Process for the preparation of tovorafenib

Abstract

The present disclosure relates to safe and efficient processes for the synthesis of Tovorafenib.

Description

PROCESS FOR THE PREPARATION OF TOVORAFENIBCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of, and priority to, Indian Provisional Application No. 202411057163 filed July 27, 2024; Indian Provisional Application No. 202411078240 filed October 15, 2024; Indian Provisional Application No. 202511042342 filed May 1, 2025; and Indian Provisional Application No. 202511057249 filed June 14, 2025. The entire contents of the foregoing applications are incorporated by reference herein.FIELD OF THE DISCLOSURE

[0002] The present disclosure relates to safe and efficient processes for the synthesis of Tovorafenib and intermediates thereof.BACKGROUND OF THE DISCLOSURE

[0003] Tovorafenib, 6-amino-5-chloro-N-((lR)-l-(5-((5-chloro-4-(trifluoromethyl)- pyridine-2-yl)carbamoyl)-l,3-thiazol-2-yl)ethyl)pyrimidine-4-carboxamide, has the following chemical structure:

[0004] Tovorafenib is a kinase inhibitor indicated for the treatment of patients 6 months of age and older with relapsed or refractory pediatric low grade glioma (LGG) harboring a BRAF fusion or rearrangement, or BRAF V600 mutation. Tovorafenib is also under investigation for the treatment of recurrent, progressive or refractory melanoma or other solid tumors with alterations in the key proteins of the RAS / RAF / MEK / ERK pathway (referred to as the MAPK pathway).

[0005] Tovorafenib and its synthesis are disclosed in U.S. Patent No. 8,293,752 (referred to herein as US '752 or the '752 patent). The entire contents of the foregoing patent areincorporated by reference herein. US ‘752 discloses that an optically active form of an amine compound is produced by subjecting the corresponding ketone compound successively to oxime formation, reduction, and optical resolution with di-p-toluoyl-(D)-tartaric acid, or an optically active form of an amine compound is produced by chiral HPLC and then Tovorafenib is produced by an amide bond formation reaction between an optically active amine compound and an acid. The process is illustrated by the following Scheme 1.Scheme 1:

[0006] U.S. Patent No. 10,988,469 discloses the synthesis of Tovorafenib. (referred to herein as US '469 or the '469 patent). The entire contents of the foregoing patent are incorporated by referenced herein. US '469 discloses that an optically active form of an amine compound is produced by subjecting the corresponding ketone compound successively to an imine preparation, asymmetric reduction using hydrogen gas as a reducing agent under pressure, employing iridium catalyst and a ligand, and optical resolution with (S)-Mandelic acid, and then Tovorafenib is produced by an amide bond formation reaction between an optically active amine compound and an acid. The process is illustrated by the following Scheme 2.Scheme-2:

[0007] Accordingly, there is a need for additional processes allowing the efficient and safe synthesis of Tovorafenib.SUMMARY OF THE DISCLOSURE

[0008] The present disclosure provides processes for preparation of Tovorafenib or any salt thereof as well as novel intermediates, their preparation and their use in the preparation of Tovorafenib and any salt thereof.

[0009] The present disclosure provides Tovorafenib produced by the processes described herein.

[0010] The present disclosure also encompasses the use of Tovorafenib produced by the processes described herein for the preparation of pharmaceutical compositions and / or formulations.

[0011] In another aspect, the present disclosure provides pharmaceutical compositions comprising Tovorafenib produced according to the present disclosure.

[0012] The present disclosure includes processes for preparing the above mentioned pharmaceutical compositions. The processes include combining Tovorafenib obtained according to the processes of the present disclosure with at least one pharmaceutically acceptable excipient.

[0013] Tovorafenib obtained according to the processes of the present disclosure as defined herein and the pharmaceutical compositions or formulations thereof may be used as medicaments, such as for the treatment of Glioma, Craniopharyngioma, Langerhans cell histiocytosis and solid tumors.

[0014] The present disclosure also provides methods for the treatment of patients with Glioma, Craniopharyngioma, Langerhans cell histiocytosis and solid tumors by administering a therapeutically effective amount of Tovorafenib obtained according to the processes of the present disclosure, or at least one of the above pharmaceutical compositions, to a subject suffering from Glioma, Craniopharyngioma, Langerhans cell histiocytosis and solid tumors or otherwise in need of the treatment.

[0015] The present disclosure also provides uses of Tovorafenib obtained according to the processes of the present disclosure, or at least one of the above pharmaceutical compositions, for the manufacture of medicaments for treating e.g., Glioma, Craniopharyngioma, Langerhans cell histiocytosis and solid tumors.BRIEF DESCRIPTION OF THE DRAWINGS

[0016] FIG. 1 shows a characteristic X-ray powder diffraction (“XRPD”) pattern of Form 1 of Compound 7; and

[0017] FIG. 2 shows a characteristic XRPD pattern of Crystalline Tovorafenib.

[0018] FIG. 3 shows a characteristic XRPD pattern of Form A of Compound 4.

[0019] FIG. 4 shows a characteristic XRPD pattern of Form B of Compound 4.

[0020] FIG. 5 shows amino acid sequences of Transaminase enzymes.DETAILED DESCRIPTION OF THE DISCLOSURE

[0021] The present disclosure relates to safe and efficient processes for the synthesis of Tovorafenib.

[0022] The processes described in the literature have significant disadvantages. These processes involve the use of hazardous, toxic and costly reagents, such as hydrogen gas, iridium (Ir) catalyst and (S)-Monophos. In addition, iridium (Ir) catalyst is used in previous processes which is sensitive to degradation by atmospheric moisture and is difficult to handle. The prior art processes involve the use of hydrogen gas which is highly flammable and require autoclave for the process.

[0023] In contrast to the prior art processes, the processes of present disclosure does not require the use of toxic reagents, or of iridium (Ir) catalyst and (S)-Monophos. Moreover, the process of the present disclosure enables a more efficient enantioselective production of Tovorafenib and its intermediates. The process of the present disclosure is simple, economic, can be completed in less time as compared to prior art processes and therefore, can be adapted for production on an industrial scale, i.e., greater than 1 kilogram scale.

[0024] As used herein, in any of the disclosed processes or compounds, and unless indicated otherwise, a compound may be referred to herein as chemically pure or purified compound or as substantially free of any other compounds. As used herein, the terms "chemically pure" or "purified" or "substantially free of any other compounds" refer to a compound that is substantially free of any impurities including enantiomers of the subject compound, or other isomers. A chemically pure or purified compound or a compound that is substantially free of any other compound will be understood to mean that it contains about 10% (w / w) or less, about 5% (w / w) or less, about 4% (w / w) or less, about 3% (w / w) or less, about 2% (w / w) or less, about 1.5% (w / w) or less, about 1% (w / w) or less, about 0.8% (w / w) or less, about 0.6% (w / w) or less, about 0.4% (w / w) or less, about 0.2% (w / w) or less, about 0.1% (w / w) or less, or about 0% of any other compound as measured, for example, by HPLC. Alternatively, a chemicallypure or purified compound or a compound that is substantially free of any other compound will be understood to mean that it contains about 10% area percent or less, about 5% area percent or less, about 4% area percent or less, about 3% area percent or less, about 2% area percent or less, about 1.5% area percent or less, about 1% area percent or less, about 0.8% area percent or less, about 0.6% area percent or less, about 0.4% area percent or less, about 0.2% area percent or less, about 0.1% area percent or less, or about 0% of any other compound as measured by HPLC.

[0025] Thus, pure or purified Tovorafenib or any of the compounds described herein as substantially free of any compounds would be understood to contain greater than about 90% (w / w), greater than about 95% (w / w), greater than about 96% (w / w), greater than about 97% (w / w), greater than about 98% (w / w), greater than about 98.5% (w / w), greater than about 99% (w / w), greater than about 99.2% (w / w), greater than about 99.4% (w / w), greater than about 99.6% (w / w), greaterthan about 99.8% (w / w), greater than about 99.9% (w / w), or about 100% of the subject Tovorafenib or any of the compounds described. Alternatively, pure or purified Tovorafenib or any of the compounds described, described herein as substantially free of any compounds would be understood to contain greater than about 90% area percent, greater than about 95% area percent, greater than about 96% area percent, greater than about 97% area percent, greater than about 98% area percent, greater than about 98.5% area percent, greater than about 99% area percent, greater than about 99.2% area percent, greater than about 99.4% area percent, greater than about 99.6% area percent, greater than about 99.8% area percent, greater than about 99.9% area percent, or about 100% of the subject Tovorafenib.

[0026] As used herein, and unless indicated otherwise, the term "protecting group" refers to a grouping of atoms that when attached to a reactive functional group in a molecule masks, reduces or prevents reactivity of the functional group. Examples of protecting groups can be found in Greene and Wuts “Greene’s Protective Groups in Organic Synthesis”, 4th Edition, publ. Wiley, 2006 and Harrison et al., "Compendium of Synthetic Organic Methods", Vols. 1- 8 (John Wiley and Sons, 1971-1996).

[0027] Representative amine protecting groups include, but are not limited to, those where the amine group is converted to carbamate or amide such as Fmoc, benzyloxycarbonyl (cbz), benzyl, trityl, Boc, trifluoroacetyl derivative, phthalic anhydride, or succinic anhydride derivative and sulfonic acid derivatives such as tosylate.

[0028] A solid state form (or polymorph) may be referred to herein as polymorphically pure or as substantially free of any other solid state (or polymorphic) forms. As used herein in this context, the expression "substantially free of any other forms" will be understood to mean thatthe solid state form contains about 20% (w / w) or less, about 10% (w / w) or less, about 5% (w / w) or less, about 2% (w / w) or less, about 1% (w / w) or less, or about 0% of any other forms of the subject compound as measured, for example, by XRPD.

[0029] Depending on which other crystalline polymorphs a comparison is made, the crystalline polymorphs of Tovorafenib or ( / ?)-2-( l -aminocthyl)- '-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 7) of the present disclosure may have advantageous properties selected from at least one of the following: chemical purity, flowability, solubility, dissolution rate, morphology or crystal habit, stability, such as chemical stability as well as thermal and mechanical stability with respect to polymorphic conversion, stability towards dehydration and / or storage stability, low content of residual solvent, a lower degree of hygroscopicity, flowability, and advantageous processing and handling characteristics such as compressibility and bulk density.

[0030] A solid state form, such as a crystal form or an amorphous form, may be referred to herein as being characterized by graphical data “as depicted in” or “as substantially depicted in” a Figure. Such data include, for example, powder X-ray diffractograms and solid state NMR spectra. As is well-known in the art, the graphical data potentially provides additional technical information to further define the respective solid state form (a so-called “fingerprint”) which cannot necessarily be described by reference to numerical values or peak positions alone. In any event, the skilled person will understand that such graphical representations of data may be subject to small variations, e.g., in peak relative intensities and peak positions due to certain factors such as, but not limited to, variations in instrument response and variations in sample concentration and purity, which are well known to the skilled person. Nonetheless, the skilled person would readily be capable of comparing the graphical data in the Figures herein with graphical data generated for an unknown crystal form and confirm whether the two sets of graphical data are characterizing the same crystal form or two different crystal forms. A crystal form of Tovorafenib or ( / ?)-2-( l -aminocthyl)-A'-(5-chloro-4-(trifluoromcthyl)pyridin-2- yl)thiazole-5 -carboxamide (compound 7) referred to herein as being characterized by graphical data “as depicted in” or “as substantially depicted in” a Figure will thus be understood to include any crystal forms of Tovorafenib or ( / ?)-2-( l -aminocthyl)- '-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 7) characterized with the graphical data having such small variations, as are well known to the skilled person, in comparison with the Figure.

[0031] As used herein, and unless stated otherwise, the term “anhydrous” in relation to crystalline forms of Tovorafenib or ( / ?)-2-( l -aminocthyl)- '-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7), relates to a crystalline form of Tovorafenib or ( / ?)-2-( I -am inocthy I )- '-(5 -chloro-4-(trifl uoromcthy I (pyridineyl )thiazolc-5 -carboxamide (compound 7) which does not include any crystalline water (or other solvents) in a defined, stoichiometric amount within the crystal. Moreover, an “anhydrous” form would generally not contain more than 1% (w / w), of either water or organic solvents as measured for example by TGA.

[0032] As used herein, and unless indicated otherwise, the term "isolated" corresponds to compounds that are physically separated from the reaction mixture in which they are formed.

[0033] The processes or steps may be referred to herein as being carried out "overnight." This refers to time intervals, e.g., for the processes or steps, that span the time during the night, when the processes or steps may not be actively observed. The time intervals are from about 8 to about 20 hours, or about 10-18 hours, or about 16 hours.

[0034] The term "solvate," as used herein and unless indicated otherwise, refers to a crystal form that incorporates a solvent in the crystal structure. When the solvent is water, the solvate is often referred to as a "hydrate." The solvent in a solvate may be present in either a stoichiometric or in a non-stoichiometric amount.

[0035] The amount of solvent employed in a chemical process, e.g., a reaction or crystallization, may be referred to herein as a number of “volumes” or “vol” or “V.” For example, a material may be referred to as being suspended in 10 volumes (or 10 vol or 10V) of a solvent. In this context, this expression would be understood to mean milliliters of the solvent per gram of the material being suspended, such that suspending a 5 grams of a material in 10 volumes of a solvent means that the solvent is used in an amount of 10 milliliters of the solvent per gram of the material that is being suspended or, in this example, 50 mb of the solvent. In another context, the term "v / v" may be used to indicate the number of volumes of a solvent that are added to a liquid mixture based on the volume of that mixture. For example, adding solvent X (1.5 v / v) to a 100 ml reaction mixture would indicate that 150 mb of solvent X was added.

