L. Rausa

The CD95/CD95 ligand system is not the major effector in anticancer drug-mediated apoptosis

Many anticancer drugs are able to induce apoptosis in tumor cells but the mechanisms underlying this phenomenon are poorly understood. Some authors reported that the p53 tumor suppressor gene may be responsible for drug-induced apoptosis; however, chemotherapy-induced apoptosis can also be observed in p53 negative cells. Recently, doxorubicin (DXR) was reported to induce CD95L expression to mediate apoptosis through the CD95/CD95L system. Thus, an impairment of such a system may be involved in drug resistance. We evaluated the in vitro antitumor activity of several cytotoxic drugs on two human p53-negative T-cell lymphoma cell lines, the HUT78-B1 CD95L-resistant cell line and the HUT78 parental CD95L-sensitive cell line. We demostrated by Western blotting assay that DXR and etoposide (VP-16) were able to induce CD95L expression after 4 h of treatment. In contrast, they were unable to induce the expression of p53. DXR, at concentrations ranging from 0.001–1 μg/ml, and VP16, at concentrations ranging from 0.05–1 μg/ml, were equally cytotoxic and induced apoptosis in both cell lines as assessed by fluorescence microscopy and flow cytometry analyses. Although we observed a slightly reduced percentage of apoptotic cells in HUT78B1 when compared with the parental HUT78 cells after few hours of drug exposure, this difference was no longer evident at 48 or 72 h. Similarly, the exposure of HUT78 cells to a CD95-blocking antibody partially reduced early apoptosis (24 h) without affecting the long-term effects of the drugs including cytotoxicity. Furthermore, as observed with DXR and VP-16, both the CD95L-sensitive and the CD95L-resistant cell lines resulted equally sensitive to the cytotoxic effects of a number of different cytotoxic drugs (vincristine, camptothecin, 5-fluorouracil and methotrexate). The treatment with the Caspase-3 tetrapeptide aldehyde inhibitor, Ac-DEVD-CHO, did not affect the DXR-induced apoptosis whereas it only modestly inhibited apoptosis and cytotoxicity of VP-16, while Z-VAD.FMK, a Caspase inhibitor that prevents the processing of Caspase-3 to its active form, was able to block DXR-induced apoptosis at 24 h but not at 48 h. Thus, our results do not confirm a crucial role for the CD95/CD95L system in drug-induced apoptosis and suggest the involvement of alternative p53-independent pathways at least in this experimental model system.

Lysosomal alterations in heart and liver of mice treated with doxorubicin

SummaryThis study was carried out to evaluate the influence of long-term treatment with doxorubicin (DXR) (4mg/kg IV for 5 weeks) on heart and liver lysosomes of mice.We evaluated the variations in both total and “sedimentable” enzyme activity of cathepsin D, which is the major endopeptidase of myocites and probably involved in physiologic and pathologic degradation of actomyosin and mitochondria, and that of acid phosphatase, which is more prominent in interstitial cells.Our results show that marked changes occur in both total and sedimentable enzyme activity of cathepsin D in the heart of treated animals and to a lesser extent in the liver.In contrast, no modification of either total or sedimentable acid phosphatase was seen in either organ.The effects we observed are much more marked for cardiac cathepsin D; this is in good agreement with the cardiac specificity of DXR-induced cardiotoxicity with long-term administration and suggests that lysosomes could play a role in the pathogenesis of this phenomenon.

Cardiac Peroxisomal Enzymes and Starvation

In mice subjected to 3-day periods of food deprivation an increase in plasma free fatty acids occurred together with a rise in the cardiac content of fatty acyl CoA-oxidase (+ 15.2%) and catalase (+ 136.2%) activities. Stimulation of hydrogen peroxide production by the heart was found after 30 hours of fasting and this phenomenon was almost completely eliminated by 6 hours of refeeding. These data suggest that high myocardial loads of free fatty acids involve the peroxisomal enzymes in the beta-oxidation process. The resulting increase in hydrogen peroxide production could be partly responsible for the myocardial injury caused by starvation.

