Hu7691 is an orally bioactive, selective Akt inhibitor (antagonist) with IC50s of 4.0 nM, 97.5 nM and 28 nM for Akt1, Akt2 and Akt3 respectively. Hu7691 inhibits tumor growth and reduces skin toxicity in mice.

Physicochemical Properties

Molecular Formula	C22H22CLF3N4O
Molecular Weight	450.88
Exact Mass	450.143
CAS #	2360523-76-6
Related CAS #	Hu7691 free base;2241232-43-7
PubChem CID	155557232
Appearance	Typically exists as solid at room temperature
Hydrogen Bond Donor Count	3
Hydrogen Bond Acceptor Count	6
Rotatable Bond Count	4
Heavy Atom Count	31
Complexity	597
Defined Atom Stereocenter Count	2
SMILES	CN1C(=CC=N1)C2=CC(=C(C=C2)C(=O)N[C@@H]3CNCC[C@H]3C4=CC(=C(C=C4)F)F)F.Cl
InChi Key	WHVFOLXIIVBJTJ-ICCPGPOKSA-N
InChi Code	InChI=1S/C22H21F3N4O.ClH/c1-29-21(7-9-27-29)14-2-4-16(18(24)11-14)22(30)28-20-12-26-8-6-15(20)13-3-5-17(23)19(25)10-13;/h2-5,7,9-11,15,20,26H,6,8,12H2,1H3,(H,28,30);1H/t15-,20+;/m0./s1
Chemical Name	N-[(3S,4S)-4-(3,4-difluorophenyl)piperidin-3-yl]-2-fluoro-4-(2-methylpyrazol-3-yl)benzamide;hydrochloride
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	Akt1 4.0 nM (IC50) Akt2 97.5 nM (IC50) Akt3 28 nM (IC50) PKA 11 nM (IC50) PKCη 629 nM (IC50) ROCK1 354 nM (IC50) RSK1 756 nM (IC50) p70S6K 229 nM (IC50)
ln Vitro	Hu7691 exhibits low inhibitory effects against the majority of kinases in the four families (PLA, IC50=11 nM; PKCη, IC50=629 nM; ROCK1, IC50=354 nM; RSK1, IC50=756 nM; P70S6K, IC50=229 nM; SGK, IC50=1009 nM);[1]. At IC50 values of 15.2 μM, B5 (10, 20, 30, 40 μM; over 72 h) demonstrates minimal toxicity towards HaCaT cells. Hu7691 exhibits a considerable, IC50 range of 0.6-27 μM, growth inhibitory effect on 18 various types of human tumor cells (U87-MG, U251, A549, HepG2, HT-29, KHOS, MDA-MB-231, PC3, SKOV3, and so on) originating from diverse tissues. With IC50 values of 5.4 and 16.1 μM, respectively, Hu7691 exhibits weak antiproliferation actions against the HPDE6-C7 and HL7702 normal cells[1].
ln Vivo	For 22 days, Hu7691 (12.5–50 mg/kg/day; ir) exhibits dose-dependent suppression of tumor growth[1]. In rats, Hu7691 (15 mg/kg; oral) exhibits an AUC of 2820.64 ng/mL·h, a Cmax of 171.17 ng/mL, and a T1/2 of 8.68 hours[1]. In rats, Hu7691 (2 mg/kg; intravenous) has an AUC of 532.87 ng/mL·h, a Cmax of 207.52 ng/mL, and a T1/2 of 6.24 hours[1]. In a beagle dog (male, 40 weeks old, 8–10 kg), Hu7691 (20 mg/kg; oral) had an AUC of 36303 ng/mL·h, a T1/2 of 16.7 hours, and a Cmax of 905.65 ng/mL[1].
Cell Assay	Western Blot Analysis[1] Cell Types: HaCaT cells Tested Concentrations: 2.25, 4.5, 9, 18, 36 μM Incubation Duration: 24 hrs (hours) Experimental Results: Induced effective decrease of the phosphorylation level of Akt (S473).
Animal Protocol	Animal/Disease Models: balb/c (Bagg ALBino) mouse: (nu/nu, female, 3-4 weeks old , 20-25 g) with 786-O and KHOS xenograft[1] Doses: 12.5, 25, 50 mg/kg Route of Administration: Oral; one time/day for 22 days Experimental Results: demonstrated dose-dependent tumor growth inhibition. Animal/Disease Models: SD rats (male, 8 weeks old, 250-300 g)[1] Doses: 15 mg/kg (pharmacokinetic/PK Analysis) Route of Administration: Oral Experimental Results: Had a T1/2 of 8.68 hrs (hours), a Cmax of 171.17 ng/mL and an AUC of 2820.64 ng/mL·h.
References	[1]. Discovery of N-((3 S,4 S)-4-(3,4-Difluorophenyl)piperidin-3-yl)-2-fluoro-4-(1-methyl-1 H-pyrazol-5-yl)benzamide (Hu7691), a Potent and Selective Akt Inhibitor That Enables Decrease of Cutaneous Toxicity. J Med Chem. 2021 Aug 26;64(16):1.

Solubility Data

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	2.2179 mL	11.0894 mL	22.1789 mL
	5 mM	0.4436 mL	2.2179 mL	4.4358 mL
	10 mM	0.2218 mL	1.1089 mL	2.2179 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.