[0036] As used herein and unless indicated otherwise, the term "ambient conditions" refer to atmospheric pressure and a temperature of 22-24°C.

[0037] As used herein, unless stated otherwise, the XRPD measurements are taken using copper Ka radiation wavelength 1.5418 A. XRPD peaks reported herein are measured using CuKa radiation, X = 1.5418 A, typically at a temperature of 25 ± 3°C.

[0038] As used herein, unless stated otherwise,13C NMR reported herein are measured at 125 MHz at a magic angle spinning frequency ®r / 27t = 11 kHz, preferably at a temperature of at 293 K ± 3°C.

[0039] As used herein, and unless indicated otherwise, the term "reduced pressure" refers to a pressure of about 10 mbar to about 500 mbar, or about 50 mbar.

[0040] The amount of solvent employed in chemical processes, e.g., reactions or crystallizations, may be referred to herein as a number of "volumes" or "vol" or "V." For example, a material may be referred to as being suspended in 10 volumes (or 10 vol or 10V) of a solvent. In this context, this expression would be understood to mean milliliters of the solvent per gram of the material being suspended, such that suspending a 5 grams of a material in 10 volumes of a solvent means that the solvent is used in an amount of 10 milliliters of the solvent per gram of the material that is being suspended or, in this example, 50 mb of the solvent. In another context, the term "v / v" may be used to indicate the number of volumes of a solvent that are added to a liquid mixture based on the volume.

[0041] "Alkyl" refers to a monoradical of a branched or unbranched saturated hydrocarbon chain. Examples include, but are not limited to, methyl, ethyl, n-propyl, n-butyl, n-pentyl, n- hexyl, n-heptyl, n-octyl, n-nonyl, n-decyl, isopropyl, tert-butyl, sec-butyl, isobutyl, etc. Alkyl groups typically contain 1-10 carbon atoms, such as 1-6 carbon atoms or 1-4 carbon atoms, and can be substituted or unsubstituted.

[0042] "Alkyloxy" refers to a linear or branched, hydrocarbon group of formula -O-alkyl, in which the term "alkyl" is defined supra, and may include, e.g., a methoxy, ethoxy, n-propoxy, iso-propoxy, n- butoxy, iso-butoxy, tert-butoxy, sec-butoxy, pentoxy, iso-pentoxy, or n-hexoxy group, or an isomer thereof.

[0043] "Aryl" refers to phenyl and 7-15 membered monoradical bicyclic or tricyclic hydrocarbon ring systems, including bridged, spiro, and / or fused ring systems, in which at least one of the rings is aromatic. Aryl groups can be substituted or unsubstituted. Examples include, but are not limited to, naphthyl, indanyl, 1,2,3,4-tetrahydronaphthalenyl, 6, 7,8,9- tetrahydro-5H-benzocyclo-heptenyl, and 6,7,8,9-tetrahydro-5H-benzocycloheptenyl. An aryl group may contain 6 (i.e., phenyl) or 9 to 15 ring atoms, such as 6 (i.e., phenyl) or 9-11 ring atoms, e.g., 6 (i.e., phenyl), 9 or 10 ring atoms.

[0044] "Alkylaryl" refers to an alkyl group in which a hydrogen atom is replaced by an aryl group, wherein alkyl group and aryl group are defined supra. Alkylaryl groups can be substituted or unsubstituted. Examples include, but are not limited to, benzyl (C6H5CH2-).

[0045] ‘ ‘Alcohol protecting groups” refers to protecting groups which are introduced into a molecule by chemical modification of hydroxyl groups, for example to obtain chemoselectivity in a subsequent reaction. The term and use of protecting groups is well known in the art and for example described in Philipp J. Kociehski: Protecting Groups, 1. Auflage, Georg Thieme Verlag, Stuttgart 1994; Peter G.M. Wuts, Theodora W. Greene: Green's Protective Groups in Organic Synthesis, Fifth Ed. John Wiley & Sons Inc., Hoboken, New Jersey. Alcohol protecting groups can be for example acetoxy groups, benzoyl groups (Bz), benzyl groups (Bn), P-methoxyethoxymethyl ether (MEM), [bis-(4-methoxyphenyl)phenylmethyl] (DMT), methoxymethyl ether (MOM), [(4-methoxyphenyl)diphenylmethyl, (MMT), p-methoxybenzyl ether (PMB), methylthiomethyl ether, pivaloyl (Piv), tetrahydropyranyl (THP), triphenylmethyl (Tr), silyl ethers, trimethylsilyl ethers (TMS), triethylsilyl ethers (TES), tertbutyldimethylsilyl ethers (TBDMS), tri-iso-propylsilyloxymethyl ethers (TOM), triisopropylsilyl (TIPS) ethers, methyl ethers and ethoxyethyl ethers (EE).

[0046] The term "amino" refers to the radical -NH2.

[0047] The term “halogen” refers to chloro, bromo or iodo.

[0048] The present disclosure provides for novel process for the synthesis of Tovorafenib.

[0049] The process of the present disclosure can be illustrated by the following Scheme 3:Scheme 3:wherein Ri and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time. Thus, in the disclosed process, the amine (II) is a chiral amine (denoted by the asterisks in the chemical formulae herein) and the resulting intermediates (III), (IV) and (7) are enantiomerically enriched, to produce Tovorafenib or a salt thereof. Without being bound by theory, the use of chiral amine (II) induces enantioselectivity at the imine carbon of compound (III) and the subsequentintermediate compound (IV), which can lead to a more enantioselective reduction reaction to form compound 7, thereby increasing the enantioselectivity for the desired isomer of compound 7.

[0050] In one aspect, the disclosure provides a process for the preparation of Tovorafenib or a salt thereof, comprising reacting 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5 -carboxamide of formula (4):with a transaminase enzyme in the presence of an amine donor, to obtain compound of formula (7) or salt thereof:; and converting the compound of formula (7) to Tovorafenib or a salt thereof. The conversion of the compound of formula (7) to Tovorafenib or a salt thereof may be carried out as described in any aspect or embodiment as described herein.

[0051] In another aspect, the present disclosure provides a process for preparation of Tovorafenib or a salt thereof, comprising: a) reacting 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide of formula (4):to obtain compound of formula (III):wherein Riand R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time; b) reducing the compound of formula (III)to compound of formula (IV) or salt thereofwherein Ri and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time. c) deprotecting a compound of formula (IV) or salt thereofto compound of formula (7) or salt thereofd) optionally purifying the compound of formula (7) or salt thereof ; and e) reacting the compound of formula (7) or salt thereofwith compound of formula (8)to get Tovorafenib or a salt thereof.

[0052] In another aspect, the disclosure relates to compound of formula (III)wherein Ri and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time.

[0053] According to any aspect or embodiment of the disclosure, Ri is preferably: phenyl; substituted phenyl (preferably selected from the group consisting of methylbenzoyl, 4- nitrophenyl, 4-methoxyphenyl, p-tolyl, 5-isopropyl-3-methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4-methoxynaphthyl); alkyl (preferably Ci-Ce alkyl,more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Cl to C2), or methoxyalkyl (Cl to C2)]; cycloalkyl (preferably C5 to Cs cycloalkyl, and particularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of C5 to Cs branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl). According to any aspect or embodiment, R2 is preferably selected from the group consisting of: phenyl; substituted phenyl (preferably methylbenzoyl, 4-nitrophenyl, 4-methoxyphenyl, p-tolyl, 5-isopropyl-3-methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4-methoxynaphthyl); alkyl (preferably Ci-Ce alkyl, more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Ci to C2), or methoxyalkyl (Ci to C2)]; cycloalkyl (preferably Cs to Cs cycloalkyl, and particularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of Cs to Cs branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl).

[0054] Particularly, according to any aspect or embodiment of the disclosure, Ri and R2 are selected from unsubstituted alkyl (preferably Ci-Ce alkyl), unsubstituted cycloalkyl (preferably Cs-ClO alkyl, more preferably Cs to Ce cycloalkyl), unsubstituted aryl (preferably Ce-Cioaryl); and more preferably Ri and R2 are selected from Ci to C3 alkyl (preferably methyl), unsubstituted phenyl; or unsubstituted naphthyl.

[0055] More particularly, according to any aspect or embodiment of the disclosure, Ri and R2 may be selected from unsubstituted Ci-Ce alkyl or unsubstituted Ce-Cio aryl; preferably wherein Ri and R2 are selected from unsubstituted C1-C3 alkyl, or unsubstituted Ce-Cs aryl; especially wherein Ri and R2 are selected from methyl and phenyl. Most preferably, the amine (II) is R-phenylethylamine.

[0056] In a preferred embodiment, step (a) comprises reacting the compound of formula (4) or a salt thereof:with R-phenylethylamine of formula (9):to obtain compound of formula (5) or a salt thereof:

[0057] According to any aspect or embodiment of the process, step (a) may be carried out in an organic solvent, preferably dichloromethane, dimethoxy ethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF, or mixtures thereof, and preferably wherein the organic solvent is a Ci to C4 alcohol, preferably methanol, ethanol or isopropanol, and particularly ethanol.

[0058] According to any aspect or embodiment of the process, step (a) may be carried out in the presence of an inorganic acid or Lewis acid (particularly hydrochloric acid, BF3, TiCh. AlCh, titanium isopropoxide, SnCh. ZnCh, or ZnBn), or an organic acid; particularly an organic acid (preferably a C2-C6 carboxylic acid, particularly a C2 to C4 carboxylic acid, and more preferably propanoic acid or acetic acid, and particularly acetic acid.

[0059] According to any aspect or embodiment of the process, step (a) may be carried out in at a temperature of: about 50°C to about 100°C, about 55 °C to about 95 °C, about 60°C to about 90°C, or about 70°C to about 85°C.

[0060] According to any aspect or embodiment of the process, step (a) may comprise isolating the compound of formula (III), preferably by cooling and precipitation from the reaction mixture, and isolating the solid, preferably by filtration.

[0061] According to any aspect or embodiment of the process, step (b) may comprise reducing the compound of formula (5):to obtain the compound of formula (6):

[0062] According to any aspect or embodiment of the process, the reducing agent in step (b) is selected from: hydrogen gas, a hydride reducing agents (preferably sodium borohydride, lithium borohydride, Alane (aluminium hydride), DiBAL-H, sodium triacetoxyborohydride, Li AIH4. sodium cyanoborohydride, borane, triethylsilane, trichlorosilane, Hantzsch ester, CBS reagent or hydride, reducing agent derived from a reaction between sodium borohydride and a chiral acid (preferably L-tartaric acid, L-proline, or S-mandelic acid etc); and preferably wherein the reducing agent is sodium borohydride.

[0063] According to any aspect or embodiment of the process, step (b) may be carried out in the presence of one or more solvents, preferably methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF, water, or mixtures thereof; more preferably wherein the solvent is selected from: methanol, ethanol, isopropyl alcohol or tetrahydrofuran.

[0064] According to any aspect or embodiment of the process, step (b) may be carried out in the presence of an inorganic acid or Lewis acid (particularly hydrochloric acid, BF3, TiCh. AlCh, titanium isopropoxide, SnCh. ZnCh, or ZnBn), or an organic acid; particularly an organic acid( preferably a C2-C6 carboxylic acid, particularly a C2 to C4 carboxylic acid, and more preferably propanoic acid or acetic acid, and particularly acetic acid.

[0065] A process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, or 27, wherein step c) comprises deprotecting the compound of formula (6):to obtain a compound of formula (7):

[0066] According to any aspect or embodiment of the process, step (c) may comprise reacting the compound of formula (IV) with an oxidizing agent, preferably in the presence of a base and a solvent. The oxidizing agent is preferably selected from: bromine gas, chlorine gas, N- chlorosuccinimide, N-bromosuccinimide, N-bromophthalimide, N-chlorophthalimide, trichloroisocyanuric acid, chlorohydantoin, borohydantoin, 2,3,4,5,6,6-hexachlorocyclohexa- 2,4-dien-l-one, or 4,4-dibromo-2,6-di-tert-butyl-cyclohexa-2,5-dienone; and preferably wherein the oxidising agent is N-bromosuccinimide. The base may be any suitable base, and is preferably an organic base, preferably selected from: triethylamine, diisopropyl ethylamine, pyridine, diethylamine, diisopropylamine, N,N-dimethylaminopyridine, 1,8- diazabicyclo[5.4.0]undec-7-ene, l,4-diazabicyclo[2.2.2]octane, piperazine, pyrrolidine, piperidine, morpholine, N-methyhnorpholine, N-methylpyrrolidine, N-methyl piperidine, or N,N-dimethylpiperazine; and more preferably wherein the base is triethylamine. The solvent is preferably selected from: methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF, water or mixture thereof; and more preferably wherein the solvent is acetonitrile, water or mixture thereof

[0067] According to any aspect or embodiment of the process, step (d)comprises purification of compound (7), preferably by fractional recrystallization or a diastereomer salt separation,preferably wherein the purification of compound (7) comprises crystallization of the diastereomer salt with an optically active di-p-toluoyl-tartaric acid or an optically active mandelic acid, preferably di-p-toluoyl-(D)-tartaric acid or (S)-mandelic acid, and more preferably (S)-mandelic acid.

[0068] According to any aspect or embodiment of the process, step e) comprises reacting the compound of formula (7):with the compound of formula (8):

[0069] According to any aspect or embodiment of the process, step (e) comprises reacting compound (7) with compound (8) in the presence of a coupling agent, a coupling additive and a solvent. Preferably, the coupling agent is 1 -hydroxybenzotriazole. Preferably, the coupling additive is 1 -(3 -dimethylaminopropyl)-3 -ethylcarbodiimide. The solvent may be any suitable solvent, and is preferably dimethylacetamide.

