The Immunotherapy of Human Cancer with Interleukin 2: Present Status and Future Directions

Interleukin-2 (IL2) is the principal soluble factor responsible for the proliferation of activated T cells. In animal models and humans, administration of IL2 can induce regressions of established cancers. These antitumor effects may be partially mediated by cytotoxic effector cells activated by IL. 2, including lymphokine-activated killer (LAK) cells and cytotoxic T lymphocytes. IL2 has additional effects on other components of the cellular immune system, including B cells and macrophages, and induces secretion of other soluble mediators, including tumor necrosis factors (TNF) alpha and beta, and interferon-gamma. These effects may contribute to the antitumor activity of IL-2 as well as its dose-related toxicity. Multiple Phase I and II trials have been completed or are ongoing evaluating the clinical and biological effects of IL2 given by diverse routes and schedules, both alone and in combination with infusions of ex vivo IL2-activated autologour LAK cells. Other studies have begun to explore the potential for antitumor synergy when IL-2 is combined with the different interferons, TNF, monoclonal antibodies, and cytotoxic drugs. The biology, toxicity, and clinical activity documented in IL-2 clinical trials to date are reviewed, and prospects for future directions outlined.     

Circulating cytokines in patients with metastatic cancer treated with recombinant interleukin 2 and lymphokine-activated killer cells

Treatment with recombinant interleukin 2 and lymphokine-activated killer cells (rIL-2/LAK) has produced a clinical antitumor effect in preliminary human trials. The cytokines gamma-interferon (IFN-gamma), tumor necrosis factor alpha (TNF-alpha), and tumor necrosis factor beta (TNF-beta, lymphotoxin) have potent in vitro antitumor activity and some clinical toxicities similar to interleukin 2 (IL-2)/LAK. This study sought to determine whether these cytokines were detectable in sera of IL-2/LAK-treated patients. Ten patients were treated with a protocol of 5-day i.v. rIL-2 bolus priming (10(5) units/kg, every 8 h), followed by 5 daily phereses with harvested lymphocytes cultured in vitro to generate LAK, and 5 days of rIL-2 bolus with infusion of LAK cells. Five patients were treated with a protocol modified to a 3-day rIL-2 prime and 6-day continuous infusion rIL-2 (3 x 10(6) units/m2/day) with infusion of LAK cells. Serum specimens were obtained prior to and 0.5, 2, 3, and 5 h after IL-2 or LAK cell administrations. IFN-gamma was detected by enzyme-linked immunosorbent assay, TNF-alpha by WEHI 164 bioassay or enzyme-linked immunosorbent assay, and TNF-beta by WEHI 164 cell bioassay. During the prime, few patients manifested in vivo detectable serum cytokines: IFN-gamma, three of ten, 5-day prime (1.03 +/- 0.46 ng/ml), and zero of five, 3-day prime; TNF-alpha, one of ten, 5-day prime, and one of three, 3-day prime; TNF-beta, one of ten, 5-day prime. The supernatants of in vitro LAK generation cultures had detectable levels of cytokines at 24 h which increased progressively until culture harvest at Day 4 (IFN-gamma, 2.56 +/- 0.34 ng/ml; TNF-alpha, 356 +/- 110 pg/ml; TNF-beta, 8.2 +/- 4.4 units/ml). The highest levels of in vivo serum cytokines occurred following LAK cell infusion and were more often elevated in patients receiving rIL-2 by bolus than by continuous infusion: IFN-gamma, four of six bolus, zero of three continuous infusion; TNF-alpha, six of six bolus (maximum 679 pg/ml) versus two of three continuous infusion (maximum, 106 pg/ml). LAK cells in vitro responded with cytokine release on stimulation by tumor cell lines (IFN-gamma, 0.88 +/- 0.06 ng/ml; TNF-alpha, 426 +/- 16 pg/ml; TNF-beta, 0.64 +/- 0.06 units/ml). In summary, this preliminary study has detected circulating cytokines in sera of patients receiving IL-2/LAK therapy. The greatest cytokine elevations followed LAK cell infusion.(ABSTRACT TRUNCATED AT 400 WORDS)     

Shedding of tnf-receptor and soluble icam-1 molecules in response to treatment with recombinant tnf-alpha and IFN-gamma in patients with colorectal-cancer

