Drug retention and distribution after intratumoral chemotherapy with fluorouracil/epinephrine injectable gel in human pancreatic cancer xenografts

Purpose: Pancreatic cancer is widespread, associated with high mortality, and rapidly fatal. Most cases are diagnosed too late for surgical treatment, and the disease responds poorly to systemic chemotherapy. Nevertheless, pancreatic cancer cells are sensitive to fluorouracil (5-FU) in a time- and dose-dependent manner, suggesting that improved retention of drug in the tumor may improve patient prognosis. In this study, we evaluated a novel drug delivery system, 5-FU/epinephrine injectable gel (5-FU/epi gel), designed to improve drug retention in tumors. Methods: We used a BxPC-3 human pancreatic cancer xenograft model in athymic mice to examine drug levels in tumor, liver, and kidney tissue following administration of: (a) 5-FU/epi gel (30 mg 5-FU/ml) intratumorally (i.t.); (b) 5-FU solution i.t.; and (c) 5-FU solution intraperitoneally (i.p.). [³H]5-FU was added as a radiolabeled marker to all test formulations. Animals were sacrificed at designated times, and the tumor, liver, and one kidney from each animal were excised and processed for radioactivity analysis. Drug concentration was quantified by both storage-phosphor autoradiography (SPA) and liquid scintillation counting (LSC). Results: Higher and sustained i.t. drug levels were achieved following i.t. administration of 5-FU/epi gel (SPA AUC 18.4 mM · h, LSC AUC 13.0 mM · h) compared with 5-FU solution i.t. (SPA AUC 2.02 mM · h, LSC AUC 1.92 mM · h) or 5-FU solution i.p. (SPA AUC 0.07 mM · h, LSC AUC 0.04 mM · h). Use of the 5-FU/gel system was associated with lower drug levels in liver and kidney, indicating that it produces far less systemic exposure. Conclusion: In the human pancreatic cancer xenografts, i.t. administration of 5-FU/epi injectable gel provided significantly higher drug and/or metabolite concentrations for extended periods than was possible with either i.t. or i.p administration of drug solution. This i.t. drug delivery system could potentially be used to treat patients with pancreatic cancer to increase tumor exposure to drug and improve the therapeutic index in comparison to systemic drug administration. Received: 24 July 1998 / Accepted: 29 January 1999     

Increased cytotoxicity of low-dose,long-duration exposure to 5-fluorouracil of V-79 cells with hyperthermia

Summary We examined the cytotoxic effects of combined low dose and long exposure to 5-fluorouracil (5-FU) and hyperthermia on Chinese hamster V-79 cells with reference to timing and sequence of administration. The survival rate following hyperthermia at 42°C for 2 h alone was 95.4%, and that after exposure to 1.0 g/ml/5-FU alone for 48 hours, 94.2%. With respect to the combination of 5-FU and heat, the survival rate of cells exposed to hyperthermia at 42°C for 2 h followed by 1.0 g/ml 5-FU treatment for 48 h followed by hyperthermia led to a survival rate of 10%. Flow cytometric analysis of V-79 cells after exposure to 1.0 g/ml 5-FU for 48h revealed an accumulation of cells in the S-phase; the percentage of S-phase exponential growing cells was 65% and the plateau phase was 38%. The former were more sensitive to heat than the latter cells according to the MTT assay. V-79 cells pretreated with 5-FU were more sensitive to hyperthermia than were those not pretreated with 5-FU. Therefore, when 5-FU plus heat is to be used to treat a patient with a malignancy, the sequence of 5-FU followed by hyperthermia may be more effective than the reverse.     

The Effect of DFMO Induced Uptake of [3H] Putrescine On Human Glioma Cells

Polyamine synthesis inhibitors, such as -difluoromethylornithine (DFMO), inhibit tumor cell growth in vitro and in vivo. However, upon cessation of treatment, tumor growth resumes. We hypothesized that incorporation of radioactive polyamines might kill the growth-arrested cells. This hypothesis was previously tested in rat 9L brain tumor cells in which DFMO increased both the uptake and the retention of [³H] putrescine. In these rat cells, DFMO-induced retention of high-specific-activity [³H] putrescine for 20 days resulted in several logs killing. In the present studies all of the 5 different human glioma cell lines tested with DFMO treatment also showed enhanced uptake of exogenous [³H] putrescine, reduced cell counts and enhanced killing of colony forming cells (CSF). Extending the time of DFMO treatment of cells that had taken up high-specific-activity (80Ci/mmol) [³H] putrescine further increased the killing. A 10-day extension resulted in a 10,000-fold reduction in cumulative cell growth. A 5-day extension resulted in a 2–3 log decrease in numbers of surviving CFC. These data further support the hypothesis and suggest that DFMO-induced cell cycle arrest enhances cellular retention of [³H] putrescine, increasing the effective internal radiation dose enough to cause proliferative death. In a clinical setting, the short ( 1m) path-length of the tritium particle should limit effects to the tumor cells and spare adjacent normal cells. These results support the concept that treatment with the combination of polyamine inhibitors and radioactive polyamines might be a useful adjunct to current therapies for glioblastoma multiforme.     