[0070] Also provided is a process according to any aspect or embodiment of the disclosure herein, further comprising combining the Tovorafenib or a pharmaceutically acceptable salt thereof, with at least one pharmaceutically acceptable excipient to form a pharmaceutical composition.

[0071] The disclosure further relates to a compound of formula (III):or a salt thereof, wherein Ri and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time. According to any aspect or embodiment, R1and R2are different and each is independently selected from: alkyl (preferably Ci-Ce alkyl), cycloalkyl (preferably C5-C10 alkyl, more preferably C5 to Ce cycloalkyl), aryl (preferably Ce-Cio aryl); and more preferably Ri and R2 are selected from Ci to C3 alkyl (preferably methyl), phenyl; or naphthyl; one of Ri or R2 is methyl, and the other is phenyl.

[0072] In another aspect, the disclosure relates to the use of the compound (III) in the preparation of Tovorafenib thereof.

[0073] In another aspect, the disclosure relates to compound of formula (IV)wherein Ri and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and Ri and R2 cannot be same at the same time. According to any aspect or embodiment, R1and R2are different and each is independently selected from: alkyl(preferably Ci-Ce alkyl), cycloalkyl (preferably C5-C10 alkyl, more preferably C5 to Ce cycloalkyl), aryl (preferably Ce-Cio aryl); and more preferably Ri and R2 are selected from Ci to C3 alkyl (preferably methyl), phenyl; or naphthyl; one of Ri or R2 is methyl, and the other is phenyl.

[0074] In another aspect, the disclosure relates to the use of the compound (IV) in the preparation of Tovorafenib thereof.

[0075] In one embodiment Ri is unsubstituted or substituted phenyl. In one embodiment Ri is a substituted phenyl selected from the group consisting of methyl benzoyl, 4-nitrophenyl, 4- methoxy phenyl, p-tolyl, 5 -isopropyl-3 -methylphenyl. In another embodiment, Ri is an unsubstituted phenyl, i.e. phenyl. In another embodiment Ri is unsubstituted and substituted naphthyl. In one embodiment Ri is substituted naphthyl selected from the group consisting of 4-nitro naphthyl, 4-methoxy naphthyl. In another embodiment, Ri is an unsubstituted naphthyl, i.e. naphthyl. In another embodiment Ri is unsubstituted or substituted alkyl. In one embodiment substituted alkyl is selected from the group consisting of Cl to C7 branched alkyl group, hydroxyl alkyl (Cl to C2), methoxy alkyl (Cl to C2). In another embodiment, Ri is an unsubstituted alkyl, i.e. methyl, ethyl, propyl or butyl. In another embodiment Ri is unsubstituted or substituted cycloalkyl. In one embodiment substituted cylcoalkyl is selected from the group consisting of, C5 to C8 branched cycloalkyl group, hydroxyl cycloalkyl, methoxy cycloalkyl. In another embodiment, Ri is an unsubstituted cycloalkyl, i.e. cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. In one embodiment R2 is unsubstituted or substituted phenyl. In one embodiment R2 is substituted phenyl selected from the group consisting of methyl benzoyl, 4-nitrophenyl, 4-methoxy phenyl, p-tolyl, 5-isopropyl-3- methylphenyl. In another embodiment, R2 is an unsubstituted phenyl, i.e. phenyl. In another embodiment R2 is substituted or substituted naphthyl. In one embodiment R2 is a substituted naphthyl selected from the group consisting of, 4-Nitro naphthyl, 4-methoxy naphthyl. In another embodiment, R2 is an unsubstituted naphthyl, i.e. naphthyl. In another embodiment R2 is unsubstituted or substituted alkyl. In one embodiment R2 is a substituted alkyl selected from the group consisting of Cl to C7 branched alkyl group, hydroxyl alkyl (Cl to C2), methoxy alkyl (C 1 to C2) . In another embodiment, R2 is an unsubstituted alkyl, i.e. methyl, ethyl, propyl or butyl. In another embodiment R2 is unsubstituted or substituted cycloalkyl. In one embodiment substituted cylcoalkyl is selected from the group consisting of, C5 to C8 branched cycloalkyl group, hydroxyl cycloalkyl, methoxy cycloalkyl. In another embodiment, R2 is an unsubstituted cycloalkyl, i.e. cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl. Ri and R2 cannot be same at the same time.

[0076] Step a) is typically carried out in the presence of one or more solvents, and a suitable catalyst. Suitable solvents may include, for example, methanol, ethanol, isopropanol, n- propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF or mixture thereof. In embodiments, the solvent is ethanol. Suitable catalyst may include, for example, Bronsted acids like hydrochloric acid, acetic acid, propanoic acid or Lewis acids like BF3, TiCh, AlCh, titanium isopropoxide, SnCE. ZnCh, ZnBn. In embodiments, the catalyst is acetic acid.

[0077] In embodiments, in step a) the compound of formula (4) is reacted with R- phenylethylamine of formula (9) to obtain compound of formula (5) , thereof, as illustrated by(9)

[0078] Step b) is typically carried out in the presence of one or more solvents, additives, and suitable reducing agent. Suitable solvents may include, for example, methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2- methyl THF, water, or mixture thereof. In embodiments, the solvent is methanol, ethanol, isopropyl alcohol or tetrahydrofuran. Additive may include, for example, Bronsted acids like hydrochloric acid, acetic acid, propanoic acid or Lewis acids like BF3, TiCI-i. AlCh, titanium isopropoxide, SnCh. ZnCh, and ZnBn. In embodiments, the additive is acetic acid. Suitable reducing agent may include, for example hydrogen gas, hydride reducing agents like sodium borohydride, lithium borohydride, Alane, DiBAL-H, sodium triacetoxyborohydride, or LiAlH- 4, sodium cyanoborohydride, borane, triethyl silane, trichloro silane, Hantzsch ester, CBS reagent or hydride, reducing agent derived from a reaction between sodium borohydride and a chiral acid like L-tartaric acid, L-proline, S-mandelic acid etc. In embodiments, the reducing agent is sodium borohydride.

[0079] In embodiments, in step b) the compound of formula (5) is reduced to get the compound of formula (6), as illustrated by the following Scheme 5 (exemplified with Ri = Ph and R2=methyl).

[0080] Step c) is typically carried out in the presence of one or more solvents, oxidizing agent, and suitable base. Suitable solvents may include, for example, methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF, water or mixture thereof. In embodiments, the solvent is acetonitrile, water or mixture thereof. Oxidising agent may include, for example, bromine gas, chlorine gas, N-chlorosuccinimide, N-bromosuccinimide, N-bromophthalimide, N-chlorophthalimide, trichloroisocyanuric acid, chlorohydantoin, borohydantoin, 2,3,4,5,6,6-hexachlorocyclohexa-2,4-dien-l-one, or 4,4- dibromo-2,6-di-tert-butyl-cyclohexa-2,5-dienone. In embodiments, Oxidising agent is N- bromosuccinimide. Suitable base may include, triethylamine, diisopropyl ethylamine, pyridine, diethylamine, diisopropylamine, N,N-dimethylaminopyridine, 1,8- diazabicyclo[5.4.0]undec-7-ene, ,4-diazabicyclo[2.2.2]octane, piperazine, pyrrolidine, piperidine, morpholine, N-methyhnorpholine, N-methylpyrrolidine, N-methyl piperidine, or N,N-dimethylpiperazine; or l,4-diazabicyclo[2.2.2]octane. In embodiments, the base is triethylamine.

[0081] In embodiments, in step c) the compound of formula (6) is deprotected to get the compound of formula (7) , as illustrated by the following Scheme 6 (exemplified with Ri = Ph and R2 = methyl).

[0082] In embodiments, in step d), in order to obtain the above-mentioned compound (7) or a salt thereof with high optical purity, it is preferably purified by a fractional recrystallization method or a diastereomer salt method, preferably crystallization of the diastereomer salt with optically active di-p-toluoyl-tartaric acid (preferably di-p-toluoyl-(D)-tartaric acid when the above-mentioned compound (7) is (R)-form) or optically active mandelic acid (preferably (S)-mandelic acid when the above-mentioned compound (7) is (R)-form), particularly preferably crystallization of the diastereomer salt with optically active mandelic acid (preferably (S)- mandelic acid when the above-mentioned compound (7) is (R)-form)

[0083] In embodiments, in step e) the compound of formula (7) is reacted with compound of formula (8) to get Tovorafenib, thereof, as illustrated by the following Scheme 7

[0084] In one embodiment, the obtained Tovorafenib is optionally converted to its salt.

[0085] Tovorafenib or salts thereof can be prepared starting from compound of formula (4) in situ without isolating the intermediates.

[0086] In another aspect, the disclosure relates to compound of formula (5)

[0087] In another aspect, the disclosure relates to the use of the compound (5) in the preparation of Tovorafenib thereof.

[0088] In another aspect, the disclosure relates to compound of formula (6)

[0089] In another aspect, the disclosure relates to the use of the compound (6) in the preparation of Tovorafenib thereof.

[0090] In one aspect, the present disclosure provides a process for preparation of Tovorafenib comprising a) reacting 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide of formula (4) with R-selective transaminase enzyme:to obtain compound of formula (7) or salt thereof; andwith compound of formula (8)to get Tovorafenib.

[0091] In another embodiment the above process further comprises the generation of a compound of formula (7) by a process that comprises treating compound of formula (4) with R-selective transaminase enzyme in presence of an amine donor.

[0092] The term "transaminase" also referred as "Amine Transaminase (ATA)" or more specifically as “co-transaminase” is used to refer to a polypeptide having an enzymatic capability of transferring an amino group (NH2) to a carbonyl group (C=O) of an acceptor molecule.

[0093] The term "amine donor" is used to refer to any amino acid or amine that will react with a transaminase and a ketone, to produce desired amine product and a ketone by product.

[0094] The transamination reaction may be carried out in presence of "pyridoxal phosphate (PLP)" that acts as a coenzyme. In transamination reactions using transaminase enzymes, the amine group of the amine donor is transferred to the coenzyme to produce a ketone as a byproduct, while pyridoxal-5 '-phosphate is converted to pyridoxamine phosphate. The transfer of the amine group from pyridoxamine phosphate to the ketone substrate produces a chiral amine and regenerates the coenzyme.

[0095] Typical amine donors that can be used with the invention include chiral and achiral amino acids, and chiral and achiral amines. Amine donors that can be used with the invention include, by way of example and not limitation, alanine, glycine, glutamate or beta-alanine as amino acids or isopropylamine, sec-butylamine, butane- 1,4-diamine, pentane- 1,5-diamine, but-2-enediamine as aliphatic amines, phenethylamine, 1 -phenylethylamine, benzylamine, o- xylylenediamine as aromatic amines and their derivatives substituted on the aryl moiety, including both (R) and (S) single isomers where possible and including all possible salts of the amines. More preferably, the amine donor is isopropylamine, sec-butylamine or alanine, thereby obtaining acetone, butanone or pyruvate as by-product.

[0096] Thus, according to any aspect or embodiment of the processes disclosed herein, the reaction of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide of formula (4) with an R-selective transaminase enzyme, may be carried out in the presence of an amine donor, wherein the amine donor is an amino acid (preferably selected from: alanine, glycine, glutamate or beta-alanine); or an amine or salt thereof, preferably an aliphatic amine (particularly a C3 to Cs alkyl monoamine or a C3 to Cs alkyl diamine), more particularly selected from: isopropylamine, sec-butylamine, butane- 1,4-diamine, pentane-l,5-diamine, or but-2- enediamine; or an aromatic amine (particularly a C5 to Cs aromatic amine), particularly phenethylamine, 1 -phenylethylamine, benzylamine, or o-xylylenediamine; or mixtures thereof. Preferably, the amine donor is selected from isopropylamine, sec-butylamine or D-alanine., or a mixture thereof, particularly sec-butylamine, or a mixture of isopropylamine and D-alanine.

[0097] Transamination of the substrates is carried out in a bioreactor using an aliquot of the enzyme with the substrate typically at a defined concentration. The reaction parameters such as pH = 7-11.5, temperature = 30-55 °C, and reaction time = 12 - 168 hours are maintained at levels that favor optimal biocatalytic activity and stability.

[0098] Suitable transaminase enzymes for this reaction include co-transaminase, preferably an R-transaminase, particularly T-185 (SEQ ID 1, FIG. 5) and T-186 (SEQ ID 2, FIG. 5), as well as transaminase (particularly co-transaminase, and preferably R-transaminase) enzymes having an amino acid sequence that comprises: at least 80%, at least 85%, 5 at least 90%, atleast 92%, at least 95%, at least 96%, at least 97%, at least 98%, and at least 99% identity, to SEQ ID 1 or SEQ ID 2 (FIG. 5). The co-transaminase enzyme T-185 originates from Shinella sp. (wild type) (see ChemBioChem 2021, 22, 1232-1242). Particularly, the transaminase enzyme (particularly co-transaminase, and preferably R-transaminase) may have an amino acid sequence corresponding to SEQ ID 2 (T-186). The co-transaminase enzyme T-186 is engineered from an co-transaminase originating from Exophiala xenobiotica (variant T273S) (see ACS Catal. 2019, 9, 1140-1148), or can be from a cell lysate containing co-transaminase, and preferably R-transaminase particularly T-185 (SEQ ID 1, FIG. 5) and T-186 (SEQ ID 2, FIG. 5), which can be prepared as per Example 16.

