Preclinical Pharmacology and in Vitro Characterization of PR-047, An Oral Inhibitor of the 20S Proteasome

The clinical utility of proteasome inhibitors as treatment for multiple myeloma and non-Hodgkin’s lymphoma (NHL) has been validated with two parenterally administered agents: bortezomib, a dipeptide boronate that was approved by the FDA in 2003; and carfilzomib, a tetrapeptide epoxyketone that has shown promising activity in Phase 1 clinical trials. The clinical development of an orally bioavailable proteasome inhibitor would offer improvement in both dosing flexibility and patience convenience over intravenous (IV) administration. Furthermore, oral administration will allow a practical approach to investigating the affect of prolonged proteasome inhibition by repeated daily dosing in patients with advanced malignancies. Here we describe the preclinical pharmacology of PR-047, a tripeptide epoxyketone that was selected through a medicinal chemistry effort designed to find a selective inhibitor of the chymotrypsin-like (CT-L) activity of the proteasome that combined the efficacy and safety of carfilzomib with the patient convenience of oral dosing. In vitro, PR-047 demonstrated potency against the proteasome CT-L activity comparable to that of carfilzomib and induced tumor cell death across multiple tumor cell types with IC50s <100 nM. In rodents and dogs, oral administration of PR-047 resulted in a prolonged dose-dependent inhibition of the proteasome in all tissues examined with the exception of brain. Proteasome inhibition following PO administration was rapid, resulting in maximal proteasome inhibition within 15 minutes. Plasma pharmacokinetics studies, also assessed in rodents and dogs, confirmed the rapid systemic exposure following oral administration. Absolute bioavailability, as measured by plasma levels of PR-047, was dose dependent and was >20% across all species tested. Doses of PR-047 that resulted in significant (>80%) proteasome inhibition in most tissues were 4–10 fold below the maximum tolerated dose (MTD) and were well tolerated when administered for five consecutive days. These data suggest that repeat daily dosing of PR-047 may be feasible. Single or repeated administrations of PR-047 did not affect kidney or liver function as measured by clinical chemistry. When assessed for anti-tumor activity, PR-047 induced consistent anti-tumor responses in human tumor xenograft models including multiple myeloma (MM1.S), NHL (RL) and solid tumors (HT-29, A549). The anti-tumor activity of PR-047 was equivalent or better than that of carfilzomib and was observed at well tolerated doses. Furthermore, TUNEL staining of tumor sections confirmed that the anti-tumor response was mediated in part by tumor cell apoptosis. PR-047 represents a promising orally bioavailable proteasome inhibitor with a favorable toxicologic profile. The results from these studies have facilitated further development of PR-047 in the treatment of malignant diseases.