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Phase I Clinical and Pharmacologic Study of Chronic Oral Administration of the Farnesyl Protein Transferase Inhibitor R115777 in Advanced Cancer

From the Netherlands Cancer Institute and Academic Medical Centre, Amsterdam; Division of Drug Toxicology, Faculty of Pharmacy, Utrecht University, Utrecht, the Netherlands; and Janssen Research Foundation, Beerse, Belgium.

Address reprint requests to M. Crul, MSc, Department of Pharmacy and Pharmacology, the Netherlands Cancer Institute/Slotervaart Hospital, Louwesweg 6, 1066 EC Amsterdam, the Netherlands; email: apmcr{at}slz.nl

	ABSTRACT

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
PURPOSE: To determine the maximum-tolerated dose, toxicities, and pharmacokinetics of R115777, a farnesyl transferase inhibitor, when administered continuously via the oral route.

PATIENTS AND METHODS: Patients with advanced solid malignancies were treated with R115777 using an interpatient dose escalation scheme starting at 50 mg bid. Pharmacokinetics were assessed on days 1, 28, and 56.

RESULTS: Twenty-eight patients were entered onto the study and the median duration of treatment was 55 days. The dose-limiting toxicities were myelosuppression and neurotoxicity. At a dose of 400 mg bid, grade 4 leukocytopenia and neutropenia were seen in two of four patients. Neurotoxicity grade 3 developed in one of five patients at 500 mg bid and in one of 13 at 300 mg bid after 8 weeks of treatment. Common nonhematologic toxicities were nausea, vomiting, and fatigue. The recommended dose for phase II/III testing in this scheme is 300 mg bid. The pharmacokinetic studies indicated dose proportionality. Little accumulation occurred and steady-state levels were reached within 2 to 3 days. Analyses of historic tumor material showed that five of 15 of patients had a K-ras mutation in codon 12. Three patients with pancreatic, colon, and cervix carcinomas had stable disease and one patient with a colon carcinoma had a minor response accompanied by a more than 50% decrease in carcinoembryonic antigen tumor marker. A fifth patient, with platinum-refractory non–small-cell lung cancer, showed a partial response that lasted for 5 months.

CONCLUSION: Continuous dosing of R115777 is feasible with an acceptable toxicity profile at a dose of 300 mg bid.

	INTRODUCTION

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
THE DEVELOPMENT of human cancer is thought to be the result of mutations in multiple genes that control normal cell proliferation, differentiation, and apoptosis. Amongst others, the ras oncogene has attracted much attention, because it is one of the genes most commonly found mutated in human tumors. Overall, mutated ras exists in 20% to 30% of all cancers, with the highest frequencies found in adenocarcinomas of the pancreas (90%), colon (50%), and lung (30%).^1,2 Ras proteins transduce growth and differentiation signals from receptor tyrosine kinases to the cell nucleus, thereby initiating gene transcription.³ In order to function, they must be farnesylated by the enzyme farnesyl protein transferase (FTase), which is the first and critical step in their posttranslational modification.^4,5 Some Ras proteins can be prenylated by geranylgeranyl transferase as well, particularly when FTase is pharmacologically inhibited.^6,7 Mutated Ras remains in its active state longer than wild-type Ras, the consequence of which is that the ras-driven cell proliferation is continuously (over)activated, possibly resulting in tumor formation.⁸ Through blocking the mutated ras gene product, such malignant transformation can be reversed.^9,10 For this aim, many different farnesyl protein transferase inhibitors (FTIs) have been developed that block the posttranslational modification of Ras, thereby preventing its activation. Evidence of the clinical utility of FTIs is rapidly increasing.^11-13 At the same time, the exact mechanism underlying the antineoplastic effect of these compounds is being unraveled. Several observations suggest that farnesylated proteins other than Ras may contribute to, or be even more important in, the mechanism of action of these drugs.^9,14 The protein RhoB, which can be either farnesylated or geranylgeranylated, is an especially likely candidate. Rho proteins regulate cytoskeletal actin organization, adhesion, and proliferation.¹⁵ As a result of treatment with FTIs, levels of geranylgeranylated RhoB increase, which in turn induces an elevation of p21^Waf1/Cip1 levels. p21^Waf1/Cip1 is a well-known cell-cycle kinase inhibitor. Furthermore, geranylgeranylated RhoB is most likely involved in induction of apoptosis.¹⁶ At least 17 other mammalian proteins require prenylation to become activated, including the nuclear lamins A and B, members of the Rac family, and several proteins involved in retinal signal transduction.¹⁷ To determine exactly which of the farnesylated proteins play a role in the mechanism of action of FTIs, further research is warranted. By blocking the activation of farnesylated proteins, FTIs interfere with growth signal transduction. Drugs that act through such a mechanism are probably more cytostatic than cytotoxic. Hence, on theoretical grounds, prolonged and continuous exposure to such an agent would provide the best results of treatment.

