17th International HIV Drug Resistance Workshop 10-14 June 2008, Sitges, Spain

IN VITRO SELECTION AND CHARACTERIZATION OF VIRUSES RESISTANT TO RO-5028, A NOVEL NON-NUCLEOSIDE REVERSE TRANSCRIPTASE INHIBITOR

Antivir Ther. 2008; 13(Suppl. 3):A23 (abstract no. 21)

H Javanbakht, Y Li, Z Sweeney, E Chow, J Hang, J Dunn, D Saito, N Cammack, K Klumpp and G Heilek
Roche Palo Alto LLC, Palo Alto, CA, USA

BACKGROUND: RO-5028 represents a new class of potent non-nucleoside reverse transcriptase inhibitors (NNRTIs). RO-5028 is broadly active against NNRTI-resistant HIV variants. Three independent sequential passaging experiments were performed to identify mutations selected by RO-5028, efavirenz and etravirine in vitro.

METHODS: The passaging experiments were performed with HIV HXB2 at a low multiplicity of infection in MT4 cells and increasing concentration of RO-5028, efavirenz and etravirine. Breakthrough viruses were characterized by pool genotypic and phenotypic analysis.

RESULTS: RO-5028 inhibited HIV-1 HXB2 replication in MT4 with an IC₅₀ value of 1.3 nM. RO-5028 is broadly active against HIV-1 strains carrying prevalent point mutations associated with resistance to current NNRTIs. In three independent sequential passaging experiments, the concentration of RO-5028 was increased from 1 nM to 10 µM within 13 passaging steps. The pool sequencing of RO-5028 breakthrough viruses in passage 13 consisted of V106A plus F227L for experiment 1, V106V/I, V108V/I plus Y188L for experiment 2 and V106A, and V108I plus F227F/L for experiment 3. Our results confirmed the published resistance pathways described previously for efavirenz and etravirine. In isogenic mutant constructs carrying the mutations identified during the passaging, we determined that development of resistance to RO-5028 required in general multiple mutant positions within the NNRTI binding pocket to loose susceptibility to RO-5028. One exception was Y188L, a mutation requiring a two nucleotide change, which by itself can confer >100-fold shift in the IC₅₀. RO-5028 passage 13 resistant viruses exhibited cross-resistance to efavirenz. In contrast, the RO-5028 resistant viruses remained sensitive to inhibition by etravirine, suggesting a complementary mutant profile between these compounds.

CONCLUSIONS: Passaging of HIV-1 HXB2 in MT4 cells under increasing drug pressure of RO-5028 in three independent experiments has identified two diverse resistant pathways independent of those seen with efavirenz and etravirine. Notably RO-5028 passage 13 resistance viruses contain mutations that are less prevalent when compared with high prevalent mutations such as K103N, Y181C and G190A. R0-5028 is a potent candidate for the potential treatment of NNRTIs naïve and pretreated patients.

2008-06-10
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