A Phase I and Pharmacokinetic Study of 1843 U 89 , a Noncompetitive Inhibitor of Thymidylate Synthase , in Patients with Advanced Solid Malignancies 1

This study was performed to assess the feasibility of administering 1843U89, a potent, noncompetitive inhibitor of thymidylate synthase that does not require polyglutamation for activity, as a 2-min i.v. infusion daily for 5 days every 3 weeks, to determine whether folic acid supplementation ameliorates the toxic effects of 1843U89 and permits further dose escalation, and to recommend doses of 1843U89 administered without and with folic acid for further clinical evaluations. The study also sought to characterize the pharmacokinetic behavior of 1843U89 and to seek preliminary evidence of anticancer activity. Patients with advanced solid malignancies were treated with escalating doses of 1843U89 as a 2-min i.v. infusion daily for 5 days every 3 weeks. Initially, patients were treated in the absence of high-dose folic acid until dose-limiting toxicity was consistently noted. Next, patients were treated with escalating doses of 1843U89 preceded by 1000 mg of folic acid administered p.o. 30 min before each of the 5 daily doses of 1843U89. Patients (32) received 101 total courses of 1843U89 at doses ranging from 1 to 6 mg/m/day with and without folic acid. At the 2 mg/m/day dose level without folic acid, 2 of 7 new patients experienced dose-limiting toxicity, principally neutropenia, mucositis, and malaise in 3 of 11 courses. 1843U89 doses were further increased with folic acid to 6 mg/m/day, but repetitive treatment was not feasible at this dose level because of an unacceptable high incidence of severe neutropenia and mucositis. Other toxicities included thrombocytopenia, rash, and fever. In contrast, repetitive treatment at the 5 mg/m/day dose level was feasible. The pharmacokinetics of 1843U89 were neither dose dependent nor affected by folic acid. On day 1, clearance, terminal half-life, and steady-state volume of distribution values averaged 47.1 6 21.7 ml/min/m, 7.72 6 4.09 h, and 16.76 8.8 liter/m/h, respectively. The results of the study indicate that the administration of 1843U89 as a 2-min infusion daily for 5 days every 3 weeks without and with folic acid is feasible at 1843U89 doses as high as 2 and 5 mg/m /day, respectively. Because folic acid pretreatment results in no diminution of the antitumor activity of 1843U89 in preclinical studies and ameliorates the toxic effects of 1843U89 in both preclinical models and cancer patients, the therapeutic index of 1843U89 may be enhanced by folic acid pretreatment and, therefore, the development of 1843U89 with folic acid is warranted. However, the question of whether to administer 1843U89 at a dose of 2 mg/m/day with folic acid, which is associated with negligible toxicity, or at its highest feasible dose with folic acid, 5 mg/m/day, should be addressed in appropriately designed trials. INTRODUCTION TS has been validated in the clinic as a strategic target for anticancer drug development (1). TS catalyzes the conversion of dUMP to thymidylate, using the reduced folate MTHF as a cofactor (1). Effective inhibition of TS depletes cells of thymidylate, which is an essential component of DNA, and ultimately results in cell death (1, 2). Malignant cells generally have higher TS levels than normal cells, which implies that cancer cells may depend onde novopyrimidine biosynthesis to a greater degree than normal cells and, therefore, may be more susceptible to TS inhibition (3, 4). The antitumor activities of FU further support the validity of TS as a therapeutic target. After metabolism to fluorodeoxyuridine, FU forms a ternary covalent, albeit reversReceived 3/5/01; revised 4/20/01; accepted 4/20/01. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisementin accordance with 18 U.S.C. Section 1734 solely to indicate this fact. 1 Supported by a grant from GlaxoSmithKline, Inc., Research Triangle Park, NC, and NIH Grant M01 RR01346 to the Frederic C. Bartter Clinical Research Unit of The Audie Murphy Veterans Administration Hospital, San Antonio, TX. 2 To whom requests for reprints should be addressed, at Institute for Drug Development, Cancer Therapy and Research Center, 8122 Datapoint Drive, Suite 700, San Antonio, TX 78229. Phone: (210) 616-5945; Fax: (210) 616-5865; E-mail: erowinsk@saci.org. 3 The abbreviations used are: TS, thymidylate synthase; ANC, absolute neutrophil count; AUC, area under the concentration-time curve; AUC0,̀ area under the concentration-time curve extrapolated to infinity; AUC0–24, area under the concentration-time curve until 24 h posttreatment;Ae0–24, total amount of drug excreted in the urine until 24 h posttreatment;Cmax, maximum plasma concentration; CL, clearance; DLT, dose-limiting toxicity;Emax, maximal effect model; FPGS, folylpolyglutamate synthetase; FU, 5-fluorouracil; MTA, multitargeted antifol; MTD, maximum tolerated dose; MTHF, 5,10-methylenetetrahydrofolate;t1/2, half-life of elimination; TDL, toxic