NF‐κB Inhibition Restores Sensitivity to Fas‐Mediated Apoptosis in Lymphoma Cell Lines

Abstract: Failure to perform the Fas‐related apoptosis pathway can account for tumor resistance both to chemotherapeutic agents and to immunological effectors. We studied the role of NK‐κB in Fas‐resistance, employing the Fas‐sensitive human T‐lymphoma HuT78 cell line and its Fas‐resistant variants HuT78B1 and HuT78G9. All these cell lines expressed high levels of constitutively activated NF‐κB. Pretreatment of cells with NF‐κB inhibitors (PDTC, MG132, or SN50) strongly enhanced CH11‐induced apoptosis in HuT78 and Hut78G9 cells, while only MG132 showed a similar potentiating effect in HuT78B1. The described synergism was significantly inhibited by pretreatment with the anti‐Fas‐blocking antibody ZB4 or with the pancapsase inhibitor Z‐VAD‐FMK, but not by capsase‐8 or ‐9 inhibitors. Overall, these data suggest that NF‐κB inhibition may restore the Fas‐pathway in Fas‐resistant NF‐κB‐overexpressing tumors.

Influence of Pharmacokinetic Variations on the Pharmacological Properties of Adriamycin

Whenever it appears impossible to modify the chemical structure of drugs with a high and established therapeutic activity but a low chemotherapeutic index, pharmacological research has to find other ways of improving the chemotherapeutic index. This problem is particularly important in the case of antitumor drugs, thus justifying research into the most suitable choice of dosage and routes of administration, as well as into the pharmacological associations which enable tumor cells to be hit at various stages of the reproductive cycle. Alternatively, the therapeutic index could be improved by the use of antagonistic compounds (like, for example, methotrexate and folinic acid) which act upon the same organic functions.

5-Fluorouracil and recombinant alpha interferon-2a in the treatment of advanced colorectal carcinoma: a dose optimization study.

A dose optimization study was carried out with the aim of identifying the maximally tolerated dose of recombinant alpha interferon-2a (raIFN-2a) in combination with 5-fluorouracil (5FU). 5FU was given at the dose of 750 mg/m2 over a 4-hour infusion on day 1- - greater than 5 followed by 750 mg/m2 weekly i.v. bolus. Recombinant aIFN-2a was started at 3 x 10(6) IU subcutaneously three times/week. 12 patients with advanced colorectal carcinoma were included in the study. 10 patients had previously received chemotherapy for advanced disease. Severe fatigue, most likely attributable to rIFN, was the dose-limiting toxicity. The dosage of raIFN-2a could not be further escalated above 12 x 10(6) IU. At this dose level all patients required dose reduction due to fatigue, fever, myalgia and severe reduction of performance status.

5-Fluorouracil plus interferon α-2a compared to 5-fluorouracil alone in the treatment of advanced colon carcinoma : A multicentric randomized study

Abstract Biochemical modulation is one of the most interesting fields in cancer chemotherapy. Interferon-α (IFNα) is a cytokine that is able to influence the pharmacodynamics of 5-fluorouracil (5FU) through a number of mechanisms. With the aim of confirming some data emerging from the literature, we initiated a multicentric randomized study comparing the combination of 5FU and IFNα-2a with 5FU alone in the treatment of advanced or metastatic colon cancer. A group of 205 colon cancer patients (104 in the 5FU arm and 101 in the 5FU+IFNα-2a arm) were included in the final intention-to-treat analysis. Rectal cancers were not considered eligible. All patients had measurable disease, were aged 75 years or less, had a Karnofsky index of at least 60 and had good bone marrow, renal, liver and cardiac functions. No previous chemo-immunotherapy was allowed. The treatment was 750 mg/m2 5FU (4 h i.v. infusion) on days 1–5 and then i.v. bolus weekly, starting from day 12, with or without IFNα-2a given s.c. three times weekly (starting dose 3 × 106 IU rising to 9 × 106 IU, if tolerated). Patients were treated until progression or, if responsive, for a maximum of 48 weeks and then observed for a period of 2 years. The primary end-point of the study was objective clinical response (OR); secondary parameters were time to progression, overall survival, and time to death after progression. WHO criteria were used for both clinical response and toxicity measurements. Dose reduction was planned a priori in the event of significant toxicity due to 5FU, IFNα-2a or both. Association between primary and secondary end-points and treatment was studied by univariate and multivariate analysis. Altogether, 47 patients achieved a documented response. A 25% OR was observed in the combination arm while a 21% OR was seen in the 5FU arm; this difference is not statistically significant (P=0.6). Patients with a small tumour burden (below 5 cm2) showed a higher probability of response in both arms. Patients in the experimental arm had a higher but not statistically significant cumulative progression-free probability. Median survival was 47.1 weeks overall, while it was 43.7 and 48.5 weeks in the control and experimental arms, respectively. The combination was clearly more toxic than 5FU alone, leukopenia being the most frequent side-effect in the experimental arm and nausea and vomiting in the control arm. In conclusion these results are quite disappointing and 5FU + IFNα-2a can not be considered a standard treatment for advanced colon cancer.

