In vivo hepatoprotective and nephroprotective effects of Stenocarpus sinuatus leaf extract against ifosfamide-induced toxicity in rats.

Ifosfamide (IFOS) is a broad-spectrum chemotherapeutic agent that has been extensively used for breast cancer and other solid tumors. Unfortunately, its use is associated with toxicities of several organs. Stenocarpus sinuatus is an Australian tree belonging to the Proteaceae family. In the current study, the phytochemical constituents of S. sinuatus methanol leaf extract (SSLE) were assessed. In addition, the protective effect of SSLE against IFOS-induced nephrotoxicity and hepatotoxicity was evaluated. Rats were randomly divided into six groups: control, IFOS (50 mg/kg), IFOS + SSLE (100 mg/kg), IFOS + SSLE (200 mg/kg), IFOS + SSLE (400 mg/kg), and SSLE (400 mg/kg). Hepatoprotective and nephroprotective potency of SSLE was assessed using different biochemical parameters. The phytochemical investigation resulted in the isolation of four flavonoid glycosides (kaempferol 3-O-ß- d-glucopyranosyl-(1→2)-α- l-rhamnopyranoside, kaempferol 3-O-α-rhamnopyranoside, isorhamnetin 3-O-ß- d-glucopyranosyl-(1→2)-α- l-rhamnopyranoside, and quercetin 3-O-ß- d-glucopyranosyl-(1→2)-α- l-rhamnopyranoside) and a coumarin (scopoletin). This is the first report on the isolated compounds from the genus Stenocarpus. SSLE showed enhancement of kidney and liver functions and reduction of oxidative stress and inflammation. The histopathology of the investigated organs confirmed the protective effect of SSLE. In conclusion, SSLE is considered as a promising candidate that can be used in defense against the toxic effects of IFOS after further clinical trials.

Inhibition of gene expression of carnitine palmitoyltransferase I and heart fatty acid binding protein in cyclophosphamide and ifosfamide-induced acute cardiotoxic rat models.

This study investigated whether cyclophosphamide (CP) and ifosfamide (IFO) therapy alters the expression of the key genes engaged in long-chain fatty acid (LCFA) oxidation outside rat heart mitochondria, and if so, whether these alterations should be viewed as a mechanism during CP- and IFO-induced cardiotoxicity. Adult male Wistar albino rats were assigned to one of the six treatment groups: Rats in group 1 (control) and group 2 (L-carnitine) were injected intraperitoneal (i.p.) with normal saline and L-carnitine (200 mg/kg/day), respectively, for 10 successive days. Animals in group 3 (CP group) were injected i.p. with normal saline for 5 days before and 5 days after a single dose of CP (200 mg/kg, i.p.). Rats in group 4 (IFO group) received normal saline for 5 successive days followed by IFO (50 mg/kg/day, i.p.) for 5 successive days. Rats in group 5 (CP-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days after a single dose of CP as group 3. Rats in group 6 (IFO-carnitine supplemented) were given the same doses of L-carnitine as group 2 for 5 days before and 5 days concomitant with IFO as group 4. Immediately, after the last dose of the treatment protocol, blood samples were withdrawn and animals were killed for biochemical, histopathological and gene expression studies. Treatment with CP and IFO significantly decreased expression of heart fatty acid binding protein (H-FABP) and carnitine palmitoyltransferase I (CPT I) genes in cardiac tissues. Moreover, CP but not IFO significantly increased acetyl-CoA carboxylase2 mRNA expression. Conversely, IFO but not CP significantly decreased mRNA expression of malonyl-CoA decarboxylase. Both CP and IFO significantly increased serum lactate dehydrogenase, creatine kinase isoenzyme MB and malonyl-CoA content and histopathological lesions in cardiac tissues. Interestingly, carnitine supplementation completely reversed all the biochemical, histopathological and gene expression changes induced by CP and IFO to the control values, except CPT I mRNA, and protein expression remained inhibited by IFO. Data from the current study suggest, for the first time, that (1) CP and IFO therapy is associated with the inhibition of the expression of H-FABP and CPT I genes in cardiac tissues with the consequent inhibition of mitochondrial transport and oxidation of LCFA. (2) The progressive increase in cardiotoxicity enzymatic indices and the decrease in H-FABP and CPT I expression may point to the possible contribution of these genes to CP- and IFO-induced cardiotoxicity.

