[0001] This invention relates to novel compounds having pharmacological activity, processes for their preparation, pharmaceutical compositions containing them and their use in the treatment of various disorders.

BACKGROUND

[0002] When a ribosome encounters a termination sequence during the process of messenger RNA (mRNA) translation no cognate transfer RNA (tRNA) can bind. This leads to a halt in protein production, release factor (RF) proteins binding to the empty site on the ribosome and conformational changes in the ribosome that lead to the break-up of the ribosomal complex. Although accurate, translational termination is not 100% effective and its efficiency depends on competition between the recognition of the stop codon by eRF1 , and the decoding of the stop codon by a near-cognate tRNA (i.e. a natural suppressor tRNA). The latter case leads to the suppression of translation termination, also called “read- through”, where an amino acid is incorporated in place of the stop (Dabrowski, M. et al., RNA Biology 2015, 12, 950-958).

[0003] If the halt in protein production occurs before the full length protein is made then the termination codon is a “premature” termination codon (PTC) and the unfinished protein may not be functional. In other words, if the terminating codon occurs before the full length protein is made then the termination codon is a “premature” termination codon (PTC) and the truncated protein may not be functional. Normal terminating stop codons are usually in a genetic and structural context that reinforces their effectiveness and are usually more resistant to read-through than PTCs arising through truncating mutations (Wangen and Green, 2020 Elife 9:e52611).

[0004] Over 1800 separate inherited genetic disorders have been identified where the causative mutation in a proportion of individuals is a nonsense mutation where an in-frame, premature termination codon (PTC) leads to a loss of gene function (Kellermayer, R. European Journal of Medical Genetics 2006, 49, 445-450). Over 9,600 PTC mutations have been published, representing around 10% of all reported gene mutations (Krawczak, M. et al., Human Mutation 2000, 15, 45-51 , Mort, M. et al., Human Mutation 2008, 29, 1037- 1047).

[0005] Nonsense mutations are involved in the pathology of a range of disorders; from polygenic pathologies such as cancer to monogenic diseases such as cystic fibrosis and Duchenne muscular dystrophy for which a PTC accounts for around 10% or 5-10% of all cases respectively (Keeling, K.M. & Bedwell, D.M., Journal of Molecular Medicine 2002, 80, 367-376, Kellermayer, R. European Journal of Medical Genetics 2006, 49, 445-450).

[0006] Whereas monogenetic diseases arise from either an inherited or de novo germline mutation, resulting in a PTC, in for example cancer, somatic mutations, can give rise to PTC mutations in tumour suppressor genes for example.

[0007] One of the surveillance systems used by the cell to remove abnormal mRNA transcripts that prematurely terminate translation is the non-sense mediated decay (NMD) pathway. It targets mRNA containing a PTC sequence more than ~50 base pairs upstream from the penultimate exon-exon junction (Maquat, L.E., Nature Reviews, Molecular Cell biology 2004, 5, 89-99).

[0008] The combination of premature termination of protein production and NMD of mRNA containing PTCs, can lead to almost total loss of protein production for mutant genes encoding a PTC.

[0009] Mechanisms by which translational read-though of PTC mutations may be achieved include but are not limited to: reducing the translational fidelity of the ribosome; reducing the efficiency of the translational termination machinery; inhibiting NMD; interfering with the regulation of gene transcription or translation or stabilising mRNA against NMD (Kellermayer, R European Journal of Medical Genetics 2006, 49, 445-450, Belgrader, P. J. C., and Maquat, L.E., Proc. Natl. Acad. Sci. USA 1993, 90, 482-486).

[0010] Agents which allow translational read-through of PTC mutations have the potential to treat any disease where the presence of a PTC in a gene is directly causative, a risk factor or an aggravating factor in disease.

[0011] Aminoglycosides (such as paromomycin, gentamicin, G418/geneticin) are a class of antibiotics able to reduce translational fidelity. They exert their antibacterial action through inhibition of prokaryotic protein synthesis, however, at sub-lethal doses they lower translational fidelity of ribosomes leading to an increased rate of mis-incorporation of amino acids and also to an increased rate of translational read-through of stop codons (Davies, J. & Gorini, L., Proc. Natl. Acad. Sci. USA 1964, 51 , 883-8, Weinstein, Proc. Natl. Acad. Sci. USA 1964, 52, 988-996).

[0012] Aminoglycosides have been shown to allow read-through of PTC mutations in the CFTR gene responsible for cystic fibrosis in vitro (Bedwell, D.M., et al., Nature Medicine 1997, 3, 1280-1284) and in vivo, including patients (Ming Du, et al., Journal of Molecular Medicine 2002, 80, 595-604, Clancy, J.P. et al., Am J Respir Crit Care Med 2001 , 163, 1683- 1692, Wilschanski, M., et al., The New England Journal of Medicine 2003, 139, 1433- 1441).

[0013] In an animal model of Duchenne muscular dystrophy (DMD), 7 of 8 male mdx mice treated with 34 mg/kg gentamicin showed increased protection from contractile injury and 10-20% restoration of wild-type dystrophin levels (Barton-Davis, E.R., et al., Journal of Clinical Investigation 1999, 104, 375-381).

[0014] Gentamicin treatment induced functional type VII collagen in recessive dystrophic epidermolysis bullosa patients (Woodley, D. T. et al. Journal of Clinical Investigation 2017, 127, 3028-3038) with in vitro evidence supporting a read-through mechanism (Has, C. et al. Journal of Investigative Dermatology 2022, 142, 1227-1230).

[0015] The read-through of genes containing PTC mutations relevant to cystinosis, Hurler syndrome, inherited blindness and cancer genes have all been demonstrated following aminoglycoside treatment (Helip- Wooley, A., et al., Molecular Genetics and Metabolism 2002, 75, 128-133, Keeling, K.M., et al., Human Molecular Genetics 2001 , 10, 291-299, Moosajee, M.K., et al., Human Molecular Genetics 2008,17, 3987-4000). However, aminoglycosides are not suitable as PTC read-through agents due to the severe toxicity observed with high doses or repeated treatments

[0016] WO2021228945A1 describes compounds, including TLN486 (TLN68), for use in preventing and/or treating a disease caused by a nonsense mutation. [0017] Certain compounds of the present invention are suitable for use in treating diseases/conditions which are associated with PTC mutations. Certain compounds of the invention show more potent read-through activity when compared to TLN468.

[0018] A structurally novel class of compounds has now been found which provides PTC read-through agents.

BRIEF SUMMARY OF THE DISCLOSURE

[0019] In accordance with a first aspect, the present invention provides a compound of formula (I), or a tautomeric form thereof, or a pharmaceutically acceptable salt or N-oxide thereof: wherein

R ¹ is independently selected from H, cyano, C1-C4-haloalkyl, C0-Cs-alkylene-R ^1a , C2-C6- alkylene-R ^{1 b} , C(O)R ^1c , S(O)R ^1c , S(O) ₂ R ^1c , and C1-C ₆ -alkylene-R ^1d ;

R ^1a is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl, 3- to 8- membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein where R ^1a is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, R ^1a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^1a is phenyl or heteroaryl, R ^1a is optionally substituted with from 1 to 5 R ⁹ groups;

R ^{1 b} is independently selected from NR ^6a R ^7a and OR ⁶ ;

R ^1c is independently selected from C1-Cs alkyl, C0-C4-alkylene-R ^1a and C0-C4-alkylene- NR ^6a R ^6a ;

R ^1d is independently selected from C(O)NR ^6a R ^6a , SO2NR ^6a R ^6a and CO2R ⁶ ;

R ² is independently selected from cyano, C1-C4-haloalkyl, C0-C6-alkylene-R ^2a , C2-C6- alkylene-R ^2b , C(O)R ^2c , S(O)R ^2c , S(O) ₂ R ^2c , and C1-C ₆ -alkylene-R ^2d ;

R ^2a is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl, 3- to 8- membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl and 5-, or 6-membered heteroaryl; wherein where R ^2a is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, R ^2a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^2a is phenyl or heteroaryl, R ^2a is optionally substituted with from 1 to 5 R ⁹ groups;

R ^2b is independently selected from NR ^6a R ^7a , and OR ⁶ ;

R ^2c is independently selected from C1-C6 alkyl, C0-C4-alkylene-R ^2a and C0-C4-alkylene- NR ^6a R ^6a ;

R ^2d is independently selected from C(O)NR ^6a R ^6a , SO2NR ^6a R ^6a and CO2R ⁶ ;

R ³ is independently selected from H and C1-C4 alkyl; or wherein R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups;

R ⁴ is independently selected from H, cyano, nitro, OR ^6b , NR ⁶ R ⁷ , CO2R ⁶ , C(O)R ⁶ , C(O)NR ⁶ R ⁶ , C1-C4-alkyl, C2-C4-alkenyl, C2-C4-alkynyl, C1-C4-haloalkyl, and cyclopropyl;

R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OH, OR ^6b , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , S(O) ₂ NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , C(O)NR ⁶ R ⁶ , C1-C ₄ -alkyl, C ₂ -C ₄ -alkenyl, C2-C4-alkynyl, C1-C4-haloalkyl, and cyclopropyl;

R ⁶ and R ^6a are each independently at each occurrence selected from H and C1-C4 alkyl; or two R ^6a groups, together with the nitrogen atom to which they are attached, form a 5- to 8- membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups;

R ^6b is independently at each occurrence selected from C1-C4 alkyl;

R ⁷ and R ^7a are each independently at each occurrence selected from H, C1-C4 alkyl and C(O)-C1-C ₄ -alkyl; or R ^6a and R ^7a , together with the nitrogen atom to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups;

R ⁸ is independently at each occurrence selected from =0, =S, halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , SO ₂ NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , CONR ⁶ R ⁶ , C1-C ₄ -alkyl, C2-C4- alkenyl, C2-C4-alkynyl, C1-C4-haloalkyl, cyclopropyl, C1-C3-alkylene-NR ⁶ R ⁷ , and C0-C3- alkylene-phenyl;

R ⁹ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , SO ₂ NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , CONR ⁶ R ⁶ , C1-C ₄ -alkyl, C ₂ -C ₄ -alkenyl, C2-C4- alkynyl, C1-C4-haloalkyl and cyclopropyl; m and x are each independently selected from 0 and 1 ; n is independently an integer selected from 0, 1 , 2, 3, and 4; wherein any of the aforementioned alkyl, alkylene, alkenyl, or cyclopropyl groups is optionally substituted, where chemically possible, by 1 to 5 substituents which are each independently at each occurrence selected from the group consisting of: C1-C4-alkyl, oxo, halo, nitro, cyano, NR ^a R ^b , OR ^a , SR ^a , CO ₂ R ^a , C(O)R ^a , CONR ^a R ^a , S(O)R ^a , and S(O) ₂ R ^a ; wherein R ^a is independently at each occurrence selected from H, and C1-C4-alkyl; and R ^b is independently at each occurrence selected from H, C1-C4-alkyl, C(O)-C1-C4-alkyl and S(O) ₂ - C1-C4-alkyl.

[0020] It may be that the compound according to formula (I) is not a compound selected from List A: List A:

5 [0021] It may be that the compound according to formula (I) is not a compound selected from List B: tautomer thereof.

[0022] It may be that the compound according to formula (I) is not a compound selected from List A and List B.

[0023] Compounds according to formula (I) may be able to tautomerise, depending on the identity of the substituent groups, to give alternative tautomeric forms of the compound depicted above. It may be that compounds of formula (I) are a mixture of these tautomeric forms. Said tautomeric forms may be interconverting in any given sample. The relative proportion of tautomeric forms in a given sample will be determined by the position of an equilibrium between those tautomeric forms under the conditions. The position of the equilibrium, and therefore the extent to which each tautomeric form of the compound is present, may be determined by factors including, but not limited to, identity of the substituent groups, temperature, pH, and/or the solvent (where compound of formula (I) is in solution).

[0024] In an embodiment, the compound of formula (I) is a compound of formula (II), or a tautomeric form thereof: wherein R ¹ , R ² , R ³ , R ⁴ , R ⁵ , and n are as defined for formula (I).

[0025] In an embodiment, the compound of formula (I) is a compound of formula (III), or a tautomeric form thereof: wherein r is selected from 2, 3, or 4;

R ¹ is independently selected from H, C(O)R ^1c , and S(O)2 ^1c ; and

R ³ , R ⁴ , R ⁵ , R ^6a , R ^7a , and n are as defined for formula (I).

[0026] In an embodiment, the compound of formula (I) is a compound of formula (IV), or a tautomeric form thereof: wherein R ^1c , R ² , R ³ , R ⁴ , R ⁵ , and n are as defined for formula (I).

[0027] In an embodiment, the compound of formula (I) is a compound of formula (V), or a tautomeric form thereof: wherein Ring A is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl, 3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6- membered heteroaryl; wherein Ring A is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups; p is selected from 0, 1 , 2, or 3;

R ¹ is independently selected from H, C(O)R ^1c , and S(O)2R ^1c ; and

R ³ , R ⁴ , R ⁵ , and n are as defined for formula (I).

[0028] In an embodiment, the compound of formula (I) is a compound of formula (VI), or a tautomeric form thereof: wherein Ring A is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl, 3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6- membered heteroaryl; wherein Ring A is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups; q is selected from 0, 1 , or 2; and R ¹ is independently selected from H, C(O)R ^1c , and S(O)2R ^1c

[0029] R ³ , R ⁴ , R ⁵ , and n are as defined for formula (I). In an embodiment, the compound of formula (I) is a compound of formula (VII), or a tautomeric form thereof: wherein R ² , R ³ , R ⁴ , R ⁵ , and n are as defined for formula (I).

[0030] In an embodiment, the compound of formula (I) is a compound of formula (VIII), or a tautomeric form thereof: wherein R ¹ is independently selected from H, C(O)R ^1c , and S(O)2 ^1c ; and

R ² and R ³ are as defined for formula (I).

[0031] In an embodiment, the compound of formula (I) is a compound of formula (IX), or a tautomeric form thereof: wherein R ² and R ³ are as defined for formula (I). [0032] In an embodiment, the compound of formula (I) is a compound of formula (X): wherein r is selected from 2, 3, or 4;

R ¹ is independently selected from H, C(O)R ^1c , and S(O)2R ^1c ; and R ³ , R ^6a , and R ^7a are as defined for formula (I).

[0033] In an embodiment, the compound of formula (I) is a compound of formula (XI), or a tautomeric form thereof: wherein R ^1c , R ² , and R ³ are as defined for formula (I). [0034] In an embodiment, the compound of formula (I) is a compound of formula (XII) or a tautomeric form thereof: wherein Ring A is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl,

3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6- membered heteroaryl; wherein Ring A is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups; p is selected from 0, 1 , 2, or 3;

R ¹ is independently selected from H, C(O)R ^1c , and S(O) ₂ R ^1c ; and

R ³ is as defined for formula (I).

[0035] In an embodiment, the compound of formula (I) is a compound of formula (XIII) or a tautomeric form thereof: wherein Ring A is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, phenyl, 3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6- membered heteroaryl; wherein Ring A is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups; q is selected from 0, 1 , or 2;

R ¹ is independently selected from H, C(O)R ^1c , and S(O)2R ^1c ; and

R ³ is as defined for formula (I).

[0036] The following embodiments apply to compounds of any of formulae (l)-(XI 11) . These embodiments are independent and interchangeable. Any one embodiment may be combined with any other embodiment, where chemically allowed. In other words, any of the features described in the following embodiments may (where chemically allowable) may be combined with the features described in one or more other embodiments. In particular, where a compound is exemplified or illustrated in this specification, any two or more of the embodiments listed below, expressed at any level of generality, which encompass that compound may be combined to provide a further embodiment which forms part of the present disclosure.

[0037] In an embodiment, R ¹ is independently selected from H, cyano, C1-C4-haloalkyl, C0- C ₄ -alkylene-R ^1a , C ₂ -C ₄ -alkylene-R ^{1 b} , C(O)R ^1c , S(O)R ^1c , S(O) ₂ R ^1c , and C1-C ₄ -alkylene-R ^1d In an embodiment, R ¹ is independently selected from H, cyano, C0-C6-alkylene-R ^1a , C ₂ -C6- alkylene-R ^{1 b} , and C(O)R ^1c . In an embodiment, R ¹ is independently selected from H, cyano, C0-C4-alkylene-R ^1a , C2-C6-alkylene-R ^{1 b} , and C(O)R ^1c . In an embodiment, R ¹ is independently selected from H, cyano, C0-C4-alkylene-R ^1a , C2-C4-alkylene-R ^{1 b} , and C(O)R ^1c . In an embodiment, R ¹ is independently selected from H, cyano, and C(O)R ^1c . In an embodiment, R ¹ is independently selected from H and C(O)R ^1c . In an embodiment, R ¹ is H. In an embodiment, R ¹ is cyano. In an embodiment, R ¹ is C(O)R ^1c . In an embodiment, R ¹ is independently selected from R ¹ is C(O)R ^1c , S(O)R ^1c and S(O)2R ^1c In an embodiment, R ¹ is S(O) ₂ R ^1C

[0038] It may be that R ¹ contains at least 1 nitrogen atom. It may be that R ¹ contains 1 , 2, or 3 nitrogen atoms. It may be that R ¹ contains 1 nitrogen atom. It may be that R ¹ contains 2 nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least one nitrogen atom is an amine. It may be that R ¹ comprises 1 , 2, or 3 amine groups, optionally 1 amine group.

[0039] In an embodiment, R ^1a is independently selected from 3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein where R ^1a is heterocycloalkyl or heterocycloalkenyl, R ^1a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^1a is heteroaryl, R ^1a is optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl, heterocycloalkenyl, or heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine. It may be that the heterocycloalkyl, heterocycloalkenyl, or heteroaryl group comprises 1 , 2, or 3 amine groups, optionally 1 amine group.

[0040] In an embodiment, R ^1a is independently selected from 3- to 8-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; wherein where R ^1a is heterocycloalkyl, R ^1a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^1a is heteroaryl, R ^1a is optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl or heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0041] In an embodiment, R ^1a is 3- to 8-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^1a is 5- or 6-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0042] In an embodiment, R ^1a is 5- or 6-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^1a is 5-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. It may be that the heterocycloalkyl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is basic. It may be that the at least 1 nitrogen atom is an amine.

[0043] In an embodiment, R ^1a is 5-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. In an embodiment, R ^1a is 6-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0044] It may be that, where a heterocycloalkyl, heterocycloalkenyl, or heteroaryl group contains a nitrogen atom, said nitrogen atom may be a ring atom, i.e. a component of the heterocycloalkyl, heterocycloalkenyl, or heteroaryl ring, or a substituent on the heterocycloalkyl, heterocycloalkenyl, or heteroaryl ring.

[0045] In an embodiment, R ^1a is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, and phenyl, wherein where R ^1a is cycloalkyl, or cycloalkenyl, R ^1a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^1a is phenyl, R ^1a is optionally substituted with from 1 to 5 R ⁹ groups.

[0046] In an embodiment, R ^1a is C3-C8 cycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^1a is C4-C6 cycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^1a is phenyl, optionally substituted with from 1 to 5 R ⁹ groups.

[0047] In an embodiment, R ^{1 b} is NR ^6a R ^7a . In an embodiment, R ^{1 b} is OR ⁶ .

[0048] In an embodiment, R ^1c is independently selected from C1-C4 alkyl, C0-C2-alkylene-

R ^1a and C0-C2-alkylene-NR ^6a R ^6a . In an embodiment, R ^1c is independently selected from C1- C4 alkyl and C0-C2-alkylene-R ^1a . In an embodiment, R ^1c is NR ^6a R ^6a .

[0049] In an embodiment, R ^1c is C1-C4 alkyl. In an embodiment, R ^1c is C1-C4 alkyl substituted with 1 to 5 halo substituents. In an embodiment, R ^1c is C1-C2 alkyl substituted with 1 to 5 halo substituents.

[0050] In an embodiment, R ^1c is C0-alkylene-R ^1a . In an embodiment, R ^1c is independently selected from C3-C8 cycloalkyl, phenyl, 3- to 8-membered heterocycloalkyl, and 5- or 6- membered heteroaryl; wherein where R ^1c is C3-C8 cycloalkyl, or heterocycloalkyl, R ^1c is optionally substituted with from 1 to 4 R ⁸ groups and where R ^1c is phenyl or heteroaryl, R ^1c is optionally substituted with from 1 to 5 R ⁹ groups.

[0051] In an embodiment, R ^1c is independently selected from C3-C8 cycloalkyl optionally substituted with from 1 to 4 R ⁸ groups, and phenyl optionally substituted with from 1 to 5 R ⁹ groups. [0052] In embodiments where R ^1d is CO2R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C2 alkyl.

[0053] In an embodiment, R ^1d is independently selected from C(O)NR ^6a R ^6a and SO ₂ NR ^6a R ^6a .

[0054] In an embodiment, R ¹ is selected from:

[0055] In an embodiment, R ² is independently selected from cyano, C1-C4-haloalkyl, C0- C ₄ -alkylene-R ^2a , C ₂ -C ₄ -alkylene-R ^2b , C(O)R ^2c , S(O)R ^2c , S(O) ₂ R ^2c , and C1-C ₄ -alkylene-R ^2d In an embodiment, R ² is independently selected from cyano, C0-C4-alkylene-R ^2a , C2-C4- alkylene-R ^2b , and C(O)R ^2c . In an embodiment, R ² is independently selected from C0-C4- alkylene-R ^2a , C2-C4-alkylene-R ^2b and C1-C4-alkylene-R ^2d .

[0056] In an embodiment, R ² is C0-C4-alkylene-R ^2a . In an embodiment, R ² is C1-C4- alkylene-R ^2a . In an embodiment, R ² is C3-alkylene-R ^2a . In an embodiment, R ² is C2-alkylene- R ^2a . In an embodiment, R ² is C1-alkylene-R ^2a . In an embodiment, R ² is C0-alkylene-R ^2a , i.e. R ² is R ^2a .

[0057] In an embodiment, R ² is C2-C4-alkylene-R ^2b . In an embodiment, R ² is C2-alkylene- R ^2b . In an embodiment, R ² is C3-alkylene-R ^2b . In an embodiment, R ² is C4-alkylene-R ^2b .

[0058] In an embodiment, R ² is C1-C4-alkylene-R ^2d . In an embodiment, R ² is C4-alkylene- R ^2d . In an embodiment, R ² is C3-alkylene-R ^2d . In an embodiment, R ² is C2-alkylene-R ^2d . In an embodiment, R ² is C1-alkylene-R ^2d .

[0059] It may be that R ² contains at least 1 nitrogen atom. It may be that R ² contains 1 , 2, or 3 nitrogen atoms. It may be that R ² contains 1 nitrogen atom. It may be that R ² contains 2 nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least one nitrogen atom is an amine. It may be that R ² comprises 1 , 2, or 3 amine groups, optionally 1 amine group.

[0060] R ^2a is independently selected from 3- to 8-membered heterocycloalkyl, 5- to 8- membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein where R ^2a is heterocycloalkyl or heterocycloalkenyl, R ^2a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^2a is heteroaryl, R ^2a is optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl, heterocycloalkenyl, or heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine. It may be that the heterocycloalkyl, heterocycloalkenyl, or heteroaryl group comprises 1 , 2, or 3 amine groups, optionally 1 amine group.

[0061] In an embodiment, R ^2a is independently selected from 3- to 8-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; wherein where R ^2a is heterocycloalkyl, R ^2a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^2a is heteroaryl, R ^2a is optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl or heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is basic. It may be that the at least 1 nitrogen atom is an amine.

[0062] In an embodiment, R ^2a is 3- to 8-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^2a is 5- or 6-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heterocycloalkyl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0063] In an embodiment, R ^2a is 5- or 6-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^2a is 5-membered heterocycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. It may be that the heterocycloalkyl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0064] In an embodiment, R ^2a is 5-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. In an embodiment, R ^2a is 6-membered heteroaryl, optionally substituted with from 1 to 5 R ⁹ groups. It may be that the heteroaryl contains at least 1 (e.g. 1 , 2, or 3) nitrogen atoms. It may be that the at least 1 nitrogen atom is a basic nitrogen. It may be that the at least 1 nitrogen atom is an amine.

[0065] It may be that, where a heterocycloalkyl, heterocycloalkenyl, or heteroaryl group contains a nitrogen atom, said nitrogen atom may be a ring atom, i.e. a component of the heterocycloalkyl, heterocycloalkenyl, or heteroaryl ring, or a substituent on the heterocycloalkyl, heterocycloalkenyl, or heteroaryl ring.

[0066] In an embodiment, R ^2a is independently selected from C3-C8 cycloalkyl, C5-C8 cycloalkenyl, and phenyl, wherein where R ^2a is cycloalkyl, or cycloalkenyl, R ^2a is optionally substituted with from 1 to 4 R ⁸ groups and where R ^2a is phenyl, R ^2a is optionally substituted with from 1 to 5 R ⁹ groups.

