New methylquinazolinone derivatives

Abstract

The invention provides a novel compound having the general formula (I), or a pharmaceutically acceptable salt thereof, and wherein X, R1-R6 are as described herein. The compound of formula (I) can be used as a medicament.

Description

[0001] F. Hoffmann-La Roche AG, CH-4070 Basel, Switzerland

[0002] Case P38576

[0003] NEW METHYLQUINAZOLINONE DERIVATIVES

[0004] The present invention provides a new compound, its manufacture, pharmaceutical compositions containing it and its use as therapeutically active substance. The compound of the invention is a paradox breaking BRAF inhibitor and has favorable pharmacokinetic properties.

[0005] The present invention provides a novel compound of formula (I) wherein

[0006] D is deuterium;

[0007] R1is selected from i) Ci-6-alkyl, and ii) Ci-6-haloalkyl;

[0008] R2is selected from iii) hydrogen, and iv) halogen;

[0009] X is selected from v) -NH-, and vi) -O-;

[0010] R3is selected from vii) H, viii) cyano, and ix) halogen; R4is selected from i) hydrogen, and ii) halogen;

[0011] R5is selected from iii) cyano, and iv) halogen;

[0012] R6is selected from v) NR7R8, and vi) CHR9R10;

[0013] R7is selected from vii) Ci-6-alkyl, viii) Cs-s-cycloalkyl, and ix) C3-8-cycloalkyl-Ci-6-alkyl;

[0014] R8is selected from x) Ci-6-alkyl, xi) C3-8-cycloalkyl, and xii) C3-8-cycloalkyl-Ci-6-alkyl; or R7and R8together with the nitrogen atom to which they are attached form an heterocycloalkyl optionally substituted with R11;

[0015] R9is selected from xiii) Ci-6-alkyl, xiv) C3-8-cycloalkyl, and xv) C3-8-cycloalkyl-Ci-6-alkyl;

[0016] R10is selected from xvi) Ci-6-alkyl, xvii) Cs-s-cycloalkyl, and xviii) C3-8-cycloalkyl-Ci-6-alkyl; or R9and R10together with the carbon atom to which they are attached form a C3-8- cycloalkyl optionally substituted with R11; and

[0017] R11is halogen; or a pharmaceutically acceptable salt thereof.

[0018] The invention further provides in particular a compound of formula (la) or a pharmaceutically acceptable salt thereof.

[0019] The invention further provides in particular a compound of formula (lb) or a pharmaceutically acceptable salt thereof

[0020] The Rapidly Accelerated Fibrosarcoma (RAF) class of serine-threonine kinases comprise three members (ARAF, BRAF, RAFI) that compose the first node of the MAP kinase signalling pathway. Despite the apparent redundancy of the three RAF isoforms in signalling propagation through phosphorylation of MEK1 and 2, frequent oncogenic activating mutations are commonly found only for BRAF. In particular, substitution of valine 600 with glutamic acid or lysine renders the kinase highly activated with consequent hyper-stimulation of the MAPK pathway, independently from external stimulations (Cell. 2015 Jun 18; 161(7): 1681-1696). Mutant BRAF is a targetable oncogenic driver and three BRAF inhibitors (vemurafenib, dabrafenib and encorafenib) reached the market up to now showing efficacy in BRAFV600E- positive melanoma. However rapid acquisition of drug resistance is almost universally observed and the duration of the therapeutic benefits for the targeted therapy remains limited.

[0021] Moreover, the developed BRAF inhibitors revealed an unexpected and “paradoxical” ability to repress MAPK signalling in BRAFV600E-driven tumours while the same inhibitors presented MAPK stimulatory activities in BRAF wild type (WT) models (N Engl J Med 2012; 366:271-273; and British Journal of Cancer volume 111, pages640-645(2014)).

[0022] Mechanistic studies on the RAF paradox then clarified that oncogenic BRAFV600E phosphorylates MEK 1 / 2 in its monomeric cytosolic form while WT BRAF and RAFI activation requires a complex step of events including cell membrane translocation and homo and / or heterodimerization promoted by activated RAS (KRAS, NRAS, HRAS) (Nature Reviews Cancer volume 14, pages455-467(2014)).

[0023] The binding of inhibitors like vemurafenib, dabrafenib or encorafenib to a WT BRAF or RAFI protomer, quickly induces RAF homo and / or hetero dimerization and membrane association of the newly formed RAF dimer. In the dimeric conformation, one RAF protomer allosterically induces conformational changes of the second resulting in a kinase active status and, importantly, in a conformation unfavourable for the binding of the inhibitor. The dimer induced by drug treatment, as a result, promotes MEK phosphorylation by the catalysis operated by the unbound protomer with hyperactivation of the pathway.

[0024] The RAF paradox results in two clinically relevant consequences:

[0025] 1) accelerated growth of secondary tumours upon BRAFi monotherapy (mainly keratochantoma and squamous-cell carcinomas) (N Engl J Med 2012; 366:271-273), and

[0026] 2) the acquisition of drug resistance in the setting of BRAFi monotherapy as well as in combinations of BRAFi+MEKi. Potential mechanisms are the activation of dimer-mediated RAF signalling by genetically driven events including RAS mutations, BRAF amplifications, expression of dimeric-acting BRAF splice variants (Nature Reviews Cancer volume 14, pages 455-467(2014)).

[0027] There is thus the need for RAF inhibitors capable of breaking that paradox.

[0028] Furthermore, the currently approved classical BRAF inhibitors Vemurafenib (Mol. Pharmaceutics 2012, 9, 11, 3236-3245), Dabrafenib (J Pharmacol Ex Ther 2013, 344 (3) 655- 664) and Encorafenib (Pharmacol Res. 2018;129:414-423) all have very poor brain permerability. This is major limitation for the use of those classical BRAF inhibitors for the treatment of brain cancer or brain metastases. There is thus the need for BRAF inhibitors having improved brain permeability and / or brain exposure.

[0029] Recently a novel generation of BRAF inhibitors has been described (WO2021116050A1 and WO2021116055A1) showing considerably less paradoxical activation of the MAPK signaling pathway while retaining high potency. These compounds can thus be referred to as paradox breakers or RAF paradox breakers, in contrast to compounds inducing the RAF paradox (and which could be referred to as paradox inducers or RAF paradox inducers). One particular compound is (3A)-A-[2-cyano-4-fluoro-3-(3-methyl-4-oxo-quinazolin-6-yl)oxy-phenyl]-3- fluoro-pyrrolidine-1 -sulfonamide, which is currently in clinical studies and which is also known as RO7276389 (herein also referred to as compound of formula (lb). In addition to being a paradox breaker, this novel generation BRAF inhibitor also has very potent brain penetration properties, thus providing an urgently needed alternative therapy for the treatment of cancers in the brain. Clinical investigation has shown that metabolism of RO7276389 is higher than predicted from animal studies, which as a consequence requires administration of comparably high doses of the drug multiple times over the day (WO2024105144A1). Our clinical pharmacokinetic and metabolite analysis have shown that one major metabolic pathway is the elimination of the active ingredient via formation of the following metabolite termed MIO (also termed RO7593626):

[0030] (MIO).

