CROSS REFERENCE TO THE RELATED APPLICATIONS

This application claims the benefit of earlier Indian provisional patent application No. IN 202241043027, filed on July 27, 2022; the entire contents of each of which are incorporated by reference herein.

FIELD OF THE INVENTION

The present disclosure generally relates to the field of pharmaceutical sciences and more specifically relates to process for the preparation of Larotrectinib sulfate polymorphic form.

BACKGROUND OF THE INVENTION

Larotrectinib sulfate, chemically known as (3S)-N-{5-[(2R)-2-(2,5-difluorophenyl)-l- pyrrolidinyl] pyrazolo[l,5-a] pyrimidin-3-yl)-3-hydroxy-l-pyrrolidinecarboxamide sulfate having the structure shown in Formula-I, is a kinase inhibitor.

Larotrectinib sulfate is approved and is being marketed under the brand name Vitrakvi® for the treatment of adult and pediatric patients with solid tumors that have a neurotrophic receptor tyrosine kinase (NTRK) gene fusion without a known acquired resistance mutation, are metastatic or where surgical resection is likely to result in severe morbidity and have no satisfactory alternative treatments .

United States Pat. No. US 8513263 discloses Larotrectinib, its pharmaceutically acceptable salt and process for the preparation of same.

The inventors of the present disclosure have developed novel crystalline form of Larotrectinib sulfate which is stable, and also provided processes for the preparation of the same. OBJECT AND SUMMARY OF THE INVENTION

In one object, the present invention provides crystalline Form Ml of Larotrectinib sulfate.

In another object, the present invention provides a process for the preparation of crystalline Form Ml of Larotrectinib sulfate comprising the steps of: a) dissolving Larotrectinib sulfate in a suitable solvent optionally in presence of a coformer; b) adding an anti-solvent; and c) isolating crystalline Form Ml of Larotrectinib sulfate.

In one more object, the present invention provides a process for the preparation of crystalline Form Ml of Larotrectinib sulfate comprising the steps of: a) taking Larotrectinib in a suitable solvent; b) adding sulfuric acid; c) adding an anti-solvent optionally in presence of a coformer; and d) isolating crystalline Form Ml of Larotrectinib sulfate.

In another object, the present invention provides a process for the preparation of crystalline Form Ml of Larotrectinib sulfate comprising the steps of: a) heating Larotrectinib sulfate in a first solvent; b) removing the solvent and adding second solvent; c) optionally seeding; d) diluting with third solvent; and e) isolating crystalline Form Ml of Larotrectinib sulfate.

BRIEF DESCRIPTION OF THE DRAWING

FIGURE 1: X-ray powder diffractogram of crystalline Form Ml of Larotrectinib sulfate.

FIGURE 2: Differential scanning calorimetry ("DSC") thermogram of crystalline Form Ml of Larotrectinib sulfate.

FIGURE 3: Thermogravimetric analysis ("TGA") thermogram of crystalline Form Ml of Larotrectinib sulfate.

FIGURE 4: ’H-NMR spectrum of crystalline Form Ml of Larotrectinib sulfate.

FIGURE 5: FT-IR spectra of crystalline Form Ml of Larotrectinib sulfate.

FIGURE 6: Overlay of FT-IR spectra of Form 1-HS (top) and Form Ml (bottom).

DETAILED DESCRIPTION OF THE INVENTION The present invention provides crystalline Form Ml of Larotrectinib sulfate and process for the preparation of the same.

Instrumentation Details:

Powder X-Ray Diffraction (PXRD)

The powder X-ray diffraction patterns of said polymorphs of the invention were measured on a Bruker D8 ADVANCE powder diffractometer equipped with a goniometer of 0/20 configuration and LYNXEYE detector. The Cu-anode X-ray tube was operated at 40kV and 30mA. The experiments were conducted over the 20 range of 2.0°-50.0°, using a 0.033° step size and 62 seconds step time.