Intercellular adhesion molecule-1 (ICAM-1) and tumor necrosis factor alpha receptor (TNF-R) have been detected in soluble forms in the serum of cancer patients. sICAM-1 can abrogate non MHC-restricted cytotoxicity mediated by NK and lymphokine-activated killer cells. Soluble TNF-alpha receptors have been shown to neutralize the cytopathic activity of recombinant TNF-alpha. Both mechanisms can facilitate neoplastic cells to escape immunosurveillance. Serum levels of sTNF-R and sICAM-1 were determined during a phase II combination trial with TNF-alpha and IFN-gamma in eight patients with colorectal cancer. Serum concentrations of sTNF-R and sICAM-1 increased significantly during the treatment period. Shedding of ICAM-1 and TNF-R induced by TNF-alpha and IFN-gamma could be responsible for the ineffectivity of TNF-alpha based treatment regimens.     

Characterization of OKT3-initiated lymphokine-activated effectors expanded with interleukin 2 and tumor necrosis factor alpha

Synergistic and cooperative effects in vitro of OKT3, interleukin 2 (IL-2), and tumor necrosis factor alpha (TNF) as stimuli in generating effectors with lymphokine-activated killer activity were studied. Activation of human peripheral blood mononuclear cells with OKT3 (10 ng/ml) for 48 h, followed by culture in low concentrations of IL-2 (10 units/ml) and TNF (250 units/ml) resulted in higher cell recovery (50- to 3300-fold) compared to the number of cells in the initial culture and enhanced lytic activity against both Raji and fresh lung tumor targets (mean 100-fold) by day 30 compared to those expanded with higher concentrations of IL-2 (100 units/ml) alone. Immunofluorescence analysis of peripheral blood mononuclear cells initiated with OKT3 and expanded with IL-2 plus TNF revealed a selective increase in CD8+ cells and a decrease in CD4+ by day 28; the opposite effect was observed when cells were incubated with 100 units/ml of IL-2 alone, resulting in a greater proportion of CD4+ cells. Almost all cells were CD3+. Studies of cytokine receptor expression indicated that OKT3 plus IL-2 plus TNF caused an earlier up-regulation of the IL-2 receptor beta chain (Tac) and higher TNF receptor expression by day 6 compared to 100 units/ml IL-2 alone. Significant TNF levels (greater than 17 units/ml) were measured in culture supernatants from peripheral blood mononuclear cells initiated with OKT3 alone. Collectively, our data demonstrate that induction of lymphokine-activated killer activity with OKT3, followed by culture in low concentrations of IL-2 plus TNF is an alternative to the use of high-dose IL-2 alone and suggest that this combination may provide potential advantages in long-term generation of cytolytic cells.     

Cytokine gene expression during the generation of human lymphokine-activated killer cells: early induction of interleukin 1 beta by interleukin 2

Culture of human peripheral blood leukocytes with interleukin 2 (IL-2) stimulates their differentiation into lymphokine-activated killer (LAK) cells, with a broad range of cytotoxicity against fresh tumor cells and tumor cell lines (Grimm et al., J. Exp. Med., 155: 1823-1841, 1982). We chose to utilize a molecular approach to determine whether IL-2 stimulates the expression of cytokine genes by the mixed cell population which may be involved in the generation or regulation of lytic activity. Northern blot analysis performed with total cellular RNA from LAK cells cultured for varying periods of time with IL-2 revealed that the genes which code for cytokines [interleukin 1 (IL-1)alpha and beta, gamma-interferon, tumor necrosis factor alpha, and lymphotoxin] were not spontaneously expressed. As soon as 2 h after IL-2 treatment, IL-1 alpha and IL-1 beta mRNAs were expressed. Both nonadherent and adherent populations of LAK cells express IL-1 beta mRNA; however, the adherent population produced more IL-1 beta mRNA and maintained its expression for a prolonged period of time. Other cytokine mRNAs (gamma-interferon, tumor necrosis factor alpha, and lymphotoxin) were expressed later than the IL-1 mRNAs with maximal levels between Days 2 through 7. Our results indicate that LAK cell populations can generate a variety of cytokines which may be involved in the generation of lytic activity.     

