Tullio Giraldi

Antineoplastic activity and toxicity of an organometallic complex of ruthenium(II) in comparison with cis-PDD in mice bearing solid malignant neoplasms

The antineoplastic activity of an organometallic complex of ruthenium(II), [cis-RuCl2(DMSO)4]o, has been examined in comparison with that of cis-PDD, using three metastasizing tumors of the mouse: Lewis lung carcinoma, B16 melanoma and MCa mammary carcinoma. [cis-RuCl2(DMSO)4]o significantly reduces primary tumor growth in all the tumors tested, and its activity is similarly pronounced at three different dosages in mice bearing Lewis lung carcinoma. On the contrary, the survival time of animals having i.v. or i.m. tumor implants are only moderately increased, and also in the case of combined treatments with surgery. The antineoplastic activity of cis-PDD appears to be less pronounced than that of [cis-RuCl2(DMSO)4]o, and is limited to mice bearing B16 melanoma, which, among the three tumors used, appears to be naturally more responsive to cis-PDD and [cis-RuCl2(DMSO)4]o. The use of [cis-RuCl2(DMSO)4]o appears advantageous over that of cis-PDD since, unlike cis-PDD, its antineoplastic effects have been obtained at dosages with reduced host toxicity, indicated by the absence of significant hematological toxicity and toxicity for normal proliferating tissues.

Physiological regulation of P‐glycoprotein, MRP1, MRP2 and cytochrome P450 3A2 during rat ontogeny

P‐glycoprotein and the multidrug resistance‐related proteins MRP1 and MRP2 belong to the ATP binding cassette family of proteins and transport a wide range of substrates. These proteins are also involved in metabolic and excretory processes of xenobiotics. The rat genes mdr1a and mdr1b code for P‐glycoproteins, while mrp1 and mrp2 genes code for MRP1 and MRP2 proteins, respectively. In this study, the physiological modulation of the level of transcript for these genes during rat ontogeny in the liver, kidney, lung, brain and heart was analyzed by reverse transcription–polymerase chain reaction. An increasing level of transcript during ontogeny was demonstrated for mdr1a and mdr1b in all tissues considered, as well as for mrp2, which was detected only in the liver and kidney. In contrast, mrp1 transcript, present in all tissues, did not show any modulation. The maximum level of expression was reached in adult animals and a significant decrease was demonstrated in aging rats. Western blot analysis with C219 and M2III‐6 monoclonal antibodies confirmed this different pattern of expression during ontogeny in the liver. The physiological regulation of cytochrome P450 3A2 was also considered: in the rat liver, an increase in the level of transcript during ontogeny, with a maximum in 60‐day‐old rats and a decrease in 8‐month‐old rats, was evident.

Antitumor effects of rhodium(I), iridium(I) and ruthenium(II) complexes in comparison with cis-dichlorodiammino platinum(II) in mice bearing Lewis lung carcinoma

The effects of square planar rhodium, [RhacacCOD]o and iridium, [IracacCOD]o complexes and of octahedral ruthenium, [cis-RuCl2 (DMSO)4]o complex have been examined in comparison with cis-dichlorodiammino platinum(II) (cis-PDD). The toxicity in BDF1 mice varies widely and decreasing LD50-values, ranging from 0.94 mg/kg to 1000 mg/kg, have been obtained for cis-PDD, [RhacacCOD]o, [IracacCOD]o and [cis-RuCl2(DMSO)4]o, respectively. All the tested complexes similarly inhibit the growth of subcutaneous Lewis lung carcinoma and the development of spontaneous as well as of artificial metastases, with the exception of [IracacCOD]o which is inactive on metastases. The antitumor activity of [RhacacCOD]o and [cis-RuCl2(DMSO)4]o appears interesting, since it is of the same magnitude as that of cis-PDD, considering also that they were found to be only marginally nephrotoxic.

Melatonin administration in tumor-bearing mice (intact and pinealectomized) in relation to stress, zinc, thymulin and IL-2.