Differential Scanning Calorimetry (DSC)

The DSC measurement was carried out on a TA Q2000 instrument. The scan was performed from 50°C to 250°C at a heating rate of 10.0°C/min with nitrogen purging at a flow rate of 50 mL/min. Standard aluminum pans covered by lids with three pinholes were used.

Thermogravimetric Analysis (TGA)

The TGA measurement was carried out on a TA Q5000 instrument. The scan was performed from ambient temperature to 350°C at a heating rate of 10.0 °C/min and purging with nitrogen at a flow rate of 25 mL/min.

Fourier Transform Infrared (FTIR) spectroscopy

The FTIR spectra were recorded on a PerkinElmer SpectrumTM One instrument in the range of 400-4000 cm-1 by using KBr pellets.

In one aspect, the present invention provides crystalline Form Ml of Larotrectinib sulfate.

In one embodiment of the present invention, crystalline Form Ml of Larotrectinib sulfate may be characterized by a PXRD pattern having significant peaks at 20 angle positions at about 7.07, 9.12, 13.96, 20.44, 20.71 and 22.45 ±0.2 °0.

In another embodiment of the present invention, crystalline Form Ml of Larotrectinib sulfate may further characterized by a PXRD pattern having significant peaks at 20 angle positions at about 7.07, 9.12, 13.96, 17.36, 19.67, 20.44, 20.71, 22.45 and 25.44 ±0.2 °0.

In one more embodiment of the present invention, the crystalline Form Ml of Larotrectinib sulfate may further characterized by a PXRD pattern having significant peaks at 20 angle In one more aspect, the present invention provides a process for the preparation of crystalline Form Ml of Larotrectinib sulfate comprising the steps of: a) taking Larotrectinib in a suitable solvent; b) adding sulfuric acid; c) adding an anti-solvent optionally in presence of a coformer; and d) isolating crystalline Form Ml of Larotrectinib sulfate.

According to the present invention, Larotrectinib is dissolved in a suitable solvent and added sulfuric acid optionally in presence of a coformer. The suitable solvent includes, but not limited to polar solvents such as alcohols selected from methanol, ethanol, propanol, isopropanol, 1-butanol, 2-butanol and 1-pentanol; preferably methanol.

Next, to the resulting Larotrectinib solution is added sulfuric acid optionally in presence of a coformer. The sulfuric acid used in the present invention may a precooled sulfuric acid solution in a suitable solvent. The addition of sulfuric acid solution is performed at 10-30 °C, preferably 20-25 °C, more preferably 15-20 °C.

The suitable solvent for the preparation of sulfuric acid solution includes, but not limited to polar solvents such as alcohols selected from methanol, ethanol, isopropanol, 1-propanol, 1- butanol, 2-butanol and 1-pentanol; preferably methanol. The suitable co- former includes, but not limited to organic acids such as maleic acid, salicylic acid, fumaric acid, adipic acid, L- pyroglutamic acid, succinic acid and the like.

Within the context of the present invention, the above resulting reaction solution may heated to 70-90 °C; preferably at 60-80 °C; more preferably at 50-55 °C. Next, the resulting reaction solution is cooled to ambient temperature and an anti-solvent is added is added to isolate the crystalline Form Ml of Larotrectinib sulfate. The suitable anti-solvent includes but not limited to ethers such as methyl tert-butyl ether, cyclopentylmethyl ether, diethyl ether and diisopropyl ether; preferably methyl tert-butyl ether.

In one embodiment, the ratio of the solvent and anti-solvent is 1:5; preferably 1:4; more preferably 1:2.5-3.

Within the context of the present invention, isolation can be done using any techniques in the art such as, decantation, filtration by gravity or suction, centrifugation. In a particular useful embodiment of the present invention the solid is isolated by filtration followed by washing and drying. In another object, the present invention provides a process for the preparation of crystalline Form Ml of Larotrectinib sulfate comprising the steps of: a) heating Larotrectinib sulfate in a first solvent; b) removing the solvent and adding second solvent; c) optionally seeding; d) diluting with third solvent; and e) isolating crystalline Form Ml of Larotrectinib sulfate.