[0031] We have found that non-CYP450 oxidases like Aldehyde Oxidase (AO) and / or Xanthine Oxidase (XO) play an important role in the generation of MIO from the parent compound. The present invention relates to the design of a new deuterated compound of formula (lb), which surprisingly maintains paradox breaking activity and brain penetration potential, but is not the subject of rapid metabolism into MIO. The metabolic instability and formation of MIO metabolite in human liver cytosol was markedly lower for deuterated compound of formula (lb) compared with the non-deuterated parent drug. In consequence, the exposure of the deuterated parent compound (herein also referred as2H-API) is expected to be higher when compared to the non-deuterated form (herein also referred as1H-API) at the same dose. The higher exposure of active parent provides for an increased coverage at clinically relevant concentrations and for a more convenient dosage regime with a lower overall pill-burden and / or a lower administration frequency.

[0032] A quinazolinone core is also used for other BRAF inhibitors currently being developed for the treatment of cancer and these compounds may similarly be metabolized via AO / XO oxidation and the formation of metabolites that are oxidized at the same position as MIO. The present invention therefore also relates to some new deuterated analogues of known BRAF inhibitors and which may present similar advantages when compared to the corresponding undeuterated parent compounds.

[0033] WO202310371 Al for instance described a compound of formula (II):

[0034] The abovementioned compound (II) is also known as ARRY-440 or PF-07799933 and we found that the corresponding AO / XO related metabolite (herein referred to as II-M10*) is present in human liver cytosol (data not shown), suggesting that blocking this metabolic pathway by deuteration would also present similar advantages as described herein for compound (lb).

[0035] WO2024017294A1 for instance described a compound of formula (III):

[0036] WO2024017294A1 for instance also described a compound of formula (IV): Brief description of Figures:

[0037] Figure 1 shows the (metabolite : parent) ratios of RO7593626 (MIO metabolite) and RO7276389 ('H-API) based on pMolar concentration versus time after a single or multiple 800 mg BID or 1200 mg TID RO7276389 ('H-API) oral administration, respectively.

[0038] Figure 2 shows the turnover of 'H-API (Fig. 2A) compared with2H-API (Fig. 2B) in human liver cytosol and the formation of M10. The turnover of the parent and the generation of the M10 metabolite are markedly lower for2H-API when compared withJH-API.

[0039] Figure 3 discloses the 5-day viability of 'H-API compared with2H-API treated A375 cell assay. Both compounds show comparable efficacy and potency in this assay.

[0040] Figure 4 discloses the P-ERK inhibition curve induced by 'H-API vs.2H-API treated A375 (Fig. 4A) and HCT-116 (Fig. 4B) cells respectively. Both compounds show comparable paradox breaking activity in that they inhibit pERK pathway in V600E mutated BRAF cell line A375 without substantial activation of the pERK pathway in wild-type BRAF cell line HCT-116.

[0041] Figure 5 discloses the growth in tumor volume (mm3) in female NOD SCID mice bearing A375 tumours treated with either vehicle, 'H-API or2H-API at a concentration of 5mg / kg QD (Fig. 5 A) or 20mg / kg QD (Fig. 5B). 'H-API and2H-API show comparable tumor growth inhibition.

[0042] The term "pharmaceutically acceptable salt" refers to those salts of the compound of formula (I), (la), lb), Ic), (II-D), (III-D) or (IV-D) as applicable, which retain the biological effectiveness and properties of the free bases or free acids, which are not biologically or otherwise undesirable. The salts are formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like, in particular hydrochloric acid, and organic acids such as acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, maleic acid, malonic acid, succinic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p- toluenesulfonic acid, salicylic acid, N-acetylcystein and the like. In addition, these salts may be prepared by addition of an inorganic base or an organic base to the free acid. Salts derived from an inorganic base include, but are not limited to, the sodium, potassium, lithium, ammonium, calcium, magnesium salts and the like. Salts derived from organic bases include, but are not limited to salts of primary, secondary, and tertiary amines, substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins, such as isopropylamine, trimethylamine, diethylamine, triethylamine, tripropylamine, ethanolamine, lysine, arginine, N-ethylpiperidine, piperidine, polyimine resins and the like.

[0043] The compounds of the present invention may comprise an asymmetric center and where applicable can be present in the form of optically pure enantiomers or mixtures of enantiomers such as, for example, racemates.

[0044] According to the Cahn-Ingold-Prelog Convention the asymmetric carbon atom can be of the "R" or "S" configuration.

[0045] Also an embodiment of the present invention is the compound according to formula (I) as described herein or a pharmaceutically acceptable salt thereof, in particular the compound according to formula (I) as described herein, more particularly the compound of formula (la), (lb) or (Ic) as described herein.

[0046] The term “Ci-6-alkyl”, alone or in combination, denotes a monovalent linear or branched saturated hydrocarbon group of 1 to 6 carbon atoms. Examples of Ci-6-alkyl include methyl, ethyl, propyl, isopropyl, n-butyl, iso-butyl, sec-butyl, tert-butyl and pentyl. Particular Ci-6-alkyl groups are methyl, ethyl, propyl and n-butyl. More particular Ci-6-alkyl groups are methyl, ethyl and propyl.

[0047] The term “Cs-s-cycloalkyl”, alone or in combination, denotes a monovalent saturated monocyclic or bicyclic hydrocarbon group of 3 to 8 ring carbon atoms. Bicyclic means a ring system consisting of two saturated carbocycles having on or two carbon atoms in common. Examples of monocyclic Cs-s-cycloalkyl are cyclopropyl, cyclobutanyl, cyclopentyl, cyclohexyl or cycloheptyl. Particular monocyclic cycloalkyl groups are cyclopropyl, cyclopentyl and cyclohexyl.

[0048] The term “C3-8-cycloalkyl-Ci-6-alkyl”, alone or in combination, denotes an -Ci-6-alkyl group wherein one of the hydrogen atoms of the Ci-6-alkyl group has been replaced by an C3-8- cycloalkyl group. Examples of C3-8-cycloalkyl-Ci-6-alkyl include cyclopropylmethyl, cyclopropylethyl, cyclopropylbutyl, cyclobutylpropyl, 2-cyclopropylbutyl, cyclopentylbutyl, cyclohexylmethyl, cyclohexylethyl, bicyclo[4.1.0]heptanylmethyl, bicyclo[4.1.0]heptanylethyl, bicyclo[2.2.2]octanylmethyl and bicyclo[2.2.2]octanylethyl. A particular example of C3-8- cycloalkyl-Ci-6-alkyl is cyclopropylmethyl.

[0049] The term “halogen” and “halo”, alone or in combination, are used interchangeably herein and denote fluoro, chloro, bromo or iodo. Particular examples of halogen are chloro and fluoro. The term “Ci-6-haloalkyl”, alone or in combination, denotes an “alkyl” group, wherein at least one carbon atom bears a “halogen” group instead of a hydrogen substituent. Examples of Ci-6-haloalkyl include chloromethyl, chloroethyl, chloropropyl, chlorobutyl, chloropentyl, chlorohexyl, fluoromehtyl, fluoroethyl, fluoropropyl, fluorobutyl, fluoropentyl and fluorohexyl. Particular Ci-6-haloalkyl are fluoromethyl, 1 -fluoroethyl, 2-fluoroetyl, 1 -fluoropropyl, 2- fluoropropyl and 3 -fluoropropyl. A more particular Ci-6-haloalkyl is 2-fluoroethyl.

