Adoptive immunotherapy of human cancer: the cytokine cascade and monocyte activation following high-dose interleukin 2 bolus treatment

Serum concentration kinetics of gamma-interferon (IFN-gamma), neopterin, 2'-5' A synthetase and tumor necrosis factor alpha were determined in five cancer patients undergoing adoptive immunotherapy with high-dose interleukin 2 (IL-2) bolus infusion and lymphokine-activated killer cells according to the National Cancer Institute, NIH protocol. In all cases a significant increase of these markers was observed after IL-2 treatment. This suggests that the antitumor effect of high-dose IL-2 bolus administration may be in part mediated by activation of a cascade of endogenous cytokines including IFN-gamma and tumor necrosis factor alpha. After IL-2 bolus injection, the kinetics of neopterin was similar but delayed when compared to that of IFN-gamma: this suggests that macrophages, the specific source of neopterin, become activated by IFN-gamma following IL-2-mediated lymphocyte induction, thus implying a possible role for macrophages in the antitumor effects mediated by IL-2 and lymphokine-activated killer cells.     

Cytokine regulation of tumor necrosis factor-alpha and -beta (lymphotoxin)-messenger RNA expression in human peripheral blood mononuclear cells.

The immune response at the molecular level is characterized by a carefully coordinated interplay of both cytokine production and receptor induction. The regulation of these molecules including the closely related tumor necrosis factors alpha (TNF) and beta (lymphotoxin, LT) is still incompletely understood. We have examined the effects of various cytokines on the expression of TNF and LT mRNA in human peripheral blood mononuclear cells (PBMC). Northern blot analysis with total cellular RNA from mixed populations of PBMC revealed that genes coding for TNF and LT were not spontaneously expressed. Treatment of PBMC with recombinant interleukin (IL)-2 resulted in a high level expression of TNF and LT mRNA. Whereas IL-1 beta was equally effective as IL-2 in inducing both TNF and LT mRNA, granulocyte-macrophage colony-stimulating factor selectively induced only TNF mRNA. Both TNF and LT mRNA were minimally induced by IL-1 alpha, IL-3, interferon (IFN)-alpha, or IFN-gamma. Similarly TNF alone had little effect on induction of TNF and LT mRNA. In conjunction with IL-2, cytokines such as IFN-alpha, IFN-gamma, or TNF did not interfere with IL-2 induction of TNF and LT mRNA. Interestingly, IL-4 in combination with IL-2 inhibited the IL-2-driven induction of TNF and LT mRNA. This inhibitory effect of IL-4 was also observed at the level of TNF and LT protein secretion. Furthermore, IL-4 was also inhibitory of IL-2-mediated induction of Tac mRNA in PBMC. These results extend the interrelationship of cytokine regulation of TNF and LT expression. In particular, they reveal the previously unrecognized function of IL-4 in antagonizing the IL-2 induction of TNF, LT, and Tac mRNA in PBMC.     

Unexpected cytokines in serum of malignant melanoma patients during sequential biochemotherapy.

Biochemotherapy, which combines traditional chemotherapy with immune modulating biologicals, produces an unexpectedly high response rate (>50%) in advanced melanoma patients. We hypothesize that immunological mechanism(s) are responsible for the increased response rate, and particularly that macrophage activation is involved in tumor reduction. Patients were randomized to receive chemotherapy, composed of cisplatin, vinblastine, and dacarbazine (CVD), or biochemotherapy, which is CVD followed by interleukin (IL)-2 and IFN-alpha2b (CVD-BIO). Laboratory analysis was performed on sera from 41 patients from each arm. Measurements of macrophage activation (neopterin), nitric oxide production (nitrite), and tumor necrosis factor-alpha (TNF-alpha), IL-1alpha, IL-1beta, IFN-gamma, IL-6, IL-10, and soluble IL-2 receptor (sIL-2R) were performed. Six of the nine biological responses (nitrite, neopterin, IFN-gamma, IL-6, soluble IL-2R, and IL-10) significantly (P < 0.0002) increased in the CVD-BIO patients but not in the CVD patients. The increased IL-6 (P = 0.04) and IL-10 (P = 0.05) correlated with patient response, but only when the minor responders were included in the analysis. Evidence of macrophage activation was found in CVD-BIO patients and not in those receiving CVD alone. In addition, an unusual cytokine elaboration composed of IL-6, IFN-gamma, IL-10, nitrite, neopterin, and sIL-2R, but not the expected TNF-alpha and IL-1, was detected. A trend of higher increase in IL-6 and IL-10 in patients having clinical response was found, suggesting an incomplete Th2 pattern of cytokine elaboration. These data show that macrophage activation does not appear to be critical in the response to CVD-BIO, but that IL-10 and IL-6 induced by the BIO component of the CVD-BIO were associated with tumor regression, and that their biology should be pursued further in the analysis of mechanism(s) of response.     