R115777 is a nonpeptidomimetic FTase inhibitor that can be administered orally. It is a quinolone analog of imidazole with the full name (B)-6-[amino(4-chlorophenyl)(1-methyl-1H-imidazol-5-yl)methyl]-4-(3-chlorophenyl)-1-methyl-2(1H)-quinolone. It has demonstrated good antineoplastic activity in both in vitro and in vivo studies (Janssen Research Foundation, Beerse, Belgium, data on file, Investigator’s Brochure, April 2001).¹⁸ The results of a phase I study with R115777 in patients with advanced cancer have been reported recently, using an intermittent schedule of 5 days of treatment followed by 7 days of rest. Observed toxicities included nausea, vomiting, headache, fatigue, anemia, and hypotension, whereas myelosuppression was rare. The recommended dose for this schedule for phase II trials of R115777 is 500 mg bid.¹⁹ Another phase I trial administered R115777 to patients with advanced leukemias for 21 days followed by 7 days of rest. Here, neurotoxicity was dose-limiting at 1,200 mg bid. Responses were observed in 29% of patients, and the advised dose for this schedule was 600 mg bid.¹³

Here, we present our results from a phase I and pharmacokinetic trial of R115777 using a continuous dosing scheme. The objectives of this study were as follows: (1) to determine the maximum-tolerated dose (MTD) of daily oral administration of R115777, (2) to characterize the dose-limiting toxicities (DLTs) associated with this regimen, and (3) to investigate the clinical pharmacology of R115777. Another aim was to assess the ras mutational status of the tumors from all treated patients and to explore possible relationships with activity of the treatment. Finally, a possible influence of R115777 treatment on lymphocyte proliferation and differentiation was investigated.

	PATIENTS AND METHODS

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Patient Population
Patients were eligible if they had a histologically or cytologically confirmed diagnosis of a solid malignant tumor not amenable to established forms of effective therapy. Other eligibility criteria included a World Health Organization performance status of 0 to 2, anticipated life expectancy of at least 3 months, and age 18 years. Previous anticancer chemotherapy had to be discontinued for at least 4 weeks before entry onto the study, or 6 weeks in case of pretreatment with nitrosourea or mitomycin. Radiation therapy should have ended at least 4 weeks before study entry. All patients had to have acceptable bone marrow function, defined by a WBC count more than 3,500/µL and a platelet count more than 100,000/µL, normal serum bilirubin, serum creatinine within 1.5 times the normal upper limit, and AST and ALT two times the normal upper limit or five times the normal upper limit in case of hepatic metastases. The study protocol was approved by the medical ethics committee of the hospital, and all patients had to give written informed consent.

Treatment Plan and Study Design
R115777 was supplied by Janssen Research Foundation (Beerse, Belgium) as 50- and 100-mg capsules. The colored hard gelatin capsules of 50 mg and 100 mg have the same qualitative composition and a dose-proportional quantitative composition. The capsules contained R115777, sugar spheres, hypromellose, and macrogol. All inactive ingredients were standard pharmaceutical excipients that comply with pharmacopeial monographs. The drug was given with or shortly after a meal. Morning and evening drug intakes were separated by intervals of 12 hours. The starting dose of R115777 was 50 mg bid. Dose escalation was performed with increments of 50 to 100 mg; within-patient dose escalation was not allowed. All patients received treatment at their assigned dose continuously, until disease progression or unacceptable toxicity intervened. At least one patient was treated at each dose level. If no evidence of cumulative toxicity and no DLTs were observed in the first 3 weeks, the dosage was escalated to the next level for the next eligible patient. The observation of evidence of cumulative toxicity that resulted in DLT with prolonged drug exposure led to the addition of two additional patients at that dose level. If accrual at the next dose level had already started, this was halted. If no DLT occurred in the next 21 days of treatment of the cohort of three patients, dose escalation was continued in the next patient. The occurrence of DLT in a patient led to the enrollment of up to five additional patients in that cohort. The dose escalation was suspended until the additional patients received treatment for at least 6 weeks at the relevant dose without additional DLT. Once the MTD had been exceeded, up to five additional patients were enrolled at the preceding dose level to ensure six patients in that cohort. Ultimately, a minimum of six patients were to be treated at the MTD for at least 6 weeks.

