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Suzhou NeuPharma (Originator)
Non-Small Cell Lung Cancer Therapy
Solid Tumors Therapy
PHASE 2 Checkpoint Therapeutics, Cancer, lung (non-small cell) (NSCLC), solid tumour
RX518(CK-101) is an orally available third-generation and selective inhibitor of certain epidermal growth factor receptor (EGFR) activating mutations, including the resistance mutation T790M, and the L858R and exon 19 deletion (del 19) mutations, with potential antineoplastic activity.
In August 2019, Suzhou Neupharma and its licensee Checkpoint Therapeutics are developing CK-101 (phase II clinical trial), a novel third-generation, covalent, EGFR inhibitor, as a capsule formulation, for the treatment of cancers including NSCLC and other advanced solid tumors. In September 2017, the FDA granted Orphan Drug designation to this compound, for the treatment of EGFR mutation-positive NSCLC; in January 2018, the capsule was being developed as a class 1 chemical drug in China.
CK-101 (RX-518), a small-molecule inhibitor of epidermal growth factor receptor (EGFR), is in early clinical development at Checkpoint Therapeutics and Suzhou NeuPharma for the potential treatment of EGFR-mutated non-small cell lung cancer (NSCLC) and other advanced solid malignancies.
In 2015, Suzhou NeuPharma granted a global development and commercialization license to its EGFR inhibitor program, excluding certain Asian countries, to Coronado Biosciences (now Fortress Biotech). Subsequently, Coronado assigned the newly acquired program to its subsidiary Checkpoint Therapeutics.
In 2017, the product was granted orphan drug designation in the U.S. for the treatment of EGFR mutation-positive NSCLC.
There are at least 400 enzymes identified as protein kinases. These enzymes catalyze the phosphorylation of target protein substrates. The phosphorylation is usually a transfer reaction of a phosphate group from ATP to the protein substrate. The specific structure in the target substrate to which the phosphate is transferred is a tyrosine, serine or threonine residue. Since these amino acid residues are the target structures for the phosphoryl transfer, these protein kinase enzymes are commonly referred to as tyrosine kinases or serine/threonine kinases.
 The phosphorylation reactions, and counteracting phosphatase reactions, at the tyrosine, serine and threonine residues are involved in countless cellular processes that underlie responses to diverse intracellular signals (typically mediated through cellular receptors), regulation of cellular functions, and activation or deactivation of cellular processes. A cascade of protein kinases often participate in intracellular signal transduction and are necessary for the realization of these cellular processes. Because of their ubiquity in these processes, the protein kinases can be found as an integral part of the plasma membrane or as cytoplasmic enzymes or localized in the nucleus, often as components of enzyme complexes. In many instances, these protein kinases are an essential element of enzyme and structural protein complexes that determine where and when a cellular process occurs within a cell.
 The identification of effective small compounds which specifically inhibit signal transduction and cellular proliferation by modulating the activity of tyrosine and serine/threonine kinases to regulate and modulate abnormal or inappropriate cell proliferation, differentiation, or metabolism is therefore desirable. In particular, the identification of compounds that specifically inhibit the function of a kinase which is essential for processes leading to cancer would be beneficial.
 While such compounds are often initially evaluated for their activity when dissolved in solution, solid state characteristics such as polymorphism are also important. Polymorphic forms of a drug substance, such as a kinase inhibitor, can have different physical properties, including melting point, apparent solubility, dissolution rate, optical and mechanical properties, vapor pressure, and density. These properties can have a direct effect on the ability to process or manufacture a drug substance and the drug product. Moreover, differences in these properties
can and often lead to different pharmacokinetics profiles for different polymorphic forms of a drug. Therefore, polymorphism is often an important factor under regulatory review of the ‘sameness’ of drug products from various manufacturers. For example, polymorphism has been evaluated in many multi-million dollar and even multi-billion dollar drugs, such as warfarin sodium, famotidine, and ranitidine. Polymorphism can affect the quality, safety, and/or efficacy of a drug product, such as a kinase inhibitor. Thus, there still remains a need for polymorphs of kinase inhibitors. The present disclosure addresses this need and provides related advantages as well.
Crystalline form II-VIII of the compound presumed to be CK-101 (first disclosed in WO2015027222 ), for treating a disorder mediated by epidermal growth factor receptor (EGFR) eg cancer.
Example 1: Preparation of the compound of Formula I (N-(3-(2-((2,3-difluoro-4-(4-(2-hydroxyethyl)piperazin-l-yl)phenyl)amino)quinazolin-8-yl)phenyl)acrylamide)
 To a solution of l,2,3-trifluoro-4-nitrobenzene (2.5 g, 14 mmol, 1.0 eq.) in DMF (20 mL) was added K2C03 (3.8 g, 28 mmol, 2.0 eq.) followed by 2-(piperazin-l-yl)ethanol (1.8 g, 14 mmol, 1.0 eq.) at 0 °C and the mixture was stirred at r.t. overnight. The mixture was poured into ice-water (200 mL), filtered and dried in vacuo to afford 2-(4-(2,3-difluoro-4-nitrophenyl)piperazin-l-yl)ethanol (2.7 g, 67.5%).