The relationship between circulating concentrations of C-reactive protein, inflammatory cytokines and cytokine receptors in patients with non-small-cell lung cancer

The relationship between circulating C-reactive protein concentrations and potential cytokine and receptor mediators (interleukin-6, leukaemia inhibitory factor (LIF), ciliary neurotrophic factor (CNTF), soluble IL-6 receptor, soluble gp130, soluble TNF receptor, interleukin-1 receptor antagonist and interleukin-8 (IL-8)) of this acute phase protein were examined in healthy subjects (n=11) and patients with non-small-cell lung cancer (n=50). Leukaemia inhibitory factor and CNTF were below detection limits in all controls and patients. C-reactive protein, interleukin-6, soluble gp130, soluble TNF receptor, interleukin-1 receptor antagonist and IL-8 concentrations were significantly elevated in cancer patients (P< or =0.001). Cancer patients with elevated C-reactive protein concentrations had greater concentrations of interleukin-6 (P<0.01) and interleukin-1 receptor antagonist (P<0.05). On regression analysis only interleukin-6 was independently associated with C-reactive protein (r=0.616, P<0.001). Interleukin-6 is an important independent mediator of elevated C-reactive protein concentrations in patients with non-small-cell lung cancer.     

Cytokine serum levels in soft tissue sarcoma patients: correlations with clinico-pathological features and prognosis

We investigated the correlations between serum levels of selected proinflammatory, hematopoietic and angiogenic cytokines and soluble cytokine receptors with the clinico-pathological features and prognosis in soft tissue sarcoma patients. Serum levels of 9 cytokines (TNFalpha, IL-1ra, IL-6, IL-8, IL-10, M-CSF, G-CSF, VEGF, bFGF) and 4 free cytokine receptors (sIL-2R alpha, sIL-6R, TNFRI, TNFRII) were measured by means of an enzyme-linked immunoadsorbent assay kit in 156 soft tissue sarcoma patients before the treatment and in 50 healthy controls. Serum levels of 10 cytokines and cytokine receptors were also assayed during patients' follow-up after the treatment. Significantly elevated pretreatment serum levels of 11/13 cytokines and cytokine receptors were found in sarcoma patients, as compared to healthy controls. In 40.4% of patients 6 or more cytokines and cytokine receptors (most frequently: TNF RI, IL-6, IL-8) were elevated in parallel. Serum levels of IL-6, sIL-2R, VEGF, M-CSF and TNF RI correlated significantly with tumor size and serum levels of IL-8 and IL-6 were significantly higher in patients with Grade 2/3 vs. Grade 1 tumors. We did not observe any significant differences in cytokine serum levels between patients with primary and recurrent tumors and patients with and without distant metastases. Using univariate analysis, overall survival (OS) in all patients was affected by tumor size (<5 cm vs. 5-10 cm vs. >10 cm), tumor grade (G1 vs. G2/3), presence of metastases, pretreatment serum levels of 8 cytokines (IL-6, IL-8, IL-10, sIL-2R, TNF RI, TNF RII, M-CSF, VEGF) and the number of cytokines increased (0-1 vs. 2-5 vs. < or = 6). Elevated serum levels of IL-6, IL-8, IL-10 and sIL-2R alpha, high tumor grade and larger tumor size strongly correlated with shorter disease-free survival (DFS). Multivariate analysis identified G2/3 tumor grade (p = 0.001), the presence of metastases (p = 0.004), elevated IL-6 serum level (p = 0.02), elevated IL-8 serum level (p = 0.048) and the number of cytokine serum levels above upper cut-off values (p = 0.01) as the independent prognostic factors related to OS, and G2/3 tumor grade (p = 0.005) and increased IL-6 serum level (p = 0.035) as independent prognostic factors related to DFS. In a group of patients without metastases (M0) higher tumor grade, elevated serum level of IL-6 and TNF RII, and the number of elevated cytokine serum levels correlated independently with poor survival. We found a significant decrease of several cytokine serum levels in patients after treatment (IL-1ra, IL-6, IL-8, IL-10, TNF RII, M-CSF) [p < 0.05]. Persistently elevated serum level of IL-6 after the treatment has also shown negative prognostic significance for OS (univariate analysis). Serum levels of some proinflammatory, hematopoietic and angiogenic cytokines and cytokine receptors are elevated, frequently in parallel, in a large percentage of soft tissue sarcoma patients. Significant correlations of serum cytokine levels with tumor size and grade suggest that some of these cytokines may be directly or indirectly involved in the progression of soft tissue sarcomas. Serum assays of IL-6, IL-8 and TNF RII before or after the treatment may be useful in establishing soft tissue sarcoma patients prognosis.     