Relationship between in vivo drug exposure of the tumor interstitium and inhibition of tumor cell growth in vitro: a study in breast cancer patients

A novel approach is described to simulate effect site pharmacodynamics of anticancer drugs. This approach is based on (i) the in vivo measurement of unbound, interstitial drug pharmacokinetics (PK) in solid tumor lesions in patients and (ii) a subsequent pharmacodynamic (PD) simulation of the time versus drug concentration profile in an in vitro setting. For this purpose, breast cancer cells (MCF-7) were exposed in vitro to the time versus interstitial tumor concentration profiles of 5-fluorouracil (5-FU) and methotrexate (MTX) from primary breast cancer lesions in patients. This led to a maximal reduction in the viable cell count of 69 on day 4, and of 71 on day 7 for 5-FU and MTX, respectively. This effect was dependent on the initial cell count and was characterized by a high interindividual variability. For 5-FU there was a significant correlation between the maximum antitumor effect and the intratumoral AUC (r=0.82, p=0.0005), whereas no correlation could be shown for MTX (r=0.05, p=0.88). We conclude, that the in-vivo-PK / in-vitro-PD model presented in this study may provide a rational approach for describing and predicting pharmacodynamics of cytotoxic drugs at the target site. Data derived from this approach support the concept that tumor penetration of 5-FU may be a response-limiting event, while the response to MTX may be determined by events beyond interstitial fluid kinetics.     

Schedule specific biochemical modulation of 5-fluorouracil in advanced colorectal cancer: A randomized study

Background:We have recently suggested that bolus 5-fluorouracil(5-FU) may work via a RNA directed mechanism while continuous infusion 5-FUmay kill cells via a thymidylate synthase related pathway. It may thus bepossible to selectively modulate each schedule biochemically. We have comparedan alternating regimen of bolus and continuous infusion 5-FU, selectivelymodulated for the schedule of administration, with modulated bolus 5-FU inadvanced colorectal cancer patients. Patients and methods:Two hundred fourteen patients from nineteenItalian centers were randomized to the control arm consisting of biweeklycycles of MTX, 200 mg/m² on day 1, followed by bolus 5-FU 600mg/m² on day 2 and 6-S-leucovorin rescue, or to the experimentalarm consisting of two biweekly cycles of the same regimen as in the controlarm alternated to three weeks of continuous infusion 5-FU (200mg/m² day) + weekly bolus 6-S-leucovorin, 20 mg/m². Results:Nine CR and twenty-seven PR were obtained on one hundredeleven evaluable patients treated in experimental arm (RR = 32%,95% confidence interval (95% CI): 24%–42%),while two CR and eleven PR were observed among one hunderd three evaluablepatients in control arm (RR = 13%, 95% CI:7%–21%). WHO grade 3–4 toxicity occurred in13% of cycles of experimental arm and in 8% of cycles in controlarm. The PFS was significantly longer in experimental arm (6.2 vs. 4.3 months,odds ratio 0.66, P = 0.003), while the overall survival was similarin both arms (14.8 months in experimental arm vs. 14.1 months in control arm);quality of life was similar as well. Eighty percent of patients receivingsecond-line chemotherapy in control arm were treated with continuous infusion5-FU. Conclusions:Alternating, schedule-specific biochemical modulationof FU is more active than MTX 5-FU as first-line treatment of advancedcolorectal cancer. However, the overall survival was similar suggesting thatalternating bolus and infusional 5-FU upfront may be as effective as givingthem in sequence as first- and second-line treatment.     

Radiosensitization by intratumoral administration of cisplatin in a sustained-release drug delivery system.

