This invention relates to compounds that inhibit or modulate the activity of VCP (also referred to as p97), pharmaceutical compositions containing the compounds and the therapeutic uses of the compounds.

Background of the Invention

Approximately a third of all human cancers are associated with mutant Ras proteins that increase Ras activity. Activating KRAS mutations are associated with pancreatic, colorectal and lung cancers. Owing to aspects of its biology, KRAS has generally been considered to be “undruggable” for many years. The discovery of so-called ‘synthetic lethal’ cellular targets whose inhibition can selectively kill cancer cells carrying oncogenic KRAS mutations is, therefore, of significant interest in cancer therapy.

KRAS (K-Ras, K-ras, Ki-ras) is a member of the highly homologous p21 Ras family of monomeric GTPases. Three Ras isoforms (HRAS, KRAS and NRAS) are expressed in all mammalian cells and function as molecular switches downstream of cell surface receptors, such as Epidermal Growth Factor Receptor (EGFR), to stimulate cell proliferation and cell survival (Quinlan and Settleman, Future Oncol. 2009, 5, 105-16). Mutations of Ras at the conserved codons 12, 13 or 61 (corresponding to amino acid residues G12, G13 or Q61) result in an impaired ability to hydrolyse GTP, either intrinsically, or in response to GTPase activating proteins (GAPs) (Prior et al., Cancer Res. 20 2012, 72, 2457-67). Oncogenic mutations of Ras at codons 12, 13 or 61, each resulting in constitutive activation of the protein, are found in -16% of all human cancer cases. Amongst the three major Ras proteins, KRAS (K-ras) is the most frequently mutated isoform in leading causes of malignant-related death in a wide range of cancer types including colorectal (colon) and pancreatic cancer. Despite a high degree of similarity, Ras isoforms display distinct codon-specific mutational profiles (Prior et al., 2012). KRAS is typically mutated at codon 12 or codon 13 and whilst mutations at both sites are activating, due to impaired GAP binding, the position of the mutation has functional and clinical relevance.

Metastatic colorectal cancer (mCRC) is one of the leading causes of cancer related death world-wide. Overall survival is relatively poor; first-line therapy for advanced colon cancer involves treatment with anti-EGFR monoclonal antibodies such as Cetuximab in combination with standard chemotherapy. Failure to respond to Cetuximab is common and a key determinant of this resistance is the presence of activating mutations in KRAS, which are present in approximately one third of CRC tumours. Consequently, mCRC tumours are routinely genotyped for KRAS status, to predict Cetuximab responsiveness and this therapy is restricted to patients with homozygous wild type KRAS alleles.

KRAS mutations occur in over 90% of human pancreatic cancers, almost always at codon 12. The KRAS mutations occur relatively early in cancer development: while they are rare in early pancreatic intraepithelial neoplasm (PIN) lesions, they are present in the majority of advanced lesions and are near universal in frank pancreatic cancers. 5 year survival rates for patients with pancreatic cancer diagnoses are very poor: the disease is one of the most lethal of all neoplasms.

Directly targeting mutant KRAS has been largely unsuccessful as a cancer therapy. Significant efforts have been made to develop therapies targeting downstream elements of the RAS signalling pathways, including RAF, MEK, ERK and PI3K inhibitors, but none of these agents have yet been approved to treat KRAS mutant colon cancer, in particular. The difficulties with targeting mutant KRAS directly or via downstream effectors mean that there remains a need to identify new therapeutic approaches to the treatment of cancers that harbour KRAS mutations, in particular colon cancers that harbour KRAS mutations. In addition, several other types of cancer are characterised by relatively high frequencies of KRAS mutation, including pancreatic and non-small cell lung cancer.

Cancer-driving KRAS mutations are colloquially termed activating mutations and increase the fraction of protein present in its signalling-promoting GTP-bound form. These mutations typically occur at codons 12 or 13 in human KRAS (but at lower frequency elsewhere in the protein, including codons 59 or 61) and impair the GTP-ase activity of the molecule.

Such activating mutations are well documented and include those described in Siena et al., 2009, JNCI, 101 :1308-1324. The K-Ras mutation may comprise one or more activating mutations in codon 10, 12, 13, 59 and/or 61.

The presence of a K-Ras mutation in a cancer can be determined by molecular diagnostic testing methods that include, but are not restricted to, mutation-specific PCR-amplification methods, tandem PCR amplification and DNA-sequencing methods (including Sangerbased, pyrosequencing- based and mass-spectrometry-based methods), whole genome sequencing, or proteomic-based technologies (including Western blotting, immuno- histochemistry and protein mass-spectrometry). Examples of such cancers are diverse, with K-Ras mutations being present in a significant proportion of many cancer types, including but not limited to colon, lung, pancreas, prostate, endometrium, ovarian, liver, thyroid, biliary tract, stomach and ovary cancers.

Valosin-containing protein (VCP), also known as p97, is a member of the AAA (ATPases Associated with diverse cellular Activities) family. VCP is associated with a diverse range of cellular functions, but a key function of VCP is as a regulator of protein homeostasis. VCP interacts with a number of cofactors in order to extract proteins for destruction by the ubiquitin/proteasome system (UPS). For example, VCP interacts with UBX-domain cofactors in order to direct VCP to protein degradation processes. VCP regulates endoplasmic reticulum (ER) associated degradation (ERAD) which is responsible for the degradation of soluble, membrane-associated proteins. VCP is also involved in the unfolded protein response (UPR) to trigger cell death in the event of large build ups of unfolded proteins.

Many cancers rely on the ERAD and UPR pathways and therefore targeting these pathways represents a potential method of treating cancers. VCP inhibitors have been shown to promote cancer cell apotosis via UPR activation (Magnaghi et al. “Covalent and allosteric inhibitors of the ATPase VCP/p97 induce cancer cell death”, Nat Chem Biol 2013, 9(9) pp. 548-556).

It has previously been found that mutations in KRAS and BRAF correlate with sensitivity to VCP inhibition (see Anderson et al., “Targetting the AAA ATPase p97 as an approach to treat cancer through disruption of protein homeostasis”, Cancer Cell., (2015), 28(5), pp. 653-665). It is therefore envisaged that VCP inhibitors will be particularly useful in the treatment of KRAS-mutant and/or BRAF-mutant cancers.

Immune checkpoint regulators (ICR) show promise in the treatment of cancer. However, to date, the benefits of ICR treatment is generally limited to those tumours having inherent genomic instability such as Microsatellite Instable (MSI) tumours, representing less than 5% of tumours. There is therefore a need to find new mechanisms for improving ICR response in those tumours that do not naturally show genomic instability.

VCP inhibition causes accumulation of misfolded proteins, a cellular insult that activates the unfolded protein response (UPR) to either resolve the insult or activate cell death pathways (Szcz^sniak PP et al. (2022) VCP inhibition induces an unfolded protein response and apoptosis in human acute myeloid leukemia cells; Doultsinos D, et al. (2017) Control of the Unfolded Protein Response in Health and Disease).

A direct consequence of VCP or proteosome inhibition is the accumulation of K-48 polyubiquitinated protein chains and immediate dissociation of BiP from its sensors - PERK, IRE-1 , and ATF6 - to bind misfolded proteins, triggering prosurvival or, in the case of overwhelming stress, prodeath mechanisms. Some of the most common markers of UPR activation are the phosphorylation of the eukaryotic translation initiation factor 2a (elF2a), which promotes the translation and transcription of CHOP; the nonconventional splicing of XBP1 mRNA, and the release of the ATF6 cytosolic domain from the golgi, which increases the translation and transcription of endoplasmic reticulum (ER) chaperones such as BiP, amongst other proteins. Accumulation of misfolded proteins is also marked by a characteristic translocation of the chaperone calreticulin (CRT) from the ER lumen to the cell membrane surface, a signal capable of eliciting activation of the immune system.

The concept of immunogenic cell death (ICD) represents a unique cell response pattern that comprises the induction of signal cascades aiming to activate both the innate and adaptive immune response. Therefore, evidence of UPR activation gives an indication of the cellular activity upon VCP inhibition, which could in principle lead to the activation of an adaptive immune response through ICD.

WO 2014/015291 A1 (Zhou et al) describes a series of fused pyrimidine compounds as inhibitors of VCP. One example of such a VCP inhibitor is CB-5083, which has the following structure: However, Phase I clinical trials of this compound were terminated due to adverse effects on vision. It was suspected that these adverse effects resulted from off-target PDE6 inhibitory activity of the compound.

Therefore, an object of the present invention is the provision of further inhibitors of VCP, particularly those which possess good selectivity for VCP over PDE6. It is envisaged that these compounds may be useful in the treatment of cancers, particularly cancers in which KRAS is mutated.

The Invention

The present invention provides a class of compounds as inhibitors of VCP with good selectivity over PDE6. It is envisaged that the compounds will be useful in the treatment of diseases such as cancer, and particularly cancers involving mutant KRAS. In addition, based on their poor PDE6 inhibitory activity, it is envisaged that the compounds will cause substantially reduced or no unwanted side effects arising from PDE6 inhibition of the type shown by compounds of the type described in WO 2014/015291 A1.

Accordingly, in a first embodiment (Embodiment 1.1) of the invention, there is provided a compound which is a compound of the formula (1): or a pharmaceutically acceptable salt or tautomer thereof, wherein:

R ¹ is a non-aromatic Ci-s hydrocarbyl group in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the Ci-s hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ ; R ⁵ is selected from fluorine, hydroxyl, C1.2 alkoxy, amino, NH(Hyd ¹ ), N(Hyd ¹ )2 and 4- to 7-membered heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic group is optionally substituted with one or more substituents selected from fluorine and a C1.4 hydrocarbyl group optionally substituted with fluorine;

R ² is a group L-Q-R ⁴ ;

L is selected from -C(=O)NR ^a -, -NR ^a C(=O)-, -NR ^a SO ₂ -, SO ₂ NR ^a -, and N(R ^a )C(=O)N(R ^b );

Q is absent or is a C1.3 alkylene linker;

R ^a and R ^b are independently selected from hydrogen and methyl;

R ⁴ is selected from:

• hydrogen;

• a C1.8 non-aromatic hydrocarbyl group in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the C1.8 hydrocarbyl group is optionally substituted with one or more substituents selected from fluorine, hydroxyl, C1.2 alkoxy, amino, Hyd ² , NH(Hyd ² ), N(Hyd ² )2; and

• Cyc ¹ ;

Cyc ¹ is a 3- to 7-membered non-aromatic carbocyclic group or a non-aromatic 4- to 7-membered heterocyclic group containing 1 , 2 or 3 heteroatom ring members selected from N, O, S and SO2 wherein Cyc ¹ is optionally substituted with one or more substituents selected from fluorine, hydroxyl, C1.2 alkoxy, amino, Hyd ² , NH(Hyd ² ), and N(Hyd ² )2; n is 0, 1 or 2;

R ³ is selected from halogen, OH, NH2, Hyd ³ , O-Hyd ³ , NH(Hyd ³ ) and N(Hyd ³ )2

Hyd ¹ , Hyd ² and Hyd ³ in each occurrence are independently selected from C1.2 alkyl groups; and wherein in each substituent consisting or containing a non-aromatic hydrocarbyl group, the hydrocarbyl group is selected from alkyl, alkenyl, alkynyl and cycloalkyl groups and combinations thereof.

Particular and preferred compounds of the formula (1) are as defined in the Embodiments 1.2 to 1.81 below.

1.2 A compound according to Embodiment 1.1 wherein R ¹ is a non-aromatic C1.8 hydrocarbyl group (for example, an acyclic non-aromatic C1.8 hydrocarbyl group) in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the Ci-s hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.3 A compound according to Embodiment 1.1 or 1.2 wherein R ¹ is a non-aromatic Ci- 6 hydrocarbyl group(for example, an acyclic non-aromatic Ci-e hydrocarbyl group) in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the Ci-e hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.4 A compound according to any one of Embodiments 1.1 to 1.3 wherein R ¹ is a non- aromatic Ci-6 hydrocarbyl group (for example, an acyclic non-aromatic Ci-e hydrocarbyl group) in which 1 of the carbon atoms but not all is optionally replaced with a heteroatom independently selected from O and N and the Ci-e hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.5 A compound according to any one of Embodiments 1.1 to 1.4 wherein R ¹ is an acyclic non-aromatic C1.4 hydrocarbyl group in which 1 of the carbon atoms but not all is optionally replaced with a heteroatom independently selected from O and N and the C1.4 hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.6 A compound according to any one of Embodiments 1.1 to 1.4 wherein R ¹ is an acyclic non-aromatic Ci-e hydrocarbyl group, wherein the Ci-e hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.7 A compound according to any one of Embodiments 1.1 to 1.6 wherein R ¹ is an acyclic non-aromatic C1.4 hydrocarbyl group, wherein the C1.4 hydrocarbyl group is optionally substituted with one or more substituents selected from R ⁵ .

1.8 A compound according to any one of Embodiments 1.1 to 1.7 wherein R ¹ is a C1.4 alkyl group optionally substituted with one or more substituents selected from R ⁵ .

1.9 A compound according to any one of Embodiments 1.1 to 1.8 wherein R ¹ is an unsubstituted C1.4 alkyl group.

1.10 A compound according to any one of Embodiments 1.1 to 1.8 wherein R ¹ is a C1.4 alkyl group optionally substituted with one substituent selected from R ⁵ . 1.11 A compound according to any one of Embodiments 1.1 to 1.8 wherein R ¹ is a C1.4 alkyl group optionally substituted with one or two substituents selected from R ⁵ .

1.12 A compound according to any one of Embodiments 1.1 to 1.8 wherein R ¹ is a C1.4 alkyl group optionally substituted with one, two or three substituents selected from R ⁵ .

1.13 A compound according to Embodiment 1.9 wherein the unsubstituted C1.4 alkyl group is selected from methyl, ethyl, propyl and iso-propyl.

1.14 A compound according to any one of Embodiments 1 .10 to 1.12 wherein the optionally substituted C1.4 alkyl group is selected from optionally substituted methyl, ethyl, propyl and iso-propyl.

1.15 A compound according to Embodiment 1.13 or Embodiment 1.14 wherein the unsubstituted C1.4 alkyl group of Embodiment 1.13 is selected from methyl and ethyl, and the optionally substituted C1.4 alkyl group of Embodiment 1.14 is selected from an optionally substituted methyl and ethyl group.

1.16 A compound according to Embodiment 1.15 wherein the unsubstituted C1.4 alkyl group is methyl, and the optionally substituted C1.4 alkyl group is an optionally substituted methyl group.

1.17 A compound according to Embodiment 1.15 wherein the unsubstituted C1.4 alkyl group is ethyl, and the optionally substituted C1.4 alkyl group is an optionally substituted ethyl group.

1 .17A A compound according to any one of Embodiments 1 .1 to 1.17 wherein R ¹ is substituted with 0, 1 , 2 or 3 substituents selected from R ⁵ .

1 .17B A compound according to any one of Embodiments 1 .1 to 1.17 wherein R ¹ is substituted with 0, 1 or 2 substituents selected from R ⁵ .

1 .17C A compound according to any one of Embodiments 1.1 to 1.17 wherein R ¹ is substituted with 0 or 1 substituents selected from R ⁵ .

1 .17D A compound according to any one of Embodiments 1 .1 to 1.17 wherein R ¹ is substituted with 1 substituent selected from R ⁵ . 1.17E A compound according to any one of Embodiments 1.1 to 1.17 wherein R ¹ is unsubstituted.

1.18 A compound according to any one of Embodiments 1.1 to 1.8, 1.10 to 1.12 and 1.14 to 1.17E wherein R ⁵ is selected from fluorine, hydroxyl, methoxyl, amino, NH(Hyd ¹ ), N(Hyd ¹ )2 and 4-to 7-membered non-aromatic heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic groups are optionally substituted with one or more substituents selected from fluorine and a C1.4 hydrocarbyl group optionally substituted with fluorine.

1.19 A compound according to any one of Embodiments 1.1 to 1.8, 1.10 to 1.12 and 1.14 to 1.18 wherein R ⁵ is selected from fluorine, hydroxyl, methoxyl, amino, NH(Hyd ¹ ), N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic groups are optionally substituted with one or more substituents selected from fluorine and a C1.4 hydrocarbyl group optionally substituted with fluorine.

1.20 A compound according to Embodiment 1.19 wherein R ⁵ is selected from fluorine, methoxyl, NH(Hyd ¹ ), N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic group is optionally substituted with one or more substituents selected from fluorine and a saturated C1.4 hydrocarbyl group optionally substituted with fluorine.

1 .21 A compound according to Embodiment 1.20 wherein R ⁵ is selected from fluorine, methoxyl, NH(Hyd ¹ ), N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic group is optionally substituted with one or more substituents selected from fluorine and a C1.4 alkyl group optionally substituted with fluorine.

1.22 A compound according to Embodiment 1.21 wherein R ⁵ is selected from fluorine, methoxyl, N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups containing a nitrogen heteroatom ring member and optionally a further heteroatom ring member selected from O and N, wherein the heterocyclic group is optionally substituted with one or more substituents selected from fluorine and a C1.4 alkyl group optionally substituted with fluorine.

1.23 A compound according to Embodiment 1.22 wherein R ⁵ is selected from fluorine, methoxyl, N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups selected from pyrrolidine, piperidine, piperazine and morpholine, wherein the heterocyclic group is optionally substituted with one or more (e.g. one or two) substituents selected from fluorine and a C1.4 alkyl group optionally substituted with fluorine.

1 .24 A compound according to Embodiment 1.23 wherein R ⁵ is selected from fluorine, methoxyl, N(Hyd ¹ )2 and 5-to 6-membered non-aromatic heterocyclic groups selected from pyrrolidine, piperidine, piperazine and morpholine, wherein the heterocyclic group is optionally substituted with one or more (e.g. one or two) substituents selected from fluorine and a Ci- ₂ alkyl group optionally substituted with fluorine.

1 .25 A compound according to Embodiment 1.24 wherein R ⁵ is selected from fluorine, methoxyl and N(Hyd ¹ )2.

1.26 A compound according to Embodiment 1.25 wherein R ⁵ is selected from methoxyl, N(CH ₃ ) ₂ and N(CH ₂ CH ₃ ) ₂ .

1 .27 A compound according to Embodiment 1.24 wherein R ⁵ is a 5-to 6-membered non-aromatic heterocyclic group selected from pyrrolidine, piperidine, piperazine and morpholine, wherein the heterocyclic group is optionally substituted with one or more substituents selected from fluorine and a methyl group optionally substituted with fluorine.

1.28 A compound according to any one of Embodiments 1.1 to 1.25 wherein Hyd ¹ is methyl.

1.29 A compound according to any one of Embodiments 1.1 to 1.25 wherein Hyd ¹ is ethyl.

1 .30 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is selected from -C(=O)NR ^a -, -NR ^a C(=O)-, -NR ^a SO ₂ - and SO ₂ NR ^a -.

1.31 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is selected from -C(=O)NR ^a -, -NR ^a C(=O)- and -NR ^a SO ₂ -. 1.32 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is selected from -C(=O)NR ^a - and -NR ^a SO ₂ -.

1.33 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is - C(=O)NR ^a -.

1.34 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is -- NR ^a SO ₂ -.

1.35 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is -- NR ^a C(=O)-.

1.36 A compound according to any one of Embodiments 1.1 to 1.29 wherein L is - SO ₂ NR ^a -.

1.37 A compound according to any one of Embodiments 1.1 to 1.36 wherein R ^a is hydrogen.

1.38 A compound according to any one of Embodiments 1.1 to 1.36 wherein R ^a is methyl.

1.39 A compound according to any one of Embodiments 1.1 to 1.29 wherein R ^b is hydrogen.

1.40 A compound according to any one of Embodiments 1.1 to 1.29 wherein R ^b is methyl.

1.41 A compound according to any one of Embodiments 1.1 to 1.40 wherein Q is absent or is selected from CH ₂ , CH ₂ CH ₂ , CH ₂ CH ₂ CH ₂ and CH(CHs).

1.42 A compound according to any one of Embodiments 1.1 to 1.40 wherein Q is absent or CH ₂ .

1.43 A compound according to any one of Embodiments 1.1 to 1.40 wherein Q is absent.

1.44 A compound according to any one of Embodiments 1.1 to 1.40 wherein Q is CH ₂ .

1.45 A compound according to any one of Embodiments 1.1 to 1.44 wherein R ⁴ is selected from: hydrogen; a Ci-6 non-aromatic hydrocarbyl group in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the Ci-e hydrocarbyl group is optionally substituted with one or more substituents selected from hydroxyl, C1.2 alkoxy, amino, NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ .

1 .46 A compound according to any one of Embodiments 1 .1 to 1.45 wherein R ⁴ is selected from: hydrogen; an acyclic Ci-e non-aromatic hydrocarbyl group in which 1 or 2 of the carbon atoms but not all are optionally replaced with a heteroatom independently selected from O and N and the Ci-e hydrocarbyl group is optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, Hyd ² NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ .

