The Influence of IFN-α on Blood Plasmacytoid Dendritic Cell in Chronic Myeloid Leukaemia

OBJECTIVE To study the mechanism of IFN on CML.METHODS Samples of 15 CML patients and 10 healthy controls were studied. The flow cytometry was performed to identify circulating pDCs. The concentration of IFN-α in serum and that in the supematant of peripheral blood mononuclear cells (PBMCs)cultured after stimulation with CpG ODN2216 were examined both in CML patients and in the healthy controls RESULTS There was significant reduction in the number of circulating pDCs, serum concentration of IFN-α and the capacity of IFN-α producing PBMCs in CML patients compared with those in healthy control individuals (P < 0.001). After the active treatment with IFN-α and hydroxyurea, the quantity and function of pDCs were increased in stabilized patients, especially the function of pDCs in 2 patients achieving major cytogenetic.response (MCR). The proportion and function of pDCs and the serum levels of IFN were inversely correlated with both WBC and age of the patients with CML, and positively correlated with the state of the illness.dysfunction of circulating pDCs. The active treatment with IFN in CML patients may be related to the restoration of pDCs.     

Immunomodulation of natural killer activity in children with acute lymphoblastic leukaemia

Modulation of NK activity of PBLs from ALL patients was studied following exposure to IFN-α, staphylococcal protein A and interleukin-2. Only 52% of ALL patients responded to IFN-α stimulation, where the majority of controls showed positive enhancement of NK activity. Protein A failed to cause a significant stimulation of ALL patient PBLs whereas all controls showed a positive response. The majority of ALL patient and child control PBLs were however able to produce significant levels of IFN-γ (protein A stimulation) and IFN-α (Sendai virus stimulation), although significantly more of both types of interferon could be induced in adult PBL samples. The ability of IL-2 to activate NK activity of ALL PBL samples showed a similar trend to IFN-α stimulation; thus, not all ALL patients showed positive augmentation of Nk activity upon IL-2 stimulation. It is clear from these results that interferons and IL-2 may not necessarily lead to activated NK cytolytic activity, and in the present study approx. 50% of ALL patients failed to respond to lymphokine stimulation.     

Studies with interferon-α in human tumors

Clinical trials with IFN-α, produced from buffy coats of leukocytes, have been going on at Radiumhemmet, Karolinska Hospital since 1969. In some of the benign and malignant tumor diseases studied, IFN has been shown to induce regressions. In the case of the benign tumor juvenile laryngeal papillomatosis, IFN has been shown to be superior to other treatment modalities. So far IFN has not been shown to be superior to conventional treatment in any of the malignant tumors studied.     

Therapeutic effects of monoclonal antibody g250, interferons and tumor necrosis factor,in mice with renal-cell carcinoma xenografts

Because renal-cell carcinoma (RCC) is considered relatively resistant to radio-and chemotherapy, RCC patients may benefit from new treatment modalities, e.g. immunotherapy. In vitro and in vivo studies suggest that combinations of cytokines such as interferon γ or interferon a (IFN-γ, IFN-α) and tumor necrosis factor a (TNF-α) may act synergistically. In this study we tested whether a monoclonal antibody (MAb) G250, reactive with a RCC-associated antigen, showed anti-tumor effects in vivoin nude mice with established s.c. human RCC xenografts, and also whether this MAb could enhance the anti-tumor effect of combinations of IFNs and TNF-α. Treatment with combinations of IFN-α/TNF-α or IFN-γ/TNF-α, or with MAb G250 alone, resulted in a significant inhibition of tumor growth. Treatment with MAb G250, in combination with IFN-γ/TNF-α, did not result in an improve anti-tumor effect as compared to that of either treatment alone. In contrast, MAb G250 combined with IFN-α/TNF-α resulted in a significantly enhanced anti-tumor response. In one experiment, 3 out of 10 mice showed complete tumor regression, with no recurrence after 90 days. Large numbers of infiltrating macrophages were found surrounding viable and necrotic tumor tissue after treatment with G250 combined with IFN-α/TNF-α. These results suggest that combination therapy, consisting of IFN-α, TNF-α and MAbs, may have therapeutic value in the treatment of RCC.     

