KRA-533 is a selective KRAS agonist. In cancer cells, constitutively active GTP-bound KRAS accumulates and initiates both apoptotic and autophagic cell death pathways. KRA-533 binds to the GTP/GDP binding pocket in the KRAS protein to prevent GTP cleavage.

Physicochemical Properties

Molecular Formula	C13H16BRNO3
Molecular Weight	314.17504
Exact Mass	313.031
Elemental Analysis	C, 49.70; H, 5.13; Br, 25.43; N, 4.46; O, 15.28
CAS #	10161-87-2
Related CAS #	10161-87-2
PubChem CID	270269
Appearance	White to off-white solid powder
Density	1.428g/cm3
Boiling Point	530.4ºC at 760 mmHg
Flash Point	274.6ºC
Vapour Pressure	4.43E-12mmHg at 25°C
Index of Refraction	1.574
LogP	2.609
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	3
Rotatable Bond Count	7
Heavy Atom Count	18
Complexity	275
Defined Atom Stereocenter Count	0
SMILES	BrCC(NCCCCC1C=CC(C(O)=O)=CC=1)=O
InChi Key	WZQJLTLURXNPQG-UHFFFAOYSA-N
InChi Code	InChI=1S/C13H16BrNO3/c14-9-12(16)15-8-2-1-3-10-4-6-11(7-5-10)13(17)18/h4-7H,1-3,8-9H2,(H,15,16)(H,17,18)
Chemical Name	4-[4-[(2-bromoacetyl)amino]butyl]benzoic acid
Synonyms	KRA-533; KRA533; KRA 533
HS Tariff Code	2934.99.9001
Storage	Powder-20°C 3 years 4°C 2 years In solvent -80°C 6 months -20°C 1 month
Shipping Condition	Room temperature (This product is stable at ambient temperature for a few days during ordinary shipping and time spent in Customs)

Biological Activity

Targets	KRAS
ln Vitro	In secondary programming, KRA-533 (10 μM; 48 hours; HCC827 cells) increases KRAS activity [1]. In a dose-dependent way, KRA-533 (0~15 μM; 48 hours; H157 cells) increases KRAS activity, which is linked to elevated pERK levels, variations in active caspase 3/pro-caspase 3, and PARP-related vulnerabilities, ultimately resulting in cell disinfection [1]. More than cells lacking KRAS mutations, KRA-533 (10 μM; 10 days; H292 cells) causes cell growth inhibition. Direct binding of KRA-533 (5~15 μM) to WT, G12C, G12D, and G13D mutant KRAS proteins is possible. KRA-533 dose-dependently raises WT KRAS activity. The activity of active KRAS mutants is further increased by -533 [1].
ln Vivo	KRA-533 (0~30 mg/kg; i.p.; 28 days) reduces tumor development in lung cancer mutant KRAS xenografts in a dose-dependent manner and increases histiocyte labeling and autophagy in a dose-dependent manner KRA-533 Doses between 7.5 mg/kg and 30 mg/kg indicate excellent therapeutic index [1].
Cell Assay	Western blot analysis [1] Cell Types: HCC827 cells Tested Concentrations: 10μM Incubation Duration: 48 hrs (hours) Experimental Results: KRAS activity was enhanced to a greater extent. Apoptosis analysis [1] Cell Types: H157 Cell Tested Concentrations: 0~15 μM Incubation Duration: 48 hrs (hours) Experimental Results: KRAS activity was enhanced in a dose-dependent manner, which was related to pERK levels, active caspase 3/procaspase 3 ratio and PARP cleavage, Cause cell apoptosis.
Animal Protocol	Animal/Disease Models: Nu/Nu nude mice (mutant KRAS xenograft) [1] Doses: 0~ [1]. 30 mg/kg Route of Administration: intraperitoneal (ip) injection; 28 days Experimental Results: Inhibited tumor growth in lung cancer mutant KRAS xenografts in a dose-dependent manner and induced apoptosis and autophagy in tumor tissue in a dose-dependent manner.
References	[1]. Small Molecule KRAS Agonist for Mutant KRAS Cancer Therapy [published correction appears in Mol Cancer. 2020 May 20;19(1):93]. Mol Cancer. 2019;18(1):85. Published 2019 Apr 10.