[0099] According to any aspect or embodiment of the processes disclosed herein, the transaminase enzyme may have an amino acid sequence according to SEQ ID 1 or SEQ ID 2 which comprises: one or more, five or more, ten or more, 15 or more, 20 or more, 25 or more, 30 or more, 35 or more, or 40, mutations at a position selected from 14, 21, 22, 25, 35, 38, 40, 43, 50, 69, 71, 73, 76, 81, 102, 108, 122, 137, 146, 150, 156, 191, 195, 197, 198, 200, 215, 224, 225, 251, 255, 262, 263, 265, 267, 269, 272, 273, 281, and 284.

[0100] Particularly, the transaminase enzyme (particularly co-transaminase, and preferably R- transaminase) may have an amino acid sequence corresponding to SEQ ID 1 (T-185). The co- transaminase enzyme T-185 is originated from Shinella sp. (wild type) (see ChemBioChem 2021, 22, 1232 1242). Particularly, the transaminase enzyme (particularly co-transaminase, and preferably R-transaminase) may have an amino acid sequence corresponding to SEQ ID 2 (T- 186). The co- transaminase enzyme T-186 is engineered from an co-transaminase originated from Exophiala xenobiotica (variant T273S) (see ACS Catal. 2019, 9, 1140-1148).

[0101] According to any aspect or embodiment of the present disclosure, the reaction of compound (4) with a transaminase enzyme in the presence of an amine donor is preferably carried out at a pH of: about 7 to about 11.5, or about 6.8 to about 11.5, about 6.8 to about 10.5, about 6.9 to about 9.5, about 7.0 to about 8.5, about 7. 1 to about 8.2, about 7.2 to about 8.0, or about 7.3 to about 7.8, about 7.4 to about 7.6 or about 7.5. According to any aspect or embodiment of the present disclosure, the pH of the reaction mixture is maintained using a phosphate buffer.

[0102] According to any aspect or embodiment of the present disclosure, the reaction of compound (4) with a transaminase enzyme in the presence of an amine donor is preferably carried out at a temperature of: about 30°C to about 55°C, about 32°C to about 50°C, about 35°C to about 45°C, about 38°C to about 42°C, or about 40°C.

[0103] According to any aspect or embodiment of the present disclosure, the reaction of compound (4) with a transaminase enzyme in the presence of an amine donor is preferably carried out in an aqueous mixture, optionally in the presence of a co-solvent, preferably wherein the co-solvent is selected from DMSO, acetonitrile, dimethylformamide, THF. DMSO is a particularly preferred co-solvent.

[0104] The present disclosure also relates to the compound of formula (7) which is obtained by enzymatic process is more than 85% enantiomerically pure, more than 90% enantiomerically pure, more than 98% enantiomerically pure or 100% enantiomerically pure.

[0105] In one aspect, the present disclosure provides a process for preparation of Tovorafenib comprising reacting 2-acetylthiazole-5 -carboxylic acid of formula (10) with R-selective transaminase enzyme:to obtain compound of formula (11) or salt thereof

[0106] The present disclosure further provides a process for the preparation of Tovorafenib comprising reacting the compound of formula (10):with R-selective transaminase enzyme, to obtain compound of formula (11) or a salt thereof:

[0107] and converting the compound of formula (11) to Tovorafenib. The compound of formula (11) may be converted to Tovorafenib by a process analogous to Scheme 2 above. In another embodiment the above process further comprises the generation of a compound of formula (11) by a process that comprises treating compound of formula (10) with R-selective transaminase enzyme in presence of an amine donor.

[0108] The term "Transaminase" also referred as "Amine Transaminase (ATA)" or more specifically “co-transaminase” is used to refer to a polypeptide having an enzymatic capability of transferring an amino group (NH2) to a carbonyl group (C=O) of an acceptor molecule.

[0109] The term "amine donor" is used to refer to any amino acid or amine that will react with a transaminase and a ketone, to produce desired amine product and a ketone by product.

[0110] The transamination reaction may be carried out in presence of "pyridoxal phosphate (PLP)" that acts as a coenzyme. In transamination reactions using transaminase enzymes, the amine group of the amine donor is transferred to the coenzyme to produce a ketone as a byproduct, while pyridoxal-5 '-phosphate is converted to pyridoxamine phosphate. The transfer of the amine group from pyridoxamine phosphate to the ketone substrate produces a chiral amine and regenerates the coenzyme.

[0111] Typical amine donors that can be used with the invention include chiral and achiral amino acids, and chiral and achiral amines. Amino donors that can be used with the invention include, by way of example and not limitation, alanine, glycine, glutamate or beta-alanine as amino acids or isopropylamine, sec-butylamine, butane- 1,4-diamine, pentane- 1,5-diamine, but-2-enediamine as aliphatic amines, phenethylamine, 1 -phenylethylamine, benzylamine, o- xylylenediamine as aromatic amines and their derivatives substituted on the aryl moiety, including both (R) and (S) single isomers where possible and including all possible salts of the amines. More preferably, the amine donor is isopropylamine, sec-butylamine or alanine, thereby obtaining acetone, butanone or pyruvate as by-product.

[0112] Transamination of the substrates is carried out in a bioreactor using an aliquot of the enzyme with the substrate typically at a defined concentration. The reaction parameters such as pH = 7 - 11.5, temperature = 30 - 55 °C, and reaction time = 12 - 168 hours are maintained at levels that favor optimal biocatalytic activity and stability.

[0113] The present disclosure also relates to the compound of formula (11) which is obtained by enzymatic process is more than 85% enantiomerically pure, more than 90% enantiomerically pure, more than 98% enantiomerically pure or 100% enantiomerically pure.

[0114] In embodiments, compound of formula (11) may be used to prepare Tovorafenib.

[0115] In one embodiment, the obtained Tovorafenib is optionally converted to its salt.

[0116] The present disclosure includes a crystalline polymorph of (R)-2-(l-aminoethyl)-N- (5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7), designated Form 1. The crystalline Form 1 of (R)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 7) may be characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 1; an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 17.6 and 24.3 degrees 2-theta ± 0.2 degrees 2-theta; and combinations of these data.

[0117] Crystalline Form 1 of (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5 -carboxamide (compound 7) may be further characterized by an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 17.6 and 24.3 degrees 2-theta ± 0.2 degrees 2- theta, and also having any one, two, three, or four additional peaks selected from 11.7, 16.3, 21.3 and 23.4 degrees 2-theta ± 0.2 degrees 2-theta. Alternatively, crystalline Form 1 of (R)-2- (l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7) may be characterized by an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 11.7, 16.3, 17.6, 21.3, 23.4, and 24.3 degrees 2-theta ± 0.2 degrees 2-theta.

[0118] In one embodiment of the present disclosure, crystalline Form 1 of (R)-2-(l- aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide is isolated. Thus, crystalline Form 1 of compound (7) as described in any aspect or embodiment of the present disclosure may be isolated.

[0119] Crystalline Form 1 of (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5 -carboxamide may be characterized by each of the above characteristics alone / or by all possible combinations, e.g., an XRPD pattern having peaks at 8.1, 10.6, 17.6 and 24.3 degrees 2-theta ± 0.2 degrees 2-theta; an XRPD pattern as depicted in FIG.l, and combinations thereof.

[0120] In another aspect, the disclosure relates to the use of the crystalline Form 1 of (R)-2- (l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7) in the preparation of Tovorafenib.

[0121] The present disclosure includes a crystalline polymorph of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4), designated Form A. The crystalline Form A of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (compound 4) may be characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 3; an X-ray powder diffraction pattern having peaks at 17.7, 20. 1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 degrees 2-theta; and combinations of these data.

[0122] Crystalline Form A of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole- 5 -carboxamide (compound 4) may be further characterized by an X-ray powder diffraction pattern having peaks at 17.7, 20.1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 degrees 2-theta, and also having any one, two, three or four additional peaks selected from at 12.4, 13.3, 14.5 and 17.3 degrees 2-theta.

[0123] Crystalline Form A of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole- 5 -carboxamide (compound 4) may be characterized by an X-ray powder diffraction pattern having peaks at 17.7, 20.1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 degrees 2-theta, and also having any one, two, three or four additional peaks selected from at 12.4, 13.3, 14.5 and 17.3 degrees 2-theta ± 0.2 degrees 2-theta. Crystalline Form A of compound 4 may be characterized by an X-ray powder diffraction pattern having peaks at 12.4, 13.3, 14.5, 17.3 17.7, 20.1, 22.1, 26.3, and 27.6 degrees 2-theta ± 0.2 degrees 2-theta.

[0124] In one embodiment of the present disclosure, crystalline Form A of (2-acetyl-N-(5- chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4) is isolated. Thus, crystalline Form A of compound 4 as described in any aspect or embodiment of the present disclosure may be isolated.

[0125] Crystalline Form A of (2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole -5 -carboxamide (compound 4) may be characterized by each of the above characteristics alone / or by all possible combinations, e.g., an XRPD pattern having peaks at 17.7, 20.1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 degrees 2-theta; an XRPD pattern as depicted in FIG. 3, and combinations thereof.

[0126] In another aspect, the disclosure relates to the use of the crystalline Form A of (2- acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4) in the preparation of Tovorafenib thereof.

[0127] The present disclosure includes a crystalline polymorph of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 4), designated Form B. The crystalline Form B of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (compound 4) may be characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 4; an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 18.6, 21.0, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta; and combinations of these data.

[0128] Crystalline Form B of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole- 5 -carboxamide (compound 4) may be further characterized by an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 18.6, 21.0, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta,and also having any one, two, or three additional peaks selected from peaks at 17.4, 22.1, and 22.7 degrees 2-theta ± 0.2 degrees 2-theta.

[0129] Crystalline Form B of compound 4 may be alternatively characterized by an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 17.4, 18.6, 21.0, 22.1, 22.7, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta.

[0130] In one embodiment of the present disclosure, crystalline Form B of (2-acetyl-N-(5- chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4) is isolated. Thus, crystalline Form B of compound (4) as described in any aspect or embodiment of the disclosure may be isolated.

[0131] Crystalline Form B of (2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole- 5 -carboxamide (compound 4) may be characterized by each of the above characteristics alone / or by all possible combinations, e.g., an XRPD pattern having peaks at 11.4, 15.2, 18.6, 21.0, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta; an XRPD pattern as depicted in FIG. 4, and combinations thereof.

[0132] In another aspect, the disclosure relates to the use of the crystalline Form B of (2- acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4) in the preparation of Tovorafenib thereof.

[0133] An overview about processes for preparation of Tovorafenib is shown in Scheme 8 below.

[0134] In another aspect the present disclosure provides Tovorafenib or salts thereof, produced by the process of the present disclosure.

[0135] In another aspect the present disclosure provides Tovorafenib produced by the processes of the present disclosure may be used to prepare salts of Tovorafenib. Such prepared Tovorafenib and Tovorafenib salts may be used to prepare pharmaceutical formulations and / ormedicaments, particularly for the treatment of Glioma, Cholangiocarcinoma; Histiocytosis; solid tumors, and Malignant melanoma.

[0136] Having described the disclosure with reference to certain preferred embodiments, other embodiments will become apparent to one skilled in the art from consideration of the specification. The disclosure is further illustrated by reference to the following examples describing in detail the preparation of the composition and methods of use of the disclosure. It will be apparent to those skilled in the art that many modifications, both to materials and methods, may be practiced without departing from the scope of the disclosure.

[0137] Powder X-ray Diffraction (XRD) method for Compound 4The sample was measured with powder X-ray diffraction technique.

[0138] Measurement was performed on a BRUKER D8 Advance X-ray powder diffractometer, CuKa radiation (X = 1.5406 A); Lynxeye XE detector, low amount PMMA sample holder with zero background plate was used. Prior to analysis, the dry sample was gently ground by means of mortar and pestle in order to obtain a fine powder. The ground sample was adjusted into a cavity of the sample holder and the surface of the sample was smoothed by means of a glass object-slide.Measurement parameters:Sample: Spin mode, rotation speed 30 rpm;Scan range: 2 - 40 degrees 2-theta;Scan mode: continuous;Step size: 0.05 ± 0.005 degrees;Time per step: 0.5 sec;Divergence slit: V20Powder X-ray Diffraction (XRD) method for Compound 7

[0139] The sample was measured with powder X-ray diffraction technique.

[0140] Measurement was performed on a BRUKER D8 Advance X-ray powder diffractometer, CuKa radiation (X = 1.5406 A); Lynxeye XE detector, low amount PMMA sample holder with zero background plate was used. Prior to analysis, the dry sample was gently ground by means of mortar and pestle in order to obtain a fine powder. The ground sample was adjusted into a cavity of the sample holder and the surface of the sample was smoothed by means of a glass object-slide.Measurement parameters:Sample: Spin mode, rotation speed 30 rpm;Scan range: 2 - 40 degrees 2-theta;Scan mode: continuous;Step size: 0.05 ± 0.005 degrees;Time per step: 0.5 sec;Divergence slit: V20Powder X-ray Powder Diffraction (“XRPD”) method for Tovorafenib

[0141] X-ray powder diffraction was performed on X-Ray powder diffractometer:Bruker D8 Advance; Copper Ka radiation (X = 1.5418 A); Lynx eye detector; laboratory temperature 22-25 °C; PMMA specimen holder ring. Prior to analysis, the samples were gently ground by means of mortar and pestle in order to obtain a fine powder. The ground sample was adjusted into a cavity of the sample holder and the surface of the sample was smoothed by means of a cover glass.Measurement parameters:Scan range: 2 - 40 degrees 2-theta;Scan mode: continuous;Step size: 0.05 degrees;Time per step: 0.5 s;Sample spin: 30 rpm;EXAMPLES

[0142] 2-acetylthiazole-5-carboxylic acid can be prepared according to methods known from the literature, for example CN117567388; 5-chloro-4-(trifluoromethyl) pyridin-2 -amine according to methods known from the literature, for example WO2020263187; 6-amino-5- chloropyrimidine-4-carboxylic acid can be prepared according to methods known from the literature, for example WO2006121648; 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5 -carboxamide can be prepared according to methods known from the literature, for example W02009006389 or Example 1 or Example 12 provided below. The entire contents of the foregoing publications are incorporated by reference herein. The entire contents of the foregoing publications is incorporated by reference herein.