[0050] The term “heterocycloalkyl” denotes a monovalent saturated or partly unsaturated mono- or bicyclic ring system of 4 to 9 ring atoms, comprising 1, 2, or 3 ring heteroatoms selected from N, O and S, the remaining ring atoms being carbon. Bicyclic means consisting of two cycles having one or two ring atoms in common. Examples for monocyclic saturated heterocycloalkyl are 4,5-dihydro-oxazolyl, oxetanyl, azetidinyl, pyrrolidinyl, 2-oxo-pyrrolidin-3-yl, tetrahydrofuranyl, tetrahydrothiophenyl, pyrazolidinyl, imidazolidinyl, oxazolidinyl, isoxazolidinyl, thiazolidinyl, piperidinyl, tetrahydropyranyl, tetrahydrothiopyranyl, piperazinyl, morpholinyl, thiomorpholinyl, l,l-dioxo-thiomorpholin-4-yl, azepanyl, diazepanyl, homopiperazinyl, or oxazepanyl. Examples for bicyclic saturated heterocycloalkyl are oxabicyclo[2.2.1]heptanyl, oxaspiro[3.3]heptanyl, 8-aza-bicyclo[3.2.1]octyl, quinuclidinyl, 8- oxa-3-aza-bicyclo[3.2.1]octyl, 9-aza-bicyclo[3.3.1]nonyl, 3-oxa-9-aza-bicyclo[3.3.1]nonyl, 3- thia-9-aza-bicyclo[3.3.1]nonyl, 2-azaspiro[2.3]hexanyl and 2-azaspiro[3.3]heptanyl. Examples for partly unsaturated heterocycloalkyl are dihydrofuryl, imidazolinyl, dihydro-oxazolyl, tetrahydro-pyridinyl, or dihydropyranyl. Particular heterocycloalkyl are pyrrolidinyl, piperidinyl, azetidinyl, 2-azaspiro[2.3]hexanyl and 2-azaspiro[3.3]heptanyl.

[0051] , D

[0052] The structural element as described herein, refers to deuterium also known as the2H isotope of hydrogen.

[0053] The present invention relates thus generally to a novel compound of formula (I) wherein

[0054] D is deuterium; R1is selected from i) Ci-6-alkyl, and ii) Ci-6-haloalkyl;

[0055] R2is selected from iii) hydrogen, and iv) halogen;

[0056] X is selected from v) -NH-, and vi) -O-;

[0057] R3is selected from vii) H, viii) cyano, and ix) halogen;

[0058] R4is selected from x) hydrogen, and xi) halogen;

[0059] R5is selected from xii) cyano, and xiii) halogen;

[0060] R6is selected from xiv) NR7R8, and xv) CHR9R10;

[0061] R7is selected from xvi) Ci-6-alkyl, xvii) Cs-s-cycloalkyl, and xviii) C3-8-cycloalkyl-Ci-6-alkyl;

[0062] R8is selected from xix) Ci-6-alkyl, xx) C3-8-cycloalkyl, and xxi) C3-8-cycloalkyl-Ci-6-alkyl; or R7and R8together with the nitrogen atom to which they are attached form an heterocycloalkyl optionally substituted with R11;

[0063] R9is selected from xxii) Ci-6-alkyl, xxiii) C3-8-cycloalkyl, and xxiv) C3-8-cycloalkyl-Ci-6-alkyl;

[0064] R10is selected from xxv) Ci-6-alkyl, xxvi) C3-8-cycloalkyl, and xxvii) C3-8-cycloalkyl-Ci-6-alkyl; or R9and R10together with the carbon atom to which they are attached form a Cs-s- cycloalkyl optionally substituted with R11; and

[0065] R11is halogen; or a pharmaceutically acceptable salt thereof.

[0066] One embodiment of the present invention relates to the compound of formula (I) wherein

[0067] D is deuterium;

[0068] R1is methyl;

[0069] R2is selected from i) hydrogen, and ii) chloro;

[0070] X is selected from iii) -NH-, and iv) -O-;

[0071] R3is selected from v) H, vi) cyano, and vii) fluoro;

[0072] R4is selected from viii) hydrogen, and ix) fluoro;

[0073] R5is selected from x) cyano, and xi) chloro;

[0074] R6is NR7R8; R7and R8together with the nitrogen atom to which they are attached form a heterocycloalkyl optionally substituted with R11;

[0075] R11is fluoro; or a pharmaceutically acceptable salt thereof. Particular compounds of the compound of formula (I) are the compounds of formulas (la), lb), Ic), (II-D), (III-D) and (IV-D) as described herein.

[0076] One embodiment of the invention relates to a novel compound of formula (la) or a pharmaceutically acceptable salt thereof. One embodiment of the invention relates to a novel compound of formula (Ic) or a pharmaceutically acceptable salt thereof.

[0077] One particular embodiment of the invention relates to a novel compound of formula (lb) or a pharmaceutically acceptable salt thereof. Also an embodiment of the present invention is the compound according to formula (lb)

[0078] (Ib).

[0079] Also an embodiment of the present invention is a pharmaceutically acceptable salt of the compound of formula (lb)

[0080] (lb).

[0081] Also an embodiment of the present invention is a compound of formula (Bl) or a salt thereof.

[0082] Also an embodiment of the present invention is a compound of formula (MIO) or a salt thereof.

[0083] Also an embodiment of the present invention is a compound of formula (II-M10*)

[0084] (II-M10*).

[0085] Also an embodiment of the present invention is a compound of formula (III-M10*)

[0086] (III-M10*).

[0087] Also an embodiment of the present invention is a compound of formula (IV-M10*)

[0088] (IV-M10*).

[0089] Also an embodiment of the present invention is a compound of formula (II-D)

[0090] (II-D) or a pharmaceutically acceptable salt thereof.

[0091] Also an embodiment of the present invention is a compound of formula (III-D)

[0092] (III-D) or a pharmaceutically acceptable salt thereof.

[0093] Also an embodiment of the present invention is a compound of formula (IV-D)

[0094] (IV-D) or a pharmaceutically acceptable salt thereof.

[0095] Further particular compounds falling under the compound of formula (I) are selected from or a pharmaceutically acceptable salt thereof.

[0096] Processes for the manufacture of the compound of formula (I) as described herein are also an object of the invention.

[0097] The preparation of the compound of formula (I) of the present invention may be carried out in sequential or convergent synthetic routes. Syntheses of the invention are shown in the following general scheme. The skills required for carrying out the reactions and purifications of the resulting products are known to those skilled in the art.

[0098] In more detail, the compound of formula (I) can be manufactured by the methods given below, by the methods given in the examples or by analogous methods. Appropriate reaction conditions for the individual reaction steps are known to a person skilled in the art. The reaction sequence is not limited to the one displayed in schemes 1, however, depending on the starting materials and their respective reactivity the sequence of reaction steps can be freely altered. Starting materials are either commercially available or can be prepared by methods analogous to the methods given below, by methods described in references cited in the description or in the examples, or by methods known in the art.

[0099] The preparation of the compound of formula (lb) of the present invention may be carried out in sequential or convergent synthetic routes. Syntheses of the invention are shown in the following general scheme. The skills required for carrying out the reactions and purifications of the resulting products are known to those skilled in the art.

[0100] In more detail, the compound of formula (lb) can be manufactured by the methods given below, by the methods given in the examples or by analogous methods. Appropriate reaction conditions for the individual reaction steps are known to a person skilled in the art. The reaction sequence is not limited to the one displayed in schemes 1, however, depending on the starting materials and their respective reactivity the sequence of reaction steps can be freely altered. Starting materials are either commercially available or can be prepared by methods analogous to the methods given below, by methods described in references cited in the description or in the examples, or by methods known in the art.