Normal breast epithelial cells produce interleukins 6 and 8 together with tumor-necrosis factor: Defective il6 expression in mammary carcinoma

Virtually pure primary cultures of normal mammary epithelial cells (MEC) obtained from healthy women were shown to release interleukin 6 and 8 (IL6, IL8) and to produce a non-secreted form of tumor-necrosis factor (TNF). No interferon (IFN), whether α, β or γ, or ILI-α or -β could be detected. Analysis of cellular RNA confirmed these findings and showed that MEC also express IL6 receptor and TNF-α-related mRNAs. Epithelial cells were selectively stained by antibodies to IL6, IL8 and TNF-α both in primary cultures and in the normal mammary gland. Samples of human milk contained sizable amounts of IL6, IL8 and IFN-γ; yet the liquid phase was consistently negative for other cytokines (i.e., TNF-α, IFN-α/-β, ILI-α/-β). Expression of IL6 (but not of IL8 and TNF-α) was abolished in ductal infiltrating carcinomas and greatly reduced in cultures of oncogene-transfected mammary cells, suggesting that alterations of IL6 expression are associated with pathogenesis in breast cancer.     

Interferon and chronic myelogenous leukaemia

Interferon (IFN) is widely employed in the therapy of chronic myelogenous leukaemia because of its ability to exert the antiproliferative activity on leukaemic haematopoietic progenitors and for the expression for IFN-α receptors by peripheral blood leukaemic cell surfaces. There is no difference between recombinant IFN α 2b and α 2a regarding their efficacy in the treatment of Ph-positive CML patients. Either no randomized studies or the randomized ones show a superior effectiveness of IFN given as single agent in the induction treatment to that one of chemotherapy regarding the complete cytogenic response percentage. The ability of IFN-γ to induce the expression of adhesion molecules such LFA 1 and ICAM 1 on peripheral blood leukaemic cell surfaces may suggest its use in the induction therapy of CML patients. Other than, a superior effectiveness of combined therapy including interferon and chemotherapy agents compared to chemotherapy alone has also been found. Finally no large series of trials to study the IFN efficacy both as second line treatment and maintenance therapy have been carried out.     

Synergistic antitumor effect of interferon-ß with gemcitabine in interferon-α-non-responsive pancreatic cancer cells

Interferon (IFN)-? is reported to have more potent antitumor effects than IFN-α. The aim of this study was to compare the synergistic antitumor activity of both IFNs when combined with gemcitabine on cultured pancreatic cancer cells expressing various levels of IFN receptor. The growth-inhibitory effects of IFN-α and IFN-? in combination with gemcitabine on three human pancreatic cancer cell lines (BxPC-3, MIAPaCa-2, Panc-1) were evaluated by MTT assay and isobolographic analysis. We also correlated their growth-inhibitory effects with the expression status of type I IFN receptor type 2 (IFNAR2). Western blot analysis indicated strong expression of IFNAR2 in BxPC-3 and MIAPaCa-2, but weak expression in Panc-1. The growth-inhibitory effect of gemcitabine was enhanced synergistically by IFN-α in BxPC-3 and MIAPaCa-2, but not in Panc-1. IFN-? exhibited more potent synergistic growth-inhibitory effects with gemcitabine in BxPC-3 and MIAPaCa-2 compared to IFN-α, and also synergistic enhancement in Panc-1. In conclusion, our results indicated that the growth-inhibitory effect of IFN-? with gemcitabine was synergistic not only in pancreatic cancer cells with strong expression of IFNAR2, but also in those with weak expression of IFNAR2.     