[0143] Transaminase enzyme can be purchased, for Example, Prozomix PRO-TRANS No. 385, or Prozomix PRO-TRANS No. 436 can be purchased from Prozomix, a biocatalysis company based in the United Kingdom.

[0144] Suitable transaminase enzymes for the reaction include co-transaminase, preferably an R-transaminase, particularly T-185 (SEQ ID 1, FIG. 5) and T-186 (SEQ ID 2, FIG. 5), as well as transaminase (particularly co-transaminase, and preferably R-transaminase) enzymes havingan amino acid sequence that comprises: at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 96%, at least 97%, at least 98%, and at least 99% identity, to SEQ ID 1 or SEQ ID 2 (FIG. 5). The co- transaminase enzyme T-185 is originated from Shinella sp. (wild type) (see ChemBioChem 2021, 22, 1232- -1242). Particularly, the transaminase enzyme (particularly co-transaminase, and preferably R-transaminase) may have an amino acid sequence corresponding to SEQ ID 2 (T-186). The co- transaminase enzyme T-186 is engineered from an co-transaminase originated from Exophiala xenobiotica (variant T273S) (see ACS Catal. 2019, 9, 1140 1148) or can be from a cell lysate containing co-transaminase, and preferably R-transaminase particularly T-185 (SEQ ID 1, FIG. 5) and T-186 (SEQ ID 2, FIG. 5), which can be prepared as per Example 16.Example 1: Preparation of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole- 5-carboxamide (Compound 4)

[0145] 2-acetylthiazole-5-carboxylic acid (35 gm), 1,2-dimethoxyethane (3.0V) and dimethylformamide (0.05V) were charged into the reactor under nitrogen. Cooled to temperature of about 0°C to about 10°C. Oxalyl chloride (1.2 mole eq.) was added dropwise to the reaction mass. Temperature was raised to about 20°C to about 30°C. Stirred for period of about 1 hour to about 2 hours at temperature of about 20°C to about 30°C. After completion of reaction, reaction mass was cooled to temperature of about 0°C to about 5°C. Meanwhile in another RBF, solution of 5-chloro-4-(trifluoromethyl) pyridin-2 -amine hydrochloride (0.9 mole eq.), acetonitrile (4.0V) and pyridine (3.4 Mole Eq.) was prepared , cooled to temperature of about 0°C to about 5°C. 5-chloro-4-(trifluoromethyl)pyridin-2-amine hydrochloride solution was added dropwise into reaction mass at temperature of about 0°C to about 5°C. Stirred the reaction mass for period of about 1 hour. After completion of reaction, Temperature was raised to about 45°C to about 55°C and stirred for 1 hour. Water (10.0V) was added into reaction mass at temperature of about 45 °C to about 55 °C and stirred for 1 hour. Cooled to temperature of about 20°C to about 30°C. Stirred for 2 hours, filtered it and washed with water (6.0V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to 8 hours. 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (Compound 4) was obtained as Brown color solid. Yield: 86%; HPLC purity >98%.1H NMR (DMSO-d6): 2.66 (s, 3H), 8.57 (s, 1H), 8.80 (s, 1H), 9.00 (s, 1H), 12.01 (s, 1H).Example 2: Preparation of (R, E)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-((l- phenylethyl)imino)ethyl)thiazole-5-carboxamide (Compound 5)

[0146] 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (20 gm) (Compound 4), ethanol (7.0V), acetic acid (0.3 mole eq.) and R-phenylethylamine (4.0 Mole Eq.) (Compound 9) were charged into reactor. Temperature was raised to 70°C to 85°C. Stirred for period of about 5 hours to about 6 hours. After completion of reaction, reaction mass was cooled to about 20°C to about 30°C, stirred for 2 hours, solid precipitation was observed. Reaction mass was cooled to about 0°C to about 5 °C and stirred for period of about 2 hours, filtered and washed with chilled ethanol (1 Vol.), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to about 8 hours to obtain (R, E)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-( 1 -(( 1 - phenylethyl)imino)ethyl)thiazole-5-carboxamide (Compound 5). Yield: 85%.1H NMR (DMSO-d6): 1.43-1.44 (d, J= 6.51 Hz, 3H), 2.39 (s, 3H), 3.41-3.46 (q, J= 6.98 Hz, 1H), 7.34-7.37 (m, 5H), 8.58 (s, 1H), 8.72 (s, 1H), 8.77 (s, 1H).Example 3: Preparation of N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-(((R)-l- phenylethyl)amino)ethyl)thiazole-5-carboxamide (Compound 6)

[0147] (R,E)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-((l- phenylethyl)imino)ethyl) thiazole -5 -carboxamide (28 gm) (compound 5), Methanol (10V), and acetic acid (2.5V) were charged into the reactor under nitrogen and was cooled to temperature of about -5°C to 15°C. Sodium Borohydride (4.0 mole eq.) was added lot wise (6-8 lots) into it at temperature of about -5 C to about 15 °C. Stirred for period of about 1 hour to about 2 hours at temperature of about -5 to 15 °C. After completion of reaction, temperature was raised to about 20°C to 30°C. Water (10 vol.) and ethyl acetate (2X10 vol.) were added to it. Layers were separated. Organic layer washed with sat. aq. sodium bicarbonate solution (10V). Organic layer was distilled under vacuum at temperature was raised to about 40°C to 50°C. N-(5-chloro- 4-(trifluoromethyl)pyridin-2-yl)-2-( 1 -(((R)- 1 -phenylethyl)amino)ethyl)thiazole-5- carboxamide (Compound 6) was obtained as red-yellow oil. Yield; 85%; HPLC purity >96%.1H NMR (DMSO-d6): 1.27-1.29 (d, J= 6.63 Hz, 6H), 3.67-3.73 (m, 2H), 7.31-7.34 (m, 5H), 8.59 (s, 1H), 8.69 (s, 1H), 8.78(s, 1H), 11.67 (s, 1H).Example 4: Preparation of N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-(((R)-l- phenylethyl)amino)ethyl)thiazole-5-carboxamide (Compound 6)

[0148] (R, E)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-( 1 -(( 1 - phenylethyl)imino)ethyl) thiazole -5 -carboxamide (1.7 gm) (Compound 5), THF (5V) and acetic acid (2.5V) were charged into the reactor under nitrogen (Solution-1). Sodium borohydride (4.0 mole eq.), THF (10 V) were charged into another reactor at 20-30°C. L- Tartaric acid (4.0 mole eq.) was added into it. Temperature was raised to 60-70°C, stirred forperiod of about 4 to 5 hours at 60-70°C and then cooled to 20-30°C. Solution- 1 was added dropwise into it at 20-30°C. Stirred for 4-5 hours at 20-30°C. After completion of reaction, water (10 vol.) and ethyl acetate (2X10 vol.) was added. Layers were separated. Organic layer was washed with sat. aq. Sodium bicarbonate solution (10V). Organic layer was distilled under vacuum at 40°C to about 50°C to get N-(5-chloro-4-(trifhroromethyl)pyridin-2-yl)-2-(l-(((R)-1-phenylethyl)amino)ethyl)thiazole-5 -carboxamide (Compound 6) as red-yellow oil. Yield (85%). HPLC purity >96%.1H NMR (DMSO-d6): 1.27-1.29 (d, J= 6.63 Hz, 6H), 3.67-3.73 (m, 2H), 7.31-7.34 (m, 5H), 8.59 (s, 1H), 8.69 (s, 1H), 8.78(s, 1H), 11.67 (s, 1H).Example 5: Preparation of (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Compound 7)

[0149] N-(5-chloro-4-(trifhioromethyl)pyridin-2-yl)-2-(l-(((R)-l-phenylethyl)amino)ethyl) thiazole-5 -carboxamide (23 gm) (Compound 6), acetonitrile(lO.OV) and water (0.5V) were charged into reactor. Heated to temperature of about 60°C to about 65°C. N-bromosuccinimide (1.6 mole eq.) was added into it at temperature of about 60°C to about 65°C. Stirred for period of about 1 hours to about 2 hours at temperature of about 60°C to about 65 °C. After completion of reaction, cooled to temperature of about 20°C to about 30°C (heterogeneous reaction mass). Triethylamine (1.6 mole eq.) was charged to get clear solution. S-mandelic acid (1.2 moleeq.) solution in acetonitrile (3.0 Vol) was charged into it. Stirred for period of about 30 min (solid precipitation observed), acetonitrile (15 vol.) was added and stirred for period of about 2 hours and filtered it and washed with acetonitrile (2.0 V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to about 8 hours to get mandelic acid salt of N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-(((R)-l- phenylethyl)amino)ethyl) thiazole-5 -carboxamide as light yellowish solid. Saturated sodium bicarbonate solution (15 vol.) and ethyl acetate (15 vol.) were charged to it at temperature of about 20°C to about 30°C. Stirred for period of about 30 min. and layers were separated. Aqueous layer was extracted with ethyl acetate (2X 10 vol.). Combined organic layer was distilled under vacuum at temperature of about 60 °C to about 70°C to obtain (R)-2-(l- aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Compound 7).Yield: 57%; HPLC purity >98% and ee >96%.1H NMR (DMSO-d6): 1.38 (d, 3H), 4.21 (q, 1H), 8.59 (s, 1H), 8.69 (s, 1H), 8.76 (s, 1H), 11.51(brs, 1H).Example 6: Preparation of (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)- N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib) (Compound 1)

[0150] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (4.2 gm) (Compound 7), 6-amino-5-chloropyrimidine-4-carboxylic acid (Compound 8) (1.1 mole eq.) and dimethylacetamide (10V) were charged into reactor. 1- hydroxybenzotriazole monohydrate (0.4 mole eq.) and dimethylamino-propyl) -ethylcarbodiimide hydrochloride (2.0 mole eq.) were charged. After completion of reaction, n- butanol (10.0V) and water (20V) were added. Stirred for 2 hour (solid precipitation observed). Filtered it and washed with water, suck dried. Dried under vacuum at 50-60°C for 7-8 h to get Tovorafenib (compound 1) as white solid. Yield: 90%; HPLC purity >99% and ee >99%.1H NMR (DMSO-d6): 1.58-1.60 (d, J= 6.89 Hz, 3H), 5.32-5.38 (m, 1H), 7.43, 7.84 (brs, 2H), 8.37 (s, 1H), 8.57 (s, 1H), 8.76, 8.77 (s, 2H), 9.52-9.54 (d, J= 7.69 Hz, 1H), 11.77 (s, 1H).Example 7: Preparation of (7?)-2-(l-aminoethyl)-JV-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5-carboxamide (compound 7)

[0151] The transaminase-catalyzed reaction of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 4) was performed in 0.18 liter volume scale, with 5 g / 1 substrate concentration in 100 mM pH = 7.5 phosphate buffer, in presence of 5 v / v% DMSO as cosolvent and 0.5 mM pyridoxal phosphate as cofactor. Secbutylamine was used as amine donor in 200 mM concentration (~9-fold molar excess). The transaminase-catalyzed reaction was conducted with 50 m / m% of Prozomix PRO-TRANS No. 385 transaminase loading relative to the substrate, at 40 °C. The reaction mixture was sparged with nitrogen. The pH was controlled manually by addition of 33% sec-butylamine aqueous solution. After 24 hours the ratio of unreacted substrate was 26.68 Area%. The ratio of the produced ( / ?)-2-( l-aminocthyl)-N-(5-chloro-4-(trifluoromcthyl)pyridin-2-yl)thiazolc-5- carboxamide (compound 7) was 52.56 Area% and the enantiomeric excess was 97.2%.Example 8: Preparation of (7?)-2-(l-aminoethyl)-JV-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5-carboxamide (compound 7)

[0152] The transaminase-catalyzed reaction of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4) was performed in 0.18 liter volume scale, with 5 g / 1 substrate concentration in 100 mM pH = 7.5 phosphate buffer, in presence of 5 v / v% DMSO as cosolvent and 0.5 mM pyridoxal phosphate as cofactor. Sec- butylamine was used as amine donor in 200 mM concentration (~9-fold molar excess). The transaminase-catalyzed reaction was conducted with 50 m / m% of Prozomix PRO-TRANS No.436 transaminase loading relative to the substrate, at 40 °C. The reaction mixture was sparged with nitrogen. The pH was controlled manually by addition of 33% sec-butylamine aqueous solution. After 24 hours the ratio of unreacted substrate was 71.55 Area%. The ratio of the produced (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide of compound 7 was 14.59 Area% and enantiomeric excess was 95.1%.Example 9: Preparation of Form 1 of (R)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7)

[0153] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (compound 7, ~ 27.5 gm) was dissolved in 5.5 L of water and aqueous solution was acidified to pH 3.0-3.5 by 1: 1 aqueous HC1 (106 ml) at 20-30°C. The obtained clear solution was stirred for 10-20 min then filtered on glass fiber filter paper. The filter was washed with ImM aqueous HC1 (280 ml). The filtrate was extracted with isopropyl acetate (2x700 ml). The pH of aqueous layer was adjusted to pH 10.5-11.0 by 5 M aqueous potassium hydroxide and extracted with isopropyl acetate (2x700 ml). The combined organic layer was evaporated on rotavapor at 40-50°C. The solid residue was dried in vacuum oven (40°C, 80 mbar). The obtained solid was analyzed by XRPD. Crystalline (R)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7) Form 1 was obtained. An XRPD pattern is shown in FIG. 1.Yield: 19.4 gm (69%); HPLC purity: 99.8 Area% and enantiomeric excess 92.8%.Example 10: Preparation of (7?)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7)