Combined effects of chemotherapy and interleukin 2 in the therapy of mice with advanced pulmonary tumors

We have evaluated the effects of chemotherapeutic agents on the toxicity and antitumor benefit of therapy of established murine tumors by high-dose interleukin 2 (IL-2). Cyclophosphamide (Cy), doxorubicin, and bischloroethylnitrosourea were given to normal mice prior to IL-2 administration to test the effects of these agents on IL-2-induced toxicity. Cy at doses of 100 mg/kg and 150 mg/kg completely protected mice from a 100% lethal dose of IL-2, and doses of 50 mg/kg and 150 mg/kg allowed the administration of a median of 4.5 and 10.0 more doses of IL-2, respectively, before death from IL-2 toxicity occurred. Doxorubicin at 8 mg/kg and bischloroethylnitrosourea at 20 mg/kg did not impact on toxicity in IL-2-treated mice. In mice bearing pulmonary metastases of the weakly immunogenic MCA-105 sarcoma, IL-2 increased median survival time from 33 (no IL-2) to greater than 60 days for all doses of IL-2 tested when combined with a single injection of Cy at 75 mg/kg (P less than 0.002). Increasing doses of either Cy or IL-2 produced increasing benefits on survival which were always greater than either treatment alone. These effects of Cy and IL-2 were also seen in mice bearing the nonimmunogenic MCA-101 sarcoma and a murine adenocarcinoma (MCA-38). Doxorubicin and bischloroethylnitrosourea did not consistently enhance the effects of IL-2 treatment. Cy appears to reduce the yield of in vivo generated lymphokine-activated killer cells, but these lymphokine-activated killer cells are still lytic for fresh tumor targets in vitro. Thus, the mechanism of this synergy does not appear to involve stimulation of lymphokine-activated killer cell activity, but may in part involve reduction of tumor burden by the chemotherapeutic agent, an increase in susceptibility of tumor to cellular immune lysis, and/or a decrease in suppressor cell activity mediated by the chemotherapy.     

Interleukin-2 in the treatment of renal cancer

Interleukin-2 (IL-2) administered in pharmacologic doses to renal cancer patients with intact organ function and good performance status induces durable complete responses in about 5% of patients and partial responses in an additional 10% to 15%. The mechanism of antitumor efficacy of IL-2 is closely related to its ability to expand and activate cytotoxic lymphocytes of the natural killer (NK)- and thymic (T)-cell subsets that express IL-2 receptors (IL-2R). There is also accumulating evidence that local or generalized effector cell dysfunction, which is characteristic of patients with advanced cancer, can be reversed with IL-2 exposure. The toxicities of IL-2 are mediated by cytokines and other small molecules secreted by IL-2R-expressing cells responding to the binding of this ligand. The common mechanism for IL-2-induced multiorgan dysfunction appears to be a capillary leak syndrome directly mediated by local production of nitric oxide by cells of the monocyte-macrophage lineage. To date, efforts to improve on the antitumor activity of IL-2 by the addition of IL-2-activated peripheral blood mononuclear cells (lymphokine-activated killer [LAK] cells) or cell subsets selected for proximity or potential antigen-specificity (tumor-infiltrating lymphocytes [TIL]) have not led to improved therapeutic outcomes. Attempts to reduce the risks of IL-2 therapy (which could potentially allow for increased IL-2 administration) by blocking one or more of the known mediators of toxicity have also been disappointing. Current research is directed at developing combination regimens with additive or synergistic antitumor effects and incompletely overlapping toxicities, as well as the identification of tumor antigens that may be the target of more focused cellular therapies. The role of high-dose IL-2 in the adjuvant therapy of resected renal cancer at high risk of relapse is also under investigation.     

Effect of macrophages on interleukin-2 (IL-2)- and IL-4-induced murine lymphokine-activated killer activity

The enhancing effect of macrophages on interleukin-2 (IL-2)- and IL-4-induced murine lymphokine-activated killer (LAK) activity was investigated in this study. Peritoneal macrophages significantly enhanced LAK activity generated from accessory cell-depleted splenic lymphocytes in both IL-2 and IL-4 cultures. This effect was dependent on the number of macrophages and was not replaced by a factor derived from macrophages or lymphocytes. Macrophages enhanced IL-2- and IL-4-induced LAK activity against both natural killer (NK)-sensitive (YAC-I, P388D1) and NK-resistant (P815) tumor cells. Negative selection of cells with antibodies and complement showed no differences in surface markers between IL-2 LAK effectors and IL-4 LAK effectors generated in the presence of macrophages. These results suggest that the same LAK effector subsets can be enhanced by macrophages in either IL-2 or IL-4 cultures.     

