Alofanib (known aslso as RPT835) is a novel, potent and selective allosteric inhibitor of fibroblast growth factor receptor 2 (FGFR2) with anticancer and antiangiogenic activity. In cancer cells expressing various FGFR2 isoforms, alofanib inhibited the phosphorylation of FRS2α with half maximal inhibitory concentrations (IC50) of 7 and 9 nmol/l. Alofanib inhibited FGF-mediated proliferation in a panel of four cell lines that represented several tumor types, including ovarian cancer, melanoma, and triple-negative breast cancer, with 50% growth inhibition (GI50) values ranging from 16 to 370 nmol/l. In comparison to brivanib and bevacizumab, alofanib dose-dependently inhibited the migration and proliferation of human and mouse endothelial cells (GI50 11-58 nmol/l). Alofanib treatment eliminated experimentally induced FGF-induced angiogenesis in vivo. Oral administration of alofanib produced strong antitumour activity and was well tolerated in a human tumour xenograft model driven by FGFR. Alofanib was noteworthy for its efficacy in models that expressed FGFR2. These findings demonstrate the effectiveness of alofanib as an FGFR2 inhibitor and offer compelling justification for its assessment in patients with FGFR2-driven malignancies.

Physicochemical Properties

Molecular Formula	C19H15N3O6S
Molecular Weight	413.403903245926
Exact Mass	413.068
Elemental Analysis	C, 55.20; H, 3.66; N, 10.16; O, 23.22; S, 7.76
CAS #	1612888-66-0
Related CAS #	1612888-66-0
PubChem CID	86280646
Appearance	Light yellow to yellow solid powder
Density	1.5±0.1 g/cm3
Boiling Point	659.6±65.0 °C at 760 mmHg
Flash Point	352.7±34.3 °C
Vapour Pressure	0.0±2.1 mmHg at 25°C
Index of Refraction	1.667
LogP	4
Hydrogen Bond Donor Count	2
Hydrogen Bond Acceptor Count	8
Rotatable Bond Count	5
Heavy Atom Count	29
Complexity	704
Defined Atom Stereocenter Count	0
SMILES	S(C1C=CC=C(C(=O)O)C=1)(NC1C(=CC(C)=C(C2C=NC=CC=2)C=1)[N+](=O)[O-])(=O)=O
InChi Key	QUQGQIASFYWKAB-UHFFFAOYSA-N
InChi Code	InChI=1S/C19H15N3O6S/c1-12-8-18(22(25)26)17(10-16(12)14-5-3-7-20-11-14)21-29(27,28)15-6-2-4-13(9-15)19(23)24/h2-11,21H,1H3,(H,23,24)
Chemical Name	3-[(4-methyl-2-nitro-5-pyridin-3-ylphenyl)sulfamoyl]benzoic acid
Synonyms	RPT835; RPT-835; RPT 835
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	FGFR2
ln Vitro	Alofanib primarily induces apoptosis through the cleavage of caspase 3, PARP, and Bcl-2 in SKOV3 cell line. The compound has little cytotoxic effect on ovarian cancer cells [1]. With IC50 values of 7 and 9 nmol/l, alofanib inhibits the phosphorylation of FRS2α in cancer cells expressing various FGFR2 isoforms. Alofanib inhibits FGF-mediated proliferation in a panel of four cell lines that represent several tumor types, including ovarian cancer, melanoma, and triple-negative breast cancer, with 50% growth inhibition (GI50) values ranging from 16 to 370 nmol/l. When compared to brivanib and bevacizumab, alofanib dose-dependently inhibits the migration and proliferation of human and mouse endothelial cells (GI50: 11-58 nmol/l)[2].
ln Vivo	The effectiveness of paclitaxel and carboplatin together was considerably enhanced by intravenous alofanib in a dose-dependent manner. In a mouse model of ovarian cancer, alofanib inhibits angiogenesis[1]. Oral alofanib administration is well tolerated and produces strong antitumour activity in a FGFR-driven human tumour xenograft model[2].
Cell Assay	After subjecting cells to alofanib (10, 100, and 1000 μM) for 72 hours, whole-cell lysates underwent immunoblotting analysis.
Animal Protocol	C57Bl/6 × DBA/2 F1 mice 15 mg/kg/d i.p.
References	[1]. Invest New Drugs . 2017 Apr;35(2):127-133. [2]. Eur J Cancer . 2016 Jul:61:20-8.
Additional Infomation	Alofanib is an inhibitor of the fibroblast growth factor receptor (FGFR) type 2 (FGFR2), with potential antineoplastic and anti-angiogenic activities. Upon administration, alofanib targets, allosterically binds to the extracellular domain of FGFR2 and inhibits the activity of FGFR2, which may result in the inhibition of basic FGF (bFGF)/FGFR2-related signal transduction pathways. This inhibits FGF-induced endothelial cell proliferation and migration, and inhibits the proliferation of FGFR2-overexpressing tumor cells. FGFR2, a receptor tyrosine kinase upregulated in many tumor cell types, plays a key role in cellular proliferation, migration and survival.

Solubility Data

Solubility (In Vitro)

DMSO: ≥ 30.1 mg/mL (~72.8 mM)

Solubility (In Vivo)

Solubility in Formulation 1: ≥ 2.08 mg/mL (5.03 mM) (saturation unknown) in 10% DMSO + 40% PEG300 +5% Tween-80 + 45% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 20.8 mg/mL clear DMSO stock solution to 400 μL PEG300 and mix evenly; then add 50 μL Tween-80 + to the above solution and mix evenly; then add 450 μL normal saline to adjust the volume to 1 mL. Preparation of saline: Dissolve 0.9 g of sodium chloride in 100 mL ddH₂ O to obtain a clear solution.  (Please use freshly prepared in vivo formulations for optimal results.)

Preparing Stock Solutions		1 mg	5 mg	10 mg
	1 mM	2.4190 mL	12.0948 mL	24.1896 mL
	5 mM	0.4838 mL	2.4190 mL	4.8379 mL
	10 mM	0.2419 mL	1.2095 mL	2.4190 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.