[0154] N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-(((R)-l-phenylethyl)amino)ethyl) thiazole-5 -carboxamide (23 gm) (compound 6), acetonitrile (10.0V) and water (0.5V) were charged into reactor. Heated to temperature of about 60°C to about 65°C. N-bromosuccinimide (1.6 mole eq.) was added into it at temperature of about 60°C to about 65°C. Stirred for period of about 1 hours to about 2 hours at temperature of about 60°C to about 65 °C. After completion of reaction, cooled to temperature of about 20°C to about 30°C (heterogeneous reaction mass). Triethylamine (1.6 mole eq.) was charged to get clear solution. S-mandelic acid (1.2 mole eq.) solution in acetonitrile (3.0V) was charged into it. Stirred for period of about 30 min (solid precipitation observed), acetonitrile (15V) was added and stirred for period of about 2 hours and filtered it and washed with acetonitrile (2.0V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to about 8 hours to get mandelic acid salt of N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l-(((R)-l- phenylethyl)amino)ethyl) thiazole -5 -carboxamide as light yellowish solid. Saturated sodiumbicarbonate solution (15V) and ethyl acetate (15V) were charged to it at temperature of about 20°C to about 30°C. Stirred for period of about 30 min. and layers were separated. Aqueous layer was extracted with ethyl acetate (2 x 10V). Combined organic layer was distilled under vacuum at temperature of about 60 °C to about 70°C. The obtained solid was analyzed by XRPD. Crystalline (R)-2-( l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole -5 -carboxamide Form 1 was obtained.Yield: 57%; HPLC purity >98% and ee >96%.1H NMR (DMSO-d6): 1.38 (d, 3H), 4.21 (q, 1H), 8.59 (s, 1H), 8.69 (s, 1H), 8.76 (s, 1H), 11.51(brs, 1H).Example 11: Preparation of (R)-2-(l-(6- amino- 5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib) (compound 1)

[0155] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (2.0 gm) (compound 7), 6-amino-5-chloropyrimidine-4-carboxylic acid (compound 8) (1.1 mole eq.) and dimethylacetamide (7V) were charged into reactor. 1- hydroxybenzotriazole monohydrate (0.5 mole eq.) and dimethylamino-propyl) -ethylcarbodiimide hydrochloride (2.2 mole eq.) were charged. After completion of reaction, n- butanol (7.0V) and water (20V) were added. Stirred for 1-2 hour (solid precipitation observed). Filtered it and washed with water, suck dried. Dried under vacuum at 50-60°C for 7-8 h. The obtained white solid was analyzed by XRPD. Crystalline Tovorafenib was obtained. An XRPD pattern is shown in FIG. 2.Dry weight: 2.5 gm ; Yield: 86.7%; HPLC purity >99% and ee >99%.1H NMR (DMSO-d6): 1.58-1.60 (d, J= 6.89 Hz, 3H), 5.32-5.38 (m, 1H), 7.43, 7.84 (brs, 2H), 8.37 (s, 1H), 8.57 (s, 1H), 8.76, 8.77 (s, 2H), 9.52-9.54 (d, J= 7.69 Hz, 1H), 11.77 (s, 1H).Example -12: Preparation of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5-carboxamide (Compound 4)

[0156] 2-acetylthiazole-5 -carboxylic acid (10 gm), Acetonitrile (6.0V) and dimethylformamide (0.05V) were charged into the reactor under nitrogen at 20-30°C. Oxalyl chloride (1.1 mol. eq.) was added dropwise to the reaction mass at temperature of about 20°C to about 30°C. Stirred for period of about 1 hour to 2 hours at temperature of about 20°C to about 30°C. After completion of reaction, 5-chloro-4-(trifluoromethyl) pyridin-2 -amine hydrochloride (1.0 mol. eq.) was added at temperature of about 20°C to about 30°C. Pyridine (3.2 mol. eq.) was added dropwise at temperature of about 20°C to about 30°C. Reaction mass was stirred for period of about 1 hour to about 2 hour. After completion of reaction, temperaturewas raised to 50°C to about 60°C. Stirred for period of about 1 hour. Water (10V) was added into reaction mass at temperature of about 50°C to about 60°C. Stirred for 1 hour. Cooled to temperature of about 20°C to about 30°C. Stirred for period of about 2 hours, fdtered it and washed with water (2V) followed by acetonitrile (2V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to about 8 hours to get 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide as off white to brown color solid. The obtained solid was analyzed by XRPD. Crystalline 2-acetyl-N-(5- chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Compound 4) Form A was obtained. An XRPD pattern is shown in FIG. 3.Weight: 17.0; Yield (83%); HPLC purity>99.76%.Example 13: Preparation of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5-carboxamide (Compound 4)

[0157] 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (5 g) and acetonitrile (3.0V) are added into the reactor at temperature of about 20°C-30°C. Stirred for period of about 1 hour at 50°C-60°C. Cooled to temperature of about 20°C to about 30°C. Stirred for about 1 hours, filtered it and washed with acetonitrile (2.0V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C get 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide as off white to brown color solid. The obtained solid was analyzed by XRPD. Crystalline 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Compound 4) Form B was obtained. An XRPD pattern is shown in FIG.4.Weight: 4.5 gm Yield (90%), HPLC purity>99.92%.Example 14: Preparation of (R)-2-(l-(6-amino-5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib) (Compound 1)

[0158] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (82 gm ) (Compound 7), 6-amino-5-chloropyrimidine-4-carboxylic acid (Compound 8) (1.1 mole eq.) and dimethylformamide (6V) were charged into reactor. 1- hydroxybenzotriazole monohydrate (0.1 mole eq.) and dimethylamino-propyl) -ethylcarbodiimide hydrochloride (1.21 mole eq.) were charged. After completion of reaction, 2- propanol (6V) was added followed by heating to temperature of about 50°C to about 60°C. Water (20V) was added at temperature of about 50°C to about 60°C. Stirred for 1 hour to about 2 hour (solid precipitation observed). Reaction mass was cooled to 20-30°C, stirred for 1-2 hours. Filtered it and washed with water, suck dried. Dried under vacuum at 50-60°Cto get Tovorafenib (compound 1) as off white to white solid. Yield: 90%; HPLC purity >99.74% and ee >99.92%.Example 15: Purification of (R)-2-(l-(6-amino-5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib) (Compound 1)

[0159] (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)-N-(5-chloro-4- (trifluoromethyl) pyridin-2-yl)thiazole-5-carboxamide (80 g) (Compound 1), 2-propanol (6V) and dimethylformamide (6V) were charged into reactor and temperature was raised to 50°C- 60°C. After carbon treatment, water (20V) was added at 50°C-60°C. Stirred for period of about 1 hour to about 2 hour (solid precipitation observed). Reaction mass was cooled to temperature of about 20°C -30°C and stirred for period of about 1 hour to about 2 hours. Filtered it and washed with water and suck dried. Dried under vacuum at 50°C-60°C to get Tovorafenib (compound 1) as off white to white solid.Yield: 90%; HPLC purity >99.79% and ee >99.94%.Example 16: Preparation of Cell lysate containing co-transaminase, and preferably R- transaminase:

[0160] The composition of main culture medium:Yeast extract 20 g / 1, com steep powder 20 g / 1, glycerol 60 g / 1, NaCl 5 g / 1, KH2PO4 2.3 g / 1, K2HPO4 12.5 g / 1. The medium was adjusted to pH 7.4-7.6, then sterilized for 25 min at 121°C. 5 ml pre-culture was transferred into 50 ml expression culture medium in Erlenmeyer flask, and incubated in a rotary shaker for 3 h at 37°C, reaching the exponential growth phase.Enzyme expression was induced by arabinose (2 g / 1). After induction, the expression was carried out in Erlenmeyer shake flask in a rotary shaker for 24 h at 28°C. ODeoo after expression (before harvest) was measured to be >20.0.

[0161] Cells were harvested by centrifugation (4000 rpm, 15 min, 4°C) and lysis buffer was added to the cell pellets. Composition of the lysis buffer: 0.1 M Triethanolamine adjusted to pH 7.2 with diluted hydrochloric acid, 0.5 g / L lysozime, 0.5 mM PLP. Lysis was carried out starting from the cell pellet from 50 ml broth, and 4 ml lysis buffer was added. Homogenization was carried out by sonication. The crude lysate was further purified by centrifugation, (4000 rpm, 15 min, 4°C) the obtained clear supernatant was used in enzymatic reactions with 2- acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4).Example 17: Preparation of (7?)-2-(l-aminoethyl)-2V-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7)

[0162] The transaminase-catalyzed reaction of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 4) was performed in 4 ml reaction volume, at 7 g / 1 substrate concentration, containing 100 mM pH = 7.5 phosphate buffer, in presence of 5 v / v% DMSO as cosolvent and without extra added amount of pyridoxal phosphate as cofactor. Isopropylamine and D-alanine were used as amine donors, each in 200- 200 mM concentration. The enzymatic reaction was started by addition of the cell lysate containing transaminase T- 185, the volume ratio of the added lysate being 50 v / v% relative to the final reaction mixture.

[0163] The reaction mixture was shaken at 35 °C for 24 hours on an orbital shaker. After 24 hours the ratio of unreacted substrate was 87.1 Area%. The ratio of the produced ( / ?)-2-( l - aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7) was 7.0 Area% and the enantiomeric excess was 91.2%.Example 18: Preparation of (7?)-2-(l-aminoethyl)-A-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 7)

[0164] The transaminase-catalyzed reaction of 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 4) was performed in 4 ml reaction volume, at 7 g / 1 substrate concentration, containing 100 mM pH = 7.5 phosphate buffer, in presence of 5 v / v% DMSO as cosolvent and without extra added amount of pyridoxal phosphate as cofactor. Isopropylamine and D-alanine were used as amine donors, each in 200- 200 mM concentration. The enzymatic reaction was started by addition of the cell lysate containing transaminase T-186, the volume ratio of the added lysate being 50 v / v% relative to the final reaction mixture. The reaction mixture was shaken at 35 °C for 24 hours on an orbital shaker. After 24 hours the ratio of unreacted substrate was 90.7 Area%. The ratio of the produced ( / ?)-2-( l-aminocthyl)-N-(5-chloro-4-(trifliioromcthyl)pyridin-2-yl)thiazolc-5- carboxamide (compound 7) was 5.3 Area% and the enantiomeric excess was 95.9%.Example 19: Preparation of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5-carboxamide (Compound 4)

[0165] 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (5 g) (Form A) and acetonitrile (3 ,0V) are added into the reactor at temperature of about 20°C-30°C. Stirred for period of about 1 hour at 50°C-60°C. Cooled to temperature of about 20°C to about 30°C. Stirred for about 1 hours, filtered it and washed with acetonitrile (2.0V), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C get 2-acetyl-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide as off white to brown color solid. The obtained solid was analyzed by XRPD. Crystalline 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Compound 4) Form B was obtained. An XRPD pattern is shown in FIG.4.Weight: 4.5 gm Yield (90%), HPLC purity>99.92%.Example 20: Preparation of (R)-2-(l-(6-amino-5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib)(Compound 1)

[0166] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (120 gm ) (Compound 7), 6-amino-5-chloropyrimidine-4-carboxylic acid (Compound 8) (1.1 mole eq.) and dimethylformamide (5V) were charged into reactor. 1- hydroxybenzotriazole monohydrate (0.1 mole eq.) and dimethylamino-propyl) -ethylcarbodiimide hydrochloride (1.21 mole eq.) were charged at 20-30°C. Reaction mass was stirred for 1-2 hrs at same temperature. After completion of reaction, 2-propanol (5V) was added followed by addition of activated carbon (10%). Reaction mass stirred for 30 min followed by filtration and washing with dimethylformamide / 2-propanol (1: 1) (2V). DM water (10V) was added at 20-30°C. Stirred for 1-2 hour (solid precipitation observed). Adjust pH ~6.5 by triethylamine (0.04V) in IPA (15 ml) at 20-30°C, stirred for 1-2 hours. Filtered it and washed with 2-propanol / water (1: 1) (5V) followed by washing with hot water (2X3 V), suck dried. Dried under vacuum at 50-60°C to get Tovorafenib (compound 1) as off white to white solid. Yield: 86%; HPLC purity: 99.87% and ee: 99.86%.Example 21: Purification of (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)- N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide(Tovorafenib) (Compound 1)

[0167] (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)-N-(5-chloro-4- (trifluoromethyl) pyridin-2-yl)thiazole-5 -carboxamide (82 g) (Compound 1), 2-propanol (6V) and Dimethylformamide (6V) were charged into reactor. Heating to temperature 50- 60°C.Water (10V) was added at 50-60°C. Stirred for 1-2 hour. Cooled reaction mass to 20- 30°C, stirred for 1-2 hours. Filtered it and washed with 2-propanol / water (1: 1) (5V) followed by washing with hot water (2X3V), suck dried. Dried under vacuum at 50-60°C to get Tovorafenib (compound 1) as off white to white solid. Yield: 85%; HPLC purity: 99.99% and ee: 99.94%.Example 22: Preparation of (R)-2-(l-(6-amino-5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib)(Compound 1)