The effect of N-acetylcysteine on ifosfamide-induced nephrotoxicity: in vitro studies in renal tubular cells.

Ifosfamide (IF) nephrotoxicity is a serious adverse effect in children undergoing chemotherapy. Previous studies have shown that, in addition to the renal production of chloroacetaldehyde, a toxic metabolite of IF, lower levels of glutathione (GSH) may predispose the kidney to damage. The antioxidant N-acetylcysteine (NAC) is used extensively as an antidote for acetaminophen poisoning in children by replenishing GSH levels. As it has been safely and effectively used clinically, the objective of this study was to test whether the reversal of ifosfamide-induced nephrotoxicity can be achieved by administering NAC. Supplementation with NAC may reduce or prevent the degree of cellular cytotoxicity induced by IF. Porcine renal proximal tubular (LLCPK-1) cells were treated with NAC (0.4 mM or 2.5 mM) concurrently with 1 mM IF and 50 microM L-buthionine sulfoximine (BSO). Cellular viability was assessed by alamarBlue assay at 96 h. Intracellular GSH and oxidized GSH (GSSG) levels were determined using a GSH/GSSG colorimetric detection kit. A significant 60% decrease in cellular viability occurred when cells were treated daily with BSO and IF for 96 h. This decrease was significantly reduced when cells were concurrently treated with NAC in a concentration-dependent manner. Intracellular and total GSH levels in cells receiving concurrent treatment of NAC were significantly higher than those without NAC treatment. NAC protects renal tubular cells from IF-induced cytotoxicity. It is likely that NAC is protecting the cells by partially acting as a precursor for GSH synthesis. This mode of therapy may allow for protecting children from life-threatening nephrotoxicity induced by IF.

Ifosfamide-induced nephrotoxicity: mechanism and prevention.

The efficacy of ifosfamide (IFO), an antineoplastic drug, is severely limited by a high incidence of nephrotoxicity of unknown etiology. We hypothesized that inhibition of complex I (C-I) by chloroacetaldehyde (CAA), a metabolite of IFO, is the chief cause of nephrotoxicity, and that agmatine (AGM), which we found to augment mitochondrial oxidative phosphorylation and beta-oxidation, would prevent nephrotoxicity. Our model system was isolated mitochondria obtained from the kidney cortex of rats treated with IFO or IFO + AGM. Oxidative phosphorylation was determined with electron donors specific to complexes I, II, III, or IV (C-I, C-II, C-III, or C-IV, respectively). A parallel study was done with (13)C-labeled pyruvate to assess metabolic dysfunction. Ifosfamide treatment significantly inhibited oxidative phosphorylation with only C-I substrates. Inhibition of C-I was associated with a significant elevation of [NADH], depletion of [NAD], and decreased flux through pyruvate dehydrogenase and the TCA cycle. However, administration of AGM with IFO increased [cyclic AMP (cAMP)] and prevented IFO-induced inhibition of C-I. In vitro studies with various metabolites of IFO showed that only CAA inhibited C-I, even with supplementation with 2-mercaptoethane sulfonic acid. Following IFO treatment daily for 5 days with 50 mg/kg, the level of CAA in the renal cortex was approximately 15 micromol/L. Taken together, these observations support the hypothesis that CAA is accumulated in renal cortex and is responsible for nephrotoxicity. AGM may be protective by increasing tissue [cAMP], which phosphorylates NADH:oxidoreductase. The current findings may have an important implication for the prevention of IFO-induced nephrotoxicity and/or mitochondrial diseases secondary to defective C-I.

Late effects surveillance system for sarcoma patients.