Does catalase play a role in Adriamycin induced cardiotoxicity

Summary Adriamycin causes an increase of lipid peroxidation in mouse cardiac homogenates that is dependent on the concentration of the antiblastic. The same phenomenon is not observed in the hearts of mice treated with an elevated dose of Adriamycin in which, conversely, an increase of the antioxidizing enzyme catalase was noticed. The significance of these findings is discussed with relationship to the hypothesis of an enhanced free radicals formation at the basis of Adriamycin induced cardiotoxicity.

Cis-diamminodichloroplatinum plus a 5-day continuous infusion of 5-fluorouracil in the treatment of locally recurrent and metastatic head and neck cancer patients

SummaryA group of 23 consecutive patients with biopsy-proven advanced or metastatic head and neck cancer were treated with cisplatinum, 100 mg/m2 i.v., on day 1 plus 5-fluorouracil, 1000 mg/m2, in continuous infusion for 5 days. Most patients (87%) had recurrent or metastatic cancer and were previously treated (78%). Out of 21 evaluable patients we obtained a 42% overall response rate (complete+partial responses) with a mean duration of more than 8 months and a 14% minimal response rate. A stabilization of disease was achieved in 28% of cases, while 14% of patients progressed. This response rate, as well as the duration of response, seems to be similar to those obtained in other series comprising previously treated patients with advanced or metastatic head and neck carcinoma. The toxicity was generally acceptable, with few cases of grade 3 (WHO criteria) toxicity. However most patients required hospitalization because of the length of treatment. In conclusion the response rate and the duration of responses obtained with cisplatinum plus a 5-day infusion of 5-FU in advanced or metastatic pretreated patients is, at present, unsatisfactory, even if the impact on survival is still not entirely clear.

High-dose folinic acid and 5-fluorouracil plus cisplatin on a weekly schedule in the treatment of advanced cancer of the head and neck

SummaryA group of 60 patients with advanced head/neck cancer were treated with high-dose folinic acid (500 mg/m−2/week−1) plus 5-fluorouracil (400 mg/m−2/week−1 on day 1, and cisplatin (20 mg/m−2/week−1) 24 h after folinic acid infusion was completed. Out of 55 evaluable patients, 10 patients (18%) experienced a complete response with a mean duration of 11.4+ months, 25 patients had a partial response (45%) of 6.7+ months, 6 patients (11%) showed a stabilization of 4.8+months, and 14 (25%) progressed. The overall response rate was 63.6% (95% confidence limits 56.5%–69.5%). Patients pretreated with radiotherapy had a 67% overall response rate, while those pretreated with chemotherapy showed a 54% overall response rate. All patients with cancer of the oropharynx had a major response, while patients with cancer of the oral cavity had the lowest response rate. The mean survival of patients who attained a complete response was 14.5+ months. Partial responders had a mean survival of 10.6+ months, while patients who progresses survived a mean of 3.6+months. The treatment has been very well tolerated with few cases of grade 3 gastrointestinal toxicity. Grade 1–2 leukopenia was recorded in 64% of cases, grade 1–2 nausea/vomiting in 85%. In one case therapy was stopped because of persistent diarrhoea.