[0101] Scheme 2

[0102] It will be appreciated that the compound of formula (lb) in this invention may be derivatised at functional groups to provide derivatives which are capable of conversion back to the parent compound in vivo.

[0103] Using the method described above in analogy and using general common knowledge, the person skilled in the art can also produce the deuterated compounds (la), (Ic), (II-D), (III-D) and (IV-D) from their known undeuterated analogues.

[0104] The invention thus also relates to a process for the preparation of a compound according to the invention, comprising the reaction of a compound of formula (Bl)

[0105] (i) with D2 in presence of a suitable base, a suitable catalyst and a suitable solvent; or

[0106] (ii) with borodeuteride in presence of cuprous iodide and a suitable solvent. The solvent in the reaction (i) can be for instance methano-di;

[0107] The base in the reaction (i) can be for instance triethylamine;

[0108] The catalyst in the reaction (i) can be for instance palladium on carbon;

[0109] The solvent in the reaction (ii) can be for instance methanol-d4;

[0110] Convenient conditions for the reaction (i) can be between around 0°C and around 40°C, particularly between around 10° and around 30 °C;

[0111] Conveniently, for the reaction (i) the solvent is methano-di, the base is triethylamine and the catalyst is palladium on carbon; wherein the reaction is maintained at room temperature for between around 1 hour and around 48 hours, in particular between around 2 hours and around 36 hours;

[0112] Convenient conditions for the reaction (ii) can be between around -40°C and around 20°C, particularly between around -10° and around 10 °C;

[0113] Conveniently, for the reaction (i) the solvent is methano-d4; wherein the reaction is maintained at around 0°C for between around 15 minutes and around 24 hours, in particular between around 30 minutes and around 4 hours.

[0114] The invention also relates to a compound according to the invention when manufactured according to a process of the invention.

[0115] The invention also relates in particular to:

[0116] A compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for use as therapeutically active substance;

[0117] A pharmaceutical composition comprising a compound of formula (I) and / or the corresponding intermediate (B2) as described herein, or acceptable salts thereof, and a therapeutically inert carrier;

[0118] A pharmaceutical composition comprising a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, and a therapeutically inert carrier;

[0119] A compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for use in the treatment or prophylaxis of cancer; A compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for use in the treatment or prophylaxis of thyroid cancer, colorectal cancer, brain cancer, melanoma or non-small cell lung cancer (NSCLC);

[0120] The use of a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for the treatment or prophylaxis of thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC;

[0121] The use of a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for the preparation of a medicament for the treatment or prophylaxis of thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC;

[0122] A method for the treatment of cancer, which method comprises administering an effective amount of a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof; and

[0123] A method for the treatment or prophylaxis of thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC, which method comprises administering an effective amount of a compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

[0124] A certain embodiment of the invention relates to the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for use in the therapeutic and / or prophylactic treatment of cancer, in particular BRAF mutation driven cancer, more particularly thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC.

[0125] A certain embodiment of the invention relates to the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the therapeutic and / or prophylactic treatment of cancer, in particular BRAF mutation driven cancer, more particularly thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC.

[0126] A certain embodiment of the invention relates to a pharmaceutical composition comprising the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0127] A certain embodiment of the invention relates to a method for the therapeutic and / or prophylactic treatment of cancer, in particular BRAF mutation driven cancer, more particularly thyroid cancer, colorectal cancer, brain cancer, melanoma or non-small cell lung cancer (NSCLC) by administering an effective amount of the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof. A certain embodiment of the invention relates to the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for the use as a medicament in therapeutic and / or prophylactic treatment of a patient with BRAF mutation driven cancers, in particular thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC, comprising determining the BRAF mutation status in said patient and then administering the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, to said patient.

[0128] A certain embodiment of the invention relates to the compound of formula (I) as described herein, or a pharmaceutically acceptable salt thereof, for the use as a medicament in therapeutic and / or prophylactic treatment of brain metastases.

[0129] A compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for use as therapeutically active substance;

[0130] A pharmaceutical composition comprising a compound of formula (lb) and / or (Bl) as described herein, or acceptable salts thereof, and a therapeutically inert carrier;

[0131] A pharmaceutical composition comprising a compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, and a therapeutically inert carrier;

[0132] A compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for use in the treatment or prophylaxis of cancer;

[0133] A compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for use in the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small-cell lung cancer (NSCLC);

[0134] The use of a compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small-cell lung cancer;

[0135] The use of a compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for the preparation of a medicament for the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small-cell lung cancer;

[0136] A method for the treatment of cancer, which method comprises administering an effective amount of a compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof; and

[0137] A method for the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small-cell lung cancer, which method comprises administering an effective amount of a compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

[0138] A certain embodiment of the invention relates to the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for use in the therapeutic treatment of cancer, in particular BRAF mutation driven cancer, more particularly BRAF mutation driven cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small-cell lung cancer.

[0139] A certain embodiment of the invention relates to the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the therapeutic treatment of cancer, in particular BRAF mutant driven cancer, more particularly thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC.

[0140] A certain embodiment of the invention relates to a pharmaceutical composition comprising the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.

[0141] A certain embodiment of the invention relates to a method for the therapeutic treatment of cancer, in particular BRAF mutation driven cancer, more particularly thyroid cancer, colorectal cancer, brain cancer, melanoma or non-small cell lung cancer (NSCLC) by administering an effective amount of the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, to a patient in need thereof.

[0142] A certain embodiment of the invention relates to the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for the use as a medicament in therapeutic treatment of a patient with BRAF mutant driven cancers, in particular thyroid cancer, colorectal cancer, brain cancer, melanoma or NSCLC, comprising determining the BRAF mutation status in said patient and then administering the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, to said patient.

[0143] A certain embodiment of the invention relates to the compound of formula (lb) as described herein, or a pharmaceutically acceptable salt thereof, for the use as a medicament in therapeutic treatment of brain metastases.

[0144] Furthermore, the invention includes all optical isomers, i.e. diastereoisomers, diastereomeric mixtures, racemic mixtures, all their corresponding enantiomers and / or tautomers as well as their solvates, wherever applicable, of the compound of formula (I). If desired, racemic mixtures of the compound of the invention may be separated so that the individual enantiomers are isolated. The separation can be carried out by methods well known in the art, such as the coupling of a racemic mixture of compounds to an enantiomerically pure compound to form a diastereomeric mixture, followed by separation of the individual diastereomers by standard methods, such as fractional crystallization or chromatography.

[0145] In the embodiments, where an optically pure enantiomer is provided, optically pure enantiomer means that the compound contains > 90 % of the desired isomer by weight, particularly > 95 % of the desired isomer by weight, or more particularly > 99 % of the desired isomer by weight, said weight percent based upon the total weight of the isomer of the compound. A chirally pure or chirally enriched compound may be prepared by chirally selective synthesis or by separation of enantiomers. The separation of enantiomers may be carried out on the final product or alternatively on a suitable intermediate.

[0146] Another embodiment of the invention provides a pharmaceutical composition or medicament containing a compound of the invention and a therapeutically inert carrier, diluent or excipient, as well as a method of using the compounds of the invention to prepare such composition and medicament. In one example, the compound of formula (I) may be formulated by mixing at ambient temperature at the appropriate pH, and at the desired degree of purity, with physiologically acceptable carriers, i.e., carriers that are non-toxic to recipients at the dosages and concentrations employed into a galenical administration form. The pH of the formulation depends mainly on the particular use and the concentration of compound, but preferably ranges anywhere from about 3 to about 8. In one example, a compound of formula (I) is formulated in an acetate buffer, at pH 5. In another embodiment, the compound of formula (I) is sterile. The compound may be stored, for example, as a solid or amorphous composition, as a lyophilized formulation or as an aqueous solution.

