Dolutegravir Interactions with HIV-1 Integrase-DNA: Structural Rationale for Drug Resistance and Dissociation Kinetics

<div><p>Signature HIV-1 integrase mutations associated with clinical raltegravir resistance involve 1 of 3 primary genetic pathways, Y143C/R, Q148H/K/R and N155H, the latter 2 of which confer cross-resistance to elvitegravir. In accord with clinical findings, in vitro drug resistance profiling studies with wild-type and site-directed integrase mutant viruses have shown significant fold increases in raltegravir and elvitegravir resistance for the specified viral mutants relative to wild-type HIV-1. Dolutegravir, in contrast, has demonstrated clinical efficacy in subjects failing raltegravir therapy due to integrase mutations at Y143, Q148 or N155, which is consistent with its distinct in vitro resistance profile as dolutegravir’s antiviral activity against these viral mutants is equivalent to its activity against wild-type HIV-1. Kinetic studies of inhibitor dissociation from wild-type and mutant integrase-viral DNA complexes have shown that dolutegravir also has a distinct off-rate profile with dissociative half-lives substantially longer than those of raltegravir and elvitegravir, suggesting that dolutegravir’s prolonged binding may be an important contributing factor to its distinct resistance profile. To provide a structural rationale for these observations, we constructed several molecular models of wild-type and clinically relevant mutant HIV-1 integrase enzymes in complex with viral DNA and dolutegravir, raltegravir or elvitegravir. Here, we discuss our structural models and the posited effects that the integrase mutations and the structural and electronic properties of the integrase inhibitors may have on the catalytic pocket and inhibitor binding and, consequently, on antiviral potency in vitro and in the clinic.</p> </div

Structural models of (A) Q148H/G140S and (B) N155H HIV-1 integrase with U5 LTR DNA and dolutegravir.

<p>(A) The Q148H/G140S mutations are predicted to disrupt the structure of the flexible active-site loop, displacing the 3₁₀ helix away from the DDE motif and weakening the H-bond interaction between the backbone CO of Q148H and the backbone NH of E152. (B) The N155H mutation is predicted to disrupt the structure of the α4 helix, widen the base of the catalytic pocket, alter the placement of at least the Mg²⁺ ion coordinated to residues D64 and E152 and alter the conformation of the terminal 3′ adenosine forming part of the pocket. Molecular representations and coloring schemes are described in Figure 2.</p

Structural models of HIV-1 integrase with U5 LTR DNA and (A, B) raltegravir, (C) elvitegravir or (D) dolutegravir.

<p>For raltegravir, the terminal 3′ adenylate is depicted in 2 distinct conformations: panel 2A shows the published conformer and panel 2B shows an alternative conformer that is also consistent with the observed electron density. Raltegravir, elvitegravir and dolutegravir are in stick representation with carbon, nitrogen, oxygen, fluorine and chlorine atoms colored gray, blue, red, cyan and green, respectively. A select subset of amino acids and nucleotides is depicted and labeled with residue type and number (numbering schemes as listed in Figure S1); all residues are in stick representation with carbon atoms colored by secondary structure/chain and nitrogen and oxygen atoms colored blue and red, respectively. The Mg²⁺ ions are represented as small yellow spheres with coordinate bonds to the inhibitors depicted as dashed yellow lines.</p

Structural model of Q148R HIV-1 integrase with U5 LTR DNA.

<p>The side chain of residue Q148R was modeled interacting with the side chain of E152 and in this conformation the residue may interfere with the binding of elvitegravir. Molecular representations and coloring schemes as described in Figure 2.</p

2D structures of (A) dolutegravir, (B) raltegravir and (C) elvitegravir.

<p>Red ovals encircle the oxygen atoms that chelate the divalent metal cations in the active site; green ovals encircle the halobenzyl groups; and blue boxes encircle the approximate regions of the scaffolds that can accommodate positive charge after chelation of the metals. The purple circles at (B) encircle raltegravir’s gem-dimethyl (small circle) and oxadiazole groups, and the purple oval at (C) encircles elvitegravir’s 1-hydroxymethyl-2-methylpropyl group.</p

Synthesis and Evaluation of 7-Substituted 4-Aminoquinoline Analogues for Antimalarial Activity

We previously reported that substituted 4-aminoquinolines with a phenyl ether substituent at the 7-position of the quinoline ring and the capability of intramolecular hydrogen bonding between the protonated amine on the side chain and a hydrogen bond acceptor on the amine’s alkyl substituents exhibited potent antimalarial activity against the multidrug resistant strain <i>P</i>. <i>falciparum</i> W2. We employed a parallel synthetic method to generate diaryl ether, biaryl, and alkylaryl 4-aminoquinoline analogues in the background of a limited number of side chain variations that had previously afforded potent 4-aminoquinolines. All subsets were evaluated for their antimalarial activity against the chloroquine-sensitive strain 3D7 and the chloroquine-resistant K1 strain as well as for cytotoxicity against mammalian cell lines. While all three arrays showed good antimalarial activity, only the biaryl-containing subset showed consistently good potency against the drug-resistant K1 strain and good selectivity with regard to mammalian cytotoxicity. Overall, our data indicate that the biaryl-containing series contains promising candidates for further study

Carbamoyl Pyridone HIV‑1 Integrase Inhibitors 3. A Diastereomeric Approach to Chiral Nonracemic Tricyclic Ring Systems and the Discovery of Dolutegravir (S/GSK1349572) and (S/GSK1265744)

We report herein the discovery of the human immunodeficiency virus type-1 (HIV-1) integrase inhibitors dolutegravir (S/GSK1349572) (<b>3</b>) and S/GSK1265744 (<b>4</b>). These drugs stem from a series of carbamoyl pyridone analogues designed using a two-metal chelation model of the integrase catalytic active site. Structure–activity studies evolved a tricyclic series of carbamoyl pyridines that demonstrated properties indicative of once-daily dosing and superior potency against resistant viral strains. An inherent hemiaminal ring fusion stereocenter within the tricyclic carbamoyl pyridone scaffold led to a critical substrate controlled diastereoselective synthetic strategy whereby chiral information from small readily available amino alcohols was employed to control relative and absolute stereochemistry of the final drug candidates. Modest to extremely high levels of stereochemical control were observed depending on ring size and position of the stereocenter. This approach resulted in the discovery of <b>3</b> and <b>4</b>, which are currently in clinical development