Phase I study of ONO-4007, a synthetic analogue of the lipid A moiety of bacterial lipopolysaccharide.

ONO-4007 is a synthetic analogue of the lipid A moiety of bacterial lipopolysaccharide, which exhibits antitumor activity by the induction of intratumoral tumor necrosis factor alpha, the potentiation of tumor-infiltrating macrophages, and the inhibition of angiogenesis. Interleukin (IL)-1 alpha, IL-6, and IL-12 induction by ONO-4007 activates cytotoxic natural killer cells to up-regulate IFN-gamma and nitric oxide synthase activity. ONO-4007 was given to 24 patients (13 males and 11 females; median age, 53 years) as a 30-min i.v. infusion on day 1, followed on day 15 by a first treatment cycle consisting of three weekly infusions at the same dose, followed by a rest period of 1 week. Cohorts of six patients received up to a maximum of four treatment cycles at increasing dose levels (75, 100, and 125 mg). The maximum tolerated dose was 125 mg, with grade 3 National Cancer Institute Common Toxicity Criteria toxicity (rigors with cyanosis) occurring in two of six patients at this dose level. An additional six patients were treated at 100 mg, the dose below the maximum tolerated dose. Other toxicities included grade 2 National Cancer Institute Common Toxicity Criteria myalgia, nausea, and hypotension. The pharmacokinetics of ONO-4007 appeared to be independent of dose and showed linearity with respect to time. ONO-4007 has a low systemic clearance (approximately 1.3 ml/min) and a small volume of distribution (5-8 liters) with a long t1/2 of 74-95 h. The administration of ONO-4007 was shown to result in a significant increase in circulating levels of tumor necrosis factor alpha and IL-6. No objective antitumor responses were observed. Seven patients maintained stable disease for at least two cycles, whereas five patients maintained stable disease for the full four-cycle duration of the study. Additional studies are required to determine the antitumor activity of ONO-4007.     

Potent graft antitumor effect in natural killer-resistant disseminated tumors by transplantation of interleukin 2-activated syngeneic bone marrow in mice

The current study is a continuation of our previous work showing that bone marrow activated in interleukin 2 has antitumor and antiviral activity in vitro. The antitumor efficacy of IL-2-activated bone marrow cells in vivo was assessed here. Our results indicated that bone marrow cells activated in IL-2 for 3 days (ABM) have antitumor activity in vivo and cause significant tumor regression in mice being treated with ABM and concurrent i.p. administration of IL-2. In mice also bearing larger tumor burdens, those receiving ABM and i.p. IL-2 showed the most significant tumor regression. The ABM seem to be more potent than conventional IL-2-activated spleen lymphokine-activated killer cells. In studies done using lower dosages of IL-2 or log lower number of cells, the ABM caused more significant tumor regression than lymphokine-activated killer cells. We also assessed the antitumor efficacy of short term (1 day) IL-2-activated bone marrow, the short term-activated bone marrow being preferred in bone marrow, transplantation because of the minimum amount of cells lost due to its shorter incubation period. We also showed that short term-activated bone marrow caused tumor regression similar to ABM and could reconstitute lethally irradiated mice similar to fresh bone marrow. Therefore, the biomodulation of bone marrow cells could be used as an active therapeutic tool in autologous bone marrow transplantation, producing graft versus tumor effects without any graft versus host effect.     

Human small-cell lung-cancer cells are cytokine-resistant but NK/LAK-sensitive.

We have studied the effects of 8 cytokines and their combinations on the in vitro growth of 10 human small-cell cancer lines (SCLC). Interferon-alpha and gamma (IFN-alpha and gamma) caused significant but slight growth inhibition over a 7-day incubation period. However, none of the other 6 cytokines, tumor necrosis factor (TNF), lymphotoxin (LT), interleukin-1 beta (IL-1 beta) interleukin-2 (IL-2), transforming growth factor-beta 1 (TGF-beta 1), or granulocyte colony-stimulating factor (G-CSF), modified SCLC cell proliferation. In contrast, all 10 lines were sensitive to lysis by natural killer (NK) and lymphokine-activated killer (LAK) cells. Sensitivity to LAK cells could be increased by pretreatment of SCLC cells with IFN-gamma. As resistance to the cytostatic/cytotoxic activity of some cytokines has been associated with autocrine production of cytokines, we screened the SCLC lines for cytokine mRNAs. Within the limits of detection of the assay we found no expression of TNF, TGF-beta 1, IL-1 beta or IL-6 mRNA in the 10 SCLC lines.     