Macrophages inhibit human osteosarcoma cell growth after activation with the bacterial cell wall derivative liposomal muramyl tripeptide in combination with interferon-γ

Background

In osteosarcoma, the presence of tumor-infiltrating macrophages positively correlates with patient survival in contrast to the negative effect of tumor-associated macrophages in patients with other tumors. Liposome-encapsulated muramyl tripeptide (L-MTP-PE) has been introduced in the treatment of osteosarcoma patients, which may enhance the potential anti-tumor activity of macrophages. Direct anti-tumor activity of human macrophages against human osteosarcoma cells has not been described so far. Hence, we assessed osteosarcoma cell growth after co-culture with human macrophages.

Methods

Monocyte-derived M1-like and M2-like macrophages were polarized with LPS + IFN-γ, L-MTP-PE +/− IFN-γ or IL-10 and incubated with osteosarcoma cells. Two days later, viable tumor cell numbers were analyzed. Antibody-dependent effects were investigated using the therapeutic anti-EGFR antibody cetuximab.

Results

M1-like macrophages inhibited osteosarcoma cell growth when activated with LPS + IFN-γ. Likewise, stimulation of M1-like macrophages with liposomal muramyl tripeptide (L-MTP-PE) inhibited tumor growth, but only when combined with IFN-γ. Addition of the tumor-reactive anti-EGFR antibody cetuximab did not further improve the anti-tumor activity of activated M1-like macrophages. The inhibition was mediated by supernatants of activated M1-like macrophages, containing TNF-α and IL-1β. However, specific blockage of these cytokines, nitric oxide or reactive oxygen species did not inhibit the anti-tumor effect, suggesting the involvement of other soluble factors released upon macrophage activation. While LPS + IFN-γ–activated M2-like macrophages had low anti-tumor activity, IL-10–polarized M2-like macrophages were able to reduce osteosarcoma cell growth in the presence of the anti-EGFR cetuximab involving antibody-dependent tumor cell phagocytosis.

Conclusion

This study demonstrates that human macrophages can be induced to exert direct anti-tumor activity against osteosarcoma cells. Our observation that the induction of macrophage anti-tumor activity by L-MTP-PE required IFN-γ may be of relevance for the optimization of L-MTP-PE therapy in osteosarcoma patients.     

Transient increase of plasma interleukin-6 after infusion of recombinant tumor necrosis factor alpha in advanced cancer patients

Tumor Necrosis Factor alpha administered in the therapy of advanced cancer influences certain hormones and cytokines secretion. In turn, these also modulate the biological activity of Tumor Necrosis Factor alpha. It has been shown in several studies that the cytokine Interleukin -6 (IL)-6 is produced in response to various hormones and other cytokines eg. Tumor Necrosis Factor (TNF alpha). In our study we focused on the Interleukin-6 (IL-6) secretion in response to TNF alpha administration in 12 patients undergoing TNF alpha biotherapy due to advanced neoplastic disease. Plasma IL-6 was estimated prior and at various time points (2, 3, 5 a,d 12 hours) after TNF alpha intravenous infusion. IL-6 level was estimated with ELISA method. In conclusion, we suggest the stimulating influence of hrec TNF alpha administered as therapy for advanced cancer on IL-6 secretion.     