[0067] In an embodiment, R ^2a is C3-C8 cycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^2a is C4-C6 cycloalkyl, optionally substituted with from 1 to 4 R ⁸ groups. In an embodiment, R ^2a is phenyl, optionally substituted with from 1 to 5 R ⁹ groups.

[0068] In an embodiment, R ^2b is independently NR ^6a R ^7a . In an embodiment, R ^2b is independently OR ⁶ .

[0069] In an embodiment, R ^2c is independently selected from C0-C4-alkylene-R ^2a and C0- C4-alkylene-NR ^6a R ^6a . In an embodiment, R ^2c is independently selected from C0-C2-alkylene- R ^2a and C0-C3-alkylene-NR ^6a R ^6a .

[0070] In an embodiment, R ^2d is independently C(O)NR ^6a R ^6a . In an embodiment, R ^2d is independently SO2NR ^6a R ^6a .

[0071] In embodiments where R ^2d is CO2R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C2 alkyl.

[0072] In an embodiment, R ² is selected from:

[0073] In an embodiment, R ³ is independently selected from H and C1-C2 alkyl. In an embodiment R ³ is H. In an embodiment, R ³ is methyl.

[0074] In an embodiment, R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4

R ⁸ groups. In an embodiment, R ² and R ³ , together with the atom or atoms to which they are attached, form a 5-membered heterocycloalkyl ring, optionally substituted with 1 to 4 R ⁸ groups. In an embodiment, R ² and R ³ , together with the atom or atoms to which they are attached, form a 6-membered heterocycloalkyl ring, optionally substituted with 1 to 4 R ⁸ groups. [0075] In an embodiment, m is 0, and R ² and R ³ , together with the nitrogen atom to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups.

[0076] In an embodiment, m is 0, and R ² and R ³ , together with the nitrogen atom to which they are attached, form a 5-membered heterocycloalkyl ring, optionally substituted with 1 to 4 R ⁸ groups. In an embodiment, m is 0, and R ² and R ³ , together with the nitrogen atom to which they are attached, form a 6-membered heterocycloalkyl ring, optionally substituted with 1 to 4 R ⁸ groups.

[0077] In an embodiment, m is 0, and R ² and R ³ , together with the nitrogen atom to which they are attached, form a structure selected from: ; optionally substituted with 1 to 4 R ⁸ groups.

[0078] In an embodiment, m is 0, and R ² and R ³ , together with the nitrogen atom to which they are attached, form a structure selected from:

[0079] In an embodiment, R ¹ is independently selected from H, cyano, C(O)R ^1c , S(O)R ^1c and S(O)2R ^1C and R ² is independently selected from C0-C4-alkylene-R ^2a , C(O)R ^2a , C2-C6-R ^2b or R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8- membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups.

[0080] In an embodiment, R ¹ is independently selected from H, cyano, C(O)R ^1c , S(O)R ^1c and S(O)2R ^1C and R ² is independently selected from C0-C4-alkylene-R ^2a , C(O)R ^2a , C2-C4-R ^2b or R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8- membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups.

[0081] In an embodiment R ¹ is independently selected from H, cyano, C(O)R ^1c , S(O)R ^1c and S(O)2R ^1c and R ² is independently selected from C0-C4-alkylene-R ^2a , C2-C6-alkylene-R ^2b , and C(0)-C0-C4-alkylene-R ^2a or R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups.

[0082] In an embodiment R ¹ is independently selected from H, cyano, C(O)R ^1c , S(O)R ^1c and S(O)2R ^1c and R ² is independently selected from C0-C4-alkylene-R ^2a , C2-C4-alkylene-R ^2b , and C(0)-C0-C4-alkylene-R ^2a or R ² and R ³ , together with the atom or atoms to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally substituted with 1 to 4 R ⁸ groups.

[0083] In an embodiment R ¹ is independently selected from H, cyano, C1-C4-haloalkyl, C0- C4-alkylene-R ^1a , C2-C6-alkylene-R ^{1 b} , C(O)R ^1c , S(O)R ^1c and S(O)2R ^1c and R ² is independently selected from C0-C4-alkylene-R ^2a , C2-C6-alkylene-R ^2b and C1-C4-alkylene-R ^2d .

[0084] In an embodiment R ¹ is independently selected from H, cyano, C1-C4-haloalkyl, C0- C4-alkylene-R ^1a , C2-C4-alkylene-R ^{1 b} , C(O)R ^1c , S(O)R ^1c and S(O)2R ^1c and R ² is independently selected from C0-C4-alkylene-R ^2a , C2-C4-alkylene-R ^2b and C1-C4-alkylene-R ^2d .

[0085] In an embodiment, R ⁴ is independently selected from H, cyano, nitro, OR ^6b , NR ⁶ R ⁷ , C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl. In an embodiment, R ⁴ is independently selected from H, cyano, nitro, NR ⁶ R ⁷ , C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl. In an embodiment, R ⁴ is independently selected from H, C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl. In an embodiment, R ⁴ is independently selected from H, and C1-C4-alkyl. In an embodiment, R ⁴ is independently selected from C1-C4-alkyl and OR ^6b . In an embodiment, R ⁴ is C1-C2-alkyl, i.e. methyl or ethyl. In an embodiment, R ⁴ is C1-alkyl, i.e. methyl. In an embodiment, R ⁴ is H.

[0086] In embodiments where R ⁴ is CO2R ⁶ or C(O)R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C2 alkyl.

[0087] In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , S(O) ₂ NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , C(O)NR ⁶ R ⁶ , C1-C4- alkyl, C2-C4-alkenyl, C2-C4-alkynyl, C1-C4-haloalkyl, and cyclopropyl.

[0088] In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OH, OR ^6b , SR ⁶ , S(O) ₂ R ⁶ , S(O) ₂ NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , C(O)NR ⁶ R ⁶ , C1-C4- alkyl, C1-C4-haloalkyl, and cyclopropyl. In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , SR ⁶ , S(O)2R ⁶ , S(O)2NR ⁶ R ⁶ , CO2R ⁶ , C(O)R ⁶ , C(O)NR ⁶ R ⁶ , C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl.

[0089] In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OH, OR ^6b , C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl. In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , C1- C2-alkyl, C1-C2-haloalkyl, and cyclopropyl. In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , C1-C4-alkyl, C1-C4-haloalkyl, and cyclopropyl. [0090] In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OH, OR ^6b , C1-C2-alkyl, C1-C2-haloalkyl, and cyclopropyl. In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , C1- C2-alkyl, C1-C2-haloalkyl, and cyclopropyl.

[0091] In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OH, OR ^6b , C1-C2-alkyl, and C1-C2-haloalkyl. In an embodiment, R ⁵ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ^6b , C1-C2- alkyl, and C1-C2-haloalkyl.

[0092] In an embodiment, R ⁵ is independently at each occurrence OR ^6b . In an embodiment, R ⁵ is independently at each occurrence OMe. In an embodiment, where n is selected from 1 , 2, 3, or 4, at least one R ⁵ is independently OR ^6b . In an embodiment, where n is selected from 1 , 2, 3, or 4, at least one R ⁵ is independently OMe. In an embodiment, n is 1 and R ⁵ is OR ^6b . In an embodiment, n is 1 and R ⁵ is OMe.

[0093] In an embodiment, R ⁵ is OH.

[0094] In embodiments where R ⁵ is CO2R ⁶ or C(O)R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C2 alkyl.

[0095] In an embodiment, R ⁶ is independently H. In an embodiment, R ⁶ is independently C1-C4 alkyl. In an embodiment, R ⁶ is independently C1-C2 alkyl.

[0096] In an embodiment, R ^6a is independently selected from H and C1-C4 alkyl. In an embodiment, R ^6a is independently selected from H and methyl.

[0097] In an embodiment, two R ^6a groups, together with the nitrogen atom to which they are attached, form a 5- to 8-membered heterocycloalkyl ring; optionally a 6-membered heterocycloalkyl ring.

[0098] In an embodiment, R ^6b is independently at each occurrence selected from C1-C3 alkyl. In an embodiment, R ^6b is independently at each occurrence selected from C1-C2 alkyl. In an embodiment, R ^6b is C1-alkyl.

[0099] In an embodiment, R ⁷ is independently H. In an embodiment, R ⁷ is independently C1-C4 alkyl. In an embodiment, R ⁷ is independently C1-C2 alkyl. In an embodiment, R ⁷ is independently C(O)-C1-C4-alkyl. In an embodiment, R ⁷ is independently C(O)-C1-C2-alkyl.

[00100] In an embodiment, R ^7a is independently H. In an embodiment, R ^7a is independently C1-C4 alkyl. In an embodiment, R ^7a is independently C1-C2 alkyl. In an embodiment, R ^7a is independently C(O)-C1-C4-alkyl. In an embodiment, R ^7a is independently C(O)-C1-C2-alkyl. [00101] In an embodiment, R ⁸ is independently at each occurrence selected from =0, =S, halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , SO ₂ NR ⁶ R ⁶ , CO ₂ R ⁶ , C(O)R ⁶ , CONR ⁶ R ⁶ , C1-C4-alkyl, C1-C4-haloalkyl, cyclopropyl, C1-C3-alkylene-NR ⁶ R ⁷ , and C0-C3-alkylene-phenyl.

[00102] In an embodiment, R ⁸ is independently at each occurrence selected from =0, halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , SR ⁶ , CO ₂ R ⁶ , C(O)R ⁶ , CONR ⁶ R ⁶ , C1-C ₄ -alkyl, C1-C ₄ -haloalkyl, cyclopropyl, C1-C3-alkylene-NR ⁶ R ⁷ , and C0-C3-alkylene-phenyl.

[00103] In an embodiment, R ⁸ is independently at each occurrence selected from =0, halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , CO ₂ R ⁶ , C1-C4-alkyl, C1-C4-haloalkyl, cyclopropyl, C1-C3-alkylene- NR ⁶ R ⁷ , and C0-C3-alkylene-phenyl.

[00104] In an embodiment, R ⁸ is independently at each occurrence selected from =0, NR ⁶ R ⁷ , CO ₂ R ⁶ , C1-C4-alkyl, C1-C3-alkylene-NR ⁶ R ⁷ , and C0-C3-alkylene-phenyl.

[00105] In an embodiment, R ⁸ is independently selected from C1-C4-alkyl, C1-C4-haloalkyl and cyclopropyl.

[00106] In embodiments where R ⁸ is OR ⁶ , CO ₂ R ⁶ , or C(O)R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C ₂ alkyl.

[00107] In an embodiment, R ⁹ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , SR ⁶ , SOR ⁶ , S(O) ₂ R ⁶ , SO ₂ NR ⁶ R ⁶ , CO ₂ R ⁶ , C(O)R ⁶ , CONR ⁶ R ⁶ , C1-C4- alkyl, C1-C4-haloalkyl and cyclopropyl.

[00108] In an embodiment, R ⁹ is independently at each occurrence selected from halo, nitro, cyano, NR ⁶ R ⁷ , OR ⁶ , C1-C4-alkyl, C1-C4-haloalkyl and cyclopropyl.

[00109] In an embodiment, R ⁹ is independently at each occurrence selected from C1-C4- alkyl, C1-C4-haloalkyl and cyclopropyl.

[00110] In an embodiment, R ⁹ is independently at each occurrence C1-C4-alkyl.

[00111] In embodiments where R ⁹ is OR ⁶ , CO ₂ R ⁶ , or C(O)R ⁶ , R ⁶ may be R ^6c , where R ^6c is independently at each occurrence selected C1-C4 alkyl; optionally wherein R ^6c is independently at each occurrence selected C1-C ₂ alkyl.

[00112] In an embodiment, m is 1. In an embodiment, m is 0.

[00113] In an embodiment, x is 1. In an embodiment, x is 0.

[00114] In an embodiment, m is 0 and x is 0.

[00115] In an embodiment, n is 4. In an embodiment, n is 3. In an embodiment, n is 2. In an embodiment, n is 1. In an embodiment, n is 0. In an embodiment, n is an integer selected from 1 , 2 and 3. In an embodiment, n is an integer selected from 1 and 2. [00116] In an embodiment, any of the alkyl, alkylene, or alkenyl groups are optionally substituted, where chemically possible, by 1 to 5 substituents which are each independently at each occurrence selected from the group consisting of: oxo, fluoro, NR ^a R ^b , OR ^a , and S(O)2R ^a ; wherein R ^a is independently at each occurrence selected from H, and C1-C4-alkyl; and R ^b is independently at each occurrence selected from H, C1-C4-alkyl, C(O)-C1-C4-alkyl and S(O)2-C1-C4-alkyl.

[00117] In an embodiment, R ² is C2-C4-alkylene-NR ^6a R ^7a . In an embodiment, R ² is (CH2)r- - NR ^6a R ^7a , wherein r is an integer from 2 to 4.

[00118] In an embodiment, R ² is C0-C4-alkylene-Ring A. In an embodiment, R ² is C0-C3- alkylene-Ring A. In an embodiment, R ² is (CH2) _P -Ring A, wherein p is selected from 0, 1 , 2, 3 or 4. In an embodiment, R ² is (CH2) _P -Ring A, wherein p is selected from 0, 1 , 2, or 3.

[00119] In an embodiment, R ² is C(0)-C0-C4-alkylene-Ring A. In an embodiment, R ² is C(O)- C0-C2-alkylene-Ring A. In an embodiment, R ² is C(O)-(CH2) _q -Ring A, wherein q is selected from 0, 1 , 2, 3 or 4. In an embodiment, R ² is C(O)-(CH2) _q -Ring A, wherein q is selected from 0, 1 , 2 or 3.

[00120] In an embodiment, Ring A is independently selected from a 3- to 8-membered heterocycloalkyl, 5- to 8-membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein Ring A is heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups.

[00121] In an embodiment, Ring A is independently selected from C3-C6 cycloalkyl, C5-Ce cycloalkenyl, phenyl, 4- to 6-membered heterocycloalkyl, 5- to 6-membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein Ring A is cycloalkyl, cycloalkenyl, heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups.

[00122] In an embodiment, Ring A is independently selected from a 4- to 6-membered heterocycloalkyl, 5- to 6-membered heterocycloalkenyl, and 5- or 6-membered heteroaryl; wherein Ring A is heterocycloalkyl or heterocycloalkenyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups.

[00123] In an embodiment, Ring A is independently selected from C3-C6 cycloalkyl, phenyl, 3- to 8-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; wherein Ring A is cycloalkyl or heterocycloalkyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is phenyl or heteroaryl, Ring A is optionally substituted with from 1 to 5 R ⁹ groups.

[00124] In an embodiment, Ring A is independently selected from a 3- to 8-membered heterocycloalkyl, and 5- or 6-membered heteroaryl; wherein Ring A is heterocycloalkyl, Ring A is optionally substituted with from 1 to 4 R ⁸ groups and where Ring A is heteroaryl, Ring

A is optionally substituted with from 1 to 5 R ⁹ groups.

[00125] In an embodiment, Ring A is independently selected from a 4-membered heterocycloalkyl. In an embodiment, Ring A is independently selected from a 5-membered heterocycloalkyl, In an embodiment, Ring A is independently selected from a 6-membered heterocycloalkyl. In an embodiment, where Ring A is a 4-, 5-, or 6-membered heterocycloalkyl, Ring A is substituted with from 1 to 4 R ⁸ groups.

[00126] In an embodiment, Ring A is 5-membered heteroaryl. In an embodiment, Ring A is 6-membered heteroaryl. In an embodiment, where Ring A is heteroaryl, Ring A is substituted with from 1 to 5 R ⁹ groups. [00127] In an embodiment, Ring A is selected from:

Optionally wherein Ring A is substituted with from 1 to 4 R ⁸ groups.

[00128] In an embodiment, Ring A is selected from:

. Optionally wherein Ring A is substituted with from 1 to 5 R ⁹ groups. [00130] In an embodiment, Ring A is selected from:

[00131] In an embodiment, p is 0. In an embodiment, p is 1. In an embodiment, p is 2. In an embodiment, p is 3. [00132] In an embodiment, q is 0. In an embodiment, q is 1. In an embodiment, q is 2.

[00133] In an embodiment, r is 2. In an embodiment, r is 3. In an embodiment, r is 4.

[00134] In an embodiment, the compound of formula (I) is selected from:

[00135] In accordance with a second aspect, the present invention provides a pharmaceutical composition comprising a compound defined in the first aspect, and one or more pharmaceutically acceptable excipients. It may be that the pharmaceutical composition does not comprise a compound selected from List A. It may be that the pharmaceutical composition does not comprise a compound selected from List B. It may be that the pharmaceutical composition does not comprise a compound selected from List A and List B.

[00136] In accordance with a third aspect, the present invention provides a compound as defined in the first aspect or a pharmaceutical composition as defined in the second aspect, for use as a medicament. It may be that the pharmaceutical composition does not comprise a compound selected from List A. It may be that the pharmaceutical composition does not comprise a compound selected from List B. It may be that the pharmaceutical composition does not comprise a compound selected from List A and List B.

[00137] In accordance with a fourth aspect, the present invention provides the use of a compound as defined in the first aspect or a pharmaceutical composition as defined in the second aspect, for the manufacture of a medicament. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B. [00138] In accordance with a fifth aspect, the present invention provides a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect, for use in a method of treatment or prophylaxis of a disease of the musculoskeletal system, a disease of the skin, a metabolic disease, a disease of the Nervous System, a Cardiovascular disease, an Endocrine disorder, a disease of the eye, a disease affecting the urogenital system, a haemic or lymphatic condition, a respiratory disease, an inflammatory or autoimmune condition, a disease of the Gastrointestinal system, a Neoplasm, cancer, or a disease or disorder selected from Amelogenesis Imperfecta, Anodontia, Odontodysplasia, Branchio-Oto-Renal Syndrome, Sotos Syndrome, and Waardenburg's Syndrome. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00139] In accordance with a sixth aspect, the present invention provides a method for the treatment or prophylaxis of a disease of the musculoskeletal system, a disease of the skin, a metabolic disease, a disease of the Nervous System, a Cardiovascular disease, an Endocrine disorder, a disease of the eye, a disease affecting the urogenital system, a haemic or lymphatic condition, a respiratory disease, an inflammatory or autoimmune condition, a disease of the Gastrointestinal system, a Neoplasm, cancer, or a disease or disorder selected from Amelogenesis Imperfecta, Anodontia, Odontodysplasia, Branchio- Oto-Renal Syndrome, Sotos Syndrome, and Waardenburg's Syndrome, said method comprising administering to a subject, an effective amount of a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00140] In accordance with a seventh aspect, the present invention provides a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect, for use in a method of treatment or prophylaxis of a disease selected from: Recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa Xeroderma pigmentosum, Netherton syndrome, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, Cystic Fibrosis, Dravet Syndrome, Aniridia, Methylmalonic Acidemia, Colorectal Cancer, Endometrium Cancer, Breast Cancer, Ovarian Cancer, Lung Squamous Cell Carcinoma, Head and Neck Squamous Cell Carcinoma, Familial adenomatous polyposis, Hemophilia A, Hemophilia B, Choroideremia, Pulmonary Artery Hypertension, Ataxia telangiectasia, Shwachman-Diamond syndrome, Mucopolysaccharidosis Type I, Mucopolysaccharidosis Type VI, Mucopolysaccharidosis type III, Niemann-Pick Disease, and Primary Ciliary Dyskinesia. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00141] In accordance with an eighth aspect, the present invention provides a method for the treatment or prophylaxis of a disease selected from: Recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa Xeroderma pigmentosum, Netherton syndrome, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, Cystic Fibrosis, Dravet Syndrome, Aniridia, Methylmalonic Acidemia, Colorectal Cancer, Endometrium Cancer, Breast Cancer, Ovarian Cancer, Lung Squamous Cell Carcinoma, Head and Neck Squamous Cell Carcinoma, Familial adenomatous polyposis, Hemophilia A, Hemophilia B, Choroideremia, Pulmonary Artery Hypertension, Ataxia telangiectasia, Shwachman- Diamond syndrome, Mucopolysaccharidosis Type I, Mucopolysaccharidosis Type VI, Mucopolysaccharidosis type III, Niemann-Pick Disease, and Primary Ciliary Dyskinesia.

[00142] In accordance with a ninth aspect, the present invention provides the use of a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect for the manufacture of a medicament for the treatment or prophylaxis of a disease of the musculoskeletal system, a disease of the skin, a metabolic disease, a disease of the Nervous System, a Cardiovascular disease, an Endocrine disorder, a disease of the eye, a disease affecting the urogenital system, a haemic or lymphatic condition, a respiratory disease, an inflammatory or autoimmune condition, a disease of the Gastrointestinal system, a Neoplasm, cancer, or a disease or disorder selected from Amelogenesis Imperfecta, Anodontia, Odontodysplasia, Branchio-Oto-Renal Syndrome, Sotos Syndrome, and Waardenburg's Syndrome. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00143] In accordance with a tenth aspect, the present invention provides a method of allowing translational read-through of PTC mutations in a subject, said method comprising administering to a subject an effective amount of a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00144] In accordance with an eleventh aspect, the present invention provides a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect, for use in a method of treating conditions or disorders which are associated with PTC mutations in a subject, said method comprising administering to a subject an effective amount of a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00145] In accordance with a twelfth aspect, the present invention provides a method of treating conditions or disorders which are associated with PTC mutations in a subject, said method comprising administering to a subject an effective amount of a compound as defined in the first aspect, or a pharmaceutical composition as defined in the second aspect. It may be that the compound is not selected from List A. It may be that the compound is not selected from List B. It may be that the compound is not selected from List A and List B.

[00146] Suitably a PTC mutation may be any mutation which generates a premature termination codon (PTC) in a gene. Suitably a PTC mutation may be any mutation which generates an in-frame premature termination codon (PTC) in a gene. Suitably a PTC mutation occurs in the nucleotide sequence of a gene. Suitably a PTC mutation may occur in the coding region or non-coding region of a gene. Suitably a PTC mutation may occur in the coding region of a gene.

[00147] Suitably the mutation may be a point mutation, for example a substitution, deletion or insertion mutation. Suitably the mutation is a substitution mutation. Suitably the substitution mutation may replace one nucleotide with another different nucleotide, suitably within the nucleotide sequence of a gene. Suitably the mutation may therefore be regarded as a nonsense mutation. Suitably the mutation may be a nonsense mutation which causes an in-frame PTC in a gene. Suitably the PTC may be any known termination codon such as TAG, TAA or TGA, suitably the PTC is TGA. Suitably therefore in some embodiments, the PTC mutation may be a substitution mutation which generates a PTC comprising TAG, TAA or TGA in a gene. Suitably therefore in some embodiments, the PTC mutation may be a substitution mutation of CGA to TGA or CAG to TAG in a gene.

[00148] A condition or disorder associated with a PTC mutation in a subject may be a condition or disorder which is directly or indirectly caused by a PTC mutation in a subject. Suitably a condition or disorder associated with a PTC mutation in a subject may be a condition or disorder which is directly caused by a PTC mutation in a subject. Suitably a condition or disorder associated with a PTC mutation in a subject may be a condition or disorder which is caused by a PTC mutation in one or more genes in a subject. Suitably a condition or disorder associated with a PTC mutation in a subject may be a condition or disorder which is directly caused by a PTC mutation in one or more genes in a subject. Suitably a condition or disorder associated with a PTC mutation in a subject may be a condition or disorder which is caused by a PTC mutation in one or more genes in a subject leading to a loss of function of the or each gene. Suitably reference herein to ‘a’ PTC mutation may be understood to refer to one or more PTC mutations, or a plurality of PTC mutations, suitably which may be present in one or more genes in a subject.

[00149] Suitably the one or more genes in which the PTC mutation is present are genes relating to one or more diseases as described herein. Suitably the one or more genes may be directly or indirectly linked with one or more diseases described herein. Suitably the one or more genes may contribute to, or cause, the phenotype of one or more diseases described herein. In some embodiments, for example in which the disease is cancer, the one or more genes in which the PTC mutation is present may be cell cycle genes, tumour suppressor genes etc. Such tumour suppressor genes may include TP53, PTEN, APC. Suitably such mutations may be regarded as driver mutations, suitably which drive a disease phenotype, such as tumour growth. Therefore, in some embodiments, the PTC mutation may be regarded as a driver mutation. In other embodiments, the one or more genes in which the PTC mutation is present may not contribute to or cause the disease phenotype. Suitably such mutations may be regarded as bystander mutations. Therefore in some embodiments, the PTC mutation may be regarded as a bystander mutation.

