RI-1

Catalog No.S8077 Batch:S807701

Technical Data

Formula

C₁₄H₁₁Cl₃N₂O₃

Molecular Weight

361.61

CAS No.

415713-60-9

Solubility (25°C)*

In vitro

DMSO

50 mg/mL (138.27 mM)

Water

Insoluble

Ethanol

Insoluble

In vivo (Add solvents to the product individually and in order)

Clear solution	5% DMSO 1 95% Corn oil	1.25mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 25 mg/ml clear DMSO stock solution to 950 μL of corn oil and mix evenly. The mixed solution should be used immediately for optimal results.
Clear solution	5%DMSO 1 40%PEG300 2 5%Tween80 3 50%ddH2O	1.25mg/ml	Taking the 1 mL working solution as an example, add 50 μL of 25 mg/ml clarified DMSO stock solution to 400 μL of PEG300, mix evenly to clarify it; add 50 μL of Tween80 to the above system, mix evenly to clarify; then continue to add 500 μL of ddH2O to adjust the volume to 1 mL. The mixed solution should be used immediately for optimal results.

* <1 mg/ml means slightly soluble or insoluble.
* Please note that Selleck tests the solubility of all compounds in-house, and the actual solubility may differ slightly from published values. This is normal and is due to slight batch-to-batch variations.
* Room temperature shipping (Stability testing shows this product can be shipped without any cooling measures.)

Preparing Stock Solutions

Biological Activity

Description

RI-1 (RAD51 inhibitor 1) is a RAD51 inhibitor with IC50 ranging from 5 to 30 μM.

Targets

RAD51 ^[1]

<30 μM

In vitro

RI-1 sensitizes cells to DNA damage by directly and specifically disrupting HsRAD51 and inhibiting the ability of RAD51 to form filaments on ssDNA. In addition, RI-1 alone generates single-agent toxicity in all three cancer cell lines (HeLa, MCF-7 and U2OS) with LD50 values in the 20–40 µM range. ^[1] RI-1 decreases the rejoining of γ-H2AX foci in G2 phase cells and results in a higher level of unrepaired DSBs 6 hours after irradiation. ^[2]

Features

A selective recombinant RAD51 protein inhibitor discovered in 2012. Valuable tool for mechanistic studies of DNA repair and potential for use in many cancers.

Protocol (from reference)

Kinase Assay:^{^[1]}	DNA binding assays All reactions are performed in black non-binding polystyrene 384-well plates with reaction volumes of 30–100 μL. Purified DNA strand exchange proteins and chemical compounds are pre-incubated at room temperature for 5 minutes; they are then further incubated at 37°C for 30 min with 100 nM of ssDNA substrate, consisting of a 45-mer poly-dT tagged with Alexa 488 at the 5’ terminus (synthesized and purified by Integrated DNA Technologies). Reactions are performed in 20 mM HEPES pH 7.5, 10 mM MgCl₂, 0.25 μM BSA, 2% glycerol, 30 mM NaCl, 4% DMSO and 2 mM ATP. Some conditions included DTT or TCEP (tris(2-carboxyethyl)phosphine) as indicated. DNA binding is measured as a function of fluorescence polarization (FP) with a Safire2 plate reader, using the following settings: excitation 470±5nm, emission 530±5nm, 10 reads/well, Z height and G factor auto-calibrated from control wells. Displayed error bars represent standard deviation. For experiments involving a titration of protein concentrations, data are fit to an equation that accounts for the cooperative nature by which recombinase proteins bind DNA. For experiments involving a titration of RI-1, protein concentrations are selected to give an ∼80% saturation of the FP signal in the absence of RI-1.
Cell Assay:^{^[1]}	Cell lines HeLa, MCF-7 and U2OS cells Concentrations ~35 μM Incubation Time 24 hours Method Cytotoxicity is determined by loss of colony-forming ability. Experiments are performed in triplicate. Crystal violet stained colonies are imaged with a CCD camera and counted using NIH Image software. Error bars denote standard error.