1 .47 A compound according to any one of Embodiments 1 .1 to 1.46 wherein R ⁴ is selected from: hydrogen; an acyclic Ci-e non-aromatic hydrocarbyl group optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ .

1 .48 A compound according to any one of Embodiments 1 .1 to 1.47 wherein R ⁴ is selected from: hydrogen; an acyclic C1.4 non-aromatic hydrocarbyl group optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ . 1 .49 A compound according to any one of Embodiments 1 .1 to 1.48 wherein R ⁴ is selected from: hydrogen; an acyclic C1.4 non-aromatic hydrocarbyl group optionally substituted with one or more substituents selected from methoxyl, NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ .

1.50 A compound according to any one of Embodiments 1 .1 to 1.49 wherein R ⁴ is selected from: hydrogen; an acyclic C1.2 non-aromatic hydrocarbyl group optionally substituted with one or more substituents selected from methoxyl, NH(Hyd ² ) and N(Hyd ² )2; and

- Cyc ¹ .

1.51 A compound according to any one of Embodiments 1.1 to 1.50 wherein R ⁴ is selected from: hydrogen; a methyl or ethyl group optionally substituted with one or more substituents selected from methoxyl, NH(CHs) and N(CHs)2; and

- Cyc ¹ .

1.52 A compound according to any one of Embodiments 1.1 to 1.51 wherein Cyc ¹ is a 3- to 7-membered non-aromatic carbocyclic group or a 4- to 7-membered non-aromatic heterocyclic group containing 1 or 2 heteroatom ring members selected from N, O, S and SO2 wherein Cyci is optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, Hyd ² , NH(Hyd ² ) and N(Hyd ² )2.

1.53 A compound according to any one of Embodiments 1 .1 to 1.52 wherein Cyc ¹ is a 3- to 7-membered non-aromatic carbocyclic group or a 4- to 7-membered non-aromatic heterocyclic group containing 1 or 2 heteroatom ring members selected from N and O wherein Cyci is optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, Hyd ² , NH(Hyd ² ) and N(Hyd ² )2. 1.54 A compound according to any one of Embodiments 1 .1 to 1.53 wherein Cyc ¹ is a 3- to 6-membered non-aromatic carbocyclic group or a 5- to 6-membered non-aromatic heterocyclic group containing 1 or 2 heteroatom ring members selected from N and O wherein Cyc ¹ is optionally substituted with one or more substituents selected from hydroxyl, methoxyl, amino, Hyd ² , NH(Hyd ² ) and N(Hyd ² )2.

1.55 A compound according to any one of Embodiments 1 .1 to 1.54 wherein Cyc ¹ is a 3- to 6-membered non-aromatic carbocyclic group or a 5- to 6-membered non-aromatic heterocyclic group containing 1 or 2 heteroatom ring members selected from N and O wherein Cyc ¹ is optionally substituted with one or more substituents selected from methoxyl, Hyd ² , NH(Hyd ² ) and N(Hyd ² ) ₂ .

1.56 A compound according to any one of Embodiments 1 .1 to 1.55 wherein Cyc ¹ is a cyclopropyl group or a 5- to 6-membered non-aromatic heterocyclic group containing 2 nitrogen heteroatom ring members wherein Cyc ¹ is optionally substituted with one or more substituents selected from methoxyl, Hyd ² , NH(Hyd ² ) and N(Hyd ² ) ₂ .

1.57 A compound according to any one of Embodiments 1 .1 to 1.56 wherein Cyc ¹ is a cyclopropyl group or a piperazine group wherein Cyc ¹ is optionally substituted with one or more substituents selected from Hyd ² .

1.58 A compound according to Embodiment 1.57 wherein Hyd ¹ is methyl.

1.59 A compound according to Embodiment 1.57 wherein Hyd ¹ is ethyl.

1.60 A compound according to any one of Embodiments 1.1 to 1.59 wherein R ⁴ is hydrogen or methyl.

1.61 A compound according to any one of Embodiments 1.1 to 1.60 wherein R ⁴ is hydrogen.

1 .62 A compound according to any one of Embodiments 1 .1 to 1.60 wherein R ⁴ is methyl.

1.62A A compound according to any one of Embodiments 1.1 to 1.29 wherein R ² is - C(O)NH ₂ or -NHSO ₂ CH ₃ .

1.62B A compound according to Embodiment 1.62A wherein R ² is -C(O)NH ₂ . 1.63 A compound according to any one of Embodiments 1.1 to 1.62B wherein n is 0 or 1.

1.64 A compound according to any one of Embodiments 1.1 to 1.63 wherein n is 0.

1.65 A compound according to any one of Embodiments 1.1 to 1.63 wherein n is i. 1.66 A compound according to any one of Embodiments 1.1 to 1.63 and 1.65 wherein

R ³ is selected from fluorine, OH, NH2, Hyd ³ , O-Hyd ³ , NH(Hyd ³ ) and N(Hyd ³ )2.

1.67 A compound according to any one of Embodiments 1.1 to 1.63 and 1.65 to 1.66 wherein Hyd ³ is methyl.

1.68 A compound according to any one of Embodiments 1.1 to 1.63 and 1.65 to 1.66 wherein Hyd ³ is ethyl.

1.69 A compound according to Embodiment 1.66 wherein R ³ is selected from fluorine, OH, NH ₂ , OCH ₃ , CH ₃ , NH(CH ₃ ) and N(CH ₃ ) ₂ .

1.70 A compound according to Embodiment 1.69 wherein R ³ is selected from fluorine, methyl and OH. 1.71 A compound according to Embodiment 1.70 wherein R ³ is fluorine.

1.72 A compound according to Embodiment 1.70 wherein R ³ is OH.

1.73 A compound according to any one of Embodiments 1.1 to 1.72 having the formula (1-A): 1.74 A compound according to any one of Embodiments 1.1 to 1.72 having the formula (1-B):

1.75 A compound selected from the title compounds of Examples 1 to 23 herein.

1.75A A compound selected from the title compounds of Examples 1 to 53 herein.

1.76 A compound according to any one of Embodiments 1.1 to 1.75A which is in the form of a salt.

1.77 A compound according to Embodiment 1.76 wherein the salt is an acid addition salt.

1.78 A compound according to Embodiment 1.76 or Embodiment 1.77 wherein the salt is a pharmaceutically acceptable salt.

1.79 A compound according to any one of Embodiments 1.1 to 1.75A which is in the form of a non-salt (e.g. free base).

1.80 A compound according to any one of Embodiments 1.1 to 1.79 which is in the form of a solvate.

1.81 A compound according to Embodiment 1.80 wherein the solvate is a hydrate.

Definitions

References herein to compounds of the formula (1) are also intended to include each of the embodiments of the compounds of formula (1) as defined herein, including subformulae such as formulae (1-A) and (1-B), unless the context indicates otherwise. The compounds of any of Embodiments 1.1 to 1.81 may be referred to herein for convenience as “compounds of the invention” or like terms.

References to “carbocyclic” and “heterocyclic” groups as used herein shall, unless the context indicates otherwise, include both aromatic and non-aromatic ring systems. Thus, for example, the term “carbocyclic and heterocyclic groups” includes within its scope aromatic, non-aromatic, unsaturated, partially saturated and fully saturated carbocyclic and heterocyclic ring systems.

The carbocyclic or heterocyclic groups can be aryl or heteroaryl groups. The aryl or heteroaryl groups can be monocyclic or bicyclic groups, as defined herein. The term “aryl” as used herein refers to a carbocyclic group having aromatic character and the term “heteroaryl” is used herein to denote a heterocyclic group having aromatic character. Where the context permits, the terms “aryl” and “heteroaryl” may embrace bicyclic ring systems wherein both rings are aromatic or one ring is non-aromatic and the other is aromatic. In such bicyclic systems containing one aromatic and one non-aromatic group, the group may be attached by the aromatic ring, or by the non-aromatic ring.

The term “non-aromatic group” refers to unsaturated ring systems without aromatic character, partially saturated and fully saturated carbocyclic and heterocyclic ring systems. The terms “unsaturated” and “partially saturated” refer to rings wherein the ring structure(s) contains atoms sharing more than one valence bond e.g. the ring contains at least one multiple bond e.g. a C=C N=C bond. The term “saturated” refers to rings where there are no multiple bonds between ring atoms. Saturated carbocyclic groups include the cycloalkyl groups cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl. Partially saturated carbocyclic groups include the cycloalkenyl groups cyclopentenyl, cyclohexenyl, cycloheptenyl and cyclooctenyl. Non-aromatic monocyclic heterocyclic groups include azetidine, pyrrolidine, piperidine, azepane, piperazine, morpholine, thiomorpholine, thiomorpholine S-oxide and S,S-dioxide, pyran (2H-pyran or 4H-pyran), dihydrothiophene, dihydropyran, dihydrofuran, dihydrothiazole, tetrahydrofuran, tetrahydrothiophene, dioxane, tetrahydropyran, imidazoline, imidazolidinone, oxazoline, thiazoline, pyrazoline and pyrazolidine. Non-aromatic bicyclic heterocyclic groups include aza-bicyclo[2.2.1]heptane, aza-bicyclo[2.2.2]octane, aza-bicyclo[3.2.1]octane, hexahydro- 1 H-isoindolyl, hexahydrocyclopenta[b]pyrrol-1 (2 H)-yl , octahydroisoquinolinyl, azaspiro[2.5]octan-4-yl and 2-oxaspiro[3.3]heptan-6-yl ring systems. The term “hydrocarbyl” as used herein refers to aliphatic (saturated or unsaturated), alicyclic (saturated or unsaturated) and aromatic groups having an all-carbon backbone and consisting of carbon and hydrogen atoms, except where otherwise stated. Examples of hydrocarbyl groups include alkyl, cycloalkyl, cycloalkenyl, carbocyclic aryl, alkenyl, alkynyl, cycloalkylalkyl, cycloalkenylalkyl, and carbocyclic aralkyl, aralkenyl and aralkynyl groups. Such groups can be unsubstituted or, where stated, substituted by one or more substituents as defined herein. In certain cases, as defined herein, one or more, but not all, of the carbon atoms of the hydrocarbyl group may be replaced by another atom or group of atoms. Any of the carbon atoms in the hydrocarbyl group may be replaced with another atom or group of atoms, where indicated, including where appropriate terminal carbon atoms in a chain (i.e. CH3 groups), linker carbon atoms in a chain (i.e. CH2 groups), branching position carbon atoms in a chain (e.g. CH groups), singly bonded carbons (i.e. sp ³ hybridised carbons) and doubly bonded carbons (i.e. sp ² hybridised carbons).

In each of the foregoing embodiments, 1.1 to 1.81, if one or more carbon atoms in a hydrocarbyl group have been replaced by a heteroatom such as oxygen or nitrogen, it is preferred that the hydrocarbyl group does not contain two adjacent oxygen atoms forming a peroxide structure. It is further preferred that the location of the heteroatoms is selected so as not to give rise to unstable structures such as acetals, ketals, hemiacetals, hemiketals, aminals and hemiaminals.

The term “alkylene” (e.g. as in C1.4 straight chain or branched chain alkylene) as used herein refers to an alkanediyl group, i.e. a divalent saturated acyclic straight chain or branched chain hydrocarbon group. Examples of straight chain alkanediyl groups include methylene (CH2), ethylene (CH2CH2) and propylene ((CH2CH2CH2). Examples of branched chain alkanediyl groups include CH(CH ₃ ), CH2CH(CH ₃ )CH2 and CH ₂ (CH ₃ )CH ₂ CH ₂ .

The term “haloalkyl” as used herein refers to alkyl group substituted with one or more halogen atom substituents, and in particular fluorine substituents. Particular examples of haloalkyl groups are trifluoromethyl and difluoromethyl.

Salts

The compounds of the invention as defined in Embodiments 1.1 to 1.75A may be presented in the form of salts. The salts referred to above (and also defined in embodiments 1.76, 1.77 and 1.78) are typically acid addition salts.

The salts can be synthesized from the parent compound by conventional chemical methods such as methods described in Pharmaceutical Salts: Properties, Selection, and Use, P. Heinrich Stahl (Editor), Camille G. Wermuth (Editor), ISBN: 3-90639-026-8, Hardcover, 388 pages, August 2002. Generally, such salts can be prepared by reacting the free base form of the compound with the acid in water or in an organic solvent, or in a mixture of the two; generally, nonaqueous media such as ether, ethyl acetate, ethanol, isopropanol, or acetonitrile are used.

Acid addition salts (as defined in Embodiment 1.77) may be formed with a wide variety of acids, both inorganic and organic. Examples of acid addition salts include salts formed with an acid selected from the group consisting of acetic, 2,2-dichloroacetic, adipic, alginic, ascorbic (e.g. L-ascorbic), L-aspartic, benzenesulphonic, benzoic, 4- acetamidobenzoic, butanoic, (+) camphoric, camphor-sulphonic, (+)-(1 S)-camphor-10- sulphonic, capric, caproic, caprylic, cinnamic, citric, cyclamic, dodecylsulphuric, ethane- 1 ,2-disulphonic, ethanesulphonic, 2-hydroxyethanesulphonic, formic, fumaric, galactaric, gentisic, glucoheptonic, D-gluconic, glucuronic (e.g. D-glucuronic), glutamic (e.g. L- glutamic), a-oxoglutaric, glycolic, hippuric, hydrobromic, hydrochloric, hydriodic, isethionic, (+)-L-lactic, (±)-DL-lactic, lactobionic, maleic, malic, (-)-L-malic, malonic, (±)- DL-mandelic, methanesulphonic, naphthalene-2-sulphonic, naphthalene-1 ,5-disulphonic, 1-hydroxy-2-naphthoic, nicotinic, nitric, oleic, orotic, oxalic, palmitic, pamoic, phosphoric, propionic, L-pyroglutamic, salicylic, 4-amino-salicylic, sebacic, stearic, succinic, sulphuric, tannic, (+)-L-tartaric, thiocyanic, p-toluenesulphonic, undecylenic and valeric acids, as well as acylated amino acids and cation exchange resins.

The salt forms of the compounds of the invention are typically pharmaceutically acceptable salts, and examples of pharmaceutically acceptable salts are discussed in Berge et al., 1977, "Pharmaceutically Acceptable Salts," J. Pharm. Sci., Vol. 66, pp. 1-19. However, salts that are not pharmaceutically acceptable may also be prepared as intermediate forms which may then be converted into pharmaceutically acceptable salts. Such non-pharmaceutically acceptable salts forms, which may be useful, for example, in the purification or separation of the compounds of the invention, also form part of the invention.

Optical Isomers Where compounds of the formula contain one or more chiral centres, and can exist in the form of two or more optical isomers, references to the compounds include all optical isomeric forms thereof (e.g. enantiomers, epimers and diastereoisomers), either as individual optical isomers, or mixtures (e.g. racemic mixtures) or two or more optical isomers, unless the context requires otherwise.

The optical isomers may be characterised and identified by their optical activity (i.e. as + and - isomers, or d and I isomers) or they may be characterised in terms of their absolute stereochemistry using the “R and S” nomenclature developed by Cahn, Ingold and Prelog, see Advanced Organic Chemistry by Jerry March, 6 ^th Edition, John Wiley & Sons, New York, 2007, pages 136-163, and see also Cahn, Ingold & Prelog, Angew. Chem. Int. Ed. Engl., 1966, 5, 385-415.

Optical isomers can be separated by a number of techniques including chiral chromatography (chromatography on a chiral support) and such techniques are well known to the person skilled in the art.

As an alternative to chiral chromatography, optical isomers can be separated by forming diastereoisomeric salts with chiral acids such as (+)-tartaric acid, (-)-pyroglutamic acid, (-)-di-toluoyl-L-tartaric acid, (+)-mandelic acid, (-)-malic acid, and (-)-camphorsulphonic, separating the diastereoisomers by preferential crystallisation, and then dissociating the salts to give the individual enantiomer of the free base.

Where compounds of the invention exist as two or more optical isomeric forms, one enantiomer in a pair of enantiomers may exhibit advantages over the other enantiomer, for example, in terms of biological activity. Thus, in certain circumstances, it may be desirable to use as a therapeutic agent only one of a pair of enantiomers, or only one of a plurality of diastereoisomers. Accordingly, the invention provides compositions containing a compound having one or more chiral centres, wherein at least 55% (e.g. at least 60%, 65%, 70%, 75%, 80%, 85%, 90% or 95%) of the compound of the formula (1) is present as a single optical isomer (e.g. enantiomer or diastereoisomer). In one general embodiment, 99% or more (e.g. substantially all) of the total amount of the compound of the formula (1) may be present as a single optical isomer (e.g. enantiomer or diastereoisomer).

The term “enantiomeric excess” as used herein is used in its conventional sense to mean the percentage excess of the enantiomer of interest (e.g. compound having formulae (1- A) or (1-B)) over any other enantiomers of the compound. Where the enantiomeric excess is 80%, this corresponds to a composition of matter containing 90% desired enantiomer + 10% other enantiomers = 100% (as 90%-10% = 80%). More usually, the compounds of the invention have optical purities (enantiomeric excesses) of at least 82%, or at least 84%, or at least 86%, or at least 88%, or at least 90%, or at least 92%, or at least 94%, or at least 96%, or at least 98%, or at least 99%, or 100%.

Accordingly, further embodiments (Embodiments 1.82 to 1.85), the invention provides:

1.82 A compound as defined in any one of Embodiments 1.1 to 1.81 wherein the compound has an optical purity of:

(i) at least 80%; or

(ii) at least 82%; or

(iii) at least 84%, or

(iv) at least 86%; or

(v) at least 88%; or

(vi) at least 90%; or

(vii) at least 92%; or

(viii) at least 94%; or

(ix) at least 96%; or

(x) at least 98%; or

(xi) at least 99%; or

(xii) 100%.

1 .83 A compound according to Embodiment 1.82 wherein the compound has an optical purity of at least 98%.

1 .84 A compound according to Embodiment 1.83 wherein the compound has an optical purity of at least 99%.

1 .85 A compound according to Embodiment 1.84 wherein the compound has an optical purity of 100%.

N-Oxides Many compounds of the Embodiments 1.1 to 1.85 may form N-oxides. Where a compound contains several amine functions, one or more than one nitrogen atom may be oxidised to form an N-oxide. Particular examples of N-oxides are the N-oxides of a tertiary amine or a nitrogen atom of a nitrogen-containing heterocycle.

N-Oxides can be formed by treatment of the corresponding amine with an oxidizing agent such as hydrogen peroxide or a per-acid (e.g. a peroxycarboxylic acid), see for example Advanced Organic Chemistry, by Jerry March, 6 ^th Edition, 2009, Wiley, pages 1776-1780. More particularly, N-oxides can be made by the procedure of L. W. Deady (Syn. Comm. 1977, 7, 509-514) in which the amine compound is reacted with m-chloroperoxybenzoic acid (MCPBA), for example, in an inert solvent such as dichloromethane.

Geometric isomers and tautomers

The compounds of the invention may exist in a number of different geometric isomeric, and tautomeric forms and references to the compounds as defined in Embodiments 1.1 to 1 .85 include all such forms. For the avoidance of doubt, where a compound can exist in one of several geometric isomeric or tautomeric forms and only one is specifically described or shown, all others are nevertheless embraced by formula (1) or subgroups, subsets, preferences and examples thereof.

Isotopes

The compounds of the invention as defined in any one of Embodiments 1.1 to 1.85 may contain one or more isotopic substitutions, and a reference to a particular element includes within its scope all isotopes of the element. For example, a reference to hydrogen includes within its scope ¹ H, ² H (D), and ³ H (T). Similarly, references to carbon, nitrogen, oxygen and fluorine include within their scope respectively ¹¹ C, ¹² C, ¹³ C and ¹⁴ C; ¹³ N and ¹⁴ N; ¹⁵ O, ¹⁶ O and ¹⁸ O; and ¹⁸ F and ¹⁹ F.

Methods of labelling organic compounds with isotopes (such as deuterium) are known to the person skilled in the art (see for example, “Deuterium Discover and Applications in Organic Chemistry”, Jaemoon Yang, 2016, Elsevier and “The Organic Chemistry of Isotopic Labelling”, James R Hanson, 2019, RSC Publishing).

Typically, the compounds of the invention do not contain isotopes (such as ² H) in amounts higher than their natural abundance. In one embodiment, the percentage of the total hydrogen atoms in the compounds of the invention that are deuterium atoms is less than 2%, more typically less than 1%, more usually less than 0.1%, preferably less than 0.05% and most preferably no more than 0.02%.

In an analogous manner, a reference to a particular functional group also includes within its scope isotopic variations, unless the context indicates otherwise.

The isotopes may be radioactive or non-radioactive. In one embodiment of the invention, the compounds contain no radioactive isotopes. Such compounds are preferred for therapeutic use. In another embodiment, however, the compound may contain one or more radioisotopes. Compounds containing such radioisotopes may be useful in a diagnostic context.