In one embodiment, Larotrectinib sulfate is heated in a first solvent. The first solvent includes but not limited alcohols such as methanol, ethanol, propanol, isopropanol, 1 -butanol, 2- butanol and 1-pentanol. In a preferred embodiment the suitable solvent is methanol.

In another embodiment, the Larotrectinib sulfate reaction mixture is heated to 45-55 °C; preferably 47 ±2 °C for 40-50 hrs; preferably 45-50 hrs; and the solvent is removed to retain 2-5 volumes with respect to the Larotrectinib sulfate.

Next, the resulting reaction mixture containing 2-5 volumes of first solvent is added a second solvent. The second solvent may in the form of mixture of solvents or a second solvent in combination with water. The second solvent includes but not limited to alcohols such as methanol, ethanol, propanol, isopropanol, 1-butanol, 2-butanol and 1-pentanol and the like; esters such as methyl acetate, ethyl acetate, propyl acetate, butyl acetate; ethers such as tertbutyl methyl ether, cyclopentylmethyl ether, diethyl ether and diisopropyl ether and the like. In a preferred embodiment the suitable second solvent is a combination of water-ethyl acetate, water-methyl tert-butyl ether and water-ethanol.

In another embodiment, the reaction mixture is optionally seeded with crystalline Larotrectinib sulfate Form Ml and diluted with a third solvent or mixtures thereof and isolated crystalline Form Ml of Larotrectinib sulfate.

In one embodiment, the second solvent of the above embodiment may also be used as a third solvent. The third solvent includes but not limited to esters such as methyl acetate, ethyl acetate, propyl acetate, butyl acetate; ethers such as tert-butyl methyl ether, cyclopentylmethyl ether, diethyl ether and diisopropyl ether and the like. In a preferred embodiment the suitable solvent is ethyl acetate, methyl-tert-butyl ether.

Within the context of the present invention, isolation can be done using any techniques in the art such as, decantation, filtration by gravity or suction, centrifugation. In a particular useful embodiment of the present invention the solid is isolated by filtration followed by washing and drying.

In one more embodiment, Larotrectinib or its pharmaceutically acceptable salt can be prepared as per the process known in US WO 2010048314; WO 2017201241 and in literature.

Yet another embodiment, the Larotrectinib sulfate may in a crystalline form or may in an amorphous form.

DSC and TGA analysis

The DSC measurements were carried out on a Q2000 DSC instrument manufactured by TA instruments. TGA data were recorded using a Q5000 TGA instrument manufactured by TA instruments. DSC of Form Ml shows a transition around 130°C, which can be attributed to a dehydration event, followed by conversion to an anhydrous form which melts around 199°C. Figure 3 shows the TGA data of Form Ml, which shows a weight loss of around 1.5% till its melting.

Moisture content analysis:

Moisture content analysis of Form Ml was carried out using a KF instrument (Metrohm, 901 titrando). Form Ml shows a moisture content in the range of 1.5-1.7%, further confirming Form Ml is a hydrate, preferably a hemihydrate.

FTIR data:

FTIR data of Larotrectinib sulfate Form Ml is significantly different in comparison with Form 1-HS (Figure 4). The characteristic IR bands for Form Ml are 3428 cm' ¹ , 1682cm' ¹ , 1251 cm' ¹ , 1060 cm' ¹ , 751 cm' ¹ (Figure 4). Significant changes are observed in the absorptions corresponding to stretching vibrations of N-H/O-H, >C=O, C-N/C-F functional groups indicating that the larotrectinib sulfate molecule is packed differently in Form Ml compared to Form LHS (Figure 5).

