Microbiology

Mechanism of Action

Darunavir is an inhibitor of the HIV-1 protease. It selectively inhibits the cleavage of HIV encoded Gag-Pol polyproteins in infected cells, thereby preventing the formation of mature virus particles.

Antiviral Activity

Darunavir exhibits activity against laboratory strains and clinical isolates of HIV-1 and laboratory strains of HIV-2 in acutely infected T-cell lines, human peripheral blood mononuclear cells and human monocytes/macrophages with median EC₅₀ values ranging from 1.2 to 8.5 nM (0.7 to 5.0 ng/mL). Darunavir demonstrates antiviral activity in cell culture against a broad panel of HIV-1 group M (A, B, C, D, E, F, G), and group O primary isolates with EC50 values ranging from < 0.1 to 4.3 nM. The EC₅₀ value of darunavir increases by a median factor of 5.4 in the presence of human serum. Darunavir did not show antagonism when studied in combination with the protease inhibitors amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, saquinavir, or tipranavir, the N(t)RTIs abacavir, didanosine, emtricitabine, lamivudine, stavudine, tenofovir, zalcitabine, or zidovudine, the NNRTIs delavirdine, efavirenz, or nevirapine, and the fusion inhibitor enfuvirtide.

Resistance

Cell Culture: HIV-1 isolates with a decreased susceptibility to darunavir have been selected in cell culture and obtained from subjects treated with darunavir/ritonavir. Darunavir-resistant virus derived in cell culture from wild-type HIV had 6- to 21-fold decreased susceptibility to darunavir and harbored 3 to 6 of the following amino acid substitutions S37N/D, R41E/S/T, K55Q, K70E, A71T, T74S, V77I, or I85V in the protease. Selection in cell culture of darunavir resistant HIV-1 from nine HIV-1 strains harboring multiple protease inhibitor resistance-associated mutations resulted in the overall emergence of 22 mutations in the protease gene, including L10F, V11I, I13V, I15V, G16E, L23I, V32I, L33F, S37N, M46I, I47V, I50V, F53L, L63P, A71V, G73S, L76V, V82I, I84V, T91A/S, and Q92R, of which L10F, V32I, L33F, S37N, M46I, I47V, I50V, L63P, A71V, and I84V were the most prevalent. These darunavir-resistant viruses had at least eight protease mutations and exhibited 50- to 641-fold decreases in darunavir susceptibility with final EC₅₀ values ranging from 125 nM to 3461 nM.

Clinical studies of darunavir/ritonavir in treatment-experienced subjects

In the Phase 2b Studies TMC114-C213 and TMC114-C202 and the TMC114-C215/C208 analysis, multiple protease inhibitor-resistant HIV-1 isolates from highly treatment-experienced subjects who received PREZISTA/rtv 600/100 mg b.i.d. and experienced virologic failure, either by rebound, or by never being suppressed, developed amino acid substitutions that were associated with a decrease in susceptibility to darunavir. The amino acid substitution V32I developed on PREZISTA/rtv 600/100 mg b.i.d. in greater than 30% of virologic failure isolates and substitutions at amino acid position I54 developed in greater than 20% of virologic failure isolates. Other substitutions that developed in 10% to 20% of PREZISTA/rtv virologic failure isolates occurred at amino acid positions I15, L33, I47, G73 and L89. The median darunavir phenotype (fold change from reference) of the virologic failure isolates was 21-fold at baseline and 94-fold at failure. Amino acid substitutions were also observed in the protease cleavage sites of some darunavir virologic failure isolates. The resistance profile in treatment-naïve subjects has not been characterized.

Cross-resistance

Cross-resistance among protease inhibitors has been observed. Darunavir has a < 10-fold decreased susceptibility in cell culture against 90% of 3309 clinical isolates resistant to amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, saquinavir and/or tipranavir showing that viruses resistant to these protease inhibitors remain susceptible to darunavir. In Studies TMC114-C213 and TMC114-C202 and the TMC114-C215/C208 analysis, 60% (88/147) of subjects on darunavir/rtv whose baseline isolates had decreased susceptibility to tipranavir (tipranavir fold change > 3) demonstrated a decrease of ≥ 1 log₁₀ in viral load at week 24, and 36% (53/147) achieved < 50 copies/mL plasma HIV RNA levels.

Darunavir-resistant viruses were not susceptible to amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir or saquinavir in cell culture. However, six of nine darunavir-resistant viruses selected in cell culture from protease inhibitor-resistant viruses showed a fold change in EC₅₀ values < 3 for tipranavir, indicative of limited cross-resistance between darunavir and tipranavir. Of the viruses isolated from subjects experiencing virologic failure on darunavir/ritonavir 600/100 mg b.i.d., greater than 50% were still susceptible to tipranavir while less than 5% were susceptible to other protease inhibitors (amprenavir, atazanavir, indinavir, lopinavir, nelfinavir, ritonavir, or saquinavir).

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