Solvates

Compounds of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 may form solvates.

Preferred solvates are solvates formed by the incorporation into the solid state structure (e.g. crystal structure) of the compounds of the invention of molecules of a non-toxic pharmaceutically acceptable solvent (referred to below as the solvating solvent).

Examples of such solvents include water, alcohols (such as ethanol, isopropanol and butanol) and dimethylsulphoxide. Solvates can be prepared by recrystallising the compounds of the invention with a solvent or mixture of solvents containing the solvating solvent. Whether or not a solvate has been formed in any given instance can be determined by subjecting crystals of the compound to analysis using well known and standard techniques such as thermogravimetric analysis (TGE), differential scanning calorimetry (DSC) and X-ray crystallography.

The solvates can be stoichiometric or non-stoichiometric solvates.

Particularly preferred solvates are hydrates, and examples of hydrates include hemihydrates, monohydrates and dihydrates.

For a more detailed discussion of solvates and the methods used to make and characterise them, see Bryn et al., Solid-State Chemistry of Drugs, Second Edition, published by SSCI, Inc of West Lafayette, IN, USA, 1999, ISBN 0-967-06710-3. Prodruqs

The compounds of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 may be presented in the form of a pro-drug.

By “prodrugs” is meant for example any compound that is converted in vivo into a biologically active compound of the formula (1), as defined in any one of Embodiments 1.1 to 1.85. Prodrugs typically comprise a biologically active compound with a biologically labile functional group that can be removed in vivo to form the biologically active compound.

For example, some prodrugs are esters of the active compound (e.g., a physiologically acceptable metabolically labile ester). During metabolism, the ester group (-C(=O)OR) is cleaved to yield the active drug, either by hydrolysis in acidic or basic aqueous conditions or by esterase enzymes. Such esters may be formed by esterification, for example, of any hydroxyl groups present in the parent compound with, where appropriate, prior protection of any other reactive groups present in the parent compound, followed by deprotection if required.

Carbonates and carbamates of active compounds can also be used as prodrugs. Similarly to esters, carbonates/carbamates can be hydrolysed in acidic/basic conditions or by esterases to form the active compound in vivo. Other examples of prodrugs are phosphonates or phosphates of the active compound.

Also, some prodrugs are activated enzymatically to yield the active compound, or a compound which, upon further chemical reaction, yields the active compound (for example, as in ADEPT, GDEPT, LIDEPT, etc.). For example, the prodrug may be a sugar derivative or other glycoside conjugate, or may be an amino acid ester derivative.

Further examples of prodrugs are described in Prodrugs Challenges and Rewards, Stella et al, 1st Edition, 2007, Sprnger-Verlag New York.

Complexes and Conjugates

The invention also provides conjugates comprising a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85. The conjugates comprise the compound of formula (1) and another therapeutic agent or binding moiety covalently attached by a chemical linker. Wherein the conjugate comprises another therapeutic agent, the therapeutic agent may be, for example, a small-molecule protein inhibitor or an antibody and may be selected from the additional therapeutic agents described herein. The binding moiety may be a small-molecule motif that binds to a protein of interest, an E3 ligase or an antibody. Examples of suitable conjugates include antibody drug conjugates (ADCs) and proteolysis targeting chimeras (PROTACs).

Also encompassed by formula (1) or subgroups, subsets, preferences and examples thereof are complexes (e.g. inclusion complexes or clathrates with compounds such as cyclodextrins, or complexes with metals) of the compounds.

Biological Activity

Compounds as defined in any one of Embodiments 1.1 to 1.85 have activity as inhibitors of valosin-containing protein (VCP). As such, they may be useful in preventing or treating disease states and conditions in which VCP or mutant forms thereof play an active part.

For example, it is envisaged that the compounds of Embodiments 1.1 to 1.85 will be useful in treating a range of proliferative disorders such as cancers.

Accordingly, in further embodiments (Embodiments 2.1 to 2.9), the invention provides:

2.1 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in medicine or therapy.

2.2 A compound as defined in any one of Embodiments 1.1 to 1.85 for use as a VCP inhibitor.

2.3 A method of inhibiting a VCP protein, said method comprising in vivo or ex vivo bringing into contact with the VCP protein a compound as defined in any one of Embodiments 1.1 to 1.85.

2.4 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in preventing or treating disease states and conditions mediated by VCP or mutant forms thereof.

2.5 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for the prevention or treatment of a disease state or condition mediated by VCP or mutant forms thereof. 2.6 A method of treating a disease state or condition mediated by VCP or mutant forms thereof in a subject in need thereof, said method comprising administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

2.7 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in preventing or treating disease states and conditions characterised by abnormal expression of VCP (e.g. over-expression or expression of a mutant form of VCP).

2.8 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for the prevention or treatment of a disease state or condition characterised by abnormal expression of VCP (e.g. over-expression or expression of a mutant form of VCP).

2.9 A method of treating a disease state or condition characterised by abnormal expression of VCP (e.g. over-expression or expression of a mutant form of VCP) in a subject in need thereof, said method comprising administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

It has previously been found that mutations in KRAS correlate with sensitivity to VCP inhibition (see Anderson et al., “Targetting the AAA ATPase p97 as an approach to treat cancer through disruption of protein homeostasis”, Cancer Cell., (2015), 28(5), pp. 653- 665). It is therefore envisaged that VCP inhibitors will be particularly useful in the treatment of KRAS-mutant cancers. On this basis, it is expected that the VCP inhibitor compounds of the invention will be useful in treating certain cancers, particular those involving KRAS or mutant KRAS. This is confirmed by the results in Example 57 below.

Accordingly, in further embodiments, there are provided:

2.10 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in the prevention or treatment of a proliferative disease.

2.11 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in the prevention or treatment of cancer.

2.12 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for the prevention or treatment of a proliferative disease. 2.13 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for the prevention or treatment of cancer.

2.14 A method of preventing or treating a proliferative disease in a subject in need thereof, said method comprising administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

2.15 A method of preventing or treating cancer in a subject in need thereof, said method comprising administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

2.16 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in enhancing a therapeutic effect of radiation therapy or chemotherapy in the treatment of a proliferative disease such as cancer.

2.17 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for enhancing a therapeutic effect of radiation therapy or chemotherapy in the treatment of a proliferative disease such as cancer.

2.18 A method for the prophylaxis or treatment of a proliferative disease such as cancer, which method comprises administering to a patient in combination with radiotherapy or chemotherapy a compound as defined in any one of Embodiments 1.1 to 1.85.

Examples of proliferative disorders (e.g. cancers) as defined in Embodiments 2.10 to 2.18 include, but are not limited to carcinomas, for example carcinomas of the bladder, breast, colon, kidney, epidermis, liver, lung, oesophagus, gall bladder, ovary, pancreas, stomach, cervix, thyroid, prostate, gastrointestinal system, or skin, hematopoieitic tumours such as leukaemia, B-cell lymphoma, T-cell lymphoma, Hodgkin’s lymphoma, non-Hodgkin’s lymphoma, hairy cell lymphoma, or Burkett's lymphoma; hematopoieitic tumours of myeloid lineage, for example acute and chronic myelogenous leukaemias, myelodysplastic syndrome, or promyelocytic leukaemia; thyroid follicular cancer; tumours of mesenchymal origin, for example fibrosarcoma or rhabdomyosarcoma ; tumours of the central or peripheral nervous system, for example astrocytoma, neuroblastoma, glioma or schwannoma; melanoma; seminoma; teratocarcinoma; osteosarcoma; xeroderma pigmentosum; keratoctanthoma; thyroid follicular cancer; or Kaposi's sarcoma. One particular subset of cancers against which the compounds of Embodiments 1.1 to 1.85 should prove particulary active consists of pancreatic, lung, colorectal, biliary tract, small intestinal, sarcoma, multiple myeloma and endometrial cancers, particularly those which are KRAS mutant.

A further particular subset of cancers against which the compounds of Embodiments 1.1 to 1.85 should prove particularly active are cancers which are characterised by VCP overexpression or elevated expression of VCP or elevated activation (phosphorylation).

The ability of the compounds of the invention to inhibit VCP can be determined by means of the protocols set out in the Examples section below.

It is believed that compounds of the invention may be effective in exploiting weaknesses in cellular pathways as a result of constitutively activating KRAS mutants and therefore the compounds of the invention may be useful for the treatment of diseases and conditions mediated by modulation of KRAS.

Mutation of KRAS, resulting from a single nucleotide substitution, has been associated with various forms of cancer. In particular, KRAS mutations are found at high rates in leukemias, colon cancer, pancreatic cancer and lung cancer.

Accordingly, in further Embodiments 2.19 to 2.38, the invention also provides:

2.19 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.18 wherein the treatment further comprises administration of another anti-cancer agent or radiation therapy.

2.20 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.19 wherein the proliferative disease is a cancer mediated by KRAS.

2.21 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.20 wherein the proliferative disease is a cancer characterised by a mutant form of KRAS.

2.22 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.21 wherein the proliferative disease is a cancer characterised by a deficiency of KRAS. 2.23 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.22 wherein the proliferative disease is a cancer which is sensitive to VCP inhibition.

2.24 A compound for use, use, or method as defined in any one of Embodiments 2.10 to 2.23 wherein the proliferative disease is a cancer which contains one or more mutations of KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61).

2.25 A method for the diagnosis and treatment of a disease state or condition mediated by KRAS which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against KRAS; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound as defined in any one of Embodiments 1.1 to 1.85 or a pharmaceutically acceptable salt thereof.

2.26 Use of a compound as defined in any one of Embodiments 1.1 to 1.85 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against KRAS.

2.27 A compound as defined in any one of Embodiments 1.1 to 1.85 or a pharmaceutically acceptable salt thereof for use in the treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against KRAS.

2.28 A method for the diagnosis and treatment of a disease state or condition characterised by the presence of a mutated form of KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61) which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against KRAS; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound as defined in any one of Embodiments 1.1 to 1.85.

2.29 A method for the treatment of a disease state or condition characterised by the presence of a mutated form of KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61), which method comprises administering a therapeutically effective amount of a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 to a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against KRAS.

2.30 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in the treatment of a cancer in a patient who has been screened and has been determined as suffering from a cancer which is characterised by mutant or deficient KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61).

2.31 The use of a compound as defined in any one of Embodiments 1.1 to 1.85 for the manufacture of a medicament for the treatment of a cancer in a patient who has been screened and has been determined as suffering from a cancer which is characterised by mutant or deficient KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61).

2.32 A method for the diagnosis and treatment of a cancer which is characterised by mutant or deficient KRAS, which method comprises (i) screening a patient to determine whether a cancer from which the patient is suffering is one which is characterised by mutant or deficient KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61); and (ii) where it is indicated that the cancer is one which is characterised mutant or deficient KRAS, thereafter administering to the patient a therapeutically effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

2.33 A method of treating a subject who has been diagnosed and has been found to be suffering from a cancer which is sensitive to VCP inhibition, which method comprises administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85. 2.34 A method of treating a subject who has been diagnosed and has been found to be suffering from a cancer which is sensitive to VCP inhibition and contains one or more mutations of KRAS (for example a K-Ras mutation comprising one or more activating mutations in codon 10, 12, 13, 59 and/or 61), which method comprises administering to the subject an effective amount of a compound as defined in any one of Embodiments 1.1 to 1.85.

2.35 A method for the diagnosis and treatment of a disease state or condition mediated by VCP which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against VCP; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound as defined in any one of Embodiments 1.1 to 1.85.

2.36 The use of a compound as defined in any one of Embodiments 1.1 to 1 .85 for the manufacture of a medicament for the treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against VCP.

2.37 A compound as defined in any one of Embodiments 1.1 to 1.85 for use in the treatment or prophylaxis of a disease state or condition in a patient who has been screened and has been determined as suffering from, or being at risk of suffering from, a disease or condition which would be susceptible to treatment with a compound having activity against VCP.

2.38 A method for the diagnosis and treatment of a disease state or condition characterised by up-regulation of VCP or the presence of a mutated form of VCP, which method comprises (i) screening a patient to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against VCP; and (ii) where it is indicated that the disease or condition from which the patient is thus susceptible, thereafter administering to the patient a compound as defined in any one of Embodiments 1.1 to 1.85. The term "treatment" as used herein in the context of treating a condition i.e. state, disorder or disease, pertains generally to treatment and therapy, whether for a human or an animal (e.g. in veterinary applications), in which some desired therapeutic effect is achieved. For example, the term “treatment” encompasses the inhibition of the progress of the condition, and includes a reduction in the rate of progress, a halt in the rate of progress, amelioration of the condition, diminishment or alleviation of at least one symptom associated or caused by the condition being treated and cure of the condition. In the context of cancers, the term “treatment” encompasses effecting a reduction in tumour size, the inhibition of tumour growth, slowing the rate of growth of a tumour or halting the rate of growth of a tumour.

The term “preventing” as used herein in the context of preventing a condition pertains generally to the prophylaxis or prevention, whether for a human or an animal (e.g. in veterinary applications), in which some desired preventative effect is achieved, for example, in preventing occurance of a disease or guarding from a disease. Prevention includes complete and total blocking of all symptoms of a disorder for an indefinite period of time, the mere slowing of the onset of one or several symptoms of the disease, or making the disease less likely to occur and does not include amelioration of the condition, diminishment or alleviation of at least one symptom associated or caused by the condition being treated and cure of the condition.

References to preventing or treating cancer include within their scope alleviating or reducing the incidence e.g. of cancer.

The diagnostic methods used to determine whether a particular cancer is susceptible to treatment with the compounds of the invention can be as described below in the section headed “Methods of Diagnosis”.

Determination of biological properties

The ability of the compounds of Embodiments 1.1 to 1.85 to inhibit cell proliferation can also be determined using the protocols set out in the Examples section below.

Preferred compounds of Embodiments 1.1 to 1.85 are those having an IC50 against VCP of less than 5 pM, or less than 1 pM and preferably less than 0.1 pM.

Accordingly, in further embodiments (Embodiments 2.39 to 2.41), the invention provides: 2.39 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against VCP of less than 5 pM.

2.40 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against VCP of or less than 1 pM.

2.41 A compound according to any one of Embodiments 1.1 to 1.85 having an an IC50 against VCP of less than 0.1 pM.

Preferred compounds are selective for VCP over PDE6.

Accordingly, in further embodiments (Embodiments 2.42 to 2.51), the invention provides:

2.42 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 1 pM.

2.43 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 5 pM.

2.44 A compound according to any one of Embodiments 1.1 to 1.85 having an an IC50 against PDE6 of greater than 10 pM.

2.45 A compound according to any one of Embodiments 1.1 to 1.85 having an an IC50 against PDE6 of greater than 20 pM.

2.46 A compound according to any one of Embodiments 1.1 to 1.85 having an an IC50 against PDE6 of greater than 30 pM.

2.47 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 100 times its IC50 against VCP.

2.48 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 500 times its IC50 against VCP.

2.49 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 1000 times its IC50 against VCP.

2.50 A compound according to any one of Embodiments 1.1 to 1.85 having an IC50 against PDE6 of greater than 1500 times its IC50 against VCP. 2.51 A compound according to any one of Embodiments 2.39 to 2.50 for use in a therapy, treatment, method or use according to any one of Embodiments 2.1 to 2.38.

Methods for the Preparation of Compounds of the Invention

The invention also provides methods for the preparation of a compound of the formula (1) and pharmaceutically acceptable salts or tautomer thereof.

Compounds of the formula (1) wherein L is CON(R ^a ) (i.e. compounds of the formula (10)) can be prepared according to the following scheme:

(12) (10)

(11)

A compound of formula (11) can be hydrolysed to give amide-containing compound (12), which corresponds to a compound of formula (1) wherein R ² is CONH2. For compounds wherein R ² is other than CONH2, the compound of formula (12) can be converted to a compound of formula (10) by alkylation or amidation.

Accordingly, in another embodiment (Embodiment 3.1), the invention provides a method for preparing a compound as defined in any one of Embodiments 1.1 to 1.85 wherein L is C(O)NR ^a , or a protected derivative thereof, which method comprises the hydrolysis of a compound of formula (11): or a protected derivative thereof to form a compound of formula (12): or a protected derivative thereof and then optionally deprotecting the protected derivative of the compound of formula (12) and/or interconverting a compound of formula (12) to a compound of formula (10).

In further embodiments, there is provided:

3.2 A method according to Embodiment 3.1 wherein the hydrolysis of the compound of formula (11) involves the reaction of the compound of formula (11) with acetaldehyde oxime in the presence of a transition metal catalyst (e.g. a Pd(ll) or Pt(ll) catalyst). 3.3 A method according to Embodiment 3.2 wherein the transition metal catalyst is

Pd(PPh) ₃ .

3.4 A method according to Embodiment 3.2 or 3.3 wherein the hydrolysis is carried out in the presence of polar, protic solvent, such as ethanol, water or mixtures thereof.

Compounds of formula (1) in which L is -NR ^a C(=O)- or -NR ^a SC>2- (i.e compounds of formula (20)) can be prepared according to the scheme below: A compound of formula (21) can be reduced to give amine-containing compound (22).

The compound of formula (22) can be converted to a compound of formula (20) by reaction with a compound of the formula LG-CO-Q-R ⁴ or LG-SO2-Q-R ⁴ , wherein LG is a suitable leaving group, such as chlorine, bromine or iodine. In compounds in which R ^a is not hydrogen, the amine group can be subsequently alkylated with a compound of the formula LG-R ^a where LG is a leaving group such as chlorine, bromine or iodine.

Accordingly, in another embodiment (Embodiment 3.5), the invention provides a method for preparing a compound as defined in any one of Embodiments 1.1 to 1.85 wherein L is -NR ^a C(=O)- or -NR ^a SO2-, or a protected derivative thereof, which method comprises the reaction of a compound of formula (21): or a protected derivative thereof with a compound of the formula LG-CO-Q-R ⁴ or LG-SO2- Q-R ⁴ wherein LG is a suitable leaving group, such as chlorine, bromine or iodine, to form a compound of formula (20): or a protected derivative thereof; and optionally deprotecting the protected derivative of compound (20) and/or interconverting a compound of formula (20) to another compound of formula (1). In further embodiments, there are provided:

3.6 A method according to Embodiment 3.5 wherein the leaving group is chlorine.

3.7 A method according to Embodiment 3.5 or 3.6 wherein the reaction is carried out in the presence of an amine base (such as N,N-diisopropylethylamine (DIPEA)).

3.8 A method according to any one of Embodiments 3.5 to 3.7 wherein the reaction is carried out in the presence of a polar, aprotic solvent, such as tetra hydrofuran (THF) or diethyl ether.

Compounds of formula (22) or a protected derivative thereof, can be prepared by the reduction of compounds of formula (21): or a protected derivative thereof, with a reducing agent.

A wide range of suitable reducing agents for reducing -NO2 to -NH2 are known and include Raney nickel, palladium-on-carbon, iron, zinc, samarium, sodium hydrosulfite, tin (II) chloride and titanium (III) chloride. As described in the Examples below, one such suitable reducing agent is iron and the reduction reaction is typically carried out under acid conditions.

Accordingly, in further embodiments there is provided:

3.10 A method of preparing a compound of the formula (22) by reacting a compound of the formula (21) or a protected derivative thereof, with a reducing agent.

3.11 A method according to Embodiment 3.10 wherein the reducing agent is selected from iron, zinc and samarium.

3.12 A method according to Embodiment 3.11 wherein the reducing agent is iron. 3.13 A method according to any one of Embodiments 3.10 to 3.12 wherein the reaction is carried out in acid conditions (i.e. at a pH of less than 7).

3.14 A method according to any one of Embodiments 3.10 to 3.13 wherein the reaction is carried out in in the presence of ammonium chloride (NH4CI). 3.15 A method according to any one of Embodiments 3.10 to 3.14 wherein the reaction is carried out in in the presence of one or more polar, protic solvents, for example methanol, ethanol, water or mixtures thereof.

3.16 A method according to any one of Embodiments 3.10 to 3.15 wherein the reaction is carried out at a temperature of 50°C or greater. As an alternative to the method described in Embodiments 3.1 above, compounds of formula (1) can be prepared according to the following scheme:

Accordingly, in another embodiment (Embodiment 3.17), there is provided a method for preparing a compound as defined in any one of Embodiments 1.1 to 1.85 or a protected derivative thereof, which method comprises the reaction of a compound of formula (31): or a protected derivative thereof, wherein LG is a suitable leaving group, with a compound of the formula (32):

H ₂ N ( ^R3)n

(32) or a protected derivative thereof to form a compound of formula (1) a protected derivative thereof; and then optionally deprotecting the protected derivative of the compound of formula (1) and/or interconverting a compound of formula (1) to another compound of formula (1).