[0147] Compositions are formulated, dosed, and administered in a fashion consistent with good medical practice. Factors for consideration in this context include the particular disorder being treated, the particular mammal being treated, the clinical condition of the individual patient, the cause of the disorder, the site of delivery of the agent, the method of administration, the scheduling of administration, and other factors known to medical practitioners.

[0148] Also an embodiment of the present invention is the compound of formula (I) as described herein, when manufactured according to any one of the described processes. Table 1: The non-deuterated compound (1H) and the deuterated compound (2H) have comparable affinity for RAF kinases.

[0149] Table 1 shows that the deuterated compound (2H-API) and the non-deuterated compound (1H- API) have comparable affinity for RAF kinases. We have further confirmed that2H-API maintains the paradoxical properties of the non-deuterated compound (data shown in Figures 4A and 4B). In addition, we have also found that2H-API maintains the high brain penetration potential of1H-API with comparable permeability, human fu,p and P-gp values (data not shown).

[0150] Pharmaceutical Compositions

[0151] The compound of formula (I) or a pharmaceutically acceptable salt thereof can be used as a medicament (e.g. in the form of a pharmaceutical preparation). The pharmaceutical preparation can be administered internally, such as orally (e.g. in the form of tablets, coated tablets, dragees, hard and soft gelatin capsules, solutions, emulsions or suspensions), nasally (e.g. in the form of nasal sprays), rectally (e.g. in the form of suppositories) or topical ocularly (e.g. in the form of solutions, ointments, gels or water soluble polymeric inserts). However, the administration can also be effected parenterally, such as intramuscularly, intravenously, or intraocularly (e.g. in the form of sterile injection solutions).

[0152] The compound of formula (I) or a pharmaceutically acceptable salt thereof can be processed with pharmaceutically inert, inorganic or organic adjuvants for the production of tablets, coated tablets, dragees, hard gelatin capsules, injection solutions or topical formulations Lactose, corn starch or derivatives thereof, talc, stearic acid or its salts etc. can be used, for example, as such adjuvants for tablets, dragees and hard gelatin capsules. Suitable adjuvants for soft gelatin capsules, are, for example, vegetable oils, waxes, fats, semi-solid substances and liquid polyols, etc.

[0153] Suitable adjuvants for the production of solutions and syrups are, for example, water, polyols, saccharose, invert sugar, glucose, etc.

[0154] Suitable adjuvants for injection solutions are, for example, water, alcohols, polyols, glycerol, vegetable oils, etc.

[0155] Suitable adjuvants for suppositories are, for example, natural or hardened oils, waxes, fats, semi-solid or liquid polyols, etc.

[0156] Suitable adjuvants for topical ocular formulations are, for example, cyclodextrins, mannitol or many other carriers and excipients known in the art.

[0157] Moreover, the pharmaceutical preparations can contain preservatives, solubilizers, viscosity -increasing substances, stabilizers, wetting agents, emulsifiers, sweeteners, colorants, flavorants, salts for varying the osmotic pressure, buffers, masking agents or antioxidants. They can also contain still other therapeutically valuable substances.

[0158] The dosage can vary in wide limits and will, of course, be fitted to the individual requirements in each particular case. In general, in the case of oral administration a daily dosage of about 1.0 mg to 80 mg per kg body weight, preferably about 10 mg to 50 mg per kg body weight (e.g. about 3750 mg per person), divided into preferably 1-3 individual doses, which can consist, for example, of the same amounts, should it be appropriate.

[0159] The compound of formula (I) or a pharmaceutically acceptable salt thereof can be used as therapeutically active substance, e.g. in the form of a pharmaceutical preparation. The pharmaceutical preparation can be administered orally, e.g. in the form of tablets, coated tablets, dragees, hard and soft gelatin capsules, solutions, emulsions or suspensions. The administration can, however, also be effected rectally, e.g. in the form of suppositories, or parenterally, e.g. in the form of injection solutions.

[0160] The compound of formula (I) and the pharmaceutically acceptable salts thereof can be processed with a pharmaceutically inert, inorganic or organic carriers for the production of a pharmaceutical preparation. Lactose, com starch or derivatives thereof, talc, stearic acids or its salts and the like can be used, for example, as such carriers for tablets, coated tablets, dragees and hard gelatin capsules. Suitable carriers for soft gelatin capsules are, for example, vegetable oils, waxes, fats, semi-solid and liquid polyols and the like. Depending on the nature of the active substance no carriers are however usually required in the case of soft gelatin capsules. Suitable carriers for the production of solutions and syrups are, for example, water, polyols, glycerol, vegetable oil and the like. Suitable carriers for suppositories are, for example, natural or hardened oils, waxes, fats, semi-liquid or liquid polyols and the like.

[0161] The pharmaceutical preparation can, moreover, contain pharmaceutically acceptable auxiliary substances such as preservatives, solubilizers, stabilizers, wetting agents, emulsifiers, sweeteners, colorants, flavorants, salts for varying the osmotic pressure, buffers, masking agents or antioxidants. They can also contain still other therapeutically valuable substances.

[0162] Medicaments containing the compound of formula (I) or a pharmaceutically acceptable salt thereof and a therapeutically inert carrier are also provided by the present invention, as is a process for their production, which comprises bringing one or more compounds of formula (I) and / or pharmaceutically acceptable salts thereof and, if desired, one or more other therapeutically valuable substances into a galenical administration form together with one or more therapeutically inert carriers.

[0163] The dosage can vary within wide limits and will, of course, have to be adjusted to the individual requirements in each particular case. In the case of oral administration the dosage for adults can vary from about 25 mg to about 4000 mg per day of a compound of general formula (I) or of the corresponding amount of a pharmaceutically acceptable salt thereof. The daily dosage may be administered as single dose or in divided doses and, in addition, the upper limit can also be exceeded when this is found to be indicated.

[0164] The following examples illustrate the present invention without limiting it, but serve merely as representative thereof. The pharmaceutical preparations conveniently contain about 5-500 mg, particularly 50-200 mg, of a compound of formula (I), in particular of a compound of formula (lb). Examples of compositions according to the invention are:

[0165] Example A

[0166] Tablets of the following composition are manufactured in the usual manner:

[0167] Table 2: possible tablet composition

[0168] Manufacturing Procedure

[0169] 1. Mix ingredients 1, 2, 3 and 4 and granulate with purified water.

[0170] 2. Dry the granules at 50°C. 3. Pass the granules through suitable milling equipment.

[0171] 4. Add ingredient 5 and mix for three minutes; compress on a suitable press.

[0172] Example B-l

[0173] Capsules of the following composition are manufactured:

[0174] Table 3: possible capsule ingredient composition Manufacturing Procedure 1. Mix ingredients 1, 2 and 3 in a suitable mixer for 30 minutes.

[0175] 2. Add ingredients 4 and 5 and mix for 3 minutes.

[0176] 3. Fill into a suitable capsule.

[0177] The compound of formula (lb), lactose and com starch are firstly mixed in a mixer and then in a comminuting machine. The mixture is returned to the mixer; the talc is added thereto and mixed thoroughly. The mixture is filled by machine into suitable capsules, e.g. hard gelatin capsules.