Differential stimulation of macrophages for tumor cytostasis and monokine production.

We have investigated whether antitumor activity could be expressed independently of cytokine production. Resident macrophages treated with interferon-gamma (IFN-gamma), lipopolysaccharide (LPS) plus muramyldipeptide (MDP) expressed a cytostatic activity against P815 tumor cells and released interleukin 6 (IL-6) and nitrite but produced neither IL-1 nor tumor necrosis factor (TNF). Thioglycollate-elicited macrophages required only LPS plus IFN-gamma for cytostatic activity which was expressed concomitantly with the release of high levels of TNF, IL-1 and IL-6, whereas C3H/HeJ macrophages produced low levels of monokines and were not cytostatic. LPS, alone, was sufficient for triggering Concanavalin A-primed macrophages leading to a full cytostatic activity, even in C3H/HeJ macrophages that was expressed, for these latter, in the absence of monokine production. TNF did not appear to play a role either in autocrine stimulation of macrophages or in the cytostatic process because anti-TNF antiserum affected neither the cytostatic activity nor the nitrite production.     

In vivo antitumor activity of interleukin 21 mediated by natural killer cells

Immunotherapy with high-dose interleukin (IL) 2 has been shown to successfully treat tumors in animal models and cause dramatic tumor regressions in some patients with metastatic melanoma, renal cell carcinoma, and non-Hodgkin's lymphoma. However, toxicity associated with IL-2 administration has compromised its widespread use in the clinic. IL-21 is a more recently discovered cytokine produced by activated CD4(+) T cells that shares significant sequence homology to IL-2, IL-4, and IL-15. Because IL-21 and IL-2 and their receptors share significant sequence similarities and both cytokines can stimulate T and natural killer (NK) cells, we sought to study whether IL-21, like IL-2, exhibits antitumor effects in vivo. In this study, we treated established s.c. tumor in mice by systemically administering plasmid DNA encoding murine IL-21 using a hydrodynamics-based gene delivery technique. Administration of IL-21 plasmid DNA resulted in high levels of circulating IL-21 in vivo. Treatment of tumor-bearing mice with IL-21 plasmid DNA significantly inhibited the growth of B16 melanoma and MCA205 fibrosarcoma in a dose-dependent manner without significant toxicity and increased the survival rate, compared with mice treated with control plasmid DNA. In vivo depletion of either CD4(+) or CD8(+) T cells did not affect IL-21-mediated antitumor activity. However, depletion of NK cells completely abolished IL-21-induced tumor inhibition. Consistent with this, the antitumor activity of IL-21 seemed to be mediated through enhanced cytolytic activity of NK cells. Our study suggests that IL-21 has significant antitumor activity and may have therapeutic potentials as an antitumor agent in the clinic.     

Induction of murine lymphokine-activated killer-like cells by Corynebacterium parvum (C. parvum) in vitro: lysis of tumor cells and macrophages by C. parvum-induced killer cells.

In vitro culture of murine spleen cells with Corynebacterium parvum (C. parvum) was found to induce lymphokine-activated killer (LAK)-like cells capable of killing both natural killer (NK)-sensitive and NK-resistant tumor cells as well as syngeneic macrophages (M phi). The induction of LAK-like activity by C. parvum was significantly inhibited by anti-interleukin-2 (IL-2) or anti-interferon (IFN) alpha, beta antibody (Ab), and it was further inhibited by the combination of two Abs, suggesting that the generation of killer cells by C. parvum was dependent on IL-2 and IFN(s) produced in the culture. It was considered that M phi were important in the induction of LAK-like cells by C. parvum because the depletion of M phi from spleen cells before culture with C. parvum significantly reduced the induction of killer activity. The majority of effectors mediating both tumor cells and M phi were Thyl+ and asialo-GM1 (aGM1)+, and the lysis of M phi by C. parvum-induced killer cells could be inhibited by the addition of cold YAC-1 tumor cells and P815 tumor cells, suggesting that the same population of effectors recognized tumor cells and M phi. These results demonstrated a possibility that the killing of M phi by C. parvum-induced killer cells might down-regulate anti-tumor effects of C. parvum.     