In further embodiments, there is provided:

3.18 A method according to Embodiment 3.17 wherein the leaving group, LG, is a halogen (e.g. chlorine, bromine or iodine) or a sulphonate group having the formula - SO2R ^S , wherein R ^s is a C1.4 alkyl group (e.g. methyl or ethyl) or a cycloproyl group.

3.19 A method according to Embodiment 3.18 wherein the leaving group, LG, is - SO2CH3.

3.20 A method according to any one of Embodiments 3.17 to 3.19 wherein the reaction is carried out in the presence of a base, for example an amine.

3.21 A method according to Embodiment 3.20 wherein the base is triethylamine.

3.22 A method according to any one of Embodiments 3.17 to 3.21 wherein the reaction is carried out in a polar, aprotic solvent.

3.23 A method according to Embodiment 3.22 wherein the solvent is acetonitrile.

In many of the reactions described above, it may be necessary to protect one or more groups to prevent reaction from taking place at an undesirable location on the molecule. Examples of protecting groups, and methods of protecting and deprotecting functional groups, can be found in Protective Groups in Organic Synthesis (T. Green and P. Wuts; 4 ^th Edition; Wiley, 2012). For example, compounds of formulae (10) and (20) can be protected as the following compounds (10A) and (20A): Compounds of formulae (11) and (12) can be protected as the following compounds

(11 A) and (12A): wherein the nucleophilic nitrogen of compound (11) is protected from reacting with electrophilic reagents. Similarly, compounds of formulae (21) and (22) can be protected as compounds (21A) and (22A) respectively:

The 2,4-dimethoxybenzyl protecting group can be removed by acidolysis typically with trifluoromethanesulfonic acid, trifluoracetic acid or p-toluenesulfonic acid (tosylic acid).

Accordingly, in the embodiments above, protected derivatives of compounds of formulae (10), (11), (12), (20), (21), (22) may be compounds of the formulae (10A), (11A), (12A), (20A), (21A) and (22A) respectively. Similarly, the step of deprotecting a protected derivative of a formula may involve removing the 2,4-dimethoxybenzyl group from a compound of formula (10A), (11 A), (12A), (20A), (21 A) or (22A), for example by acidolysis typically with trifluoromethanesulfonic acid, trifluoracetic acid or or p- toluenesulfonic acid (tosylic acid). Also provided herein are compounds of the formulae (10), (11), (12), (20), (21), (22), (10A), (11A), (12A), (20A), (21A) or (22A) as defined herein, which may be used as synthetic intermediates in the preparation of compounds of formula (1).

Once formed, one compound of the formula (1), or a protected derivative thereof, can be converted into another compound of the formula (1) by methods well known to the skilled person. Examples of synthetic procedures for converting one functional group into another functional group are set out in standard texts such as Advanced Organic Chemistry, by Jerry March, 6 ^th edition, 2009,, Wiley; Fiesers' Reagents for Organic Synthesis, Volumes 1-17, John Wiley, edited by Mary Fieser (ISBN: 0-471-58283-2); and Organic Syntheses, Volumes 1-8, John Wiley, edited by Jeremiah P. Freeman (ISBN: 0- 471-31192-8)).

Compounds made by the foregoing methods may be isolated and purified by any of a variety of methods well known to those skilled in the art and examples of such methods include recrystallisation and chromatographic techniques such as column chromatography (e.g. flash chromatography) and HPLC. Pharmaceutical Formulations

The compounds of the invention are typically administered to patients in the form of a pharmaceutical composition. Accordingly, in another Embodiment of the invention (Embodiment 4.1), the invention provides a pharmaceutical composition comprising a compound according to any one of Embodiments 1.1 to 1.85 and a pharmaceutically acceptable excipient.

In further embodiments, there are provided:

4.2 A pharmaceutical composition according to Embodiment 4.1 which comprises from approximately 1% (w/w) to approximately 95% (w/w) of a compound of any one of Embodiments 1.1 to 1.85 and from 99% (w/w) to 5% (w/w) of a pharmaceutically acceptable excipient or combination of excipients and optionally one or more further therapeutically active ingredients.

4.3 A pharmaceutical composition according to Embodiment 4.2 which comprises from approximately 5% (w/w) to approximately 90% (w/w) of a compound of any one of Embodiments 1.1 to 1.85 and from 95% (w/w) to 10% of a pharmaceutically excipient or combination of excipients and optionally one or more further therapeutically active ingredients.

4.4 A pharmaceutical composition according to Embodiment 4.3 which comprises from approximately 10% (w/w) to approximately 90% (w/w) of a compound of any one of Embodiments 1.1 to 1.85 and from 90% (w/w) to 10% of a pharmaceutically excipient or combination of excipients.

4.5 A pharmaceutical composition according to Embodiment 4.4 which comprises from approximately 20% (w/w) to approximately 90% (w/w) of a compound of any one of Embodiments 1.1 to 1.85 and from 80% (w/w) to 10% of a pharmaceutically excipient or combination of excipients.

4.6 A pharmaceutical composition according to Embodiment 4.5 which comprises from approximately 25% (w/w) to approximately 80% (w/w) of a compound of any one of Embodiments 1.1 to 1.85 and from 75% (w/w) to 20% of a pharmaceutically excipient or combination of excipients. The pharmaceutical compositions of the invention can be in any form suitable for oral, parenteral, topical, intranasal, intrabronchial, ophthalmic, otic, rectal, intra-vaginal, or transdermal administration. Where the compositions are intended for parenteral administration, they can be formulated for intravenous, intramuscular, intraperitoneal, subcutaneous administration or for direct delivery into a target organ or tissue by injection, infusion or other means of delivery.

Pharmaceutical dosage forms suitable for oral administration include tablets, capsules, caplets, pills, lozenges, syrups, solutions, sprays, powders, granules, elixirs and suspensions, sublingual tablets, sprays, wafers or patches and buccal patches.

Accordingly, in further embodiments, the invention provides:

4.7 A pharmaceutical composition according to any one of Embodiments 4.1 to 4.6 which is suitable for oral administration.

4.8 A pharmaceutical composition according to Embodiment 4.7 which is selected from tablets, capsules, caplets, pills, lozenges, syrups, solutions, sprays, powders, granules, elixirs and suspensions, sublingual tablets, sprays, wafers or patches and buccal patches.

4.9 A pharmaceutical composition according to Embodiment 4.8 which is selected from tablets and capsules.

4.10 A pharmaceutical composition according to any one of Embodiments 4.1 to 4.6 which is suitable for parenteral administration.

4.11 A pharmaceutical composition according to Embodiment 4.10 which is formulated for intravenous, intramuscular, intraperitoneal, subcutaneous administration or for direct delivery into a target organ or tissue by injection, infusion or other means of delivery.

4.12 A pharmaceutical composition according to Embodiment 4.11 which is a solution or suspension for injection or infusion.

Pharmaceutical compositions (e.g. as defined in any one of Embodiments 4.1 to 4.12) containing a compound according to Embodiments 1.1 to 1.85 of the invention can be formulated in accordance with known techniques, see for example, Remington’s Pharmaceutical Sciences, Mack Publishing Company, Easton, PA, USA. Thus, tablet compositions (as in Embodiment 4.9) can contain a unit dosage of active compound together with an inert diluent or carrier such as a sugar or sugar alcohol, e.g.; lactose, sucrose, sorbitol or mannitol; and/or a non-sugar derived diluent such as sodium carbonate, calcium phosphate, talc, calcium carbonate, or a cellulose or derivative thereof such as methyl cellulose, ethyl cellulose, hydroxypropyl methyl cellulose, and starches such as corn starch. Tablets may also contain such standard ingredients as binding and granulating agents such as polyvinylpyrrolidone, disintegrants (e.g. swellable crosslinked polymers such as crosslinked carboxymethylcellulose), lubricating agents (e.g. stearates), preservatives (e.g. parabens), antioxidants (e.g. BHT), buffering agents (for example phosphate or citrate buffers), and effervescent agents such as citrate/bicarbonate mixtures. Such excipients are well known and do not need to be discussed in detail here.

Capsule formulations (as in Embodiment 4.9) may be of the hard gelatin or soft gelatin variety and can contain the active component in solid, semi-solid, or liquid form. Gelatin capsules can be formed from animal gelatin or synthetic or plant derived equivalents thereof.

The solid dosage forms (e.g.; tablets, capsules etc.) can be coated or un-coated, but typically have a coating, for example a protective film coating (e.g. a wax or varnish) or a release controlling coating. The coating (e.g. a Eudragit ™ type polymer) can be designed to release the active component at a desired location within the gastro-intestinal tract. Thus, the coating can be selected so as to degrade under certain pH conditions within the gastrointestinal tract, thereby selectively release the compound in the stomach or in the ileum or duodenum.

Instead of, or in addition to, a coating, the drug can be presented in a solid matrix comprising a release controlling agent, for example a release delaying agent which may be adapted to selectively release the compound under conditions of varying acidity or alkalinity in the gastrointestinal tract. Alternatively, the matrix material or release retarding coating can take the form of an erodible polymer (e.g. a maleic anhydride polymer) which is substantially continuously eroded as the dosage form passes through the gastrointestinal tract.

Compositions for topical use include ointments, creams, sprays, patches, gels, liquid drops and inserts (for example intraocular inserts). Such compositions can be formulated in accordance with known methods. Compositions for parenteral administration (as in Embodiments 4.10 to 4.12) are typically presented as sterile aqueous or oily solutions or fine suspensions, or may be provided in finely divided sterile powder form for making up extemporaneously with sterile water for injection.

Examples of formulations for rectal or intra-vaginal administration include pessaries and suppositories which may be, for example, formed from a shaped mouldable or waxy material containing the active compound.

Compositions for administration by inhalation may take the form of inhalable powder compositions or liquid or powder sprays, and can be administrated in standard form using powder inhaler devices or aerosol dispensing devices. Such devices are well known. For administration by inhalation, the powdered formulations typically comprise the active compound together with an inert solid powdered diluent such as lactose.

The compounds of the inventions will generally be presented in unit dosage form and, as such, will typically contain sufficient compound to provide a desired level of biological activity. For example, according to any one of Embodiments 4.1 to 4.9), a composition intended for oral administration may contain from 2 milligrams to 200 milligrams of active ingredient, more usually from 10 milligrams to 100 milligrams, for example, 12.5 milligrams, 25 milligrams and 50 milligrams.

Combination Therapy

It is envisaged that the compounds of Embodiments 1.1 to 1.85 will be useful either as sole chemotherapeutic agents or, more usually, in combination therapy with chemotherapeutic agents or radiation therapy in the prophylaxis or treatment of a range of proliferative disease states or conditions. Examples of such disease states and conditions are set out above.

Particular examples of chemotherapeutic agents that may be co-administered with the compounds of Embodiments 1.1 to 1.85 include:

• Topoisomerase I inhibitors e.g. Topotecan, Irinotecan

• Antimetabolites e.g. 5-fluoro uracil (5-Fll)

• Tubulin targeting agents

• Topoisomerase II inhibitors e.g. etoposide • EGFR inhibitors (e.g. Gefitinib - see Biochemical Pharmacology 78 2009460- 468)

• mTOR inhibitors (e.g. Everolimus)

• PI3K pathway inhibitors (e.g. PI3K, PDK1)

• Akt inhibitors

• Alkylating Agents (e.g. temozolomide, cyclophosphamide)

• Monoclonal Antibodies (e.g. antibodies targeting CTLA-4, PD-1 , PD-L1, 0X40, CD52, CD40 or CD20, TIGIT, LAG3). Examples include ipilimumab, nivolumab, pembrolizumab, avelumab, durvalumab, atezolizumab, tiragolumab, relatlimab)

Anti-Hormones

Signal Transduction Inhibitors

Proteasome Inhibitors

DNA methyl transferases

Cytokines and retinoids

Hypoxia triggered DNA damaging agents (e.g. Tirapazamine)

Aromatase inhibitors

Anti Her2 antibodies, (PCT Patent Application WO 2007/056118)

Inhibitors of angiogenesis

HDAC inhibitors

MEK inhibitors

B-Raf inhibitors

ERK inhibitors

HER2 small molecule inhibitors (e.g. lapatinib)

Bcr-Abl tyrosine-kinase inhibitors (e.g. imatinib)

CDK4/6 inhibitors e.g. Ibrance

VEGFR inhibitors

IGFR-1 inhibitors • Inhibitors of the Hedgehog signalling pathway

• PARP inhibitors e.g. Olaparib

• Immune Checkpoint inhibitors

• Gemcitabine

Further examples of chemotherapeutic agents that may be co-administered with a compound as defined in any one Embodiments 1.1 to 1.85 include:

Tore 1 inhibitors

Taxanes (e.g. paclitaxel, docetaxel, cabazitaxel)

Platinum agents (e.g. cisplatin, carboplatin, oxaliplatin)

Anthracyclines (e.g. Doxorubicin)

Inhibitors of Bcl-2 family proteins e.g. ABT263 (navitoclax), a Bcl-2/Bcl-extra large (Bcl-xL) inhibitor

Cytarabine

Belinostat

EGFR inhibitors (e.g. erlotinib, osimertinib, cetuximab, neratinib, lapatinib)

MEK inhibitors (e.g. trametinib, binimetinib, selumetinib)

IGF1R inhibitors (e.g. linsitinib, ceritinib)

BRAF inhibitors (e.g. vemurafenib, dabrafenib)

PI3K inhibitors (e.g. copanlisib) mTOR inhibitors (e.g. temsirolimus)

Ipilimumab

Mifepristone

Nivolumab

Pembrolizumab • Avelumab

• Durvalumab

• Atezolizumab

• Tiragolumab

• Tebentafusp

• Relatimab

The compounds may also be administered in conjunction with radiotherapy.

Posology

The compounds may be administered over a prolonged term to maintain beneficial therapeutic effects or may be administered for a short period only. Alternatively, they may be administered in a pulsatile or continuous manner.

The compounds of the invention will be administered in an effective amount, i.e. an amount which is effective to bring about the desired therapeutic effect. For example, the "effective amount" can be a quantity of compound which, when administered to a subject suffering from cancer, slows tumour growth, ameliorates the symptoms of the disease and/or increases longevity.

The amount of VCP inhibitor compound of the invention administered to the subject will depend on the type and severity of the disease or condition and on the characteristics of the subject, such as general health, age, sex, body weight and tolerance to drugs. The skilled person will be able to determine appropriate dosages depending on these and other factors.

The compounds are generally administered to a subject in need of such administration, for example a human or animal subject (patient), preferably a human.

A typical daily dose of the compound of any of Embodiments 1.1 to 1.85 can be in the range from 100 picograms to 100 milligrams per kilogram of body weight, more typically 5 nanograms to 25 milligrams per kilogram of bodyweight, and more usually 10 nanograms to 15 milligrams per kilogram of bodyweight (e.g. 10 nanograms to 10 milligrams, and more typically 1 microgram per kilogram to 20 milligrams per kilogram, for example 1 microgram to 10 milligrams per kilogram) although higher or lower doses may be administered where required. The compound can be administered on a daily basis or on a repeat basis every 2, or 3, or 4, or 5, or 6, or 7, or 10 or 14, or 21, or 28 days for example.

In one particular dosing schedule, a patient will be given an infusion of a compound for periods of one hour daily for up to ten days in particular up to five days for one week, and the treatment repeated at a desired interval such as two to four weeks, in particular every three weeks.

More particularly, a patient may be given an infusion of a compound for periods of one hour daily for 5 days and the treatment repeated every three weeks.

In another particular dosing schedule, a patient is given an infusion over 30 minutes to 1 hour followed by maintenance infusions of variable duration, for example 1 to 5 hours, e.g. 3 hours.

In a further particular dosing schedule, a patient is given a continuous infusion for a period of 12 hours to 5 days, an in particular a continuous infusion of 24 hours to 72 hours.

Ultimately, however, the quantity of compound administered and the type of composition used will be commensurate with the nature of the disease or physiological condition being treated and will be at the discretion of the physician.

Methods of Diagnosis

Prior to administration of a compound of any one of Embodiments 1.1 to 1.85, a patient may be screened to determine whether a disease or condition from which the patient is or may be suffering is one which would be susceptible to treatment with a compound having activity against VCP. Such patient can then be treated according to the methods described above.

For example, a biological sample taken from a patient may be analysed to determine whether a condition or disease, such as cancer, that the patient is or may be suffering from is one which is characterised by a genetic abnormality or abnormal protein expression which leads to up-regulation of VCP or to sensitisation of a pathway to normal VCP activity or to over-expression of VCP. The term up-regulation includes elevated expression or over-expression, including gene amplification (i.e. multiple gene copies) and increased expression by a transcriptional effect, and hyperactivity and activation, including activation by mutations. Thus, the patient may be subjected to a diagnostic test to detect a marker characteristic of up-regulation of VCP. The term diagnosis includes screening. By marker we include genetic markers including, for example, the measurement of DNA composition to identify mutations of VCP. The term marker also includes markers which are characteristic of up-regulation of VCP, including enzyme activity, enzyme levels, enzyme state (e.g. phosphorylated or not) and mRNA levels of the aforementioned proteins.

Tumours with upregulation of VCP may be particularly sensitive to VCP inhibitors. Tumours may preferentially be screened for upregulation of VCP. Thus, the patient may be subjected to a diagnostic test to detect a marker characteristic of up-regulation of VCP. The diagnostic tests are typically conducted on a biological sample selected from tumour biopsy samples, blood samples (isolation and enrichment of shed tumour cells), stool biopsies, sputum, chromosome analysis, pleural fluid, peritoneal fluid.

Methods of identifying expression levels of VCP by immunoblotting and immunoprecipitation are described in Valle et al., “Critical Role of VCP/p97 in the Pathogenesis and Progression of Non-Small Cell Lung Carcinoma”, PLoS One, (2011), 6(12), e29073.

Testing for KRAS mutations is also known, see for example Perincheri et al., “KRAS mutation testing in clinical practice”, Expert Rev. Mol. Diagn., (2015) Mar;15(3):375-84. Common mutations include mutations at the 34, 35 and 38 positions of the KRAS gene, specifically 34G>A, 34G>C, 34G>T, 35G>A, 35G>C, 35G>T, 38G>A.

Other methods of identification and analysis of mutations and up-regulation of proteins are known to a person skilled in the art. Screening methods could include, but are not limited to, standard methods such as reverse-transcriptase polymerase chain reaction (RT-PCR) or in-situ hybridisation.

In screening by RT-PCR, the level of mRNA in the tumour is assessed by creating a cDNA copy of the mRNA followed by amplification of the cDNA by PCR. Methods of PCR amplification, the selection of primers, and conditions for amplification, are known to a person skilled in the art. Nucleic acid manipulations and PCR are carried out by standard methods, as described for example in Ausubel, F.M. et al., eds. Current Protocols in Molecular Biology, 2004, John Wiley & Sons Inc., or Innis, M.A. et-al., eds. PCR Protocols: a guide to methods and applications, 1990, Academic Press, San Diego. Reactions and manipulations involving nucleic acid techniques are also described in Sambrook et al., 2001 , 3 ^rd Ed, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press. Alternatively a commercially available kit for RT-PCR (for example Roche Molecular Biochemicals) may be used, or methodology as set forth in United States patents 4,666,828; 4,683,202; 4,801 ,531 ; 5,192,659, 5,272,057, 5,882,864, and 6,218,529 and incorporated herein by reference.

An example of an in-situ hybridisation technique for assessing mRNA expression would be fluorescence in-situ hybridisation (FISH) (see Angerer, 1987 Meth. Enzymol., 152: 649).

Generally, in situ hybridization comprises the following major steps: (1) fixation of tissue to be analyzed; (2) pre-hybridization treatment of the sample to increase accessibility of target nucleic acid, and to reduce nonspecific binding; (3) hybridization of the mixture of nucleic acids to the nucleic acid in the biological structure or tissue; (4) post-hybridization washes to remove nucleic acid fragments not bound in the hybridization, and (5) detection of the hybridized nucleic acid fragments. The probes used in such applications are typically labeled, for example, with radioisotopes or fluorescent reporters. Preferred probes are sufficiently long, for example, from about 50, 100, or 200 nucleotides to about 1000 or more nucleotides, to enable specific hybridization with the target nucleic acid(s) under stringent conditions. Standard methods for carrying out FISH are described in Ausubel, F.M. et al., eds. Current Protocols in Molecular Biology, 2004, John Wiley & Sons Inc and Fluorescence In Situ Hybridization: Technical Overview by John M. S. Bartlett in Molecular Diagnosis of Cancer, Methods and Protocols, 2nd ed.; ISBN: 1- 59259-760-2; March 2004, pps. 077-088; Series: Methods in Molecular Medicine.

Alternatively, the protein products expressed from the mRNAs may be assayed by immunohistochemistry of tumour samples, solid phase immunoassay with microtiter plates, Western blotting, 2-dimensional SDS-polyacrylamide gel electrophoresis, ELISA, flow cytometry and other methods known in the art for detection of specific proteins. Detection methods would include the use of site-specific antibodies. The skilled person will recognize that all such well-known techniques for detection of up-regulation of VCP could be applicable in the present case. Brief Description of the Drawings

Figure 1 shows the anticipated effect of VCP inihibition on a cellular expressed frame shift reporter construct where inhibition of VCP will lead to a luciferase signal, as described in Example 56.