dose low. 1901 Vol. 7, 1901–1911, July 2001 Clinical Cancer Research Research. on October 2, 2017. © 2001 American Association for Cancer clincancerres.aacrjournals.org Downloaded from ible, complex with both MTHF and TS, which, in turn, inhibits synthesis of thymidylate (5, 6). Measures that inhibit dissociation of the ternary complex in vitro, such as increasing intracellular concentrations of reduced folate cofactors and coadministration of reduced folate or folinic acid, generally enhance the antitumor effects of FU (7). Furthermore, factors that affect TS activity, such as TS kinetics, rate of TS renewal, TS gene amplification, stabilization of the ternary complex, intracellular folate concentrations, and folate polyglutamation, are also determinants of cellular sensitivity to FU (1, 8–10). The requirement of TS activity on folate cofactors has served as the rationale for therapeutic evaluations of synthetic analogues of folate cofactors. The 3-methyl-substituted benzoquinazolone folate analogue 1843U89 (GlaxoWellcome, Inc, Research Triangle Park, NC; Fig. 1) is a very potent, noncompetitive inhibitor of human TS ( Ki ' 90 pM; Refs. 11–14). 1843U89 was developed through chemical optimization of an initial series of benzo[f]quinazolin-1(2H)-one compounds, which inhibit human TS withKi values of;20 nM (11, 12). Although structurally similar to the natural folate cofactor of the TS reaction, cotreatment with MTHF does not alter the binding of 1843U89 to TS, which indicates that 1843U89 is a noncompetitive inhibitor of TS (11–14). Because 1843U89 does not require polyglutamation to effectively inhibit TS, tumors with acquired resistance to polyglutamable folate analogue attributable to either low FPGS or high folylpolyglutamyl hydrolase may hypothetically retain sensitivity to the agent (14–17). On the other hand, 1843U89 is an efficient FPGS substrate for synthesis of the diglutamate (14–17). Unlike most folate-based TS inhibitors, 1843U89 does not require additional polyglutamation beyond its diglutamate species to be retained intracellularly, and the diglutamate species has been demonstrated to accumulate intracellularly, even in cells with reduced FPGS (17, 18). Furthermore, there is evidence indicating that 1843U89 and other compounds with high affinities for the reduced folate carrier may produce a tumor-selective advantage because some tumors have been demonstrated to predominately use this mechanism for folate transport (14). 1843U89 has demonstrated impressive cytotoxicity against a broad range of human tumor xenografts implanted in nude mice, when the high circulating thymidine levels of mice are reduced (14, 19). The agent has been shown to inhibit the growth or induce regression of s.c. or subrenal capsule implants of MOLT-4 leukemia, SW480 and WiDr colon, HCT-8 ileocecal, and MCF-7 breast carcinomas with IC 50 values ranging from 0.2 to 0.7 nM (14, 17, 19). Regressions of advanced mammary and spinal nerve sheath malignancies have also been noted after treatment of dogs with 1843U89 and folic acid (14). Thymidine reverses the cytotoxicity of 1843U89 against human tumor xenografts, whereas folic acid has either no or a negligible effect on drug activity (10). In one study, the cytotoxicity of 1843U89 was not reduced by high folic acid concentrations (100 mM) in most cancer cell lines evaluated (14). In addition, the antitumor activity of 1843U89 was not altered in vitro after treatment with more clinically relevant folic acid concentrations ('5 mM; Ref. 14). In contrast, treatment with folinic acid increased the IC 50 values for 1843U89, but only by 10to 50-fold, compared with 10 to 10-fold for other folate-based agents (13). In preclinical toxicology studies, 1843U89 administered daily for 5 days principally affected the gastrointestinal mucosa in dogs and monkeys (14). At high doses, vomiting, diarrhea, anorexia, and decreased activity were the most common toxicities, whereas myelosuppression was uncommon. Treatment of dogs with oral folic acid (50 mg/kg) 40 min before lethal (6 mg/kg) and twice-lethal doses of 1843U89 resulted in minimal clinical toxicity and histopathological effects; however, severe gastrointestinal and hematological toxicities were observed after treatment with three times the lethal dose of 1843U89, despite pretreatment with an identical dose of folic acid (14). The minimal folic acid dose given once daily that was necessary to provide complete protection from a lethal dose of 1843U89 was between 10 and 50 mg/kg/day. The impressive preclinical antitumor activity of 1843U89 and its unique noncompetitive mechanism of TS inhibition served as the impetus for its clinical development. The principal objectives of this study were to: (a) determine the principal toxicities of 1843U89 administered as a 2-min i.v. infusion daily for 5 days every 3 weeks without and with folic acid pretreatment; (b) determine the MTD of 1843U89 without and with folic acid and recommend doses for subsequent disease-directed evalua