lymphocytosis related to rIL-2 occurred and flow cytometry demonstrated significant increases in the following subsets: CD3+CD25+HLADr+ and CD11b-CD16c+CD8-. Natural killer cell activity and lymphokine-activated killer (LAK) cell precursors were increased in patients treated at dose levels 1A and 1B (40 mg/m2 doxorubicin), but no consistent changes in LAK activity were noted. No clinical responses were seen and the overall toxicity of this combination was moderate to severe. Administration of doxorubicin prior to rIL-2 does not enhance the immunologic effects of rIL-2.

In these studies, the effects of recombinant human interleukin 2 (rHuIL-2) were examined on in vitro tumor cytotoxicity by canine blood lymphocytes obtained from peripheral vessels through use of a chromium release microcytotoxicity assay. Cytotoxic activity by lymphokine-activated killer cells was significantly increased, compared with that by untreated lymphocytes in a dose-dependent manner. The maximal effect was attained with 300,000 IU of rHuIL-2/ml. Lymphokine-activated killing also was dependent on the duration of incubation with rHuIL-2. After 1 day of rHuIL-2 incubation, cytotoxicity was significantly increased, compared with that of untreated lymphocytes. Of the 3 times examined, cytotoxicity peaked after 3 days of rHuIL-2 incubation. High levels of cytotoxic activity were still present at 7 days of incubation. Numbers of granular lymphocytes increased over the times examined. These results demonstrate functional and morphologic changes in canine peripheral blood lymphocytes obtained from peripheral vessels after incubation with rHuIL-2 in a dose- and time-dependent manner.

A Phase I trial of tumor-infiltrating lymphocytes (TIL) expanded in vitro and administered on Days 1 and 8, with or without continuous infusion recombinant interleukin 2 (rIL-2) in 25 patients with metastatic renal cell carcinoma, was conducted. Eighteen of the 25 eligible patients were treated with TIL and escalating doses of rIL-2 (0.0, 3.0, 4.5 x 10(6) units/m2) on Days 1 to 5 and 8 to 12. Dose-limiting toxicity was pulmonary, and the maximum tolerated dose of rIL-2 was 3.0 x 10(6) units/m2. No clinical responses were observed. Immunological monitoring of peripheral blood lymphocytes demonstrated significant increases in CD3+ and CD56+ cells, including the activated T-cell subsets. Phenotypic analysis of cultured TILs demonstrated significant heterogeneity and the presence of CD3+CD4+ and CD3+CD8+ T-cells, with CD3-CD56+ and CD3+CD56+ populations also present. The majority of cultured TILs expressed HLA-DR and CD45RO, with a variable number expressing CD25. The rIL-2-expanded TILs possessed cytotoxicity against allogeneic and autologous tumor, with cytolytic activity against only autologous tumor seen in one patient. Results demonstrate that in vitro expansion of TILs is possible, but further studies are needed to define the biology of TILs in renal cancer and to isolate and expand tumor-specific T-cells.

Interleukin-2 (IL-2) mediates the regression of metastatic cancer, but clinical application has been limited by the induction of dose-dependent toxicities in normal tissues. The most clinically significant toxicities occur secondary to a vascular leak syndrome and include acute respiratory failure and hemodynamic instability. Because previous studies suggested a role for pentoxifylline in attenuating the toxic effects of IL-2, we hypothesized that pentoxifylline would inhibit alterations in the microvasculature induced by IL-2 and would ultimately reduce IL-2-induced toxicity. To determine the validity of this hypothesis, we prepared four groups of rats for in vivo microvascular observation. In the first group, a bolus intravenous injection of IL-2 (1 x 10(6) units/kg) acutely induced hypotension, tachypnea, hypoxia, increased lung water, decreased microvascular blood flow, and increased leukocyte-endothelial adherence. No significant changes occurred in animals treated by pentoxifylline alone or the control IL-2 vehicle-alone group. However, pentoxifylline inhibited many of the IL-2-induced systemic and microvascular effects, such as hypotension, tachypnea, increased lung water, hypoxia, and increased leukocyte-endothelial adherence, but not tachycardia or increased microvascular protein leakage. These data support our hypothesis that systemic toxicities induced by IL-2 are associated with alterations in the microcirculation, which may be ameliorated by pentoxifylline.