Melatonin (MEL) may counteract tumors through a direct oncostatic role. MEL is also an antistress agent with immunoenhancing properties against tumors due to a suppressive role of MEL on corticosterone release. Rotational stress (RS) (spatial disorientation) facilitates metastasis progression in mice. Also, MEL counteracts tumors because of its influence on immune responses via the metabolic zinc pool, which, is reduced in tumors and stress. Zinc is required for normal thymic endocrine activity (i.e. thymulin) and interleukin-2 (IL-2) production. Because in vivo data is still controversial, exogenous MEL treatment (22 days in drinking water) in both intact and pinealectomized (px) mice bearing Lewis lung carcinoma leads to significant decrements of metastasis volume, restoration of the negative crude zinc balance, recovery of thymulin activity and increment of IL-2 exclusively in intact and px tumor bearing mice subjected to RS. Significant inverse correlations are found in both stressed intact and px tumor bearing mice after MEL treatment between zinc and corticosterone (r = 0.78, P < 0.01; r = 0.80, P < 0.01, respectively). Positive correlations between zinc and IL-2 (r = 0.75, P < 0.01; r = 0.73, P < 0.01, respectively) or thymulin (r = 0.75, P < 0.01; r = 0.82, P < 0.01, respectively) are observed in same models of mice. These findings suggest a MEL action to decrease metastasis mediated by a possible interplay between zinc and MEL, via corticosterone, with consequent restoration of thymic efficiency and IL-2 production. MEL as an antistress agent with immunoenhancing properties in cancer deserves further consideration.nuclear factor-kb; POMC, proopiomelanocortin; Px, pinealectomized mice; RIA, radioimmunoassay; RS, rotational stress; SDI, stressed intact mice; SDPx, stressed pinealectomized mice; TNF-alpha, tumor necrosis factor-alpha; ZnFTS, active zinc-bound thymulin; ZnFTS + FTS, total thymulin.

Antitumour activity of equinatoxin

Equinatoxin, a highly basic protein extracted from Actinia equina, causes an increase in the survival time of mice bearing the ascitic form of Ehrlich carcinoma, whereas it has no effect on L1210 leukaemia. When tested for in vitro cytotoxicity by the dye exclusion test, it shows a potent activity on both tumour cell lines, with ED50 of a few ng/ml. Higher concentrations produce an extensive lysis of the cells. The cytotoxic effects of Equinatoxin are inhibited by phospholipids, thus suggesting that its mechanism of action may be related to interactions with lipids or other charged components of cell membrane. The observed lack of in vivo activity against L1210 leukaemia presumably is due to poor systemic absorption of the protein and/or neutralization by serum factors.

Lysosomal enzyme inhibitors and antimetastatic activity in the mouse

Abstract Saccharo- 1,4 -lactone, a β-glucuronidase inhibitor, and three protease inhibitors did not affect primary tumour growth when tested against Lewis lung carcinoma in mice. The number of lung metastases was significantly reduced by aprotinin only. The inactivity of leupeptin and pepstatin seems to indicate that cathepsin B and D are not involved in metastasis formation. These results also indicate that aprotinin, a broad spectrum protease inhibitor, reduces tumour dissemination, in addition to the tumour invasion already reported by others.

Toxicologic and pharmacokinetic study of low doses of verapamil combined with doxorubicin.

The effect of a chronic treatment with low oral doses of verapamil, a calcium channel blocker commonly employed in cardiovascular therapy, on doxorubicin toxicity, was evaluated in CD1 mice. Verapamil, administered at a dosage corresponding to a typical cardiovascular posology in humans, significantly increased doxorubicin toxicity. In particular the mortality was significantly higher and earlier and histological analysis revealed an increase in the severity of lesions in the liver, kidney and small bowel of verapamil pretreated animals. The pharmacokinetic profiles revealed that verapamil treated group had higher doxorubicin peak plasma and tissue levels and AUCs. This study shows that verapamil, administered at low doses, dramatically increases doxorubicin toxicity, probably through an interaction between the two drugs, both P-glycoprotein substrates, on the protein expressed in normal tissues, and suggests caution in the use of the calcium channel blocker for cardiovascular pathologies in patients who have to be treated with antineoplastic agents, substrates of P-glycoprotein.

