Mauro Moroni

In vitro antioxidant properties of amifostine (WR-2721, ethyol)

Purpose: Amifostine (WR-2721), a phosphorylated aminothiol pro-drug which is an analogue of cysteamine, is a selective cytoprotective agent for normal tissues from the toxicities associated with chemotherapy and irradiation. Despite a growing number of reports strongly supporting amifostines clinical efficacy, few authors have focused on the biochemical basis of amifostines antioxidant activity. Methods: We report on amifostines free-radical scavenging activity against superoxide (O2˙−), hydroxyl (OH−) and lipoperoxyl radicals in an in vitro model, using pure chemical systems. Amifostine was dephosphorylated to its active metabolite, WR-1065, by adding 10% non-heat-inactivated serum; different amifostine concentrations (1, 10, 50, 100 μM and 200 μM) and pH conditions (pH 5, 7.4 and 9) were tested. Results: Independent of the concentration, amifostine exhibited no major activity against O2˙− ions, neither did any pH variations in the experimental model provide any scavenger effects of the drug against O2˙− radicals. On the other hand, the protective effect of amifostine against OH− radicals was confirmed, yielding an EC50 of 255 μM at pH 7.4 and 230 μM at pH 5. Finally, amifostine exhibited scavenging activity against spontaneous lipoperoxidation, but no apparent antioxidant effect on iron ascorbate-induced lipoperoxidation. Conclusions: With this in vitro study, we are able to confirm the scavenging activity of the chemo- and radioprotector amifostine, whose activity seems to be particularly important from a biological point of view, since it is exerted mainly against highly reactive OH−.

Epirubicin and etoposide combination chemotherapy to treat hepatocellular carcinoma patients : a phase II study

Approximately half the patients affected with hepatocellular carcinoma (HCC) present with unresectable disease, so that efficacious systemic chemotherapy protocols are badly needed. We report the results of a phase-II study aimed at testing the efficacy and toxicity of a combination of epirubicin and VP-16. Thirty six patients (80 men and 6 women) received epirubicin (40 mg/m2, on day 1) and VP-16 (120 mg/m2, on days 1, 3 and 5) every 28th day. Chemotherapy was stopped in case of disease progression, while the patients who achieved an objective response or who had stable disease continued treatment for a maximum of 10 cycles. One patient (3%) achieved a complete response, while 13 patients (36%) achieved partial response, i.e. 14 objective responses in all (39%, 95% CI: 23-55%). 11 patients (31%) exhibited stable disease, while in the other 11 patients (31%) the disease progressed. Median overall survival time was 10 months and 13.5 months in the subgroup of patients responding to treatment. Significant, especially haematological, toxicity was documented, but in no case was it so severe as to require discontinuation of treatment or reduction of the dosage. In conclusion, this combination appears to be an active and tolerable therapeutic option for HCC patients who are not candidates for surgical or locoregional procedures, and in our opinion it deserves further exploration within a randomised controlled trial versus best supportive therapy.

5-Fluorouracil and d, l-Leucovorin Calcium Are Active to Treat Unresectable Hepatocellular Carcinoma Patients: Preliminary Results of a Phase II Study

Twenty-five patients affected with unresectable hepatocellular carcinomas (HCCs) were treated with 370 mg/m2 5-fluorouracil (5-FU) plus 200 mg/m2 racemic leucovorin both for 5 consecutive days. The chemotherapy cycle was repeated every 4 weeks until disease progression. One complete remission (4%) and 6 partial responses (24%) were obtained, that is 7 objective responses in all (28%, ninety-five confidence interval: 10.1-45.9%). All responders had a good PS and only 1 of them presented with bulky disease. Five patients (20%) exhibited stable disease, while the remaining 13 patients (52%) progressed. Toxicity was mild: 11 patients (44%) had grade II/III mucositis, 10 patients (40%) grade II diarrhea, 7 patients (28%) grade II nausea, 2 patients grade III granulocytopenia and 1 patient only (4%) grade I skin toxicity. This regimen made it possible to obtain a high rate of objective responses even in inoperable HCCs which are commonly considered as chemoresistant lesions.

Interleukin‐2 induces cell cycle perturbations leading to cell growth inhibition and death in malignant mesothelioma cells in vitro

Previous report indicated that Interleukin‐2 (IL‐2) is able to inhibit the growth of IL‐2‐receptor‐positive cancer cell lines without any involvement of the immune system, through IL‐2‐induced alterations of the cell cycle kinetics. In this study we provide evidence that IL‐2 exerts anti‐proliferative effect on three human malignant mesothelioma (MMe) cells in vitro, while no effects were observed on normal human mesothelial cell (HMC) primary cultures. The growth inhibitory effect of IL‐2 on neoplastic cells appeared to depend on the baseline proliferative status of these cells. Indeed, in highly proliferating MMe cells, we observed a reduction of malignant cells in the S‐phase of the cell cycle, with an accumulation in G0/G1, followed by apotosis for longer incubations or exposure to higher doses. On the contrary, in MMe cells proliferating at lower rate, IL‐2 induces only a late cytotoxic effect, leading to apoptosis, without significantly affecting the cell cycle. IL‐2Rβ mRNA was detectable by RT‐PCR in all MMe cells, IL‐2Rα mRNA in one only out the three assayed and IL‐2Rγ mRNA in none. In addition, mRNA specific for the IL‐2Rβ‐associated Jak‐1 tyrosine kinase was expressed in all MMe cell lines, further suggesting that IL‐2Rβ may play a role in the observed effects. Very low, albeit detectable, levels of IL‐2Rβ chain appeared to be expressed at the cell surface of MMe cells by indirect immunofluorescence and FACS analyses. Finally, Ca++ fluxes were rapidly induced when MMe cells were exposed to exogenous IL‐2. J. Cell. Physiol. 185:126–134, 2000.

5-Methyltetrahydrofolate or 5-formyltetrahydrofolic acid to modulate 5-fluorouracil’s cytotoxic activity in vivo? A phase II study in patients with advanced colon cancer

Abstract The purpose of this study was to test the hypothesis that 5-methyltetrahydrofolate (Me-THF), a source of reduced folates alternative to leucovorin, could effectively modulate 5-fluorouracil’s (5-FU) cytotoxic activity in patients with advanced colon cancer. A total of 23 patients were enrolled in a phase II trial; they received 5-FU as a 30-min infusion at a dose of 370 mg/m2 following a rapid i.v. push of 200 mg/m2 Me-THF, both drugs being given for 5 consecutive days. Cycles were repeated every 4 weeks until disease progression. No patient achieved a complete response. In all, 4 patients showed a partial response (17.4%), 7 developed stable disease (30.4%), and the remaining 12 (52.2%) progressed. Toxicity was acceptable and never exceeded WHO grade III intensity. According to our experience, the MeTHF/5-FU combination does not appear to be an effective treatment for advanced colon cancer. Despite its low toxic profile, in our opinion its wider use should be discouraged.