Stability Study

In yet another embodiment, the physical and chemical stability of Larotrectinib sulfate Form Ml were determined by storing the samples at 40°C/75% relative humidity (RH) and at 25°C/60% RH conditions for six months as shown in Table 1. The samples were analyzed by PXRD and HPLC. Larotrectinib sulfate Form Ml was found to be physically and chemically stable at 40°C/75% relative humidity (RH) and at 25°C/60% relative humidity (RH) conditions for up to six months.

Table 1. Summary of stability studies on Form Ml

In view of the above description and the examples below, one of ordinary skill in the art will be able to practice the invention as claimed without undue experimentation. The foregoing will be better understood with reference to the following examples that detail certain procedures for the preparation of molecules according to the present invention. All references made to these examples are for the purposes of illustration. The following examples should not be considered exhaustive, but merely illustrative of only a few of the many aspects and embodiments contemplated by the present disclosure.

Examples:

Example 1:

In an RBF, charged Larotrectinib sulfate (50 mg), maleic acid (17 mg) and methanol (0.9mL) at 25±5°C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C and was added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form Ml . Yield: 20 mg. Example 2:

In an RBF, charged Larotrectinib sulfate (50 mg), salicylic acid (20 mg) and methanol (0.6 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5°C, and was added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form Ml . Yield:25 mg.

Example 3:

In an RBF, charged Larotrectinib sulfate (50 mg), fumaric acid (16.5 mg) and methanol (0.9 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C, and was added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 35 mg.

Example 4:

In an RBF, charged Larotrectinib sulfate (50 mg), adipic acid (21 mg) and methanol (0.6 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C, and was added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 30 mg.

Example 5:

In an RBF, charged Larotrectinib sulfate (50 mg), L-pyroglutamic acid (18 mg) and methanol (0.6 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C and was added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 25 mg. Example 6:

In an RBF, charged Larotrectinib sulfate (50 mg), succinic acid (17 mg) and methanol (0.6 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C, added methyl tert-butyl ether (3 mL) and maintained under stirring at 25±5 °C for 4 hrs. The product obtained was filtered at 25-30 °C and suck-dried for 15 mins. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 28 mg.

Example 7:

In an RBF, charged Larotrectinib sulfate (1 g) and methanol (15 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C and was added methyl tert-butyl ether (75 mL) and maintained under stirring at 25±5 °C for 3-4 hrs. The product obtained was filtered, suck-dried for 15-20 min at 25-30 °C and dried under vacuum at 40 °C for 14 hrs. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 0.9 g.

Example 8:

In an RBF, charged Larotrectinib sulfate (5 g) and methanol (75 mL) at 25±5 °C. Heated the contents to 50-55 °C to obtain clear solution and maintained under stirring for 48 hrs at 50-55 °C. The reaction mass was slowly cooled to 25±5 °C and was added methyl tert-butyl ether (300 mL) and maintained under stirring at 25±5 °C for 3-4 hrs. The product obtained was filtered, suck-dried for 15-20 min at 25-30 °C and dried under vacuum at 40 °C for 10 hrs. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form ML Yield: 4.85 g.

Example 9:

In an RBF, dissolve Larotrectinib base (300 mg) in Methanol (4.5 mL) at 25±5 °C and clear solution was cooled to 15±5 °C. To this was added a pre-cooled solution of sulfuric acid (67 mg) in methanol (0.6 mL) at 15±5 °C. The obtained clear solution was stirred for 90 mins at 20-25 °C, then heated to 50-55 °C and maintained under stirring for 15 hrs at 50-55 °C. The reaction mass was cooled to 25±5 °C and was added methyl tert-butyl ether (12 mL) and maintained under stirring at 25±5°C for 16 hrs. The product obtained was filtered, suck-dried for 15-20 mins at 25-30 °C. The solid obtained was identified by PXRD as crystalline form of Larotrectinib sulfate Form Ml. Yield: 225 mg.