Figures 2 and 3 are Western Blots showing an unfolded protein response upon VCP inihibition by Example 2, Thapsigargin and Bortezomib, as described in Example 58.

Figures 4 and 5 show the gating strategy used in Example 59.

Figures 6 and 7 show the Calreticulin levels in HCT116 and MC38 cells respectively, following treatment with Example 2, Example 4, Bortezomib or Cisplatin, as described in Example 59.

Figures 8 and 9 show extracellular ATP levels in HCT 116 cells treated with Example 2 or Bortezomib, as described in Example 60.

EXAMPLES

The invention will now be illustrated, but not limited, by reference to the specific embodiments described in the following examples.

In the examples, the following abbreviations are used.

AcOH acetic acid

ACN acetonitrile

ADP adenosine diphosphate aq, aqueous

ATP adenosine triphosphate

BnNH2 benzylamine

Boc tert-butyloxycarbonyl

BOC2O di-tert-butyl dicarbonate cGMP cyclic guanosine monophosphate dba dibenzylideneacetone

DCM dichloromethane DI PEA diisopropylethylamine

DMEM Dulbecco's modified Eagle medium

DMF dimethylformamide

DMP Dess-Martin periodinane

DMS dimethylsulfide

DMSO dimethylsulfoxide

DPEN 1 ,2-diphenyl- 1 ,2-ethylenediamine

DTT dithiothreitol

Et20 diethyl ether

EtOAc ethyl acetate

EtOH ethanol

HATLI 1-[bis(dimethylamino)methylene]-1 H-1,2,3-triazolo[4,5-b]pyridinium 3-oxid hexafluorophosphate

H BTU N , N , N N'-tetramethyl-O-(1 H-benzotriazol-1 -yl)uronium hexafluorophosphate

HCI hydrogen chloride

HCOOH formic acid

HEPES 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid

HPLC high performace liquid chromatography

ImH imidazole

I PA isopropyl alcohol

LCMS liquid chromatography-mass spectrometry

MeCN acetonitrile

MeOH methanol min/mins minutes

MS mass spectrometry

MsCI methanesulfonyl chloride NaOMe sodium methoxide

Na ₂ SO ₄ sodium sulfate

NCCHP(Bu)s cyanomethyltributylphosphorane

NH3 ammonia

NH4CI ammonium chloride

NMR nuclear magnetic resonance

OAc acetate

Pd ₂ (dba)3 T ris(dibenzylideneacetone)dipalladium(0)

Pd(OAc) ₂ palladium acetate

PhMe toluene

PPhs triphenylphosphine

PS-PPhs polymer-supported triphenylphosphine

RuCI(p-cymene)[(s,s)-Ts-DPEN] [(S,S)-N-(2-amino-1 ,2-diphenylethyl)-p- toluenesulfonamide]chloro(p-cymene)ruthenium(ll)

RuPhos dicyclohexylphosphino-2',6'-diisopropoxybiphenyl

TBAF tetra-n-butylammonium fluoride

TBDMS tert-butyldimethylsilyl

TBS-CI tert-butyldimethylsilyl chloride

TEA triethylamine

TFA trifluoroacetic acid

THF tetra hydrofuran

TLC thin layer chromatography

TMA trimethylaluminium

Tris-HCI tris(hydroxymethyl)aminomethane hydrochloride

Ts tosyl

UV ultraviolet XPhos 2-dicyclohexylphosphino-2',4',6'-triisopropylbiphenyl

Proton magnetic resonance ( ¹ H NMR) spectra were recorded on a Bruker 400 instrument operating at 400 MHz, in DMSO-de or MeOH-d4 (as indicated) at 27°C, unless otherwise stated and are reported as follows: chemical shift b/ppm (multiplicity where s=singlet, d=doublet, t=triplet, q=quartet, m=multiplet, br=broad, number of protons). The residual protic solvent was used as the internal reference.

Liquid chromatography and mass spectroscopy analyses were carried out using the system and operating conditions set out below. Where atoms with different isotopes are present and a single mass quoted, the mass quoted for the compound is the monoisotopic mass (i.e. ³⁵ CI; ⁷⁹ Br etc.)

ANALYTICAL HPLC/MS CONDITIONS

The LCMS data given in the following examples were obtained using one of Methods A to E below.

LCMS Method A

Samples were analysed by reverse phase HPLC-MS using a Waters 2795 Alliance HT HPLC, a Micromass ZQ mass spectrometer and a Waters 996 photodiode array UV detector. The LCMS used electrospray ionisation and one of six different chromatography systems, as follows:

Mass Spectrometer:

Ionization mode: Positive Negative

Capillary Voltage: 3.20kV -3.00kV

Cone Voltage: 30V -30V

Source Temperature: 110 °C 110 °C

Desolvation Temperature: 350°C 350°C Cone Gas Flow: 30 L/Hr 30 L/Hr

Desolvation Gas Flow: 400 L/Hr 400 L/Hr

Scan duration: 0.50 seconds 0.50 seconds

Interscan delay: 0.20 seconds 0.20 seconds

Mass range: 80 to 1000 AMU 80 to 1000 AMU LCMS was carried out using a YMC C18 50 X 2.0 mm, 1.9 micron column at 211 nm.

Column flow was 0.4 mL/min and the solvents used were 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B), with an injection volume of 1 pL.

LCMS Method B

LCMS was carried out using a ACQUITY BEH C18 50 X 2.1 mm, 1.7 micron column at 254 nm. Column flow was 0.5 mL/min and the solvents used were 0.05% TFA in water (A) and 0.05% TFA in acetonitrile (B) with an injection volume of 10 pL.

LCMS Method C

Samples were analysed by reverse phase HPLC-MS using an Agilent 1200 Series HPLC system equipped with Pumps, Degasser, Column oven, Diode array detector and a 6140 Single quadrupole Mass spectrometer. The LCMS used a combination of electrospray ionisation and atmospheric pressure chemical ionisation.

Mass Spectrometer:

Ionisation mode: Positive Negative

Cycle time: 50% 50%

Fragmentor: 75 75

Gain EMV: 1.5 1.5

Threshold: 150 150 Step-size: 0.20 0.20

Capillary voltage: 4000 V -4000 V

Charging voltage: 2000 V -2000 V

Corona current: 4 pA 4 pA

Mass range: 113 to 1000 AMU 113 to 1000 AMU

Speed: 5200 p/sec 5200 p/sec

Gas temperature: 350 C

Vaporizer temperature: 250 °C

Drying gas flow: 13.0 L/min

Nebuliser Pressure: 60 psig Polarity Switch Delay: 25 ms Ionization Switch Delay: 50 ms Cycle time: 0.43 sec/cycle Peak width: 0.03 min

LCMS was carried out using a Phenomenex Luna® C18 50 x 2.0 mm, 2.5 pm column held at 45°C with a flowrate of 1.0 mL/min. Samples were analysed at 254 nm. Samples were eluted using 0.1 % formic acid in water (A) and 0.1% formic acid in acetonitrile (B), with an injection volume of 10 pL.

LCMS Method D

Samples were analysed by reverse phase UPLC-MS using a Waters Acquity Ultra performance LC equipped with Acquity PDA detector and QDA detector. The LCMS used electrospray ionisation using the following settings:

Mass Spectrometer:

Ionization mode: Positive Negative Capillary Voltage: 0.8 kV -0.8 kV

Cone Voltage: 10 V -10 V

Source Temperature: 120 °C 120 °C

Desolvation Temperature: Default Default

Cone Gas Flow: Default Default

Desolvation Gas Flow: Default Default

Mass range: 60 to 1250 AMU 60 to 1250 AMU

LCMS was carried out using a X-Bridge C18 50 X 2.1 mm, 2.5 pm column held at 35°C with a variable flowrate. Samples were analysed at 210 nm & 254 nm. Samples were eluted using 0.1% formic acid in water (A) and 0.1% formic acid in Water : Acetonitrile (10:90) (B) with an injection volume of 2 pL.

LCMS Method E

Samples were analysed by reverse phase LC-MS using an Agilent 1260 Series HPLC system equipped with Pumps, Degasser, Column oven, Diode array detector and a 6110 Single quadrupole Mass spectrometer. The LCMS used electrospray ionisation using the following settings:

Mass Spectrometer:

Ionization mode: Positive Negative

Capillary Voltage: 3 kV -3 kV

Cone Voltage: N/A N/A

Source Temperature: N/A N/A Desolvation Temperature: 350°C 350°C

Cone Gas Flow: N/A N/A

Desolvation Gas Flow: 12 L/min 12 L/min

Mass range: 10 to 1350 AMU 10 to 1350 AMU

LCMS was carried out using a Waters CORTECS C18 30 X 4.6 mm, 2.7 pm column held at 40°C with a flowrate of 1.8 mL/min. Samples were analysed at 214 nm & 254 nm. Samples were eluted using 0.05% formic acid in water (A) and 0.05% formic acid in acetonitrile (B) with an injection volume of 1 pL.

ANALYTICAL HPLC CONDITIONS

The HPLC analytical data given in the following examples were obtained using one of Methods A to C below.

HPLC Method A

HPLC was carried out using a XB C18 150 x 4.6 mm, 5 micron column held at 30°C at

244 nm. Column flow was 1 mL/min and the solvents used were 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B) with an injection volume of 10 pL.

HPLC Method B HPLC was carried out using an Atlantis C18 150 x 4.6 mm, 5 micron column held at 30°C at 254 nm. Column flow was 1 mL/min and the solvents used were 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B) with an injection volume of 10 pL. HPLC Method C

HPLC was carried out using a Welch Xtimate C18 150 x 4.6 mm, 5 pm column held at 30°C and a flowrate of 1.0 mL/min. Samples were analysed at 210 nm and 254 nm. Samples were eluted using 0.05% trifluoroacetic acid in water (A) and acetonitrile (B) with an injection volume of 10 pL.

PREPARATIVE HPLC CONDITIONS

The preparative HPLC conditions used in the following examples were obtained using Method A or B below, as indicated.

Preparative HPLC Method A Samples were purified using a Shimadzu LC-20AP with an SPD-20A UV-Vis detector utilizing a SHIM-PACK GIST C18, 20 x 250 mm, 5 pm column and a flowrate of 20 mL/min. Compounds were eluted using 0.1% formic acid in water (A) and acetonitrile (B) with sample injections up to 1000 pL.

Preparative HPLC Method B Preparative HPLC was carried out on a Teledyne ACCQPrep HP150 Prep HPLC system with 200-400 nm UV variable wavelength detector and an ACCQPrep HP150 AS 2x2 - AutoSampler utilizing a Waters XBridge BEH C18 OBD Prep column, 5 pM 19 mm x 50 mm i.d. column and a flow rate of 24 mL/min and an injection volume of 0.2 mL to 2.5 mL. Compounds were eluted using 0.1% formic acid in water (A) and 0.1% formic acid in acetonitrile (B) over the course of 10 minutes.

PREPARATIVE CHIRAL HPLC CONDITIONS

The preparative chiral HPLC conditions used in the following examples were obtained using one of Methods A to C below, as indicated. Preparative Chiral HPLC method A

Samples were purified using an Agilent 1260 Series I nfinity-l I Preparative LC system, utilizing a Chromega Chiral CCO, 250 x 20 mm, 5pm, 1000A column and a flowrate of 28 mL/min. Compounds were eluted using 0.1% NHs.MeOH in heptane (A) and isopropanol (B) held in an isocratic manner for 40 minutes.

Preparative Chiral HPLC method B

Preparative HPLC was carried out using a Gilson Preparative LC system, using a Gilson 333 pump, a Gilson 151 UV detector and a Gilson valvemate 6 position fraction collector. Samples were purified utilizing a Phenomenex Lux Cellulose-1 , 21.2 mm x 250 mm, 5 pm column and a flowrate of 21 mL/min. Compounds were eluted with 0.2% NH3 in hexane (A) and 0.2% NH3 in EtOH (B) held in an isocratic manner for 20 minutes.

Preparative Chiral HPLC method C Preparative HPLC was carried out using a Gilson Preparative LC system, using a Gilson 333 pump, a Gilson 151 UV detector and a Gilson valvemate 6 position fraction collector. Samples were purified utilizing a Phenomenex Lux Cellulose-1 , 21.2 mm x 250 mm, 5 pm column and a flowrate of 21 mL/min. Compounds were eluted with 0.2% NH3 in hexane (A) and 0.2% NH3 in EtOH (B) held in an isocratic manner for 20 minutes.

PREPARATIVE CHIRAL SFC CONDITIONS

The preparative chiral SFC conditions used in the following examples were obtained using Method A or B below, as indicated.

Preparative Chiral SFC method A Supercritical fluid chromatography (SFC) was carried out on a Waters Investigator SFC comprising of a Waters 05962 fluid delivery module, Waters 07419 autosampler, Waters 2489 UV/Vis detector, Waters 08005 column oven, Waters 279002192 heat exchanger, Waters ABPR-20A back pressure regulator and a Waters 08127 fraction collection module utilizing a Chiralpak® AS-H (10 x 250 mm) column and repeated injections of 150 pL. UV detection was at 254 nm. The compounds were eluted using liquid CO2 (Airproducts) and 40% methanol with 0.1% diethylamine using a flowrate of 15 mL/min at 40°C and 120 bar in an isocratic manner.

Preparative Chiral SFC method B

Supercritical fluid chromatography (SFC) was carried out on a Waters Investigator SFC comprising of a Waters 05962 fluid delivery module, Waters 07419 autosampler, Waters 2489 UV/Vis detector, Waters 08005 column oven, Waters 279002192 heat exchanger, Waters ABPR-20A back pressure regulator and a Waters 08127 fraction collection module utilizing a Chiralcel® OD-H (10 x 250 mm) column and repeated injections of 150 pL. UV detection was at 254 nm. The compounds were eluted using liquid CO2 (Airproducts) and 30% methanol with 0.1% diethylamine using a flowrate of 15 mL/min at 40°C and 120 bar in an isocratic manner.

EXAMPLES 1 TO 53

The compounds of Examples 1 to 53 shown in Table 1 below have been prepared. Their NMR, HPLC and LCMS properties and the methods used to prepare them are set out in Table 2.

Examples A, B and C are provided as comparative examples.

Table 1

Synthetic Routes A to II are described below.

Synthetic Route A

(Illustrated with reference to Example 1 : 1-[4-(benzylamino)-8-methoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE 1

1A. Ethyl 3-methoxy-2-oxocyclohexane-1 -carboxylate

Potassium hydride (9.34 g, 70.3 mmol, 30% in mineral oil) was added in portions over 10 mins to a stirred solution of diethyl carbonate (19.0 mL, 156.2 mmol) in dry THF (80 mL) under a nitrogen atmosphere. The mixture was stirred for 15 mins then a solution of 2- methoxycyclohexan-1-one (2.0 g, 15.6 mmol) in dry THF (10 mL) was added dropwise over 10 mins. The reaction mixture was stirred at 70°C for 5 hours under a nitrogen atmosphere, then cooled to 0°C and a 3M solution of acetic acid was added carefully to adjust the pH to 6. The mixture was saturated with NaCI and extracted with DCM (3 x 200 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by flash column chromatography (silica gel, EtOAc: Hexane, 1 :1) to give the title compound (3.5 g, 100%) as a colourless liquid.

1 B: 8-methoxy-5,6,7,8-tetrahydroquinazoline-2,4-diol

Urea (2.2 g, 37.62 mmol) and 30% NaOMe in MeOH (6.7 mL, 37.62 mmol) were added to a solution of ethyl 3-methoxy-2-oxocyclohexane-1 -carboxylate (3.5 g, 18.81 mmol) in MeOH (17.5 mL) at room temperature under a nitrogen atmosphere. The reaction mixture was stirred at 60°C for 4 hours then cooled to room temperature and the solvents removed under reduced pressure. The residue was partitioned between water (100 mL) and EtOAc (100 mL) and the separated aqueous phase was extracted with EtOAc (3 x 100 mL). The combined organic extracts were dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (2.0 g, 70%) as an off- white solid.

1 C: 2,4-dichloro-8-methoxy-5,6,7,8-tetrahydroquinazoline

DI PEA (3.60 mL, 20.4 mmol) was added dropwise to a stirred solution of 8-methoxy- 5,6,7,8-tetrahydroquinazoline-2,4-diol (2.0 g, 10.2 mmol) in POOL (20 mL) at 0°C under a nitrogen atmosphere. The reaction mixture was heated to 100°C and stirring continued for 3 hours. The mixture was allowed to cool to room temperature and concentrated under reduced pressure. The resulting residue was added to ice cold water (50 ml) and basified (pH ~8) with aq. NaOH solution. The mixture was extracted with EtOAc (3 x 50 mL), then the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by flash column chromatography (silica gel, 0- 100% EtOAc in Hexane) to give the title compound (700 mg, 30%) as a brown solid.

1 D: N-benzyl-2-chloro-8-methoxy-5,6,7,8-tetrahydroquinazolin-4-a mine

Benzylamine (0.36 g, 3.44 mmol) and TEA (0.70 mL, 5.16 mmol) were added to a stirred solution of 2,4-dichloro-8-methoxy-5,6,7,8-tetrahydroquinazoline (0.40 g, 1.72 mmol) in MeCN (5 mL) under a nitrogen atmosphere. The mixture was stirred at 40°C for 16 hours then allowed to cool to room temperature and concentrated under reduced pressure. The crude product was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (300 mg, 57%) as a yellow solid.

1 E: 1-(4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinazolin-2- yl)-2-methyl-indole-4- carbonitrile

A stirred mixture of N-benzyl-2-chloro-8-methoxy-5,6,7,8-tetrahydroquinazolin-4-a mine (0.30 g, 0.99 mmol), 2-methyl-1 H-indole-4-carbonitrile (0.23 g, 1.48 mmol) and CS2CO3 (0.65 g, 1.98 mmol) in 1 ,4-dioxane (6 mL) was degassed with nitrogen for 15 mins. Pd2(dba)s (135 mg, 0.15 mmol) and XPhos (70 mg, 0.15 mmol) were added to the reaction mixture which was then heated to 100°C for 16 hours under a nitrogen atmosphere. The cooled mixture was partitioned between water (30 mL) and EtOAc (30 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SC and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (380 mg, 80%) as an off-white solid. 1 F: 1-r4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinazolin-2- yl1-2-methyl-indole-4- carboxamide)

A stirred mixture of 1-(4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinazolin-2- yl)-2- methyl-indole-4-carbonitrile (0.37 g, 0.87 mmol), acetaldehyde oxime (102 mg, 1.74 mmol), Pd(OAc)2 (19 mg, 0.09 mmol) and solid (polystyrene) supported PPhs (74 mg, 0.2 w/w) in EtOH (5 ml) and water (1 mL) was heated to 100°C for 16 hours then cooled to room temperature and the solvents removed under reduced pressure. The resulting residue was partitioned between EtOAc (20 mL) and water (20 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to give the title compound (150 mg, 38%) as a light yellow solid.

Synthetic Route B

(Illustrated with reference to Example 2: 1-[(8S)-4-(benzylamino)-8-methoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE 2

2A: 2,4-dichloro-5,6,7,8-tetrahydroquinazolin-8-ol

BC (2.57 g, 21.9 mmol) was added dropwise to a stirred solution of 2,4-dichloro-8- methoxy-5,6,7,8-tetrahydroquinazoline (1.6 g, 7.30 mmol) in DCM (30 mL) at 0°C under a nitrogen atmosphere. The mixture was allowed to warm to room temperature and stirred at room temperature for a further 4 hours. The reaction mixture was basified (pH ~8) with saturated aq. NaHCCh solution and was extracted with EtOAc (3 x 50 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (900 mg, 68%) as a white solid. 2B: N-f(2,4-dimethoxyphenyl)methyl1-1-phenyl-methanamine

A stirred solution of 2,4-dimethoxybenzaldehyde (5.0 g, 30.1 mmol), benzylamine (3.2 mL, 30.1 mmol) and MgSC (1.0 g, 0.2 w/w) in EtOH (23 mL) was heated to 80°C for 4 hours. The reaction mixture was cooled to 0°C and NaBH4 (1.25 g, 33.1 mmol) was added in portions over 10 mins. The mixture was allowed to warm to room temperature and stirred for a further 16 hours. The solvents were removed under reduced pressure and the resulting residue partitioned between saturated aq. NH4CI solution (50 mL) and EtOAc (50 mL). The separated aqueous phase was extracted with EtOAc (3 x 50 mL) and the combined organic extracts dried over Na2SO4 and concentrated under reduced pressure to give the title compound (5.5 g, 77%) as an off-white solid.