A variety of side effects have been reported with the use of interleukin-2 alone or in combination with lymphokine-activated killer cells in patients with disseminated neoplasms. The present study was undertaken to determine the effects of high-dose interleukin-2 administration in normal rats. Sprague-Dawley rats were treated with intravenous recombinant interleukin-2 (900,000 IU/kg/day) for 9 consecutive days. Animals were placed in individual metabolic cages, and arterial blood pressure, food intake, body weight, and urine output were monitored. On day 10, animals were killed by exsanguination, various tissues were harvested, and a variety of hematologic and chemical assays were performed. The results were compared with those of placebo-injected normal control and pair-fed groups. The interleukin-2-treated group exhibited anorexia, weight loss, hypotension, anemia, leukocytosis, lymphocytosis, eosinophilia, hypercalcemia, azotemia, and a marked urinary concentration defect. Histologic examination of various tissues revealed widespread infiltration with mono-nuclear cells and eosinophils in most organs, especially in the lungs and liver of interleukin-2-treated animals. Other abnormalities included severe panlobular hepatitis, hepatocellular necrosis, and thymic involution. Renal involvement was mild and consisted of focal interstitial infiltration by mononuclear cells. According to these observations, administration of high-dose interleukin-2 in normal rats results in a score of significant functional, biochemical, and histologic abnormalities.

Interleukin 2 (IL-2) treatment of malignancies is often associated with severe toxicity, and the alterations observed after high dose administration of IL-2 are similar to those induced by recombinant tumor necrosis factor (TNF). We therefore examined the hypothesis that IL-2 induces TNF gene expression in vivo. Purified, recombinant human IL-2 was injected intraperitoneally into mice which had been previously primed with complete Freund's adjuvant (CFA). Biologically-active TNF was detected in the ascites fluid of CD-1 mice; it was detectable 30 minutes after IL-2 and peaked at 1 hour (500 +/- 158 units/ml). Plasma levels of TNF also peaked at 1 hour at 32 +/- 4 units/ml. Similar kinetics were observed in CBA/J mice. TNF specific mRNA was also present in the ascites cells, and peaked 30 minutes after IL-2 injection into CBA/J mice. Injection of vehicle containing 10 times the maximum contaminating dose of endotoxin did not induce TNF above background levels. As a further control for potential endotoxin contamination, IL-2 was injected into endotoxin hyporesponsive C3H/HeJ mice. These mice also demonstrated the rapid upregulation of biologically-active TNF in the ascites, with peak production occurring at 1 hour (125 +/- 47 units/ml). The induction of biologically-active TNF in the C3H/HeJ mice was associated with a peripheral blood neutrophilia and lymphopenia, pathophysiologic alterations that have been attributed to TNF. These data show that a single injection of purified, recombinant IL-2 induces TNF gene expression in vivo.

The level of intracellular glutathione (GSH) in mitogen-stimulated mouse lymphocytes is increased in the presence of 2-mercaptoethanol (2-ME), an enhancer of lymphocyte activation and proliferation. Since proliferation of lymphocytes in response to mitogens involves direct activation by a mitogen followed by continued proliferation in response to interleukin-2 (IL-2), we have investigated the effect of 2-ME and exogenous IL-2 on the GSH content and cell proliferation of rat lymphocytes stimulated with phytohemagglutinin (PHA). PHA stimulation increased both GSH content and the magnitude of the proliferative response, as measured by thymidine incorporation into cellular DNA. However, incubation of stimulated lymphocytes with 2-ME or IL-2 for 72 hr produced a significant further elevation of GSH levels and thymidine incorporation. 2-ME also increased the GSH content in unstimulated cultures, but it had little effect on thymidine incorporation. IL-2 increased GSH content and decreased thymidine incorporation in unstimulated lymphocytes. Exposure of cells to DL-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of GSH biosynthesis, significantly depleted GSH and lowered the proliferative response, suggesting a crucial role of de novo GSH synthesis for lymphocyte activation. The data suggest that both 2-ME and IL-2 promote lymphocyte proliferation, although the mechanisms by which intracellular GSH levels are increased by the agents are apparently different.

Interleukin-2 (IL2), as a modifier of the biological response, has been intravenously used in patients with advanced cancer associated or not to LAK cells or tumor infiltrating lymphocytes. In different neoplasias positive results have been obtained, being effective in melanoma and renal cancer. There are still, at present, many questions to be answered and multiple research lines are currently open. The association with other cytokines and new chemotherapy protocols grant new therapeutic possibilities.