PURPOSE: Effects of combining local irradiation and intratumoral (i.t.) administration of cisplatin (CDDP) in a sustained-release drug delivery system (epi gel) were studied in a murine SCCVII squamous cell carcinoma model in mice. MATERIALS AND METHODS: The epinephrine injectable gel was used as a drug delivery system. Intratumoral pharmacokinetics of CDDP was studied by using 195mPt-CDDP. The tumor volume quadrupling time (TVQT) and tumor growth delay (TGD) time were used to evaluate the antitumor efficacy of treatment regimens. RESULTS: The concentration and residence of 195mPt-CDDP was significantly higher in tumors treated with 195mpt-CDDP/epi gel than in tumors treated with 195mPt CDDP gel or 195mPt-CDDP suspension. Intratumoral administration of CDDP/epi gel (4 mg/kg) produced an average TGD time of 15.5 +/- 2.8 days, which was 5.2 - 7.4 times longer than CDDP suspension i.t. or i.p. When combined with a single dose of radiation (10 Gy), i.t. administration of CDDP/epi gel was 2.0 - 3.6-fold as effective as administered i.t. in suspension (39.2 +/- 4.1 vs. 19.8 +/- 3.9 days of TGD, P < 0.05) or i.p. in solution (39.2 +/- 4.1 vs. 11.0 +/- 1.6 days, P < 0.001) in inhibiting tumor growth and produced 20-60% complete remission of tumors. When combined with fractionated irradiation, pre-irradiation CDDP administration was more effective than post-radiation administration (26.7 vs. 12.1 days of TGD, P < 0.05). Mice treated with CDDP/epi gel i.t. alone or in combination with irradiation, had little systemic toxicity. CONCLUSIONS: Intratumoral administration of CDDP using the sustained-release drug delivery system is an efficient and safe method to maximize the drug concentration in tumor, minimize the systemic toxicity and enhance antitumor efficacy of irradiation.     

Pharmacokinetic study of doxifluridine given by 5-day stepped-dose infusion

Summary Doxifluridine (5-deoxy-5-fluorouridine, 5-dFUR) metabolism has been reported to be saturable and associated with a fall in clearance of the drug as the dose is increased. The aim of the present study was to determine the disposition of 5-dFUR and 5-fluorouracil (5-FU) when 5-dFUR was given as a 5-day infusion, with the infusion rate increased stepwise every 24 h. Measurement of plasma and urinary levels of 5-dFUR and 5-FU at steadystate for each infusion rate enabled the estimation of 5-dFUR renal (Cl_R) and nonrenal (Cl_NR) clearance and 5-FU renal clearance. A total of 28 patients with histologically proven malignancy received 5-day courses of 5-dFUR ranging in dose from 3.75 to 20 g/m² per 120 h. The lowest dose given over 24 h was 0.25 g/m², and the highest was 5 g/m². Steady-state plasma levels of 5-dFUR ranged from 167 to 6.519 ng/ml. At these plasma levels there was no evidence of significant saturation of 5-dFUR metabolism; steady-state plasma levels of 5-dFUR increased approximately linearly with dose, and nonrenal clearance did not change significantly with dose. There was also no evidence of nonlinearity in 5-dFUR renal clearance. The mean (±SD) Cl_R of 5-dFUR was 108.9±53.6 ml/min per m² (range, 45.7–210 ml/min per m²), and the Cl_NR was 728±181 ml/min per m² (range, 444–1,119 ml/min per m²). Renal clearance comprised 13% of the total 5-dFUR clearance. The mean renal clearance of 5-FU was 100.8±48.6 ml/min per m² (range, 23.5–198 ml/min per m²). There was considerable interpatient variability in 5-dFUR renal and nonrenal clearance, event at the same dose level. We concluded that the administration of 5-dFUR by the infusion method described avoided the saturation of nonrenal elimination processes reported to occur with shorter infusion schedules.This study was supported by a grant from F. Hoffmann-La Roche, Basel, Switzerland     

Effect of atorvastatin on 5-fluorouracil-induced experimental oral mucositis

Purpose

Oral mucositis (OM) is a frequent side effect in patients with cancer. We investigate the effect of atorvastatin (ATV), a cholesterol-lowering drug, on OM induced by 5-fluorouracil (5-FU) in hamsters.

Methods

OM was induced by the i.p. administration of 5-FU, with excoriations of the cheek pouch mucosa. The animals were pretreated with i.p. ATV 1, 5 or 10?mg/kg or vehicle (saline and 5% (vol/vol) ethanol) 30?min before 5-FU injection and daily for 5 or 10?days. Samples of cheek pouches and main organs were removed for histopathological analysis, determination of TNF-??, IL-1??, nitrite, non-protein sulfhydryl group (NP-SH) levels, myeloperoxidase (MPO) assay and immunohistochemistry for induced nitric oxide synthase (iNOS). Blood was collected for a leukogram analysis of biochemical parameters and analysis of bacteremia.

Results

ATV at doses of 1 and 5?mg/kg reduced mucosal damage and inflammation, as well as the levels of cytokines, nitrite and myeloperoxidase activity on the 5th and 10th day of OM and immunostaining for iNOS on the 5th day of OM.ATV at 1?mg/kg increased cheek pouch NP-SH when compared to 5-FU groups on the 10th day of OM. The association between ATV 5?mg/kg and 5-FU decreased the survival rate, amplified the leukopenia of animals, increased transaminase serum levels and caused liver lesions. We also detected the presence of Gram-negative bacillus in the blood of 100% of the animals treated with ATV 5?mg/kg?+?5-FU.

