OC144093 a novel P glycoprotein inhibitor for the enhancement of antiepileptic therapy
Michael J. Newman, Ross Dixon and Barry Toyonaga
Ontogen Corporation, 6451 El Camino Real, Carlsbad, CA 92009, USA
Abstract. Inhibitors of P glycoprotein (Pgp) may be useful for the enhancement of blood-brain barrier penetration of anti-epileptic drugs (AEDs). Due to polypharmacy and the need for chronic treatment, Pgp inhibitors used in epilepsy should be highly specific and non-toxic. In particular, it may be essential to use compounds that produce minimal inhibition of enzymes involved in metabolism of AEDs and other drugs used by epilepsy patients. 0C144-093 is a novel substituted diarylimidazole generated using the OntoBLOCK® system, a solid-phase combinatorial chemistry technology, in combination with high-throughput cell-based screening. The compound is an extremely potent inhibitor of Pgp-mediated multidrug resistance (MDR) in cancer with an average EC50 of 32 nM, but does not inhibit multidrug resistance-associated protein (MRP1). OC144-093 is the least non-specifically toxic Pgp inhibitor described to date, with an average cytostatic IC50 of 460 mM in 15 cell types. It is not metabolized by cytochrome P450s CYP3A4, 2C8 or 2C9 enzymes involved in AED metabolism. 0C144-093 does not produce a pharmacokinetic (PK) interaction with paclitaxel and has exhibited an excellent PK and safety profile in phase I clinical trials. Our results suggest that 0C144-093 may represent an ideal candidate for use in enhancement of AED blood-brain barrier penetration.
2002 Mechanisms of drug resistance in epilepsy: lessons from oncology. Wiley, Chichester (Novartis Foundation Symposium 243) p 213-230
Ontogen is a chemical drug discovery and development company that is integrating modern tools of biology, chemistry, engineering, and information technologies to discover and optimize novel, small molecules for a variety of therapeutic indications. The company's technology is based on the application of state-of-the art engineering approaches to the evolving needs of biology and chemistry. As a consequence of these efforts Ontogen has developed and patented the OntoBLOCK1 System, a comprehensive array of software and hardware modules to plan, conduct and track the high-speed synthesis of small molecule libraries. Such parallel syntheses on solid support have provided large, proprietary, spatially diverse combinatorial libraries, in milligram quantities per compound, which have been used in a variety of drug discovery and lead optimization programmes.
Recent engineering efforts have been focused on instrumentation for high-throughput chromatographic purification. This technology, called OntoCHROM®, is now used in our laboratories for the practical and efficient high-throughput purification of moderately large libraries (10 000-40 000 compounds). OntoCHROM® not only produces high quality, purified libraries with minimal amounts of solvent, but allows, for the first time, the realization of one of the original promises of combinatorial libraries, that is, high quality structure-activity relationships from the high-throughput biological screening of the libraries. The OntoCHROM® system is a roboticized instrument employing multichannel super critical fluid chromatography, in which target compounds are identified in real time by mass spectrometry and directly collected in a 96-well microtitre plate with well-to-well mapping from the original plate. The development and use of novel high throughput combinatorial chemistry technologies has been validated at Ontogen as a result of the discovery and early development of OC144-093, an inhibitor of P glycoprotein (Pgp).
Currently, over 75% of cancer chemotherapy patients exhibit intrinsic or develop acquired resistance to treatment. Multidrug resistance (MDR) is now recognized as the most common cause of failure of cancer chemotherapy. The MDR phenotype results from cross-resistance to a variety of structurally and functionally unrelated natural products used for cytotoxic anti-tumour therapy. These include anthracyclines, vinca alkaloids, epipodophyllotoxins and taxanes. Ling and co-workers identified a 170 kDa membrane glycoprotein (Pgp), which was subsequently found to mediate ATP-dependent efflux of each of these cancer therapeutics from multidrug-resistant tumour cells (Sikic 1997, van Zuylen et al 2000, for reviews). Additional mechanisms contributing to MDR have been described, including expression of the multidrug resistance-associated protein (MRP) class of transporters. MRP-related proteins appear to be able to transport certain anthracyclines, vinca alkaloids and epipodophyllotoxins, but not taxanes. Members of this family are also involved in normal biliary transport (Borst et al 2000, for review).