Studies were conducted in cats to determine whether this species could serve as an animal model for the therapeutic toxicity of recombinant interleukin-2 in humans, and to establish the role of the area postrema in the causation of recombinant interleukin-2-induced vomiting. Injections of recombinant interleukin-2 (3.6 x 10(6) IU/kg, i.m.), given once every 24 hr for one to three days, evoked repeated vomiting in 4 out of 6 area postrema intact cats and in 3 out of 3 area postrema-ablated cats. These results suggest that the area postrema is not essential for the emetic action of recombinant interleukin-2. In anesthetized intact cats, no remarkable changes in ventilation, blood pressure, heart rate or blood pH were observed over 4.5 to 54 hr of continuous physiological recording after i.v. injections of recombinant interleukin-2 to a total dose as high as 27 x 10(6) IU/kg. Cat lymphocytes responded appropriately to the cytokinetic action of human recombinant interleukin-2.

Interleukin 2 (IL-2) is a potent cytokine with diverse effects, including the ability to stimulate lymphocyte differentiation into cells capable of lysing tumor. Its therapeutic efficacy is limited because of side effects such as breakdown of the microvascular barrier and edema. Control of the microvascular barrier is in part regulated by endothelial cell cytoskeletal contractile proteins. This study tests whether the cyclopeptides that maintain actin filament organization and distribution and reduce macromolecular flux across the endothelial cell junction in vitro would similarly maintain barrier tightness and prevent early edema produced by IL-2 in vivo. Anesthetized rats were treated at 30-min periods with intravenous saline (0.5 ml, n = 41), phalloidin (20 micrograms in 0.5 ml, n = 21), or antamanide, (20 micrograms in 0.5 ml, n = 21), starting 30 min before the 1-h infusion of 10(6) U of recombinant human IL-2 or saline. Six hours after the start of IL-2, there was edema in the saline/IL-2 group, as measured by increased wet-to-dry ratios (W/D) in the lungs, heart, and kidney. With saline/IL-2, bronchoalveolar lavage (BAL) fluid contained an elevated protein concentration and higher plasma thromboxane levels compared with controls. The number of neutrophils sequestered in the lungs was more than twice that of saline controls. Phalloidin significantly attenuated edema in lung and reduced BAL protein leak. Antamanide treatment was as effective in limiting lung and heart edema, but, in contrast to phalloidin, antamanide prevented kidney edema and did not lead to an alteration in the liver W/D. Antamanide also prevented BAL fluid protein leak.(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-2 therapy leads to respiratory dysfunction caused by increased vascular permeability. This study examines the role of oxygen-derived free radicals (OFR). Sheep (n = 6) with chronic lung lymph fistulae were given interleukin-2, 10(5) units/kg, as an intravenous bolus. The mean pulmonary artery pressure rose from 13 to 23 mm Hg (p less than 0.05) at 1 hour and remained elevated for 4 hours, although the pulmonary artery wedge pressure was unchanged at 4 mm Hg. Arterial oxygen tension fell from 88 to 77 mm Hg (p less than 0.05). Lung lymph flow rose from 2.2 to 6.4 ml/30 min (p less than 0.05) at 3 hours. This rise coincided with an increase in the lymph/plasma protein ratio from 0.67 to 0.77 (p less than 0.05) and lymph protein clearance from 1.5 to 4.4 ml/30 min (p less than 0.05), indicating increased lung microvascular permeability. Interleukin-2 led to transient increases in plasma thromboxane B2 from 168 to 388 pg/ml (p less than 0.05) and lung lymph thromboxane B2 from 235 to 694 pg/ml (p less than 0.05). The leukocyte count fell from 8156 to 4375/mm3 (p less than 0.05) primarily caused by a 78% drop in lymphocyte count. Platelet count declined from 292 to 184 X 10(3)/mm3 (p less than 0.05). Pretreatment with the hydroxyl radical scavenger dimethylthiourea, 1 gm/kg, intravenously, (n = 6) prevented the interleukin-2-induced increase in mean pulmonary artery pressure, lung lymph flow, lymph/plasma protein ration, lymph protein clearance, and thromboxane B2 levels in plasma and lung lymph. The arterial oxygen tension decreased from 85 to 80 mm Hg (p less than 0.05). The leukocyte count declined from 7854 to 6229/mm3 (p less than 0.05), but this was not as low nor as prolonged as the interleukin-2 group. Further, the decrease in platelet count was prevented (p less than 0.05). Interleukin-2 incubated with sheep or human leukocytes led to a dose-dependent increase in intracellular hydrogen peroxide production by neutrophils as measured by flow cytometry of dichlorofluorescein oxidation. These data indicate that interleukin-2 stimulates OFR generation and that OFR moderate the interleukin-2-induced increased lung permeability.