Combined immunotherapy encompassing intratumoral poly-ICLC,dendritic-cell vaccination and radiotherapy in advanced cancer patients

BackgroundCombination immunotherapy has the potential to achieve additive or synergistic effects. Combined local injections of dsRNA analogues (mimicking viral RNA) and repeated vaccinations with tumor-lysate loaded dendritic cells shows efficacy against colon cancer mouse models. In the context of immunotherapy, radiotherapy can exert beneficial abscopal effects.Patients and methodsIn this two-cohort pilot phase I study, 15 advanced cancer patients received two 4-week cycles of four intradermal daily doses of monocyte-derived dendritic cells preloaded with autologous tumor lysate and matured for 24 h with poly-ICLC (Hiltonol), TNF-α and IFN-α. On days +8 and +10 of each cycle, patients received intratumoral image-guided 0.25 mg injections of the dsRNA-analogue Hiltonol. Cyclophosphamide 600 mg/m² was administered 1 week before. Six patients received stereotactic ablative radiotherapy (SABR) on selected tumor lesions, including those injected with Hiltonol. Expression of 25 immune-relevant genes was sequentially monitored by RT-PCR on circulating peripheral blood mononuclear cell (PBMCs) and serum concentrations of a cytokine panel were sequentially determined before and during treatment. Pre- and post-treatment PBMC from patients achieving durable stable disease (SD) were studied by IFNγ ELISPOT-assays responding to tumor-lysate loaded DC and by TCRβ sequencing.ResultsCombined treatment was, safe and well tolerated. One heavily pretreated castration-resistant prostate cancer patient experienced a remarkable mixed abscopal response to SABR+ immunotherapy. No objective responses were observed, while nine patients presented SD (five of them in the six-patient radiotherapy cohort). Intratumoral Hiltonol increased IFN-β and IFN-α mRNA in circulating PBMC. DC vaccination increased serum IL-12 and IL-1β concentrations, especially in patients presenting SD. IFNγ-ELISPOT reactivity to tumor lysates was observed in two patients experiencing durable SD.ConclusionsThis radio-immunotherapy combination strategy, aimed at resembling viral infection in tumor tissue in combination with a dendritic-cell vaccine and SABR, is safe and shows immune-associated activity and signs of preliminary clinical efficacy.     

A long-term survival case of hepatocellular carcinoma with portal vein and inferior vena cava tumor thrombi

We report a case of advanced hepatocellular carcinoma (HCC) successfully treated by hepatic arterial infusion of 5-fluorouracil (5-FU) combined with systemic administration of interferon (IFN)-α and trans-arterial infusion (TAI) therapy of cisplatin (CDDP). A case was a 60-year-old man who had right upper abdominal pain and back pain. The abdominal CT revealed an early enhanced lesion in the posterior segment of the liver with portal vein and inferior vena caval tumor thrombi and multiple intrahepatic metastases. Tumor markers were elevated, AFP 2,480 ng/mL, PIVKA-II 31,900 mAU/mL. The patient underwent 4 courses of IFN-α/5-FU combination therapy and 8 times of TAI therapy of CDDP. After these therapies, tumors in the liver disappeared and tumor markers returned to the normal range. The patient is alive more than 58 months after the initial treatment. This case suggests that some patients with advanced HCC with tumor thrombus can get a long-term survival when intrahepatic lesions are controlled by various therapies including IFN-α/5-FU combination therapy.     

Elevated circulating levels of tumor necrosis factor predict unresponsiveness to treatment with interferon alfa-2b in chronic myelogenous leukemia.

PURPOSE: The study was undertaken to analyze circulating tumor necrosis factor (TNF) levels in patients with chronic-phase chronic myelogenous leukemia (CML) undergoing interferon (IFN) alfa-2b therapy, and to correlate pretreatment serum levels of TNF with response to IFN alfa-2b therapy. PATIENTS AND METHODS: Fourteen patients with CML in chronic phase were treated with recombinant human IFN alfa-2b for 7 to 39 months. RESULTS: In eight patients IFN alfa-2b treatment failed due to lack of hematologic response. A complete or partial hematologic remission was achieved in the remaining six patients, of whom two patients experienced a complete cytogenetic response. Retrospective analysis of serum samples obtained from all patients before the onset of IFN alfa-2b administration revealed that levels (mean +/- SEM) of circulating TNF were higher (P less than .001) in the group of patients who did not respond to IFN alfa-2b treatment (157 +/- 15 U/mL) than in the responders (10.3 +/- 4 U/mL) or healthy control subjects (9.1 +/- 3 U/mL). However, there was no correlation between TNF serum levels and other patient characteristics at study enrollment including age, sex, duration of disease, performance status, splenomegaly, WBC count, platelet count, hemoglobin value, prior therapy, and prognostic category. CONCLUSION: These findings indicate that circulating levels of TNF are increased in a subset of patients with chronic-phase CML and that this elevation is associated with poor response to IFN alfa-2b therapy.     