[00150] Conditions or disorders associated with a PTC mutation are well known in the art, and suitable such disorders which may be treated or prevented by the present invention are described hereinbelow. However, suitably, such disorders that are associated with PTC mutations may also be identified or determined by the use of standard molecular biology techniques. Suitably a condition or disorder may be determined as being associated with a PTC mutation by the use of molecular biology techniques, such as sequencing, single strand conformational polymorphism, denaturing gradient gel electrophoresis, heteroduplex analysis, or restriction fragment length polymorphism. Suitably a PTC mutation present in one or more genes of a subject which may cause a disease or condition can be identified using such techniques. Suitably, whole genome sequencing or whole exome sequencing may be used to identify a PTC mutation in one or more genes of a subject, suitably which may cause a disease or condition. Suitable methodologies may be described in Karagiannakos et al. Cancers (Basel) 2022, 14, 664, Stark et al. ‘A prospective evaluation of whole-exome sequencing as a first-tier molecular test in infants with suspected monogenic disorders’ Genetics in Medicine, Volume 18, Issue 11 , 2016, North et al. ‘Approach to the diagnosis of congenital myopathies’ Neuromuscular Disorders, Volume 24, Issue 2, 2014, for example.

[00151] Suitably such techniques may be applied to a sample obtained from a subject suffering from a disease or condition or suspected of suffering from a disease or condition, suitably which may be caused by a PTC mutation. Suitably such techniques may comprise comparing a result from said sample, to a result obtained from a reference sample. Suitably a reference sample may be a sample of the same type from a healthy subject, suitably from an equivalent healthy subject of the same age, nationality, race, height, weight etc. Suitable samples may include: a blood sample, serum sample, CNS fluid sample, tissue sample, cell sample, and the like. Suitably comparing a result in the context of the techniques above may comprise comparing the sequence of one or more genes from the subject suffering from a disease or condition to the sequence of the same one or more genes of the healthy subject. Suitably to identify PTC mutations therein. Suitably such sequence comparisons may be carried out by available software, such as alignment software.

[00152] PTC read-through agents, such as the compounds disclosed herein, may in one or more embodiments, be of value and used in the treatment or amelioration of the following non-limiting examples of disorders and diseases. Suitably any of the following disorders and diseases may be regarded as a disease or disorder associated with a PTC mutation in a subject. Suitably therefore the present invention provides a method of treating conditions or disorders which are associated with PTC mutations in a subject, wherein the conditions or disorders are selected from any listed in the following paragraphs.

[00153] The disease may be a disease of the musculoskeletal system. The disease of the musculoskeletal system may be a disease selected from Shwachman-Diamond Syndrome, Rickets, Laron Syndrome, Muscular Dystrophies (e.g. Duchenne muscular dystrophy (DMD)), Microcephaly, congenital limb deformities, Muscle Spasticity, Dwarfism, Gigantism, Osteopoikilosis, Cleidocranial Dysplasia, Synostosis, Mitochondrial Myopathies and Encephalomyopathies, Craniosynostoses, Mandibulofacial Dysostosis, Scoliosis, Osteoporosis, Osteopetrosis, Hyperostosis, Osteosclerosis, Osteogenesis Imperfecta, Holoprosencephaly, Acromegaly, Arthrogryposis, and Campomelic Dysplasia.

[00154] The disease may be a disease of the skin. The disease of the skin may be a disease selected from Ectodermal Dysplasia, Epidermolysis Bullosa, Focal Dermal Hypoplasia, Ichthyosis Vulgaris, Autosomal Recessive Congenital Ichthyosis (ARCI), Recessive X-linked Ichthyosis, Lamellar Ichthyosis, Congenital Ichthyosiform Erythroderma, Harlequin Ichthyosis, Epidermolytic Ichthyosis, Superficial Epidermolytic Ichthyosis, CHILD syndrome, Netherton syndrome, MEDNIK syndrome, Neutral lipid storage disease with ichthyosis, Atopic Dermatitis, Alopecia, Atrichia with papular lesions, Hypotrichosis, Incontinentia Pigmenti, Epidermolysis Bullosa Simplex, Lipodystrophy, Hidradenitis Suppurativa, Hyperpigmentation, Junctional Epidermolysis Bullosa, Oculocutaneous Albinism, Chronic Mucocutaneous Candidiasis, Chronic Mucocutaneous Ichthyosis, X-Linked Hypopigmentation, Cutis Laxa, Menkes Kinky Hair Syndrome, Palmoplantar Keratoderma, Ehlers-Danlos Syndrome, Congenital Monilethrix, Onychomycosis, Benign Familial Pemphigus, Albinism; Kindler syndrome, Cowden Syndrome, Xeroderma pigmentosum, epidermodysplasia verruciformis, Lipoid proteinosis, dyschromatosis symmetrica hereditaria, Striate keratodermas, Autosomal Recessive Congenital Anonychia, Focal dermal hypoplasia, and Skin fragility/woolly hair syndrome.

[00155] The disease may be a metabolic disease. The metabolic disease may be selected from Congenital Adrenal Hyperplasia, Inborn errors of Amino Acid Metabolism, Hypertriglyceridemia, Diabetes Mellitus, Type 1 and Type 2, Glycogen Storage Disease, Optic Atrophy, Calcinosis, Multiple Sulfatase Deficiency Disease, Fabry Disease, Hyperinsulinism, Familial Hypophosphatemia, Pseudohypoaldosteronism, Tangier Disease, Amyotrophic Lateral Sclerosis, Lactic Acidosis, Familial Amyloidosis, Mucopolysaccharidosis, Anemia, Sandhoff Disease, Cytochrome-c Oxidase Deficiency, Pyruvate Dehydrogenase Complex Deficiency Disease, Hypoglycemia, Neuronal Ceroid- Lipofuscinoses, Severe Combined Immunodeficiency, , Chronic Idiopathic Jaundice, Progeria, Hyponatremia, Wasting Syndrome, Cystinuria, Glycogen Storage Disease, Gaucher Disease, Hypolipoproteinemias, Oculocerebrorenal Syndrome, Smith-Lemli-Opitz Syndrome, Adrenoleukodystrophy, Ataxia Telangiectasia, Canavan Disease, Carbamoyl- Phosphate Synthase I Deficiency Disease, beta-Mannosidosis, and lyosomal storage diseases, wherein the lyosomal storage disease may be selected from Niemann-Pick Disease Mucopolysaccharidosis type 1 (Hurler syndrome), Mucopolysaccharidosis type 6, Mucopolysaccharidosis type 7, CLN1 disease, and CL3 disease.

[00156] The disease may be a disease of the Nervous System. The disease of the Nervous System may be selected from Rett Syndrome, Ataxias, Sensorineural Hearing Loss, Epilepsy, Charcot-Marie-Tooth Disease, Spinal Muscular Atrophy, Spastic Paraplegia, Hydrocephalus, Migraine with Aura, Chorea, Tremor, Usher Syndromes, De Lange Syndrome, Duane Retraction Syndrome, Dementia, Myoclonus, Hereditary Sensory and Autonomic Neuropathies, Intellectual Disability, X-linked Mental Retardation, Fragile X Syndrome, Hypopituitarism, Leukoencephalopathies, Dystonia, Congenital Pain Insensitivity, Tourette Syndrome, Alzheimers Disease, Parkinsons Disease, Angelman Syndrome, Apraxias, Cerebral Palsy, and Frontotemporal Dementia.

[00157] The disease may be a Cardiovascular disease. The cardiovascular disease may be selected from Coronary Disease, Ventricular Fibrillation, Telangiectasis, Kartagener Syndrome, Alagille Syndrome, Andersen Syndrome, Atrioventricular Block, and Cardiomyopathies.

[00158] The disorder may be an Endocrine disorder. The Endocrine disorder may be selected from Hypogonadism, Goiter, Fetal Macrosomia, Thyroid Hormone Resistance Syndrome, Gonadal Dysgenesis, Hypoparathyroidism, Neurogenic Diabetes Insipidus, and Androgen-Insensitivity Syndrome. [00159] The disease may be a disease of the eye. The disease of the eye may be selected from Leber syndrome, Hereditary Optic Atrophies, Ectopia Lentis, Coloboma, Aphakia, and Choroideremia.

[00160] The disease may be a disease affecting the urogenital system. The disease affecting the urogenital system may be selected from Hypospadias, Hydrops Fetalis, Interstitial Nephritis, Polycystic Kidney Diseases, and Azoospermia.

[00162] The disease or disorder may be a haemic or lymphatic condition. The haemic or lymphatic condition may be selected from alpha- and beta-Thalassemia, Afibrinogenemia, Hemophagocytic Lymphohistiocytosis, Factor XI Deficiency, Hemophilia A, von Willebrand Diseases, Factor V Deficiency, Sideroblastic Anemia, Hereditary Elliptocytosis, Neutropenia, Chronic Granulomatous Disease, Hereditary Spherocytosis, Polycythemia, Hemophilia B, Factor VII Deficiency, Bernard-Soulier Syndrome, Dyserythropoietic Anemia, Hemolytic Anemia, Idiopathic Thrombocytopenic Purpura, Thrombasthenia, Factor XIII Deficiency, Hepatoerythropoietic Porphyria, and Acute Intermittent Porphyria.

[00163] The disease may be a respiratory disease. The respiratory disease may be selected from Cystic Fibrosis, Pulmonary Hypertension, Lipoid Proteinosis of Urbach and Wiethe, Newborn Respiratory Distress Syndrome, Chronic Obstructive Pulmonary Disease, Chronic Obstructive, Asthma, and Choanal Atresia.

[00164] The disease or disorder may be an inflammatory or autoimmune condition. The inflammatory or autoimmune condition may be selected from Immunologic Deficiency Syndromes, and Leukocyte-Adhesion Deficiency Syndrome.

[00165] The disease may be a disease of the Gastrointestinal system. Diseases of the gastrointestinal system may be selected from Chronic Hepatitis B, Colitis, Intestinal Polyposis, Inflammatory Bowel Diseases, Hirschsprung Disease, Exocrine Pancreatic Insufficiency, Crohn’s Disease, and Cholestasis.

[00166] The disease or disorder may be a Neoplasm. The Neoplasm may be selected from Acute Myeloid Leukemia, Paraganglioma, Rhabdoid Tumor, Rhabdomyoma, Adenoid Cystic Carcinoma, Large Cell Carcinoma, Lobular Carcinoma, Skin Appendage Carcinoma, Squamous Cell Carcinoma, Alveolar Rhabdomyosarcoma, Neuroectodermal Tumors, Multiple Hamartoma Syndrome, Pheochromocytoma, Nevus, Osteosarcoma, Teratoma, and Adenoma.

[00167] The disease or disorder may be cancer. The cancer may be selected from acoustic neuroma, anal cancer, bladder cancer, Bowen's disease, brain cancer, breast cancer, carcinomas including basal cell carcinoma, bile duct carcinoma, bronchogenic carcinoma, choriocarcinoma, embryonal carcinoma, cystadenocarcinoma, epithelial carcinoma, medullary carcinoma, NUT midline carcinoma (NMC), papillary carcinoma, papillary adenocarcinomas, renal cell carcinoma, sebaceous gland carcinoma, small cell lung carcinoma, squamous cell carcinoma, and sweat gland carcinoma, cervical cancer, chordoma, colon cancer, colorectal cancer, craniopharyngioma, dysproliferative changes (dysplasias and metaplasias), endometrial cancer, ependymoma, esophageal cancer, essential thrombocythemia, estrogen-receptor positive breast cancer, Ewing’s tumour, genital cancer, cancer of the cervix, cancer of the vulva, vulvar intraepithelial neoplasia (VI N), cancer of the vagina, germ cell testicular cancer, gastrointestinal cancers, gastric cancer, glioblastoma, glioma, heavy chain disease, hemangioblastoma, hepatocellular cancer, hepatoma, hormone insensitive prostate cancer, keratinocyte carcinomas, kidney cancer, leukaemias including acute leukaemia, acute lymphocytic leukaemia, acute myeloid leukaemia, acute myelocytic leukaemia (monocyctic, myeloblastic, adenocarcinoma, angiosarcoma, astrocytoma, myelomonocytic and promyelocytic), acute t-cell leukaemia, chronic leukaemia, chronic lymphocytic leukaemia, chronic myelocytic (granulocytic) leukaemia, chronic myelogenous leukaemia, erythroleukemia, lymphoblastic leukaemia, and myelogenous leukaemia, liver cancer, lung cancer, lymphoid malignancies of T-cell or B-cell origin, lymphomas (Hodgkin’s and non-Hodgkin’s) including cutaneous T-cell lymphoma, diffuse large B-cell lymphoma, and follicular lymphoma, cutaneous (skin) lymphomas, malignancies and hyperproliferative disorders including of the bladder, breast, colon, lung, ovaries, pancreas, prostate, skin and uterus, advanced malignancies, medulloblastoma, melanoma, meningioma, Merkel cell cancer mesothelioma, metastatic cancer, multiple myeloma, myeloma, pancreatic cancer, myelofibrosis, myeloproliferative neoplasms, neuroblastoma, non-small cell lung cancer, head and neck cancer, oligodendroglioma, oral cancer, ovarian cancer, pancreatic cancer, pinealoma, polycythemia vera, prostate cancer, rectal cancer, retinoblastoma, sarcomas including chondrosarcoma, endotheliosarcoma, fibrosarcoma, gliosarcoma, leiomyosarcoma, liposarcoma, lymphagioendotheliosarcoma, lymphangiosarcoma, myxosarcoma, Castleman's disease and Kaposi's sarcoma, osteogenic sarcoma, and rhabdomyosarcoma, seminoma, skin cancer, skin adnexal tumors, and sarcomas, small cell lung cancer, solid tumors, stomach cancer, synovioma, testicular tumours, thyroid cancer, uterine cancer, Waldenstrom’s macroglobulinemia, and Wilms’ tumour.

[00168] The disease or disorder may be selected from Amelogenesis Imperfecta, Anodontia, Odontodysplasia, Branchio-Oto-Renal Syndrome, Sotos Syndrome, and Waardenburg's Syndrome (all of which will be referred to as “ PTC conditions”).

[00169] The disease or disorder may be selected from: Recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa, Xeroderma pigmentosum, Netherton syndrome, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, Cystic Fibrosis, Dravet Syndrome, Aniridia, Methylmalonic Acidemia, Colorectal Cancer, Endometrium Cancer, Breast Cancer, Ovarian Cancer, Lung Squamous Cell Carcinoma, Head and Neck Squamous Cell Carcinoma, Familial adenomatous polyposis, Hemophilia A, Hemophilia B, Choroideremia, Pulmonary Artery Hypertension, Ataxia telangiectasia, Shwachman- Diamond syndrome, Mucopolysaccharidosis Type I, Mucopolysaccharidosis Type VI, Mucopolysaccharidosis type III, Niemann-Pick Disease, and Primary Ciliary Dyskinesia.

[00170] The disease or disorder may be selected from: Recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa, Duchenne Muscular Dystrophy, Cystic Fibrosis, Colorectal Cancer, Endometrium Cancer, Breast Cancer, Ovarian Cancer, Lung Squamous Cell Carcinoma, Head and Neck Squamous Cell Carcinoma, and Familial adenomatous polyposis. Suitably such disorders may be associated with a PTC mutation in a subject.

[00171] Suitably therefore the present invention provides a method of treating conditions or disorders which are associated with PTC mutations in a subject, wherein the condition or disorder associated with PTC mutations in a subject is selected from recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa, Xeroderma pigmentosum, Netherton syndrome, Duchenne Muscular Dystrophy, Becker Muscular Dystrophy, Cystic Fibrosis, Dravet Syndrome, Aniridia, Methylmalonic Acidemia, Familial adenomatous polyposis, Hemophilia A, Hemophilia B, Choroideremia, Pulmonary Artery Hypertension, Ataxia telangiectasia, Shwachman-Diamond syndrome, Mucopolysaccharidosis Type I, Mucopolysaccharidosis Type VI, Mucopolysaccharidosis type III, Niemann-Pick Disease, Primary Ciliary Dyskinesia and Alport Syndrome. In some embodiments, the present invention provides a method of treating conditions or disorders which are associated with PTC mutations in a subject, wherein the condition or disorder associated with PTC mutations in a subject is selected from recessive dystrophic epidermolysis bullosa, Junctional epidermolysis bullosa, Duchenne Muscular Dystrophy, Cystic Fibrosis, and Familial adenomatous polyposis.

[00172] PTC read-through agents, such as the compounds disclosed herein, may in one or more embodiments, also be of value and used in the palliation, diagnosis or prevention of any disease, disorder or condition in humans of one or more of the aforesaid non-limiting examples of disorders and diseases. Suitably any of the aforesaid disorders and diseases may be regarded as a disease or disorder associated with a PTC mutation in a subject. Suitably therefore the present invention further provides a method of palliation, diagnosis or prevention of conditions or disorders which are associated with PTC mutations in a subject, wherein the conditions or disorders are selected from any listed in the aforesaid paragraphs. [00173] Treatment or amelioration with PTC read-through agents, such as compositions comprising the compounds disclosed herein or salts thereof (or combinations thereof), in some embodiments may be effective if administered orally. In some other embodiments may be effective if applied topically, and in some further embodiments may be effective if applied topically and orally.

[00174] In some embodiments, compositions comprising a novel compound disclosed herein or salt thereof (or combinations thereof) may be administered to young children. In some embodiments, compositions comprising a compound of the invention or salt thereof (or combinations thereof) may be administered to adolescents or teenagers. In some embodiments, compositions comprising a compound of the invention or salt thereof (or combinations thereof) may be administered to adults.

BRIEF DESCRIPTION OF THE DRAWINGS

[00175] Embodiments of the invention are further described hereinafter with reference to the accompanying drawings, in which:

Figure 1 shows the effect of treatment of HDQ-P1 cells with Example 3 (0.41 pM, 1.23 pM, 3.7 pM, 11 pM, or 33 pM) compared to DMSO (0.3%), and G418 (200 pM).

Figure 2 shows the effect of treatment of HDQ-P1 cells with Example 4 (0.41 pM, 1.23 pM, 3.7 pM, 11 pM, or 33 pM) compared to DMSO (0.3%), and G418 (200 pM).

[00176]

DETAILED DESCRIPTION

[00177] The term C _m -C _n refers to a group with m to n carbon atoms. For the absence of doubt, the term “Co” refers to a group with 0 carbon atoms.

[00178] The term “alkyl” refers to a monovalent linear or branched saturated hydrocarbon chain. For example, C1-C6-alkyl may refer to methyl, ethyl, n-propyl, /so-propyl, n-butyl, sec- butyl, tert-butyl, n-pentyl and n-hexyl. The alkyl groups may be unsubstituted or substituted by one or more substituents.

[00179] The term “alkylene” refers to a bivalent linear saturated hydrocarbon chain. For example, C1-C3-alkylene may refer to methylene, ethylene or propylene. The alkylene groups may be unsubstituted or substituted by one or more substituents. For the absence of doubt, the term “C0-alkylene” refers to a group in which an alkylene chain is absent. For example, “C0-alkylene-R ^a ” refers to an R ^a .

[00180] The term “haloalkyl” refers to a hydrocarbon chain substituted with at least one halogen atom independently chosen at each occurrence from: fluorine, chlorine, bromine and iodine. The halogen atom may be present at any position on the hydrocarbon chain. For example, C1-C6-haloalkyl may refer to chloromethyl, fluoromethyl, trifluoromethyl, chloroethyl e.g. 1 -chloromethyl and 2-chloroethyl, trichloroethyl e.g. 1 ,2,2-trichloroethyl, 2,2,2-trichloroethyl, fluoroethyl e.g. 1 -fluoromethyl and 2-fluoroethyl, trifluoroethyl e.g. 1 ,2,2- trifluoroethyl and 2,2,2-trifluoroethyl, chloropropyl, trichloropropyl, fluoropropyl, trifluoropropyl. A haloalkyl group may be a fluoroalkyl group, i.e. a hydrocarbon chain substituted with at least one fluorine atom. Thus, a haloalkyl group may have any amount of halogen substituents. The group may contain a single halogen substituent, it may have two or three halogen substituents, or it may be saturated with halogen substituents.

[00181] The term “alkenyl” refers to a branched or linear hydrocarbon chain containing at least one double bond. The double bond(s) may be present as the E or Z isomer. The double bond may be at any possible position of the hydrocarbon chain. For example, “C2-C6-alkenyl” may refer to ethenyl, propenyl, butenyl, butadienyl, pentenyl, pentadienyl, hexenyl and hexadienyl. The alkenyl groups may be unsubstituted or substituted by one or more substituents.

[00182] The term “alkynyl” refers to a branched or linear hydrocarbon chain containing at least one triple bond. The triple bond may be at any possible position of the hydrocarbon chain. For example, “C2-C6-alkynyl” may refer to ethynyl, propynyl, butynyl, pentynyl and hexynyl. The alkynyl groups may be unsubstituted or substituted by one or more substituents.

[00183] The term “cycloalkyl” refers to a saturated hydrocarbon ring system containing 3, 4, 5 or 6 carbon atoms. For example, “C3-C6-cycloalkyl” may refer to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl. The cycloalkyl groups may be unsubstituted or substituted by one or more substituents.

[00184] The term “y- to z-membered heterocycloalkyl” refers to a y- to z- membered heterocycloalkyl group. Thus it may refer to a monocyclic or bicyclic saturated or partially saturated group having from y to z atoms in the ring system and comprising 1 or 2 heteroatoms independently selected from O, S and N in the ring system (in other words 1 or 2 of the atoms forming the ring system are selected from O, S and N). By partially saturated it is meant that the ring may comprise one or two double bonds. This applies particularly to monocyclic rings with from 5 to 6 members. The double bond will typically be between two carbon atoms but may be between a carbon atom and a nitrogen atom. Examples of heterocycloalkyl groups include; piperidine, piperazine, morpholine, thiomorpholine, pyrrolidine, tetra hydrofuran, tetrahydrothiophene, dihydrofuran, tetrahydropyran, dihydropyran, dioxane, azepine. A heterocycloalkyl group may be unsubstituted or substituted by one or more substituents. [00185] Aryl groups may be any aromatic carbocyclic ring system (i.e. a ring system containing 2(2n + 1)TT electrons). Aryl groups may have from 6 to 10 carbon atoms in the ring system. Aryl groups will typically be phenyl groups. Aryl groups may be naphthyl groups or biphenyl groups.

[00186] The term ‘heterocyclyl’ group refers to rings comprising from 1 to 4 heteroatoms independently selected from O, S and N. The rings may be heterocycloalkyl rings (including both saturated and partially saturated rings) or heteroaryl rings. The term “heterocyclyl” also encompasses groups that are tautomers of hydroxy heteroaryl groups, such pyridones, and tautomers of hydroxy heteroaryl groups that are substituted on the nitrogen, such as N-alkyl pyridones.

[00187] The term ‘heterocycloalkenyl’ refers to partially saturated rings comprising from 1 to 2 heteroatoms independently selected from O, S and N.

[00188] The term “heteroaryl” refers to any aromatic (i.e. a ring system containing 2(2n +

1 )TT electrons) 5 or 6 membered ring system comprising from 1 to 4 heteroatoms independently selected from O, S and N (in other words from 1 to 4 of the atoms forming the ring system are selected from O, S and N). Thus, any heteroaryl groups may be independently selected from: 5 membered heteroaryl groups in which the heteroaromatic ring is substituted with 1-4 heteroatoms independently selected from O, S and N; and 6- membered heteroaryl groups in which the heteroaromatic ring is substituted with 1-3 (e.g.1-

2) nitrogen atoms. Specifically, heteroaryl groups may be independently selected from: pyrrole, furan, thiophene, pyrazole, imidazole, oxazole, isoxazole, triazole, oxadiazole, thiadiazole, tetrazole; pyridine, pyridazine, pyrimidine, pyrazine, triazine.

[00189] Certain compounds of the invention comprise a ‘basic nitrogen’. A basic nitrogen may be an amine (including primary, secondary and tertiary amines) or it may be a nitrogen in a heteroaromatic ring (e.g. a pyridine, pyrimidine, pyridazine, pyrazine, imidazole or pyrazole). Amines include cyclic amines. The term ‘basic nitrogen’ does not include the nitrogen of an amide or a sulfonamide.

[00190] Compounds of the invention containing one or more asymmetric carbon atoms can exist as two or more stereoisomers. Where a compound of the invention contains a double bond such as a C=C or C=N group, geometric cis/trans (or Z/E) isomers are possible. Where structural isomers are interconvertible via a low energy barrier, tautomeric isomerism (‘tautomerism’) can occur. This can take the form of proton tautomerism in compounds of the invention containing, for example, an imino, keto, or oxime group, or so-called valence tautomerism in compounds which contain an aromatic moiety. It follows that a single compound may exhibit more than one type of isomerism. [00191] Included within the scope of the present invention are all stereoisomers, geometric isomers and tautomeric forms of the compounds of the invention, including compounds exhibiting more than one type of isomerism, and mixtures of one or more thereof. Also included are acid addition or base salts wherein the counter ion is optically active, for example, d-lactate or l-lysine, or racemic, for example, dl-tartrate or dl-arginine.

[00192] It will be apparent to those skilled in the art that the compounds of Formula I can by drawn in various tautomeric forms, non-exhaustive examples of which are illustrated below. Equally, it will be apparent to those skilled in the art that where substituent R groups are capable of resonance (for example OR, NRR, CN, NO2, C(O)R etc.), additional tautomers may be possible. All possible tautomeric or resonance forms are encompassed by the present invention.