2C: 4-rbenzyl-r(2,4-dimethoxyphenyl)methyl1amino1-2-chloro-5,6,7 ,8-tetrahydroquinazolin- 8-0I

TEA (1 .70 mL, 12.3 mmol) was added dropwise to a stirred solution of 2,4-dichloro- 5,6,7,8-tetrahydroquinazolin-8-ol (0.90 g, 4.11 mmol) and N-[(2,4- dimethoxyphenyl)methyl]-1-phenyl-methanamine (1.16 g, 4.52 mmol) in MeCN (10 mL) under a nitrogen atmosphere. The stirred reaction mixture was heated to 40°C for 16 hours then allowed to cool to room temperature. The solvents were removed under reduced pressure to leave a residue that was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (800 mg, 70%) as a beige solid.

2D: 1-r4-rbenzyl-r(2,4-dimethoxyphenyl)methyl1amino1-8-hydroxy-5 , 6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carbonitrile

A stirred mixture of 4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-2-chloro-5, 6,7,8- tetrahydroquinazolin-8-ol (0.75 g, 1.70 mmol), 2-methyl-1 H-indole-4-carbonitrile (0.35 g, 2.22 mmol) and CS2CO3 (1.1 g, 3.40 mmol) in 1 ,4-dioxane (15 mL) was degassed with nitrogen for 15 mins. Pd2(dba)s (233 mg, 0.25 mmol) and XPhos (121 mg, 0.25 mmol) were added to the reaction mixture and then heated to 100°C for 16 hours under a nitrogen atmosphere. The mixture was partitioned between water (50 mL) and EtOAc (50 mL) and the separated aqueous phase was extracted with EtOAc (3 x 50 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (700 mg, 70%) as an off-white solid. 2E: 1-1 |-8-oxo-6,7-di in-2-

|-2-methyl-indole-4-carbonitrile

Dess-Martin periodinane (6.06 g, 14.29 mmol) was added in portions over 10 minutes to a stirred solution of 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydroxy-5 , 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carbonitrile (2.00 g, 3.57 mmol) in DCM (20 mL) at 0°C under a nitrogen atmosphere. The mixture was allowed to warm to room temperature and stirring continued for 16 hours. The solution was slowly added to saturated NaHCCh solution (100 mL) and was extracted with EtOAc (3 x 100 mL). The combined organic extracts were washed with saturated NaHCCh solution (50 mL), brine (40 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (1.0 g, 50%) as an off-white solid.

2F: 1-[(8S)-4-[benzyl-[(2,4-di |-8-hydroxy-5, 6,7,8- l-indole-4-carbonitrile

RuCI(p-cymene)[(S,S)-Ts-DPEN] (11 mg, 0.02 mmol) was added to a solution of 1-[4- [benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-oxo-6,7-dihydr o-5H-quinazolin-2-yl]-2- methyl-indole-4-carbonitrile (1.00 g, 1.79 mmol) in DMF (10 mL) at O°C under a nitrogen atmosphere. The solution was degassed with nitrogen for 15 mins then 5:2 HCOOH:TEA complex (0.75 g, 8.95 mmol) was added to the reaction mixture and stirring continued at 0°C for 1 hour. The reaction mixture was slowly added to chilled water (50 mL) and extracted with EtOAc (3 x 50 mL) then the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (560 mg, 60%, 97.6% ee) as an off-white solid.

2G: 1-[(8S)-4-[benzyl-[(2,4-di |-8-methoxy-5,6,7,8- l-indole-4-carbonitrile

NaH (120 mg, 3.0 mmol, 60% in mineral oil) was added in portions over 10 mins to a stirred solution of 1-[(8S)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydr oxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (0.56 g, 1.00 mmol) in THF (10 mL) at 0°C under a nitrogen atmosphere. The mixture was stirred for 15 mins then Mel (0.23 mL, 2.00 mmol) was added dropwise over 5 mins and the mixture heated at reflux for 5 hours. The mixture was allowed to cool to room temperature, partitioned between water (50 mL) and EtOAc (50 mL) and the separated aqueous phase extracted with EtOAc (3 x 50 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (300 mg, 64%, 94.6% ee) as an off-white solid.

2H: 1-r(8S)-4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinazol in-2-yl1-2-methyl- indole-4-carboxamide

A stirred mixture of 1-[(8S)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-meth oxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (0.30 g, 0.52 mmol), acetaldehyde oxime (62 mg, 1.04 mmol), Pd(OAc)2 (11 mg, 0.05 mmol) and solid (polystyrene) supported PPhs (60 mg, 0.2 w/w) in EtOH (5 ml) and water (1 mL) was heated to 100°C for 16 hours then cooled to room temperature and the solvents removed under reduced pressure. The resulting residue was partitioned between EtOAc (20 mL) and water (20 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to give the title compound (550 mg, 21%, 95% ee) as an off-white solid.

Synthetic Route C

(Illustrated with reference to Example 21: N-[1-[(8S)-4-(benzylamino)-8-methoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indol-4-yl]methanesulfon amide)

EXAMPLE 20

20A: 4-rbenzyl-r(2,4-dimethoxyphenyl)methyl1amino1-2-(2-methyl-4- nitro-indol-1-yl)-

5,6,7,8-tetrahydroquinazolin-8-ol A stirred solution of 4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-2-chloro-5, 6,7,8- tetrahydroquinazolin-8-ol (3.5 g, 7.9 mmol), 2-methyl-4-nitro-1 H-indole (1.6 g, 9.5 mmol) and CS2CO3 (5.16 g, 15.9 mmol) in 1 ,4-dioxane (13 mL) was degassed with nitrogen for 15 mins. Pd2(dba)s (0.72 g, 0.79 mmol) and RuPhos (0.724 g, 1.5 mmol) were added to the solution and the mixture heated to 100°C for 16 hours. The reaction mixture was allowed to cool to room temperature and partitioned between water (100 mL) and EtOAc (100 mL). The separated aqueous phase was extracted with EtOAc (3 x 150 mL), then the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by normal phase column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (3.9 g, 84%) as an off- white solid. l-4-nitro-indol-1 -vl)-6,7-

Dess-Martin periodinane (8.56 g, 20.1 mmol) was added in portions over 10 minutes to a stirred solution of 4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-2-(2-methyl-4- nitro- indol-1-yl)-5,6,7,8-tetrahydroquinazolin-8-ol (3.90 g, 6.72 mmol) in DCM (30 mL) at 0°C under a nitrogen atmosphere. The mixture was allowed to warm to room temperature and stirring continued for 16 hours. The solution was slowly added to saturated NaHCCh solution (100 mL) and was extracted with EtOAc (3 x 75 mL). The combined organic extracts were washed with saturated NaHCCh solution (75 mL), brine (40 mL), dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (2.3 g, 59%) as a light brown solid.

20C: (8S)-4-[benzyl-[(2,4-i l-4-nitro-indol-1 -yl)-

5,6, 7,8-' in-8-ol

RuCI(p-cymene)[(S,S)-Ts-DPEN] (25.3 mg, 0.039 mmol) was added to a solution of 4- [benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-2-(2-methyl-4-ni tro-indol-1-yl)-6,7-dihydro- 5H-quinazolin-8-one (2.30 g, 3.9 mmol) in DMF (70 mL) at 0°C under a nitrogen atmosphere. The solution was degassed with nitrogen for 15 mins then 5:2 HCOOH:TEA complex (1.72 g, 19.9 mmol) was added to the reaction mixture and stirring continued at 0°C for 1 hour. The reaction mixture was slowly added to chilled water (100 mL) and extracted with EtOAc (3 x 75 mL) then the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (2.10 g, 90%, 97.6% ee) as a yellow solid.

20D: (8S)-N-benzyl-N-[(2,4-di l-4-nitro-indol-

1-yl)-5,6,7,8-1 in-4-amine

NaH (517 mg, 12.9 mmol, 60% in mineral oil) was added in portions over 10 mins to a stirred solution of (8S)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-2-(2-meth yl-4- nitro-indol-1-yl)-5,6,7,8-tetrahydroquinazolin-8-ol (1.50 g, 2.50 mmol) in THF (15 mL) at 0°C under a nitrogen atmosphere. The mixture was stirred for 15 mins then Mel (1.46 g, 10.3 mmol) was added dropwise over 5 mins and the mixture heated at reflux for 5 hours. The mixture was allowed to cool to room temperature, partitioned between water (100 mL) and EtOAc (100 mL) and the separated aqueous phase extracted with EtOAc (3 x 80 mL). The combined organic extracts were dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (0.74 g, 47%) as a yellow solid.

20E: (8S)-2-(4-amino-2-methyl-indol-1-yl)-N-benzyl-N-r(2,4-dimeth oxyphenyl)methyl1-8- methoxy-5,6,7,8-tetrahydroquinazolin-4-amine

A mixture of (8S)-N-benzyl-N-[(2,4-dimethoxyphenyl)methyl]-8-methoxy-2-(2 -methyl-4- nitro-indol-1-yl)-5,6,7,8-tetrahydroquinazolin-4-amine (1.25 g, 2.1 mmol) and Raney nickel (0.25 g) in MeOH (25 mL) was stirred at room temperature under a hydrogen atmosphere for 3 hours. The reaction mixture was filtered through a bed of celite washing with MeOH (20 mL) and the filtrate was concentrated under reduced pressure to give the title compound (1.1 g, 93%) as a pale yellow solid.

20F: (8S)-2-(4-amino-2-methyl-indol-1-yl)-N-benzyl-8-methoxy-5, 6,7,8- tetrahydroquinazolin-4-amine

4 N HOI in 1,4-dioxane (8 mL) was added to a stirred solution of (8S)-2-(4-amino-2- methyl-indol-1-yl)-N-benzyl-N-[(2,4-dimethoxyphenyl)methyl]- 8-methoxy-5, 6,7,8- tetrahydroquinazolin-4-amine (400 mg, 0.70 mmol) in 1,4-dioxane (4 mL) at 0°C under a nitrogen atmosphere. The solution was stirred at 0°C for 3 hours then concentrated under reduced pressure. The resulting solid was triturated with Et20 (2 x 10 mL) and dried under reduced pressure to give the title compound (420 mg, 100%) as an off-white solid.

20G: N-n-r(8S)-4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinaz olin-2-yl1-2-methyl- indol-4-yllmethanesulfonamide

Methanesulfonyl chloride (68 uL, 0.88 mmol) was added dropwise over 5 mins to a stirred solution of (8S)-2-(4-amino-2-methyl-indol-1-yl)-N-benzyl-8-methoxy-5, 6,7,8- tetrahydroquinazolin-4-amine (200 mg, 0.47 mmol) and DIPEA (0.38 mL, 2.3 mmol) in THF (10 mL) at 0°C under a nitrogen atmosphere. The solution was allowed to warm to room temperature and stirring continued for 5 hours. The mixture was partitioned between water (50 mL) and EtOAc (50 mL), the separated aqueous phase extracted with EtOAc (3 x 50 mL) and the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC (Method A) to give the title compound (10 mg, 5%) as an off-white solid.

Synthetic Route D

(Illustrated with reference to Example 10: N-[1-[4-(benzylamino)-8-methoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indol-4-yl]propanamide)

EXAMPLE 10

10A: N-benzyl-8-methoxy-2-(2-methyl-4-nitro-indol-1-yl)-5,6,7,8-t etrahydroquinazolin-4- amine A stirred solution of N-benzyl-2-chloro-8-methoxy-5,6,7,8-tetrahydroquinazolin-4-a mine (0.20 g, 0.65 mmol), 2-methyl-4-nitro-1 H-indole (0.17 g, 0.98 mmol) and CS2CO3 (0.43 g, 1.31 mmol) in 1 ,4-dioxane (5 mL) was degassed with nitrogen 15 mins. Pd2(dba)s (90 mg, 0.10 mmol) and XPhos (47 mg, 0.10 mmol) were added to the solution and the mixture heated to 100°C for 6 hours. The reaction mixture was allowed to cool to room temperature and partitioned between water (50 mL) and EtOAc (50 mL). The separated aqueous phase was extracted with EtOAc (3 x 50 mL), then the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (240 mg, 47%) as a yellow solid. 10B: 2-(4-amino-2-methyl-indol-1-yl)-N-benzyl-8-methoxy-5,6,7,8-t etrahydroquinazolin-4- amine

A mixture of N-benzyl-8-methoxy-2-(2-methyl-4-nitro-indol-1-yl)-5, 6,7,8- tetrahydroquinazolin-4-amine (0.36 g, 0.81 mmol), Fe powder (90 mg, 1.62 mmol), NH4CI (130 mg, 2.43 mmol) in MeOH (7 mL) and water (7 mL) was stirred at 60°C for 16 hours. The mixture was allowed to cool to room temperature, diluted with MeOH (20 mL) and filtered through a bed of celite. The filtrate was concentrated under reduced pressure and the resulting residue was partitioned between water (50 mL) and EtOAc (50 mL). The separated aqueous phase was extracted with EtOAc (3 x 50 mL) and the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure to give the title compound (100 mg, 33%) as a dark brown solid.

10C: N-n-r4-(benzylamino)-8-methoxy-5,6,7,8-tetrahvdroquinazolin- 2-yl1-2-methyl-indol- 4-yllpropanamide

Propionyl chloride (10 pL, 0.13 mmol) was added dropwise over 5 mins to a stirred of 2- (4-amino-2-methyl-indol-1-yl)-N-benzyl-8-methoxy-5,6,7,8-tet rahydroquinazolin-4-amine (50 mg, 0.12 mmol) and DIPEA (70 pL, 0.36 mmol) in THF (1 mL) at 0°C under a nitrogen atmosphere. The solution was allowed to warm to room temperature and stirring continued for 3 hours. The mixture was partitioied between water (20 mL) and EtOAc (20 mL), the separated aqueous phase extracted with EtOAc (3 x 20 mL) and the combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to give the title compound (19 mg, 17%) as a yellow solid.

Synthetic Route E

(Illustrated with reference to Example 19: 1-[(8R)-4-(benzylamino)-8-methoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE 19

19A: 1-r(8R)-4-rbenzyl-r(2,4-dimethoxyphenyl)methyl1amino1-8-hydr oxy-5,6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carbonitrile RuCI(p-cymene)[(R,R)-Ts-DPEN] (7 mg, 0.01 mmol) was added to a solution of 1-[4- [benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-oxo-6,7-dihydr o-5H-quinazolin-2-yl]-2- methyl-indole-4-carbonitrile (0.60 g, 1.07 mmol) in DMF (10 mL) at 0°C under a nitrogen atmosphere. The solution was degassed with nitrogen for 15 mins then 5:2 HCOOH:TEA complex (0.45 g, 5.38 mmol) was added to the reaction mixture and stirring continued at 0°C for 3 hours. The reaction mixture was slowly added to chilled water (50 mL) and extracted with EtOAc (3 x 50 mL) then the combined organic extracts were dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (310 mg, 46%, 97.9% ee) as an off-white solid. 19B: 1-r(8R)-4-fbenzyl-r(2,4-dimethoxyphenyl)methyl1amino1-8-meth oxy-5, 6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carbonitrile

NaH (120 mg, 1.66 mmol, 60% in mineral oil) was added in portions over 10 mins to a stirred solution of 1-[(8R)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydr oxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (0.31 g, 0.55 mmol) in THF (10 mL) at 0°C under a nitrogen atmosphere. The mixture was stirred for 15 mins then Mel (0.13 mL, 1.11 mmol) was added dropwise over 5 mins and the mixture heated at reflux for 5 hours. The mixture was allowed to cool to room temperature, partitioned between water (50 mL) and EtOAc (50 mL) and the separated aqueous phase extracted with EtOAc (3 x 50 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (250 mg, 61%, 94.6% ee) as an off-white solid.

19C: 1-r(8R)-4-(benzylamino)-8-methoxy-5,6,7,8-tetrahydroquinazol in-2-yl1-2-methyl- indole-4-carboxamide

A stirred mixture of 1-[(8R)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-meth oxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (0.25 g, 0.44 mmol), acetaldehyde oxime (52 mg, 0.87 mmol), Pd(OAc)2 (10 mg, 0.04 mmol) and solid (polystyrene) supported PPhs (50 mg, 0.2 w/w) in EtOH (10 mL) and water (5 mL) was heated to 100°C for 16 hours then cooled to room temperature and the solvents removed under reduced pressure. The resulting residue was partitioned between EtOAc (30 mL) and water (30 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to give the title compound (35 mg, 24%, 96.5% ee) as an off-white solid.

Synthetic Route H

(Illustrated with reference to Example 36: 1-[(8S)-4-(benzylamino)-8-ethoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE 36 36A: 1-(4-(benzyl(2,4-dimethoxybenzyl) amino)-8-ethoxy-5,6,7,8-tetrahydroquinazolin-2- yl)-2-methyl-indole-4-carbonitrile

To a stirred suspension of NaH (110 mg, 2.68 mmol, 60% in mineral oil) in THF (5 mL) at 0°C under nitrogen was added 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8- hydroxy-5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4 -carbonitrile (500 mg, 0.89 mmol). The reaction mixture was stirred for 30 minutes before iodoethane (170 mg, 1.07 mmol) was added. The reaction mixture was heated to 60°C for 16 hours before cooling to room temperature and pouring into water (30 mL). The resulting mixture was extracted with EtOAc (2 x 50 mL) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in hexane) to afford the title compound (290 mg, 28%) as an off white solid.

36B: 1-(4-(benzylamino)-8-ethoxy-5,6,7,8-tetrahydroquinazolin-2-y l)-2-methyl-indole-4- carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl) amino)-8-ethoxy-5,6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carbonitrile (290 mg, 0.49 mmol) and acetaldehyde oxime (60 mg, 0.99 mmol) in EtOH (6 mL) at room temperature was added Pd(OAc)2 (11 mg, 0.05 mmol), polystyrene-supported PPhs (58 mg, 0.2% w/w) and water (1 mL). The reaction mixture was heated to 100°C for 16 hours before cooling to room temperature and filtering through a pad of celite washing with EtOH (10 mL). The filtrate was concentrated under reduced pressure before water (30 mL) was added, and the resulting mixture was extracted with EtOAc (2 x 30mL). The combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (60 mg, 27%) as a yellow solid.

36C: 1-r(8S)-4-(benzylamino)-8-ethoxy-5,6,7,8-tetrahvdroquinazoli n-2-yl1-2-methyl-indole- 4-carboxamide

A racemic mixture of 1-(4-(benzylamino)-8-ethoxy-5,6,7,8-tetrahydroquinazolin-2-y l)-2- methyl-indole-4-carboxamide (60 mg) was subjected to chiral preparative HPLC (method A) to afford 1-[(8S)-4-(benzylamino)-8-ethoxy-5,6,7,8-tetrahydroquinazoli n-2-yl]-2-methyl- indole-4-carboxamide (3 mg) as an off white solid. Synthetic Route I

(Illustrated with reference to Example 28: 1-(4-(benzylamino)-8-(2-morpholinoethoxy)-

5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carb oxamide)

28A: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-morpholinoethox y)-5, 6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carbonitrile

To a stirred solution of 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydroxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (500 mg, 0.89 mmol) in THF (10 mL) at 0°C under nitrogen was added NaH (180 mg, 4.47 mmol, 60% in mineral oil) portion-wise. The reaction mixture was stirred for 10 mins and then 4-(2-iodoethyl) morpholine (650 mg, 2.68 mmol) was added. The reaction mixture was then heated to 80°C for 16 hours before cooling to room temperature and pouring into water (50 mL). followed by extraction with EtOAc (3 x 50 mL). The combined organics were dried over Na2SC>4 before being concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (260 mg, 43%) as a yellow sticky solid.

28B: 1-(4-(benzyl(2,4-i i-5,6,7,8- l-indole-4-carboxamide To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-morpholinoethox y)- 5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboni trile (100 mg, 0.15 mmol) in EtOH (12 mL) and water (3 mL) was added acetaldehyde oxime (20 mg, 0.30 mmol), Pd(OAc)2 (7 mg, 0.03 mmol) and polystyrene supported PPhs (20 mg, 0.2% w/w). The resulting mixture was heated to 100°C for 12 hours and then cooled to room temperature before being concentrated under reduced pressure to afford the title compound (200 mg, assumed quant.) which was used directly without further purification.

28C: 1-(4-(benzylamino)-8-(2-morpholinoethoxy)-5,6,7,8-tetrahydro quinazolin-2-yl)-2- methyl-indole-4-carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-morpholinoethox y)- 5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboxa mide (200 mg, 0.29 mmol) in DCM (1 mL) at 0°C was added TFA (2 mL). The reaction mixture was stirred at room temperature for 1 hour before concentrating under reduced pressure. The resulting residue was purified by preparative HPLC (method A) to afford the title compound (90 mg, 57%) as a white solid.

