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Loratinib (formerly also known as Lorlatinib, PF-06463922; trade name:Lorbrena) is a potent, orally bioavailable, brain-penetrant,ATP-competitive, anddual ALK/ROS1 inhibitor with potential antitumor activity. With Ki values of less than 0.02 nM, 0.07 nM, and 0.7 nM, respectively, it inhibits ROS1, ALK (WT), and ALK (L1196M).The FDA approved loratinib for the treatment of patients with metastatic non-small cell lung cancer that is positive for anaplastic lymphoma kinase (ALK). After being administered, PF-06463922 binds to and inhibits ROS1 kinases as well as ALK kinases. This disrupts ALK and ROS1-mediated signaling and ultimately stops tumor cell growth. In addition to treating ROS1 fusion-positive cancers, including those that need drugs with CNS-penetrating capabilities, PF-06463922 may be able to reverse the effects of crizotinib resistance caused by ROS1 mutation.
Physicochemical Properties
| Molecular Formula | C21H19FN6O2 |
| Molecular Weight | 406.41 |
| Exact Mass | 406.155 |
| Elemental Analysis | C, 62.06; H, 4.71; F, 4.67; N, 20.68; O, 7.87 |
| CAS # | 1454846-35-5 |
| Related CAS # | 1924207-18-0 (acetate);2135926-03-1;1454846-35-5;2306217-6 (hydrate); |
| PubChem CID | 71731823 |
| Appearance | White to off-white solid powder |
| Density | 1.4±0.1 g/cm3 |
| Boiling Point | 675.0±55.0 °C at 760 mmHg |
| Flash Point | 362.1±31.5 °C |
| Vapour Pressure | 0.0±2.1 mmHg at 25°C |
| Index of Refraction | 1.687 |
| LogP | 1.24 |
| Hydrogen Bond Donor Count | 1 |
| Hydrogen Bond Acceptor Count | 7 |
| Rotatable Bond Count | 0 |
| Heavy Atom Count | 30 |
| Complexity | 700 |
| Defined Atom Stereocenter Count | 1 |
| SMILES | FC1C([H])=C([H])C2C(N(C([H])([H])[H])C([H])([H])C3C(=C(C#N)N(C([H])([H])[H])N=3)C3C([H])=NC(=C(C=3[H])O[C@]([H])(C([H])([H])[H])C=2C=1[H])N([H])[H])=O |
| InChi Key | IIXWYSCJSQVBQM-LLVKDONJSA-N |
| InChi Code | InChI=1S/C21H19FN6O2/c1-11-15-7-13(22)4-5-14(15)21(29)27(2)10-16-19(17(8-23)28(3)26-16)12-6-18(30-11)20(24)25-9-12/h4-7,9,11H,10H2,1-3H3,(H2,24,25)/t11-/m1/s1 |
| Chemical Name | (16R)-19-amino-13-fluoro-4,8,16-trimethyl-9-oxo-17-oxa-4,5,8,20-tetrazatetracyclo[16.3.1.02,6.010,15]docosa-1(22),2,5,10(15),11,13,18,20-octaene-3-carbonitrile |
| Synonyms | Lorbrena; PF06463922; PF-6463922; PF6463922; PF 6463922; PF 06463922; PF-06463922;Lorlatinib; 1454846-35-5; Lorbrena; Lorviqua; lorlatinibum; PF06463922 |
| HS Tariff Code | 2934.99.09.01 |
| 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 | ALKL1196 (IC50 = 15-43 nM); ALKG1269A (IC50 = 14-80 nM); ALK1151Tins (IC50 = 38-50 nM); ALKG1202R (IC50 = 77-113 nM); ALKWT (IC50 <0.07 nM); ALKL1996M (IC50 = 0.6 nM); ALKG1269A (IC50 = 0.9 nM); ALK1151Tins (IC50 = 0.1 nM); ALKL1152R (IC50 <0.1 nM); ALKS1206Y (IC50 = 0.2 nM); ALKC1156Y (IC50 <0.1 nM); ALKF1174L (IC50 <0.1nM) |
| ln Vitro | PF-06463922 exhibits a broad range of ALK clinical mutations with IC50 values between 0.2 and 77 nM, as well as notable cell activity against ALK. [1] In HCC78 human NSCLC cells carrying SLC34A2-ROS1 fusions and BaF3-CD74-ROS1 cells expressing human CD74-ROS1, PF-06463922 dramatically reduces cell proliferation and induces cell apoptosis. **[2]** In NSCLC cells expressing either non-mutant ALK or mutant ALK fusions, PF-06463922 also exhibits strong growth inhibitory activity and causes apoptosis.[3] |
| ln Vivo | PF-06463922 exhibits a low propensity for p-glycoprotein 1-mediated efflux, a moderate volume of distribution, a reasonable half-life, low plasma clearance, and 100% bioavailability in rats.[1] PF-06463922 exhibits cytoreductive antitumor efficacy in NIH3T3 xenograft models that express human CD74-ROS1 and Fig-ROS1 through the inhibition of downstream signaling molecules and ROS1 phosphorylation, in addition to inhibiting the cell cycle protein Cyclin D1 in tumors.[2] In mice with tumor xenografts expressing EML4-ALK, EML4-ALK-L1196M, EML4-ALK-G1269A, EML4-ALK-G1202R, or NPM-ALK, PF-06463922 also exhibits strong antitumor activity in vivo.[3] |