[00193] Cis/trans isomers may be separated by conventional techniques well known to those skilled in the art, for example, chromatography and fractional crystallisation.

[00194] Conventional techniques for the preparation/isolation of individual enantiomers when necessary include chiral synthesis from a suitable optically pure precursor or resolution of the racemate (or the racemate of a salt or derivative) using, for example, chiral high pressure liquid chromatography (HPLC). Thus, chiral compounds of the invention (and chiral precursors thereof) may be obtained in enantiomerically-enriched form using chromatography, typically HPLC, on an asymmetric resin with a mobile phase consisting of a hydrocarbon, typically heptane or hexane, containing from 0 to 50% by volume of isopropanol, typically from 2% to 20%, and for specific examples, 0 to 5% by volume of an alkylamine e.g. 0.1 % diethylamine. Concentration of the eluate affords the enriched mixture.

[00195] Alternatively, the racemate (or a racemic precursor) may be reacted with a suitable optically active compound, for example, an alcohol, or, in the case where the compound of the invention contains an acidic or basic moiety, a base or acid such as 1 -phenylethylamine or tartaric acid. The resulting diastereomeric mixture may be separated by chromatography and/or fractional crystallization and one or both of the diastereoisomers converted to the corresponding pure enantiomer(s) by means well known to a skilled person.

[00196] When any racemate crystallises, crystals of two different types are possible. The first type is the racemic compound (true racemate) referred to above wherein one homogeneous form of crystal is produced containing both enantiomers in equimolar amounts. The second type is the racemic mixture or conglomerate wherein two forms of crystal are produced in equimolar amounts each comprising a single enantiomer.

[00197] While both of the crystal forms present in a racemic mixture have identical physical properties, they may have different physical properties compared to the true racemate. Racemic mixtures may be separated by conventional techniques known to those skilled in the art - see, for example, “Stereochemistry of Organic Compounds” by E. L. Eliel and S. H. Wilen (Wiley, 1994).

[00198] Suitable pharmaceutically acceptable salts include, but are not limited to, salts of pharmaceutically acceptable inorganic acids such as hydrochloric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic, and hydrobromic acids, or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, malic, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, toluenesulphonic, benzenesulphonic, salicylic, sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids. Suitable base salts are formed from bases which form non-toxic salts. Examples include the aluminium, arginine, benzathine, calcium, choline, diethylamine, diolamine, glycine, lysine, magnesium, meglumine, olamine, potassium, sodium, tromethamine and zinc salts. Hemisalts of acids and bases may also be formed, for example, hemisulfate and hemicalcium salts.

[00199] The activity of the compounds of the present invention can be assessed by a variety of in silico, in vitro and in vivo assays. In silico analysis of a variety of compounds has been demonstrated to be predictive of ultimate in vitro and even in vivo activity.

[00200] It is to be appreciated that references to “treating” or “treatment” include prophylaxis as well as the alleviation of established symptoms of a condition. “Treating” or “treatment” of a state, disorder or condition therefore includes: (1) preventing or delaying the appearance of clinical symptoms of the state, disorder or condition developing in a human that may be afflicted with or predisposed to the state, disorder or condition but does not yet experience or display clinical or subclinical symptoms of the state, disorder or condition, (2) inhibiting the state, disorder or condition, /.e., arresting, reducing or delaying the development of the disease or a relapse thereof (in case of maintenance treatment) or at least one clinical or subclinical symptom thereof, or (3) relieving or attenuating the disease, /.e., causing regression of the state, disorder or condition or at least one of its clinical or subclinical symptoms.

[00201] A “therapeutically effective amount” includes the amount of a compound that, when administered to a mammal for treating a disease, is sufficient to affect such treatment for the disease. The "therapeutically effective amount" will vary depending on the compound, the disease and its severity and the age, weight, etc., of the mammal to be treated.

[00202] A compound of the invention, or pharmaceutically acceptable salt thereof, may be used on their own but will generally be administered in the form of a pharmaceutical composition in which the compounds of the invention, or pharmaceutically acceptable salt thereof, is in association with a pharmaceutically acceptable adjuvant, diluent or carrier.

[00203] Conventional procedures for the selection and preparation of suitable pharmaceutical formulations are described in, for example, "Pharmaceuticals - The Science of Dosage Form Designs", M. E. Aulton, Churchill Livingstone, 1988.

[00204] Depending on the mode of administration of the compounds of the invention, the pharmaceutical composition which is used to administer the compounds of the invention will preferably comprise from 0.05 to 99 % w/w compounds of the invention, more preferably from 0.05 to 80 % w/w compounds of the invention, still more preferably from 0.10 to 70 % w/w compounds of the invention, and even more preferably from 0.10 to 50 % w/w compounds of the invention (all percentages by weight being based on total composition).

[00205] The pharmaceutical compositions may be administered topically (e.g. to the skin) in the form, e.g., of creams, ointments, gels, lotions, solutions, suspensions; or systemically, e.g. by oral administration in the form of tablets, lozenges, hard or soft capsules, aqueous or oily suspensions, emulsions, dispersible powders or granules, syrups or elixirs; or by parenteral administration in the form of a sterile aqueous or oily solution, suspension or emulsion for injection (including intravenous, intracoronary, subcutaneous, intramyocardial, intraperitoneal, intramuscular, intravascular or infusion); by rectal administration in the form of suppositories or enemas; by inhalation for example as a finely divided powder or a liquid aerosol; or for administration by insufflation (for example as a finely divided powder).

[00206] For oral administration the compounds of the invention may be admixed with an adjuvant or a carrier, for example, lactose, saccharose, sorbitol, mannitol; a starch, for example, potato starch, corn starch or amylopectin; a cellulose derivative; a binder, for example, gelatine or polyvinylpyrrolidone; and/or a lubricant, for example, magnesium stearate, calcium stearate, polyethylene glycol, a wax, paraffin, and the like, and then compressed into tablets. If coated tablets are required, the cores, prepared as described above, may be coated with a concentrated sugar solution which may contain, for example, gum arabic, gelatine, talcum and titanium dioxide. Alternatively, the tablet may be coated with a suitable polymer dissolved in a readily volatile organic solvent. Thus, compositions intended for oral use may contain, for example, one or more colouring, sweetening, flavouring and/or preservative agents.

[00207] For the preparation of soft gelatine capsules, the compounds of the invention may be admixed with, for example, a vegetable oil or polyethylene glycol. Hard gelatine capsules may contain granules of the compound using either the above-mentioned excipients for tablets. Also liquid or semisolid formulations of the compound of the invention may be filled into hard gelatine capsules. Liquid preparations for oral application may be in the form of syrups or suspensions, for example, solutions containing the compound of the invention, the balance being sugar and a mixture of ethanol, water, glycerol and propylene glycol. Optionally such liquid preparations may contain colouring agents, flavouring agents, sweetening agents (such as saccharine), preservative agents and/or carboxymethylcellulose as a thickening agent or other excipients known to those skilled in art.

[00208] For intravenous (parenteral) administration the compounds of the invention may be administered as a sterile aqueous or oily solution.

[00209] The size of the dose for therapeutic or prophylactic purposes of a compound of the invention will naturally vary according to the nature and severity of the conditions, the concentration of the compound required for effectiveness in isolated cells, the concentration of the compound required for effectiveness in experimental animals, the age and sex of the animal or patient and the route of administration, according to well known principles of medicine.

[00210] Dosage levels, dose frequency, and treatment durations of compounds of the invention are expected to differ depending on the formulation and clinical indication, age, and co-morbid medical conditions of the patient.

[00211] An effective amount of a compound of the present invention for use in therapy of a condition is an amount sufficient to achieve symptomatic relief in a warm-blooded animal, particularly a human of the symptoms of the condition, to mitigate the physical manifestations of the condition, or to slow the progression of the condition.

[00212] The amount of active ingredient that is combined with one or more excipients to produce a single dosage form will necessarily vary depending upon the host treated and the particular route of administration. For example, a formulation intended for oral administration to humans will generally contain, for example, from 0.5 mg to 0.5 g of active agent (more suitably from 0.5 to 100 mg, for example from 1 to 30 mg) compounded with an appropriate and convenient amount of excipients which may vary from about 5 to about 98 percent by weight of the total composition.

[00213] For the above-mentioned compounds of the invention the dosage administered will, of course, vary with the compound employed, the mode of administration, the treatment desired and the disorder indicated. In using a compound of the invention for therapeutic or prophylactic purposes it will generally be administered so that a daily dose in the range, for example, a daily dose selected from 0.1 mg/kg to 100 mg/kg, 1 mg/kg to 75mg/kg, 1 mg/kg to 50 mg/kg, 1 mg/kg to 20 mg/kg or 5 mg/kg to 10 mg/kg body weight is received, given if required in divided doses. In general lower doses will be administered when a parenteral route is employed. Thus, for example, for intravenous or intraperitoneal administration, a dose in the range, for example, 0.1 mg/kg to 30 mg/kg body weight will generally be used. Similarly, for administration by inhalation, a dose in the range, for example, 0.05 mg/kg to 25 mg/kg body weight will be used. Suitably the compound of the invention is administered orally, for example in the form of a tablet, or capsule dosage form. The daily dose administered orally may be, for example a total daily dose selected from 1 mg to 1000 mg, 5 mg to 1000 mg, 10 mg to 750 mg or 25 mg to 500 mg. Typically, unit dosage forms will contain about 0.5 mg to 0.5 g of a compound of this invention.

[00214] The compounds of the invention may be administered along with other active compounds as part of a treatment regime. The other active compounds may be administered simultaneously with, subsequently to or previously to the administration of the compounds of the invention. It may be that the pharmaceutical formulation comprising the compounds of the invention also comprises one or more other active compounds. The other active compounds may be anticancer, anti-inflammatory, antibacterial, antiviral, antiemetic, antithrombotic or compounds that alter the metabolism.

[00215] Throughout the description and claims of this specification, the words “comprise” and “contain” and variations of them mean “including but not limited to”, and they are not intended to (and do not) exclude other moieties, additives, components, integers or steps. Throughout the description and claims of this specification, the singular encompasses the plural unless the context otherwise requires. In particular, where the indefinite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.

[00216] Features, integers, characteristics, compounds, chemical moieties or groups described in conjunction with a particular aspect, embodiment or example of the invention are to be understood to be applicable to any other aspect, embodiment or example described herein unless incompatible therewith. All of the features disclosed in this specification (including any accompanying claims, abstract and drawings), and/or all of the steps of any method or process so disclosed, may be combined in any combination, except combinations where at least some of such features and/or steps are mutually exclusive. The invention is not restricted to the details of any foregoing embodiments. The invention extends to any novel one, or any novel combination, of the features disclosed in this specification (including any accompanying claims, abstract and drawings), or to any novel one, or any novel combination, of the steps of any method or process so disclosed.

[00217] It will be understood that various modifications may be made to the embodiments disclosed herein. Therefore the above description should not be construed as limiting, but merely as exemplifications of preferred embodiments. For example, the functions described above and implemented as the best mode for operating the present invention are for illustration purposes only. Other arrangements and methods may be implemented by those skilled in the art without departing from the scope and spirit of this invention. Moreover, those skilled in the art will envision other modifications within the scope and spirit of the specification appended hereto.

[00218] The reader's attention is directed to all papers and documents which are filed concurrently with or previous to this specification in connection with this application and which are open to public inspection with this specification, and the contents of all such papers and documents are incorporated herein by reference.

EXAMPLES

Table of Abbreviations:

Purification Methods:

Purification by preparative HPLC (prep-HPLC) employed the following instruments and conditions:

A: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 19*150 mm, 5 pm particle size; Mobile phase, water+0.05% NH ₃ .H ₂ O/acetonitrile, linear gradient: 80:20 to 40:60 over 8 min, flow rate: 25 mLmin ^-1 .

B: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH F-Phenyl OBD column, 19*150 mm, 5 pm particle size; Mobile phase, water+0.1 %FA /acetonitrile, linear gradient: 3:97 to 11 :89 over 8 min, flow rate: 25 mLmin ^-1 .

C: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Prep OBD C18 Column, 30*150 mm, 5 pm particle size; Mobile phase, water +10 mmol/L NH4HCO3+0.1 %NH3.H2O/MeOH; linear gradient from 66:34 to 38:62 in 8 min, flow rate: 25 mLmin ^-1 .

D: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shieid RP18 OBD Column 30 *150 mm 5 pm; Mobile phase, water+10 mmol/L NH^COs/acetonitrile, linear gradient from 75:25 to 60:40 over 7 min, flow rate: 60 mLmin' ¹ .

E: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Sunfire prep C18 Column 30 *150 mm 5 pm; Mobile phase, water+0.1% TFA/acetonitrile, linear gradient from 93:7 to 80:20 over 8 min, flow rate: 60 mLmin' ¹ .

F: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XB-C18 Column 50 *250 mm 10 pm; Mobile phase, water+0.1 % FA/acetonitrile, linear gradient from 62:38 to 42:58 over 20 min, flow rate: 100 mLmin' ¹ .

G: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column 30*150 mm 5 pm; Mobile phase, Water+0.1 %FA/ acetonitrile; linear gradient, 74:26 to 42:58 in 7 min, flow rate: 60 mLmin' ¹ .

H: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column 30*150 mm 5 pm; Mobile phase, Water+0.1 %FA/ acetonitrile; linear gradient, 95:5 to 80:20 in 7 min, flow rate: 60 mLmin' ¹ .

I: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column 30*150 mm 5 pm; Mobile phase, Water+0.1 %FA/ acetonitrile; linear gradient, 98:2 to 92:8 in 7 min, flow rate: 60 mL/ min ^-1 .

J: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column 30*150 mm 5 pm; Mobile Phase: Water+0.05%TFA/acetonitrile; linear gradient, 90:10 to 50:50 in 7 min, flow rate: 60 mL/ min ^-1 .

K: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+10 mmol/L NH4HCOs+0.1%NH3 H2O /acetonitrile, liner gradient: 75:25 to 45:55 over 8 min, flow rate: 60 mLmin' ¹ . L: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+0.05%TFA/MeOH, liner gradient: 84:16 to 57:43 over 7 min, flow rate: 60 mLmin' ¹ .

M: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 50*250 mm, 5 pm particle size; Mobile phase, water+0.1 %FA I MeOH, liner gradient: 80:20 to 50:50 over 7 min, flow rate: 60 mLmin' ¹ .

N: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+0.1 %FA I acetonitrile, liner gradient: 95:5 to 85:15 over 7 min, flow rate: 60 mLmin' ¹ .

O: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XB-C18 Column, 50*250 mm, 10 pm particle size; Mobile phase, 5 mmol NH4HCO3 + water/acetonitrile, linear gradient: 67:33 to 47:53 over 20 min, flow rate: 100 mLmin' ¹ .

P: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+0.05 %TFA /acetonitrile, liner gradient: 82:18 to 58:42 over 7 min, flow rate: 60 mLmin' ¹ .

Q: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xbridge Shied RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+10 mmol/L NH4HCO3 /acetonitrile, liner gradient: 85:15 to 55:45 over 7 min, flow rate: 60 mLmin' ¹ .

R: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a YMC-Actus Triart C18, 30*150 mm, 5 pm particle size; Mobile Phase, water+0.1 %FA/acetonitrile, linear gradient from 98:2 to 83:17 in 7 min, flow rate: 60 mLmin' ¹ .

S: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD, 30*150 mm, 5 pm particle size; Mobile Phase, water+0.05%TFA/MeOH, linear gradient from 83:17 to 57:43 in 7 min, flow rate: 60 mLmin' ¹ .

T: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column, 30*150mm, 5 pm particle size; Mobile phase, water+0.1 %FA /acetonitrile, linear gradient: 95:5 to 70:30 over 7 min, flow rate: 60 mLmin' ¹ .

U: Preparative HPLC separations were performed with a Gilson HPLC (321 pumps, 819 injection module, 215 liquid handler/injector) connected to a Gilson 155 UV/vis detector. HPLC chromatographic separations were conducted using a Waters XBridge C18 column, 19 x 100 mm, 5 pm particle size; mobile phase, water I acetonitrile +0.1% NH3; linear gradient 95:5 to 5:95 over 6.5min and then held for 1min; flow rate 18 mLmin' ¹ .

V: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water + 10 mmol/L NH4HCO31 acetonitrile, liner gradient: 85:15 to 75:25 over 7 min, flow rate: 60 mLmin' ¹ .

W: Preparative HPLC separations were performed with a Biotage isolera. HPLC chromatographic separations were conducted using C18 silica gel; Mobile phase, water+0.1% FA/acetonitrile, liner gradient: 95:5 to 0:100 over 20 min, flow rate: 60 mLmin' ¹ . X: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Xselect CSH C18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+0.05% HCI/acetonitrile, linear gradient: 98:2 to 88:12 over 7 min, flow rate: 60 mLmin' ¹ .

Y: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Prep C18 OBD Column, 30*100 mm, 5 pm particle size; Mobile phase: water+0.1% NH3 H2O/ acetonitrile, linear gradient: 90:10 to 72:28 over 7 min; flow rate: 60 mLmin' ¹ .

Z: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water + 0.05%TFA/ MeOH, liner gradient: 79:21 to 70:30 over 9 min, flow rate: 60 mLmin' ¹ . AA: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Prep C18 OBD Column, 30*100 mm, 5 pm; Mobile phase: water+10 mmol/L NH4HCO3/ MeOH, linear gradient: 55:45 to 45:55 over 8 min; flow rate: 60 mLmin ^-1 .

AB: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a YMC-Actus Triart C18, 30*150 mm, 5 pm particle size; Mobile Phase, water+10 mmol/L NH^COs/acetonitrile, linear gradient from 94:6 to 74:26 in 8 min, flow rate: 60 mLmin ^-1 .

AC: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a Sunfire prep C18 Column 30 *150 mm 5 pm; Mobile phase, water+0.05% TFA/acetonitrile, linear gradient from 92:8 to 78:22 over 8 min, flow rate: 60 mLmin ^-1 .

AD: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a YMC-Actus Triart C18, 30*150 mm, 5 pm particle size; Mobile Phase, water+10 mmol/L NH4HCO3+0.1% NH ₃ H ₂ O/acetonitrile, linear gradient from 70:30 to 53:47 in 8 min, flow rate: 60 mLmin ^-1 .

AE: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a YMC-Actus Triart C18, 30*150 mm, 5 pm particle size; Mobile Phase, water+10 mmol/L NH4HCO3+0.1% NH ₃ H ₂ O/acetonitrile, linear gradient from 77:33 to 50:50 in 9 min, flow rate: 60 mLmin ^-1 .

AF: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Prep C18 OBD Column, 30*100 mm, 5 pm; Mobile phase: water+0.05% NH3 H2O/ acetonitrile, linear gradient: 98:2 to 83:17 over 10 min; flow rate: 60 mLmin ^-1 .

AG: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Shield RP18 OBD Column, 30*150 mm, 5 pm particle size; Mobile phase, water+10 mmol/L NH4HCO3+0.1%NH3 H2O /acetonitrile, liner gradient: 98:2 to 80:20 over 9 min, flow rate: 60 mLmin ^-1 .

AH: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a XBridge Prep C18 OBD Column, 30*100 mm, 5 pm; Mobile phase: water+10 mmol/L NH4HCO3/ MeOH, linear gradient: 66:34 to 48:52 over 10 min; flow rate: 60 mLmin ^-1 .

Al: Preparative HPLC separations were performed with a Waters 2545 Binary Gradient Module connected to a Waters 2489 UV/visible detector. HPLC chromatographic separations were conducted using a YMC-Actus Triart C18, 30*150 mm, 5 pm particle size; Mobile Phase, water+10 mmol/L NH4HCC>3+0.1% NHs ^O/acetonitrile, linear gradient from 69:31 to 55:45 in 9 min, flow rate: 60 mLmin ^-1 .

Analytical Methods:

¹ H NMR spectra were recorded on Bruker AVANCE III HD 300, Bruker AVANCE NEO 400 or Bruker AVANCE II 500 spectrometers. Chemical shifts are denoted in ppm (5) relative to residual protonated solvent as an internal standard as described in, for example, Gottlieb et al. Journal of Organic Chemistry (1997) 62 7512. The splitting pattern for NMR spectra are denoted as follows: s (singlet), br (broad), d (doublet), t (triplet), m (multiplet) or combinations thereof. Coupling constants ( ) are designated in Hz and reported to one decimal place.

Liquid chromatography-mass spectra were recorded using the following systems and running conditions:

A: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.05% TFA/acetonitrile +0.05% TFA, linear gradient from 95:5 to 30:70 over 1.90 min and then linear gradient to 0:100 over 0.35 min, held for 0.35 min; flow rate: 1.5 mLmin ^-1 .

B: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.05 %TFA/acetonitrile+0.05 %TFA, linear gradient from 95:5 to 40:60 over 1.90 min and then linear gradient to 0:100 over 0.35 min, held for 0.35 min; flow rate: 1.5 mLmin ^-1 .

C: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.1 %FA/acetonitrile+0.1%FA, linear gradient from 95:5 to 30:70 over 1.79 min and then linear gradient to 0:100 over 0.20 min, held for 0.7 min; flow rate: 1.5 mLmin ^-1 .

D: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.1 %FA/acetonitrile+0.1 %FA, linear gradient from 95:5 to 30:70 over 1.79 min, then linear gradient to 0:100 over 0.20 min, and then held for 0.70 min. flow rate: 1.50 mLmin ^-1 .

E: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.1 %FA/acetonitrile+0.1 %FA, linear gradient from 95:5 to 30:70 over 1.79 min, then linear gradient to 0:100 over 0.20 min, and then held for 0.70 min. flow rate: 1.50 mLmin' ¹ .

F: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 urn 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2 min; flow rate: 1.0 mLmin' ¹ .

G: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.05 %TFA/acetonitrile +0.05 %TFA, linear gradient to 0:100 over 2:00 min and then held for 0.70 min; flow rate 1.5 mLmin' ¹ .

H: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, Water+ 0.05 %TFA/Acetonitrile+0.05 %TFA, linear gradient from 95:5 to 40:60 over 1.90 min and then linear gradient to 0:100 over 0.35 min, held for 0.35 min; flow rate: 1.5 mLmin' ¹ .

I: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 urn 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2 min; flow rate: 1.0 mLmin' ¹ .

J: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 urn 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2 min; flow rate: 1.0 mLmin' ¹ .

K: Shimadzu LCMS-2020 connected to Ascentis Express C18 50 mmx3.0 nm. mobile phase, water+0.1 %FA/acetonitrile+0.1 %FA, linear gradient from 95:5 to 30:70 over 1.80 min, and then linear gradient to 0:100 over 0.2 min, and then held for 0.70 min. flow rate: 1.50 mLmin' ¹ .

L: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 urn 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2 min; flow rate: 1.0 mLmin' ¹ .

M: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 urn 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2 min; flow rate: 1.0 mLmin' ¹ .

N: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.1 %TFA/ acetonitrile+0.1 %TFA, linear gradient from 95:5 to 40:60 over 2.00 min, then linear gradient to 0:100 over 0.20 min, and then held for 0.40 min. flow rate: 1.50 mLmin' ¹ .

O: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.1 %TFA/acetonitrile+0.1 %TFA, linear gradient from 95:5 to 40:60 over 2.00 min, then linear gradient to 0:100 over 0.20 min, and then held for 0.40 min. flow rate: 1.50 mLmin' ¹ . P: Shimadzu LCMS-2020 connected to Atlantis™ T3 3 um 4.6x100 mm. mobile phase, water+0.1 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 8 min and held for 2.0 min; flow rate: 1.0 mLmin' ¹ .

Q: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.05 %TFA/acetonitrile +0.05 %TFA, 95:5 for 0 min, then linear gradient to 40:60 over 1.90 min and then linear gradient to 0:100 for 0.35 min; flow rate 1.5 mLmin' ¹ .

R: Shimadzu LCMS-2020 connected to Ascentis Express 90 A 10 cm x 4.6 mm. mobile phase, water+0.05 %TFA /acetonitrile, linear gradient from 95:5 to 5:95 over 12 min; flow rate: 1.5 mLmin' ¹ .

S: Shimadzu LCMS-2020 connected to Ascentis Express C18 30 mmx3.0 nm. mobile phase, water+0.05%TFA/acetonitrile +0.05%TFA, linear gradient from 95:5 to 40:60 over 1.90 min and then linear gradient to 0:100 over 0.35 min, held for 0.35 min; flow rate: 1.5 mLmin' ¹ .

T: Shimadzu LCMS-2020 connected to Ascentis Express 90 A C18 4.6 mm x 100 mm. mobile phase, water+0.1 %FA /acetonitrile+0.1 % FA, linear gradient from 95:5 to 5:95 over 6 min and held for 2 min; flow rate: 0.8 mLmin' ¹ .