[0168] (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5- carboxamide (80 gm ) (Compound 7), 6-amino-5-chloropyrimidine-4-carboxylic acid (Compound 8) (1.1 Mole Eq.) and Dimethylformamide (6V) were charged into reactor. 1- hydroxybenzotriazole monohydrate (0.1 Mole Eq.) and dimethylamino-propyl)-ethyl- carbodiimide Hydrochloride (1.21 Mole Eq.) were charged at 20-30°C. Reaction mass was stirred for 1-2 hrs at same temperature. After completion of reaction, 2-propanol (6V) was added followed by addition of Water (20V) was added at 20-30°C. Stirred for 1-2 hour (solid precipitation observed). Adjust pH ~6.5 by Triethylamine (0.04V) in IPA (16 ml) at 20-30°C, stirred for 1-2 hours. Filtered it and washed with water (2X3V), suck dried. Dried under vacuum at 50-60°C to get Tovorafenib (compound 1) as off white to white solid. Yield: 90.40%; HPLC purity 99.54 % and ee 99.88 %Example 23: Purification of (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)- N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide(Tovorafenib) (Compoundl)

[0169] (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)-N-(5-chloro-4- (trifluoromethyl) pyridin-2-yl)thiazole-5-carboxamide (80 g) (Compound 1), 2-propanol (5V) and Dimethylformamide (5V) were charged into reactor at temperature 20°C-30°C. Stirred for 30-60 min. Charged Eno carbon (5% w / w) at temperature 20°C-30°C. Stirred for 30 min to 60 min at temperature 20°C-30°C.Fiiltered over Hyflo bed and washed with mixture of 2- propanol and Dimethylformamide (1: 1)(2V) at temperature 20°C-30°C.Heated filtered reaction mass to temperature 50-60°C.Added water (6V) in 30-60 min at temperature 50°C- 60°C. Stirred for 30-60 min temperature 50°C-60°C. Added water (4V) in 30-60 min at temperature 50°C-60°C. Stirred for 60-120 min temperature 50°C-60°C. Cooled reaction mass to temperature 20-30°C, stirred for 1-2 hours at temperature 20-30°C. Filtered it temperature 20-30°C and washed with Acetonitrile / 2-propanol / water (3: 1: 1) (5V) followed by washing with hot water (2X3V), suck dried. Dried under vacuum at 50-60°C to get Tovorafenib (compound 1) as off white to white solid. Yield: 83.40%; HPLC purity 99.92% and ee 99.94%.Example 24: Preparation of (R, E)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)-2-(l- ((l-phenylethyl)imino)ethyl)thiazole-5-carboxamide (Compound-5)

[0170] 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (20 gm) (Compound 4), ethanol (6.0V), Acetic acid (0.3 Mole Eq.) and R-phenyl ethyl amine (4.0 Mole Eq.) were charged into reactor. Temperature was raised to about 70°C to about 85°C. Stirred for period of about 5 hours to about 6 hour. After completion of reaction. Reaction mass was cooled to about 20°C to about 30°C, Stirred for 2h, Solid precipitationwas observed. Reaction mass was cooled to about 0°C to about 5°C and stirred for period of about 2 hours, fdtered and washed with chilled ethanol (1 Vol.), suck dried. Dried under vacuum at temperature of about 50°C to about 60°C for period of about 7 hours to about 8 hours. Yield 89.14%; 1H NMR (DMS0-d6): 1.43-1.44 (d, J= 6.51 Hz, 3H), 2.39 (s, 3H), 3.41-3.46 (q, J= 6.98 Hz, 1H), 7.34-7.37 (m, 5H), 8.58 (s, 1H), 8.72 (s, 1H), 8.77 (s, 1H).Example 25: Purification of (R)-2-(l-(6-amino-5-chloropyrimidine-4- carboxamido)ethyl)-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (Tovorafenib) (Compound 1)

[0171] (R)-2-(l-(6-amino-5-chloropyrimidine-4-carboxamido)ethyl)-N-(5-chloro-4- (trifluoromethyl) pyridin-2-yl)thiazole-5-carboxamide (80 g) (Compound 1), 2-propanol (6V) and dimethylformamide (6V) were charged into reactor and temperature was raised to 50°C- 60°C. Activated carbon (5% w / w) was added, and the mixture stirred at 50°C-60°C for 30-60 minutes. The reaction mass was filtered over a Hyflo bed at 50°C-60°C. Water (20V) was slowly added to the filtrate over a period of about 60-90 minutes at 50°C-60°C. The mixture was stirred for period of about 1 hour to about 2 hours at 50°C-60°C (solid precipitation observed). Reaction mass was cooled to temperature of about 20°C -30°C and stirred for period of about 1 hour to about 2 hours. Filtered it and washed with water and suck dried. Dried under vacuum at 50°C-60°C to get Tovorafenib (compound 1) as off white to white solid.Yield: 90%; HPLC purity >99.79% and ee >99.94%.Example 26: Preparation of Cell lysate containing T-185 enzyme

[0172] The composition of main culture medium:Yeast extract 20 g / 1, com steep powder 20 g / 1, glycerol 60 g / 1, NaCl 5 g / 1, KH2PO4 2.3 g / 1, K2HPO4 12.5 g / 1. The medium was adjusted to pH 7.4-7.6, then sterilized for 25 min at 121°C.

[0173] Escherichia coli TOP 10 recombinant strain was used for the enzyme expression. The strain contained a plasmid which harbored the gene sequence encoding T-185 enzyme and pBad promoter. The plasmid was assembled by molecular cloning by a method similar to that disclosed in ChemBioChem 2021, 22, 1232-1242.5 ml preculture of the recombinant strain was transferred into 50 ml expression culture medium in Erlenmeyer flask, and incubated in a rotary shaker for 3 h at 37°C, reaching the exponential growth phase.

[0174] Enzyme expression was induced by arabinose (2 g / 1). After induction, the expression was carried out in Erlenmeyer shake flask in a rotary shaker for 24 h at 28°C. ODeoo after expression (before harvest) was measured to be >20.0.Cells were harvested by centrifugation (4000 rpm, 15 min, 4°C) and lysis buffer was added to the cell pellets. Composition of the lysis buffer: 0.1 M Triethanolamine adjusted to pH 7.2 with diluted hydrochloric acid, 0.5 g / L lysozime, 0.5 mM PLP. Lysis was carried out starting from the cell pellet from 50 ml broth, and 4 ml lysis buffer was added. Homogenization was carried out by sonication. The crude lysate was further purified by centrifugation, (4000 rpm, 15 min, 4°C) the obtained clear supernatant was used in enzymatic reactions with 2-acetyl-N- (5 -chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 4) .Example 27: Preparation of Cell lysate containing T-186 enzyme

[0175] The composition of main culture medium:Yeast extract 20 g / 1, com steep powder 20 g / 1, glycerol 60 g / 1, NaCl 5 g / 1, KH2PO4 2.3 g / 1, K2HPO4 12.5 g / 1. The medium was adjusted to pH 7.4-7.6, then sterilized for 25 min at 121°C.

[0176] Escherichia coli TOP 10 recombinant strain was used for the enzyme expression. The strain contained a plasmid which harbored the gene sequence encoding T-186 enzyme and pBad promoter. The plasmid was assembled by molecular cloning by a method similar to that disclosed in ChemBioChem 2021, 22, 1232-1242.

[0177] 5 ml pre-culture of the recombinant strain was transferred into 50 ml expression culture medium in Erlenmeyer flask, and incubated in a rotary shaker for 3 h at 37°C, reaching the exponential growth phase.

[0178] Enzyme expression was induced by arabinose (2 g / 1). After induction, the expression was carried out in Erlenmeyer shake flask in a rotary shaker for 24 h at 28°C. ODeoo after expression (before harvest) was measured to be >20.0.

[0179] Cells were harvested by centrifugation (4000 rpm, 15 min, 4°C) and lysis buffer was added to the cell pellets. Composition of the lysis buffer: 0. 1 M Triethanolamine adjusted to pH 7.2 with diluted hydrochloric acid, 0.5 g / L lysozime, 0.5 mM PLP. Lysis was carried out starting from the cell pellet from 50 ml broth, and 4 ml lysis buffer was added.Homogenization was carried out by sonication. The crude lysate was further purified by centrifugation, (4000 rpm, 15 min, 4°C) the obtained clear supernatant was used in enzymatic reactions with 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide (compound 4).

Claims

CLAIMS1. A process for the preparation of Tovorafenib or a salt thereof, comprising reacting 2- acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide of formula (4):with a transaminase enzyme in the presence of an amine donor, to obtain compound of formula (7) or salt thereof:; and converting the compound of formula (7) to Tovorafenib or a salt thereof.

2. The process forthe preparation of Tovorafenib or a salt thereof, according to Claim 1, comprising: a) reacting 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide of formula (4):with R-selective transaminase enzyme in the presence of an amine donor, to obtain compound of formula (7) or a salt thereof; and(7) ; and b) reacting the compound of formula (7) or a salt thereof:with the compound of formula (8):to obtain Tovorafenib, or a salt thereof.

3. The process according to any of Claim 1 or Claim 2, wherein the reaction is carried out in the presence of an enzyme cofactor, preferably wherein the enzyme cofactor is pyridoxal phosphate.

4. The process according to any of Claims 1, 2 or 3, wherein the amine donor is selected from: a chiral or achiral amino acid and a chiral or achiral amine.

5. The process according to any of Claims 1, 2, 3, or 4, wherein the amine donor is an amino acid (preferably selected from: alanine, glycine, glutamate or beta-alanine); or an amine or salt thereof, preferably an aliphatic amine (particularly a C3 to C8 “alkyl monoamine or a C3 to C8 “■alkyl diamine), more particularly selected from: isopropylamine, sec-butylamine, butane- 1,4-diamine, pentane-l,5-diamine, or but-2-enediamine; or an aromatic amine (particularly a C5^ to C8 “■aromatic amine), particularly phenethylamine, 1 -phenylethylamine, benzylamine, or o-xylylenediamine; or mixtures thereof.

6. The process according to any of Claims 1, 2, 3, 4, or 5, wherein the amine donor is selected from isopropylamine, sec-butylamine or D-alanine., or a mixture thereof, particularly sec-butylamine, or a mixture of isopropylamine and D-alanine.

7. The process according to any of Claims 1, 2, 3, 4, 5, or 6, wherein the transaminase enzyme is an co transaminase, preferably an R-transaminase, more particularly wherein the transaminase has an amino acid sequence having an amino acid sequence that has: at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 96%, at least 97%, at least 98%, and at least 99% identity, to SEQ ID 1 or SEQ ID 2:

8. The process according to any of Claims 1, 2, 3, 4, 5, 6, or 7, wherein the transaminase enzyme has an amino acid sequence that comprises at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 96%, at least 97%, at least 98%, and at least 99% identity to: at least 50, at least 100, at least 150, at least 200, or at least 250, contiguous amino acid residues of SEQ ID 1 or SEQ ID 2.

9. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, or 8, wherein the transaminase enzyme has an amino acid sequence according to SEQ ID 1 or SEQ ID 2 which comprises: one or more, five or more, ten or more, 15 or more, 20 or more, 25 or more, 30 or more, 35 or more, or 40, mutations at a position selected from 14, 21, 22, 25, 35, 38, 40, 43, 50, 69, 71, 73, 76, 81, 102, 108, 122, 137, 146, 150, 156, 191, 195, 197, 198, 200, 215, 224, 225, 251, 255, 262, 263, 265, 267, 269, 272, 273, 281, and 284.

10. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, 8, or 9, wherein the transaminase enzyme has an amino acid sequence corresponding to: SEQ ID 1 or SEQ ID 2.

11. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10, wherein the reaction of compound (4) with a transaminase enzyme in the presence of an amine donor is carried out at a pH of: about 7 to about 11.5, or about 6.8 to about 11.5, about 6.8 to about 10.5, about 6.9 to about 9.5, about 7.0 to about 8.5, about 7.1 to about 8.2, about 7.2 to about 8.0, or about 7.3 to about 7.8, about 7.4 to about 7.6 or about 7.5.

12. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11, wherein the reaction of compound (4) with a transaminase enzyme in the presence of an amine donor is carried out at a temperature of: about 30°C to about 55°C, about 32°C to about 50°C, about 35°C to about 45°C, about 38°C to about 42°C, or about 40°C.

13. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12, wherein the reaction is carried in the presence of a phosphate buffer.

14. The process according to any of Claims 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, or 13, wherein the reaction is carried out in an aqueous mixture, optionally in the presence of a co-solvent, preferably wherein the co-solvent is selected from DMSO, acetonitrile, dimethylformamide, THF, and more preferably DMSO.

15. A process for preparation of Tovorafenib or a salt thereof, comprising a) reacting 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2-yl)thiazole-5-carboxamide of formula (4):with an amine of formula (II):wherein R1 and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and R1 and R2 cannot be same at the same time,to obtain compound of formula (III):b) reducing the compound of formula (III):to obtain a compound of formula (IV) or salt thereof:c) deprotecting the compound of formula (IV) or salt thereof:to obtain a compound of formula (7) or salt thereof:d) optionally purifying the compound of formula (7) or salt thereof; and e) reacting the compound of formula (7) or salt thereof:with a compound of formula (8):to obtain Tovorafenib or a salt thereof.

16. The process according to Claim 15, wherein R1 is: phenyl; substituted phenyl (preferably selected from the group consisting of methylbenzoyl, 4-nitrophenyl, 4- methoxyphenyl, p-tolyl, 5 -isopropyl-3 -methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4-methoxynaphthyl); alkyl (preferably C1-C6-'-' alkyl, more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Cl to C2), or methoxyalkyl (Cl to C2)]; cycloalkyl (preferably C5 to C8 cycloalkyl, and particularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of C5 to C8 branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl).

