Purpose Triptolide induces cancer cell apoptosis by inhibiting RNA synthesis and signaling pathways like NF-B. In a rat toxicology comparison using a new intravenous emulsion formulation, the MRx102 MTD was 4.5 mg/kg for males, 3 mg/kg for females; the triptolide MTD was 0.63 mg/kg for males, 0.317 mg/kg for females. The MRx102 NOAEL was 1.5C3.0 mg/kg, and the triptolide NOAEL was 0.05C0.15 mg/kg. Mean plasma concentrations for both MRx102 and triptolide decreased rapidly from a high Cmax following i.v. injection. Plasma triptolide levels stabilized at a consistent level through 2 hours after MRx102 injection. Triptolide T1/2,e values for MRx102-injected SB-262470 rats (~0.85 to ~3.7 hours) were markedly greater than triptolide injected rats (~0.15 to ~0.39 hours), indicating more extended triptolide exposure with MRx102. MRx102 dog toxicology and toxicokinetics results are presented. Conclusions MRx102 was 20? to 60-fold safer than triptolide comparing rat NOAELs. This may be due to the improved toxicokinetic profile of MRx102 compared to triptolide using the emulsion formulation, with no high Cmax and more consistent early exposure to triptolide. Keywords: AML, prodrug, leukemia, triptolide, MRx102, toxicology, toxicokinetics Introduction Triptolide is a natural product extracted from the medicinal plant, Tripterygium wilfordii,  used traditionally to treat inflammatory and autoimmune diseases. Triptolide has been used clinically to treat rheumatoid arthritis , nephritis  and certain SB-262470 cancers. Despite complete remissions for some patients with triptolide in acute leukemia  and the F60008 triptolide prodrug in acute myeloid leukemia (AML) , significant safety challenges have been encountered . F60008 underwent incomplete conversion in human plasma. High interindividual variability of pharmacokinetic parameters between patients reflected widely Rabbit Polyclonal to Myb. divergent circulating triptolide levels, and the most severe adverse events were due to overexposure (evident as a high Cmax) . Thus, the major problem for further development of triptolide and triptolide-related compounds is the toxicity profile that appears to result from poor pharmacokinetics. Our development of triptolide prodrug MRx102 is intended to optimize the activity for clinical application and produce, essentially, an improved and safer triptolide derivative. Our requirements for an optimized compound include complete prodrug triptolide conversion, and avoiding very high initial exposure to triptolide in preclinical animal studies by maintenance of relatively constant triptolide blood levels early after i.v. injection. A new formulation was developed to help achieve the goal of improved pharmacokinetics for the lipophilic MRx102 prodrug and to avoid excipient-related side effects. The activity of MRx102 was compared to SB-262470 triptolide for in vitro cytotoxicity with the MV4C11 AML cell line. We evaluated MRx102 and triptolide activity with AML patient cells, using flow cytometric characterization of CD34+CD38? stem/progenitor cells that are thought to be responsible for AML disease progression, resistance to therapy and relapse. The new formulation was developed for intravenous studies, and was utilized in toxicology and toxicokinetic comparisons. The combination of the MRx102 triptolide prodrug SB-262470 and the emulsion formulation produced an altered and improved triptolide exposure profile, resulting in an apparently much safer triptolide. Materials and Methods Animals Male and female Sprague-Dawley rats from Harlan (Frederick, MD) were 7C8 weeks old at the start of dosing in the toxicology and toxicokinetics studies. Rats were individually housed upon assignment to the studies in compliance with National Research Council Guide for the Care and Use of Laboratory Animals. Male and female beagle dogs from Marshall BioResources USA (North Rose, NY) were 6.6C8.8 kilograms in the 7-day repeat dose toxicology and toxicokinetics studies. Dogs were individually housed in compliance with USDA guidelines. Cells, cell culture and treatment of cells The MV4C11 human AML cell line was obtained from the American Type Culture Collection (Manassas VA, CRL-9591) SB-262470 for studies at Murigenics Inc. (Vallejo, CA) and cultured in Iscoves Modified Dulbeccos Medium (Invitrogen, Inc, Carlsbad CA) containing glutamine, penicillin/streptomycin and 10% heat inactivated fetal calf serum (FCS). Fresh peripheral blood samples from AML patients with high blast counts (95% or higher) were acquired after written informed consent had been obtained according to the Declaration of Helsinki, and the study protocol was.