Phase II study of liposomal muramyl tripeptide in osteosarcoma: the cytokine cascade and monocyte activation following administration.

PURPOSE: A phase II trial that uses liposome-encapsulated muramyl tripeptide phosphatidylethanolamine (L-MTP-PE) in patients with relapsed osteosarcoma is underway. To determine if in vivo cytokine induction plays a role in the mechanism of action of L-MTP-PE, we investigated the circulating cytokine levels of 16 patients who were undergoing therapy. PATIENTS AND METHODS: Patients had histologically proven osteosarcoma and pulmonary metastases that developed either during adjuvant chemotherapy or that were present at diagnosis and persisted despite chemotherapy. Patients were rendered disease-free by surgery. The major goal of the study was to improve the disease-free interval in this high-risk group. L-MTP-PE 2 mg/m2 was infused during a 1-hour period twice a week for 12 weeks, then once a week for 12 weeks. Serial blood samples were collected after L-MTP-PE administration and were assayed for cytokine levels (tumor necrosis factor-alpha [TNF alpha] interleukin-1 alpha [IL-1 alpha], IL-1 beta, IL-6, interferon-gamma [IFN-gamma], neopterin, C-reactive protein). RESULTS: After the infusion of L-MTP-PE, there was rapid induction of circulating TNF alpha and IL-6. TNF alpha levels peaked 1 to 2 hours after infusion in 10 of 16 patients, whereas peak IL-6 levels were detected at 2 to 3 hours in all patients. Induction of circulating TNF alpha and IL-6 was evident only after the first dose of L-MTP-PE. Neither IL-1 alpha nor IL-1 beta was detected in the plasma. Neopterin levels increased at 24 hours postinfusion, which indicated macrophage activation, and were not related to the induction of circulating IFN-gamma. C-reactive protein was elevated in all patients at 24 hours and decreased by 72 hours. Unlike circulating TNF alpha and IL-6, elevations in C-reactive protein and neopterin could be detected throughout the treatment course. CONCLUSION: It is concluded that L-MTP-PE has specific biologic effects in patients with osteosarcoma that may be important to the drug's immunostimulatory capacity and its effectiveness as an antitumor agent.     

Down-regulation by interleukin 4 of activation of human alveolar macrophages to the tumoricidal state

The effect of recombinant human interleukin 4 (IL-4) on the expression of antitumor activity of human alveolar macrophages (AM) obtained by bronchoalveolar lavage from healthy donors was examined. AM were incubated for 16 h in medium with various macrophage activators [lipopolysaccharide, des-methyl muramyldipeptide, Nocardia rubra cell wall skeleton, and heptanoyl-gamma-D-Glu-(L)-meso-alpha,epsilon-A2pm(L)-D-Al aOH] in the presence or absence of IL-4, and then their tumoricidal activity was assayed by measuring 125I-UdR release from human melanoma (A375) cells. The spontaneous tumoricidal activity of AM was slightly suppressed by IL-4 in 3 of 7 donors. Addition of IL-4 to cultures of AM with the activators resulted in dose-dependent suppression of AM-mediated cytotoxicity against A375 cells. IL-4 also inhibited AM-mediated cytotoxicity against A375-R cells, which are resistant to interleukin 1 (IL-1) and tumor necrosis factor alpha, HT-29 colon cancer cells, and KB cells. IL-4 inhibited the early induction phase of AM activation. Pretreatment of AM with IL-4 also suppressed their expression of antitumor activity in response to lipopolysaccharide. IL-4 inhibited the production of monokines (IL-1 and tumor necrosis factor alpha) by AM at the protein and mRNA levels. These findings suggest that IL-4 may be important in vivo in the down-regulation of antitumor expression of AM in the lung by inhibiting the production of monokines and other killing mechanisms.     

