Novel stapled peptides serving as KRASG12C/SOS1 inhibitor and application of novel stapled peptide

A technology of A0B0C0D0E0F0G0A1B1C1D1, stapled peptides, applied in the direction of animal/human peptides, peptides, peptide sources, etc., can solve problems such as inability, reduced solubility, and poor stability

Active Publication Date: 2020-07-10
CHINA PHARM UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, after the polypeptide fragment leaves the overall structure of the protein, it will not be able to stably form the secondary structure required for binding, but will easily form a random coil conformation, resulting in a decrease in binding activity, and it is more susceptible to degradation by peptidases, so it cannot be directly used as a drug.
Elizaveta S. Leshchinera et al. designed and synthesized SAH-SOS1A with the sequence Ac-RRFFGIXLTNXLKTEEGN-NH 2 , X stands for (S)-α-methyl-α-p

Method used

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  • Novel stapled peptides serving as KRASG12C/SOS1 inhibitor and application of novel stapled peptide
  • Novel stapled peptides serving as KRASG12C/SOS1 inhibitor and application of novel stapled peptide
  • Novel stapled peptides serving as KRASG12C/SOS1 inhibitor and application of novel stapled peptide

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0110] Example 1: Preparation and Characterization of Stapled Peptides:

[0111] 1.1. Synthesis of stapled peptide

[0112] Synthesis of stapled peptides was artificially carried out by SPPS method; the synthesis scale was 0.1 mmol, 40-RAM amphiphilic Rink amide resin was loaded at 0.4 mmol / g, and an excess of 5 equivalents was used for protected amino acids; all protected amino acids and even The joint agent HATU was pre-dissolved in DMF to prepare a stock solution with a concentration of 0.5M; under vortex stirring, the resin was first swelled in 6mlDMF for 15 minutes, and after the DMF was removed by filtration, the solution was added by adding 20% ​​piperidine / DMF solution ( 6ml) and vortexed for 1 minute to deprotect the Fmoc group; the above steps were carried out twice; finally, the resin was washed 3 times with DMF, and then the required amino acid (1ml, 0.5mmol), N, N-diisopropyl Diethylamine (DIEA) (0.164ml, 1mmol) and HATU coupling reagent (1ml, 0.5mmol) and stirre...

Embodiment 2

[0120] Example 2: Evaluation of whether stapled peptides maintain α-helical structure:

[0121] In order to further understand the effect of shortening the length of the polypeptide on the α-helical structure, and using trifluoroethanol to promote the formation of the α-helical secondary structure, a circular dichroism test was performed on the stapled peptide; on the CD spectrum The typical secondary structure of polypeptides in the far ultraviolet region (185–245nm) has its own characteristic absorption peaks; among them, the α-helix has double negative peaks at 208nm and 222nm, and a positive peak at 195nm; A weaker negative peak appears; random curl has a negative peak at 200nm, and a small and broad positive peak at 220nm.

[0122] Determination of the secondary structure of stapled peptides by circular dichroism spectroscopy, CD spectra were measured using a J-810 spectrometer at 25 °C using a quartz cell with an optical path length of 1.0 mm and recorded by scanning in th...

Embodiment 3

[0131] Example 3: Evaluation of stapled peptides and KRAS G12C Protein binding affinity:

[0132] For detection of SAH-SOS1A and Peptide-DB-1 with KRAS G12C For direct protein binding, real-time binding assays were performed using label-free biomolecules combined with the ForteBio Octet Red 96 system and biolayer interferometry (BLI) technology; biomolecules bind to the sensor surface to form biofilms; biofilms interfere with the light transmitted through the sensor; Interference phenomena are detected by phase shifts and thus changes in the number of molecules bound to the sensor surface; binding affinity is typically assessed using the Kd (equilibrium dissociation constant), which is used to assess the strength of bimolecular interactions.

[0133] Assessment of stapled peptides and KRAS by biofilm interferometry (BLI) G12C Binding affinity of protein; KRAS assessed using biolayer interferometric biosensor in OctetRed 96 instrument G12C Binding affinity between proteins a...

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Abstract

The invention discloses novel stapled peptides serving as a KRASG12C/SOS1 inhibitor and application of the novel stapled peptides. Also provided are a composition comprising these stapled peptides anda method for using such peptides in the treatment of cancers. The stapled peptides are more stable in alpha-helical conformation to some extent; the stapled peptides are higher in affinity with KRASG12C protein; the enzymolysis capability of resisting trypsin and alpha-chymotrypsin is better; besides, the plasma stability of the stapled peptides is greatly enhanced, and the stapled peptides are derived from an alpha-helical binding region of SOS1 protein and KRAS protein and inhibit the activation of the KRAS protein. The invention also relates to a preparation method of the novel stapled peptides, a pharmaceutical composition containing these stapled peptides, and application of the stapled peptides independently used or in combination with other compounds for the prevention or treatmentof cancers (such as non-small cell lung cancer).

Description

technical field [0001] The present invention relates to a staple peptide, which is derived from the α-helix binding region of SOS1 protein and KRAS protein. Background technique [0002] Lung cancer is the main cause of cancer death in the world (Cheng and Planken 2018), which seriously endangers human life and health. In the past few decades, the treatment of lung cancer has received great attention (Wang, Huang et al. 2019). Non-small cell lung cancer (NSCLC) accounts for 80% of all lung cancer cases (Jemal, Bray et al.2011), and the Ras proto-oncogene is the most frequently mutated gene in NSCLC (Prior, Lewis et al.2012, Li, Liu et al. 2018), mutations were detected in approximately 25% of all tumors (de Castro Carpeno and Belda-Iniesta 2013), and the v-Ki-ras2Kirsten rat sarcoma virus oncogene (KRAS) accounted for 90% of RAS mutations in lung adenocarcinomas (John C. Hunter and Sudershan Gondi2015, Rothschild2015). Most oncogenic forms of RAS impair their intrinsic GTP...

Claims

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Application Information

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IPC IPC(8): C07K14/47A61K38/17A61P35/00A61P11/00
CPCC07K14/4705A61P35/00A61P11/00A61K38/00
Inventor 徐莉莉狄斌李翠翠安橹燕
Owner CHINA PHARM UNIV
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