Patient Evaluation
Pretreatment evaluation included a complete medical history and complete physical examination. Indicator lesions were measured before start of treatment and every two cycles during the study, as a basis for the assessment of efficacy of the treatment. Because the retina contains several proteins that require farnesylation to function, ophthalmic examination was performed at baseline, and monthly thereafter. This included ocular history, best visual acuity, ocular examination including dilatation, visual field screening, D-15 color testing, contrast sensitivity testing performed by means of Pelli-Robson charts, and, if indicated, fundus photographs. In addition, electro-oculography was performed, and if indicated, this was followed by electroretinography. All toxicities observed were graded according to the common toxicity criteria (CTC).²⁰ DLT was defined as grade 3 toxicity, with the exception of neurotoxicity (for which the limit defining toxicity was grade 2 severity). A required interruption of treatment more than 2 weeks because of toxicity also defined DLT. The MTD was taken at one dose level below the level at which two or more of six patients experienced DLT.

Pharmacokinetics
Plasma concentrations of R115777 were determined by a validated high-performance liquid chromatography method. In brief, samples were alkalinized with sodium hydroxide 0.1 mol/L, extracted with heptane-isoamyl alcohol (90:10 vol/vol), and analyzed on a C18 BDS-Hypersil column (10 cm x 4.6 mm, particle size 3 µmol/L; Hypersil, Cheshire, United Kingdom). The mobile phase consisted of ammonium acetate-acetonitrile 0.01 mol/L (52:48), and the peaks were quantified by ultraviolet detection at 240 nm. On days 1, 28, and 56, a predose sample was drawn immediately before administration of study medication. On those days, additional blood samples for the determination of R115777 plasma concentrations were drawn at 1, 2, 4, 6, 8, and 12 hours after the morning dose (the last sample at 12 hours was taken immediately before the evening dose). On days 2, 4, and 8, a predose sample was drawn immediately before the morning intake of study medication. At least 6 mL of heparinized blood was collected for each sample. Samples were placed on ice immediately and centrifuged within 2 hours after collection (10 minutes, at 1,000 x g or 2,500 rpm). Separated plasma was transferred to polyethylene cups and frozen by immersion in a dry ice/ethanol bath and stored at -20°C for subsequent drug analysis. The following pharmacokinetic parameters were determined by either direct observation or calculation from the R115777 plasma concentrations: the time to reach the maximum concentration (t_max), the maximum plasma concentration (C_max), the area under the plasma concentration-time curve from 0 to 12 hours (AUC_0-12h), the average concentration calculated by the AUC_0-12h divided by 12 (C_avg), and the predose or trough plasma concentration (C_min) obtained just before the intake of medication. For calculation of AUCs, a noncompartmental model was used. The predose concentrations in the morning of days 2, 4, and 8 were used for evaluation of steady-state conditions. The pharmacokinetic parameters AUC_0-12h, C_max, and C_avg were plotted versus dose to visually evaluate dose proportionality. Furthermore, the apparent oral clearance was calculated per patient by dividing the dose by the AUC_0-12h. Subsequently, these last data were tested for absence of correlation with dose. Differences in pharmacokinetic parameters on days 1, 28, and 56 were evaluated using the paired Student’s t test. The level of significance was set at P < .05. All statistical analyses were performed using the software package SPSS for Windows (Version 6.1, SPSS, Inc, Chicago, IL).