Preoperative and postoperative cytokines in patients with cancer.

BACKGROUND. Changes in tumor necrosis factor-alpha (TNF-alpha), interleukin-1 alpha (IL-1 alpha), interleukin-1 beta (IL-1 beta), interleukin-6 (IL-6), and granulocyte macrophage-colony-stimulating factor (GM-CSF) were investigated before and after operation in patients with hepatocellular carcinoma (HCC), metastatic liver carcinoma, and gastrointestinal carcinoma. RESULTS. Serum levels of TNF-alpha, IL-1 alpha, and IL-1 beta were high in patients with liver carcinoma (HCC and metastatic liver carcinoma) before operation in comparison with those of normal controls (P less than 0.01). In patients with gastrointestinal carcinoma, serum levels of cytokines, except those of TNF-alpha, were the same as in patients with liver carcinoma. The level of TNF-alpha in patients with gastrointestinal carcinoma was low compared with that in patients with liver carcinoma. Within 1 day after operation, the peak level of TNF-alpha was observed at 15 hours, IL-1 alpha at 18 hours, IL-1 beta at 21 hours, and IL-6 at 24 hours after operation. Subsequently, these cytokine levels peaked again: TNF-alpha at 48 hours, IL-1 alpha at 72 hours, IL-1 beta at 120 hours, and IL-6 at 168 hours after operation. GM-CSF levels increased gradually after operation. Moreover, in HCC, serum levels of TNF-alpha were high in patients with recurrence compared with those without recurrence (P less than 0.01). The difference in IL-1 alpha and IL-1 beta levels between patients with recurrence and those without recurrence can be regarded as significant (P less than 0.01). CONCLUSION. These results suggest that TNF-alpha, IL-1, IL-6, and GM-CSF may play an important role in the pathogenesis of cancers; TNF-alpha may be especially important as a tumor marker in HCC and metastatic liver carcinoma.     

In vitro and in vivo enhancement of canine pulmonary alveolar macrophage cytotoxic activity against canine osteosarcoma cells

The combination of chemotherapy with immunotherapy may offer an advantage over either therapy alone and provide a greater potential for total tumor eradication. Monocyte/macrophage-mediated tumor cell killing is a major mechanism of the host's defense against primary and/or metastatic neoplasia. We evaluated the tumoricidal activity against canine osteosarcoma cells of canine pulmonary alveolar macrophages (PAM) exposed in vitro to two recombinant canine (rc) cytokines (rcTNF alpha and rcIFN gamma). We also evaluated the in vivo tumoricidal activity of PAM from dogs treated with the macrophage activator, liposome-encapsulated muramyl tripeptide-phosphatidyl-ethanolamine (L-MTP-PE) alone or in combination with doxorubicin (DOX). This study demonstrated that rcTNF alpha and rcIFN gamma significantly enhance in vitro canine PAM cytotoxicity against canine osteosarcoma cells, and that PAM from dogs treated with DOX + L-MTP-PE have enhanced cytotoxic activity against osteosarcoma cells when compared to dogs treated with DOX or L-MTP-PE alone. These findings support the rationale for combining a chemotherapy agent with an immunotherapy agent for the treatment of metastatic disease, and suggest a role for TNF alpha and IFN gamma as agents for stimulating the antitumor activity of macrophages.     

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.     

Inflammatory biomarkers,geriatric assessment,and treatment outcomes in acute myeloid leukemia

ObjectivesTo investigate changes in inflammatory biomarkers during induction therapy for older adults with acute myeloid leukemia (AML) and their associations with geriatric assessment (GA) measures and outcomes.MethodsThis was a single institution ancillary study to a prospective observational study (N = 20 consecutive adults aged ≥60 with newly diagnosed AML who received induction chemotherapy). Biomarkers (Interleukin-6 [IL-6], IL-6 soluble receptor [IL-6 sR], tumor necrosis factor alpha [TNFα], TNFα soluble receptor 1 [TNFα sR1], interleukin-3 [IL-3], C-reactive protein [CRP]) were collected at start of induction, weekly for three weeks, and post-induction and were compared over time using paired t-tests. GA was administered at baseline and post-induction, and correlated with biomarker levels using Spearman correlations. Survival was estimated using Kaplan-Meier and compared by categorized biomarker level using Wilcoxon tests.ResultsBiomarker levels were stable during induction, except for CRP and IL-6 sR. Declines in objectively measured physical function [Short Physical Performance Battery (SPPB); r = 0.71, p < 0.01] and increases in self-reported limitation in instrumental activities of daily living (r = 0.81, p < 0.01) were correlated with increased TNFα sR1. Declines in SPPB were correlated with increased CRP (r = −0.73, p < 0.01). Improvement in depression was correlated with increased IL-6 sR (r = −0.59 p = 0.02). Survival was shorter in those with baseline TNFα or CRP levels above the median (6.1 vs. 40.2 months and 5.5 vs. 27.6 months respectively, p = 0.04 for both).ConclusionAmong older adults with AML, the relationships between TNFα sR1, CRP, and IL-6 sR with change in physical and emotional health during treatment warrants further investigation.     