Therapy of renal cell carcinoma with interleukin-2 and lymphokine-activated killer cells: phase II experience with a hybrid bolus and continuous infusion interleukin-2 regimen

Forty-seven patients with metastatic or unresectable renal cell carcinoma were treated with interleukin-2 (IL-2) and lymphokine-activated killer (LAK)-cell therapy, using a hybrid IL-2 regimen. IL-2 was administered initially by intravenous bolus (10(5) U/kg [Cetus Corp, Emeryville, CA] every 8 hours for 3 days) during the priming phase, and subsequently by continuous infusion (3 x 10(6) U/m2 for 6 days); during this second treatment period, in vitro-generated LAK cells were administered. Despite selection of patients for good performance status (PS) (29, PS 0; 18, PS 1) prior nephrectomy (43 of the 47 patients), and low tumor burden, the response rate was low (two complete [CRs] and two partial responses [PRs], for an overall objective response rate of 9%). Toxicity was comparable to that experienced with the high-dose bolus regimen. These results suggest that the dose and schedule of IL-2 administration may influence the likelihood of response to IL-2 in renal cell carcinoma.     

The role of autologous tumor cells in preventing lymphokine-activated killer cell induction in vitro

Peripheral blood lymphocytes (PBL), when cultured in vitro in the presence of autologous irradiated tumor and interleukin-2 (IL-2), become more restricted in the spectrum of their cytotoxicity. The cells continue to exhibit cytotoxicity for autologous tumor cells and major histocompatibility complex (MHC)-concordant allogeneic tumor cells of similar histologic type but not for the natural killer target cell line, K562. Furthermore, the addition of autologous tumor at different time points after the initiation with IL-2 alone of conventional lymphokine-activated killer cell cultures modifies both the specificity and the degree of cytotoxicity of these lymphocytes for tumor targets. By varying the culture conditions it may be possible to generate killer cells that will exhibit similarly enhanced and more restricted antitumor effects in vivo.     

The use of interleukin-2 and lymphokine-activated killer cells for the treatment of patients with non-Hodgkin's lymphoma.

PURPOSE: The study was undertaken to assess whether immunotherapy regimens with bolus high-dose interleukin-2 (IL-2) alone or with lymphokine-activated killer (LAK) cells are active in previously treated, relapsed patients with non-Hodgkin's lymphoma. PATIENTS AND METHODS: Nineteen patients with low- or intermediate-grade lymphomas were treated with bolus high-dose IL-2 alone (11 patients) or IL-2 with LAK cells (eight patients). IL-2 was administered by intravenous bolus infusion at 720,000 IU/kg every 8 hours. Eight patients had low-grade histologies; 11 patients were intermediate-grade. Eighteen patients had received second- or third-generation combination chemotherapy, and eight had also received radiation. All 19 relapsed after a median of two chemotherapy regimens. RESULTS: Four responses were observed, three partial and one complete, in patients with follicular histologies who received IL-2 with LAK cells. Response durations were 10, 16, 16, and 26 months, and three responders were re-treated after relapse with subsequent disease control for an additional 16, 39+, and 2+ months, respectively. CONCLUSION: High-dose, bolus IL-2-based immunotherapy with LAK cells may be an effective treatment for patients with non-Hodgkin's lymphoma and merits further testing with larger numbers of patients in phase II trials.     

Direct in vitro evidence and in vivo analysis of the antiangiogenesis effects of interleukin 12

As an antitumor agent, interleukin-12 (IL-12) has been revealed to be a key regulator of the immune response, particularly that involving CTL and natural killer (NK) cells. We report herein the antiangiogenesis effect of IL-12 on human as well as murine tumors in NK-depleted severe-combined immunodeficient mice using fibroblasts genetically engineered to secrete this cytokine. Although the in vitro growth of tumor cells was not affected by the presence of IL-12, coinoculation of IL-12-secreting fibroblasts strongly inhibited tumor growth in immunodeficient mice. The neovascularization surrounding the tumor was remarkably inhibited in the area in which the IL-12-secreting fibroblasts were implanted, resulting in the suppression of tumor growth. Lectin staining in tumor sample sections also showed a significant reduction in the number of vessels. The RNA expression of IFN-gamma and its inducible antiangiogenic chemokine IFN gamma-inducible protein 10 was stimulated in endothelial cells cultured with IL-12. It was also found that IL-12 down-regulated the expression of the endothelial cell mitogens vascular endothelial growth factor and basic fibroblast growth factor. The antitumor effects of IL-12 were accompanied by interesting histological changes consisting of a high degree of keratinization and apoptosis and a decrease in the proliferation rate of human tumors and extensive necrosis in the murine ones.