Conclusions

Atorvastatin prevented mucosal damage and inflammation associated with 5-FU-induced OM, but the association of a higher dose of ATV with 5-FU induced hepatotoxicity and amplified leukopenia.     

Sequence-dependent cytotoxic effects of the combination of a new nitrosourea,fotemustine, with 5-fluorouracil plus folinic acid

Summary The present study was designed to analyse the cytotoxic effects of the combination of fotemustine with 5-fluorouracil (5-FU) plus folinic acid (FA). Two human tumor cell lines were used; one line was derived from colon cancer (WIDR) and the other, from a non-small-cell lung cancer (CAL 12). Cytotoxic effects were assessed using the MTT (tetrazolium bromide) semi-automated test in 96-well incubation plates. The effects of various drug combinations were evaluated by the isobologram method. The drug combinations tested included fotemustine concentrations of 20, 30, 40, 50 and 70 g/ml. 5-FU concentrations of 5, 15 and 30 g/ml, and a constant FA concentration of 10^–5 m. A total of 180 different experimental conditions were tested. When cells were exposed to fotemustine prior to treatment with 5-FU, the final cytotoxic effects on both cell lines were additive or synergistic in the majority of cases (P<0.001). The 5-FU concentration was a determinant factor that modified the effects of the drug combination from antagonism (at low 5-FU concentrations) to synergism (high 5-FU concentrations;P<0.001). The addition of FA (10^–5 m) resulted in a significant shift towards synergistic associations in both cell lines. Administration of 5-FU prior to treatment with fotemustine caused marked antagonism, which 10^–5 m FA could not significantly shift towards simple additivity.     

Measurement of in vitro cellular pharmacokinetics of 5-fluorouracil in human and rat cancer cell lines and rat hepatocytes using a flow-through system

Summary A flow-throught system was used to study the cellular pharmacokinetics of 5-fluorouracil (5-FU) in four human cell lines (squamous-cell carcinoma HEp-2, colon carcinoma WiDr, hepatoma Hep G2, and breast carcinoma MCF-7) as well as in the rat hepatoma H35 cell line and in freshly isolated rat hepatocytes. The system made it possible to restrict the decrease in the concentration of 5-FU in the medium, to keep the volume in which the metabolites accumulated relatively small, and to study the dynamics of a response during and after a change in the composition of the eluent. Clearance of 5-FU from the eluent was achieved predominantly (>95%) by its catabolism to dihydrofluorouracil in the tumor cell lines and to 2-fluoro--alanine in the hepatocytes. Not only rat hepatocytes but also HEp-2 cells showed relatively high clearance values. A concentration-dependent 5-FU elimination was observed, indicating saturation of 5-FU elimination according to Michaelis-Menten kinetics (K_m 14–22 M). The maximal velocity (V_max) values ranged from 0.025 to 0.13 nmol 5-FU/10⁶ cells per minute. For HEp-2 cells, high-concentration pulse injections of 5-FU, thymine, uridine, or uracil immediately led to a reduction in 5-FU conversion, followed by recovery within 5 min. The flow-through system proved to be adequate for the study of the non-linear pharmacokinetics of 5-FU in different intact cells and for the comparison of various manipulations of these pharmacokinetics.Abbreviations 5-FU 5-fluorouracil - FUR 5-fluorouridine - F-DHU dihydrofluorouracil - F--ala 2-fluoro--alanine - F-UPA -fluoroureidopropionic acid - HEPES 4-(2-hydroethyl)-1-piperazine-ethane sulfonic acid - MEM modified minimal essential medium - HBSS Hanks' balanced salt solution - HPLC high-performance liquid chromatography Supported by grant IKA 87-16 from the Netherlands Cancer Foundation. One author (G. J. P.) is the recipient of a senior research fellowship from the Royal Netherlands Academy of Arts and Sciences (KNAW)     

Modification of the antitumor activity of chemotherapeutic drugs by the hypoxic cytotoxic agent tirapazamine