 To a solution of 2-(4-(2,3-difluoro-4-nitrophenyl)piperazin-l-yl)ethanol (2.7 g, 9.0 mmol) in MeOH (30 mL) was added Pd/C (270 mg) and the resulting mixture was stirred at r.t.
overnight. The Pd/C was removed by filtration and the filtrate was concentrated to afford 2-(4-(4-amino-2,3-difluorophenyl)piperazin-l-yl)ethanol (2.39 g, 99% yield) as off-white solid.
 To a solution of 8-bromo-2-chloroquinazoline (15.4 g, 63.6 mmol, 1 eq. ) and (3-aminophenyl)boronic acid (8.7 g, 63.6 mmol, 1 eq.) in dioxane/H20 (200 mL/20 mL) was added Na2C03 (13.5 g, 127.2 mmol, 2 eq.), followed by Pd(dppf)Cl2 (2.6 g, 3.2 mmol, 0.05 eq.) under N2, then the mixture was stirred at 80 °C for 12 h. Then the solution was cooled to r.t.,
concentrated and the residue was purified via column chromatography (PE/EA=3 :2, v/v) to afford 3-(2-chloroquinazolin-8-yl)aniline as yellow solid (8.7 g, 53.7% yield).
 To a solution of 3-(2-chloroquinazolin-8-yl)aniline (8.7 g, 34 mmol, 1 eq.) in DCM ( 200 mL ) cooled in ice-bath was added TEA (9.5 mL, 68 mmol, 2 eq. ), followed by acryloyl chloride (4.1 mL, 51 mmol, 1.5 eq.) dropwise. The resulting mixture was stirred at r.t. for 1 h, then washed with brine, dried over anhydrous N2S04 concentrated and the residue was purified via column chromatography (PE/EA=l : 1, v:v) to afford N-(3-(2-chloroquinazolin-8-yl)phenyl)acryl amide as yellow solid(6.6 g, 65% yield).
 To a suspension of 2-(4-(4-amino-2,3-difluorophenyl)piperazin-l-yl)ethanol (83 mg,
0.32 mmol, 1 eq.) and N-(3-(2-chloroquinazolin-8-yl)phenyl)acrylamide (100 mg, 0.32 mmol, 1 eq.) in n-BuOH (5 mL) was added TFA (68 mg, 0.64 mmol, 2 eq.) and the resulting mixture was stirred at 90 °C overnight. The mixture was concentrated, diluted with DCM (20 mL) , washed with Na2C03 solution (20 mL), dried over anhydrous Na2S04, concentrated and the residue was purified via column chromatography (MeOH/DCM=l/30, v:v) to afford N-(3-(2-((2,3-difluoro-4-(4-(2-hydroxyethyl)piperazin-l-yl)phenyl)amino)quinazolin-8-yl)phenyl)acrylamide as a yellow solid(l6.3 mg, 9.5% yield). LRMS (M+H+) m/z calculated 531.2, found 531.2. 1H NMR
(CD3OD, 400 MHz) d 9.21 (s, 1 H), 7.19-8.01 (m, 10 H), 8.90 (s, 1 H), 6.41-6.49 (m, 3 H), 5.86 (m, 1 H), 3.98-4.01 (m, 3 H), 3.70-3.76 (m, 3 H), 3.40-3.49 (m, 2 H), 3.37-3.39 (m, 4 H), 3.18 (m, 2H).
Example 2. Preparation of Form I of the compound of Formula I
 Crude compound of Formula I (~30 g, 75% of weight based assay) was dissolved in ethyl acetate (3 L) at 55-65 °C under nitrogen. The resulting solution was filtered via silica gel pad and washed with ethyl acetate (3 L><2) at 55-65 °C. The filtrate was concentrated via vacuum at 30-40 °C to ~2.4 L. The mixture was heated up to 75-85 °C and maintained about 1 hour.
Then cooled down to 50-60 °C and maintained about 2 hours. The heat-cooling operation was repeated again and the mixture was then cooled down to 20-30 °C and stirred for 3 hours. The resulting mixture was filtered and washed with ethyl acetate (60 mL><2). The wet cake was dried via vacuum at 30-40 °C to get (about 16 g) of the purified Form I of the compound of Formula I.