Synthetic Route J

(Illustrated with reference to Example 30: 1-(4-(benzylamino)-8-(2-(4-methylpiperazin-1- yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indol e-4-carboxamide)

5,6, 7,8- in-2-yl)-2-i l-indole-4-carbonitrile

To a stirred solution of 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydroxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (500 mg, 0.89 mmol) in THF (5 mL) was added NaH (290 mg, 7.15 mmol, 60% in mineral oil) followed by 1-(2- chloroethyl)-4-methylpiperazine dihydrochloride (630 mg, 2.68 mmol). The reaction mixture was heated to 70°C for 16 hours and was then cooled to room temperature before a saturated solution of NH4CI (60 mL) was added. The resulting mixture was extracted with EtOAc (3 x 60 mL) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (800 mg, 66%) as a white sticky solid.

5,6, 7,8- in-2-yl)-2- l-indole-4-carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(4-methylpipera zin- 1-yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-ind ole-4-carbonitrile (400 mg, 0.58 mmol) in EtOH (10 mL) was added acetaldehyde oxime (70 mg, 1.17 mmol), Pd(OAc)2 (13 mg, 0.06 mmol), polystyrene-supported PPhs (80 mg, 0.2 w/w) and water (2 mL). The reaction mixture was heated to 100°C for 16 hours and then cooled to room temperature before being concentrated under reduced pressure. Water (60 mL) was added to the residue before being extracted with EtOAc (3 x 50m L) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting mixture was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (320 mg, 39%) as a colourless oil. l-indole-4-carboxamide

To a solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(4-methylpipera zin-1- yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indol e-4-carboxamide (320 mg, 0.45 mmol) in DCM (5 mL) was added 4 M HCI in dioxane (3 mL, 12 mmol). The reaction mixture was stirred at room temperature for 2 hours and then concentrated under reduced pressure. The resulting residue was purified by preparative HPLC (method A) to afford the title compound (28 mg, 11 %) as an off white solid. Synthetic Route K

(Illustrated with reference to Example 33: 1-(4-(benzylamino)-8-(2-(3,3-difluoropyrrolidin- 1-yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-ind ole-4-carboxamide)

Example 33: 33A: methyl 2-((4-(benzyl(2,4-dimethoxybenzyl)amino)-2-(4-cyano-2-methyl -indol-1-yl)-

5.6.7.8-tetrahydroquinazolin-8-yl)oxy)acetate

To a stirred solution of 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydroxy-

5.6.7.8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carb onitrile (2.50 g, 4.47 mmol) in THF (50 mL) at 0°C was added NaH (890 mg, 22.23 mmol, 60% in mineral oil) before stirring for 10 minutes. Methyl bromoacetate (1.3 mL, 13.40 mmol) was added and the reaction mixture was heated to 80°C for 16 hours before cooling to room temperature and pouring into water (100 mL). The resulting mixture was extracted with EtOAc (3 x 100 mL). Combined organics were dried over Na2SO4 and concentrated under reduced pressure. The residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in hexane) to afford the title compound (1.70 g, 60%) as a light yellow solid. i-8-(2-(3,3-difluoropyrrolidin-1 -yl)-2- -2-methyl-indole-4-carbonitrile

To a stirred solution of 3,3-difluoropyrrolidine (90 mg, 0.85 mmol) in toluene (0.9 mL) at 0°C was added trimethylaluminum solution (2 M in toluene) (0.57 mL 1.14 mmol). The reaction mixture was stirred for 10 minutes before methyl 2-((4-(benzyl(2,4- dimethoxybenzyl)amino)-2-(4-cyano-2-methyl-indol-1-yl)-5,6,7 ,8-tetrahydroquinazolin-8- yl)oxy)acetate (360 mg, 0.57 mmol) was added. The reaction mixture was then heated to 60°C for 5 hours before being cooled to room temperature and then poured into water (50 mL). The reaction mixture was extracted with EtOAc (3 x 50 mL) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (300 mg, 75%) as a light yellow solid.

33C: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3,3-difluoropy rrolidin-1-yl)ethoxy)- 5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboni trile

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3,3- difluoropyrrolidin-1-yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquin azolin-2-yl)-2-methyl-indole-4- carbonitrile (300 mg, 0.42 mmol) in THF (10 mL) was added borane-DMS (2 M in THF) (0.64 mL, 1.27 mmol). The reaction mixture was heated to 60°C for 5 hours and then cooled to 0°C. MeOH (2 mL) was added and the reaction mixture was heated to 50°C for 2 hours. The reaction mixture was cooled to room temperature, water (50 mL) was added and the mixture was extracted with EtOAc (3 x 50 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (150 mg, 51 %) as a light yellow solid.

'-5,6,7, 8-

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3,3- difluoropyrrolidin-1-yl)ethoxy)-5,6,7,8-tetrahydroquinazolin -2-yl)-2-methyl-indole-4- carbonitrile (150 mg, 0.22 mmol) in EtOH (1.5 mL) was added acetaldehyde oxime (30 mg, 0.43 mmol), Pd(0Ac)2 (5 mg, 0.02 mmol), polystyrene supported PPhs (30 mg, 0.2 w/w) and water (0.5 mL). The reaction mixture was heated to 100°C for 16 hours. The reaction mixture was then cooled to room temperature and concentrated under reduced pressure. Water (60 mL) was added to residue before being extracted with EtOAc (3 x 50 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (7.0 mg, 6%) as an off white solid.

Synthetic Route L

(Illustrated with reference to Example 34: 1-(4-(benzylamino)-8-(2-(3,3-difluoropyrrolidin-

1-yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-m ethyl-indole-4-carboxamide)

Example 34:

34A: 1-(4-(benzylamino)-8-(2-(3,3-difluoropyrrolidin-1-yl)-2-oxoe thoxy)-5,6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3,3- difluoropyrrolidin-1-yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquin azolin-2-yl)-2-methyl-indole-4- carbonitrile (160 mg, 0.23 mmol) in EtOH (1.5 mL) was added acetaldehyde oxime (26 mg, 0.45 mmol), Pd(OAc)2 (7.6 mg, 0.03 mmol), polystyrene supported PPhs (32 mg, 0.2 w/w) and water (0.5 mL). The reaction mixture was heated to 100°C for 16 hours. The reaction mixture was then cooled to room temperature and concentrated under reduced pressure. Water (50 mL) was added to the residue before being extracted with EtOAc (3 x 50mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (20 mg, 15%) as an off white solid.

:ic Route M

(Illustrated with reference to Example 38: 1-(4-(benzylamino)-8-(2-(3-fluoroazetidin-1-yl) ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4 -carboxamide)

To a stirred solution of methyl 2-((4-(benzyl(2,4-dimethoxybenzyl)amino)-2-(4-cyano-2- methyl-indol-1-yl)-5,6,7,8-tetrahydroquinazolin-8-yl)oxy)ace tate (0.55 g, 0.87 mmol) in THF (5 mL), LiOH (180 mg, 4.35 mmol) in water (1 mL) was added and reaction mixture heated to 60°C for 2 hours. The reaction mixture was cooled to room temperature and diluted with water (10 mL). 1 N HCI (2 mL) was added to bring the mixture to pH 3, before being extracted with EtOAc (3 x 10 mL). The combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in hexane) to afford the title compound (300 mg, 56%) as a white solid. 38B: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1-yl)-2-oxoethoxy)- 5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboni trile

To a stirred solution of 2-((4-(benzyl(2,4-dimethoxybenzyl)amino)-2-(4-cyano-2-methyl - indol-1-yl)-5,6,7,8-tetrahydroquinazolin-8-yl)oxy)acetic acid (500 mg, 0.81 mmol) in DMF (3 mL) at 0°C was added DIPEA (0.42 mL, 2.43 mmol) and HATU (460 mg, 1.21 mmol). After 30 minutes, 3-fluoroazetidine hydrochloride (110 mg, 0.97 mmol) was added and the reaction mixture was allowed to warm to room temperature and stirred for 4 hours. The reaction mixture was then poured into water (25 mL) and extracted with EtOAc (3 x 50 mL). The combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-10% MeOH in DCM) to afford the title compound (140 mg, 26%) as a white solid.

38C: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1-yl)ethoxy)-5,6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carbonitrile

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1- yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl -indole-4-carbonitrile (70 mg, 0.11 mmol) in THF (2 mL) at 0°C was added borane-DMS (2 M in THF) (0.26 mL, 0.53 mmol). The reaction mixture was allowed to warm to room temperature and stirred for 6 hours. MeOH (1 .4 mL) was added before the reaction mixture was heated to 60°C for 2 hours. The reaction mixture was then cooled to room temperature and poured into water (10 mL) before being extracted with EtOAc (3 x 15 mL). The combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by preparative TLC (10% MeOH in DCM) to afford the title compound (80 mg, quant.) as a yellow solid.

38D: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1-yl)ethoxy)-5,6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1- yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indol e-4-carbonitrile (80 mg, 0.12 mmol) in EtOH (1 mL) and water (0.1 mL) was added acetaldehyde oxime (14 mg, 0.24 mmol), polystyrene supported PPhs (16 mg, 0.2% w/w) and Pd(OAc)2 (3 mg, 0.01 mmol). The reaction mixture was heated to 100°C for 4 hours and then cooled to room temperature before water (10 mL) was added. The mixture was extracted with EtOAc (3 x 15 mL) and the combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by preparative TLC (10% MeOH in DCM) to afford the title compound (30 mg, 37%) as an off white solid.

38E: 1-(4-(benzylamino)-8-(2-(3-fluoroazetidin-1-yl)ethoxy)-5,6,7 ,8-tetrahydroquinazolin- 2-yl)-2-methyl-indole-4-carboxamide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1- yl)ethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indol e-4-carboxamide (30 mg, 0.04 mmol) in DCM (1 mL) at 0°C was added TFA (0.1 mL). The reaction mixture was then stirred for 2 hours. Saturated NaHCCh solution (25 mL) was added to bring the solution to pH 8 before being extracted with EtOAc (3 x 20 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by preparative TLC (10% MeOH in DCM) to afford the title compound (8 mg, 34%) as an off white solid.

Synthetic Route N

(Illustrated with reference to Example 39: 1-(4-(benzylamino)-8-(2-(3-fluoroazetidin-1-yl)-

2-oxoethoxy)-5,6,7,8-tetrahydro quinazolin-2-yl)-2-methyl-indole-4-carboxamide

39A: 1- ;-fluoroazetidin-1 -yl)-2-i

5, 6,7,8-- in-2-yl)-2- l-indole-4-carboxamide To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1- yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl -indole-4-carbonitrile (50 mg, 0.07 mmol) in EtOH (1 mL) was added acetaldehyde oxime (10 mg, 0.15 mmol), Pd(OAc)2 (2 mg, 0.0075 mmol), polystyrene supported PPhs (10 mg, 0.2% w/w.) and water (0.5 mL). The reaction mixture was heated to 100°C for 6 hours. The reaction mixture was then cooled to room temperature and poured into water (25 mL) before being extracted with EtOAc (2 x 50 mL). The combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by preparative TLC (5% MeOH in DCM) to afford the title compound (35 mg, 68%) as a white solid.

39B: 1-(4-(benzyl(2,4-dimethoxybenzyl)amino)-8-(2-(3-fluoroazetid in-1-yl)-2-oxoethoxy)- 5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carboxa mide

To a stirred solution of 1-(4-(benzyl(2,4-dimethoxybenzyl) amino)-8-(2-(3-fluoroazetidin-1- yl)-2-oxoethoxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl -indole-4-carboxamide (0.035 g, 0.05 mmol) in DCM (2 mL) at 0°C was added TFA (0.21 mL, 2.74 mmol) drop-wise. The reaction mixture was stirred for 2 hours before being basified using saturated NaHCOs solution (10 mL). The mixture was extracted with DCM (2 x 10 mL) and the combined organics were dried over Na2SO4 before being concentrated under reduced pressure. The resulting residue was purified by preparative TLC (5% MeOH in DCM) to afford the title compound (10 mg, 36%) as a white solid.

Synthetic Route O

(Illustrated with reference to Example 37: 1-[(8S)-4-(benzylamino)-8-isopropoxy-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide)

Example 37: '-5,6, 7,8-

To a stirred solution of 1-[(8S)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydr oxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (150 mg, 0.27 mmol) in 2- iodopropane (3 mL) under nitrogen was added Ag2<D (0.18 g, 0.80 mmol). The reaction mixture was stirred at room temperature for 84 hours and then filtered through a pad of celite. Water (30 mL) was added to the filtrate before the mixture was extracted with EtOAc (4 x 50 mL) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure. The crude product was purified by flash column chromatography (silica gel, 0-100% EtOAc in hexane) to afford the title compound (80 mg, 50%) as a light yellow solid.

37B: 1-[(8S)-4- i-8-isopropoxy-5, 6,7,8- in-2-yll-2-i indole-4-carboxamide

To a stirred solution of 1-[(8S)-4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8- isopropoxy-5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indol e-4-carbonitrile (80 mg, 0.13 mmol) in EtOH (3 mL) was added acetaldehyde oxime (16 mg, 0.27 mmol), Pd(OAc)2 (6 mg, 0.03 mmol), polystyrene supported PPhs (16 mg, 0.2 w/w) and water (0.5 mL). The reaction mixture was heated to 100°C for 16 hours and then concentrated under reduced pressure. Water (60 mL) was added to the residue and extracted with EtOAc (3 x 50 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by reverse phase column chromatography (C18, 5-95% MeCN in water) to afford the title compound (15 mg, 24%) as a light yellow solid.

:ic Route P

(Illustrated with reference to Example 32: 1-(4-(benzylamino)-8-((1-methylpiperidin-4- yl)oxy)-5,6,7,8-tetrahydroquinazolin-2-yl)-2-methyl-indole-4 -carboxamide)

Example 32:

32A: 1-(4-(benzylamino)-8-hvdroxy-5,6,7,8-tetrahvdroquinazolin-2- yl)-2-methyl-indole-4- carbonitrile To a stirred solution of 1-[4-[benzyl-[(2,4-dimethoxyphenyl)methyl]amino]-8-hydroxy- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (3.00 g, 5.36 mmol) in 1 ,4-dioxane (5 mL) at 0°C was added 4M HCI in 1 ,4-dioxane (20 mL, 80 mmol). The reaction mixture was allowed to warm to room temperature and then stirred for 4 hours, before being concentrated under reduced pressure. Saturated NaHCCh solution (60 mL) was added before the mixture was extracted with EtOAc (3 x 100 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (1.80 g, 82%) as a yellow solid.

32B: 1-(4-(benzylamino)-8-chloro-5,6,7,8-tetrahvdroquinazolin-2-y l)-2-methyl-indole-4- carbonitrile

To a stirred solution of 1-(4-(benzylamino)-8-hydroxy-5,6,7,8-tetrahydroquinazolin-2- yl)-2- methyl-indole-4-carbonitrile (1.80 g, 4.40 mmol) in DCM (20 mL) and DMF (1 mL) at 0°C was added thionyl chloride (0.64 ml, 8.79 mmol) drop-wise. The reaction mixture was allowed to come to room temperature and stirred for 16 hours, before being concentrated under reduced pressure. Saturated NaHCCh solution (40 mL) was added before the mixture was extracted with EtOAc (4 x 50 mL). The combined organics were dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in hexane) to afford the title compound (400 mg, 21%) as a yellow solid.

32C: 1-(4-(benzylamino)-8-((1-methylpiperidin-4-yl)oxy)-5,6,7,8-t etrahydroquinazolin-2- yl)-2-methyl-indole-4-carbonitrile

To a stirred solution in 1-methylpiperidin-4-ol (540 mg, 4.67 mmol) in THF (10 mL) under nitrogen was added NaH (0.30 g, 7.48 mmol, 60% in mineral oil) portion wise. 1-(4- (benzylamino)-8-chloro-5,6,7,8-tetrahydroquinazolin-2-yl)-2- methyl-indole-4-carbonitrile (400 mg, 0.93 mmol) was added before the reaction mixture was heated to 90°C for 16 hours. The reaction mixture was then cooled to room temperature before water (30 mL) was added and the mixture was extracted with EtOAc (3 x 30 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-10% MeOH in DCM) to afford the title compound (80 mg, 17%) as a yellow solid.

32D: 1-(4-(benzylamino)-8-((1-methylpiperidin-4-yl)oxy)-5,6,7,8-t etrahydroquinazolin-2- yl)-2-methyl-indole-4-carboxamide

To a stirred solution of 1-(4-(benzylamino)-8-((1-methylpiperidin-4-yl)oxy)-5, 6,7,8- tetrahydroquinazolin-2-yl)-2-methyl-indole-4-carbonitrile (75 mg, 0.15 mmol) in EtOH (1 mL) was added acetaldehyde oxime (20 mg, 0.30 mmol), Pd(OAc)2 (3.33 mg, 0.015 mmol), polystyrene supported PPhs (7.75 mg, 0.2 w/w) and water (0.1 mL). The reaction mixture was heated to 100°C for 2 hours before being concentrated under reduced pressure. The resulting residue was added to water (7 mL) before being extracted with EtOAc (3 x 5 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by preparative TLC (10% MeOH in DCM) to afford the title compound (15 mg, 19%) as a white solid.

Synthetic Route Q (Illustrated with reference to Example 42: 1-[(8S)-4-[(3-fluorophenyl)methylamino]-8- methoxy-5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4 -carboxamide)

Example 42: 42A: tert-Butyl-f(2,4-dichloro-5,6,7,8-tetrahydroquinazolin-8-yl) oxy1-dimethyl-silane

To a stirred solution of 2,4-dichloro-5,6,7,8-tetrahydroquinazolin-8-ol (5.00 g, 22.8 mmol) in DMF (114 mL) at room temperature was added tert-butyldimethylsilyl chloride (7.57 g, 50.2 mmol) and imidazole (3.26 g, 47.9 mmol). The resulting mixture was stirred for 60 hours, then diluted with EtOAc (500 mL) and brine:water (1 :1 , 600 mL). The layers were separated, and the organic extracts washed with brine (400 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-10% EtOAc in heptane) to afford the title compound (6.54 g, 82%) as a colourless oil.

42B: tert-Butyl-r(2-chloro-4-methylsulfanyl-5,6,7,8-tetrahydroqui nazolin-8-yl)oxy1-dimethyl- silane To a stirred solution of tert-butyl-[(2,4-dichloro-5,6,7,8-tetrahydroquinazolin-8-yl) oxy]- dimethyl-silane (6.04 g, 17.2 mmol) in THF (29 mL) at -15°C was added sodium thiomethoxide (21% in water, 6.89 mL, 20.7 mmol). The resulting mixture was warmed to room temperature and stirred for 60 hours, The reaction mixture was poured into water (50 mL) before being extracted with EtOAc (3 x 50 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The oil obtained was purified by flash column chromatography (silica gel, 0-10% EtOAc in heptane) to afford the title compound (5.77 g, 92%) as a colourless oil that solidified to a white solid on standing.

42C: 1-r8-rtert-butyl(dimethyl)silyl1oxy-4-methylsulfanyl-5,6,7,8 -tetrahydroquinazolin-2-yl1- 2-methyl-indole-4-carbonitrile

To a degassed stirred solution of tert-Butyl-[(2-chloro-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-8-yl)oxy]-dimethyl-silane (2.64 g, 7.27 mmol), 2-methyl-1 H-indole-4- carbonitrile (1.48 g, 9.45 mmol) and cesium carbonate (4.74 g, 14.5 mmol) in 1 ,4-dioxane (73 mL) was added Pd2(dba)s (998 mg, 1.09 mmol) and XPhos (520 mg, 1.09 mmol). The reaction mixture was then heated to 100°C for 18 hours. The resulting mixture was allowed to cool to room temperature and poured into water (200 mL) before being extracted with EtOAc (2 x 200 mL). The combined organics were washed with brine (100 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0 to 20% EtOAc in heptane) to afford an off white solid. The solid obtained was suspended in heptane. This suspension was filtered and the solid obtained was washed with further heptane and Et20 to afford the title compound (5.67 g, 88%) as a yellow solid.

42D: 1-(8-Hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazolin-2 -yl)-2-methyl-indole-4- carbonitrile

To a stirred solution of 1 -[8-[tert-butyl(dimethyl)silyl]oxy-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carbonitrile (3.36 g, 7.23 mmol) in THF (72 mL) was added tetrabutylammonium fluoride trihydrate (1.0 M in THF, 14.5 mL, 14.5 mmol). The reaction mixture was stirred for 3.5 hours, before being poured into saturated NH4CI solution (50 mL) and being extracted with EtOAc (2 x 50 mL). The combined organics were washed with water (50 mL), dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-50% EtOAc in heptane) to afford the title compound (2.03 g, 80%) as a bronze coloured foam.