 Purpose: Preclinical studies were performed to examine the interaction of the hypoxic cell toxin tirapazamine (TPZ), a benzotriazine di-N-oxide, with several chemotherapeutic agents, including carboplatin, cyclophosphamide, doxorubicin, etoposide, 5-fluorouracil (5-FU), taxol, and navelbine. Methods: The modification by TPZ of the antitumor drug activity and the effect of schedule were determined with an in vivo/in vitro clonogenic assay using well-established RIF-1 murine tumors transplanted into C3H mice. Results: Additive, or greater than additive, tumor cell killing was observed when TPZ was combined with carboplatin, cyclophosphamide, doxorubicin, etoposide, 5-FU and taxol. With the exception of 5-FU there were only small, or no, enhancements of the systemic toxicities of the drugs by TPZ. The greatest enhancement of antitumor activity was with carboplatin, with the maximum effectiveness when TPZ was given 2–3 h before the carboplatin. The activity of cyclophosphamide, doxorubicin, etoposide and taxol were most enhanced when TPZ was given 24 h before the drug. Additional investigations with three-drug combination treatments using cisplatin and TPZ with either etoposide or navelbine indicated a substantial therapeutic gain from the addition of TPZ. Conclusions: The data for each of the drugs tested in combination with TPZ, with the exception of 5-FU, indicate that potential clinical benefit may be obtained from therapies combining TPZ with conventional chemotherapy. Received: 6 March 1996 / Accepted: 30 June 1996     

Does cisplatin (CDDP) function as a modulator of 5-fluorouracil (5-FU) antitumor action? A study based on a clinical trial

Objective To clarify whether CDDP acts as a modulator of 5-FU antitumor action in gastric cancer, patients were treated preoperatively with 5-FU+CDDP (FP) chemotherapy.Patients and methods From September 2000 to November 2001 at Takarazuka Municipal Hospital, 29 patients preoperatively diagnosed with stages II–IV gastric cancer were enrolled. Written informed consent was obtained from all patients. The patients were randomly assigned to two groups: the FU group, in which patients received a continuous intravenous infusion of 5-FU 320 mg/m² per day over 24 h a day for 5 days beginning 5 days prior to surgery, and the FP group, in which patients received bolus intravenous injections of CDDP 3.5 mg/m² per day for 5 days prior to surgery in addition to the same infusion of 5-FU as the FU group. As indicators of the intracellular effect of 5-FU treatment, thymidylate synthase (TS) inhibition rates, TS protein levels, TS and dihydropyrimidine dehydrogenase (DPD) activity, and F-RNA concentrations were measured.Results Using Scheffes multiple comparison test, in both treatment groups the tumor regions were found to have significantly higher TS inhibition rates than the nontumor regions (P<0.05). No significant differences in TS protein levels, TS activity, DPD activity or F-RNA concentrations were found between the four regions.Conclusions Our results show that CDDP clinically may act to enhance the antitumor effects of 5-FU in terms of the inhibition of DNA synthesis and could therefore act as a modulator of 5-FU.     

A mathematical model of the kinetics of 5-fluorouracil and its metabolites in cancer patients

Summary A compartmental model of the kinetics of 5-fluorouracil (5-FU) and its catabolites in humans is proposed. This model was developed using data from a previous study in which plasma levels and urinary amounts of unchanged drug and metabolites were quantitated after i.v. bolus injection of 500 mg/m² 5-FU in ten patients. Biliary excretion was also quantified in two subjects. The different processes, biochemical transformations, and urinary and biliary excretion were adequately described by first-order kinetics. The technique of multiresponse modelling was used for global fitting of all data for each patient. Satisfactory agreement was achieved between measured and predicted values. This model enabled accurate avaluation of pharmacokinetic parameters that could not be adequately calculated using a model-free analysis. The total clearance and elimination half-life of 5-FU and its catabolites are reported for all subjects. The estimated mean half-life was 6.9±3.9 min for unchanged 5-FU and 225±352, 7.6±4, and 9.6±7.7 min, respectively, for the three measured catabolites dihydrofluorouracil (FUH₂), -fluoro--ureidopropionic acid (FUPA), and -fluoro--alanine (FBAL). The percentage of anabolic, catabolic, urinary, and biliary elimination in total clearance was also quantitated. Anabolic clearance accounted for 39%±14% of total 5-FU clearance, with substantial variation occurring among patients. Urinary clearance represented 6.5%±3.2%, 0.8%±0.9%, 13.2%±4.7%, and 98.2%±2.5% of total clearance for 5-FU, FUH₂, FUPA, and FBAL, respectively. The model was also satisfactorily fitted to the data of a patient deficient in dihydropyrimidine dehydrogenase, an enzyme previously thought to be the rate-limiting step for 5-FU catabolism. In this case, catabolism was highly reduced and urinary excretion of 5-FU increased up to 64% of total drug clearance. This first global model of the kinetics of 5-FU and all of its catabolites in patients given an i.v. bolus infusion of 500 mg/m² 5-FU represents a further step toward detailed comprehensive modeling of the kinetics of this drug.     