Solubility Data

Solubility (In Vitro)

DMSO: ~250 mg/mL (~795.7 mM）

Solubility (In Vivo)

Note: Listed below are some common formulations that may be used to formulate products with low water solubility (e.g. < 1 mg/mL), you may test these formulations using a minute amount of products to avoid loss of samples. Injection Formulations (e.g. IP/IV/IM/SC) Injection Formulation 1: DMSO : Tween 80： Saline = 10 : 5 : 85 (i.e. 100 μL DMSO stock solution → 50 μL Tween 80 → 850 μL Saline) *Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH ₂ O to obtain a clear solution. Injection Formulation 2: DMSO : PEG300 ：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL DMSO → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Injection Formulation 3: DMSO : Corn oil = 10 : 90 (i.e. 100 μL DMSO → 900 μL Corn oil) Example: Take the Injection Formulation 3 (DMSO : Corn oil = 10 : 90) as an example, if 1 mL of 2.5 mg/mL working solution is to be prepared, you can take 100 μL 25 mg/mL DMSO stock solution and add to 900 μL corn oil, mix well to obtain a clear or suspension solution (2.5 mg/mL, ready for use in animals). Injection Formulation 4: DMSO : 20% SBE-β-CD in saline = 10 : 90 [i.e. 100 μL DMSO → 900 μL (20% SBE-β-CD in saline)] *Preparation of 20% SBE-β-CD in Saline (4°C，1 week): Dissolve 2 g SBE-β-CD in 10 mL saline to obtain a clear solution. Injection Formulation 5: 2-Hydroxypropyl-β-cyclodextrin : Saline = 50 : 50 (i.e. 500 μL 2-Hydroxypropyl-β-cyclodextrin → 500 μL Saline) Injection Formulation 6: DMSO : PEG300 : castor oil : Saline = 5 : 10 : 20 : 65 (i.e. 50 μL DMSO → 100 μLPEG300 → 200 μL castor oil → 650 μL Saline) Injection Formulation 7: Ethanol : Cremophor : Saline = 10： 10 : 80 (i.e. 100 μL Ethanol → 100 μL Cremophor → 800 μL Saline) Injection Formulation 8: Dissolve in Cremophor/Ethanol (50 : 50), then diluted by Saline Injection Formulation 9: EtOH : Corn oil = 10 : 90 (i.e. 100 μL EtOH → 900 μL Corn oil) Injection Formulation 10: EtOH : PEG300：Tween 80 : Saline = 10 : 40 : 5 : 45 (i.e. 100 μL EtOH → 400 μLPEG300 → 50 μL Tween 80 → 450 μL Saline) Oral Formulations Oral Formulation 1: Suspend in 0.5% CMC Na (carboxymethylcellulose sodium) Oral Formulation 2: Suspend in 0.5% Carboxymethyl cellulose Example: Take the Oral Formulation 1 (Suspend in 0.5% CMC Na) as an example, if 100 mL of 2.5 mg/mL working solution is to be prepared, you can first prepare 0.5% CMC Na solution by measuring 0.5 g CMC Na and dissolve it in 100 mL ddH2O to obtain a clear solution; then add 250 mg of the product to 100 mL 0.5% CMC Na solution, to make the suspension solution (2.5 mg/mL, ready for use in animals). Oral Formulation 3: Dissolved in PEG400 Oral Formulation 4: Suspend in 0.2% Carboxymethyl cellulose Oral Formulation 5: Dissolve in 0.25% Tween 80 and 0.5% Carboxymethyl cellulose Oral Formulation 6: Mixing with food powders Note: Please be aware that the above formulations are for reference only. InvivoChem strongly recommends customers to read literature methods/protocols carefully before determining which formulation you should use for in vivo studies, as different compounds have different solubility properties and have to be formulated differently.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	3.1829 mL	15.9144 mL	31.8289 mL
	5 mM	0.6366 mL	3.1829 mL	6.3658 mL
	10 mM	0.3183 mL	1.5914 mL	3.1829 mL

*Note: Please select an appropriate solvent for the preparation of stock solution based on your experiment needs. For most products, DMSO can be used for preparing stock solutions (e.g. 5 mM, 10 mM, or 20 mM concentration); some products with high aqueous solubility may be dissolved in water directly. Solubility information is available at the above Solubility Data section. Once the stock solution is prepared, aliquot it to routine usage volumes and store at -20°C or -80°C. Avoid repeated freeze and thaw cycles.