Analysis of changes in serum levels of TNF, IL-1 and IL-6 during administration of IL-2. Correlation with clinical response to treatment]

We have investigated the serum concentrations of TNF, IL-1 and IL-6 in 49 patients with metastatic renal carcinoma receiving interleukin 2 (IL-2) or a combination of IL-2 and interferon alpha (IFN). Our results demonstrate that IL-2 and/or IFN induce an increase of serum concentrations of IL-1 and TNF in 95% and 75% of the patients respectively. Serum IL-6 levels increase in 44% of the patients. Serum concentrations of IL-1 and TNF remain elevated 48 hours after the end of IL-2 infusion. IL-1 and TNF levels are higher in patients receiving a combination of IL-2 and IFN. TNF and IL-1 levels in serum are significantly higher in responders to IL-2 treatment 48 hours after the end of IL-2 infusion. These two biological criteria enable a subgroup of patients with a very low response rate to IL-2 to be defined. The persistent increase of these cytokines in serum indicates a persistent activation of the immune system lasting after the end of IL-2 treatment which could be involved in the antitumor response.     

Clinical significance of serum cytokine measurements in untreated colorectal cancer patients: soluble tumor necrosis factor receptor type I--an independent prognostic factor.

The aim of this study was to exploit the potential clinical use of circulating cytokine measurements in colorectal cancer (CRC) patients. The levels of cytokines and cytokine receptors were assessed by ELISA in the sera of 50 healthy volunteers and 157 patients with previously untreated CRC and then related to clinicopathological features and prognosis. All tumors were verified histologically as colorectal adenocarcinomas and staged according to TNM classification. The levels of circulating interleukin (IL)-6, IL-8, macrophage colony-stimulating factor (M-CSF) and interleukin 1 receptor antagonist (IL-1ra) significantly increased with the clinical stage of CRC, and the levels of IL-6, soluble tumor necrosis factor (sTNF) receptor type I (RI), soluble interleukin 2 receptor alpha and TNFalpha with tumor grade, while IL-6, IL-8, M-CSF, IL-1ra and sTNF RI levels significantly rose with bowel wall invasion. None of the cytokine or soluble cytokine receptor levels were influenced by age, gender and colon versus rectum localization. sTNF RI, IL-8, IL-6 and vascular endothelial growth factor measurements demonstrated the highest diagnostic sensitivity. sTNF RI was found elevated in the greatest percentage of all CRC patients, in the greatest proportion of stage I patients and presented the best diagnostic sensitivity. In addition, the sTNF RI level strongly correlated with tumor grade and invasion and proved to be an independent prognostic factor.     

Immune changes in patients with advanced breast cancer undergoing chemotherapy with taxanes