Clinical implications of dihydropyrimidine dehydrogenase (DPD) activity in 5-FU-based chemotherapy: mutations in the <Emphasis Type="Italic">DPD</Emphasis> gene,and DPD inhibitory fluoropyrimidines

Dihydropyrimidine dehydrogenase (DPD) is the rate-limiting enzyme in the degradation of pyrimidine bases. DPD is also responsible for the degradation of 5-fluorouracil (5-FU), which is the most frequently prescribed anticancer drug for the treatment of malignancies of the gastrointestinal tract. DPD could influence the antitumor effect and the adverse effects of 5-FU. High intratumoral DPD activity markedly decreases the cytotoxic effect of 5-FU. More than 80% of administered 5-FU is detoxified and excreted as F--alanine in urine. In 5-FU-based chemotherapy, escape from the degradation catalyzed by DPD is important. Recently, the dihydropyrimidine dehydrogenase gene (DPYD) was isolated, and its physical map and exon-intron organization were determined. To date, many DPYD variant alleles associated with a lack of DPD activity have been identified. In 5-FU-based cancer chemotherapy, severe toxicities were observed at higher rates in patients who were heterozygous for a mutant DPYD allele, compared with toxicities in patients who were homozygous for the wild DPYD allele. Furthermore, the adverse effects of 5-FU are often lethal for patients homozygous for the mutant DPYD allele. The apparently high prevalence of the DPYD mutation associated with lack of DPD activity in the normal population warrants genetic screening for the presence of these mutations in cancer patients before the administration of 5-FU. DPD inhibitory fluoropyrimidines (DIFs), including uracil plus tegafur (UFT) and tegafur plus 5-chloro-2,4-dihydroxypyridine plus potassium oxonate, in a molar ratio of 1:0.4:1 (TS-1), have recently been used in clinical settings. DIFs should provide chemotherapy that improves both quality of life and duration of survival.     

Effect of lonidamine on the cytotoxicity of four alkylating agents in vitro

Summary We examined the ability of lonidamine, which has been described as an inhibitor of cellular respiration and glycolysis, to enhance the cytotoxicity of alkylating agents to MCF-7 human breast-carcinoma cells. Lonidamine was increasingly cytotoxic to MCF-7 cells with increasing time of exposure. With a 12-h exposure, the IC₅₀ for lonidamine was about 365 M, and with a 24-h exposure it was about 170 M. A drug concentration of 250 M was chosen for use in the drug combination studies. Lonidamine appeared to have a dose-modifying effect on cisplatin (CDDP), producing increasingly supraadditive cell kill with increasing CDDP concentration. When simultaneously incubated with lonidamine for 1 h, 500 M CDDP yielded a cell kill that was 2 log greater than additive cytotoxicity. Extending the exposure to lonidamine for 12 h after CDDP treatment led to a small, additional aliquot of cell kill of about 2.5-fold over the CDDP concentration range. Lonidamine also appeared to have a dose-modifying effect on melphalan cytotoxicity in the melphalan concentration range of 100–500 M. Between concentrations of 10 and 100 M melphalan, the drug combination survival after 1 h exposure fell within the envelope of additivity for the two agents. However, maintaining the presence of lonidamine for an additional 12 h increased the effect such that the combination was supraadditive over the entire concentration range of melphalan. Simultaneous exposure to 4-hydroperoxycyclophosphamide (4-HC) and lonidamine for 1 h resulted in greater than additive cell kill, and extending the lonidamine exposure period such that lonidamine was present during and 12 h after 4-HC treatment further increased this effect. Lonidamine had a moderate effect on the cytotoxicity of carmustine (BCNU) with a 1 h simultaneous exposure; however, this treatment combination reached greater than additive cytotoxicity only at the highest concentration of BCNU tested. Extending the lonidamine exposure time for an additional 12 h resulted in supraadditive cell kill over the BCNU concentration range. Therefore, when lonidamine was present during exposure to the alkylating agent and its presence was then extended for an additional 12 h, a synergistic cell kill was produced with all four alkylating agents tested.This work was supported by a grant from DeSanctis Consultants, Montreal, Canada and National Cancer Institute Grant IPOI-CA38493     

5-Fluorouracil-metronidazole combination therapy in metastatic colorectal cancer

Summary We have investigated the role of metronidazole (MND) combined with 5-fluorouracil (5-FU) in the treatment of metastatic colorectal cancer. MND (750 mg/m²) was administered i.v. 1 h before 5-FU (600 mg/m²) i.v., daily for 5 consecutive days. Treatment was repeated every 4 weeks until disease progression or prohibitive toxicity occurred. Of the 27 patients entered in the study, 4 (15%) had an objective complete or partial response lasting an average of 7 months. 5-FU toxicity was greatly enhanced by the administration of MND, however, 74% of patients having granulocytopenia (<1500/l). We investigated the possible mechanisms underlying this enhanced 5-FU toxicity by examining whether MND modified 5-FU pharmacokinetics or whether the two drugs had a synergistic effect in vitro against the HCT-8 colon cancer cell line. While the in vitro studies failed to reveal any synergism between 5-FU and MND, pharmacokinetic evaluation revealed that 5-FU clearance was significantly reduced (26.9%, P<0.001) by prior MND administration. MND reduces 5-FU's therapeutic index in the treatment of colorectal cancer by impairing its clearance, which leads to increased toxicity without enhanced therapeutic efficacy.     

