16th International HIV Drug Resistance Workshop 12-16 June 2007, Barbados

HIGHLY POTENT PROTEASE INHIBITORS WITH NOVEL ESCAPE PATHWAYS

Antivir Ther. 2007; 12:S21 (abstract no. 19)

E Afonina¹, SV Gulnik¹, M Eissenstat¹, H Yokoe¹, B Yu¹, NT Parkin² and JW Erickson^1
1Sequoia Pharmaceuticals, Inc., Gaithersburg, MD, USA; ²Monogram Biosciences, Inc., San Francisco, CA, USA

BACKGROUND: We have previously described the discovery of SPI-256, an HIV protease inhibitor (PI) with high potency and broad spectrum activity against multidrug-resistant (MDR) HIV strains. We report here the evaluation of two next generation PIs, SPI-390 and SPI-457 with high potency against worst case scenario MDR HIV, slower rate of in vitro resistance development and novel escape pathways compared to SPI-256.

METHODS: The enzyme inhibition was characterized using recombinant WT and MDR HIV PRs and fluorogenic substrate. Antiviral activity was evaluated in cellbased assay in MT4 cells. The PhenoSense HIV assay (Monogram Biosciences) was used to evaluate the antiviral activity of inhibitors against a panel of 10 highly PI-resistant MDR HIV strains. In vitro selection experiments were performed by passaging WT-HXB2 HIV in MT4 cells.

RESULTS: Both inhibitors were highly active against purified WT and MDR-HIV proteases with K_i in the picomolar range. In MT4 cells, both compounds demonstrated potent antiviral activity against subtype B HIV and retained low nanomolar potency (<50 nM) against a selected panel of highly PI resistant MDR strains derived from clinical isolates. In the PhenoSense assay, SPI-390 and SPI-457 exhibited IC₅₀ values of 0.6 nM and of 0.3 nM, respectively against wt HIV, and were more potent than FDA-approved PIs including darunavir (IC₅₀ 0.8–74 nM). For the selected panel of 10 isolates with >50-fold resistance to reference PIs (IC₅₀ range: 4.5 to >2,600 nM) and/or six primary PI mutations the average SPI-390 and SPI-457 IC₅₀ was 8.4 nM (range: 1.5–21 nM), and 6.8 nM (range: 1.2–13 nM), respectively. Resistance selection experiments were initiated at drug concentrations equal to IC₅₀ values, with subsequent incremental increase of less than 30%. After more than 130 days and 12 passages, genotypic analysis of virus growing at 80 nM of SPI-390 (nine times IC₅₀) and 37 nM of SPI-457 (five times IC₅₀) revealed the presence of R41I, L63P, K70E, and A71V mutations respectively.

CONCLUSIONS: SPI-390 and SPI-457 represent a group of highly potent protease inhibitors and maintain nanomolar potency against worst-case scenario MDR HIV isolates. Selection of virus resistant to both compounds was a very slow process and required multiple passages with small incremental increases of concentration with the novel escape pathways.

2007-06-12
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