Preclinical characterization of the antiviral activity of SCH 900518 (narlaprevir), a novel mechanism-based inhibitor of hepatitis C virus NS3 protease.

Small-molecule hepatitis C virus (HCV) NS3 protease inhibitors such as boceprevir (SCH 503034) have been shown to have antiviral activity when they are used as monotherapy and in combination with pegylated alpha interferon and ribavirin in clinical trials. Improvements in inhibitor potency and pharmacokinetic properties offer opportunities to increase drug exposure and to further increase the sustained virological response. Exploration of the structure-activity relationships of ketoamide inhibitors related to boceprevir has led to the discovery of SCH 900518, a novel ketoamide protease inhibitor which forms a reversible covalent bond with the active-site serine. It has an overall inhibition constant (K*(i)) of 7 nM and a dissociation half-life of 1 to 2 h. SCH 900518 inhibited replicon RNA at a 90% effective concentration (EC(90)) of 40 nM. In biochemical assays, SCH 900518 was active against proteases of genotypes 1 to 3. A 2-week treatment with 5x EC(90) of the inhibitor reduced the replicon RNA level by 3 log units. Selection of replicon cells with SCH 900518 resulted in the outgrowth of several resistant mutants (with the T54A/S and A156S/T/V mutations). Cross-resistance studies demonstrated that the majority of mutations for resistance to boceprevir and telaprevir caused similar fold losses of activity against all three inhibitors; however, SCH 900518 retained more activity against these mutants due to its higher intrinsic potency. Combination treatment with alpha interferon enhanced the inhibition of replicon RNA and suppressed the emergence of resistant replicon colonies, supporting the use of SCH 900518-pegylated alpha interferon combination therapy in the clinic. In summary, the results of the preclinical characterization of the antiviral activity of SCH 900518 support its evaluation in clinical studies.

Combination surgery and nonviral interleukin 2 gene therapy for head and neck cancer.

We have developed a novel nonviral interleukin 2 (IL-2) gene therapy that demonstrates significant treatment-specific, antitumor efficacy in combination with subtotal surgical resection in a head and neck cancer murine model. Treatment of established head and neck tumors in immunocompetent mice was performed via direct injection with a cationic liposome composed of DOTMA and cholesterol formulation carrying DNA plasmid for human IL-2 (hIL-2) gene expression. ELISA assays of tumor extracts 24 h after treatment of hIL-2 gene therapy revealed increased local hIL-2 production as well as a formulation-specific secondary induction of murine IFN-gamma and IL-12. We hypothesize that the paracrine production of multiple cytokines after IL-2 single gene transfer is important for generating a therapeutic effect, and that this strategy will be well tolerated and effective in combination with surgery for head and neck cancer. In animal experiments where surgery was performed in conjunction with an operative site injection of hIL-2 plasmid formulation, no pre-, intra-, or postoperative toxicity or compromise to wound healing was identified. In murine experiments combining partial surgical resection with the nonviral gene therapy, significant antitumor efficacy was demonstrated in the hIL-2 plasmid formulation group compared with empty plasmid formulation and lactose-injected controls. In a separate experiment using smaller tumor sizes, we also demonstrated that treatment outcomes were dependent on the technical aspect of the actual treatment injection as well as visualization with surgical access. The hIL-2 plasmid formulation gene therapy induces local expression of multiple cytokines, results in treatment-specific antitumor effects, and circumvents many of the concerns and toxicity encountered with viral gene transfer. These data support the need for continued preclinical investigation and the consideration of human clinical trials for combination nonviral hIL-2 gene therapy and surgery for head and neck cancer.