U: Shimadzu LCMS-2020 connected to Ascentis Express C18 50 mmx3.0 nm. mobile phase, water+0.05%TFA/acetonitrile+0.05%TFA, linear gradient from 95:5 to 30:70 over 1 .5 min, and then linear gradient to 0: 100 over 1 .5 min. flow rate: 1.50 mLmin' ¹ .

V: Agilent Technologies 1200 series HPLC connected to an Agilent Technologies 6130 spectrometer using a Waters Xbridge C18 column, 50 mm x 2.1 mm, 3.5 pm particle size; mobile phase, water/acetonitrile +0.1% HCOOH; 80:20 for 0.5min then linear gradient to 5:95 over 3.0min and then held for 1.5min; flow rate 0.5 mLmin' ¹ .

W: Agilent HPLC 1100 series connected to a Bruker Daltonics MicrOTOF spectrometer using a Waters Xbridge C18 column, 50 mm x 2.1 mm, 3.5 pm particle size; mobile phase, water/acetonitrile + 0.1% NH3; 95:5 for 0.5min then linear gradient to 5:95 over 3.5min and then held for 2.0min; flow rate 0.5 mLmin' ¹ .

X: Agilent Technologies 1200 series HPLC connected to an Agilent Technologies 6130 spectrometer using a Waters Xbridge C18 column, 50 mm x 2.1 mm, 3.5 pm particle size; mobile phase, water/acetonitrile +0.1% NH3; 80:20 for 0.5min then linear gradient to 5:95 over 3.0min and then held for 1.5min; flow rate 0.5 mLmin' ¹ .

Y: Agilent Technologies 1200 series HPLC connected to an Agilent Technologies 6130 spectrometer using a Waters Xbridge C18 column, 50 mm x 2.1 mm, 3.5 pm particle size; mobile phase, water/acetonitrile +0.1% NH3; 80:20 for 0.5min then linear gradient to 5:95 over 5.0min and then held for 1.5min; flow rate 0.5 mLmin' ¹ .

Z: Shimadzu LCMS-2020 connected to HALO, 50 mmx3.0 mm, 4.0 pm particle size. Mobile phase, water+0.1 % FA/acetonitrile+0.1% FA, linear gradient from 95:5 to 40:60 over 1.80 min, and then linear gradient to 0:100 over 0.2 min and then held for 0.70 min. flow rate: 1.50 mLmin ^-1 .

AA: Shimadzu LCMS-2020 connected to EC-C18, 30 mmx3.0 mm, 1.9 pm particle size. Mobile phase, water+0.05%TFA/acetonitrile+0.05%TFA, linear gradient from 95:5 to 40:60 over 1.80 min, , then linear gradient to 0:100 over 0.2 min, and then held for 0.70 min. flow rate: 1.50 mLmin' ¹ .

AB: Shimadzu LCMS-2020 connected to HPH, 50 mmx3.0 mm, 4.0 pm particle size. Mobile phase, water+0.1% FA/acetonitrile+0.1 % FA, linear gradient from 100:0 to 0:100 over 1.20 min, and then held for 0.60 min. flow rate: 1.50 mLmin' ¹ .

AC: Shimadzu LCMS-2020 connected to HPH, 50 mmx3.0 mm, 2.7 pm particle size. Mobile phase, water+0.04% NH ₃ H ₂ O/acetonitrile, linear gradient from 100:0 to 30:70 over2.20 min, and linear gradient to 5:95 over 0.20 min, then held for 0.40 min. flow rate: 1.50 mLmin ^-1 .

AD: AC: Shimadzu LCMS-2020 connected to HPH, 50 mmx3.0 mm, 2.7 pm particle size. Mobile phase, water+0.04% NH ₃ H ₂ O/acetonitrile, linear gradient from 100:0 to 30:70 over 2.00 min, and linear gradient to 5:95 over 0.20 min, then held for 0.40 min. flow rate: 1 .50 mLmin ^-1 .

AE: Shimadzu LCMS-2020 connected to Ascentis Express C18 50 mmx3.0 mm. mobile phase, water+0.05%TFA/acetonitrile+0.05%TFA, linear gradient from 100:0 to 30:70 over 1.90 min, and then linear gradient to 0:100 over 0.35 min, and then held for 0.35 min. flow rate: 1.50 mLmin ^-1 .

AF: Shimadzu LCMS-2020 connected to Ascentis Express C18 50 mmx3.0 mm. mobile phase, water+0.05%TFA/acetonitrile+0.05%TFA, linear gradient from 100:0 to 0:100 over 2.00 min, and then held for 0.70 min. flow rate: 1.50 mLmin ^-1 .

AG: Shimadzu LC-MS 2020 with photodiode array detector using a Thermo Hypersil Gold C18 column, 50mm x 2.1 mm 1.9pm particle size; mobile phase water/acetonitrile +0.1% formic acid; 95:5 for 0.2min then linear gradient to 5:95 over 2.0min and then held for 0.5min; flow rate 0.8 mLmin ^-1 .

Process for Preparation:

[00219] Certain compounds of the invention may be synthesised according to the general methods disclosed herein. Certain compounds of the invention may be synthesised according to general schemes 1 to 4. Certain compounds of the invention may be synthesised according to or analogously to the syntheses provided in examples 1 to 54 and 97 to 110. General Scheme 1 - Synthesis of Intermediates A and B

Synthetic routes to Intermediates A and B have been reported in, for example, W02003026657 A1. Alternative routes to Intermediates A and B are illustrated in General Scheme 1 and described in the following examples.

General Scheme 2 - Synthesis of Intermediates C

Synthetic routes to Intermediate C have been reported in, for example, WO2021228945 A1.

A route to Intermediate C is illustrated in General Scheme 2 and described in the following examples.

General Scheme 3 Compounds of Formula (I) can be synthesised from Intermediate A or B in one to two steps as illustrated in General Scheme 3. First, the reaction of the Intermediate A with an appropriate nucleophile (for example an amine or hydroxylamine), using an appropriate base if necessary (for example triethylamine, DI PEA, potassium carbonate), in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane, ethanol, methanol) with heating (conventional or by microwave irradiation) if necessary.

Subsequently, alkylation can be achieved by using an appropriate reagent (for example an alkyl halide, mesylate, tosylate or cyanogen halide), with an appropriate base (for example triethylamine, DIPEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

Acylation can be achieved by using an appropriate reagent (for example an acyl halide or anhydride), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary. Acylation may also be achieved by activating a carboxylic acid with common coupling reagents (for example EDC, HATLI, T3P), with an appropriate base (for example triethylamine, DIPEA, DMAP) in an appropriate solvent (for example DCM DMF, NMP, THF, ethyl acetate) with heating (conventional or by microwave irradiation) if necessary.

Sulfonylation can be achieved by using an appropriate reagent (for example a sulfonyl halide), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

Ureas may be formed by using an appropriate reagent (for example an isocyanate), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary. Ureas may also be formed by first activating an amine with a reagent such as GDI, phosgene or triphosgene with an appropriate base if necessary (for example triethylamine, DI PEA, potassium carbonate, DMAP), in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

General Scheme 4 Compounds of Formula (I) can be synthesised from Intermediate C in a single step as illustrated in General Scheme 4. Alkylation can be achieved by using an appropriate reagent (for example an alkyl halide, mesylate, tosylate or cyanogen halide), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

Acylation can be achieved by using an appropriate reagent (for example an acyl halide or anhydride), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary. Acylation may also be achieved by activating a carboxylic acid with common coupling reagents (for example EDC, HATU, T3P), with an appropriate base (for example triethylamine, DIPEA, DMAP) in an appropriate solvent (for example DCM DMF, NMP, THF, ethyl acetate) with heating (conventional or by microwave irradiation) if necessary.

Sulfonylation can be achieved by using an appropriate reagent (for example a sulfonyl halide), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

Ureas may be formed by using an appropriate reagent (for example an isocyanate), with an appropriate base (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary. Ureas may also be formed by first activating an amine with a reagent such as GDI, phosgene or triphosgene with an appropriate base if necessary (for example triethylamine, DI PEA, potassium carbonate, DMAP) in an appropriate solvent (for example DCM, DMSO, DMF, NMP, THF, 1 ,4-dioxane) with heating (conventional or by microwave irradiation) if necessary.

Intermediate 1 : N-(4-methylquinazolin-2-yl) cyanamide

To a mixture of 1-(2-aminophenyl)ethan-1-one hydrochloride (29.0 g, 0.168 mol) in EtOH (290 mL) was added sodium dicyanoazanide (37.6 g, 0.420 mol). The resulting mixture was stirred at 80 °C for overnight. The precipitated solids were collected by filtration and washed with water (100 mLx3). The filter cake was washed with ethyl ether (3 x 100 ml) and dried under vacuum to give N-(4-methylquinazolin-2-yl) cyanamide, Intermediate 1 (11.0 g, 35%) as a yellow solid. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 12.9 (br, 1 H), 8.11 - 8.09 (m, 1 H), 7.88 - 7.83 (m, 1 H), 7.69 - 7.44 (m, 2H), 2.81 (s, 3H). LCMS ^e : Rt 0.79 min, MH ⁺ 185.

Intermediate 2: N-(Quinazolin-2-yl)cyanamide

Sodium dicyanamide (735 mg, 8.25 mmol) and 2-aminobenzaldehyde hydrochloride (1.30 g, 8.25 mmol) were heated at reflux in ethanol (15 mL). After 2h, the mixture was cooled and filtered. The collected solid was washed with ethanol, water and finally ethanol. The resulting yellow solid was lyophilised to give 2-cyanoamidoquinazoline (168 mg, 12%). ¹ H NMR (500 MHz, DMSO-d ₆ ) δ 13.15 (br s, 1 H), 9.42 (s, 1 H), 8.04 (d, 1 H, J = 7.8 Hz), 7.91 (t, 1 H, J = 7.5 Hz), 7.55 (d, 1 H, J = 8.4 Hz), 7.47 (t, 1 H, J = 7.6 Hz).

Intermediate 3: 1-(4-methylquinazolin-2-yl) guanidine (TLN468)

To a stirred solution of 2,2,4-trimethyl-1 H-quinoline (200 mg, 1.15 mmol) in aq. HCI (0.4 mL, 2.0 M) was added 2-cyanoguanidine (110 mg, 1 .27 mmol). The resulting mixture was stirred for 2 h at 100 °C. The precipitated solids were then collected by filtration and washed with water (2.0 mL) and EtOH (2.0 mL). The crude product (150 mg) was purified by Prep-HPLC ^a to afford 1-(4-methylquinazolin-2-yl) guanidine (46 mg, 20 %) as a white solid. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 7.95 (dd, J = 8.2, 1.4 Hz, 1 H), 7.68 - 7.65 (m, 1 H), 7.56 - 7.50 (m, 1 H), 7.29 - 7.25 (m, 1 H), 7.11 (br, 3H), 2.71 (s, 3H). LCMS ^a : Rt 0.73 min, MH ⁺ 202.

Example 1 : 1-(2-(Dimethylamino)ethyl)-3-(4-methylquinazolin-2-yl)guanid ine

Intermediate 1 (100 mg, 0.543 mmol) and /V,/V-dimethylethylenediamine (63 pl, 0.585 mmol) were mixed in ethanol (3 mL) and heated for 15 min at 160 °C. The resulting mixture was evaporated. The residue was purified by flash column chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to afford 1-(2-(dimethylamino)ethyl)-3-(4- methylquinazolin-2-yl)guanidine (54 mg, 36%). LCMSw: Rt 4.5 min, MH ⁺ 273. Example 2: 1 -(4-Methylquinazolin-2-yl)-3-(2-(pyridin-3-yl)ethyl)guanidin e

Intermediate 1 (100 mg, 0.543 mmol), 3-(2-aminoethyl) pyridine hydrochloride (91 mg, 0.574 mmol) and triethylamine (84 pl, 0.603 mmol) were mixed in ethanol (3 mL) and heated for 15 min at 160 °C. The resulting mixture was evaporated. The residue was purified by chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give 1-(4- methylquinazolin-2-yl)-3-(2-(pyridin-3-yl)ethyl)guanidine (133 mg, 80%). LCMSw: Rt 4.2 min, MH ⁺ 307.

Example 3: 1 -(1 -Methylpiperidin-4-yl)-3-(4-methylquinazolin-2-yl)guanidine

To a solution of Intermediate 1 (180 mg, 0.978 mmol) in EtOH (2.0 mL) was added 1- methylpiperidin-4-amine (223 mg, 1 .96 mmol). The reaction mixture was stirred at 80 °C for 3 days. The reaction mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC ^C to afford 1-(1-methylpiperidin-4-yl)-3-(4-methylquinazolin-2- yl)guanidine (90 mg, 30%) as a light yellow solid. ¹ H NMR (400 MHz, DMSO-cfe) δ 7.96 (d, J = 8.2 Hz, 1 H), 7.68 - 7.56 (m, 2H), 7.52 (d, J = 8.4 Hz, 1 H), 7.28 (t, J = 7.5 Hz, 1 H), 3.75 (s, 1 H), 2.71 - 2.66 (m, 5H), 2.17 (s, 3H), 2.07 - 2.02 (m, 2H), 1.91 - 1.80 (m, 2H), 1.45 - 1 .44 (m, 2H). LCMS ^d : Rt 0.62 min, MH ⁺ 299.

Example 4: N-((( 1 -methylpiperidin-4-yl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

To a solution of Example 3 (37 mg, 0.124 mmol) in DMSO (1.0 mL) was added AC2O (12.6 mg, 0.124 mmol). The resulting mixture was stirred for overnight at room temperature. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC ^d to afford N-(((1-methylpiperidin-4-yl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide (20 mg, 45%) as a white solid. ¹ H NMR (300 MHz, Chloroform-d, mix of rotamers) δ 14.50 (s, 0.5H), 13.07 (s, 0.4H), 10.44 (s, 0.3H), 8.84 (s, 0.5 H), 8.07 - 7.86 (m, 1 H), 7.80 - 7.78 (m, 0.4 H), 7.74 - 7.68 (m, 1 H), 7.63 - 7.60 (m, 0.5H), 7.50 - 7.44 (m, 0.5H), 7.40 - 7.35 (m, 0.5H), 4.42 - 4.41 (m, 0.6H), 4.27 - 4.17 (m, 0.4H), 2.90 (s, 1.6H), 2.87(s, 1.4H), 2.82 - 2.66 (m, 2H), 2.36 (s, 1.5H), 2.32 -2.30 (m, 5H), 2.17- 2.08 (m, 3.5H), 1.87 - 1.65 (m, 2.0H). LCMS ^e : Rt 0.604 min, MH ⁺ 341 .

Example 5: 1 -(4-Methylquinazolin-2-yl)-3-(2-(piperidin-1 -yl)ethyl)guanidine

Intermediate 1 (100 mg, 0.543 mmol) and 1-(2-aminoethyl)piperidine (0.19 mL, 1.33 mmol) were mixed in ethanol (4 mL) and heated for 10 min at 160 °C. The resulting mixture was evaporated. The residue was purified by chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give 1-(4-methylquinazolin-2-yl)-3-(2-(piperidin-1- yl)ethyl)guanidine (74 mg, 44%). LCMS ^V : Rt 0.33 min, MH ⁺ 313.

Example 6: 1 -(4-Methylquinazolin-2-yl)-3-(3-(piperidin-1 -yl)propyl)guanidine

By analogous method to that described for Example 1 , with 1-(3-aminopropyl)piperidine (91 μL) as the nucleophilic amine. Afforded 1-(4-methylquinazolin-2-yl)-3-(3-(piperidin-1- yl)propyl)guanidine (46 mg, 26%). LCMSw: Rt 4.9 min, MH ⁺ 327.

Example 7: 1-(1-Methylpiperidin-3-yl)-3-(4-methylquinazolin-2-yl)guanid ine

Intermediate 1 (125 mg, 0.68 mmol), 3-amino-1-methylpiperidine dihydrochloride (191 mg 1.02 mmol) and DI PEA (0.20 mL, 1.15 mmol) were mixed in ethanol (4 mL) and heated for 10 min at 160 °C. The resulting mixture was evaporated. The residue was purified by chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give racemic 1-(1-methylpiperidin-3-yl)-3-(4-methylquinazolin-2-yl)guanid ine (118 mg, 58%). LCMS ^V : Rt 0.4 min, MH ⁺ 299. Example 8 1 -(3-(Dimethylamino)propyl)-3-(4-methylquinazolin-2-yl)guanid ine

Intermediate 1 (200 mg, 1.09 mmol) in ethanol (3 mL) had dimethylaminopropylamine (444 mg, 4.34 mmol) added at room temperature. The resulting mixture was stirred for overnight at 80 °C. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC ^e to afford 1-(3-(dimethylamino)propyl)-3-(4- methylquinazolin-2-yl)guanidine (128 mg, 22%) as a yellow solid TFA salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 11.22 (s, 1 H), 9.97 (s, 1H), 9.62 (s, 1 H), 8.88 (s, 2H), 8.28 (d, J = 8.3 Hz, 1 H), 8.09 - 7.88 (m, 2H), 7.68 - 7.66 (m, 1 H), 3.52 - 3.50 (m, 2H), 3.29 - 3.08 (m, 2H), 2.94 (s, 3H), 2.82 (d, J = 4.4 Hz, 6H), 2.13 - 1.83 (m, 2H). LCMS ^f : Rt 3.21 min, MH ⁺ 287.

Example 9 N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

Example 8 (348 mg, 1.21 mmol), triethylamine (0.37 mL, 2.65 mmol) and 4- (dimethylamino)pyridine (16 mg, 0.13 mmol) were dissolved in DCM (15 mL) and cooled in an ice bath. Acetyl chloride (0.10 mL) was added drop-wise and the mixture stirred for 2 h. The mixture was evaporated and the residue purified by chromatography by chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give N-(((3- (dimethylamino)propyl)amino)((4-methylquinazolin-2-yl)amino) methylene)acetamide (326 mg, 82%). LCMS ^V : Rt 0.3 min, MH ⁺ 329.

Example 10: 1-(4-Methylquinazolin-2-yl)-3-(2-(pyrrolidin-1-yl)ethyl)guan idine

By analogous method to that described for Example 1, with 1-(2- aminoethyl)pyrrolidine (73 μL) as the nucleophilic base. Afforded 1-(4-methylquinazolin-2-yl)-3-(2-(pyrrolidin-1- yl)ethyl)guanidine (30 mg, 19%). LCMS ^V : Rt 0.5 min, MH ⁺ 299. Example 11 : 1 -(2-( 1 -Methylpyrrolidin-2-yl)ethyl)-3-(4-methylquinazolin-2-yl)gua nidine

By analogous method to that described for Example 1 , with 2-(2-aminoethyl)-1- methylpyrrolidine (83 μL) as the nucleophilic base. Afforded racemic 1-(2-(1- methylpyrrolidin-2-yl)ethyl)-3-(4-methylquinazolin-2-yl)guan idine (58 mg, 34%). LCMSw: Rt 4.5 min, MH ⁺ 313.

Example 12: N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)cyclobutanecarboxamide

Example 8 (75 mg, 0.26 mmol), triethylamine (73 pl, 0.52 mmol) and 4- (dimethylamino)pyridine (3 mg, 0.02 mmol) were dissolved in 1 ,4-dioxane (4 mL). Cyclobutanecarbonyl chloride (26 pl, 0.23 mmol) added drop-wise and the mixture stirred for 2 h. The mixture was evaporated and the residue purified by chromatography (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give /V-(((3- (dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)cyclobutanecarboxamide (45 mg, 48%). LCMS ^W : Rt 5.7 min, MH ⁺ 369. Example 13: N-((( 1 -methylpiperidin-4-yl)amino)((4-methylquinazolin-2- yl)amino)methylene)cyclobutanecarboxamide

By analogous method to that described for Example 12, using Example 2. Afforded /V-(/V-(1- methylpiperidin-4-yl)-/V-(4-methylquinazolin-2-yl)carbamimid oyl)cyclobutanecarboxamide

(37 mg, 78%). LOVIS'": Rt 5.4 min, MH 381. Example 14 1-(8-Methyl-8-azabicyclo[3.2.1]octan-3-yl)-3-(4-methylquinaz olin-2- yl)guanidine

By analogous method to that described for Example 1 , with 8-methyl-8- azabicyclo[3.2.1]octan-3-amine (81 L) as the nucleophilic base. Afforded 1-(8-methyl-8- azabicyclo[3.2.1]octan-3-yl)-3-(4-methylquinazolin-2-yl)guan idine (87 mg, 49%). LCMS ^W : Rt 4.3 min, MH ⁺ 325.

Example 15 1-(4-Methylquinazolin-2-yl)-3-(3-morpholinopropyl)guanidine

Intermediate 1 (200 mg, 1.09 mmmol) in ethanol (3 mL) had 4-morpholinepropanamine (626 mg, 4.34 mmol) at room temperature .The resulting mixture was stirred for overnight at 80 °C. The resulting mixture was concentrated under reduced pressure. The crude product (200 mg) was purified by Prep-HPLC ^k to afford 1-(4-methylquinazolin-2-yl)-3-(3- morpholinopropyl)guanidine (86 mg, 23%) as a white solid. ¹ H NMR (400 MHz, Chloroform- d) δ 8.00 - 7.81 (m, 1 H), 7.67 - 7.62 (m, 1 H), 7.61 - 7.52 (m, 1 H), 7.35 - 7.30 (m, 1 H), 3.76 - 3.74 (m, 4H), 3.44 - 3.41 (m, 2H), 2.83 (s, 3H), 2.50 - 2.48 (m, 6H), 1.84 - 1.82 (m, 2H). LCMS ¹ : Rt 3.39 min, MH ⁺ 329.

Example 16 1-(4-Methylquinazolin-2-yl)-3-(pyridin-4-ylmethyl)guanidine

Intermediate 1 (200 mg, 1.09 mmol) in ethanol (3 mL) had 4-pyridinemethaneamine (470 mg, 4.34 mmol) added at room temperature. The resulting mixture was stirred for overnight at 80 °C. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC ^h to afford 1-(4-methylquinazolin-2-yl)-3-(pyridin-4- ylmethyl)guanidine (170 mg, 28%) as a yellow solid TFA salt. ¹ H NMR (400 MHz, DMSO- d ⁶ ) δ 11.35 (s, 1 H), 9.94 (s, 1 H), 8.97 (s, 2H), 8.73 - 8.65 (m, 2H), 8.29 (d, J = 8.3 Hz, 1 H), 8.01 (d, J = 3.8 Hz, 2H), 7.67 - 7.66 (m, 3H), 4.91 (d, J = 6.5 Hz, 2H), 2.95 (s, 3H). HPLC: Rt 3.27 min, MH ⁺ 293. Example 17 1-(4-Methylquinazolin-2-yl)-3-(2-(pyridin-4-yl)ethyl)guanidi ne

Intermediate 1 (100 mg, 0.55 mmol) and 4-(2-aminoethyl)pyridine (69 μL) were mixed in ethanol (3 mL) and heated for 15 min at 160 °C. The resulting mixture was evaporated. The residue was purified (Biotage®KP-NH silica) eluted with methanol I DCM (0-5%) to give 1- (4-methylquinazolin-2-yl)-3-(2-(pyridin-4-yl)ethyl)guanidine (97 mg, 58%). LCMS ^W : Rt 4.2 min, MH ⁺ 307.

Example 18 1-(2-(Diethylamino)ethyl)-3-(4-methylquinazolin-2-yl)guanidi ne

Intermediate 1 (125 mg, 0.679 mmol) and A/,N-diethylethylenediamine (0.14 mL, 1.00 mmol) were added to ethanol (4 mL) and heated for 10 min at 160 °C. The resulting mixture was evaporated. The residue was dissolved in diethyl ether and allowed to stand overnight. The resulting mixture was filtered and washed with diethyl ether. The collected solid was lyophilized to give 1-(2-(diethylamino)ethyl)-3-(4-methylquinazolin-2-yl)guanidi ne (129 mg, 63%). LCMS ^V : Rt 0.3 min, MH ⁺ 301.

Example 19 N-(((2-(diethylamino)ethyl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

Example 18 (50 mg, 0.17 mmol), triethylamine (69 μL, 0.50 mmol) and 4- (dimethylamino)pyridine (2 mg, 0.01 mmol) were dissolved in 1 ,4-dioxane (2 mL). Acetyl chloride (15 μL, 0.21 mmol) was added drop-wise and the mixture stirred 2h. The mixture was evaporated and the residue purified by preparative HPLC ^e to give N-(((2- (diethylamino)ethyl)amino)((4-methylquinazolin-2-yl)amino)me thylene)acetamide (57 mg, 98%). LCMS ^V : Rt 0.3 min, MH ⁺ 343. Example 20 N-(((4-methylquinazolin-2-yl)amino)((pyridin-4- ylmethyl)amino)methylene)acetamide

By analogous method to that described for Example 19, using Example 16. Afforded N-(((4- methylquinazolin-2-yl)amino)((pyridin-4-ylmethyl)amino)methy lene)acetamide (28 mg, 49%). LCMS ^V : Rt 4.8 min, MH ⁺ 335.

