Biological Activity

GDC-0068 (RG7440) is a novel, ATP-competitive and orally bioavailable Akt inhibitor with IC50 values of 5 nM, 18 nM and 8 nM for Akt1, Akt2 and Akt3, respectively. Ipatasertib (GDC-0068) demonstrates potent inhibition of all three Akt isoforms in biochemical assays, but poor inhibition of other AGC family kinases. Moreover, GDC-0068 selectively inhibits cell cycle progression and viability of cancer cell lines driven by PI3K/Akt signaling pathway, including those with defects in the tumor suppressor PTEN, oncogenic mutations in PIK3CA, and amplification of HER2. GDC-0068 (Ipatasertib) blocks the phosphorylation of multiple downstream targets of Akt in human cancer cell lines. GDC-0068 (RG7440) shows good oral exposure resulting in dose-dependent pharmacodynamic effects on downstream biomarkers and a robust anti-tumor response in xenograft models in which the PI3K-Akt-mTOR pathway is activated.

Customer Product Validations & Biological Datas

	Source	Clin Cancer Res (2013). Figure 1.GDC-0068
	Method	Western blot
	Cell Lines	IGROV-1 cell
	Concentrations	200 nmol/L
	Incubation Time	4 h
	Results	GDC-0068 showed more than 600 and more than 100-fold selectivity for Akt1 in IC50 against the closely related kinases PKA and p70S6K, respectively

Protocol (for reference only)

Cell Experiment
Cell lines	MCF10A cells with or without PTEN knockout (KO)
Preparation method	Cell viability assays The 384-well plates were seeded with 2,000 cells per well in a volume of 54 μL per well followed by incubation at 37°C under 5% CO2 overnight (∼16 hours). Compounds were diluted in dimethyl sulfoxide (DMSO) to generate the desired stock concentrations then added in a volume of 6 μL per well. All treatments were tested in quadruplicates. After 4 days incubation, relative numbers of viable cells were estimated using CellTiter-Glo (Promega) and total luminescence was measured on a Wallac Multilabel Reader (PerkinElmer). The concentration of drug resulting in IC50 was calculated from a 4-parameter curve analysis (XLfit, IDBS software) and was determined from a minimum of 3 experiments. For cell lines that failed to achieve an IC50, the highest concentration tested (10 μmol/L) is listed.
Concentrations	0~10µM
Incubation time	4 days

Animal Experiment
Animal models	MCF7-neo/HER2 tumor xenograft model
Formulation	0.5% methylcellulose/0.2% Tween-80 (MCT)
Dosages	0,12.5, 25, 50, 75, 100mg/kg
Administration	oral gavage

Chemical Information

Molecular Weight	458
Formula	C24H32ClN5O2
CAS Number	1001264-89-6
Solubility (25°C)	DMSO 90 mg/mL Ethanol 60 mg/mL
Storage	Powder          -20°C   3 years ;  4°C   2 years In solvent       -80°C   6 months ;  -20°C   1 month

Conversion of different model animals based on BSA (PMID: 27057123)

Species	Mouse	Rat	Rabbit	Guinea pig	Hamster	Dog
Weight (kg)	0.02	0.15	1.8	0.4	0.08	10
Body Surface Area (m2)	0.007	0.025	0.15	0.05	0.02	0.5
Km factor	3	6	12	8	5	20

Animal A (mg/kg) = Animal B (mg/kg) multiplied by	Animal B Km
	Animal A Km

For example, to modify the dose of Compound A used for a mouse (20 mg/kg) to a dose based on the BSA for a rat, multiply 20 mg/kg by the K_m factor for a mouse and then divide by the K_m factor for a rat. This calculation results in a rat equivalent dose for Compound A of 10 mg/kg.

References

[1] Blake et al. J Med Chem. Discovery and Preclinical Pharmacology of a Selective ATP-competitive Akt Inhibitor (GDC-0068) for the Treatment of Human Tumors.