BACKGROUND: In 1998, a prospective multicenter pilot study of the 'Late Effects Surveillance System' (LESS) was started to investigate late effects of patients with Ewing, osteo- or soft-tissue sarcoma. PROCEDURE: Two hundred thirty patients were included in this pilot study. The patients were treated between 1/1/1998 and 6/30/1999 according to the sarcoma protocols COSS-96, CWS-96, and EICESS-92, the median cumulative doses of the focussed drugs were for cisplatin: 360 mg/m(2), for doxorubicin: 270 mg/m(2), and for ifosfamide: 24 g/m(2). The patients were investigated using an organ related standardized screening methodology. We report on toxicities in the first year after cessation of therapy-the beginning of the patient follow-up-and the feasibility of LESS. RESULTS: Cardiotoxicity: 16/129 (12%) patients treated with doxorubicin exhibited a reduced systolic heart function (fractional shortening (FS) <29%). Altogether three patients required cardiac drug therapy. Ototoxicity: In 5/73 (7%) patients treated with cisplatin a hearing deficit <4 kHz (>20 dB) was found. One patient needed a hearing aid. Nephrotoxicity: 2 of 214 (1%) patients treated with ifosfamide suffered from a tubulopathy, which required supplementation therapy. 10/50 (20%) showed a reduced fractional phosphate reabsorption. Incidence of hypomagnesemia was significantly increased in patients additionally treated with cisplatin. CONCLUSIONS: Some relevant impairments are noted in the first year after antineoplastic therapy. We expect to detect more major late sequelae in our prospective study during the increasing posttherapeutic interval. Our pilot study shows the feasibility of the methodology.

Thymoquinone attenuates ifosfamide-induced Fanconi syndrome in rats and enhances its antitumor activity in mice.

The effect of thymoquinone (TQ), the main constituent of the Nigella sativa L. oil, on ifosfamide (IFO)-induced Fanconi syndrome (FS) and its antitumor activity were investigated in rats and mice, respectively. In rats, a daily injection of IFO (50 mg/kg per day, i.p.) for 5 days induced a FS characterized by wasting off glucose, electrolytes and organic acids, along with elevated serum creatinine and urea, as well as decreased creatinine clearance rate. Administration of TQ with the drinking water of rats, (5 mg/kg per day) for 5 days before and during IFO treatment, ameliorated the severity of IFO-induced renal damage. TQ significantly improved IFO-induced phosphaturia, glucosuria, elevated serum creatinine and urea, and significantly normalized creatinine clearance rate. Moreover, TQ significantly prevented IFO-induced renal glutathione (GSH) depletion and lipid peroxide accumulation. In mice bearing Ehrlich ascites carcinoma (EAC) xenograft, TQ (10 mg/kg per day) administered in drinking water significantly enhanced the antitumor effect of IFO (50 mg/kg per day, i.p. on days 1-4 and 15-18). Furthermore, mice treated with IFO in combination with TQ showed less body weight loss and mortality rate compared to IFO single therapy. These observations demonstrate that TQ may improve the therapeutic efficacy of IFO by decreasing IFO-induced nephrotoxicity and improving its antitumor activity.

Whole-body hyperthermia combined with ifosfamide and carboplatin causes hypotension and nephrotoxicity.

It was previously postulated on the basis of clinical data that the cardiovascular sequelae of extracorporeal whole-body hyperthermia (e-WBH), i.e., hypotension (which requires catecholamine support) results in unique nephrotoxicity in combination with select chemotherapeutic agents. In an attempt to explain this phenomenon, we mimicked e-WBH physiological conditions in a rat model. Animals were treated with and without ifosfamide (IFO) and/or carboplatin (CBDCA) at 37 degrees C or 41.5-41.8 degrees C, with blood pressure monitoring and catecholamine support comparable to the clinical setting. Ex vivo post-treatment data (24 h) from artificially perfused kidneys (i.e., histology, urine volume, perfusion rate, glomerular filtration rate, and the reabsorption of sodium, glucose, and water) demonstrated unique toxicity including proximal tubular necrosis for the combination of WBH and IFO, for WBH and CBDCA and for WBH and IFO plus CBDCA, but not for IFO and CBDCA without WBH. These data, considered together with results derived from a subsequent clinical trial and the laboratory work of others were consistent with the hypothesis.