Example 21 : N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)isobutyramide

Example 8 (50 mg, 0.17 mmol), triethylamine (68 μL, 0.48 mmol) and 4- (dimethylamino)pyridine (2 mg, 0.01 mmol) were dissolved in 1 ,4-dioxane (2 mL). Isobutyryl chloride (21 μL, 0.20 mmol) was added drop-wise and the mixture stirred for 2 h. The mixture was evaporated and the residue purified by preparative HPLC ^e to give N-(((3- (dimethylamino)propyl)amino)((4-methylquinazolin-2-yl)amino) methylene)isobutyramide (48 mg, 79%). LCMS ^V : Rt 0.5 min, MH ⁺ 357.

Example 22: N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)cyclopentanecarboxamide

By analogous method to that described for Example 21 , with cyclopentanecarbonyl chloride (25 μL) as the acid chloride. Afforded N-(((3-(dimethylamino)propyl)amino)((4- methylquinazolin-2-yl)amino)methylene)cyclopentanecarboxamid e (48 mg, 74%). LCMS ^X : Rt 5.4 min, MH ⁺ 383. Example 23: N-(((4-methylquinazolin-2-yl)amino)((2-(piperidin-1 - yl)ethyl)amino)methylene)acetamide

Example 5 (50 mg, 0.16 mmol), triethylamine (65 μL, 0.48 mmol) and 4- (dimethylamino)pyridine (2 mg, 0.01 mmol) were dissolved in 1 ,4-dioxane (2 mL). Acetyl chloride (14 μL, 0.21 mmol) was added drop-wise and the mixture stirred for 2 h. The mixture was evaporated and the residue purified by preparative HPLC ^e to give N-(((4- methylquinazolin-2-yl)amino)((2-(piperidin-1 -yl)ethyl)amino)methylene)acetamide (34 mg, 60%). LCMS ^V : Rt 0.4 min, MH ⁺ 355.

Example 24: N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)-3-methylbutanamide

By analogous method to that described for Example 21 , with isovaleryl chloride (25 μL) as the acid chloride. Afforded N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)-3-methylbutanamide (35 mg, 55%). LCMS ^V : Rt 5.5 min, MH ⁺ 371.

Example 25 1 -(4-Methylquinazolin-2-yl)-3-(2-morpholinoethyl)guanidine

Intermediate 1 (125 mg, 0.679 mmol) and 2-(aminoethyl)morpholine (0.12 mL, 0.91 mmol) added to ethanol (4 mL) and heated for 10 min at 160 °C. The resulting mixture was evaporated. The residue was triturated with propan-2-ol and filtered. The collected solid was washed with propan-2-ol and lyophilized to give 1-(4-methylquinazolin-2-yl)-3-(2- morpholinoethyl)guanidine (116 mg, 54%). LCMS ^X : Rt 3.3 min, MH ⁺ 315. Example N-(((4-methylquinazolin-2-yl)amino)((2- morpholinoethyl)amino)methylene)acetamide

By analogous method to that described for Example 21 , with Example 25 (50 mg) and acetyl chloride (14 μL). Afforded N-(((4-methylquinazolin-2-yl)amino)((2- morpholinoethyl)amino)methylene)acetamide (34 mg, 56%). LCMS ^V : Rt 0.3 min, MH ⁺ 357.

Example 27: N-(((4-methylquinazolin-2-yl)amino)((3- morpholinopropyl)amino)methylene)acetamide

By analogous method to that described for Example 21 , with Example 15 (50 mg, 0.15 mmol) and acetyl chloride (14 μL). Afforded N-(((4-methylquinazolin-2-yl)amino)((3- morpholinopropyl)amino)methylene)acetamide (43 mg, 77%). LCMS ^X : Rt 3.9 min, MH ⁺ 371.

Example 28 N-(((4-methylquinazolin-2-yl)amino)(piperidin-4- ylamino)methylene)acetamide

Intermediate 1 (150 mg, 0.81 mmol) and 4-amino-1-boc-piperidine (195 mg, 0.97 mmol) were added to ethanol (4 mL) and heated for 15 min at 160 °C. The resulting mixture was evaporated. The residue was purified by chromatography (alumina) eluting with 2 M methanolic ammonia in DCM (0-6%) to give tert-butyl 4-(3-(4-methylquinazolin-2- yl)guanidino)piperidine-1-carboxylate (190 mg). The latter was dissolved in DCM (4 mL), 2- (dimethylamino)pyridine (2 mg, 0.01 mmol) and triethylamine (0.17 mL, 1.22 mmol) were added, then acetyl chloride (42 μL, 0.63 mmol) was added drop-wise. The mixture was stirred for 2 h then evaporated. The residue was purified by chromatography (silica gel) eluted with 2 M methanolic ammonia in DCM (0-10%) to give tert-butyl-4-(2-acetyl-3-(4- methylquinazolin-2-yl)guanidino)piperidine-1 -carboxylate (132 mg). The latter was dissolved in DCM (1.5 mL) and trifluoroacetic acid (0.5 mL) was added. The mixture was stirred for 2 h then evaporated. The residue was purified by chromatography (alumina) eluting with 2 M methanolic ammonia in DCM (0-5%) to give N-(((4-methylquinazolin-2- yl)amino)(piperidin-4-ylamino)methylene)acetamide (69 mg, 26% over 3 steps). LCMS ^W : Rt 4.9 min, MH ⁺ 327.

Example 29: 3,3,3-Trifluoro-/V-(((1-methylpiperidin-4-yl)amino)((4-methy lquinazolin-2- yl)amino)methylene)propanamide

By analogous method to that described for Example 21 , with Example 3 (50 mg, 0.17 mmol) and 3,3,3-trifluoropropionyl chloride (19 μL). Afforded 3,3,3-trifluoro-N-(((1 -methylpiperidin- 4-yl)amino)((4-methylquinazolin-2-yl)amino)methylene)propana mide (8 mg, 11%). LCMS ^W : Rt 5.5min, MH ⁺ 409.

Example 30: 2,2-Dichloro-/V-(((1 -methylpiperidin-4-yl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

By analogous method to that described for Example 21 , with Example 3 (50 mg, 0.17 mmol) and 3,3,3-dichloroacetyl chloride (17 μL). Afforded 2,2-dichloro-/V-(((1-methylpiperidin-4- yl)amino)((4-methylquinazolin-2-yl)amino)methylene)acetamide (14 mg, 20%). LCMS ^W : Rt 5.6 min, MH ⁺ 409. Example 31 : 2,2-Difluoro-N-(((1-methylpiperidin-4-yl)amino)((4-methylqui nazolin-2- yl)amino)methylene)acetamide

ABy analogous method to that described for Example 21 , with Example 3 (50 mg, 0.17 mmol) and difluoroacetic anhydride (20 μL). Afforded 2,2-difluoro-/V-(((1-methylpiperidin-4- yl)amino)((4-methylquinazolin-2-yl)amino)methylene)acetamide (22 mg, 34%). LCMS ^W : Rt 5.2 min, MH ⁺ 377.

Example 32: N-(((3-(Dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)-3,3,3-trifluoropropanamide

By analogous method to that described for Example 21 , with Example 8 (75 mg, 0.26 mmol), triethylamine (73 μL) and 3,3,3-trifluoropropionyl chloride (28 μL). Afforded N-(((3- (dimethylamino)propyl)amino)((4-methylquinazolin-2-yl)amino) methylene)-3,3,3- trifluoropropanamide (16 mg, 16%). LCMS ^W : Rt 5.1 min, MH ⁺ 397.

Example 33: 2,2-Dichloro-/V-(((3-(dimethylamino)propyl)amino)((4-methylq uinazolin- 2-yl)amino)methylene)acetamide

By analogous method to that described for Example 32, with di chloroacetyl chloride (25 μL).

Afforded 2,2-dichloro-/V-(((3-(dimethylamino)propyl)amino)((4-methylq uinazolin-2- yl)amino)methylene)acetamide (23 mg, 22%). LCMS ^W : Rt 5.3min, MH ⁺ 397. Example 34: 2,2,2-Trifluoro-/V-(((1-methylpiperidin-4-yl)amino)((4-methy lquinazolin-2- yl)amino)methylene)acetamide

By analogous method to that described for Example 21 , with Example 3 (50 mg, 0.17 mmol) and trifluoroacetic anhydride (22 μL). Afforded 2,2,2-trifluoro-N-(((1-methylpiperidin-4- yl)amino)((4-methylquinazolin-2-yl)amino)methylene)acetamide (21 mg, 31 %). LCMS ^W : 5.6 min, MH ⁺ 395.

Example 35: N-(((3-(Dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)-2,2,2-trifluoroacetamide

By analogous method to that described for Example 32, with trifluoroacetic anhydride (35 μL). Afforded N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)-2,2,2-trifluoroacetamide (8 mg, 8%). LCMS ^W : Rt 5.7 min, MH ⁺ 383.

Example 36: 1 -(2-(Dimethylamino)ethyl)-3-(quinazolin-2-yl)guanidine

Intermediate 2 (165 mg, 0.97 mmol) and /V,/V-dimethylethylenediamine (0.16 mL, 1 .46 mmol) were added to ethanol (4 mL) and heated for 10 min at 160 °C. The resulting mixture was evaporated. The residue was purified by flash column chromatography (alumina) eluting with 2 M methanolic ammonia in DCM (0-6%) to give 1-(2-(dimethylamino)ethyl)-3-(quinazolin- 2-yl)guanidine (105 mg, 42%). LCMS ^W : Rt 4.8 min, MH+ 259. Example 37: N-(((2-(dimethylamino)ethyl)amino)(quinazolin-2- ylamino)methylene)acetamide

By analogous method to that described for Example 21 , with Example 36 (50 mg, 0.19 mmol) and acetyl chloride (14 μL). Afforded N-(((2-(dimethylamino)ethyl)amino)(quinazolin-2- ylamino)methylene)acetamide (20 mg, 35%). LCMS ^b : Rt 4.7 min, MH ⁺ 301.

Example 38 1 -(4-Methylquinazolin-2-yl)-3-((tetrahydrofuran-2-yl)methyl)g uanidine

Intermediate 1 (80 mg, 0.43 mmol) in ethanol (2 mL) had tetrahydrofuran-2-ylmethanamine (48 mg, 0.48 mmol added and heated for 30 min at 160 °C in a microwave. The mixture was concentrated and the residue was purified by Prep-HPLC ^U to afford 1-(4- methylquinazolin-2-yl)-3-((tetrahydrofuran-2-yl)methyl)guani dine (28 mg, 22%). ¹ H NMR (500 MHz, DMSO-d ⁶ ) δ 8.25-8.15 (m, 1 H), 7.95-7.80 (m, 1 H), 7.75-7.65 (m, 1 H), 7.50-7.40 (m, 1 H), 4.30-4.15 (m, 1 H), 4.15-4.00 (m, 1 H), 3.90-3.85 (m, 1 H), 3.75-3.55 (m, 1 H), 3.50- 3.40 (m, 1 H), 2.90 (s, 3H), 2.30-1.90 (m, 3H), 1.90-1.60 (m, 1 H). LCMS ^W : Rt 4.3 min, MH ⁺ 286.

Example 39 1-Benzyl-3-(4-methylquinazolin-2-yl)guanidine

Intermediate 1 (200 mg, 1.09 mmol) in ethanol (3 mL) had benzylamine (465 mg, 4.34 mmol) at room temperature. The resulting mixture was stirred for overnight at 80 °C. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep- HPLC ^j to afford 1-benzyl-3-(4-methylquinazolin-2-yl)guanidine (100 mg, 23%) as a white solid TFA salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 11.12 (s, 1 H), 9.81 (s, 1 H), 8.90 - 8.70 (m, 2H), 8.27 (d, J = 8.3 Hz, 1 H), 8.04 - 7.85 (m, 2H), 7.71 - 7.57 (m, 1 H), 7.44 - 7.18 (m, 5H), 4.71 (d, = 6.1 Hz, 2H), 2.93 (s, 3H). LCMS ^k : Rt 1.11 min, MH ⁺ 292.

Example 40 1 -(Cyclohexylmethyl)-3-(4-methylquinazolin-2-yl)guanidine

By analogous method to that described for Example 38, with cyclohexylmethylamine. Afforded 1-(cyclohexylmethyl)-3-(4-methylquinazolin-2-yl)guanidine (8 mg, 6%). LCMS ^y : Rt 6.1 min, MH ⁺ 298.

Example 41 1 -(4-Methylquinazolin-2-yl)-3-(tetrahydro-2H-pyran-4-yl)guani dine

To a suspension of Intermediate 1 (80 mg, 0.43 mmol) in ethanol (2 mL) was added tetrahydropyran-4-amine hydrochloride (66 mg, 0.48 mmol) and DIPEA (67 mg, 0.52 mmol) and the reaction heated at 160 °C under microwave irradiation for 30 minutes. The mixture was concentrated and the residue was purified by Prep-HPLC ^U to afford 1-(4- methylquinazolin-2-yl)-3-(tetrahydro-2H-pyran-4-yl)guanidine (14 mg, 11%). ¹ H NMR (500 MHz, DMSO-d ⁶ ) δ 8.05-7.95 (m, 1 H), 7.75-7.65 (m, 1 H), 7.60-7.50 (m, 1 H), 7.35-7.25 (m, 1 H), 4.10-3.75 (m, 3H), 3.50-3.35 (m, 2H), 2.75 (s, 3H), 2.00-1.80 (m, 2H), 1.50-1.30 (m, 2H). LCMS ^W : Rt 4.1 min, MH ⁺ 286.

Example 42 1 -(4-Methylquinazolin-2-yl)-3-(2-(morpholinosulfonyl)ethyl)gu anidine

By analogous method to that described for Example 38, with 2- morpholinosulfonylethanamine hydrochloride. Afforded 1-(4-methylquinazolin-2-yl)-3-(2- (morpholinosulfonyl)ethyl)guanidine (7 mg, 4%). ¹ H NMR (500 MHz, DMSO-d ⁶ ) δ 8.15-8.05 (m, 1 H), 7.80-7.70 (m, 1 H), 7.65-7.55 (m, 1 H), 7.35-7.25 (m, 1 H), 4.15-4.00 (m, 6H), 3.50- 3.35 (m, 2H), 3.30-3.10 (m, 4H), 2.75 (s, 3H). LCMS ^W : Rt 4.1 min, MH ⁺ 379. Example 43 1 -(2-( 1 H-Pyrazol-1 -yl)ethyl)-3-(4-methylquinazolin-2-yl)guanidine

By analogous method to that described for Example 38, with 2-pyrazol-1-ylethanamine. Afforded 1-(2-(1 H-pyrazol-1-yl)ethyl)-3-(4-methylquinazolin-2-yl)guanidine (25 mg, 19%). ¹ H NMR (500 MHz, DMSO-d ⁶ ) δ 8.05-7.95 (m, 1 H), 7.75 (s, 1 H), 7.75-7.65 (m, 1 H), 7.65-7.55 (m, 1 H), 7.50 (s, 1 H), 7.35-7.25 (m, 1 H), 6.25 (s, 1 H), 4.45-4.25 (m, 2H), 3.80-3.60 (m, 2H), 2.75 (s, 3H). LCMS ^W : Rt 4.1 min, MH ⁺ 296.

Example 44 1 -(3-(Dimethylamino)propyl)-1 -methyl-3-(4-methylquinazolin-2- yl)guanidine

A mixture of Intermediate 1 (100 mg, 0.54 mmol) and 3-(dimethylamino) propyl](methyl)amine (95 mg, 0.82 mmol) in DMSO (1.0 ml ) was stirred for 5 min at room temperature. The reaction mixture was irradiated with microwave radiation for 20 min at 160 °C. The crude product was purified by Prep-HPLC ^q to afford 1-(3-(dimethylamino)propyl)-1- methyl-3-(4-methylquinazolin-2-yl)guanidine (42 mg, 26%) as a light yellow solid. ¹ H NMR (400 MHz, Chloroform-d) δ 8.99 (br, 2H), 7.89 (d, J = 8.2 Hz, 1 H), 7.64 - 7.63 (m, 2H), 7.27 (s, 1 H), 3.49 - 3.40 (m, 2H), 3.20 (s, 3H), 2.84 (s, 3H), 2.32 - 2.30 (m, 2H), 2.24 (s, 6H), 1.77 - 1 .75 (m, 2H). LCMS ^r : Rt 2.68 min, MH ⁺ 301.

Example 45 1 -(2,6-Dichlorobenzyl)-3-(4-methylquinazolin-2-yl)guanidine

Intermediate 1 (500 mg, 2.71 mmol) in ethanol (5 mL) had 1-(2,6- dichlorophenyl)methanamine (717 mg, 4.07 mmol) added at room temperature. The resulting mixture was stirred for overnight at 80 °C. The resulting mixture was concentrated under reduced pressure. The crude product (200 mg) was purified by Prep-HPLC ^f to afford 1-(2,6-Dichlorobenzyl)-3-(4-methylquinazolin-2-yl)guanidine (101 mg, 9%) as a white solid formic acid salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 11.93 (s, 1 H), 8.67 (s, 1 H), 8.02 (d, J = 8.4 Hz, 1 H), 7.83 - 7.81 (m, 1 H), 7.66 (d, J = 8.5 Hz, 1 H), 7.56 - 7.47 (m, 1 H), 7.42 (d, J = 8.1 Hz, 2H), 7.34 - 7.28 (m, 1 H), 4.89 (s, 2H), 2.92 (s, 3H). LCMS ^g: Rt 1.11 min, MH ⁺ 360. Example 46 N-(((2,6-dichlorobenzyl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

Example 45 (120 mg, 0.33 mmol) in DMSO (4.0 mL) was treated with AC2O (31.5 μL, 0.33 mmol) for 4 h at room temperature. Evaporated to dryness and the residue was purified by Prep-HPLC ⁹ to afford N-(((2,6-dichlorobenzyl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide (62 mg, 46%) as a white solid. ¹ H NMR (300 MHz, Chloroform-d, mix of rotamers) δ 14.53 (s, 0.5H), 13.04 (s, 0.5H), 10.85 (s, 0.5H), 8.75 (s, 0.5H), 8.05 - 7.96 (m, 1 H), 7.86 - 7.74 (m, 1 H), 7.67 (m, 1 H), 7.55 - 7.34 (m, 3H), 7.23 (m, 1 H), 5.19 - 5.07 (m, 2H), 2.91 (m, 3H), 2.28 (m, 3H). LCMS ^h : Rt 1.40 min, MH ⁺ 402.

Example 47 1 -(4-(Dimethylamino)butyl)-3-(4-methylquinazolin-2-yl)guanidi ne

Intermediate 1 (170 mg, 0.92 mmol) in DMSO (1.5 mL) was added (4- aminobutyl)dimethylamine (161 mg, 1.39 mmol) at room temperature. The reaction mixture was irradiated with microwave radiation for 20 min at 160 °C. The crude product was purified by Prep-HPLC ^P to afford 1-(4-(dimethylamino)butyl)-3-(4-methylquinazolin-2-yl)guanid ine (160 mg, 38.1 %) as a yellow solid TFA salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 11 .67 (s, 1 H), 10.17 (s, 1 H), 7.99 - 7.97 (m, 1 H), 7.82 - 7.80 (m, 1 H), 7.73 - 7.72 (m, 1 H), 7.52 - 7.50 (m, 1 H), 3.49 - 3.46 (m, 2H), 3.16 - 3.15 (m, 2H), 2.88 - 2.66 (m, 9H), 2.02 - 1.79 (m, 4H). LCMS ^q : Rt 0.64 min, MH ⁺ 301.

Example 48 1 -(7-Methoxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine

Step 1: 1 -(2-((2,4-dimethoxybenzyl)amino)-4-methoxyphenyl)ethan-1 -one

To a stirred solution of 1-(2-fluoro-4-methoxyphenyl) ethanone (30.0 g, 178 mmol) and CS2CO3 (116 g, 356 mmol) in DMSO (200 mL) was added (2,4-dimethoxyphenyl) methanamine (50 mL, 267 mmol) dropwise at room temperature. The resulting mixture was stirred for 3 h at 120 °C. The resulting mixture was diluted with H2O (500 mL). The resulting mixture was extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (3 x 100 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 :1) to afford 1-(2-((2,4-dimethoxybenzyl)amino)-4- methoxyphenyl)ethan-1-one (40.0 g, 79%) as a yellow solid. LCMS: MH ⁺ 284.

Step 2: 1-(2-amino-4-methoxyphenyl) ethan-1-one hydrochloride

To a stirred solution of 1-(2-((2,4-dimethoxybenzyl) amino)-4-methoxyphenyl)ethan-1-one (20.0 g, 74.3 mmol) in DCM (200 mL) were added TFA (40 mL) dropwise at room temperature. The resulting mixture was stirred for overnight at room temperature. The reaction was quenched with sat. NaHCO3 (aq.) at 0 °C. The aqueous layer was extracted with DCM (3 x 150 mL). The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 :1) to afford 1-(2-amino-4-methoxyphenyl) ethanone as a yellow solid. The product was further treated with 4 N HCI in 1 ,4-dioxane (25 mL) and stirred for 30 min at 0 °C. The precipitated solids were collected by filtration and washed with MTBE (3 x 10 mL). The filter cake was dried under vacuum to give 1-(2-amino-4-methoxyphenyl) ethan-1-one hydrochloride (4.60 g, 31%). LCMS: MH ⁺ 166.

Step 3: N-(7-methoxy-4-methylquinazolin-2-yl)cyanamide 1-(2-amino-4-methoxyphenyl)ethan-1-one hydrochloride (4.60 g, 22.9 mmol) was dissolved in ethyl alcohol (46 mL). To the above solution was added sodium dicyanoazanide (6.20 g, 69.6 mmol) in portions at room temperature. The resulting mixture was stirred for overnight at 80 °C. The precipitated solids were collected by filtration and washed with water and MTBE (3 x 100 mL). This resulted in N-(7-methoxy-4-methylquinazolin-2-yl) cyanamide (1.8 g, 34%) as a red solid. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 12.9 (br, 1 H), 8.11 - 8.09 (m, 1 H), 7.08 - 7.01 (m, 1 H), 6.92 - 7.89 (m, 1 H), 3.90 (s, 3H), 2.75 (s, 3H). LCMS: MH ⁺ 215.

Step 4: 1-(7-Methoxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine

To a stirred mixture of N-(7-methoxy-4-methylquinazolin-2-yl)cyanamide (200 mg, 0.934 mmol) in dimethylformamide (2.0 mL) was added 1-methylpiperidin-4-amine (160 mg, 1.40 mmol) at room temperature. The resulting mixture was stirred for overnight at 120 °C under nitrogen atmosphere. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC ^m to afford 1-(7-methoxy-4-methylquinazolin-2-yl)- 3-(1-methylpiperidin-4-yl)guanidine (35 mg, 10%) as a red solid formic acid salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 8.33 (s, 2H), 8.10 (d, J = 9.1 Hz, 1 H), 7.30 -7.07 (m, 2H), 3.94 (s, 3H), 3.72 (s, 1 H), 2.80 (s, 3H), 2.72 (d, J = 10.9 Hz, 2H), 2.22 (s, 3H), 2.13 - 2.11 (m, 2H), 1.92 - 1.89 (m, 2H), 1.66 - 1 .63 (m, 2H). LCMS ⁿ : Rt 1 .42 min, MH ⁺ 329.

Example 49 1 -(8-Methoxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine

Step 1: 1 -(3-methoxy-2-nitrophenyl) ethanone m-Methoxyacetophenone (50.0 g, 0.33 mol) had HNO3 (250 mL) added at 0 °C. The resulting mixture was stirred for 3 days at room temperature. The reaction was quenched by the addition of water (1.00 L) at 0 °C. The precipitated solid was collected by filtration, the filter cake was washed with water (100 mL) and ethanol (50 mL). The solid was further dried under vacuum to give 1-(3-methoxy-2-nitrophenyl) ethanone (30 g) as a yellow solid which was used directly in the next step. LCMS: MH ⁺ 196.

Step 2: 1-(2-amino-3-methoxyphenyl)ethanone

To a stirred solution of 1-(3-methoxy-2-nitrophenyl)ethanone (27.0 g, 138 mmol) and Zn (27.1 g, 415 mmol) in EtOH (270 mL) was added AcOH (55.5 mL) dropwise at room temperature. The resulting mixture was stirred for 2 h at 80 °C. The precipitated solids were collected by filtration and washed with MeOH (50 mL). The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (1 :3) to afford 1-(2-amino-3-methoxyphenyl)ethanone (23 g) as a yellow solid which was used directly in the next step. LCMS: MH ⁺ 166.

