Rational Drug Design : HIV Integrase

A process for drug design which bases the design of the drug upon the structure of its protein target.

1. Structural mapping of the receptor (protein, P) active site

2. Identification of ligands (L) of complementary shape and appropriate functionality

3. Docking of the ligand to the receptor site - predicting a range of PL complexes with different GPL values

4. Scoring i.e. ranking GPL and correlating with experimentally determined properties such as IC50 values

In the first step of the integration process, two nucleotides are removed fromeach 3’-end of the viral DNA. This reaction exposes the terminal 3’-hydroxylgroup that is to be joined to target DNA (Fig. 1B). In the second step, DNAstrand transfer, a pair of processed viral DNA ends is inserted into thetarget DNA (Fig. 1C). Integrase is responsible for 3’-processing and DNAstrand transfer, but the latter repair steps are likely to be carried out bycellular enzymes.

The catalytic domainhas an RNaseH-type fold and belongs to the superfamily of polynucleotidyl transferases. The active site is comprised of two Asp residues and one Glu, in the typicalD,D(35)E motif, each of which is required for catalysis.

de novo Ligand Design

DCQ acids; DCT acids

DKAs

Quinolone derived

PDP SQL

four criteria to conclude that integrase is theinhibitor target:1. found to be active against recombinant integrase.2. infected cells treated with the drug must show an accumulation of 2-LTR circles, resulting from the accumulation of viral cDNA and decreased HIV integration into host3. integrase mutations must be found in drug-resistant viruses4, the drug should be inactive in biochemical assays against recombinantintegrases bearing the mutations identified in the drug-resistant viruses

Issues in Protein Setup

Crystal structure available for Integrase but :I. Limitations of crystal structure:

only catalytic domain DNA binding not revealed cystal structure vs. physiologically

active structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Protein Setup

Crystal structure available for Integrase Catalytic Domain but :

I. Crystal reveals trimeric structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Protein Setup

Crystal structure available for Integrase Catalytic Domain but :

I. Crystal reveals trimeric structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Protein Setup

Crystal structure available for Integrase Catalytic Domain but :

I. Crystal reveals trimeric structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Protein Setup

Crystal structure available for Integrase Catalytic Domain but :

I. Crystal reveals trimeric structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Protein Setup

Crystal structure available for Integrase Catalytic Domain but :

I. Crystal reveals trimeric structureII. Position of hydrogens undetermined III. Residues missing or ill-definedIV. Protonation of His undeterminedV. Solvation

Issues in Ligand Design

Crystal structure available for CITEP bound to catalytic core but :

I. Position of hydrogens undeterminedII. Tautomeric structures possibleIII. Influence of pHIV. Need to limit conformational flexibility based

on experimental and theoretical crteria

Issues in Ligand Design

Crystal structure available for CITEP bound to catalytic core but :

I. Position of hydrogens undeterminedII. Tautomeric structures possibleIII. Influence of pHIV. Need to limit conformational flexibility based

on experimental and theoretical crteria

Issues in Ligand Design

Crystal structure available for CITEP bound to catalytic core but :

I. Position of hydrogens undeterminedII. Tautomeric structures possibleIII. Influence of pHIV. Need to limit conformational flexibility based

on experimental and theoretical crteria

Issues in Ligand Design

Crystal structure available for CITEP bound to catalytic core but :

I. Position of hydrogens undeterminedII. Tautomeric structures possibleIII. Influence of pHIV. Need to limit conformational flexibility based

on experimental and theoretical crteria

Tetrazole pKa=5

Issues in Ligand Design

Crystal structure available for CITEP bound to catalytic core but :

I. Position of hydrogens undeterminedII. Tautomeric structures possibleIII. Influence of pHIV. Need to limit conformational flexibility based

on experimental and theoretical crteria

Fixed and planar

Based on HF/6-31G* calculationsLimited to +/- 45 degrees

Issues in Docking

The prediction of the ligand conformation and orientation within a targeted binding site involves:I. Positioning ligand and

evaluating quality of binding

II. Manually refining ligand position

III. Energy minimization (electrostatic, steric, strain and h-bond)

Issues in Docking

The prediction of the ligand conformation and orientation within a targeted binding site involves:I. Positioning ligand and

evaluating quality of binding

II. Manually refining ligand position

III. Energy minimization (electrostatic, steric, strain and h-bond)

Issues in Docking

The prediction of the ligand conformation and orientation within a targeted binding site involves:I. Positioning ligand and

evaluating quality of binding

II. Manually refining ligand position

III. Energy minimization (electrostatic, steric, strain and h-bond)

Issues in Scoring

The prediction of the optimum ligand conformation and orientation within a targeted binding site involves:I. Posing : Determining the fit of

the ligandII. Conformational SearchingIII. Scoring and Ranking

Results

Results

Criterion for Ligand Selection:

I. Theoretical and experimental structuresII. Fill active siteIII. Conformational structures

Ligand Design

Criterion for Ligand Selection:

I. Theoretical and experimental structuresII. Fill active siteIII. Conformational structures

Ligand Design

Criterion for Ligand Selection:

I. Theoretical and experimental structuresII. Fill active siteIII. Conformational structures

Ligand Design

The prediction of the affects of mutations within thebinding site on the effects of the ligands involves:

I. Identifying possible sights of mutationsII. Determining effect of mutations

Site Mutations and Drug Resistance

The prediction of the affects of mutations within thebinding site on the effects of the ligands involves:

I. Identifying possible sights of mutationsII. Determining effect of mutations

Site Mutations and Drug Resistance

Site Mutations and Drug Resistance

Problem with Protein Flexibility

http://folding.stanford.edu/villin/S300x300.105.56.95.mpg