Synergistic enhancement by interleukin-1 α of cisplatin-mediated antitumor activity in RIF-1 tumor-bearing C3H/HeJ mice

Administration of interleukin-1 (IL-1 ) plus certain cytotoxic drugs causes substantially greater clonogenic tumor-cell kill and tumor-regrowth delay than does treatment with either agent alone. IL-1 itself has little effect on tumor growth despite its ability to induce acute hemorrhagic necrosis, restrict tumor blood flow, and cause microvascular injury in a variety of murine model systems. To investigate further IL-1 's ability to enhance the antitumor activity of cytotoxic drugs, we initiated studies to examine the effect of IL-1 on cisplatin (cDDP)-mediated cytotoxicity using the RIF-1 tumor system. The antitumor activity of IL-1 and cDDP was quantitated through standard clonogenic tumor-cell survival assays, a tumor hemorrhagic necrosis assay and tumor-regrowth delay studies, with the interaction between IL-1 and cDDP being analyzed through median dose-effect. In vitro, IL-1 had no enhancing effect on the cDDP-mediated tumorcell kill. For examination of the in vivo efficacy of this regimen. RIF-1 tumor-bearing C3H/HeJ mice (14 days postimplantation) were treated concurrently with single i.p. injections of IL-1 and/or cDDP at various doses. The increased clonogenic tumor-cell kill obtained with IL-1 /cDDP was dose-dependent, with significant enhancement by IL-1 being observed (P<0.001), even at the lowest doses tested (2 mg/kg and 6 g/kg for cDDP and IL-1 , respectively), but it did not correlate with an increase in tumor hemorrhage. Using median dose-effect analysis, this interaction was determined to be strongly synergistic. When treated animals were monitored for long-term antitumor effects, combinations with IL-1 significantly increased the tumor-regrowth delay and decreased the fractional tumor volume (P<0.001). These results demonstrate that IL-1 synergistically enhances cDDP mediated in vivo antitumor activity and suggest that the combination of IL-1 and cDDP may have potential therapeutic application in the design of effective treatment modalities for cancer.Abbreviations IL-1 interleukin-1 - cDDP cis-diamminedichloroplatinum (cisplatin) - BRMs biological response modifiers - TNF tumor necrosis factor - IFN interferon - Fa traction affected - D_m or ED₅₀ drug concentration necessary to produce Fa=0.5 as compared with untreated controls - CI combination index; SF, surviving fraction - ANOVA oneway analysis of variance This work was supported in part by Public Health Service grant CA-48077 from the National Cancer Institute, National Institutes of Health, Department of Health and Human Services, by the Mary Hillman Jennings Foundation, and by the Ramona DeSantis Cancer Research Fund     

Taxol and taxotere in bladder cancer: in vitro activity and urine stability

In this study the antimicrotubular agents taxol, taxotere, and vinblastine were compared for their ability to inhibit the clonal growth of human bladder tumor cell lines using a soft-agar clonogenic assay. The stability of taxol and taxotere was evaluated by high-performance liquid chromatography over a range of pH in human urine. Both taxol and taxotere were shown to maximally inhibit the clonal growth of human bladder cell lines within 1 h of drug incubation. The most active agent in the panel of tumor lines was taxotere, with 6 of 12 lines being sensitive to the agent at 0.01 M and all cell lines being sensitive at 0.1 M. Taxol was active in 1 of 12 lines at 0.01 M and in 11 of 12 at 0.1 M. Only 2 of 12 cell lines were sensitive to vinblastine over the 0.01- to 0.1-M dose range. Taxol and taxotere were found to be stable in human urine for 4 h over a pH range of 5–7. At least 85% of both drugs were present during this period of drug incubation. Our findings suggest that both taxol and taxotere may be clinically useful agents for systemic and intravesical use in bladder cancer.This work was supported in part by the Veterans Administration Research Service     

Pharmacokinetics and bioequivalence of a combined oral formulation of eniluracil, an inactivator of dihydropyrimidine dehydrogenase, and 5-fluorouracil in patients with advanced solid malignancies