Pharmacokinetic-Pharmacodynamic Analysis
Relationships between systemic exposure and pharmacodynamics were investigated using scatterplots of the AUC_12h versus the maximal percentage decrease in WBC count, absolute neutrophil count (ANC), and thrombocytes per patient. The data were fit using a sigmoidal maximum effect (E_max) model, as described by the Hill equation²¹:

equation

where E_max is the maximal effect, DE is a measure of drug exposure (in this case, AUC), DE₅₀ is the drug exposure at 50% of E_max, and y is the Hill coefficient, describing the sigmoidicity of the curve.

K-ras Mutation Analysis
From each patient, representative formalin-fixed paraffin-embedded tumor tissue blocks were collected from referring hospitals or from the files of the pathology laboratory of the Netherlands Cancer Institute. All available material was reviewed and tested for K-ras point mutations within codons 12 and 13,^22,23 representing the majority of all K-ras mutations.²⁴ From each paraffin-embedded tumor sample, 5-µm sections were collected in 1.5-mL Eppendorf vials. An additional 5-µm section stained with hematoxylin and eosin was used to classify the tumor and to determine the percentage of tumor cells in the specimen. To increase the percentage of tumor cells in each sample, the areas with the highest density of tumor cells were selected using the hematoxylin and eosin–stained section as a guide. These tumor cell–rich areas were used for DNA isolation. DNA was isolated from the tumor sections in 200 µL of lysis buffer (potassium chloride 50 mmol/L, Tris-HCl 10 mmol/L [pH 8.0], magnesium chloride 2.5 mmol/L, gelatin 0.1 mg/mL, 0.45% Tween, and proteinase K 10 µg/mL). Remaining debris was removed by centrifugation. Five microliters of the supernatant was used for the polymerase chain reaction. Amplification with K-ras–specific primers encompassing codons 12 and 13 included 35 cycles of denaturation at 94°C for 30 seconds, annealing at 54°C for 30 seconds, and extension at 72°C for 60 seconds in a Dri-block device (Techne, Cambridge, United Kingdom) with 1 unit of Taq polymerase (Perkin Elmer Cetus, Emeryville, CA) in a standard buffer (Tris-HCL10 mmol/L [pH 8.3], potassium chloride 50 mmol/L, magnesium chloride 2.5 mmol/L, and deoxynucleoside triphosphate 0.1 µmol/L). In addition, a mutant-enriched amplification for K-ras 12 was performed as described previously.¹⁹ From each reaction, 1 µL of the polymerase chain reaction product was spotted on nylon membranes (Hybon, Amersham, United Kingdom) for hybridization analysis. Point mutations were detected by hybridization with P32-labeled mutation-specific oligonucleotides and visualized by x-ray autoradiography films.

Immunologic Monitoring
Blood samples were obtained at baseline and every 4 weeks during treatment. For each sample, 10 mL of whole blood was collected in EDTA. WBCs were isolated by lysis of red cells with ammonium chloride. Triple staining was performed as previously described²⁵ by the incubation of cell samples in a mixture of fluorescein R-phycoerythrin (PE), and PE-cychrome (Cy5)–conjugated mouse monoclonal antibodies, directed to cellular differentiation markers, for 20 minutes at room temperature followed by washing in phosphate-buffered saline containing 0.2% bovine serum albumin and 0.02% azide. Fluorescein and PE conjugates were obtained from Becton Dickinson Immunocytometry Systems (San Jose, CA) and CyChrome conjugates were obtained from Coulter Immunotech (Mijdrecht, the Netherlands). To prevent aspecific binding by means of Fc receptors, incubation was carried out in the presence of 1% normal mouse serum (Central Laboratory of the Netherlands Red Cross Blood Transfusion Service, Amsterdam, the Netherlands). Fluorochrome-labeled isotype control antibodies were included in each assay to determine background staining. Lymphocyte (CD3⁺, CD19⁺, and CD16⁺56⁺), and monocyte (CD14⁺) populations were gated according to their differential forward and side scatter, which was confirmed by back-gating. Fluorescence was measured on a FACScan flow cytometer (Becton Dickinson, Mountain View, CA) and data were analyzed using CellQuest software (Becton Dickinson). Differences in levels of circulating lymphocytes at different time points were compared by Wilcoxon’s rank test for paired data. P values less than .05 were considered significant.