| Enzyme Assay | Microfluidic mobility shift assay is used to measure kinase activity in recombinant human wild-type and mutant ALK kinase domain proteins (amino acids 1093–1411), which are produced in-house via baculoviral expression and autophosphorylation with MgATP. The reactions contained 3 μM 5-FAM-KKSRGDYMTMQIG-CONH2), 5 mM MgCl2, 1.3 nM wild-type ALK or 0.5 nM mutant ALK (suitable to produce 15-20% phosphorylation of peptide substrate after 1 hour of reaction), and the Kmlevel of ATP in 25 mM Hepes, pH 7.1. The results of kinetic and crystallographic investigations demonstrate that the inhibitors are ATP-competitive. Fitting the conversion (%) to a competitive inhibition equation yields the Kivalues. The procedure for assaying ROS1 enzyme is the same as that for ALK, with the exception that 0.25 nM recombinant human ROS1 catalytic domain (amino acids 1883-2347) is used. A 206-kinase panel is utilized to assess the selectivity of kinase inhibitors. |
| Cell Assay | In 96-well plates, cells are sown in growth medium with 10% FBS, and they are incubated at 37°C for the entire night. The cells are incubated at 37°C for 72 hours after serial dilutions of Lorlatinib or suitable controls are added to the assigned wells the following day. To ascertain the relative cell numbers, a CellTiter-Glo assay is conducted. A four-parameter analytical method is used to fit a concentration-response curve and determine IC50 values. |
| Animal Protocol | In LSL-FIG-ROS1;Cdkn2a−/−;LSL-Luc mice, de novoGBM tumorigenesis is induced by intracranial stereotactic injections of Adeno-Cre, as previously reported. BLI is used to track the development of tumors as will be discussed below. Animals are randomly assigned to either vehicle control or 3-, 7-, or 14-day treatments with the prescribed doses of lerlatinib once tumors reach a specific size (107 p -1·s -1·cm -2·sr -1). The medication is delivered via s.c. implanted Alzet osmotic pumps. Following therapy, GBM tumors are microdissected, tissues are flash-frozen in liquid N2, and mice are killed. For histology, the remaining brains are processed. |
| ADME/Pharmacokinetics |
Absorption, Distribution and Excretion The median lorlatinib Tmax was 1.2 hours (0.5 to 4 hours) following a single oral 100 mg dose and 2 hours (0.5 to 23 hours) following 100 mg orally once daily at steady state. The mean absolute bioavailability is 81% (90% CI 75.7%, 86.2%) after oral administration compared to intravenous administration. Administration of lorlatinib with a high fat, high-calorie meal (approximately 1000 calories with 150 calories from protein, 250 calories from carbohydrate, and 500 to 600 calories from fat) had no clinically meaningful effect on lorlatinib pharmacokinetics. Following a single oral 100 mg dose of radiolabeled lorlatinib, 48% of the radioactivity was recovered in urine (<1% as unchanged) and 41% in feces (about 9% as unchanged). The mean (CV%) steady-state volume of distribution (Vss) was 305 L (28%) following a single intravenous dose. The mean oral clearance (CL/F) was 11 L/h (35%) following a single oral 100 mg dose and increased to 18 L/h (39%) at steady state, suggesting autoinduction. Metabolism / Metabolites In vitro, lorlatinib is metabolized primarily by CYP3A4 and UGT1A4, with minor contribution from CYP2C8, CYP2C19, CYP3A5, and UGT1A3. In plasma, a benzoic acid metabolite (M8) of lorlatinib resulting from the oxidative cleavage of the amide and aromatic ether bonds of lorlatinib accounted for 21% of the circulating radioactivity in a human [14C] mass balance study. The oxidative cleavage metabolite, M8, is pharmacologically inactive. Biological Half-Life The mean plasma half-life (t½) of lorlatinib was 24 hours (40%) after a single oral 100 mg dose of lorlatinib. |
| Toxicity/Toxicokinetics |