Example 10:

In an RBF, charged Larotrectinib sulfate (2 g) and methanol (34 mL) at 25±5°C. Heated the contents to 50-55°C to obtain a clear solution and maintained under stirring at 50-55°C for 16-18h. Distilled the reaction mass up to 3-4mL under vacuum at 50-55°C. Cooled the reaction mass to 25±2°C and added Methyl tert-butyl ether (10 mL), added Larotrectinib sulfate Form Ml seeds (10 mg) and maintained under stirring at 25±2°C for l-2h. Added methyl tert-butyl ether (10 mL) and stirred for l-2h at 25±2°C. The product obtained was filtered, washed with Methyl tert-butyl ether (5 mL) suck-dried for 10-15min at 25-30°C and dried under vacuum at 30°C for 16h. The solid obtained was identified by PXRD as novel crystalline form of Larotrectinib sulfate Form ML Yield: 1.75 g.

Example 11:

In an RBF, charged Larotrectinib sulfate (1 g) and methanol (25 mL) at 25±2°C. Heated the contents to 45-50 °C and stirred for 48 hrs. Distilled the reaction mass up to 2 mL under vacuum at 47±2 °C. Cooled the reaction mass to 42±2 °C, added water (40 mg), added ethyl acetate (3 mL) and stir the reaction mass for 30 minutes. To the obtained clear solution added seeds of Larotrectinib sulfate Form Ml (-1%) and maintained under stirring at 42±2°C for lOminutes. To the hazy reaction mass slowly added ethyl acetate (7 mL) at 42±2°C and stir the reaction mass for 2 h at 42±2 °C. Slowly cooled to 27±2°C and maintained for 14h. The product obtained was filtered, washed with mixture of methanol and ethyl acetate (l:5v/v, 2 mL), suck-dried for 10-15min at 25-30 °C. The solid obtained was further dried under vacuum at 25-30 °C for 16 hrs and identified by PXRD as novel crystalline form of Larotrectinib sulfate Form ML Yield: 900 mg.

Example 12:

In an RBF, charged Larotrectinib sulfate (1g) and methanol (25 mL) at 25±2 °C. Heated the contents to 45-50 °C and stirred for 48 hrs. Distilled the reaction mass up to 2mL under vacuum at 47±2 °C. Cooled the reaction mass to 42±2°C, added water(40mg), added methyl tert-butyl ether (3 mL) and stir the reaction mass for 30minutes. To the obtained clear solution added seeds of Larotrectinib sulfate Form Ml (-1%) and maintained under stirring at 42±2 °C for lOminutes. To the hazy reaction mass slowly added methyl tert-butyl ether (7 mL) at 42±2 °C and stir the reaction mass for 2 hrs at 42±2 °C. The product obtained was filtered, washed with mixture of methanol and methyl tert-butyl ether (l:5v/v, 2 mL), suck-dried for 10-15min at 25-30 °C. The solid obtained was further dried under vacuum at 25-30 °C for 16 hrs and identified by PXRD as novel crystalline form of Larotrectinib sulfate Form ML Yield: 930 mg.

Example 13:

In an RBF, charged Larotrectinib sulfate ( 1g) and methanol (25 mL) at 25±2 °C. Heated the contents to 45-50 °C and stirred for 48 hrs. Distilled the reaction mass completely under vacuum at 47±2 °C. Cooled the reaction mass to 42±2°C, added ethanol (2 mL) and water (40 mg) at 42±2 °C. Slowly added ethyl acetate (9 mL) at 42±2 °C. To the resulting clear solution added seeds of Larotrectinib sulfate Form Ml (5 mg) and stirred the reaction mass for 2h at 42±2 °C. The product obtained was filtered, washed with mixture of ethanol and ethyl acetate (l:5v/v, ImL) and suck-dried for 10-15min at 25-30 °C. The solid obtained was further dried at 25-30 °C under vacuum for 16 hrs and identified by PXRD as novel crystalline form of Larotrectinib sulfate Form ML Yield: 910 mg.