Example 3. Preparation of Form III of the compound of Formula I
 The compound of Formula I (2 g) was dissolved in EtOH (40 mL) at 75-85 °C under nitrogen. n-Heptane (40 mL) was added dropwise into reaction at 75-85 °C. The mixture was stirred at 75-85 °C for 1 hour. Then cooled down to 50-60 °C and maintained about 2 hours. The heat-cooling operation was repeated again and continued to cool the mixture down to 20-30 °C and stirred for 3 hours. The resulting mixture was filtered and washed with EtOH/n-Heptane (1/1, 5 mL><2). The wet cake was dried via vacuum at 30-40 °C to get the purified Form III of the compound of Formula I (1.7 g).
Example 4. Preparation of Form IV of the compound of Formula I The crude compound of Formula I (15 g) was dissolved in ethyl acetate (600 mL) at 75-85 °C under nitrogen and treated with anhydrous Na2S04, activated carbon, silica metal scavenger for 1 hour. The resulting mixture was filtered via neutral Al203 and washed with ethyl acetate (300 mL><2) at 75-85 °C. The filtrate was concentrated under vacuum at 30-40 °C and swapped with DCM (150 mL). n-Heptane (75 mL) was added into this DCM solution at 35-45 °C, and then the mixture was cooled down to 20-30 °C slowly. The resulting mixture was filtered and washed with DCM/n-Heptane (2/1, 10 mL><3). The wet cake was dried via vacuum at 35-40 °C to get the purified Form IV of the compound of Formula I (9.6 g).
Example 5. Preparation of Form V of the compound of Formula I
 Polymorph Form III of the compound of Formula I was dried in oven at 80 °C for 2 days to obtain the polymorph Form V.
Example 6. Preparation of Form VI of the compound of Formula I
 The compound of Formula I (1 g) was dissolved in IPA (20 mL) at 75-85 °C under nitrogen. n-Heptane (20 mL) was added dropwise into reaction at 75-85 °C. The mixture was stirred at 45-55 °C for 16 hours. Then heated up to 75-85 °C and maintained about 0.5 hour.
Then cooled down to 45-55 °C for 0.5 hour and continued to cool the mixture down to 20-30 °C and stirred for 3 hours. Filtered and washed with IPA/n-Heptane (1/1, 3 mL><2). The wet cake was dried via vacuum at 75-80 °C for 2 hours to get the purified Form VI of the compound of Formula I.
Example 7. Preparation of Form VIII of the compound of Formula I
 The polymorph Form VI of the compound of Formula I was dried in oven at 80 °C for 2 days to obtain the polymorph Form VIII.
Example 8. X-ray powder diffraction (XRD)
 X-ray powder diffraction (XRD) patterns were obtained on a Bruker D8 Advance. A CuK source (=1.54056 angstrom) operating minimally at 40 kV and 40 mA scans each sample between 4 and 40 degrees 2-theta. The step size is 0.05°C and scan speed is 0.5 second per step.
Example 9. Thermogravimetric Analyses (TGA)
 Thermogravimetric analyses were carried out on a TA Instrument TGA unit (Model TGA 500). Samples were heated in platinum pans from ambient to 300 °C at 10 °C/min with a nitrogen purge of 60mL/min (sample purge) and 40mL/min (balance purge). The TGA temperature was calibrated with nickel standard, MP=354.4 °C. The weight calibration was performed with manufacturer-supplied standards and verified against sodium citrate dihydrate desolvation.
Example 10. Differential scanning calorimetry (DSC)
 Differential scanning calorimetry analyses were carried out on a TA Instrument DSC unit (Model DSC 1000 or 2000). Samples were heated in non-hermetic aluminum pans from ambient to 300 °C at 10 °C/min with a nitrogen purge of 50mL/min. The DSC temperature was calibrated with indium standard, onset of l56-l58°C, enthalpy of 25-29J/g.
Example 11. Hygroscopicity (DVS)
 The moisture sorption profile was generated at 25°C using a DVS Moisture Balance Flow System (Model Advantage) with the following conditions: sample size approximately 5 to 10 mg, drying 25°C for 60 minutes, adsorption range 0% to 95% RH, desorption range 95% to 0% RH, and step interval 5%. The equilibrium criterion was <0.01% weight change in 5 minutes for a maximum of 120 minutes.
Example 12: Microscopy
 Microscopy was performed using a Leica DMLP polarized light microscope equipped with 2.5X, 10X and 20X objectives and a digital camera to capture images showing particle shape, size, and crystallinity. Crossed polars were used to show birefringence and crystal habit for the samples dispersed in immersion oil.
Example 13: HPLC
 HPLCs were preformed using the following instrument and/or conditions.
///////////////CK-101 , CK 101 , CK101 , phase II , Suzhou Neupharma, Checkpoint Therapeutics , Orphan Drug designation, EGFR mutation-positive NSCLC, NSCLC, CANCER, SOLID TUMOUR, China, RX-518, AK543910