Taurine attenuates fanconi syndrome induced by ifosfamide without compromising its antitumor activity.

Nephrotoxicity is an important clinical side effect of the chemotherapeutic agent ifosfamide (IFO). Taurine (TAU), an antioxidant amino acid, was used in the present study to evaluate its beneficial effects against the Fanconi syndrome (FS) induced by IFO. The rationale of use of TAU was based on its reported antioxidant effect and the fact that the kidney tends to waste amino acid in IFO-induced FS. Rats received daily injection of IFO (50 mg/kg, IP) for 5 days with or without oral supplementation of 1% TAU in the drinking water for 7 days before IFO and daily thereafter. The results demonstrated that IFO induced a FS characterized by wasting off glucose, electrolytes, and organic acids, along with elevated serum creatinine and urea, and decreased creatinine clearance rate. TAU markedly ameliorated the severity of renal dysfunction induced by IFO, with a significant decrease in total and fractional excretion of Na+, K+, PO4(-3), and glucose, decreased serum creatinine, urea, and albumin, and increased creatinine clearance rate. TAU significantly improved the IFO-induced renal glutathione (GSH) depletion, renal malondialdehyde accumulation, and body weight loss. On the other hand, in Ehrlich ascites carcinoma (EAC)-bearing mice, IFO (50 mg/kg/day, IP on days 1-4 and 15-18) demonstrated antitumor activity as a single agent. No reduction in IFO activity was observed with the supplementation of TAU (1%) in the drinking water. Furthermore, the use of TAU not only maintained high IFO antitumor activity but also was associated with lower toxicity as manifested by less body weight loss and less mortality rate of IFO therapy compared with IFO when given alone. These observations demonstrate that oral supplementation of TAU can protect against IFO-induced renal dysfunction and maintain the antitumor activity of IFO.

Emesis induced in domestic pigs: a new experimental tool for detection of antiemetic drugs and for evaluation of emetogenic potential of new anticancer agents.

The domestic pig was used to develop a new model for evaluating the emetogenic potential of anticancer drugs and determining the antiemetic activity of drugs. Emesis was characterized by expulsion of solid or liquid material. In each animal, the number of vomits after infusion of the emetogenic drug (infusion in ketamine and xylazine anesthesia) was recorded in 1-hr periods during the first 4 hr and then in a 4- and a 16-hr period. Intravenous infusion of cisplatin caused a concentration-dependent emetic response. Anti-cancer drugs other than cisplatin such as carboplatin, dactinomycin, cyclophosphamide, and ifosfamide, also induced emesis, indicating that the domestic pig is suitable to detect the emetogenic potential of chemotherapeutic agents. A cisplatin dose of 2 mg/kg i.v. proved to be most suitable for studying the effect of potential antiemetic drugs (applied as i.v. injection), because this cisplatin dose caused consistent emetic responses without other toxic signs in the 24 hr following its infusion. Emesis induced by cisplatin was reduced by high doses of metoclopramide (25 mg/pig; approximately 0.8 mg/kg). The more selective dopamine D2 receptor antagonists, alizapride and domperidone, even at high doses (25-50 mg/pig; approximately 0.8-1.6 mg/kg), did not inhibit cisplatin-induced emesis, nor did haloperidol up to 20 mg/pig (approximately 0.6 mg/kg). Sulpride (50 mg/pig; approximately 1.6 mg/kg) halved the occurrence of vomits in the first 4 hr after cisplatin, but this effect was followed by an increase in the frequency of vomits; thus, no change in the total number of vomits was observed in the 24-hr observation period.(ABSTRACT TRUNCATED AT 250 WORDS)

Short duration, high dose, alternating chemotherapy in metastatic neuroblastoma. (ENSG 3C induction regimen). The European Neuroblastoma Study Group.