Combination Doxorubicin and Interferon-α Therapy Stimulates Immunogenicity of Murine Pancreatic Cancer Panc02 Cells via Up-regulation of NKG2D ligands and MHC Class Ӏ

Background: Pancreatic adenocarcinoma is a malignant gastrointestinal cancer with significant morbidity andmortality. Despite severe side effects of chemotherapy, the use of immunotherapy combined with chemotherapy hasemerged as a common clinical treatment. In this study, we investigated the efficacy of the combined doxorubicinand interferon-α (IFN-α) therapy on murine pancreatic cancer Panc02 cells in vitro and in vivo and underlyingmechanisms. Materials and Methods: A Panc02-bearing mouse model was established to determine whetherdoxorubicin and interferon-α (IFN-α) could effectively inhibit tumor growth in vivo. Cytotoxicity of naturalkiller (NK) cells and cytotoxic T lymphocytes (CTLs) was evaluated using a standard LDH release assay. Toevaluate the relevance of NK cells and CD8 T cells to the combination therapy-mediated anti-tumor effects,they were depleted in tumor-bearing mice by injecting anti-asialo-GM-1 antibodies or anti-CD8 antibodies,respectively. Finally, the influence of doxorubicin+interferon-α (IFN-α) on the ligands of NK and T cells wasassessed by flow cytometry. Results: The combination therapy group demonstrated a significant inhibition ofgrowth of Panc02 in vivo, resulting from activated cytotoxicity of NK cells and CTLs. Depleting CD8 T cells orNK cells reduced the anticancer effects mediated by immunochemotherapy. Furthermore, the doxorubicin+IFN-atreatment increased the expression of major histocompatibility complex class Ӏ (MHC Ӏ) and NKG2D ligands onPanc02 cells, suggesting that the combined therapy may be a potential strategy for enhancing immunogenicityof tumors. All these data indicate that the combination therapy using doxorubicin and interferon-α (IFN-α) maybe a potential strategy for treating pancreatic adenocarcinoma.     

Pharmacokinetic interaction of 5-fluorouracil and interferon alpha-2b with or without folinic acid

The purpose of this study was to evaluate a potential pharmacokinetic (PK) interaction between fluorouracil (5-FU) and the biomodulating agent interferon alpha (IFN-α) in patients with metastatic colorectal carcinoma. 5-FU was applied as an intravenous bolus injection of 750 mg m^?2 weekly and IFN-α 2b (IFN) 5 MU was injected 3 times weekly (TIW) subcutaneously. In the first study, 13 patients were treated by this schedule, 5-FU plasma levels were determined by HPLC on day (d) one as baseline before starting IFN; the analysis was repeated at the second or third cycle of 5-FU administration 1 hour after the last IFN injection respectively. In the second study, 10 patients additionally received folinic acid (FA) 200 mg m^?2 as a short time infusion immediately before 5-FU, and a third analysis of FU kinetics was performed in order to compare the influence of a double modulation of IFN and FA to IFN alone. Combination of 5-FU and IFN resulted in a significant increase of the AUC of 5-FU (80%) and the fictive initial concentration (C₀, 65%) obviously caused by a reduction of 5-FU clearance by 50%. However, when FA was added to this schedule, no significant changes of FU kinetics compared to 5-FU alone could be documented. Finally, in two pilot patients 5-FU 750 mg was given as a continuous infusion (CI) over 5 days and IFN 5 × 10⁶ u was added daily from d2 to d5. Analysis of 5-FU plasma levels was performed over 24 hours on d1 as baseline, on d2 after the first IFN injection and on d5 after the 4th IFN application. Compared to 5-FU alone, already on d2 there was a slight, but on d5 a marked increase of 5-FU plasma levels. This PK effect may contribute to the cytostatic action of 5-FU combined with IFN, especially in patients with enhanced 5-FU catabolism. Abrogation of this effect by using a double modulation of IFN and FA must be considered as disadvantageous despite several theoretical and preclinical synergism.     