Significant progress has been made on determination of the role of Pgp and related proteins in normal physiology. One Pgp gene (MDR1) in human and two genes (Mdrla, Mdrlb) in rodents have been shown to play a significant role in drug resistance. Additional members of the Pgp gene family are involved in phospholipid and bile salt transport. While simultaneous genetic knockout of Mdrla and Mdrlb resulted in healthy mice, indicating that Pgp is not essential for basic physiological functions, the mice exhibited significant alterations in the pharmacological handling of drugs. Blood-brain barrier (BBB) function was decreased and intestinal absorption of drugs was increased (Schinkel 1998, for review). The involvement of Pgp in BBB function suggests that it may play a role in resistance to drugs targeting the CNS. For example, a significant percentage of epilepsy patients fail to respond to conventional anti-epileptic drug (AED) therapy. Several studies have demonstrated elevated levels of Pgp in brain tissue obtained from patients with drug-resistant epilepsy (Tishler et al 1995, Lazarowski et al 1999, Sisodiya et al 1999). In addition, at least one widely prescribed AED, phenytoin, has been shown to be a Pgp substrate (Schinkel et al 1996).
Studies carried out over the last several years have demonstrated that intrinsic and acquired expression of Pgp play a major role in clinical MDR. Tumour types that frequently express Pgp in the absence of exposure to chemotherapy include colorectal, renal cell, hepatocellular, and adrenocortical cancers, as well as chronic leukaemia. Several additional tumour types express Pgp at diagnosis in approximately 10—30% of cases. Examples include breast carcinoma, acute myelogenous leukaemia and ovarian carcinoma (Ramachandran & Melnick 1999, van Zuylen et al 2000, Newman et al 2000, for reviews). Pgp expression at diagnosis in these tumour types can play a significant role in treatment outcome. For example, patients with breast carcinomas expressing Pgp are three times more likely to fail to respond to chemotherapy than patients whose tumours are Pgp negative (Trock et al 1997).
Chemotherapy can induce Pgp expression and/or select for expansion of Pgp-expressing MDR tumour cells. In addition, Pgp inhibitors have been found to decrease the mutation rate for resistance to doxorubicin, and suppress activation of Mdr1 and the appearance of MDR mutants in tissue culture models. In light of these findings, it appears that the most effective way to use chemotherapeutic agents that are Pgp substrates will be in conjunction with a Pgp inhibitor at the time of tumour diagnosis (Sikic 1997, van Zuylen et al 2000, Bates 1999, for reviews).
The first attempts to reverse Pgp-mediated MDR in cell lines, tumour-bearing animals, and in the clinic, took advantage of the observation by Tsuruo and coworkers that Ca2+ channel blockers such as verapamil are inhibitors of MDR. Similar observations were subsequently made with cyclosporin A (Sikic 1997, van Zuylen et al 2000, for reviews). While having some efficacy, these agents are relatively weak Pgp inhibitors (EC5qs = 2—10 mM), are often substrates for Pgp, and exhibit dose-limiting side effects that severely restrict their clinical utility. To address the problems described above there has been considerable interest in second-generation Pgp inhibitors. PSC 833 (Atadja et al 1998), VX-710 (Germann et al 1997) and XR9051 (Dale et al 1998) are 3—100-fold more potent than the first generation compounds and typically do not elicit significant toxicity at doses required for Pgp inhibition. Common dose-limiting toxicities for these types of compounds are ataxia and hyperbilirubinaemia, which are reversible upon cessation of drug treatment.
An additional problem with many Pgp inhibitors is that they are metabolized by enzymes, such as cytochromes P450 CYP3A4 and CYP2C8, involved in the metabolism of a wide variety of therapeutics, including anticancer drugs (Sonnichsen et al 1995, Desai et al 1998) and AEDs (Kerr et al 1994, Komatsu et al 2000). This can result in significant pharmacokinetic (PK) interactions, necessitating close monitoring and reductions in the dose of the co-administered therapeutic agent. For example, PSC 833 and VX-710 have been reported to produce significant PK interactions with agents such as paclitaxel (Sikic 1997, van Zuylen et al 2000, for reviews). Although dose reduction in cancer therapy is feasible, interpatient variability in PK interaction makes this approach problematic, particularly in the 'up-front' therapy setting, designed to kill cancer cells already expressing Pgp and prevent the appearance of MDR before multiple resistance mechanisms evolve. In epilepsy, where chronic treatment and polypharmacy must be considered, it may not be possible to adequately anticipate potential drug interactions when using a Pgp inhibitor that is metabolized by the most abundant and broadly acting P450, CYP3A4. These considerations have led to a search for third generation Pgp inhibitors, which combine high potency with minimal potential for PK interactions.
The Pgp inhibitor properties likely to be required for safe and effective MDR cancer therapy and enhancement of AED CNS penetration are low nanomolar potency, Pgp specificity, lack of non-specific toxicity, relatively long duration of action with reversibility, good oral bioavailability, and lack of PK interaction via metabolism by major P450s. 0C144-093 appears to be one of the first inhibitors to meet all of these requirements.
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