17. The process according to any of Claim 15 or Claim 16, wherein R2 is: phenyl; substituted phenyl (preferably methylbenzoyl, 4-nitrophenyl, 4-methoxyphenyl, p-tolyl, 5-isopropyl-3-methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4- methoxynaphthyl); alkyl (preferably C1-C6-'-' alkyl, more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Cl to C2), or methoxyalkyl (Cl to C2)]; cycloalkyl (preferably C5 to C8 cycloalkyl, and particularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of C5 to C8 branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl).

18. The process according to any of Claims 15, 16, or 17, wherein R-4 and R2 are selected from unsubstituted C1-C6 alkyl or unsubstituted C_,6-C10 aryl; preferably wherein R-1! and R2 are selected from unsubstituted C 1 -_,C3 alkyl, or unsubstituted C6-C8 aryl, and more preferably wherein the amine (II) is (R)-phenylethylamine.

19. The process according to any of Claims 15, 16, 17, or 18, wherein step a) comprises reacting the compound of formula (4) or a salt thereof:with R-phenylethylamine of formula (9):NH2^"Ph(9)to obtain compound of formula (5) or a salt thereof:

20. The process according to any of Claims 15, 16, 17, 18, or 19, wherein step (a) is carried out in an organic solvent, preferably dichloromethane, dimethoxy ethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n- heptane, 2-methyl THF, or mixtures thereof, and preferably wherein the organic solvent is a Cl to C~4 alcohol, preferably methanol, ethanol or isopropanol, and particularly ethanol.

21. The process according to any of Claims 15, 16, 17, 18, 19, or 20, wherein step (a) is carried out in the presence of an inorganic acid or Lewis acid (particularly hydrochloric acid, BF3, TiC14, A1C13, titanium isopropoxide, SnC-1, ZnC12, or ZnBr^2^). or an organic acid; particularly an organic acid (preferably a C2-C6 carboxylic acid, particularly a C2 to C4 carboxylic acid, and more preferably propanoic acid or acetic acid, and particularly acetic acid.

22. The process according to any of Claims 15, 16, 17, 18, 19, 20, or 21, wherein step (a) is carried out at a temperature of: about 50°C to about 100°C, about 55°C to about 95°C, about 60°C to about 90°C, or about 70°C to about 85°C.

23. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, or 22, wherein step (a) comprises isolating the compound of formula (III), preferably by cooling and precipitation from the reaction mixture, and isolating the solid, preferably by fdtration.

24. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, or 23, wherein step b) comprises reducing the compound of formula (5):to obtain the compound of formula (6):

25. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, or 24, wherein the reducing agent in step (b) is selected from: hydrogen gas, a hydride reducing agents (preferably sodium borohydride, lithium borohydride, Alane, DiBAL-H, sodium triacetoxyborohydride, LiAlH_,4, sodium cyanoborohydride, borane, triethylsilane, trichlorosilane, Hantzsch ester, CBS reagent or hydride, reducing agent derived from a reaction between sodium borohydride and a chiral acid (preferably L-tartaric acid, L-proline, or S- mandelic acid etc); and preferably wherein the reducing agent is sodium borohydride.

26. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or 25, wherein step b) is carried out in the presence of one or more solvents, preferably methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxy ethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2- methyl THF, water, or mixtures thereof; more preferably wherein the solvent is selected from: methanol, ethanol, isopropyl alcohol or tetrahydrofuran.

27. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, or 26, wherein step (b) is carried out in the presence of an inorganic acid or Lewis acid (particularly hydrochloric acid, BF3, TiC14, A1C13, titanium isopropoxide, SnC-1, ZnC12, or ZnBr-,2-'), or an organic acid; particularly an organic acid( preferably a C2-C6 carboxylicacid, particularly a C2 to C4 carboxylic acid, and more preferably propanoic acid or acetic acid, and particularly acetic acid.

28. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, or 27, wherein step c) comprises deprotecting the compound of formula (6):to obtain a compound of formula (7):

29. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26,27, or 28, wherein step (c) comprises reacting the compound of formula (IV) with an oxidizing agent, preferably in the presence of a base and a solvent.

30. The process according to Claim 29, wherein the oxidizing agent is selected from: bromine gas, chlorine gas, N-chlorosuccinimide, N-bromosuccinimide, N-bromophthalimide, N-chlorophthalimide, trichloroisocyanuric acid, chlorohydantoin, borohydantoin, 2, 3, 4, 5, 6, 6- hexachlorocyclohexa-2,4-dien-l-one, or 4,4-dibromo-2,6-di-tert-butyl-cyclohexa-2,5- dienone; and preferably wherein the oxidising agent is N-bromosuccinimide.

31. The process according to any of Claim 29 or Claim 30, wherein the base is an organic base, preferably selected from: triethylamine, diisopropyl ethylamine, pyridine, diethylamine, diisopropylamine, N,N-dimethylaminopyridine, l,8-diazabicyclo[5.4.0]undec-7-ene, 1,4-diazabicyclo[2.2.2]octane, piperazine, pyrrolidine, piperidine, morpholine, N- methylmorpholine, N-methylpyrrolidine, N-methyl piperidine, or N,N-dimethylpiperazine; and more preferably wherein the base is triethylamine.

32. The process according to any of Claims 29, 30 or 31, wherein the solvent is selected from: methanol, ethanol, isopropanol, n-propanol, n-butanol, dichloromethane, dimethoxyethane, ethyl acetate, isopropyl acetate, methyl acetate, acetonitrile, tetrahydrofuran, MTBE, toluene, n-heptane, 2-methyl THF, water or mixture thereof; and preferably wherein the solvent is acetonitrile, water or mixture thereof.

33. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, or 32, wherein step d) comprises purification of compound (7), preferably by fractional recrystallization or a diastereomer salt separation, preferably wherein the purification of compound (7) comprises crystallization of the diastereomer salt with an optically active di-p-toluoyl-tartaric acid or an optically active mandelic acid, preferably di-p- toluoyl-(D)-tartaric acid or (S)-mandelic acid, and more preferably (S)-mandelic acid.

34. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26,27, 28, 29, 30, 31, 32, or 33, wherein step e) comprises reacting the compound of formula (7):with the compound of formula (8):

35. The process according to any of Claims 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, or 34, wherein step (e) comprises reacting compound (7) with compound (8) in the presence of a coupling agent, a coupling additive and a solvent.

36. The process according to Claim 35, wherein the coupling agent is 1- hydroxybenzotriazole .

37. The process according to Claim 35 or Claim 36, wherein the coupling additive is l-(3- dimethylaminopropyl)-3 -ethylcarbodiimide .

38. The process according to any of Claims 35, 36, or 37, wherein the solvent is dimethylacetamide .

39. The process according to any preceding claim, further comprising combining the Tovorafenib or a pharmaceutically acceptable salt thereof, with at least one pharmaceutically acceptable excipient to form a pharmaceutical composition.

40. A compound of formula (III):or a salt thereof, wherein R1 and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and R1 and R2 cannot be same at the same time.

41. Use of a compound of formula (III) as defined in Claim 40, in the preparation of Tovorafenib or a salt thereof.

42. A compound of formula (IV):or a salt thereof, wherein R1 and R2 are substituted or unsubstituted alkyl, substituted or unsubstituted cycloalkyl, substituted or unsubstituted aryl; substituted or unsubstituted phenyl; or substituted or unsubstituted naphthyl and R1 and R2 cannot be same at the same time.

43. Use of a compound (IV) as defined in Claim 42, in the preparation of Tovorafenib or a salt thereof.

44. The compound or use thereof, according to any of Claims 40, 41, 42, or 43, wherein R1 is phenyl; substituted phenyl (preferably selected from the group consisting of methylbenzoyl, 4-nitrophenyl, 4-methoxyphenyl, p-tolyl, 5-isopropyl-3-methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4-methoxynaphthyl); alkyl (preferably C1-C6-’-' alkyl, more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Cl to C2), or methoxyalkyl (Cl to C2)]; cycloalkyl (preferably C5 to C8 cycloalkyl, and particularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of C5 to C8 branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl) .

45. The compound or use thereof, according to any of Claims 40, 41, 42, 43, or 44, wherein R2 is selected from the group consisting of: phenyl; substituted phenyl (preferably methylbenzoyl, 4-nitrophenyl, 4-methoxyphenyl, p-tolyl, 5-isopropyl-3-methylphenyl); naphthyl; substituted naphthyl (preferably 4-nitronaphthyl or 4-methoxynaphthyl); alkyl (preferably C1-C6-’-' alkyl, more preferably methyl, ethyl, propyl or butyl); substituted alkyl [preferably selected from the group consisting of C1-C7 branched alkyl group, hydroxylalkyl (Cl to C2), or methoxyalkyl (Cl to C2)]; cycloalkyl (preferably C5 to C8 cycloalkyl, andparticularly cyclopropyl, cyclobutyl, cyclopentyl, or cyclohexyl), or substituted cycloalkyl (preferably selected from the group consisting of C5 to C8 branched cycloalkyl group, hydroxylcycloalkyl, or methoxy cycloalkyl).

46. The compound or use thereof, according to any of Claims 40, 41, 42, 43, 44, or 45, wherein R1 and R2 are selected from unsubstituted alkyl (preferably C1-C6 alkyl), unsubstituted cycloalkyl (preferably C5-C10 alkyl, more preferably C5 to C6 cycloalkyl), unsubstituted aryl (preferably C6-C10 aryl); and more preferably R1 and R2 are selected from Cl to C3 alkyl (preferably methyl), unsubstituted phenyl; or unsubstituted naphthyl.

47. The compound or use thereof, according to any of Claims 40, 41, 42, 43, 44, 45, or 46, wherein R-11 and R2 are selected from unsubstituted C1-C6 alkyl or unsubstituted C_,6- C10 aryl; preferably wherein R^ l and R2 are selected from unsubstituted C l -^C3 alkyl, or unsubstituted C6-C8 aryl; particularly wherein R1 and R2 are selected from methyl and phenyl.

48. A compound of formula (5):

49. Use of the compound (5) according to Claim 48 in the preparation of Tovorafenib or a salt thereof.

50. A compound of formula (6):

51. Use of the compound (6) as defined in Claim 50 in the preparation of Tovorafenib or a salt thereof.

52. A crystalline polymorph of (R)-2-(l-aminoethyl)-N-(5-chloro-4-(trifluoromethyl)- pyridin-2-yl)thiazole-5-carboxamide (compound 7), designated Form 1, which is characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 1; or an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 17.6 and 24.3 degrees 2-theta ± 0.2 degrees 2-theta; and combinations of these data.

53. The crystalline polymorph of (R)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)pyridin-2-yl)thiazole-5 -carboxamide (compound 7) according to Claim 52, which is further characterized by an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 17.6 and 24.3 degrees 2-theta ± 0.2 degrees 2-theta, and also having any one, two, three, or four additional peaks selected from 11.7, 16.3, 21.3 and 23.4 degrees 2-theta ± 0.2 degrees 2- theta; or which is characterized by an X-ray powder diffraction pattern having peaks at 8.1, 10.6, 11.7, 16.3, 17.6, 21.3, 23.4, and 24.3 degrees 2-theta ± 0.2 degrees 2-theta..

54. Use of the crystalline polymorph of (R)-2-(l-aminoethyl)-N-(5-chloro-4- (trifluoromethyl)-pyridin-2-yl)thiazole-5 -carboxamide (compound 7) according to any of Claims 52 and Claim 53, in the preparation of Tovorafenib or a salt thereof.

55. A crystalline polymorph of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5 -carboxamide (compound 4), designated Form A, which is characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 3; or an X-ray powder diffraction pattern having peaks at 17.7, 20.1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 at degrees 2-theta; and combinations of these data.

56. The crystalline polymorph of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole -5 -carboxamide (compound 4) according to Claim 55, which is further characterized by an X-ray powder diffraction pattern having peaks at 17.7, 20.1, 22.1, 26.3 and 27.6 degrees 2-theta ± 0.2 degrees 2-theta, and also having any one, two, three or four additional peaks selected from at 12.4, 13.3, 14.5 and 17.3 degrees 2-theta ± 0.2 degrees 2-theta; or which is characterized by an X-ray powder diffraction pattern having peaks at 12.4, 13.3, 14.5, 17.3 17.7, 20.1, 22.1, 26.3, and 27.6 degrees 2-theta ± 0.2 degrees 2-theta.

57. Use of the crystalline polymorph of (2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5 -carboxamide (compound 4) according to any of Claims 55 and 56, in the preparation of Tovorafenib or a salt thereof.

58. A crystalline polymorph of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole-5 -carboxamide (compound 4), designated Form B, which is characterized by data selected from one or more of the following: an X-ray powder diffraction pattern substantially as depicted in FIG. 4; and an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 18.6, 21.0, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta; and combinations of these data.

59. The crystalline polymorph of 2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin-2- yl)thiazole -5 -carboxamide (compound 4) according to Claim 58, which is further characterized by an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 18.6, 21.0, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta, and also having any one, two, or three additional peaks selected from peaks at 17.4, 22.1, and 22.7 degrees 2-theta ± 0.2 degrees 2-theta; or which is characterized by an X-ray powder diffraction pattern having peaks at 11.4, 15.2, 17.4, 18.6, 21.0, 22.1, 22.7, and 27.3 degrees 2-theta ± 0.2 degrees 2-theta.

60. Use of the crystalline polymorph of (2-acetyl-N-(5-chloro-4-(trifluoromethyl)pyridin- 2-yl)thiazole-5 -carboxamide (compound 4) according to any of Claims 58 and 59, in the preparation of Tovorafenib or a salt thereof.

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