Wojciech Feleszko

Potentiated antitumour effects of cisplatin and lovastatin against MmB16 melanoma in mice

Lovastatin, the drug used in the treatment of hypercholesterolaemia, has previously been reported to exert synergistic antitumour activity in a melanoma model in mice when used together with some immune response modifiers. In this study, we examined the antitumour effect of cisplatin augmented by its combined application with lovastatin, both in vitro and in vivo, in a murine melanoma model. The results of this study suggest that lovastatin may enhance the therapeutic effects of cisplatin in the treatment of malignant melanomas.

Antitumor effects of the combination immunotherapy with interleukin-12 and tumor necrosis factor α in mice

Abstract There is strong evidence that antitumor activity of interleukin-12 (IL-12) in vivo is mediated, in part, through interferon (IFNγ) produced by IL-12-stimulated natural killer and T cells. Since IFNγ and tumor necrosis factor α (TNFα) have been reported to synergize in antitumor effects in a number of models, we decided to examine whether the combined treatment with recombinant mouse IL-12 and recombinant human TNFα would produce similar effects. The efficacy of the combined IL-12/TNFα immunotherapy was evaluated in three tumor models in mice: B16F10 melanoma, Lewis lung (LL/2) carcinoma and L1 sarcoma. Intratumoral daily injections of 1 μg IL-12 in combination with 5 μg TNFα into B16F10-melanoma-bearing mice resulted in a significant retardation of the tumor growth as compared with that in controls and in mice treated with either cytokine alone. Similar effects were obtained using 0.1 μg IL-12 and 5 μg TNFα in LL/2 carcinoma and L1 sarcoma models. Antitumor activity against L1 sarcoma was still preserved when TNFα at a low dose (1 μg) was combined with 0.1 μg IL-12 and applied for a prolonged time. Potentiation of antitumor effects, which was observed in IL-12/TNFα-based immunotherapy, could result from at least three different mechanisms, partly related to stimulation of IFNγ and TNFα production in treated mice: (a) direct cytostatic/cytotoxic effects on tumor cells, (b) induction of antitumor activity of macrophages, and (c) inhibition of blood vessel formation in the tumor. Our studies demonstrate that combination tumor immunotherapy with IL-12 and TNFα may be more effective than single-cytokine treatment, and suggest possible mechanisms by which IL-12 and TNFα may exert potentiated therapeutic effects against locally growing tumors.

Effective chemo-immunotherapy of L1210 leukemia in vivo using interleukin-12 combined with doxorubicin but not with cyclophosphamide, paclitaxel or cisplatin

It has been well established that chemo‐immunotherapy using cytotoxic drugs and appropriate cytokines offers a new approach to increasing the therapeutic index in the treatment of neoplastic diseases. This study investigates the efficacy of combinations of interleukin‐12 with cyclophosphamide, paclitaxel, cisplatin or doxorubicin in the murine L1210 leukemia model. Mice inoculated i.p. with 1 × 103 or 1 × 105 leukemia cells were treated with interleukin‐12 and/or chemotherapeutics, and were observed daily for survival. Immunosuppression with X‐irradiation or macrophage depletion with injections of silica were used to examine the dependence of the therapeutic effects on the efficiency of the immune system. Treatment with interleukin‐12 or one of the studied chemotherapeutics given alone resulted in moderate anti‐leukemic effects. Combination of interleukin‐12 with cyclophosphamide or paclitaxel produced no augmentation of anti‐leukemic effects in comparison with these agents given alone. Combination of interleukin‐12 with cisplatin resulted in prolongation of the survival time; however, in the experiment with mice inoculated with 1 × 105 leukemia cells, no long‐term survivors (>60 days) were observed; on the contrary, combination of interleukin‐12 with doxorubicin resulted in 100% long‐term survivors. This effect was completely abrogated either by X‐irradiation of mice or by macrophage depletion. We also found that doxorubicin augments IL‐12‐stimulated production of interferon‐γ in vivo. Our observations demonstrating potentiation of the anti‐leukemic effects of the IL‐12 and doxorubicin combination suggest that the combined use of these 2 agents could be beneficial in leukemia therapy. Int. J. Cancer 77:720–727, 1998.