	RESULTS

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

 
Patient Characteristics
Twenty-eight patients were included on the study, 16 men and 12 women with a median age of 55.5 years. Twenty-seven patients (96%) had a performance status of 0 or 1, and one patient had a performance status of 2. All but four patients received prior cytotoxic therapies. Additional patient characteristics are listed in Table 1. Initially, one patient was included per dose level. At dose level 3 (150 mg bid), the first included patient developed exanthema, which recurred after a rechallenge with R115777 treatment. Even though this was not a DLT by definition, two extra patients were included at this dose level. No further significant toxicities were observed in the additional patients and therefore normal dose escalation was resumed at the next level of 200 mg bid. The median duration of treatment was 55 days. In total, six patients went off study during or at the end of the first month of treatment, because of progressive disease or because of patients’ refusal to continue treatment. Seven patients remained on treatment for 1 to 2 months, seven for 2 to 3 months, and six for 3 to 4 months. Finally, two patients remained on study for 5 and 7 months, respectively. They were treated at dose levels 6 (400 mg bid) and 5 (300 mg bid). Seven patients required a dose reduction after initial inclusion, and in three additional patients the course was interrupted for 1 to 2 weeks because of side effects. In general, treatment interruptions did not exceed 20 days, and toxicity did not recur on restart. The dose-escalation schedule and the number of patients included at each dose level are listed in Table 2.

View larger version (16K): [in this window] [in a new window]   Fig 1. Leukocyte levels of three patients (, patient no. 7; , patient no. 8; , patient no. 19) treated at 300 mg bid during the first 2 months of treatment.

Because several farnesylated proteins are involved in retinal signal transduction, we monitored all patients for ophthalmologic abnormalities. In only one patient (with no history of ocular disease), an asymptomatic decrease of electroretinography values was found after 1 month of treatment. In particular, the scotopic B waves had decreased from 57 µV to 26 µV in the right eye, and from 57 µV to 25 µV in the left. Also, the photopic B waves showed a slight decrease: from 64 µV to 52 µV and from 76 µV to 50 µV, in the right eye and left eye, respectively. This patient was initially included on the study at dose level 7 (500 mg bid) but developed leukocytopenia after 2 weeks, necessitating an interruption of treatment. On recovery, treatment was resumed at a lower dose (300 mg bid). At the next assessment (2 months after study entry and 1 month after the dose reduction), all electroretinography values were in the normal range.

Antitumor Response
Twenty-eight patients were treated on this phase I trial. Six were withdrawn from the study before completing 6 weeks of medication, because of adverse events (three patients, two with DLT) or early progression (three patients). These patients were not evaluated for antitumor response. Of the remaining 22, one patient with platinum-refractory metastatic non–small-cell lung cancer showed a partial response (75% decrease in tumor size), which lasted for 5 months. In addition, a patient with colon carcinoma, which was both fluorouracil- and irinotecan-resistant, had a minor response of large liver metastases and a more than 50% decrease in CEA tumor marker. Finally, three patients had stable disease lasting for 2 to 3 months confirmed by computed tomography scans. One of these patients had a pancreatic tumor, the second had a colon tumor, and the third had a cervix carcinoma. The patient with the colon carcinoma also had a CEA marker decrease of nearly 50% (from 220 µg/L to 128 µg/L).

Pharmacokinetics
Blood samples for pharmacokinetic analyses were obtained from all patients. After a single dose of R115777, peak plasma concentrations were obtained after 2 to 4 hours. C_max, AUC_0-12h, and C_avg increased with increasing doses, which appeared to occur linearly on visual inspection. However, interpatient variability was considerable. In addition, the apparent clearance showed no correlation with the dose (correlation coefficient < .1). The pharmacokinetic parameters according to dose level are summarized in Table 4. Little accumulation occurred: comparing pharmacokinetics per patient on days 1, 28, and 56 yielded accumulation ratios of the AUCs close to 1. The predose concentrations determined on days 2, 4, and 8 demonstrated that steady-state concentrations were reached quickly (within 2 to 3 days) and maintained throughout the dosing period. On days 28 and 56, trough levels (C_min) were determined as well. On day 28, the values ranged from 9.8 to 254 ng/mL and on day 56 from 15.4 to 240 ng/mL. Intrapatient variation in AUC was limited (Fig 2).

View this table: [in this window] [in a new window]   Table 4.  Pharmacokinetic Parameters of R115777 Represented as Means ± SD (SDs only represented if n 3)

View larger version (17K): [in this window] [in a new window]   Fig 2. AUC12h on day 1 versus day 28 ( or day 56 (). The dotted line represents the line of identity.