Fifty-one children, aged from 15 months to 13 years 5 months with metastatic neuroblastoma presenting sequentially at the participating institutions received four 3 to 4 weekly courses of high dose multiagent chemotherapy. High dose cisplatin (200 mg m-2) combined with etoposide (500 mg m-2), HIPE, was alternated with ifosfamide (9 g m-2), vincristine (1.5 mg m-2), and adriamycin (60 mg m-1), IVAd. Disease status was re-evaluated 3 to 4 weeks after the fourth course and the response classified according to the International Neuroblastoma Response Criteria (INRC). The overall response rate in evaluable patients was 55% and response rates by site were: bone marrow 67% (complete response 47%); bone scan 68%; primary tumour 61%, and urinary catecholamine metabolites (VMA/HVA) 95%. Serial 51Cr EDTA renal clearance studies showed a glomerular filtration rate (GFR) decline in 40% of patients but in only seven cases to below 50% of the pretreatment value. There was no instance of renal failure during induction, though two patients developed severe renal failure following 'megatherapy' given to consolidate remission. Serial audiometry showed a significant decline in hearing at frequencies above 2,000 Hz in 37% of children but at or below 2,000 Hz in only 17%. Neutropenia and thrombocytopenia were severe and intravenous antibiotics were required after 30% of courses. Each of two treatment-related deaths occurred during pancytopenia following courses of IVAd. Complete, or greater than 90%, removal of primary site tumour was possible in 70% of cases following this induction regimen and 75% of patients proceeded to elective megatherapy within a median time of 24 weeks after diagnosis. This short intensive induction programme is highly effective at achieving cytoreduction, enabling early surgery and early megatherapy procedures. It is, however, too early to draw firm conclusions about the impact of this approach to treatment on the cure rate.

Preclinical phase II study of ifosfamide in human tumour xenografts in vivo.

The in vivo effects of the oxazaphosphorine compound ifosfamide (IFO) on human tumour xenografts were assessed in thymus aplastic nude mice. The human origin of the tumours was confirmed by isoenzymatic and immunohistochemical methods. Tumour models were selected from a panel of 180 regularly growing, well-characterized xenografts. The maximum tolerated dose in tumour-bearing nude mice was determined to be 130 mg/kg per day given on days 1-3 and 15-17. After 21 days, lethality was 14% after i.p. and 6% after s.c. administration. A total of 43 human tumours were tested for antineoplastic activity, 15 of which (36%) showed regression: 4/5 breast cancer xenografts, 1/3 colon, 1/1 gastric, 2/7 non-small-cell lung cancers (NSCLC), 3/4 small-cell lung cancers (SCLC), 1/2 sarcomas and 3/3 testicular cancers. Two ovarian, two uterine and six renal cancer xenografts as well as three melanomas and five tumours of various histologies were resistant. In 30 human tumour xenografts, the antineoplastic efficacy of the two oxazaphosphorine derivatives cyclophosphamide and IFO was compared. The maximum tolerated dose of cyclophosphamide was 200 mg/kg per day given i.p. on days 1 and 15; it led to 17% lethality after 21 days. Cyclophosphamide induced tumour regression or remission in 10/30 xenografts (33%) and IFO in 13/30 (43%). In conclusion, the observed efficacy of IFO parallels the clinical situation. Breast, lung and testicular cancer and sarcomas proved to be responsive. The antitumoural activity of IFO shows similarities to that of cyclophosphamide; however, a higher response rate and lower toxicity were noted for the former. Preclinical phase II studies in nude mice seem to offer an effective way of identifying active drugs as well as sensitive tumour types for further clinical development.

Antitumor activity of optical isomers of cyclophosphamide, ifosfamide and trofosfamide as compared to clinically used racemates.

The relationship between enantiomeric homogeneity of three oxazaphosphorine drugs: cyclophosphamide, ifosfamide and trofosfamide and their antitumor activity was evaluated by standard screening tests against four in vivo transplantable tumor models: L 1210 and P 388 lymphoid leukemias, Lewis lung carcinoma and 16/C line of mouse mammary adenocarcinoma. It was shown that the stereodifferentiation of anti-tumor effect of enantiomers was not outstanding although quite consistently in favour of levorotatory forms. The only exception was seen for cyclophosphamide enantiomers tested against leukemias where R/+/form was more effective than S/-/or racemate.