[0178] Examples

[0179] Abbreviations

[0180] AO = aldehyde oxidase; API = active pharmaceutical ingredient; BID = twice per day; DCM = dichloromethane; DMSO = diemethyl sulfoxide; ESI = electrospray ionization; EtOAc = ethyl acetate; HMDS = hexamethyldisilane; HPLC = high-performance liquid chromatography; MS = mass spectrometry; RT = room temperature; PEG400 = polyethylene glycol 400; P-gp = P- glycoprotein; PG-Dx = post grouping day x; PK = pharmacokinetics; p.o. = per os (by mouth); QD = quaque die (once every day); RP-HPLC = reversed phase-HPLC; Solutol HS15 = polyethylene glycol 15 hydroxystearate; TBME = tert-butyl methyl ether; THF = tetrahydrofuran; TID = three times per day; TLC = thin-layer chromatography; UV = ultraviolet; XO = xanthine oxidase.

[0181] Example 1 - Pharmacokinetics of Metabolites:

[0182] Preliminary metabolite data of M10 were available for 24 human patients who received either 800 mg BID (12 patients) or 1200 mg TID (12 patients) RO7276389 ('H-API) as single agent at Cycle 1 Day 1. For M10, Tmax values were ranging from approximately 1.00-8.02 hours (with a median of 3.94 hours) with geometric mean (coefficient of variation [CV]) Cmax-values of 15.5 pg / mL (42.8%) and 21.9 pg / mL (31.6%) for the 800 mg BID and 1200 mg TID RO7276389 (JH-API) oral doses, respectively. Preliminary metabolite M10 PK data were also available after multiple dosing of either 800 mg BID (12 patients) or 1200 mg TID (10 patients) RO7276389 ('H-API) as single agent at Cycle 1 Day 15. The geometric mean (CV) Cmax of M10 was 16.2 pg / mL (44.1%) and 14.8 pg / mL (56.2%) for the 800 mg BID and 1200 mg TID RO7276389, respectively. Figure 1 shows the ratios of M10 metabolite and RO7276389 ('H-API) based on pMolar concentration versus time after a single or multiple 800 mg BID or 1200 mg TID RO7276389 oral administration suggesting linear and time-independent PK characteristics for the M10 metabolite as for the parent RO7276389. The preliminary MIO metabolite data indicate that the exposure of patients to this MIO metabolite appears to be substantial compared to that of parent drug.

[0183] Step 1 : Iodination reaction

[0184] (3A)-7V-[2-cyano-4-fluoro-3-(2-iodo-4-keto-3-methyl-quinazolin-6-yl)oxy-phenyl]-3-fluoro- pyrroli dine- 1 -sulfonami de

[0185] A solution of (3A)-7V-[2-cyano-4-fluoro-3-(3-methyl-4-oxo-quinazolin-6-yl)oxy-phenyl]-3- fluoro-pyrrolidine-1 -sulfonamide (6.00 g, 12.95 mmol, 1.00 eq.; method described in W02021116050A1) in dry THF (60 mL) was cooled to -70 °C (dry ice / acetone), and a solution of LiHMDS (IM in THF, 60 mL, 60 mmol, 4.63 eq.) was added dropwise over the course of 25 minutes. After addition, the mixture was stirred for another 35 minutes at -70 °C, then a solution of iodine (6.6 g, 26 mmol, 2.01 eq.) in dry THF (30 mL) was added dropwise over the course of 15 minutes. A brown suspension was formed, which was allowed to warm up to room temperature to give a clear brown solution. Analysis by RP-HPLC revealed a conversion of 99% after reaching room temperature. The reaction mixture was diluted with 2-MeTHF (250 mL), washed with IM citric acid (2 x 50 mL), brine (2 x 50 mL), saturated aqueous sodium bicarbonate solution (2 x 50 mL), 10% sodium thiosulfate solution (2 x 50 mL) and finally again with brine (50 mL). The organic layer was separated, dried over anhydrous sodium sulfate and concentrated to a slurry under reduced pressure and elevated temperature (50 °C). Coevaporation with absolute ethanol (2 x 100 mL) and concentration to about 60 mL total volume gave a beige suspension. The solid was filtered off, washed with a minimal amount of absolute ethanol, then dried under reduced pressure and elevated temperature (55 °C, 5 mbar). 6.83 g of the crude product was obtained as brown crystalline powder, purity 93% (RP-HPLC). For further purification, the crude product was dissolved in warm (55 °C) THF (70 mL), and concentrated to about half the volume. Methanol (analytical grade, 100 mL) was added, and the mixture was concentrated until crystallization started to begin. Another 50 mL of methanol (analytical grade) were added, and again ca. 30 mL of solvent was removed under reduced pressure to give a beige suspension. It was allowed to cool to room temperature, the precipitate filtered off and washed with methanol. The collected solid was dried under reduced pressure and elevated temperature (55 °C, 5 mbar) to furnish (3A)-7V-[2- cyano-4-fluoro-3-(2-iodo-4-keto- 3-methyl-quinazolin-6-yl)oxy-phenyl]-3-fluoro-pyrrolidine-l- sulfonamide (5.0 g, 8.11 mmol, 63%) as light-brown solid (purity 95%, RP-HPLC, detection at 225 nm).

[0186] 1H NMR (400 MHz, DMSO-d6): 8 = 10.47 - 10.18 (m, 1H), 7.86 (dd, J = 9.4, 10.2 Hz, 1H), 7.72 - 7.66 (m, 1H), 7.66 - 7.61 (m, 1H), 7.52 (dd, J = 4.0, 9.1 Hz, 1H), 7.32 (d, J = 3.0 Hz, 1H), 5.44 - 5.21 (m, 1H), 3.65 (s, 3H), 3.57 - 3.30 (m, 4H), 2.20 - 1.97 (m, 2H) ppm.

[0187] Step 2: Deutero-deiodination

[0188] (3A)-A-[2-cyano-4-fluoro-3-(2-deuterio-4-keto-3-methyl-quinazolin-6-yl)oxy-phenyl]-3-fluoro- pyrroli dine- 1 -sulfonami de

[0189] Method 1 :

[0190] To a solution of (3A)-7V-[2-cyano-4-fluoro-3-(2-iodo-4-keto-3-methyl-quinazolin-6-yl)oxy- phenyl]- 3 -fluoro-pyrrolidine-1 -sulfonamide (5.00 g, 8.11 mmol, 1.00 eq.) in methanol-di (100 mL) was added palladium on carbon (10% w / w, 1.00 g, 0.94 mmol, 0.12 eq.), followed by triethylamine (3.4 mL, 24.4 mmol, 3.00 eq.) at room temperature. A balloon containing D2 gas (1 atm) was attached, and the suspension was deuterogenated for 23 hours at room temperature under vigorous stirring. After the indicated time, analysis by RP-HPLC revealed a conversion of 79%. Another portion of palladium on carbon (10% w / w, 1.00 g, 0.94 mmol, 0.12 eq.) and methanol-di (25 mL) was added, and after continued deuteration for another 2 hours, full conversion was achieved. The catalyst was filtered off through a glass fiber filter, the residue rinsed with abs. ethanol, and the combined filtrates concentrated under reduced pressure (rotovap, 50 °C bath temperature). Repeated solvent exchange with absolute ethanol (3 x 50 mL) yielded a yellow solution, which after further concentration (to 20 - 25 mL) gave a white suspension. It was allowed to settle for 30 minutes at room temperature, then the precipitate was filtered off, washed with a minimal amount of absolute ethanol and / / -pentane and dried under reduced pressure (55 °C, 5 mbar). The desired product (3A)-7V-[2-cyano-4-fluoro-3-(2-deuterio-4-keto-3-methyl-quinazolin- 6-yl)oxy-phenyl]-3- fluoro-pyrrolidine-1 -sulfonamide was obtained as white crystals (2.20 g, 2.90 mmol, 58%), with a deuterium incorporation of 99.6% (MS) and an optical purity of 99.5% (chiral HPLC).