Hepatotoxicity In large early clinical trials, elevations in serum aminotransferase levels occurred in up to 28% of patients treated with standard doses of lorlatinib but were above 5 times ULN in only 2% of patients and rarely led to early discontinuation of therapy. The abnormalities were typically transient, asymptomatic and not associated with jaundice. In prelicensure clinical trials there were no instances of clinically apparent liver injury. In contrast, most other ALK inhibitors have been linked to instances of acute liver injury which can be severe and even fatal. The liver injury typically arises within 4 to 12 weeks of starting therapy and presents with marked elevations in serum aminotransferase levels followed by jaundice and progressive hepatic dysfunction. While lorlatinib has not been associated with instances of severe liver injury, it has had limited clinical use. Likelihood score: E* (unproven but suspected rare cause of clinically apparent liver injury). Effects During Pregnancy and Lactation ◉ Summary of Use during Lactation No information is available on the clinical use of lorlatinib during breastfeeding. The manufacturer recommends that breastfeeding be discontinued during lorlatinib therapy and for 7 days after the last dose. ◉ Effects in Breastfed Infants Relevant published information was not found as of the revision date. ◉ Effects on Lactation and Breastmilk Relevant published information was not found as of the revision date. Protein Binding In vitro, lorlatinib was 66% bound to plasma proteins at a concentration of 2.4 µM. The blood-to-plasma ratio was 0.99. |
| References |
[1]. J Med Chem . 2014 Jun 12;57(11):4720-44. [2].Mol Cancer Ther (2013) 12 (11_Supplement): A277. [3]. Mol Cancer Ther (2013) 12 (11_Supplement): C253. |
| Additional Infomation |
Pharmacodynamics Based on data from Study B7461001, exposure-response relationships for Grade 3 or 4 hypercholesterolemia and for any Grade 3 or 4 adverse reaction were observed at steady-state exposures achieved at the recommended dosage, with higher probability of the occurrence of adverse reactions with increasing lorlatinib exposure. In 295 patients who received lorlatinib at the recommended dosage of 100 mg once daily and had an ECG measurement in the same Study B7461001, the maximum mean change from baseline for their PR interval was 16.4 ms (2-sided 90% upper confidence interval [CI] 19.4 ms). Among the 284 patients with PR interval <200 ms at baseline, 14% had PR interval prolongation ≥200 ms after starting use with lorlatinib. The prolongation of PR interval occurred in a concentration-dependent manner and atrioventricular block occurred in 1% of patients. Finally, in 275 patients who received lorlatinib at the recommended dosage in the activity-estimating portion of Study B7461001, no large mean increases from baseline in the QTcF interval (i.e., >20 ms) were detected. |
Solubility Data
| Solubility (In Vitro) |
DMSO: ~81 mg/mL (~199.3 mM) Water: <1 mg/mL Ethanol: ~30 mg/mL warmed (~73.8 mM) |
| Solubility (In Vivo) |
Solubility in Formulation 1: ≥ 2.5 mg/mL (6.15 mM) (saturation unknown) in 10% DMSO + 40% PEG300 + 5% Tween80 + 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 25.0 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. Solubility in Formulation 2: 2.5 mg/mL (6.15 mM) in 10% DMSO + 90% (20% SBE-β-CD in Saline) (add these co-solvents sequentially from left to right, and one by one), suspension solution; with ultrasonication. For example, if 1 mL of working solution is to be prepared, you can add 100 μL of 25.0 mg/mL clear DMSO stock solution to 900 μL of 20% SBE-β-CD physiological saline solution and mix evenly. 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. Solubility in Formulation 3: ≥ 2.5 mg/mL (6.15 mM) (saturation unknown) in 10% DMSO + 90% Corn Oil (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 25.0 mg/mL clear DMSO stock solution to 900 μL of corn oil and mix evenly. Solubility in Formulation 4: ≥ 2.5 mg/mL (6.15 mM) (saturation unknown) in 5% DMSO + 40% PEG300 + 5% Tween80 + 50% Saline (add these co-solvents sequentially from left to right, and one by one), clear solution. 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. Solubility in Formulation 5: 2% DMSO+30% PEG 300+ddH2O: 5mg/mL (Please use freshly prepared in vivo formulations for optimal results.) |
| Preparing Stock Solutions | 1 mg | 5 mg | 10 mg | |
| 1 mM | 2.4606 mL | 12.3028 mL | 24.6057 mL | |
| 5 mM | 0.4921 mL | 2.4606 mL | 4.9211 mL | |
| 10 mM | 0.2461 mL | 1.2303 mL | 2.4606 mL |