Kinase Assay:^{^[1]}

DNA binding assays
All reactions are performed in black non-binding polystyrene 384-well plates with reaction volumes of 30–100 μL. Purified DNA strand exchange proteins and chemical compounds are pre-incubated at room temperature for 5 minutes; they are then further incubated at 37°C for 30 min with 100 nM of ssDNA substrate, consisting of a 45-mer poly-dT tagged with Alexa 488 at the 5’ terminus (synthesized and purified by Integrated DNA Technologies). Reactions are performed in 20 mM HEPES pH 7.5, 10 mM MgCl₂, 0.25 μM BSA, 2% glycerol, 30 mM NaCl, 4% DMSO and 2 mM ATP. Some conditions included DTT or TCEP (tris(2-carboxyethyl)phosphine) as indicated. DNA binding is measured as a function of fluorescence polarization (FP) with a Safire2 plate reader, using the following settings: excitation 470±5nm, emission 530±5nm, 10 reads/well, Z height and G factor auto-calibrated from control wells. Displayed error bars represent standard deviation. For experiments involving a titration of protein concentrations, data are fit to an equation that accounts for the cooperative nature by which recombinase proteins bind DNA. For experiments involving a titration of RI-1, protein concentrations are selected to give an ∼80% saturation of the FP signal in the absence of RI-1.

Cell Assay:^{^[1]}

Cell lines
HeLa, MCF-7 and U2OS cells
Concentrations
~35 μM
Incubation Time
24 hours
Method
Cytotoxicity is determined by loss of colony-forming ability. Experiments are performed in triplicate. Crystal violet stained colonies are imaged with a CCD camera and counted using NIH Image software. Error bars denote standard error.

References

Customer Product Validation

: Data from [Data independently produced by , , Biomed Pharmacother, 2017, 94:165-168]

Selleck's RI-1 has been cited by 18 publications

Short-Term Starvation Weakens the Efficacy of Cell Cycle Specific Chemotherapy Drugs through G1 Arrest [ Int J Mol Sci, 2023, 24(3)2498]	PubMed: 36768821
Short-Term Starvation Weakens the Efficacy of Cell Cycle Specific Chemotherapy Drugs through G1 Arrest [ Int J Mol Sci, 2023, 24(3)2498]	PubMed: 36768821
DNA‑PKcs phosphorylation specific inhibitor, NU7441, enhances the radiosensitivity of clinically relevant radioresistant oral squamous cell carcinoma cells [ Biomed Rep, 2023, 18(4):28]	PubMed: 36926187
U2AF1 mutation connects DNA damage to the alternative splicing of RAD51 in lung adenocarcinomas [ Clin Exp Pharmacol Physiol, 2022, 10.1111/1440-1681.13646]	PubMed: 35434831
CRIP1 cooperates with BRCA2 to drive the nuclear enrichment of RAD51 and to facilitate homologous repair upon DNA damage induced by chemotherapy [ Oncogene, 2021, 10.1038/s41388-021-01932-0]	PubMed: 34262130
A non-genetic, cell cycle-dependent mechanism of platinum resistance in lung adenocarcinoma [ Elife, 2021, 10e65234]	PubMed: 33983115
The Valproate Mediates Radio-Bidirectional Regulation Through RFWD3-Dependent Ubiquitination on Rad51 [ Front Oncol, 2021, 11:646256]	PubMed: 33842359
Upregulation of FoxO6 in nucleus pulposus cells promotes DNA damage repair via activation of RAD51 [ Eur Rev Med Pharmacol Sci, 2021, 25(17):5392-5401]	PubMed: 34533813
Comprehensive optimization of a reporter assay toolbox for three distinct CRISPR-Cas systems [ FEBS Open Bio, 2021, 10.1002/2211-5463.13198]	PubMed: 33999508
Targeting Mutant PPM1D Sensitizes Diffuse Intrinsic Pontine Glioma Cells to the PARP Inhibitor Olaparib. [ Mol Cancer Res, 2020, 10.1158/1541-7786.MCR-19-0507]	PubMed: 32229503

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SHIPPING AND STORAGE
Selleck products are transported at room temperature. If you receive the product at room temperature, please rest assured, the Selleck Quality Inspection Department has conducted experiments to verify that the normal temperature placement of one month will not affect the biological activity of powder products. After collecting, please store the product according to the requirements described in the datasheet. Most Selleck products are stable under the recommended conditions.

NOT FOR HUMAN, VETERINARY DIAGNOSTIC OR THERAPEUTIC USE.