P53R3 is a potent p53 reactivator that restores sequence-specific DNA binding of p53 hotspot mutants such as p53R175H, p53R248W, and p53R273H. P53R3 induces p53-dependent antiproliferation effects with much higher specificity than PRIMA-1. P53R3 enhances the recruitment of wild-type p53 and p53M237I to the promoters of several target genes. P53R3 strongly enhances the mRNA, total protein and cell surface expression of the death receptor death receptor 5 (DR5). P53R3 could be used in cancer research.

Physicochemical Properties

Molecular Formula	C32H35CL2N5O2
Molecular Weight	592.55860543251
Exact Mass	591.216
CAS #	922150-12-7
PubChem CID	58809185
Appearance	White to off-white solid powder
LogP	7.3
Hydrogen Bond Donor Count	1
Hydrogen Bond Acceptor Count	7
Rotatable Bond Count	10
Heavy Atom Count	41
Complexity	790
Defined Atom Stereocenter Count	1
SMILES	C(OC)(=O)[C@H](C(C)C)NC1=C2C(=NC(CN3CCN(C(C4=CC=C(Cl)C=C4)C4=CC=C(Cl)C=C4)CC3)=N1)C=CC=C2
InChi Key	DFBRKGUTHGRQMH-LJAQVGFWSA-N
InChi Code	InChI=1S/C32H35Cl2N5O2/c1-21(2)29(32(40)41-3)37-31-26-6-4-5-7-27(26)35-28(36-31)20-38-16-18-39(19-17-38)30(22-8-12-24(33)13-9-22)23-10-14-25(34)15-11-23/h4-15,21,29-30H,16-20H2,1-3H3,(H,35,36,37)/t29-/m0/s1
Chemical Name	methyl (2S)-2-[[2-[[4-[bis(4-chlorophenyl)methyl]piperazin-1-yl]methyl]quinazolin-4-yl]amino]-3-methylbutanoate
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 Note: Please store this product in a sealed and protected environment (e.g. under nitrogen), avoid exposure to moisture.
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

ln Vitro	In WiDr colon cancer cells harbouring p53R273H and KLE cells with p53R175H, P53R3 (10 μg/ml; 24 hours; in the absence or presence of the unlabelled p53 consensus oligonucleotide) restores p53-specific DNA binding activity to p53R273H (a DNA contact mutant) and p53R175H (a structural mutant)[1]. In a p53-dependent manner, P53R3 (1–33 μg/ml; 24 hours) prevents the LN-308 sublines carrying mutant p53 plasmids from proliferating. While p53R273H-dependent effects are less and necessitate high concentrations of P53R3, p53R175H-dependent effects are significant across a wide concentration range[1]. Proliferation inhibition caused by P53R3 is more pronounced than that of P53R273H when it comes to p53R248W reactivation. Not even at concentrations near its solubility limit (33 μg/ml) does P53R3 show cytotoxic effects[1]. In LN-308 p53R175H and LN-308 p53R273H cells, P53R3 (33 μg/ml; 18 hours) causes a significant reduction in S phase cells and a G0/G1 cell cycle arrest. However, it had no effect on the LN-308 p53R248W cells' cell cycle distribution[1].
Cell Assay	Cell Viability Assay[1] Cell Types: p53 null LN -308 human glioma cells with a control plasmid or plasmids encoding the mutants p53R175H, p53R248W and p53R273H Tested Concentrations: 1-33 μg/mL Incubation Duration: 24 hrs (hours) Experimental Results: Induced p53-dependent and -independent antiproliferative and cytotoxic effects in vitro.
References	[1]. Targeting Oncogenic Mutant p53 for Cancer Therapy. Front Oncol.

Solubility Data

Solubility (In Vitro)

DMSO : ≥ 100 mg/mL (168.76 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	1.6876 mL	8.4380 mL	16.8759 mL
	5 mM	0.3375 mL	1.6876 mL	3.3752 mL
	10 mM	0.1688 mL	0.8438 mL	1.6876 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.