Potentiation of antitumor effects of tumor necrosis factor α and interferon γ by macrophage-colony-stimulating factor in a MmB16 melanoma model in mice

The efficacy of systemic infusion of recombinant human macrophage-colony-stimulating factor (M-CSF) in combination with local treatment with human recombinant tumor necrosis factor (TNF) α and mouse recombinant interferon (IFN) γ was studied in vivo on a subclone of B16 melanoma (MmB16) in mice. Short-term intravenous administration of M-CSF at a dose of 106 units daily had no antitumor effect in vivo. Similarly, local treatment of tumor with TNFα (5 μg daily) did not produce any therapeutic effect. However, simultaneous administration of the same dose of TNFα with IFNγ (1000 units daily) resulted in a synergistic effects manifested by the retardation of tumor growth. Addition of systemic infusion of M-CSF to the local therapy with TNFα and IFNγ induced further augmentation of antitumor efficacy and delayed progression of MmB16 melanoma. The strengthened antitumor effect of combination therapy including M-CSF, TNFα and IFNγ was most probably due to the increased release of monocytes from the bone marrow, their recruitment into the site of tumor growth and subsequent local stimulation of their antitumor activity.

Environmental tobacco smoke exposure and risk of allergic sensitisation in children: a systematic review and meta-analysis

Background Environmental tobacco smoke (ETS) exposure in children is linked with the development of allergic asthma. However, its influence on allergic sensitisation in children has not been conclusively determined. Objective To systematically review existing evidence of ETS exposures impact on markers of allergic sensitisation in children. Methods CENTRAL, MEDLINE and EMBASE databases were searched. Included studies assessed following markers of atopic sensitisation: total immunoglobulin E (tIgE) concentrations, at least one specific IgE (sIgE+), and positive skin-prick tests (SPTs+) in ETS-exposed and non-exposed children. Results 8 studies on the influence of ETS on tIgE concentration (2603 patients), 6 studies on ETS and sIgE+ (9230 participants) and 14 papers on ETS and SPT (14 150 patients) met our inclusion criteria. ETS was shown to raise tIgE concentrations by 27.7 IU/mL (95% CI 7.8 to 47.7; I2=58%; results based on 3 studies) and to increase the risk of atopic sensitisation, as assessed by sIgE+ (OR=1.12, 95%CI 1.00 to 1.25; I2=54%; results based on 4 studies) and SPT+ (OR=1.15; 95% CI 1.04 to 1.28; I2=0%; results based on 10 studies). In a subgroup analysis, this effect was most pronounced in children <7 years (preschoolers) by OR=1.20; (95% CI 1.05 to 1.38) and OR=1.30 (95% CI 1.05 to 1.61), (for sIgE+ and SPT+, respectively). Conclusions Current analysis supports an association between ETS exposure in early childhood and the increased risk of allergic sensitisation. Subgroup meta-analyses demonstrate that younger children suffer the most from detrimental immunomodulating effects of ETS exposure. This study underscores ETS as an important but avoidable risk factor for the development of allergic disease in children.

Potentiation of the anti-tumor effect of actinomycin D by tumor necrosis factor α in mice : Correlation between in vitro and in vivo results

The anti‐tumor effects of actinomycin D (Act D) and recombinant human tumor necrosis factor (TNF)‐α have been studied on 4 established murine tumor cell lines: MmB16 melanoma, Lewis lung (LL/2) carcinoma, L1 sarcoma and L1210 leukemia. During short‐term incubation (24 hr) Act D produced dose‐dependent cytostatic/cytotoxic effects against MmB16, LL/2 and L1 tumor cells but did not reduce the viability of these cells even at high concentration (10 μg/ml), below a threshold of 30–60%. However, L1210 leukemic cells were highly susceptible to Act D, and no viable cells were detected in cultures incubated with 1 μg/ml of Act D. TNF‐α alone, when used under the same culture conditions, had only a negligible effect on all cell lines tested. However, the combination of this cytokine with Act D produced synergistic cytotoxic effects against MmB16, LL/2 and L1 cells but not against L1210 leukemia cells. In an in vivo model of regional therapy in which tumor‐bearing mice were treated with Act D and TNF‐α, a correlation with in vitro results was observed. In mice bearing MmB16 melanoma, LL/2 carcinoma and L1 sarcoma, the most potent anti‐tumor effects were observed in mice treated with Act D and TNF‐α together. This treatment led to a delay of tumor growth and induced complete tumor regression in some cases. On the contrary, TNF‐α did not enhance the effect of Act D in mice injected with L1210 leukemia cells. Our results show that TNF‐α can potentiate the anti‐tumor effects of Act D against tumors weakly susceptible to Act D and may be a useful adjuvant to chemotherapy in the local treatment of neoplasia.