Step 3: N-(2-acetyl-6-methoxyphenyl)-2, 2, 2-trichloroacetamide

To a stirred solution of 1-(2-amino-3-methoxyphenyl)ethanone (22.0 g, 139 mmol) in DCM (34.0 mL) was added trichloroacetyl chloride (20.0 mL, 161 mmol) dropwise at 0 °C. The resulting mixture was stirred for 2 h at room temperature. The reaction was quenched with aq. HCI (1 N) at 0 °C. The mixture was neutralized to pH 7 with saturated NaHCO3. The aqueous layer was extracted with DCM (3 x 200 mL). The resulting mixture was concentrated under reduced pressure. This resulted in N-(2-acetyl-6-methoxyphenyl)-2,2,2- trichloroacetamide (35 g) as a yellow solid which was used directly in the next step. LCMS: MH ⁺ 310.

Step 4: 8-methoxy-4-methyl-1H-quinazolin-2-one

A stirred solution of N-(2-acetyl-6-methoxyphenyl)-2, 2, 2-trichloroacetamide (34.5 g, 111 mmol) in MeOH (324 mL) had ammonium carbamate (43.4 g, 555 mmol) added at room temperature. The resulting mixture was stirred for overnight at 40 °C. The resulting mixture was filtered; the filter cake was washed with MeOH (2 x 50 mL). The filtrate was concentrated under reduced pressure. The residue was purified by trituration with tert-butyl methyl ether (50 mL). This resulted in 8-methoxy-4-methyl-1 H-quinazolin-2-one (16 g) as a reddish brown solid which was used directly in the next step. LCMS: MH ⁺ 191.

Step 5: 2-chloro-7-methoxy-4-methylquinazoline

A stirred solution of 8-methoxy-4-methyl-1 H-quinazolin-2-one (10.0 g, 52.6 mmol) in phosphorus oxychloride (30 mL) at 0 °C was stirred for 3 h at 60 °C. The resulting mixture was concentrated under reduced pressure. The residue was dissolved in EtOAc (200 mL) and water (200 mL). The resulting mixture was extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (3 x 100 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 :1) to afford 2- chloro-7-methoxy-4-methylquinazoline (4 g, 36%) as a yellow solid. LCMS: MH ⁺ 209.

Step 6: N-(8-methoxy-4-methylquinazolin-2-yl) cyanamide

To a stirred solution of 2-chloro-8-methoxy-4-methylquinazoline (2.80 g, 13.4 mmol, 1.00 eq.) and cyanamide, monosodium salt (1.29 g, 20.1 mmol, 1.50 equiv) in NMP (15 mL) at room temperature. The resulting mixture was stirred for overnight at 60 °C. The residue was purified by reversed-phase flash chromatography (C18, MeCN in Water (0.1 % FA), 10 % to 100 % gradient) This resulted in N-(8-methoxy-4-methylquinazolin-2-yl) cyanamide (1.10 g, 38%) as a grey solid. ¹ H NMR (400 MHz, DMSO-d ⁶ ) 7.48 - 7.42 (m, 1 H), 7.12 - 6.95 (m, 2H), 3.87 (s, 3H), 2.58 (s, 3H). LCMS: MH ⁺ 215.

Step 7: 1-(8-Methoxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine

To a stirred mixture of N-(8-methoxy-4-methylquinazolin-2-yl) cyanamide (200 mg, 0.93 mmol) in dimethylformamide (2.0 mL) was added 1-methylpiperidin-4-amine (128 mg, 1.12 mmol) at room temperature. The resulting mixture was stirred for overnight at 120 °C. The resulting mixture was concentrated under reduced pressure. The crude product was purified by Prep-HPLC to afford 1-(8-methoxy-4-methylquinazolin-2-yl)-3-(1-methylpiperidin-4 - yl)guanidine (14 mg, 3%) as a green solid TFA salt. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 11.48 (br, 1 H), 8.31 (s, 2H), 8.08 (d, J= 8.8 Hz, 1 H), 7.18 - 7.14 (m, 2H), 3.94 (s, 3H), 3.85 (s, 1 H), 2.79 (s, 3H), 2.72 - 2.69 (m, 2H), 2.21 (s, 3H), 2.19 - 2.13 (m, 2H), 1.91 - 1.89 (m, 2H), 1.64 - 1 .60 (m, 2H). LCMS ^m : Rt 6.62 min, MH ⁺ 329.

Example 50 1 -(6-Methoxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine Step 1: 1 -(2-amino-5-methoxyphenyl)ethan-1 -one hydrochloride

To a stirred solution of 2-bromo-4-methoxyaniline (20.0 g, 99.9 mmol) and (1- butoxyethenyl)tributylstannane (116 g, 297 mmol) in toluene (200 mL) was added Pd(PPh ₃ ) ₂ CI ₂ (6.95 g, 9.90 mmol) in portions under nitrogen atmosphere. The resulting mixture was stirred for overnight at 100 °C under nitrogen atmosphere. The reaction was quenched with sat. HCI (aq.) and sat. KF (aq.) at 0 °C. The precipitated solids were collected by filtration and washed with EtOAc (3 x 10 mL). The resulting mixture was extracted with EtOAc (3 x 10 mL). The combined organic layers were washed with brine (3 x 10 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (1 :1) to afford 1-(2-amino-5-methoxyphenyl) ethanone as a brown solid. The product was treated with 4 N HCI in 1 ,4-dioxane (25 mL) and stirred for 30 mins at room temperature. The precipitated solids were collected by filtration and washed with MTBE (3 x 10 mL). This result in 1-(2-amino-5-methoxyphenyl)ethan-1-one hydrochloride (5.0 g, 25%). LCMS: MH ⁺ 166.

Step 2: N-(6-methoxy-4-methylquinazolin-2-yl)cyanamide

To a stirred solution of 1-(2-amino-5-methoxyphenyl)ethanone hydrochloride (5.00 g, 24.8 mmol) in ethyl alcohol (50.0 mL) was added sodium dicyanoazanide (4.42 g, 49.6 mmol) in portions. The resulting mixture was stirred for 4 h at 80 °C. The precipitated solid was collected by filtration, the filter cake was washed with EtOH, water and Et20. The solid was further dried under vacuum to give N-(6-methoxy-4-methylquinazolin-2-yl)cyanamide (3.00 g, 56%) as a white solid. ¹ H NMR (300 MHz, DMSO-d ⁶ ) δ 12.78 (br, 1 H), 5 7.50 (dd, J = 9.1 , 2.6 Hz, 1 H), 7.45 - 7.36 (m, 2H), 3.88 (s, 3H), 2.80 (s, 3H). LCMS: MH ⁺ 215.

Step 3: 1-(6-Methoxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine

To a stirred solution of N-(6-methoxy-4-methylquinazolin-2-yl)cyanamide (200 mg, 0.934 mmol) in DMF (2.0 mL) was added 1-methylpiperidin-4-amine (160 mg, 1.40 mmol) at room temperature. The resulting mixture was stirred for overnight at 120 °C. The reaction was monitored by LCMS. The crude product was purified by Prep-HPLC ⁿ to afford 1-(6-methoxy- 4-methylquinazolin-2-yl)-3-(1-methylpiperidin-4-yl)guanidine (156 mg, 39.7 %) as a brown solid formic acid salt. ¹ H NMR (400 MHz, DMSO-d ⁶ , mix of rotamers) δ 11.10 (br, 0.8H), 9.58 (br, 0.2H), 8.66 (s, 2H), 7.67 (d, J = 9.2 Hz, 1 H), 7.48 - 7.46 (m, 1 H), 7.17 (d, J = 2.7 Hz, 1 H), 4.17 (br, 1 H), 3.94 (s, 3H), 3.30 (br, 2H), 2.87 (s, 4H), 2.64 - 2.62 (m, 4H), 2.41 - 2.30 (m, 2H), 2.07 - 2.00 (m, 2H). LCMS ⁰ : Rt 1.38 min, MH ⁺ 329. Example 51 1 -(5-Methoxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine

Step 1: 1 -(2-amino-6-methoxyphenyl)ethan-1 -one hydrochloride

To a stirred solution of 2-bromo-3-methoxyaniline (20.0 g, 99.1 mmol) and (1- butoxyethenyl)tributylstannane (57.8 g, 149 mmol) in toluene (200 mL) was added Pd(PPhs)4 (11.4 g, 9.91 mmol) in portions under nitrogen atmosphere. The resulting mixture was stirred for overnight at 100 °C under nitrogen atmosphere. The reaction was quenched with sat. HCI (aq.) and sat. KF (aq.) at 0 °C. The precipitated solids were collected by filtration and washed with EtOAc (3 x 10 mL). The resulting mixture was extracted with EtOAc (3 x 10 mL). The combined organic layers were washed with NaCI (aq.) (3 x 10 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 :1) to afford a brown solid which was further treated with 4 N HCI in 1 ,4-dioxane (25 mL), The precipitated solids were collected by filtration and washed with MTBE (3 x 10 mL). The filter cake was dried under vacuum to give 1-(2-amino-6-methoxyphenyl)ethanone hydrochloride (2.46 g, 15%). LCMS: MH ⁺ 166.

Step 2: N-(5-methoxy-4-methylquinazolin-2-yl)cyanamide

To a stirred solution of 1-(2-amino-6-methoxyphenyl)ethanone hydrochloride (2.46 g, 12.2 mmol) in ethyl alcohol (24.0 mL) was added sodium dicyanoazanide (1.63 g, 18.3 mmol). The resulting mixture was stirred for 4 h at 80 °C. The precipitated solid was collected by filtration, the filter cake was washed with EtOH, water and Et20. The solid was further dried under vacuum to give N-(5-methoxy-4-methylquinazolin-2-yl)cyanamide (1.20 g, 46%) as a yellow solid. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 12.89 (s, 1 H), δ 7.74 (m, 1 H), 7.27 - 6.75 (m, 2H), 3.96 (s, 3H), 2.84 (s, 3H). LCMS: MH ⁺ 215.

Step 3: 1-(5-Methoxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine

To a stirred solution of N-(5-methoxy-4-methylquinazolin-2-yl)cyanamide (200 mg, 0.93 mmol) in DMSO (4.00 mL) was added 1-methylpiperidin-4-amine (117 mg, 1.03 mmol). The reaction mixture was irradiated with microwave radiation for 20 min at 160 °C. The crude product was purified by Prep-HPLC ⁰ to afford 1-(5-Methoxy-4-methylquinazolin-2-yl)-3-(1- methylpiperidin-4-yl)guanidine (120 mg, 28%) as a brown formic acid solid salt. ¹ H NMR (400 MHz, DMSO-d ⁶ , mix of roatmers) δ 10.55 (br, 0.5H), 8.32 (s, 2.5H), 7.82 (t, J = 8.2 Hz, 1 H), 7.36 (d, J = 8.3 Hz, 1 H), 7.03 (d, J = 8.1 Hz, 1 H), 3.98 (s, 3H), 3.71 (s, 1 H), 2.93 (s, 3H), 2.79 (d, J = 10.7 Hz, 2H), 2.27 (s, 3H), 2.16 - 2.00 (m, 2H), 1.95 - 1.93 (m, 2H), 1.68 - 1 .66 (m, 2H). LCMS ^P : Rt 3.51 min, MH ⁺ 329.

Example 52 1 -((1 -Methyl-1 H-imidazol-4-yl)methyl)-3-(4-methylquinazolin-2- yl)guanidine

By analogous method to Example 39. Afforded 1-((1-methyl-1 H-imidazol-4-yl)methyl)-3-(4- methylquinazolin-2-yl)guanidine (39 mg, 10%) as a red liquid formic acid salt. ¹ H NMR (400 MHz, Chloroform-d) δ 11.85 (br, 1 H), 5 10.61 (br, 1 H), 9.34 (br, 1 H), 8.72 (s, 1 H), 8.01 (d, J = 8.4 Hz, 1 H), 7.78 - 7.74 (m, 2H), 7.58 - 7.35 (m, 2H), 6.99 (s, 1 H), 4.42 (s, 2H), 3.70 (s, 3H), 2.92 (s, 3H). LCMS ^j : Rt 6.43 min, MH ⁺ 296.

Example 53 1 -(5-(Dimethylamino)pentyl)-3-(4-methylquinazolin-2-yl)guanid ine

Intermediate 1 (100 mg, 0.54 mmol) in DMSO (2 mL) was treated with (5- aminopentyl)dimethylamine (126 μL, 0.82 mmol) for 5 min at room temperature. The final reaction mixture was irradiated with microwave radiation for 20 min at 160 °C. The resulting mixture was purified by Prep-HPLC ^S to afford 1-(5-(dimethylamino)pentyl)-3-(4- methylquinazolin-2-yl)guanidine (25 mg, 10%) as a red oil TFA salt. ¹ H NMR (400 MHz, CDCh) δ 10.4 - 10.3 (m, 2H), 8.63 - 8.45 (m, 1 H), 7.98 - 7.96 (m, 1 H), 7.79 - 7.83 (m, 1 H), 7.56 - 7.49 (m, 1 H), 7.46 - 7.38 (m, 1 H), 3.48 - 3.26 (m, 2H), 3.21 - 3.08 (m, 2H), 2.88 - 2.81 (m, 9H), 1.89 - 1.77 (m, 4H), 1.58 - 1.56 (m, 2H). LCMS‘: Rt 3.22 min, MH ⁺ 315.

Example 54 N-(amino((4-methylquinazolin-2-yl) amino) methylene) acetamide

Intermediate 3 (150 mg, 0.75 mmol) in DSMO (1.5 mL) had acetic anhydride (92 mg, 0.89 mmol) added. The resulting mixture was stirred for overnight at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-HPLC ^b to afford N-(amino((4-methylquinazolin-2-yl) amino) methylene) acetamide (17.4 mg, 10 %) as a white solid. ¹ H NMR (400 MHz, DMSO-d ⁶ ) δ 12.81 - 9.02 (m, 3H), 8.22 - 8.07 (m, 1 H), 7.96 - 7.79 (m, 2H), 7.54 (apparent t, J = 7.5 Hz, 1H), 2.86 (s, 3H), 2.12 (s, 3H). LCMS ^b : Rt 0.87 min, MH ⁺ 244.

Examples 55-96

Each of Examples 55-96 can be synthesised by way of the methods described in General Schemes 1 to 4. Example 97 1 -(4-Methoxyquinazolin-2-yl)-3-(1 -methylpiperidin-4-yl)guanidine

Step 1: 2-aminobenzoic acid

To a stirred solution of 2-nitrobenzoic acid (3.00 g, 18.0 mmol) in EtOH (30 mL) was added Pd/C (3.0 g, 10% wt). The mixture was stirred for overnight under hydrogen atmosphere. The resulting mixture was filtered; the filter cake was washed with MeOH (3 x 30 mL). The filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1 :1) to afford 2-aminobenzoic acid (2.1 g, 85%) as a yellow oil. LCMS: MH ⁺ 138.

Step 2: N'-cyano-N-(1-methylpiperidin-4-yl)guanidine

To a stirred solution of sodium dicyanamide (10.0 g, 112 mmol) and 1-methylpiperidin-4- amine (12.1 g, 106 mmol) in n-BuOH (100 mL) was added cone. HCI (9.6 mL) dropwise at room temperature. The resulting mixture was stirred overnight at 110 °C. The resulting mixture was filtered; the filter cake was washed with water (3 x 10 mL). The filtrate was dried under vacuum. The resulting /V'-cyano-/V-(1-methylpiperidin-4-yl) guanidine (9.2 g, 45%) was used for the next step without further purification.

Step 3: 1-( 1-methylpiperidin-4-yl)-3-(4-oxo- 1, 4-dihydroquinazolin-2-yl)guanidine

To a stirred solution of A/'-cyano-N-(1-methylpiperidin-4-yl)guanidine (4.00 g, 22.1 mmol) and 2-aminobenzoic acid (1.01 g, 7.35 mmol) in H2SO4 (10% wt in H2O, 8.0 mL) at room temperature. The resulting mixture was stirred for 20 min at 100 °C. The reaction was quenched with sat. NaHCO3 (aq.) at room temperature. The resulting mixture was filtered. The filter cake was washed with EtOH and dried under vacuum to afford 1-(4- chloroquinazolin-2-yl)-3-(1-methylpiperidin-4-yl)guanidine (1.9 g, 85 %) as a yellow solid. LCMS: MH ⁺ 301

Step 4: 1-(4-chloroquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine

A solution of N-(1-methylpiperidin-4-yl)-A/'-(4-oxo-1 H-quinazolin-2-yl)guanidine (300 mg, 0.99 mmol) in POCI3 (3.0 mL, 32.2 mmol) was stirred for 1 h at room temperature. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with water (20 mL). The aqueous layer was extracted with EtOAc (2 x 20 mL). The organic phases were combined and dried under Na2SO4. filtered, and concentrated under reduced pressure to afford crude 1-(4-chloroquinazolin-2-yl)-3-(1-methylpiperidin-4- yl)guanidine (200 mg) as a yellow oil which was used for the next step without purification. Step 5: 1-(4-Methoxyquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine A solution of /V'-(4-chloroquinazolin-2-yl)-/V-(1-methylpiperidin-4-yl)gua nidine (100 mg, 0.31 mmol) and MeONa (20.3 mg, 0.37 mmol) in MeOH (12.1 mL) was stirred at room temperature for 2 h. The mixture was neutralized to pH 7 with aq. HCI (1 N). The resulting mixture was directly concentrated under vacuum. The crude product (100 mg) was purified by Prep-HPLC‘ to afford 1-(4-methoxyquinazolin-2-yl)-3-(1-methylpiperidin-4-yl)guani dine (22 mg, 18 %) as a colorless oil formate salt. ¹ H NMR (400 MHz, Chloroform-d) δ 11.4 - 9.10 (br, 2H), 8.67 (s, 1 H), 8.04 (d, J = 8.1 Hz, 1 H), 7.78-7.76 (m, 1 H), 7.68 - 7.62 (m, 1 H), 7.43 (t, J = 7.6 Hz, 1 H), 4.40 - 4.21 (m, 1 H), 4.19 (s, 3H), 3.68 (d, J = 10.8 Hz, 3H), 3.40 - 3.18 (m, 2H), 2.70 - 2.51 (m, 2H), 2.38 - 2.22 (m, 2H), 2.13 - 1.95 (m, 2H). LCMS ^U : Rt 0.883 min, MH ⁺ 315.

Example 98 1 -(8-hydroxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4-yl) guanidine

Step 1: N-(8-hydroxy-4-methylquinazolin-2-yl)cyanamide

To a solution of N-(8-methoxy-4-methylquinazolin-2-yl) cyanamide (300 mg, 1.40 mmol) in DCE (3.00 mL) at 0 °C was added a 1 M solution of BBr3 in DCM (2.94 mL, 2.94 mmol) dropwise. The mixture was stirred for 2 h at 80 °C. The mixture was allowed to cool down to room temperature. The reaction was quenched by the addition of water/ice (5.00 mL) at 0 °C. The resulting mixture was concentrated under vacuum. The residue was purified by Prep-HPLC ^W to afford the title compound (80.0 mg, 28.6%) as a brown solid. LCMS: MH ⁺ 201.

Step 2: 1-(8-hydroxy-4-methylquinazolin-2-yl)-3-(1-methylpiperidin-4 -yl) guanidine

A mixture of N-(8-hydroxy-4-methylquinazolin-2-yl) cyanamide (80.0 mg, 0.400 mmol) and 1-methylpiperidin-4-amine (300 uL, 2.40 mmol) in DMSO (1.00 mL) was stirred for 5 min at room temperature. The reaction mixture was irradiated with microwave radiation for 1 h at 160 °C. The mixture was allowed to cool down to room temperature. The solution was purified by Prep-HPLC ^X to afford the title compound (22.4 mg, 17.5%) as a brown oil. ¹ H- NMR (400 MHz, Methanol-d ₄ ) δ 7.74 - 7.63 (m, 1 H), 7.52 - 7.42 (m, 1 H), 7.39 - 7.30 (m, 1 H), 4.29 - 4.00 (m, 1 H), 3.72 - 3.64 (m, 1 H), 3.61 - 3.54 (m, 1 H), 3.30 - 3.20 (m, 2H), 3.00 - 2.87 (m, 6H), 2.48 - 2.30 (m, 2H), 2.29 - 1.99 (m, 2H). LCMS ^Z : Rt 0.452 min; MH ⁺ 315.

Example 99 1 -(7-hydroxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine

To a solution of Example 48 (100 mg, 0.304 mmol) in DCE (1.00 mL) was added a 1 M solution of BBrs in DCM (1.82 ml, 1.82 mmol) dropwise at 0 °C. The resulting mixture was stirred overnight at 80 °C. The mixture was allowed to cool down to room temperature and quenched by the addition of water/lce (10.0 mL) at 0 °C. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-HPLC ^Y to afford the title compound (18.6 mg, 19.4%) as a white solid. ¹ H NMR (400 MHz, Methanol-cL) δ 7.77 (d, J = 9.2 Hz, 1 H), 6.87 - 6.76 (m, 1 H), 6.54 (s, 1 H), 3.75 (s, 1 H), 2.82 (s, 2H), 2.68 (s, 3H), 2.33 (s, 5H), 2.11 - 2.01 (m, 2H), 1.68 (d, J = 11.6 Hz, 2H). LCMS ^AA : Rt 0.691 min; MH ⁺ 315.

Example 100 1-(6-hydroxy-4-methylquinazolin-2-yl)-3-(1-methylpiperidin-4 - yl)guanidine

By an analogous method to that described in Example 99 using Example 50 (60.0 mg, 0.183 mmol). Purification by Prep-HPLC ^R afforded title compound (12.3 mg, 16.7 %) as a reddish brown formate salt. ¹ H NMR (400 MHz, Methanol-d ₄ ) δ 8.48 (br, 2H), 7.76 (d, J = 9.0 Hz, 1 H), 7.56 - 7.49 (m, 1 H), 7.38 (d, J = 2.Q Hz, 1 H), 3.84 (s, 1 H), 3.29 - 3.18 (m, 2H), 2.85 (s, 3H), 2.80 - 2.70 (m, 2H), 2.64 (s, 3H), 2.29 - 2.14 (m, 2H), 1.98 - 1.84 (m, 2H). LCMS ^B : Rt 0.586 min, MH ⁺ 315.

Example 101 1 -(5-hydroxy-4-methylquinazolin-2-yl)-3-(1 -methylpiperidin-4- yl)guanidine

By an analogous method to that described in Example 99 using Example 51 (150 mg, 0.457 mmol). Purification by Prep-HPLC ^Z afforded title compound (17.6 mg, 9.0%) as a reddish brown trifluoroacetate salt. ¹ H NMR (400 MHz, Methanol-^) δ 7.69 (t, J = 8.1 Hz, 1 H), 7.27 (d, J = 8.2 Hz, 1 H), 6.92 (dd, J = 8.0, 1.0 Hz, 1 H), 3.95 (s, 1 H), 3.68 (d, J = 12.8 Hz, 2H), 3.15 (t, J = 12.9 Hz, 2H), 3.07 (s, 3H), 2.94 (s, 3H), 2.44 - 1.91 (m, 4H). LCMS ^B , Rt 0.547 min, MH ⁺ 315.

Example 102 1 -methyl-1 -(1 -methylpiperidin-4-yl)-3-(4-methylquinazolin-2-yl) guanidine

To a solution of N-(4-methylquinazolin-2-yl) cyanamide (200 mg, 1.09 mmol) in DMSO (4.00 mL) was added A/,1-dimethylpiperidin-4-amine (209 mg, 1.63 mmol). The mixture was irradiated with microwave radiation for 30 min at 160 °C. The mixture was allowed to cool down to room temperature. The mixture was purified by Prep-HPLC^ to afford the title compound (110 mg, 32.4%) as a red solid. ¹ H NMR (400 MHz, Chloroform-d) δ 8.12 - 7.76 (m, 3H), 7.73 - 7.46 (m, 2H), 7.30 - 7.29 (m, 1 H), 4.86 (s, 1 H), 3.10 - 2.91 (m, 5H), 2.86 (s, 3H), 2.33 (s, 3H), 2.30 - 2.20 (m, 2H), 1.94 - 1.82 (m, 2H), 1.80 - 1.71 (m, 2H). LCMS ^AC : Rt 1.213 min, MH ⁺ 313.

Example 103 N-((methyl(1-methylpiperidin-4-yl)amino)((4-methylquinazolin -2- yl)amino)methylene)acetamide

To a solution of Example 102 (100 mg, 0.320 mmol) in DMSO (3.0 mL) was added acetic anhydride (32.7 mg, 0.320 mmol). The mixture was stirred for 8 h at 60 °C. The mixture was allowed to cool down to room temperature. The mixture was purified by Prep-HPLC ^AB to afford the title compound (40 mg, 35.4%) as an orange solid. ¹ H NMR (400 MHz, DMSO-cfe)

6 12.51 (br, 1 H), 7.98 - 7.96 (d, J = 8.0 Hz, 1 H), 7.77 - 7.70 (m, 1 H), 7.70 - 7.68 (m, 1 H),

7.52 - 7.40 (m, 1 H), 4.65 (br, 1 H), 3.19 - 3.00 (m, 5H), 2.92 (s, 3H), 2.51 - 2.29 (m, 5H), 2.18 - 2.02 (m, 5H), 2.00 - 1.89 (m, 2H). LCMS ^AD : Rt 0.977 min, MH ⁺ 355.