Besides cytotoxicity, taxanes induce other biological effects, especially in the immune system. Taxanes have demonstrated immunostimulatory effects against neoplasms, supporting the idea that these agents suppress cancer through several mechanisms and not solely through inhibiting cell division. The purpose of the present study was to evaluate the effect of taxanes (paclitaxel and docetaxel) and investigate their ability in alterating important immunological parameters in breast cancer patients. Thirty women with advanced breast cancer undergoing chemotherapy were randomly assigned into two groups treated with either single agent Paclitaxel or Docetaxel. Sera from patients before the first and after the last treatment cycle and from normal donors were assayed by ELISA for IL-2, IL-1beta, IFN-gamma, GM-CSF, IL-6, TNF-alpha, and PGE2 levels. In these same blood samples, NK and LAK cell activity was tested in the total PBMC population against NK-sensitive K562 tumour targets, respectively, and autologous mixed lymphocyte reaction was tested by (3)H-thymidine proliferation assays. All patients in both groups responded to therapy. Significant differences were observed in the following immune parameters between the control group of healthy blood donors and the pretreatment values of both taxane groups; IL-2, GM-CSF, IFN-gamma levels and NK and LAK cell cytotoxicity were depressed, whereas TNF-alpha and IL-6 levels were raised in breast cancer patients before treatment compared to controls. There were no significant differences between the two treatment groups regarding any of the parameters studied. Both drugs led to increases in MLR values, NK and LAK cell cytotoxicity, and IL-6, GM-CSF, IFN-gamma levels, and decreases for IL-1, TNF, and PGE2 levels. The percentage of these differences was greater for docetaxel in comparison to paclitaxel (P<0.0001). More specifically, docetaxel demonstrated a more pronounced effect on enhancing MLR, NK, LAK activity and IFN-gamma, IL-2, IL-6, and GM-CSF levels, as well as caused more potent reduction in IL-1 and TNF-alpha levels when compared to paclitaxel. The present study indicates that patients responded to treatment of advanced breast cancer with single-agent paclitaxel or docetaxel leads to an increase in serum IFN-gamma, IL-2, IL-6, GM-CSF cytokine levels and enhancement of PBMC NK and LAK cell activity, while they both lead to a decrease of acute phase serum cytokine levels of IL-1 and TNF-alpha. Moreover, the effects of docetaxel are in all the above parameters more pronounced than those of paclitaxel.     

Interleukin-6 and its relationships to acute phase proteins in serous effusion differentiation

The aim of the study was to assess the discriminative power of cytokine interleukin 6 (IL-6) between transudative and exudative pleural and peritoneal effusions, and to compare IL-6 with common acute phase proteins in serous effusion differentiation. One hundred and forty-five consecutive patients with pleural or peritoneal effusion underwent diagnostic parecentesis. Patients were categorized in three groups. Malignant effusion (group A) 56 patients, non-malignant effusion (group B) 46 patients and transudate (group C) 43 patients. Serum and effusion levels of IL-6, C-reactive protein (CRP), alpha2-macroglobuline (alpha2-MG), alpha1-antitrypsin (alpha1-AT) and alpha1-acid glycoprotein (alpha1-AG) were determined. Serum IL-6 levels were significantly higher in groups A and B in comparison to group C (p<0.001 and 0.001, respectively). In addition, serum IL-6 levels were higher in group A compared to group B (p<0.001), while the studied acute phase proteins were not significantly different. All the studied parameters were higher in the effusions of groups A and B compared to group C. At a cut-off value of 72.1 fmol/ml IL-6 had a sensitivity of 82.6-89.3%, specificity of 88.4-90.7% and positive predictive value of 90.7-94.6% among the three groups. Our results suggest that IL-6 at levels > or =72.1 fmol/ml, alpha1-AT at > or =170 mg/dl and alpha1-AG at > or =52.3 mg/ml give strong evidences for malignancy in exudates.     

Early variations of circulating interleukin-6 and interleukin-10 levels during thoracic radiotherapy are predictive for radiation pneumonitis.

PURPOSE: To investigate variations of circulating serum levels of interleukin-6 (IL-6), tumor necrosis factor alpha (TNFalpha), and interleukin-10 (IL-10) during three-dimensional conformal radiation therapy (3D-CRT) in patients with non-small-cell lung cancer and correlate these variations with the occurrence of radiation pneumonitis. PATIENTS AND METHODS: Ninety-six patients receiving 3D-CRT for stage I to III disease were evaluated prospectively. Circulating cytokine levels were determined before, every 2 weeks during, and at the end of treatment. Radiation pneumonitis was evaluated prospectively between 6 and 8 weeks after 3D-CRT. The predictive value of clinical, dosimetric, and biologic (cytokine levels) factors was evaluated both in univariate and multivariate analyses. RESULTS: Forty patients (44%) experienced score 1 or more radiation pneumonitis. No association was found between baseline cytokine levels and the risk of radiation pneumonitis. In the whole population, mean levels of TNFalpha, IL-6, and IL-10 remained stable during radiotherapy. IL-6 levels were significantly higher (P = .047) during 3D-CRT in patients with radiation pneumonitis. In the multivariate analysis, covariations of IL-6 and IL-10 levels during the first 2 weeks of 3D-CRT were evidenced as independently predictive of radiation pneumonitis in this series (P = .011). CONCLUSION: Early variations of circulating IL-6 and IL-10 levels during 3D-CRT are significantly associated with the risk of radiation pneumonitis. Variations of circulating IL-6 and IL-10 levels during 3D-CRT may serve as independent predictive factors for this complication.     

