KRAS G12C/SOS1 binding kit HTRF®

Associated products

The KRAS G12C/SOS1 binding kit is designed to identify KRAS G12C/SOS1 inhibitors.

No-wash
All inclusive kit
Accurate pharmacology

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The KRAS G12C/SOS1 binding kit is designed to identify KRAS G12C/SOS1 inhibitors.

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Overview

A fast and easy way to identify new inhibitors of KRAS G12C/SOS1 protein interactions.

KRAS is a small GTPase implicated in various biological processes, such as cell proliferation, cell survival, and cell metabolism. This proto-oncogene is well known to be mutated in many cancer subtypes, inducing an uncontrolled proliferation and cell metabolism modifications. It thereby contributes to the Warburg effect in cancer cells. Like the majority of small GTPases, KRAS binds to GDP in its inactive form or binds to GTP to switch into active form. KRAS G12C is one of the most commonly present mutant forms in cancer which leads to a permanently active state of KRAS. The Ras guanine nucleotide exchange factor, also called SOS1, is a GEF protein promoting the active form of KRAS. The upregulation of the KRAS/SOS1 interaction leads to cancer phenotypes. Identifying new KRAS/SOS1 inhibitors or GTP competitors are therefore the two major strategies to control biological processes involved in cancer growth, by reducing the KRAS activity as well as the associated pathways.

Benefits

CANCER RESEARCH
KRAS INHIBITORS

Assay principle

The KRAS G12C / SOS1 PPI assay includes tagged human recombinant partners (KRAS G12C and SOS1) and labeled anti-tag reagents for HTRF detection. Without any inhibitor, KRAS G12C loaded with GTP binds to SOS1, and the binding of each detection reagent to its tagged target generates an HTRF signal. In the presence of KRAS G12C / SOS1 inhibitors or GTP competitors, the HTRF signal decreases.

Principle of the HTRF KRAS G12C/SOS1 binding assay

Assay protocol

The Human KRAS G12C / SOS1 binding assay can be run in a 96- or 384-well low volume white plate (20 µL final). As described here, compounds or standards are dispensed directly into the assay plate. The pre-mixed GTP and human Tag1-KRAS G12C protein is then added, together with the human Tag2-SOS1 protein, followed by the dispensing of the HTRF reagents: The anti- Tag2 antibody labeled with Terbium cryptate and the anti-Tag1 antibody labeled with XL665. The reagents labeled with HTRF fluorophores may be pre-mixed and added in a single dispensing step. No washing steps are needed. The protocol can be further miniaturized or upscaled by simply resizing each addition volume proportionally.

KRAS G12C/SOS1 inhibitor screening

Various compounds known to be KRAS or SOS1 inhibitors were added to the assay. BAY-293, BI-3406, BI-2852, MRTX-1257, and the two KRAS G12C selective compounds, ARS-1620 and AMG-510, displayed the right potency and pharmacological ranking in good correlation with the literature. ARS-1620 and AMG-510 results confirm that the KRAS G12C/SOS1 binding kit is specific to G12C mutated form of KRAS.

Validation of the HTRF KRAS G12C/SOS1 binding kit with inhibitors

Competitive or non competitive nucleotide screening

The two nucleotides GDP and ATP were added to the assay. GDP, a GTP competitor well known to bind KRAS G12C, displayed the right potency in good correlation with the literature. Meanwhile ATP, a non GTP competitive nucleotide, had no effect on the assay, thus confirming the specificity of the kit in detecting only GTP competitors or KRAS G12C/SOS1 inhibitors.

Validation with competitive or non-competitive nucleotides

Best practices for pharmacological characterization of PPI inhibitors

Easy pharmacological characterization of PPI modulators. - Technical Notes

HTRF assays handle low- to high affinity protein-protein interactions

Deciphering low- and high affinity interactions - Application Notes

Nuclear receptor ligand identification with HTRF

Monitoring nuclear receptor binding with HTRF assays - Application Notes

HTRF addresses large protein-protein interaction complexes

Challenge large complexes with HTRF assays - Application Notes

HTRF PPI your dream assay served on a plate

Sandwiches aren't just for eating - Infographics

A brief history of Protein-Protein Interactions

How well do you know PPI? - Infographics

Virology research solutions using HTRF Protein-Protein Interaction assays

See how peer researchers challenge the viral life cycle with PPI assays - Application Notes

Product Insert KRAS G12C/SOS1 kit / 64KRASG12PEG-64KRASG12PEH

64KRASG12PEG-64KRASG12PEH - Product Insert

KRAS GTP competitors and KRAS/SOS1 inhibitors screening with new HTRF biochemical-based assays

Screening for KRAS competitors and inhibitors with HTRF technology - Application Notes

Safety Data Sheet (DEU) KRAS G12C/SOS1 kit / 64KRASG12PEG