Example 104 1-(6-(dimethylamino)hexyl)-3-(4-methylquinazolin-2-yl)guanid ine

To a mixture of N-(4-methylquinazolin-2-yl) cyanamide (200 mg, 1.18 mmol) in DMSO (2.00 mL) was added (6-aminohexyl)dimethylamine (188 mg, 1.30 mmol). The mixture was irradiated with microwave radiation for 30 min at 160 °C. The mixture was allowed to cool down to room temperature and purified by Prep-HPLC ^AC to afford the title compound (63.0 mg, 11.8%) as an orange trifluoroacetate salt. ¹ H NMR (400 MHz, Chloroform-d) δ 11.61 (s, 1 H), 10.05 (s, 1 H), 7.99 (d, J = 8.3 Hz, 1 H), 7.83 (t, J = 7.6 Hz, 1 H), 7.72 (d, J = 8.3 Hz, 1 H), 7.51 (t, J = 7.6 Hz, 1 H), 3.41 (d, J = 7.4 Hz, 2H), 3.07 (s, 2H), 2.92 - 2.79 (m, 9H), 2.01 - 1 .79 (m, 4H), 1.76 - 1.48 (m, 4H). LCMS ^AE : Rt 0.645 min, MH ⁺ 329.

Example 105 N-(((3-(dimethylamino)propyl)amino)((4-methylquinazolin-2- yl)amino)methylene)acetamide

To a solution of Example 8 (180 mg, 0.629 mmol) in DMSO (1.5 mL) was added AC2O (64.2 mg, 0.629 mmol) dropwise at 0 °C. The resulting mixture was stirred for 3 h at room temperature. The mixture was purified by Prep-HPLC ^AD directly to afford the title compound (55 mg, 26.7%) as a yellow solid. ¹ H NMR (400 MHz, Methanol-d ₄ ) δ 8.13 (dd, J = 8.5, 4.8 Hz, 1 H), 7.97 - 7.65 (m, 2H), 7.62 - 7.45 (m, 1 H), 3.82 - 3.60 (m, 2H), 3.25 - 3.02 (m, 2H), 3.00 - 2.60 (m, 9H), 2.33 (d, J = 2.7 Hz, 2H), 2.22 - 1.89 (m, 3H); LCMS°: Rt 0.807 min, MH ⁺ 329.1.

Example 106 1-(3-(dimethylamino)propyl)-3-(8-hydroxy-4-methylquinazolin- 2- yl)guanidine

Step 1: N-(8-methoxy-4-methylquinazolin-2-yl)cyanamide

To a solution of 1-(2-amino-3-methoxyphenyl)ethan-1-one hydrochloride (1.12 g, 5.55 mmol) in EtOH (12.0 mL) was added 1 M HCI (4.0 mL). The reaction mixture was stirred for 2 min and then followed by the addition of sodium dicyanamide (1.24 g, 13.9 mmol). The resulting mixture was stirred for additional 1 h at 80 °C. The mixture was allowed to cool down to room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-HPLC ^W to afford the title compound (490 mg, 40.8%) as a light-yellow solid. LCMS: MH ⁺ 215.

Step 2: N-(8-hydroxy-4-methylquinazolin-2-yl)cyanamide

To a solution of N-(8-methoxy-4-methylquinazolin-2-yl) cyanamide (300 mg, 1.40 mmol) in DCM (3.0 mL) was added 1 M BBrs in DCM (2.80 mL, 2.80 mmol). The resulting mixture was stirred for additional 2 h at room temperature. The reaction was quenched by the addition of water/lce (30 mL) and extracted with EtOAc (3 x 40 mL). The combined organic layers were washed with brine (3 x 5 mL), dried over anhydrous Na2SO ₄ , filtered, and concentrated under reduced pressure. The residue was purified by Prep-HPLC ^W to afford the title compound (90.0 mg, 32.1%) as a yellow solid. LCMS: MH ⁺ 201.

Step 3: 1-(3-(dimethylamino)propyl)-3-(8-hydroxy-4-methylquinazolin- 2-yl)guanidine To a solution of N-(8-hydroxy-4-methylquinazolin-2-yl) cyanamide (70.0 mg, 0.350 mmol) in DMSO (1.50 mL) was added dimethylaminopropylamine (66.0 μL, 0.525 mmol). The reaction mixture was irradiated with microwave radiation for 1 h at 160 °C. The mixture was allowed to cool down to room temperature. The mixture was purified by Prep-HPLC ^AE to afford the title compound (4.7 mg, 4.45%) as a yellow solid. ¹ HNMR (400 MHz, Chloroform - d) δ 7.37 (d, J = 7.8 Hz, 1 H), 7.21 - 7.10 (m, 2H), 3.41- 3.38 (m, 2H), 2.81 (s, 3H), 2.40 (t, J = 5.9 Hz, 2H), 2.25 (s, 6H), 1.80 -1 .70 (m, 2H). LCMS ^AF : Rt 0.479 min, 303 [M+H] ⁺

Example 107 1-(8-hydroxy-4-methylquinazolin-2-yl)-3-(3-morpholinopropyl) guanidine

To a solution of N-(8-hydroxy-4-methylquinazolin-2-yl) cyanamide (130 mg, 0.649 mmol) in DMSO (1.5 mL) was added 3-morpholinopropan-1-amine (112 mg, 0.779 mmol). The reaction mixture was irradiated with microwave radiation for 30 min at 160 °C. The mixture was allowed to cool down to room temperature. The mixture was purified by Prep- HPLC ^AI to afford the title compound (91 mg, 40.7%) as an off-white solid. ¹ H NMR (400 MHz, DMSO-d6) δ 7.71 (br, 1 H), 7.42 - 7.38 (m, 1 H), 7.13 - 7.00 (m, 2H), 3.63 - 3.54 (m, 4H), 3.30 - 3.28 (m, 2H), 2.68 (s, 3H), 2.39 - 2.19 (m, 6H), 1.78 - 1.58 (m, 2H). LCMS ^N : Rt 0.672 min, 345.

Example 108 1-(7-fluoro-8-hydroxy-4-methylquinazolin-2-yl)-3-(1-methylpi peridin-4- yl)guanidine

Step 1: 1-(4-fluoro-3-methoxy-2-nitrophenyl) ethenone

To a solution of 1-(4-fluoro-3-methoxyphenyl) ethanone (4.00 g, 23.8 mmol) in AC2O (45.0 mL) was added cupric nitrate (72.5 mg, 0.386 mmol). The reaction mixture was stirred for 30 min at 0 °C. The reaction mixture was quenched by the addition of water (10 mL) and extracted with EtOAc (10 mLx3). The combined organic phases were washed with brine, dried over Na2SC>4, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (6:1) to afford the title compound (2.70 g, 53.3%) as a light-yellow solid. LCMS: MH ⁺ 214.

Step 2 : 1-(2-amino-4-fluoro-3-methoxyphenyl) ethanone To a solution of 1-(4-fluoro-3-methoxy-2-nitrophenyl) ethanone (2.79 g, 13.1 mmol) in the mixture of EtOH (20.0 mL), THF (20.0 mL) and H ₂ O (10.0 mL) were added Fe (2.93 g, 52.4 mmol) and NH4CI (1.40 g, 26.2 mmol). The resulting mixture was stirred for 2 h at 60 °C. The mixture was allowed to cool down to room temperature. The mixture was filtered through a short pad of Celite. The pad was washed with EtOH (10 mL). The combined filtered was concentrated under reduce pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (6:1) to afford the title compound (890 mg, 37.1 %) as a light-yellow oil. LCMS: MH ⁺ 184.

Step 3 : [(7-fluoro-8-methoxy-4-methylquinazolin-2-yl) amino] formonitrile

To a solution of 1-(2-amino-4-fluoro-3-methoxyphenyl) ethanone (440 mg, 2.40 mmol) in 1 ,4-dioxane (4.0 mL) was added 4 M HCI in 1 ,4-dioxane (1 .0 mL). The resulting mixture was stirred for 10 min at room temperature and concentrated under reduce pressure. The residue was dissolved in EtOH (5.0 mL) and followed by the addition of sodium dicyanoazanide (428 mg, 4.80 mmol). The resulting mixture was stirred for additional 2 h at 80 °C and followed by the addition of 2 ^nd batch of 4 M HCI in 1 ,4-dioxane (1.0 mL). The reaction mixture was stirred for additional 30 min at 80 °C. The mixture was allowed to cool down to room temperature. The reaction mixture was neutralized to pH ~7 with aq. NaHCOs. The resulting mixture was extracted with EtOAc (30 mLx3). The combined organic phases were washed with brine, dried over Na ₂ SO4, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (7:1) to afford the title compound (253 mg, 45.36%) as a yellow solid. LCMS: MH ⁺ 233. Step 4 : [(7-fluoro-8-hydroxy-4-methylquinazolin-2-yl) amino] formonitrile

To a solution of [(7-fluoro-8-methoxy-4-methylquinazolin-2-yl) amino] formonitrile (190 mg, 0.818 mmol) in DCM (0.5 mL) was added 1 M BBr ₂ in DCM (2.0 mL). The resulting mixture was stirred for additional 1 h at 40 °C. The mixture was allowed to cool down to room temperature and concentrated under reduced pressure. The residue was purified by Prep- HPLC ^w to afford the title compound (28 mg, 15.7%) as a yellow solid. LCMS: MH ⁺ 219. Step 5 : 1-(7-fluoro-8-hydroxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine To a solution of [(7-fluoro-8-hydroxy-4-methylquinazolin-2-yl) amino] formonitrile (21.0 mg, 0.096 mmol, 1.00 eq.) in DMSO (0.5 mL) was added 1-methylpiperidin-4-amine (16.5 mg, 0.144 mmol, 1 .50 eq.). The reaction mixture was irradiated with microwave radiation for 30 min at 160 °C. The residue was purified Prep-HPLC ^AF to afford the title compound (8.8 mg, 27.51 %) as a yellow solid. ¹ H NMR (400 MHz, DMSO-d6) δ 7.65 (br, 2H), 7.48 - 7.38 (m, 1 H), 7.21 - 7.07 (m, 1 H), 3.75 (s, 1 H), 2.68 (s, 5H), 2.17 (s, 3H), 2.07 - 1.95 (m, 2H), 1.90 - 1 .80 (m, 2H), 1.54 - 1.37 (m, 2H). LCMS ^N : Rt 0.647 min MH ⁺ 333.

Example 109 1-(6-fluoro-8-hydroxy-4-methylquinazolin-2-yl)-3-(1-methylpi peridin-4- yl)guanidine

Step 1 : 2-bromo-4-fluoro-6-methoxyaniline

To a solution of 4-fluoro-2-methoxyaniline (30.0 g, 212 mmol) in DCM (500 mL) was added NBS (41.6 g, 234 mmol) in portions at -78 °C. The resulting mixture was stirred for 2 h at - 78 °C. The resulting mixture was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (9:1) to afford the title compound (15.0 g, 32.2%) as a red oil. LCMS: MH ⁺ 222.

Step 2 : 1-(2-amino-5-fluoro-3-methoxyphenyl) ethanone

To a solution of 2-bromo-4-fluoro-6-methoxyaniline (15.0 g, 68.2 mmo) and tributyl(1- ethoxyethenyl) stannane (49.2 g, 136 mmol) in toluene (225 mL) were added Pd(dppf)Cl2.CH2Cl2 (5.55 g, 6.82 mmol) in portions at room temperature. The resulting mixture was stirred for 1 h at 100 °C under nitrogen atmosphere. The reaction was allowed to cool down to room temperature and quenched by the addition of 1 M aq. HCI (225 mL). The resulting mixture was extracted with EtOAc (1.0 Lx3). The combined organic phases were washed with brine (600 mLx3), dried over Na2SO4, filtered, and concentrated under reduced pressure. The residue was purified by Prep-HPLC ^W to afford the title compound (4.5 g, 36.0%) as a yellow solid. LCMS: MH ⁺ 184.

Step 3 : [(6-fluoro-8-methoxy-4-methylquinazolin-2-yl) amino] formonitrile

To a solution of 1-(2-amino-5-fluoro-3-methoxyphenyl) ethanone (1.00 g, 5.45 mmol) in 1 ,4- dixoane (5.0 mL) was added 4 M HCI in 1 ,4-dioxane (5.00 mL). The reaction mixture was stirred for 30 min at 0 °C. The precipitated solids were collected by filtration, the filter cake was washed with MTBE (50 mLx3) and redissolved in EtOH (10.0 mL). To the above mixture was added sodium dicyanamide (0.97 g, 10.9 mmol). The resulting mixture was stirred for additional 3 h at 80 °C. The resulting mixture was concentrated under reduced pressure. The residue was further purified by trituration with MTBE (50.0 mL) to afford the title compound (1.0 g, crude) as an off-white solid, which was used for the next step without purification. LCMS: MH ⁺ 233.

Step 4 : [(6-fluoro-8-hydroxy-4-methylquinazolin-2-yl)amino]formonitr ile

To a solution of [(6-fluoro-8-methoxy-4-methylquinazolin-2-yl)amino]formonitr ile (900 mg, 3.88 mmol) in DCM (18.0 mL) was added 1 M BBr3 in DCM (7.76 mL, 7.76 mmol) dropwise at 0 °C .The resulting mixture was stirred for 2 h at 0 °C. The mixture was neutralized to pH ~10 with sat. NaHCO3 (aq.). The resulting mixture was concentrated under reduced pressure. The residue was purified by Prep-HPLC ^W to afford the title compound (300 mg, 35.5%) as an off-white solid. LCMS: MH ⁺ 219. Step 5 : 1-(6-fluoro-8-hydroxy-4-methylquinazolin-2-yl)-3-( 1-methylpiperidin-4-yl)guanidine To a solution of [(6-fluoro-8-hydroxy-4-methylquinazolin-2-yl) amino] formonitrile (100 mg, 0.458 mmol, 1.0 eq.) in DMSO (2.0 mL) was added 1-methylpiperidin-4-amine (105 mg, 0.916 mmol, 2.0 eq.) at room temperature. The mixture was irradiated with microwave radiation for 30 min at 160 °C. The mixture was allowed to cool down to room temperature. The mixture was purified by Prep-HPLC ^AG to afford the title compound (10.3 mg, 6.7%) as a yellow solid. ¹ H NMR (300 MHz, Methanol-d ₄ ) δ 6.83 - 6.74 (m, 2H), 3.65 (br, 1 H), 3.05 - 2.58 (m, 5H), 2.49 - 2.13 (m, 5H), 2.12 - 1.90 (m, 2H), 1.89 -1.40 (m, 2H), LCMS ^N : Rt 0.879 min, MH ⁺ 333.

Example 110 N-((( 1 -methylpiperidin-4-yl)amino)((4-methylquinazolin-2- yl)amino)methylene)pivalamide

To a solution of Example 3 (100 mg, 0.335 mmol) in DCM (1.0 mL) was added Et ₃ N (116 μL, 0.838 mmol) and 2,2-dimethylpropanoyl chloride (45.4 μL, 0.369 mmol) at 0 °C. The resulting mixture was stirred for additional 1 h at 0 °C. The resulting mixture was diluted with H2O (2.0 mL) and extracted with DCM (3 x 10 mL). The combined organic layers were washed with brine, dried over anhydrous Na2SO ₄ , filtered, and concentrated under reduced pressure. The residue was purified by Prep-HPLC ^AH to afford the title compound (43.8 mg, 34.2%) as a white solid. ¹ H NMR (400 MHz, Chloroform-d) δ 14.20 (s, 0.7H), 13.11 (s, 0.3H), 10.25 (s, 0.3H), 9.07 (s, 0.7H), 7.99 - 7.95 (m, 1 H), 7.81 - 7.65 (m, 2H), 7.48 - 7.36 (1 H), 4.43 (s, 0.7H), 4.18 (s, 0.3H), 2.90 (s, 2H), 2.84 (s, 1 H), 2.80 (br s, 2H), 2.38 - 2.34 (m, 5H), 2.14 - 2.10 (m, 2H), 1.75 - 1.70 (m, 2H), 1.39 (s, 6H), 1.25 (s, 3H). Rotamers observed. LCMS ^G : Rt 0.659 min MH ⁺ 383.

Examples 111-138

Each of Examples 111-138 can be synthesised by way of the methods described in General Schemes 1 to 4. BIOLOGY

Assays

Luciferase read-through assay

The read-through activity of compounds was assessed using the ADXC8 luciferase reporter cell line (generated as described in McElroy et al. PLoS Biology (2013), 11 , e1001593). Active compounds promote translational read-through of the R223X stop codon in the mutated luciferase gene which leads to generation of full-length luciferase in the compound- treated cells. The amount of luciferase is measured by adding a lysis buffer containing luciferin substrate and detecting luminescence using one of the following protocols: Protocol A

ADXC8 cells in assay media (DMEM+10% FBS) were seeded into 96-well tissue culture treated white plates (Greiner) to a density of 13,000 cells per well and incubated for 24 hours at 37 °C in an atmosphere of 5% CO2. Compounds were solubilised in DMSO at a top concentration of 10 mM, serially diluted with assay media to 25 times final concentration and added to the plate to achieve a range of final concentrations of 100 pM to 0.045 pM in a final volume of 100 μL. Plates were incubated for a further 24 hours at 37 °C in an atmosphere of 5% CO2. Cells treated with 200 pM G418 (with 1% DMSO) were used as a positive control.

Cell viability and firefly luciferase activity were measured in the same assay wells. CellTiter- Fluor™ reagent was prepared as per manufacturer’s instructions (Promega E7120) and 20 pl of 5X CellTiter-Fluor™ reagent was added to all wells, briefly mixed by orbital shaking (300-500 rpm for ~30 seconds) and the plates incubated for 30-45 minutes at 37 °C in an atmosphere of 5% CO2. Cell viability was measured by monitoring fluorescence on a CLARIOstar® plate reader (BMG Labtech). Cells containing 1% DMSO were used as a positive control and cells without addition of reagent was used as a negative control to define 100 and 0% respectively. ONE-Glo™ luciferase assay buffer (Promega E7120) was prepared according to manufacturer’s instructions and 100 pl of luciferase reagent was added to each well and the plate incubated at RT for 3 mins before measuring the luminescence on the CLARIOstar® plate reader (BMG Labtech). Cells treated with 200 pM G418 (with 1% DMSO) were used as a positive control and arbitrarily assigned a value of 100%. Cells treated with 1 % DMSO were used as the negative control to define 0% response.

Concentration effect curves were analysed using non-linear regression using the CDD Vault from Collaborative Drug Discovery (Burlingame, CA). Where datapoints at the highest concentrations tested showed bell-shaped dose-response behaviour (due to cell toxicity confirmed by viability assay), they were removed from the analysis. The bottom and top of the curves were constrained to 0 and the maximum measured response respectively. From the fitted curves, the software derived C100% values (the concentration giving the same response as 200 pM G418) for each compound. The negative logarithm of C100% is reported as pC100%. In addition to pC100% values, the maximum fold response relative to 200 pM G418 was also captured.

Protocol B

As described in McElroy et al. PLoS Biology (2013), 11 , e1001593, ADXC8 cells were seeded into white 96 well plates (Greiner) in a volume of 100 pl and incubated for 24 hours at 37 °C in an atmosphere of 5% CO2. Compounds were solubilised in DMSO at a top concentration of 10 mM and serially diluted with media in half log steps to achieve a range of final assay concentrations of 100 pM to 1 nM. Compound at each concentration (100- fold final) was added to plates and they were incubated for a further 24 hours. Luciferase activity was measured by monitoring luminescence on a TopCount luminometer (PerkinElmer) following addition of lysis/detection buffer and calculating fold change in signal relative to DMSO only controls.

Concentration effect curves were fitted using non-linear regression with Activitybase XE (I DBS). Where datapoints at the highest concentrations tested showed bell-shaped dose- response behaviour, they were removed from the analysis. From the fitted curves, the software derived C10F values (the concentration giving 10-fold response) for each compound. The negative logarithm of C10F is reported as pC10F. In addition to pC10F values, the maximum fold response over vehicle control was also captured.

Protocol C

Cells suspended in DM EM + glutamax media containing 10% FCS were seeded into clear 384 well plates (Greiner) to a density of 3200 cells per well in a volume of 25 pl and incubated for 24 hours at 37 °C in an atmosphere of 5% CO2. Compounds were solubilised in DMSO at a top concentration of 10 mM and serially diluted with media in half log steps to achieve a range of final assay concentrations of 100 pM to 1 nM. 5 μL compound at each concentration (6-fold final) was added to plates so that the final assay volume was 30 μL and they were incubated for 24 hours at 37 °C in an atmosphere of 5% CO2. Steady Gio® Luciferase Assay System (Promega) was prepared according to manufacturers’ instructions. 30 pl of Steady Gio® Luciferase Assay System was added to each well and the plates were shaken on platform at 450 rpm for 5 minutes before reading on a PHERAstar® plate reader using LUM plus protocol and calculating fold change in signal relative to DMSO only controls. Concentration effect curves were fitted using non-linear regression with Activitybase XE (I DBS). Where datapoints at the highest concentrations tested showed bell-shaped dose- response behaviour, they were removed from the analysis. From the fitted curves, the software derived C10F values (the concentration giving 10-fold response) for each compound. The negative logarithm of C10F is reported as pC10F. In addition to pC10F values, the maximum fold response over vehicle control was also captured.

Key

- pC100%/pC10F > 3.7 < 4.5 # Fold over Ctrl > 0 < 1 X Max Fold > 1 < 5

+ pC100%/pC10F > 4.5 < 5.0 ## Fold over Ctrl >1 < 5 XX Max Fold >5 < 20

++ pC100%/pC10F > 5.0 < 5.5 ### Fold over Ctrl >5 < 10 XXX Max Fold >20 < 50

+++ pC100%/pC10F > 5.5 < 6.0 #### Fold over Ctrl >10 < 15 XXXX Max Fold >50 < 100 ++++ pC100%/pC10F > 6.0 ##### Fold over Ctrl >15 XXXXX Max Fold >100

Read-through of R213X mutation in HDQ-P1 cells by Western Blot The ability of compounds to promote read-through in cells was assessed using the HDQ-P1 cell line which contain the R213X mutation in the p53 gene. Active compounds promote translational read-through of the premature stop codon encoded by this mutation, resulting in the detection of full length p53 protein by Western blot. An increase in truncated p53 may also be observed as a result of compounds suppressing nonsense mediated decay of mRNA.

HDQ-P1 cells (R213X) were treated with DMSO (0.3%), or test article for 24 h at various concentrations. Cells were washed once with ice cold 1X PBS and lysed in MRC lysis buffer (1 M Tris-HCI, pH 7.4, 0.5 M EDTA, 0.25 mM EGTA (Ethylene glycol-bis(2- aminoethylether)-N,N,N',N'-tetraacetic acid), 0.5 M sodium fluoride, 0.1 M sodium pyrophosphate, 0.2 M sodium 2-glycerol 1 -phosphate, 0.2 M sodium orthovanadate, 0.27 M sucrose, 10% (v/v) Triton X-100, 0.1 mM phenylmethylsulphonyl fluoride, 1 M dithiothreitol) and protease inhibitor mixture (Roche) centrifuged at 4 °C for 15 min at 16000xg, the supernatants, termed cell extracts, were collected and total proteins were quantified with Bradford protein assay kit (ThermoFisher). Cell extracts were denatured by incubation in NuPAGE LDS sample buffer for 10 min at 95 °C. 25 pg total protein from HDQ-1 cells were subjected to sodium dodecyl sulfatepolyacrylamide-gel electrophoresis in 4-12% pre-cast Bis-Tris gels (ThermoFisher). Proteins were transferred onto PVDF membranes, and membranes were blocked for 1 hr at room temperature in 5% skimmed milk powder in Tris- buffered saline with 0.1 % Tween® 20 Detergent (TBST) and incubated overnight with the primary monoclonal antibody DO-1 (sc-126, Santa Cruz Biotechnology, 1/1000). After three washes in TBST, the membranes were incubated with the secondary antibody (horseradish peroxidase-conjugated anti-mouse IgG [1/10,000]) for 1 hr at room temperature. The membranes were washed three times with TBST and chemiluminescence was detected with ECL Western Blot reagent kit (Cytiva-VWR) or Clarity Max ECL substrate (Bio-rad). The signal was quantified with Imaged software (NIH).

Figure 1 shows the effect of treatment of HDQ-P1 cells with Example 3 (0.41 pM, 1.23 pM, 3.7 pM, 11 pM, or 33 pM) compared to DMSO (0.3%), and G418 (200 pM).

Figure 2 shows the effect of treatment of HDQ-P1 cells with Example 4 (0.41 pM, 1.23 pM, 3.7 pM, 11 pM, or 33 pM) compared to DMSO (0.3%), and G418 (200 pM).