Julie Naud

Discovery of a potent and selective noncovalent linear inhibitor of the hepatitis C virus NS3 protease (BI 201335).

C-Terminal carboxylic acid containing inhibitors of the NS3 protease are reported. A novel series of linear tripeptide inhibitors that are very potent and selective against the NS3 protease are described. A substantial contribution to the potency of these linear inhibitors arises from the introduction of a C8 substituent on the B-ring of the quinoline moiety found on the P2 of these inhibitors. The introduction of a C8 methyl group results not only in a modest increase in the cell-based potency of these inhibitors but more importantly in a much better pharmacokinetic profile in rats as well. Exploration of C8-substitutions led to the identification of the bromo derivative as the best group at this position, resulting in a significant increase in the cell-based potency of this class of inhibitors. Structure-activity studies on the C8-bromo derivatives ultimately led to the discovery of clinical candidate 29 (BI 201335), a very potent and selective inhibitor of genotype1 NS3 protease with a promising PK profile in rats.

Discovery of the first series of inhibitors of human papillomavirus type 11: inhibition of the assembly of the E1-E2-Origin DNA complex.

We have discovered a series of inhibitors of the assembly of the HPV11 E1-E2-origin DNA complex, which incorporate an indandione fused to a substituted tetrahydrofuran.

Synthesis and antiviral activity of monobactams inhibiting the human cytomegalovirus protease.

A series of monobactam inhibitors of HCMV (N(o)) protease bearing a heterocycle linked by a methylene group at C-4 is described. Inhibitors containing a heterocycle such as a 2-furyl, 2-thiophenyl, 4-methyl-2-tetrazole and 2-benzothiazole were found to be active in a plaque reduction assay. Furthermore, 2-benzothiazole derivatives were shown to inhibit the HCMV protease activity inside cells by using a cell transfection assay, indicating that their antiviral activity in the plaque reduction assay could be attributed to protease inhibition.

Potent triazolyl-proline-based inhibitors of HCV NS3 protease

The design and synthesis of tripeptide-based inhibitors of the HCV NS3 protease containing a novel P2-triazole is described. Replacement of the P2 quinoline with a triazole moiety provided a versatile handle which could be expediently modified to generate a diverse series of inhibitors. Further refinement by the incorporation of an aryl-substituted triazole and replacement of the P1 acid with an acyl sulfonamide ultimately provided inhibitors with interesting cellular activity.

Potent β-Lactam Inhibitors of Human Cytomegalovirus Protease:

A series of novel monobactam inhibitors of human cytomegalovirus (HCMV) protease has been described that possess a heterocyclic thiomethyl side chain at C-4. Changes to the heterocycle did not significantly change the inhibitory activity of these compounds in an enzymatic assay, although improvements in solubility and cell culture activity were noted. A number of permutations between C-4 substitutions and N-1 derivatives led to the identification of several β-lactams with antiviral activity in a plaque reduction assay. N-methyl thiotetrazole-containing compounds were found to be the most potent inhibitors in the enzymatic assay.

Structure-based design of novel HCV NS5B thumb pocket 2 allosteric inhibitors with submicromolar gt1 replicon potency: Discovery of a quinazolinone chemotype

We describe the structure-based design of a novel lead chemotype that binds to thumb pocket 2 of HCV NS5B polymerase and inhibits cell-based gt1 subgenomic reporter replicons at sub-micromolar concentrations (EC50<200nM). This new class of potent thumb pocket 2 inhibitors features a 1H-quinazolin-4-one scaffold derived from hybridization of a previously reported, low affinity thiazolone chemotype with our recently described anthranilic acid series. Guided by X-ray structural information, a key NS5B-ligand interaction involving the carboxylate group of anthranilic acid based inhibitors was replaced by a neutral two-point hydrogen bonding interaction between the quinazolinone scaffold and the protein backbone. The in vitro ADME and in vivo rat PK profile of representative analogs are also presented and provide areas for future optimization of this new class of HCV polymerase inhibitors.

The use of chemical double-mutant cycles in biomolecular recognition studies: application to HCV NS3 protease inhibitors.

Chemical double-mutant cycles (DMCs) have been elegantly used to quantify the energies of noncovalent interactions between specific pairs of functional groups in the molecular recognition of abiotic systems. The method has also been employed to estimate the free energy of a novel noncovalent interaction in an intramolecular context. The analysis assumes that there are no significant structural differences between the four components of the cycle and that the free energies of the individual interactions are additive. It can then be used to determine the energetic contribution of a specific interaction in the context of a more complex, global binding event in such a manner that secondary interactions are effectively cancelled out. Here, we describe the novel use of a particular type of chemical DMC, which serves as a useful formalism to characterize the mutual effect of different substructures within a molecule on a biomolecular recognition process, namely, binding to an enzyme active site. This treatment varies from those described above in that two “deletion mutations” occupy different positions on the same chemical species while the binding partner is kept constant. Contrary to the DMC whose validity rests on the conditions of unchanging structure and the additive contributions of subsites to the total binding energy, the present analysis is useful in providing a clear indication of whether two portions of a ligand contribute to the total binding energy in an additive or cooperative (synergistic or antagonistic) manner, thus affording insight into the binding phenomenon and/or aiding in the understanding of structure–activity relationships (SAR) when structural data is lacking. The treatment is exemplified by thermodynamic cycles constructed from peptidic inhibitors of the hepatitis C virus NS3 protease (HCV protease) where the binding event is competitive inhibition of the enzyme. Examples of all three cases (additive, synergistic and antagonistic) are described. HCV protease represents an important target in the quest for a specific anti ACHTUNGTRENNUNGviral agent against hepatitis C infection, a serious global health problem for which the therapeutic need has not been met. Of central importance in our early efforts was the discovery that this novel serine protease suffers product inhibition by oligopeptide N-terminal cleavage products. Optimization of product-based hexapepACHTUNGTRENNUNGtides entailed N-truncation, elaboration of a novel P1 residue and the incorporation of an aryloxy substituent onto the P2 residue, which led to potent tripeptides bearing a free Cterminal carboxylate. This series ultimately provided the first HCV protease inhibitor demonstrated to reduce viral load in ACHTUNGTRENNUNGinfected patients. A molecule central to this optimization effort was inhibitor 4, which has become a key standard and biological tool in the study of this class of ligands and the enzyme target. As is generally observed for peptidyl inhibitors of serine proteases, compound 4 is bound to HCV protease in an extended conformation (Figure 1), canonical H-bonds involving the P1-NH, P3-

Synthesis and optimization of a novel series of HCV NS3 protease inhibitors: 4-Arylproline analogs

In this report we describe the synthesis and evaluation of diverse 4-arylproline analogs as HCV NS3 protease inhibitors. Introduction of this novel P2 moiety opened up new SAR and, in combination with a synthetic approach providing a versatile handle, allowed for efficient exploitation of this novel series of NS3 protease inhibitors. Multiple structural modifications of the aryl group at the 4-proline, guided by structural analysis, led to the identification of analogs which were very potent in both enzymatic and cell based assays. The impact of this systematic SAR on different drug properties is reported.