Neutrophil Number After Interferon-Alfa Treatment is an Independent Predictive Marker of Overall Survival in Metastatic Renal Cell Carcinoma

BackgroundThe purpose of this study was to assess the outcome in patients treated by immunotherapy using interferon-alpha (IFN-α) and to evaluate the significance of the neutrophil count after IFN-α immunotherapy as a predictive marker for metastatic renal cell carcinoma (RCC).Patients and MethodsWe identified 84 patients with metastatic RCC who underwent immunotherapy with IFN-α between 1998 and 2006. The predictive values of the neutrophil count before and after IFN-α treatment as well as other clinical and laboratory parameters were assessed retrospectively.ResultsOn univariate analysis, the significant correlation with overall survival (OS) was recognized in the Eastern Cooperative Oncology Group (ECOG) performance score (PS), lactate dehydrogenase (LDH) levels, corrected calcium levels, interval from diagnosis to treatment, and the ratio of neutrophil number before and after treatment with INF-α. Multivariate analysis showed that ECOG PS, corrected calcium levels, interval from diagnosis to treatment and neutrophil number after IFN-α treatment were independent factors for OS. Using the number of neutrophils after IFN-α treatment, subgroups were identified using the Memorial Sloan-Kettering Cancer Center (MSKCC) model. The 1-year survival rate was 93% vs. 63% in the intermediate-risk group and 34% vs. 8% in the poor-risk group. In the favorable-risk group, all patients had a good decrease in neutrophil number after treatment with IFN-α.ConclusionNeutrophil number after IFN-α treatment can be a good predictive marker for OS in metastatic RCC. By combining MSKCC score with neutrophil number after treatment with IFN-α, we can subdivide each group.     

In vitro radiosensitization of human cervical carcinoma cells by combined use of 13-cis-retinoic acid and interferon-α2a

Background: Significant antitumor activity has been reported with the combined use of 13-cis-retinoic acid (cRA) and interferon-α2a (IFN-α) in the treatment of advanced-stage cervical cancers and skin cancers. Since IFN-α has been shown to be a modest radiation enhancer for selected malignant tumor cells and the cytotoxic activity is more enhanced by combining cRA and IFN-α, we hypothesized that the exposure of selected human carcinoma cells to combined cRA and IFN-α would render the cells highly radiosensitive.Methods and Materials: Two human cervical carcinoma cell lines, ME-180 and HeLa-S3, were chosen for the present study because of the different characteristics of the retinoic acid receptor status of the cell lines. To demonstrate the effects of combined cRA and IFN-α treatment on radiation response, we exposed the cells to cRA, IFN-α, or a combination of the drugs for 72 h before radiation. Experiments were carried out at minimally cytotoxic concentrations of the drug for radiation studies. End points of the study were cell growth inhibition and clonogenic ability of the single-plated cells. Effects of cRA and IFN-α on radiation response were quantitatively analyzed by constructing the radiation cell survival curves of ME-180 and HeLa cells.Results: ME-180 cells exhibited varying degrees of cytotoxicity with cRA and IFN-α, while HeLa cells showed no toxic effects with the same treatment. Combined treatment of cRA and IFN-α produced an additive cytotoxic effect in ME-180 cells. Radiosensitization was minimal when ME-180 cells were treated with either cRA or IFN-α before radiation. When ME-180 cells were exposed to 10 μM cRA for 48 h and 1000 U/ml IFN-α for 24 h prior to radiation, there was a significant enhancement in radiation-induced cell killing; the dose modification factor was 2.1 ± 0.9 at the 1% cell-survival level. On the other hand, HeLa-S3 cells exhibited no increased cytotoxicity or radiation enhancement under the same experimental conditions.Conclusion: The present data provide a radiobiological basis for using cRA and IFN-α as a combination radiosensitizer in selected human carcinoma cells.