Solution-Phase Peptide Selection for P53-MDM2 Inhibition
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Solution Overview
Problem
Current methods for generating peptide scaffolds with non-proteinogenic side chains are time-consuming and limited in their ability to rapidly discover potent peptide-based protein-protein interaction inhibitors, lacking efficiency in structural diversity and pharmacological properties.
Innovation Solution
Development of specific peptide sequences, such as LTFX1HX2WAX3LTSK and LTX1EHYX2AQX3TSK, incorporating non-canonical amino acids and macrocyclic structures, which are synthesized using advanced techniques like split and pool methods and validated through high-pressure size exclusion chromatography and mass spectrometry for enhanced binding affinity.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If conventional display-based combinatorial approaches are used, then structural diversity of peptide libraries can be achieved, but screen complexity and false positives increase
Solution Approach 1:
The patent replaces conventional display-based combinatorial approaches (mechanical/biological systems) with solution-phase affinity selection using high-pressure size exclusion chromatography (physical/chemical system). This substitution eliminates the complexity of display systems while maintaining the ability to generate structurally diverse peptide libraries with non-proteinogenic side chains, thereby reducing false positives and screen complexity while preserving structural diversity.
2Reliability
If systematic studies and in silico guided efforts are used, then metabolic stability and physicochemical properties can be enhanced, but time consumption increases
Solution Approach 1:
The patent incorporates non-canonical amino acids and macrocyclic structures directly into the peptide library design from the outset, rather than requiring subsequent systematic optimization studies. The high-pressure size exclusion chromatography method enables direct selection of peptides with desired metabolic stability and physicochemical properties, eliminating time-consuming iterative optimization while achieving enhanced reliability in drug candidate selection.
3Adaptability or versatility
If non-canonical amino acids are incorporated, then structural diversity and pharmacological properties are improved, but synthesis complexity increases
Solution Approach 1:
The patent changes the fundamental parameter of peptide synthesis by using solution-phase affinity selection with high-pressure size exclusion chromatography instead of conventional solid-phase synthesis followed by display-based screening. This parameter change allows direct incorporation of non-canonical amino acids and macrocyclic structures into the library, achieving enhanced structural diversity while simplifying the overall synthesis and selection process through a single integrated method.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
These peptides demonstrate improved metabolic stability and pharmacological properties, enabling the rapid identification of potent protein-protein interaction inhibitors, particularly effective in targeting interactions like p53-MDM2 and HIV capsid formation.
Implementation Method 1
fractionating the mixture comprising the one or more peptide-protein target complexes using high-pressure size exclusion chromatography (HPSEC) into a plurality of fractions
Implementation Method 2
analyzing the protein fraction directly by liquid chromatography-tandem mass spectrometry (LC-MS/MS) to obtain one or more peptide sequences
Data Source
AI summary
The present invention provides novel peptides (e.g., peptides, macrocyclic peptides, mini-proteins) that modulate protein-protein interactions or salts thereof, and methods of making and using the inventive peptides. In some embodiments, the peptides are high affinity inhibitors (e.g., KD of at most 100 nM, at most 10 nM, at most 1 nM) of a protein-protein interaction. In certain embodiments, these peptides interfere with p53-MDM2 binding interactions (e.g., by binding to MDM2 (GenBank® Gene ID: 4193)). In some embodiments, the peptides interfere with the dimerization of the C-terminal domain of the human immunodeficiency virus (HIV) capsid protein (C-CA), comprising residues 146-231 of the HIV capsid protein (e.g., by binding to the C-terminal domain of the HIV capsid protein (C-CA), thereby inhibiting the dimeric interface of HIV capsid protein, thereby inhibiting viral assembly). These inventive peptides were rapidly generated and identified using novel methods described herein comprising combinatorial peptide synthesis and/or solution affinity selection.