View larger version (16K): [in this window] [in a new window]   Fig 3. The maximal percentage decrease in WBCs ( and ANCs () versus the AUC12h of R115777. The solid line indicates the best fit of the data to the sigmoidal maximum effect pharmacodynamic model.

	DISCUSSION

TOP ABSTRACT INTRODUCTION PATIENTS AND METHODS RESULTS DISCUSSION REFERENCES

Because lymphopenia was seen as a significant side effect in preclinical studies,¹⁸ and because Ras plays an important role in the activation of T cells and NK cells,²⁶ we hypothesized that the number of circulating lymphocytes might be influenced by R115777 treatment. The results of our immunologic monitoring are difficult to interpret in view of the significant lymphocytopenia at baseline in the majority of patients. The absence of opportunistic infections is noteworthy. The observed effects on cytotoxic T cells and NK cells could be a consequence of treatment with R115777, prior extensive chemotherapy, the underlying disease, or any combination of such factors. The clinical significance of these findings cannot be fully addressed in the context of this study. In a previous phase I trial, with R115777 applied as single agent five times daily every 2 weeks, no immunosuppressive activity of the treatment could be detected.¹⁹ Hence, this effect may be schedule-related and warrants additional investigation in future studies.

The pharmacokinetic studies showed a positive correlation between R115777 dose and the parameters AUC, C_max, and C_avg on visual examination of the data. However, interpatient variability was considerable. This has been demonstrated as well in another trial, where R115777 was administered to patients with leukemia in doses ranging from 100 to 1,200 mg bid for up to 21 days.¹¹ In our trial, the trough levels measured on days 2, 4, 8, 28, and 56 demonstrated that steady-state levels were reached quickly and maintained throughout the treatment period. Within the 50- to 500-mg dose range, the steady-state trough levels ranged between 9.8 and 254 ng/mL. In vitro, R115777 inhibits proliferation of tumor cells with median inhibitory concentrations between 1.7 and 22 nmol/L, which corresponds to 0.8 to 10.8 ng/mL.¹⁸ Considering a plasma protein binding of 99% (Janssen Research Foundation, data on file, Investigator’s Brochure, April 2001), these in vitro concentrations correspond to plasma concentrations of 123 and 1,636 ng/mL. Hence, the trough and peak plasma levels obtained in our study are within the active range. Furthermore, the R115777 AUC_12h could be adequately correlated to the extent of myelosuppression with a sigmoidal E_max model. The estimated DE₅₀ levels for the myelosuppressive effect represent the AUCs that will lower the pretreatment baseline WBC count and ANC by 50%.²¹ These DE₅₀s correspond to average daily concentrations of 450 and 350 ng/mL, respectively, both also within the in vitro bioactive concentration range. When a larger number of patients has been treated, it would be of in-terest to investigate the pharmacokinetics of R115777 more extensively with regard to the influence of demographic factors such as age, weight, and patho-logic characteristics such as liver function and their influence on the kinetics. Furthermore, it would be of interest to study the metabolism and excretion of R115777 in cancer patients.

The analyses of K-ras mutations showed that five of the tumor biopsy specimens that were investigated harbored a mutant gene. Either the first or the second guanine base in the GGT sequence at codon 12 was altered, which results in a loss of the glycine normally present in this position. All of the observed cases (GGT to GCT, GGT to GAT, GGT to GTT, and GGT to AGT) lead to an activated form of the Ras protein.¹ All patients with stable disease on R115777 treatment had wild-type ras. Two of these had clearly been progressive on previous therapy. In preclinical experiments, it has been shown that a ras mutation is not a prerequisite for response to FTI treatment.¹⁴ Here, we present the first indication in humans that tumors with wild-type ras could be susceptible to R115777.

In conclusion, we have demonstrated that chronic oral dosing with R115777 is feasible. The recommended dose of this schedule is 300 mg bid. The hints of antitumor effects observed on this phase I trial especially warrant more extensive phase II and phase III clinical testing.

	ACKNOWLEDGMENTS

 
We thank Johan Sein and Colette van den Bogaard for their assistance in the immunology assays.

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Submitted September 25, 2001; accepted March 1, 2002.

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