[0191] 'H NMR (400 MHz, DMSO-d6): 6 = 10.67 - 9.84 (m, 1H), 8.17 - 6.78 (m, 5H), 5.73 - 4.62 (m, 1H), 3.84 (s, 8H), 2.29 - 1.79 (m, 2H) ppm. MS: [M+l]+= 463 m / z

[0192] Method 2:

[0193] To a solution of (3A)-A-[2-cyano-4-fluoro-3-(2-iodo-4-keto-3-methyl-quinazolin-6-yl)oxy- phenyl]- 3 -fluoro-pyrrolidine-1 -sulfonamide (450 mg, 0.73 mmol, 1.00 eq.) in methanol-d4 (7.5 mL) was added cuprous iodide (145 mg, 0.76 mmol, 1.04 eq.), and the resulting suspension was cooled to 0 °C (mantle temperature). Sodium borodeuteride (90 mg, 2.14 mmol, 2.94 eq.) was added portionwise over the course of 15 minutes, and the stirring was continued for 30 minutes at 0 °C after completion of the addition. Analysis by RP-HPLC revealed a conversion of 89%. Another portion of methanol-d4 (0.5 mL) and sodium borodeuteride (45 mg, 1.08 mmol, 1.48 eq.) was added over the course of 5 min, and the stirring was continued for another 30 minutes at 0 °C, after which 98% conversion was detected (RP-HPLC). The mixture was diluted with ethyl acetate (100 mL) and washed subsequently with IM citric acid (15 mL), saturated aqueous sodium bicarbonate solution (15 mL) and brine (15 mL). The organic layer was separated and dried over anhydrous sodium sulfate, then concentrated under reduced pressure (50 °C) to yield the crude product as yellow foam. TLC analysis (silica, ethyl acetate, detection at 254 nm UV) revealed the presence of a slightly less polar impurity. Purification by flash column chromatography (40 g silicycle column, eluent: ethyl acetate, 30 mL / min flow rate, wet loading in ethyl acetate) gave the fractions of pure product, which were concentrated under reduced pressure (50 °C), taken up in a small amount of methanol and coevaporated with TBME. The resulting white suspension was filtered, the collected precipitate washed with TBME and dried under reduced pressure (50 °C, 5 mbar) to furnish the desired product (3A)-A-[2-cyano-4-fluoro-3-(2-deuterio-4-keto-3-methyl- quinazolin-6-yl)oxy-phenyl]-3-fluoro-pyrrolidine-l-sulfonamide as white crystals (140 mg, 0.30 mmol, 41%), with a deuterium incorporation of 98.7% (MS) and an optical purity of 99.8% (chiral HPLC).

[0194] Example 3 - Human liver cytosol assay

[0195] Human liver cytosol is a liver fraction that is richer in cytosolic enzymes such as AO compared with hepatocytes. Therefore, we compared the turnover of 'H-API versus2H-API and the formation of M10 metabolite in human liver cytosol. The results are shown in Figure 2. The turnover of RO7276389 ('H-API) and formation of M10 after 45 min incubation at 37 °C in human liver cytosol was markedly higher than for2H-API. We also found that selective deuteration of RO7276389 reduced conversion to M10 in this human liver cytosol assay. The % peak area ratio of metabolites was estimated by comparison of peak areas of MS ion intensities. This semi- quantitative approach is based on several assumptions (equimolar response of different analytes, no matrix effect, etc.) and consequently, it cannot be excluded that in some cases this semi- quantitative data will be different to data generated by structure independent radioactivity measurements or other validated bioanalytical methods using individual reference compounds.

[0196] Example 4 - Cell Viability Assays

[0197] To evaluate the cytotoxic activity of the tested compounds (results shown in Figure 3), cell viability was assessed using the CellTiter-Glo® 2.0 Luminescent Cell Viability assay (Promega #G9242). Cells were seeded in 96-well plates at a density of 3000 cells / well. 24 h later, cells were treated with 0.001-10 micromolar BRAF inhibitor for 120 h. For each compound, results are expressed as relative percent luminescence compared to controls. Cells were lysed by adding one volume of the CellTiter-Glo® reagent and incubated for 10 minutes on a shaker. Luminescence signal was measured using the PHERAstar FSX plate reader (Gain 3600). IC50s were calculated in Prism GraphPad Software Inc., San Diego, CA, by using the following curve fitting functions: “log(inhibitor) vs. response - Variable slope (four parameters)”

[0198] Example 5 - HTRF assay for P-ERK determination in A375 or HCT116 cells

[0199] A375 is a cellular cancer model expressing V600E mutated BRAF and HCT116 a cellular cancer model expressing WT BRAF. First generation BRAF inhibitors such as e.g. dabrafenib induce a paradox effect on tumour cells in that they inhibit the growth of V600E mutated BRAF cells (such as e.g. A375), while they activate growth in WT BRAF cells (such as e.g. HCT 116). ERK 1,2 phosphorylation (terminal member of the phosphorylation cascade of the MAPK pathway) is hereafter reported as main readout for the activation status of the MAPK pathway. Prior to the assay, A375 and HCT116 cell lines (originally obtained from ATCC and banked by the Roche repository) are maintained in DMEM no-phenol red medium (Gibco, cat# 21063029) supplemented with 10% fetal bovine serum (FBS) (Gibco, cat# 10270-106). Following compound treatment, P-ERK levels are determined by measuring FRET fluorescence signal induced by selective binding of 2 antibodies provided in the mentioned kit (Cisbio, cat# 64AERPEH) on ERK protein when phosphorylated at Thr202 / Tyr204. Briefly, 8000 cells / well in 12 pl media / well are plated in the 384-well plate (Greiner Bio-One, With Lid, HiBase, Low volume, cat# 784-080) and left overnight in the incubator (at 37 °C with 5% CO2-humidified atmosphere), the following day the plate is treated in duplicate with test compounds, dabrafenib and PLX8394 (the latter two as controls). Compound treatment was performed for 1 hour with an 11 -point dilution curve starting from the concentration of 10 micromolar. Then, 4pl of a 4X lysis buffer supplied with the kit are added to the wells, the plate is then centrifuged for 30 second (300 ref) and incubated on a plate shaker for 1 hour at RT.

[0200] At the end of the incubation 4pL / well of advanced P-ERK antibody solution (prepared according to manufacturer’s instruction) followed by 4pL / well of criptate P-ERK antibody solution (prepared according to manufacturer’s instruction) (Cisbio cat# 64AERPEH) are added to test wells.

[0201] In order to allow proper data normalization control wells non drug treated reported in the following table are always included in each plate (according to manufacturer’s instruction): p-ERK HTRF well compositions (pl):

[0202] The plate is then centrifuged at 300 ref for 30 second, sealed to prevent evaporation and incubated overnight in the dark at room temperature.

[0203] The plate is then analyzed and fluorescence emission value collected through a Pherastast FSX (BMG Labtech) apparatus at 665 and 620 nM.

