Disposition and Metabolism of C-rifapentine in Healthy Volunteers

Rifapentine is a long-acting cyclopentyl-derivative of rifampin. This study was designed to investigate the mass balance and biotransformation of C-rifapentine in humans. Four healthy male volunteers received a single 600-mg oral dose of C-rifapentine in a hydroalcoholic solution. Whole blood, urine, and fecal samples were collected before and at frequent intervals after drug administration. Amount of radioactivity recovered in urine and feces was assessed for up to 18 days postdose. Metabolite characterization in urine and feces was conducted using high-performance liquid chromatography with radiometric detection and liquid chromatography/mass spectroscopy. The total recovery of radioactive dose was 86.8%, with the majority of the radioactive dose recovered in feces (70.2%). Urine was a minor pathway for excretion (16.6% of the dose recovered). More than 90% of the excreted radioactivity was profiled as C chromatographic peaks and 50% was structurally characterized. These characterized compounds found in feces and urine were rifapentine, 25-desacetyl-rifapentine, 3-formyl-rifapentine, and 3-formyl-25-desacetyl-rifapentine. The 25-desacetyl metabolite, formed by esterase enzymes found in blood, liver, and other tissues, was the most abundant compound in feces and urine, contributing 22% to the profiled radioactivity in feces and 54% in urine. The 3-formyl derivatives of rifapentine and 25-desacetyl-rifapentine, formed by nonenzymatic hydrolysis, were also prominent in feces and, to a lesser extent, in urine. In contrast to the feces and urine, rifapentine and 25-desacetyl-rifapentine accounted for essentially all of the plasma radioactivity (99% of the C area under the concentration-time curve), indicating that 25-desacetyl-rifapentine is the primary metabolite in plasma. It appears, therefore, that the nonenzymatic hydrolysis of rifapentine to 3-formyl byproducts occurs primarily in the gut and the acidic environment of the urine. Rifapentine is a rifamycin-derivative antibiotic (Arioli et al., 1981), differing from rifampin by the presence of a cyclopentyl ring instead of a methyl group at the piperazinyl moiety (fig. 1). Rifapentine possesses the same broad spectrum of antimicrobial activity as rifampin and is particularly active against Mycobacterium tuberculosis (Arioli et al., 1981; Dickinson and Mitchison, 1987; Heifets et al., 1990; Yates and Collins, 1982). A distinguishing difference between the two rifamycin antibiotics, however, is rifapentine’s prolonged duration of activity. The elimination half-lives (t1/2) of rifapentine and rifampin differ by about 4-fold based on animal (Assandri et al., 1978, 1984) and human (Birmingham et al., 1978; Buniva et al., 1983; Keung et al., 1996; Biya et al., 1987; Chan et al., 1994; Qing et al., 1994; Zheng et al., 1987; He et al., 1996) pharmacokinetic studies. The only rifapentine metabolite identified in humans to date is 25-desacetyl-rifapentine (Keung et al., 1996; Biya et al., 1987). The other rifamycin derivatives used clinically, rifampin (Acocella, 1978; Loos et al., 1985) and rifabutin (Battaglia et al., 1990; Cocchiara et al., 1989; Blashke and Skinner, 1996), also undergo desacetylation in humans to a 25-desacetyl derivative. Similar to 25-desacetyl metabolites of rifampin and rifabutin, the 25-desacetyl metabolite of rifapentine has in vitro activity against M. tuberculosis (Heifets et al., 1990; Mor et al., 1995). Additional rifapentine-related byproducts identified in recent mouse, rat, and monkey studies included 3-formyl rifapentine and 3-formyl-25-desacetyl-rifapentine (Emary et al., 1998); the 3-formyl derivatives have also been found after rifampin administration in animals and humans (Acocella, 1978; Loos et al., 1985). The purpose of our study was to characterize the disposition of C-rifapentine in man and to isolate and identify metabolites formed after single oral doses. Materials and Methods Chemicals. Rifapentine and C-rifapentine were synthesized by Hoechst Marion Roussel (Kansas City, MO). Acetonitrile and methanol, HPLC grade, were used as received from Burdick and Jackson (Muskegon, MI). Heptafluorobutyric acid (HFB), ascorbic acid, Trizma buffer, sulfatase (type VI), b-glucuronidase (type VII-A), p-nitrocatechol sulfate, and phenolphthalein glucuronic acid were purchased from Sigma Chemical Co. Potassium phosphate (dibasic) was received from Mallinckrodt (Paris, KY). FloScint III, Ultima Gold, and Permafluor E scintillation fluids were obtained from Packard (Meriden, CT). Human serum was obtained from Valley Biomedical, Inc. (Winchester, VA), human plasma from Rockland, Inc. (Gilbertsville, PA), and human serum albumin and a1-acid glycoprotein from Sigma Chemical Co. (St. Louis, MO). Dose Formulation. The labeled drug was prepared with C in the 38position (fig. 1). Radioactive doses were prepared by mixing 25 ml alcohol U.S.P. and 10 ml of potassium phosphate buffer (pH 7.4) in vials containing 600 mg (108 mCi; specific activity 0.18 mCi/mg) C-rifapentine. The radiochemical purity was 98.41%, and the chemical purity was 97.71%. Study Design. Four healthy male volunteers between the ages of 18 and 45 1 Abbreviations used are: HPLC, high performance liquid chromatography; RBC, red blood cell; HSA, human serum albumin; AUC, area under the plasma concentration-time curve; HFB, heptafluorobutyric acid; AAG, a1-acid glycoprotein; MS, mass spectroscopy; LC, liquid chromatography; dpm, disintegrations