Effects of stress on tumor growth and metastasis in mice bearing lewis lung carcinoma

The progression of Lewis lung carcinoma has been examined in mice under the stress of different housing and experimental conditions. The maintenance of the animals in a low stress environment decreased the weight of spontaneous lung metastases in comparison with conventional housing. The handling of mice in the low stress environment for intraperitoneal saline administration increased metastasis formation, whereas the application of a psychological stressor (spatial disorientation) to these animals increased both primary tumor growth and metastasis formation. These results indicate that psychological and experimental stressors can modulate, presumably via neuroendocrine mechanisms, the hosts antitumor responses which can control metastases and primary tumor independently from each other.

Effects of melatonin on doxorubicin cytotoxicity in sensitive and pleiotropically resistant tumor cells

Melatonin has been reported to attenuate the oxidative damage caused by doxorubicin on kidney, brain, heart and bone marrow, whereas the in vivo antitumor effects of doxorubicin were not attenuated. The effects of melatonin on doxorubicin cytotoxicity have, therefore, been examined on human normal mammary epithelium HBL‐100, on mammary adenocarcinoma MCF‐7, on colon carcinoma LoVo, and on mouse P388 leukemia cell lines, and on tumor cell sublines pleiotropically resistant to anthracyclines. Melatonin in the concentration range 10–2000 pg/mL causes an inhibition of the growth of the human cell lines examined which is not clearly dose‐dependent and less than 25% when significant. Melatonin similarly causes minor effects on doxorubicin cytotoxicity either on the parental human cell lines or on their resistant sublines. On the contrary, 200–1000 pg/mL melatonin cause a significant and dose‐dependent partial sensitization to doxorubicin of resistant P388 mouse leukemia (P388/ADR), which occurs also in vivo, as indicated by a significant increase in survival time of the hosts. Doxorubicin intracellular concentrations in P388/ADR cells are increased by melatonin, suggesting that melatonin might inhibit P‐glycoprotein‐mediated doxorubicin efflux from the cells. These results indicate that the use of melatonin in clinical cancer treatment should not pose the risk of an attenuation of the effectiveness of doxorubicin, and encourage the further examination of the possible reduction by melatonin of the host toxicity of antitumor chemotherapy.

RESTRAINT STRESS REDUCES THE ANTITUMOR EFFICACY OF CYCLOPHOSPHAMIDE IN TUMOR-BEARING MICE

Treatment with the cytotoxic antitumor drug cyclophosphamide is highly effective in mice bearing Lewis lung carcinoma, causing the absence of macroscopically detectable tumors at necroscopy after sacrifice. When the effects of the treatment on survival are determined, a significant increase in survival time and in the proportion of long-term survivors is observed. When restraint stress is further applied, tumors develop in all of the mice treated with cyclophosphamide, and survival time and the fraction of long-term survivors are significantly reduced. Flow cytometry of splenic T-lymphocyte subsets in normal mice indicates a significant decrease in the number of CD3+, CD4+, and CD8+ subsets after treatment with cyclophosphamide and after application of restraint stress; the interaction of the two treatments is significant for CD3+ and marginally significant for CD4+ subsets. The attenuation by restraint stress which was observed for the effects of cyclophosphamide on the presence of tumors at necroscopy and for the survival of the treated mice might thus be interpreted as follows: restraint stress attenuates the immune functions of the host directed toward the weakly immunogenic tumor, an effect which, in the absence of restraint stress, interacts effectively with the cytotoxic action of cyclophosphamide toward tumor cells. The results obtained using this animal model thus indicate that experimental stress reduces the therapeutic efficacy of a cytotoxic antitumor drug; experimental and clinical implications are discussed.