[0204] The obtained fluorescence values are processed according to the formula Ratio=Signal(620nm) / Signal(625nm)* 10000 then the average of the ratio on the blank is subtracted to all values. Data are normalized in the case of A375 cells (BRAF inhibition) considering the average of the ratio (blank subtracted) derived by DMSO only treated cells as 100% and by considering the average of the ratio (blank subtracted) derived by lOpM dabrafenib treated cells as 0%. Mean of the normalized points are fitted with sigmoidal curve and IC50 determined. The results are shown in Figure 4 A.

[0205] Data are normalized in the case of HCT116 cells (BRAF activation,) considering the average of the ratio (blank subtracted) derived by DMSO only treated cells as 0% and by considering the average of the ratio (blank subtracted) derived by dabrafenib treated cells at the concentration which provides the highest signal as 100%. Individual points are fitted with either sigmoidal or bell shape curves, and the percentage of activation compared to maximum dabrafenib-mediated activation is determined. The EC50 is the concentration at which activation equal to 50% of the maximum achieved by dabrafenib is obtained. The results are shown in Figure 4B.

[0206] In case the activation does not reach 50% of the maximum achieved by dabrafenib, then the EC50 calculation is not applicable.

[0207] The Percentage of Maximum paradox inducing effect from dabrafenib is determined by evaluating the percentage at which the test compound induce its maximum P-ERK signal as percentage of the highest signal produced by dabrafenib within the dose range tested.

[0208] Compound concentrations that yielded 50% pERK inhibition (IC50) in A375 cells were calculated in Prism GraphPad Software Inc., San Diego, CA, by using the following curve fitting function: “log(inhibitor) vs. response, compounds variable slope (four parameters)” For HCT- 116 cells, pERKl / 2 signal intensity was plotted using the following curve fitting function: “Bellshaped, X is log(concentration)”.

[0209] Example 6 in vivo study in mice bearing A375 tumours

[0210] A375 tumor cells were maintained in vitro in DMEM medium supplemented with 10% fetal bovine serum and 1% Antibiotic- Antimycotic at 37 °C in an atmosphere of 5% CO2 in air. The tumor cells were routinely subcultured twice weekly, the cells growing in an exponential growth phase were harvested and counted for tumor inoculation.

[0211] The details of reagents are shown in Table 4:

[0212] Female NOD SCID mice (supplied by Zhejiang Vital River Laboratory Animal Technology Co, LTD) with an age of 6 to 8 weeks were subjected to A375 tumour inoculation. Each mouse was inoculated subcutaneously at the right flank with the tumor cells (7.5 x 106) in 0.1 mL of PBS for tumor development. The treatments were started when the geometric average tumor volume reached approximately 118 mm3.

[0213] The test article administration and the animal numbers in each group are shown in Table 5:

[0214] Note:

[0215] 1. N: animal number. 2. 'H-API and2H-API were formulated in “10% PEG400 and 10% Solutol HS15 in water”. Animals in vehicle group were dosed with “10% PEG400 and 10% Solutol HS15 in water”.

[0216] 3. The dosing of all compounds in mice bearing A375 tumors was started when the geometric mean tumor volume reached 118 mm3. The day of randomization was assigned to PG-DO and the treatment was started from PG-DO.

[0217] 4. On PG-D20, plasma was collected from 5 animals at 0 h (pre-dose), 1 h, 2 h, 4 h, 6 h, and 24 h in groups 2-5 for PK analysis.

[0218] Prior to the onset of drug treatment, mice were measured for tumor size in two dimensions using a caliper, and the tumor volume (mm3) was calculated using formula V = 0.5 a x b2where a and b are the long and short diameters of the tumor in mm, respectively. Mice were stratified and randomized into different treatment groups based on the tumor volume.

[0219] Tumor growth curves (GM ± SEM) are shown in Figure 5. No significant difference was observed for tumour growth inhibition in mice treated with 'H-API compared to2H-API. The pharmacokinetics of 'H-API were comparable to2H-API in this mouse model which can be attributed to the minor contribution of AO / XO metabolism in mice compared to humans.

Claims

1. -37-Claims1. A compound of formula (I)whereinD is deuterium;R1is selected from i) Ci-6-alkyl, and ii) Ci-6-haloalkyl;R2is selected from iii) hydrogen, and iv) halogen;X is selected from v) -NH-, and vi) -O-;R3is selected from vii) H, viii) cyano, and ix) halogen;R4is selected from x) hydrogen, and xi) halogen;-38-R5is selected from xii) cyano, and xiii) halogen;R6is selected from xiv) NR7R8, and xv) CHR9R10;R7is selected from xvi) Ci-6-alkyl, xvii) Cs-s-cycloalkyl, and xviii) C3-8-cycloalkyl-Ci-6-alkyl;R8is selected from xix) Ci-6-alkyl, xx) C3-8-cycloalkyl, and xxi) C3-8-cycloalkyl-Ci-6-alkyl; or R7and R8together with the nitrogen atom to which they are attached form an heterocycloalkyl optionally substituted with R11;R9is selected from xxii) Ci-6-alkyl, xxiii) C3-8-cycloalkyl, and xxiv) C3-8-cycloalkyl-Ci-6-alkyl;R10is selected from xxv) Ci-6-alkyl, xxvi) C3-8-cycloalkyl, and xxvii) C3-8-cycloalkyl-Ci-6-alkyl;or R9and R10together with the carbon atom to which they are attached form a C3-8- cycloalkyl optionally substituted with R11; andR11is halogen; or a pharmaceutically acceptable salt thereof.

2. A compound of formula (lb)or a pharmaceutically acceptable salt thereof.

3. The compound according to claim 1 or 2, wherein the compound is the compound of formula (lb).

4. The compound according to any one of claims 1 to 3, wherein the compound is (3R)-N-[2- cyano-4-fluoro-3-(2-deuterio-4-keto-3-methyl-quinazolin-6-yl)oxy-phenyl]-3-fluoro- pyrrolidine-l-sulfonamide.

5. A pharmaceutically acceptable salt of the compound of formula (lb)(lb).

6. A process for the preparation of a compound according to any one of claims 2 to 5, comprising the reaction of a compound of formula (Bl)with D2 in presence of a suitable base, a suitable catalyst and a suitable solvent; orwith borodeuteride in presence of cuprous iodide and a suitable solvent.

7. A compound of formula (Bl) or a salt thereof.

8. A compound of formulaor a salt thereof.

9. A compound according to any one of claims 2 to 5 when manufactured according to a process of claim 6.

10. A pharmaceutical composition comprising a compound according to any one of claims 1 to 5 and / or 7 and a therapeutically inert carrier.

11. A compound according to any one of claims 1 to 5 for use as therapeutically active substance.

12. A compound according to any one of claims 1 to 5 for use in the treatment of cancer.

13. A compound according to any one of claims 1 to 5 or a composition according to claim 10, for use in the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small cell lung cancer.

14. The use of a compound according to any one of claims 1 to 5 or a composition according to claim 10, for the treatment cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small cell lung cancer.

15. The use of a compound according to any one of claims 1 to 5 for the preparation of a medicament for the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small cell lung cancer.

16. A method for the treatment of cancer, which method comprises administering an effective amount of a compound as defined in any one of claims 1 to 5 to a patient in need thereof.

17. A method for the treatment of cancer selected from the group of thyroid cancer, colorectal cancer, brain cancer, melanoma and non-small cell lung cancer, which method comprises administering an effective amount of a compound as defined in any one of claims 1 to 5 to a patient in need thereof.

18. The invention as hereinbefore described.***

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