Serum cytokine levels in response to hepatic cryoablation

BACKGROUND AND OBJECTIVES: Cryogenic treatment sometimes stimulates the immune system by releasing intracellular antigens. We evaluated anti-tumor immune response by analyzing alterations in serum cytokine levels. METHODS: Percutaneous cryosurgery was performed in 13 patients with unresectable tumors. Serum levels of interleukin (IL) -4, -6, and -10, tumor necrosis factor (TNF)-alpha, and interferon (IFN)-gamma were measured by enzyme-linked immunosorbent assay (ELISA). The Th1/Th2 ratio was estimated from the IFN-gamma/IL-4 ratio. RESULTS: Levels of serum factors in the immune reaction (IR) group, in which tumor necrosis was identified not only in the treated area but also away from the treated area, were compared with those in the local effect (LE) group. Serum amyloid A (SAA), C-reactive protein (CRP), and IL-6 levels were increased in both groups after three treatments. The serum IL-10 level tended to increase with the number of treatments. Pretreatment IL-10 levels in the LE group were significantly greater than those in the IR group, and the maximum value in the LE group (59.5 +/- 13.2 pg/ml) was greater than that in the IR group (47.0 +/- 15.0 pg/ml). The TNF-alpha level was increased in the IR group. Pretreatment TNF-alpha levels and maximum levels in response to treatment were significantly greater in the IR group than in the LE group (P = 0.0313). The Th1/Th2 ratio was increased in the IR group, and the maximum ratio was significantly greater in the IR group than in the LE group. CONCLUSION: It might be possible to evaluate the appearance of immune responses to cryosurgery by monitoring serum cytokine levels.     

Down-regulation of IL-18 receptor in cancer patients: its clinical significance

Interleukin-18 (IL-18) is a powerful inducer of interferon-gamma (IFN-gamma), a key immunoregulatory cytokine. Cellular immune responsiveness, as measured by IL-18-induced IFN-gamma production from peripheral blood mononuclear cells (PBMCs) in ELISA assay, was evaluated in 10 patients with advanced cancer and in 10 normal controls. Supernatant levels of IFN-gamma were detected at 2 hours after PBMCs culture and markedly increased thereafter in healthy volunteers. In contrast, IFN-gamma production in cancer patients was not detected during the culture period (0-72 hours). We also measured IL-18-stimulated IL-12 production in healthy volunteers and null response was observed in cancer-bearing patients. Next, we studied mRNA expressions of IL-18 receptor (IL-18R) and IFN-gamma in PBMCs in cancer patients and healthy volunteers by RT-PCR assay. Both mRNA levels of IL-18R and IFN-gamma were significantly decreased in cancer-bearing patients compared with normal controls. These results suggested that IL-18 responsiveness for IFN-gamma production in cancer-bearing patients was impaired. Using flow cytometric analysis, we studied T-cell subsets, CD3- CD56+ (NK cell), CD3+ CD45RO+ (memory T-cell), CD3+ CD95+ (Fas+ T-cell), CD3+ CD4+ (helper T-cell), CD3+ CD8+ (cytotoxic T-cell: CTL) and CD3+ V alpha24+ (NKT-cell), in cancer patients and normal controls. The NK and cytotoxic T-cells significantly decreased and NKT-cells had decreased tendency in cancer patients compared with normal controls. In contrast, memory T cells, Fas+ T-cells and helper T-cells were all significantly increased in cancer patients compared with normal controls. These results suggested that the underlying mechanism of impaired IL-18 responsiveness in PBMCs from cancer-bearing patients was, at least in part, ascribed to a drastic decrease of NK cells and CTL which constitutively and highly express IL-18R and also attributed to null production of IL-12 which up-regulates IL-18R.