E-Cigarette use among children and young people: the need for regulation

Electronic cigarettes (e-cigarettes) are devices designed to deliver nicotine to the body via the route of inhalation. The principle of operation is based on heating a nicotine solution in propylene glycol and/or glycerine (e-liquid), turning it into aerosol (commonly called ‘vapour’), which is then inhaled by the user. The scientific evidence on the health consequences of long-term e-cigarette use is sparse and currently inconclusive. Young people are the most vulnerable group to initiate use of e-cigarettes. The novelty of the e-cigarette, perceptions about the harmlessness of the product, a wide variety of flavours (fruit, chocolate, peanut butter, bubble gum, gummy bear, amongst others), and peer-influence are just a few examples of factors contributing to the e-cigarette popularity among youth. The comprehensive e-cigarette regulations need to include rules on marketing, safety of newly introduced products (nicotine dosage, packaging, and labelling), marketing limitations, and banning the sale of e-cigarettes to minors.

In vitro antitumor activity of cerivastatin, a novel and potent HMG-CoA reductase inhibitor

Angiogenesis, the growth of new blood vessels from a preexisting microvascular bed, is of crucial importance for the growth, maintenance, and metastasis of solid tumors. Various anti-angiogenic factors are currently being investigated, indicating that angiogenesis may soon become a suitable target for novel antitumor therapies. However, most of the currently available potent angiostatic factors (angiostatin, endostatin) are small protein fragments and their clinical application may be associated with an unusual cost expense. In their recent report Vincent and colleagues [1] demonstrate an interesting, anti-angiogenic activity of 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitor (HMG-CoA RI), cerivastatin. Inhibitors of HMG-CoA reductase (or statins) represent a newly discovered family of chemically related molecules, selected for their lipid-lowering eiect. Statins are extensively used in medical practice, and large clinical trials have demonstrated that this class of lipid-lowering drugs greatly reduces cardiovascular-related morbidity and mortality in patients with and without coronary disease ([2] ; for review see [3]). According to recent studies, the bene¢cial eiect of statins may also be attributed to their favorable eiects on vasculature [3]. In fact, we have demonstrated for the ¢rst time that lovastatin, another HMG-CoA RI, inhibits angiogenesis, and this eiect was due to the decreased production of VEGF imposed by lovastatin in tumor cells [4]. Interestingly, the results of our study and the recent paper of Vincent et al. do not match that of Kureishi et al. [5], who show that statins promote angiogenesis. This discrepancy most likely depends of the type of the examined cells and the diierence of the experimental model used. In our study, lovastatin eiectively suppressed VEGF production by tumor cells harboring ras mutations. This resulted in the inhibition of bloodvessel formation, and ¢nally in the retardation of tumor growth. These results suggest statins remarkable anti-angiogenic and antitumor eiects, particularly towards tumors harboring ras mutations. Until recently, the bene¢cial antitumor eiects of treatment with statins have been attributed only to their direct antiproliferative eiects on tumor cells. Evidence was provided that statins induce cell cycle block in the G1 phase, interfere with the function of the Ras oncoprotein, and induce a potent apoptotic response. Some of the statins are being tested in clinical trials as potential novel antitumor agents, as they have been widely used and have well-de¢ned pharmacokinetics at the clinical level, displaying negligible adverse side eiects. Strikingly, a large clinical trial with lovastatin (a total of 6605 patients), designed to study prevention of acute coronary events with lovastatin, demonstrates a signi¢cant reduction in the incidence of melanoma among lovastatintreated patients [2]. The results of our previous study [4] and the paper of Vincent et al. [1] underline the feasibility of utilizing statins as anti-angiogenic agents in tumor therapy, especially if it is taken into account that they may be safely used to in£uence tumor bloodvessels on a daily basis at levels well below the maximum tolerated dose. Cerivastatin may be of particular interest, since it possesses superior lipid-lowering activity at doses equivalent to 1^3% of the doses of other statins (for review see [6]). Although recent reports showed that cerivastatin exerts the most potent antiproliferative activity in comparison to simvastatin, lovastatin and atorvastatin against smooth muscle and endothelial cells, no data exist yet about its direct antiproliferative activity against tumor cells. In order to further evaluate the antiproliferative eiects of cerivastatin on tumor cell growth, we compared its cytostatic/cytotoxic activity against various tumor cells to that exerted by lovastatin and simvastatin. We tested our hypothesis in a standard 3-[4,5-dimethylthiazol-2-yl]diphenyltetrazolium bromide assay (MTT) which was successfully applied in our previous studies [4]. In this study we tested cerivastatin on a panel of human and murine tumor cell lines, as shown in Table 1. The results of these experiments were plotted as dose^response curves and then subjected to median eiect analysis using the CalcuSyn software (Biosoft, www.biosoft.com). Subsequently, the IC50 values (fraction of aiected cells = 0.5) were calculated for each drug and cell line.

