5 & 6 - HIV Protease Inhibitors Flashcards
What class of viruses doe HIV belong to?
Retroviruses
How does HIV infect host T cells?
HIV infects host T cells that have the CD4 antigen on their surface
Viral DNA enters host cell nucleus
- it is integrated into the genetic material by an Integrase enzyme
Activation of host cell then results in transcription of viral DNA into messenger RNA
- which is then translated into viral proteins
What’re the steps of the HIV life cycle?
After HIV mRNA has been translated into viral proteins what key step happens next?
Viral proteins are cleaved to active forms by HIV PROTEASE
- crucial for virion development
What’s an issue when treating HIV proteases with inhibitors?
Like reverse transcriptase inhibitors, resistance is quickly developed to protease inhibitors
Requires combination therapy for treating HIV infections
What enzymes can be targeted by med chemists for HIV
Reverse transcriptase
Integrase
Transcriptase
HIV protease
How do protease inhibitors differ to reverse transcriptase inhibitors?
Protease inhibitors are not prodrugs and do not require activation
What family is the HIV protease enzyme a part of?
It is an example of an enzyme family called the:
Aspartyl proteases-enzymes
- they contain aspartic acid in the active site
- crucial for catalytic mechanism
What do apartyl protease-enzymes do?
They catalyse the cleavage of peptide bonds
What is the structure of the HIV protease enzyme?
It is a dimer made up of 2 identical protein units, each consisting of 99 amino acids
In the sense of assays and analysis, why is HIV protease a good drug target?
It is relatively small can can be obtained by synthesis
It’s can be cloned and expressed in fast-growing cells
It is easily crystallised with & without inhibitor bound
What amino acids lie on the floor of the active site of HIV protease?
Aspa-25
Thr-26
Gly-27
What makes the structure of HIV protease a good target?
The active site is at the interface between the protein units
This means that drugs can interact on all sides = very high affinities
How many binding sites are there in HIV protease enzyme?
8
- 4 on each protein unit located on either side of the catalytic region
What do the sub-sites accept for binding?
The sub-sites accept the amino acids residues of the substrate
What’re the key interactions seen in the HIV protease active site?
Asp-25 & Asp-25’ are involved in the catalytic mechanism with a bridging water molecule
What is the cleavage mechanism of HIV Protease?
Tetrahedral intermediate is key
What is an issue (& its solution) when designing TS inhibitors of HIV protease?
The intermediate is inherently unstable
Hence, an inhibitor must contain a TS isostere which has a tetrahedral centre to mimic the TS.
- it also must be stable to hydrolyis
Which functional group shows the most success in mimicking the TS?
What is the best approach to designing transition inhibitors for HIV protease?
Begin with a core unit spanning S1 to S1’ sub-sites, then grow the molecule outwards (either end) to fit into S2/S3 and S2’/S3’ subsites.
How did the design of saquinavir begin?
A pentapeptide sequence was identified and the amide link between Phe—Pro was replaced by the hydroxyethylamine isostere
What is the key binding site in HIV protease, how was this found?
Inclusion of asparagine at P2 gave 40 fold increase in activity
Shows that S2/S2’ is key binding site
Is the S3 site hydrophilic or hydrophobic?
Hydrophobic
Why was the bicyclic ring system added?
Because it occupies the S1’ site more fully
The R-configuration is CRITICAL
Why was the ester substituted with an amide?
Due to metabolic stability - amides are more stable than esters to metabolism
What is the important about the placement of the carbonyl groups either side of the hydroxyethylamine?
The carbonyl groups can act as H-bond acceptors to the bridging water molecule found in the transition state in HIV Protease active site
Where does the OH of the isostere replacement bind to in the active site?
It forms hydrogen bonds to both aspartic acids in the floor of the active site
- Asp-25 & Asp-25’
What’s the point of adding Valine here?
It extends the molecules to S2 and S2’ pockets to be occupied
Gave large increase to inhibitory activity
What’s the point of adding Z-protecting groups to valine’s amine?
Aside from a major increase in potency, it improved the drugs ability to cross membranes
Why were polar groups introduced at the ends of the molecule here?
Because, it was found that the terminal portions fo the molecule were exposed to solvation
This meant that more polar groups could be added at those positions without affecting binding
What’s a problem within the body with adding these polar groups?
The pyridine groups were extensively metabolised to N-oxides that are excreted into bile
Why was the S-OH removed?
Because it only formed 1 H-bond
R-OH forms 2 H-bonds with enzyme
Removing it may enhance solubility?
Why was the pyridine ring replaced with a thiazole ring system?
Done to improve pharmacokinetics
- replacement of pyridine with a more metabolically stable ring system
Describe and explain the changes made to Ritonavir after resistance rose
The isopropyl group, at S3, made important hydrophobic interactions with enzyme
- resistant viral strains reduced this interaction
In Lopinavir, the S3 residue is removed and cyclic urea has been included to maximise interactions at S2 site
Lopinavir is active against ritonavir resistant viruses
Give the mechanism for the synthesis of 5