Background:This study was performed to evaluate thepharmacokinetics, bioequivalence, and feasibility of a combined oralformulation of 5-flurouracil (5-FU) and eniluracil (Glaxo Wellcome Inc.,Research Triangle Park, North Carolina), an inactivator of dihydropyrimidinedehydrogenase (DPD). The rationale for developing a combined eniluracil/5-FUformulation oral dosing form is to simplify treatment with these agents, whichhas been performed using separate dosing forms, and decrease the probabilityof severe toxicity and/or suboptimal therapeutic results caused byinadvertently high or conversely insufficient 5-FU dosing. Patients and methods:The trial was a randomized, three-waycrossover bioequivalence study of three oral dosing forms of eniluracil/5-FUtablets in adults with solid malignancies. Each period consisted of two daysof treatment and a five- to seven-day washout phase. Eniluracil at a dose of20 mg, which results in maximal DPD inactivation, was administered twice dailyon the first day and in the evening on the second day of each of the threetreatments. On the morning of the second day, all patients received a totaleniluracil dose of 20 mg orally and a total 5-FU dose of 2 mg orally as eitherseparate tablets (treatment A) or combined eniluracil/5-FU tablets in twodifferent strengths (2 tablets of eniluracil/5-FU at a strength (mg/mg) of10/1 (treatment B) or 8 tablets at a strength of 2.5/0.25 (treatment C)). Thepharmacokinetics of plasma 5-FU, eniluracil, and uracil, and the urinaryexcretion of eniluracil, 5-FU, uracil, and -fluoro--alanine (FBAL),were studied. To determine the bioequivalence of the combined eniluracil/5-FUdosing forms compared to the separate tablets, an analysis of variance onpharmacokinetic parameters reflecting eniluracil and 5-FU exposure wasperformed. Results:Thirty-nine patients with advanced solid malignancies hadcomplete pharmacokinetic studies performed during treatments A, B, and C. Thepharmacokinetics of eniluracil and 5-FU were similar among the three types oftreatment. Both strengths of the combined eniluracil/5-FU dosing form and theseparate dosing forms were bioequivalent. Mean values for terminal half-life,systemic clearance, and apparent volume of distribution for oral 5-FU duringtreatments A/B/C were 5.5/5.6/5.6 hours, 6.6/6.6/6.5 liters/hour, and50.7/51.5/50.0 liters, respectively. The intersubject coefficient of variationfor pharmacokinetic variables reflecting 5-FU exposure and clearance intreatments ranged from 23% to 33%. The urinary excretion ofunchanged 5-FU over 24 hours following treatments A, B, and C averaged52.2%, 56.1%, and 50.8% of the administered dose of 5-FU,respectively. Parameters reflecting DPD inhibition, including plasma uraciland urinary FBAL excretion following treatments A, B, and C were similar.Toxicity was generally mild and similar following all three types oftreatments. Conclusions:The pharmacokinetics of 5-FU and eniluracil weresimilar and met bioequivalence criteria following treatment with the separateoral formulations of 5-FU and eniluracil and two strengths of the combinedformulation. The availability of a combined eniluracil/5-FU oral dosing formwill likely simplify dosing and decrease the probability of severe toxicityor suboptimal therapeutic results caused by an inadvertent 5-FU overdose orinsufficient 5-FU dosing in the case of separate oral formulations, therebyenhancing the overall feasibility and therapeutic index of oral 5-FU therapy.     

Postload plasma glucose concentration and 27-year prostate cancer mortality (United States)

Objective: Findings from epidemiologic studies on the association between diabetes and prostate cancer risk are inconsistent. However, data from at least three studies suggest that the direction and strength of this association differs according to duration of diabetes. To determine the potential effects of early-stage abnormal glucose metabolism on risk, we assessed the relationship of postload glycemia in the absence of self-reported diabetes with risk of prostate cancer mortality. Methods: Data from the Chicago Heart Association Detection Project in Industry were used to examine this relationship. Between 1967 and 1973 some employees of 84 Chicago area organizations underwent a health screening examination. Blood was drawn for measurement of plasma glucose concentration 1 h after a 50-g oral glucose load among 20,433 men. After a mean length of follow-up of 27 years, 176 men died of prostate cancer. Cox regression was used to compute adjusted relative risks (RRs) and 95% confidence intervals (CIs). Results: After controlling for age, body mass index, heart rate, education, and race, the RRs of prostate cancer mortality for postload plasma glucose levels of 6.7–8.8, 8.9–11, and 11.1 mmol/L compared to 6.6 mmol/L were 1.64, 1.37, and 1.64, respectively (p for trend = 0.19). The RR (95% CI) associated with a 2.2 mmol/L (1 standard deviation) higher glucose concentration was 1.1 (0.95–1.2). Conclusions: These results provide weak evidence of an association between hyperglycemia and prostate cancer mortality.