Granulocyte-macrophage colony-stimulating factor potentiates antitumor activity of interleukin-12 in melanoma model in mice.

To study the antitumor activity of the combination immunotherapy with interleukin-12 (IL-12) and granulocyte-macrophage colony-stimulating factor (GM-CSF), a murine MmB16 melanoma tumor model was used. Seven days after inoculation of MmB16 melanoma cells into the footpad of the right hind limb, mice were treated with IL-12 and/or GM-CSF administered intratumorally for 7 consecutive days. IL-12 used both at a high (1 µg) and at a low (0.01 µg) dose per day produced retardation of tumor growth, although neither treatment resulted in any significant prolongation of the survival of tumor-bearing mice. GM-CSF did not by itself exert antitumor activity in this model; however, it potentiated antitumor effects of IL-12. In particular, survival of tumor-bearing mice treated with IL-12 (0.01 µg per day) and GM-CSF was significantly prolonged compared with that in mice treated with either IL-12 or GM-CSF alone.

Augmentation of Antitumor Efficacy by the Combination of Actinomycin D with Tumor Necrosis Factor-Alpha and Interferon-Gamma on a Melanoma Model in Mice

The efficacy of combination treatment with actinomycin D (Act D), recombinant human tumor necrosis factor-alpha (TNF-alpha), and recombinant murine interferon-gamma (IFN-gamma) was examined on established MmB16 melanoma in mice. TNF-alpha alone had marginal effect in vitro on melanoma cells. However, when this cytokine was combined with either Act D or IFN-gamma, synergistic cytostatic/cytotoxic effects were observed. The highest cytotoxicity was demonstrated in cultures of melanoma cells in which all three agents together were added. In mice inoculated with 10(6) melanoma cells (into the footpad of the hind limb) and treated locally with Act D, TNF-alpha and IFN-gamma, beneficial therapeutic effects were found. When initiated 1 week after tumor cell inoculation, the 7-day treatment with all these agents administered together at daily doses: 0.2 microgram (Act D), 1 microgram (TNF-alpha), and 200 U (IFN-gamma) resulted in a significant delay of tumor progression in comparison to the therapy that included either Act D alone or TNF-alpha in combination with IFN-gamma. Side effects of such a treatment, both local and systemic, were negligible. The results of this study demonstrate that combination of regional chemotherapy (actinomycin D) and immunotherapy (TNF-alpha/IFN-gamma) may display higher efficacy than either treatment alone and may increase therapeutic index without augmenting toxic effects.