42E: 1-(4-methylsulfanyl-8-oxo-6,7-dihydro-5H-quinazolin-2-yl)-2- methyl-indole-4- carbonitrile

To a stirred solution of 1-(8-hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazolin-2 -yl)-2- methyl-indole-4-carbonitrile (1.72 g, 4.91 mmol) in DCM (49 mL) was added Dess-Martin periodinane (3.12 g, 7.36 mmol). The resulting solution was stirred for 18 hours, then diluted with DCM (100 mL) and a solution of NaS2<D4 (10% w/w, 100 mL) before being vigorously stirred for 30 minutes. The biphasic mixture that formed was passed through a phase separator, and the organic extracts were washed with saturated NaHCCh solution (100 mL). The combined organics were concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in heptane, followed by 50% EtOAc in DCM) to afford the title compound (1.46 g, 81%) as a light yellow solid.

42F: 1-r(8S)-8-hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazo lin-2-yl1-2-methyl- indole-4-carbonitrile

A degassed stirred solution of 1-(4-methylsulfanyl-8-oxo-6,7-dihydro-5H-quinazolin-2-yl)- 2-methyl-indole-4-carbonitrile (1.13 g, 3.23 mmol) and RuCI(p-cymene)[(S,S)-Ts-DPEN] (20.5 mg, 0.0300 mmol) in DMF (40 mL) was cooled to 0°C before a solution of HCOOH:TEA complex (5:2) (1.42 g, 16.5 mmol) in DMF (14 mL) was added. The resulting suspension was maintained at 0°C for 10 minutes, warmed to RT and stirred for 4 hours. Saturated NaHCCh solution (100 mL) and water (100 mL) were added, before the resulting mixture was extracted with EtOAc (2 x 100 mL). The combined organics were washed with brine (2 x 75 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-50% EtOAc in heptane) to afford the title compound (956 mg, 84%) as bronze coloured solid. 42G: 1-r(8S)-8-methoxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazo lin-2-yl1-2-methyl- indole-4-carbonitrile

To a stirred solution of 1-[(8S)-8-hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazo lin-2- yl]-2-methyl-indole-4-carbonitrile (1.51 g, 4.10 mmol) in methyl iodide (10.0 mL, 161 mmol) in darkness was added Ag2<D (2.85 g, 12.3 mmol) before being stirred for 48 hours at room temperature. The reaction mixture was diluted with EtOAc (10 mL) and filtered through a plug of celite, washing further with EtOAc (20 mL). The filtrate was concentrated under reduced pressure to afford the title compound (1.46 g, 93%) as a pale yellow solid.

42H: 1-f(8S)-8-methoxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazo lin-2-yl1-2-methyl- indole-4-carboxamide

To a stirred suspension of 1 -[(8S)-8-methoxy-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carbonitrile (69.3 mg, 0.190 mmol) in EtOH (1.6 mL) was added acetaldehyde oxime (64.0 mg, 1.08 mmol), polymer supported PPhs (1.4-2 mmol/g, 20.0 mg), Pd(OAc)2 (6.00 mg, 0.030 mmol) and water (0.4 mL). The resulting mixture was heated to 100°C for 90 minutes. The reaction mixture was then cooled to room temperature and filtered through a plug of celite, washing with further EtOAc (5 mL). The filtrate was poured into water (5 mL) and extracted with EtOAc (10 mL). The combined organics were washed with brine (5 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 25-100% EtOAc in heptane) to afford the title compound (81 mg, 76%) as a yellow solid.

42I: 1-r(8S)-8-methoxy-4-methylsulfonyl-5,6,7,8-tetrahydroquinazo lin-2-yl1-2-methyl- indole-4-carboxamide

To a stirred solution of 1-[(8S)-8-methoxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazo lin- 2-yl]-2-methyl-indole-4-carboxamide (165 mg, 0.400 mmol) in DCM:MeOH (4:1 , 2.5 mL) was added sodium tungstate dihydrate (39 mg, 0.120 mmol), AcOH (110 pL, 1.98 mmol) and H2O2 (30% w/w in water, 0.48 mL, 4.77 mmol). The resulting mixture was heated to 40°C for 24 hours. The reaction mixture was then cooled to room temperature and poured into saturated NaHCCh solution (20 mL) before being extracted with DCM (3 x 20 mL). The combined organics were dried over Na2SC>4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 50-100% EtOAc in heptane) to afford the title compound (130 mg, 75%) as a white solid.

42 J: 1-[(8S)-4- |-8-methoxy-5,6,7,8- in-2-

|-2-methyl-indole-4-carboxamide

To a stirred suspension of 1 -[(8S)-8-methoxy-4-methylsulfonyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide (50 mg, 0.110 mmol) in MeCN (1.0 mL) was added TEA (48 pL, 0.340 mmol) and (3-fluorophenyl)methanamine (78 pL, 0.690 mmol). The reaction mixture was heated to 70°C for 45 hours. The mixture was then cooled to room temperature before being concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 50-100% EtOAc in heptane) followed by preparative HPLC (method B) to afford the title compound (21 mg, 39%) as a white solid.

:ic Route R

(Illustrated with reference to Example 46: 1-[4-(benzylamino)-8-(2,2-difluoroethoxy)- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxa mide)

46A: Methyl 1-r8-(2,2-difluoroethoxy)-4-methylsulfanyl-5,6,7,8-tetrahydr oquinazolin-2-yl1-

2-methyl-indole-4-carboxylate To a stirred suspension of cyanomethyltributylphosphorane (1.23 mL, 4.69 mmol) and methyl 1-(8-hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydroquinazolin-2 -yl)-2-methyl-indole- 4-carboxylate (300 mg, 0.780 mmol) in toluene (3.9 mL) was added 2,2-difluoroethanol (350 pL, 5.48 mmol). The resulting mixture was heated to 110°C for 4 hours, then cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-50% EtOAc in heptane) to afford the title compound (217 mg, 62%) as a yellow sticky gum.

46B: 1-f8-(2,2-Difluoroethoxy)-4-methylsulfanyl-5,6,7,8-tetrahydr oquinazolin-2-yl1-2- methyl-indole-4-carboxamide

To a stirred solution of methyl 1-[8-(2,2-difluoroethoxy)-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxylate (217 mg, 0.480 mmol) in 1 ,4- dioxane:water (1 :1 , 4.8 mL) was added lithium hydroxide monohydrate (122 mg, 2.90 mmol). The resulting mixture was stirred at room temperature for 17 hours before being heated to 40 °C for 7.5 hours. The reaction mixture was cooled to room temperature before being concentrated under reduced pressure. The resulting residue was added to water (10 mL) before 1 M HCI was added to bring the solution to pH 3. The mixture was extracted with EtOAc (2 x 20 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The obtained solid was dissolved in DMF (4.2 mL), cooled to 0°C and treated with HBTU (241 mg, 0.640 mmol), NH4CI (114 mg, 2.12 mmol) and TEA (30 pL, 2.12 mmol) before being warmed to room temperature and stirred for 65 hours. The reaction was partitioned between EtOAc:DCM (1 :1 , 20 mL) and brine (20 mL). The layers were separated, and the organic extracts washed with water (2 x 20 mL) and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-5% MeOH in DCM) to afford the title compound (140 mg, 76%) as a white solid.

46C: 1-r8-(2,2-Difluoroethoxy)-4-methylsulfonyl-5,6,7,8-tetrahydr oquinazolin-2-yl1-2- methyl-indole-4-carboxamide

To a stirred solution of 1-[8-(2,2-difluoroethoxy)-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide (140 mg, 0.320 mmol) in DCM:MeOH (4:1 , 3.2 mL) was added sodium tungstate dihydrate (32 mg, 0.100 mmol), AcOH (93 pL, 1.62 mmol) and H2O2 (30% w/w in water, 264 pL, 2.58 mmol). The resulting mixture was heated to 45°C for 23 hours and then cooled to room temperature before being diluted with saturated NaHCCh solution (20 mL) and extracted with DCM:MeOH (4:1 , 20 mL). The combined organics were then washed with brine (20 mL), passed through a phase separator and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in heptane) to afford the title compound (110 mg, 67%) as a bright yellow solid. in-2-yll-2-

To a stirred solution of 1-[8-(2,2-difluoroethoxy)-4-methylsulfonyl-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxamide (55 mg, 0.120 mmol) in MeCN (0.8 mL) was added benzylamine (71 pL, 0.650 mmol) and TEA (50 pL, 0.360 mmol). The reaction mixture was heated to 70°C for 20 hours and then concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 20-100% EtOAc in heptane) followed by preparative HPLC (method B) to afford the title compound (18 mg, 30%) as a yellow solid.

:ic Route S

(Illustrated with reference to Example 45: 1-[(8S)-4-(benzylamino)-8-(2-hydroxyethoxy)- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxa mide)

Example 45

45A: 1-r(8S)-8-f2-rtert-butyl(dimethyl)silyl1oxyethoxy1-4-methyls ulfanyl-5,6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carbonitrile

A stirred suspension of 1-[(8S)-8-hydroxy-4-methylsulfanyl-5,6,7,8-tetrahydro quinazolin- 2-yl]-2-methyl-indole-4-carbonitrile (500 mg, 1.43 mmol), tert-butyl-(2-iodoethoxy)- dimethyl-silane (2.45 g, 8.56 mmol) and Ag2<D (992 mg, 4.28 mmol) was heated to 110°C in the dark for 40 hours. The resulting mixture was diluted with EtOAc (10 mL) and filtered through celite, washing with further EtOAc (30 mL). The filtrate was concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-50% EtOAc in heptane) to afford the title compound as a light brown oil, which solidified on standing to a cream waxy solid (643 mg).

45B: 1-r(8S)-8-[2-rtert-Butyl(dimethyl)silyl1oxyethoxy1-4-methyls ulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carboxamide

A stirred suspension of 1-[(8S)-8-[2-[tert-butyl(dimethyl)silyl]oxyethoxy]-4-methyl sulfanyl- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboni trile (600 mg, 1.18 mmol), acetaldehyde oxime (279 mg, 4.72 mmol), Pd(0Ac)2 (39.7 mg, 0.180 mmol) and PS- triphenylphosphine (1.4-2 mmol/g, 120 mg) in EtOH:water (4:1 , 7.2 mL) was heated to 85°C for 2.5 hours. The resulting suspension was cooled to room temperature and partitioned between MeOH:DCM (1 :9, 20 mL) and water (20 mL). The biphasic mixture was passed through a phase separator and the combined organics were concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in heptane) to afford the title compound as an orange solid (320 mg, 52% over two steps).

45C. 1-r(8S)-8-r2-rtert-Butyl(dimethyl)silyl1oxyethoxy1-4-methyls ulfonyl-5, 6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carboxamide

To a stirred solution of 1-[(8S)-8-[2-[tert-butyl(dimethyl)silyl]oxyethoxy]-4-methyls ulfanyl- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxa mide (320 mg, 0.610 mmol) in DCM:MeOH (4:1 , 6.1 mL) was added sodium tungstate dihydrate (120.2 mg, 0.360 mmol), AcOH (348 pL, 3.04 mmol) and H2O2 (30% w/w in H2O, 992 pL, 9.72 mmol). The resulting mixture was heated to 45 °C for 40 hours. This mixture was cooled to room temperature, diluted with DCM (30 mL) and washed with saturated NaHCCh solution (2 x 20 mL). The aqueous extracts were extracted with further DCM (10 mL), and the combined organics were washed with brine (10 mL), dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 25-100% EtOAc in heptane) to afford the title compound (189 mg, 50%) as a yellow solid.

45D. 1-r(8S)-4-(benzylamino)-8-r2-rtert-butyl(dimethyl)silyl1oxye thoxy1-5, 6,7,8- tetrahydroquinazolin-2-yl1-2-methyl-indole-4-carboxamide

To a stirred solution of 1-[(8S)-8-[2-[tert-butyl(dimethyl)silyl]oxyethoxy]-4-methyl sulfonyl- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxa mide (189 mg, 0.300 mmol) in MeCN (1.5 mL) was added TEA (127 pL, 0.910 mmol) and phenylmethanamine (67 pL, 0.610 mmol). The resulting mixture was heated to 50°C for 70 hours, then cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 20-100% EtOAc in heptane) to afford the title compound (163 mg, 82%) as a pale yellow gum. 45E. 1-r(8S)-4-(benzylamino)-8-(2-hydroxyethoxy)-5,6,7,8-tetrahyd ro quinazolin-2-yl1-2- methyl-indole-4-carboxamide)

To a stirred solution of 1-[(8S)-4-(benzylamino)-8-[2-[tert-butyl(dimethyl)silyl] oxyethoxy]- 5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carboxa mide (311 mg, 0.478 mmol) in THF (2.4 mL) at 0°C was added TBAF.3H ₂ O (1.0 M solution in THF, 1.43 mL, 1.43 mmol) dropwise. The resulting mixture was maintained at 0°C for 10 minutes, warmed to room temperature and stirred for 4 hours, then partitioned between EtOAcJPA (9:1 , 20 mL) and water (10 mL). The layers were separated, and the combined organics were washed with brine (5 mL), dried over Na ₂ SO4 and concentrated under reduced pressure.

The resulting residue was purified by flash column chromatography (silica gel, 0-10% MeOH in DCM). The resulting partitally purified gummy solid was further purified by preparative HPLC (Method B) to afford the title compound (57 mg, 25%) as a white solid. Synthetic Route T

(Illustrated with reference to Example 24: 1-[(8S)-8-(azetidin-3-ylmethoxy)-4-

(benzylamino)-5,6,7,8-tetrahydroquinazolin-2-yl]-2-methyl -indole-4-carboxamide) EXAMPLE 24

24A: tert-Butyl 3-H(8S)-2-(4-cyano-2-methyl-indol-1-yl)-4-methylsulfanyl-5,6 ,7,8- tetrahydroquinazolin-8-yl1oxymethyl1azetidine-1 -carboxylate

A stirred suspension of 2-methyl-1-[(8S)-8-hydroxy-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-2-yl]indole-4-carbonitrile (150 mg, 0.430 mmol), tert-butyl 3- (iodomethyl)azetidine-l -carboxylate (763 mg, 2.57 mmol) and Ag2<D (298 mg, 1.28 mmol) was heated to 110°C in darkness for 65 hours. The reaction mixture was cooled to room temperature before being filtered through a plug of celite and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in heptane) to afford the title compound (164 mg, 88%) as a light-yellow gum, which was used directly without further purification.

24B: tert-Butyl 3-ff(8S)-2-(4-carbamoyl-2-methyl-indol-1 -yl)-4-methylsulfanyl-5, 6,7,8- tetrahydroquinazolin-8-yl1oxymethyl1azetidine-1 -carboxylate

To a stirred solution of tert-butyl 3-[[(8S)-2-(4-cyano-2-methyl-indol-1-yl)-4-methylsulfanyl- 5,6,7,8-tetrahydroquinazolin-8-yl]oxymethyl]azetidine-1-carb oxylate (163 mg, 0.17 mmol) in EtOH (1.4 mL) and water (0.4 mL) was added PS-PPhs (1.4-2 mmol/g, 32 mg), acetaldehyde oxime (84 mg, 1.42 mmol) and Pd(OAc)2 (12 mg, 0.05 mmol). The resulting suspension was heated to 100°C for 3 hours. The resulting suspension was cooled to room temperature, diluted with EtOAc (15 mL) and filtered through celite. The filtrate was concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 20-100% EtOAc in heptane) to afford the title compound (90 mg, 53%) as a light-yellow gum.

24C: tert-Butyl 3-ff(8S)-2-(4-carbamoyl-2-methyl-indol-1 -yl)-4-methylsulfonyl-5, 6,7,8- tetrahydroquinazolin-8-yl1oxymethyl1azetidine-1 -carboxylate

To a stirred solution of tert-butyl 3-[[(8S)-2-(4-carbamoyl-2-methyl-indol-1-yl)-4-methyl sulfanyl-5, 6, 7, 8-tetrahydroquinazolin-8-yl]oxymethyl]azetidine-1 -carboxylate (90.0 mg, 0.0993 mmol) in DCM:MeOH (2:1 , 3.0 mL) was added sodium tungstate dihydrate (20.0 mg, 0.0600 mmol), AcOH (46 pL, 0.795 mmol) and H2O2 (30% w/w in H2O, 202 pL, 1.99 mmol). The resulting mixture was heated to 40°C for 24 hours. The reaction mixture was cooled to room temperature, diluted with DCM (20 mL) and washed with saturated NaHCCh solution (10 mL). The aqueous extracts were extracted with further DCM (10 mL), and the combined organics were washed with brine (10 mL), dried over Na2SO4 and concentrated under reduced pressure to afford the title compound (95 mg) as a brown film, which was used directly without further purification.

27D: tert-Butyl 3-H(8S)-4-(benzylamino)-2-(4-carbamoyl-2-methyl-indol-1 -yl)-5, 6,7,8- tetrahvdroquinazolin-8-yl1oxymethyl1azetidine-1 -carboxylate

To a stirred solution of tert-butyl 3-[[(8S)-2-(4-carbamoyl-2-methyl-indol-1-yl)-4-methyl sulfonyl-5, 6, 7, 8-tetrahydroquinazolin-8-yl]oxymethyl]azetidine-1 -carboxylate (95.0 mg, 0.070 mmol) in MeCN (0.4 mL) was added TEA (61 pL, 0.441 mmol) and phenylmethanamine (32 pL, 0.294 mmol). The resulting mixture was heated to 50°C for 70 hours. The reaction was cooled to room temperature and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 30-100% EtOAc in heptane) to afford the title compound (40 mg, 39% over 2 steps) as a yellow film.

24E: 1-r(8S)-8-(Azetidin-3-ylmethoxy)-4-(benzylamino)-5,6,7,8-tet rahydroquinazolin-2-yl1- 2-methyl-indole-4-carboxamide

To a stiurred solution of tert-butyl 3-[[(8S)-4-(benzylamino)-2-(4-carbamoyl-2-methyl- indol-1 -yl)-5, 6, 7, 8-tetrahydroquinazolin-8-yl]oxymethyl]azetidine-1 -carboxylate (41 .0 mg, 0.050 mmol) in DCM (2.0 mL) was added TFA (500 pL, 6.53 mmol). The resulting mixture was stirred at room temperature for 1 hour, then concentrated under reduced pressure. The resulting residue was dissolved in DCM (5 mL) and washed with saturated NaHCCh solution (5 mL). The aqueous was extracted with EtOAcJPA (9:1 , 2 x 10 mL) and the combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC (Method B). The resulting solid was taken into saturated NaHCCh solution (2 mL) before being extracted with DCM (3 x 2.0 mL). The combined organics were dried using a phase separator and concentrated under reduced pressure to afford the title compound (2.6 mg, 7%) as a white solid.

Synthetic Route U (Illustrated with reference to Example 50: 1-[(8S)-4-(Benzylamino)-8-(2-hydroxy-2-methyl- propoxy)-5,6,7,8-tetrahydro quinazolin-2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE 50

50A: 1-f(8S)-4-(benzylamino)-8-(2-hydroxy-2-methyl-propoxy)-5,6,7 ,8-tetrahydro quinazolin-2-yl1-2-methyl-indole-4-carbonitrile

To a stirred solution of methyl 2-[[(8S)-4-(benzylamino)-2-(4-cyano-2-methyl-indol-1-yl)- 5,6,7,8-tetrahydroquinazolin-8-yl]oxy]acetate (71.0 mg, 0.0800 mmol) in THF (1.0 mL), at 0 °C was added methyl magnesium bromide (3.0 M in diethyl ether, 270 pL, 0.802 mmol) dropwise. The resulting mixture was maintained at 0 °C for 4 hours. The crude mixture was quenched through dropwise addition of water and saturated NH4CI solution (10 mL). The resulting mixture was extracted with EtOAc (3 x 10 mL). The combined organics were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 20-100% EtOAc in heptane). Further purification by preparative HPLC (Method B) afforded the title compound (8.0 mg, 18%) as a white solid. 50B. 1-f(8S)-4-(Benzylamino)-8-(2-hydroxy-2-methyl-propoxy)-5,6,7 ,8-tetrahydro quinazolin-2-yl1-2-methyl-indole-4-carboxamide

To a stirred solution of 1-[(8S)-4-(benzylamino)-8-(2-hydroxy-2-methyl-propoxy)-5, 6,7,8- tetrahydroquinazolin-2-yl]-2-methyl-indole-4-carbonitrile (8.0 mg, 0.0100 mmol) in EtOH:water (5:1 , 600 L) was added acetaldehyde oxime (4.0 mg, 0.0677 mmol), Pd(OAc)2 (1.0 mg, 0.0045 mmol) and PS-PPhs (1.4-2 mmol/g, 2.0 mg). The resulting mixture was heated at 80°C for 90 minutes, then cooled to room temperature and diluted with DCM (5 mL). The resulting biphasic suspension was passed through a phase separator and concentrated under reduced pressure. The resulting residue was purified by preparative HPLC (Method B) to afford the title compound (3.5 mg, 47%) as a white solid.

Synthetic Route F

(Illustrated with reference to Example A: 1-[4-(benzylamino)-8-methoxy-quinazolin-2-yl]-2- methyl-indole-4-carboxamide)

EXAMPLE A

A-A: N-benzyl-2-chloro-8-methoxy-quinazolin-4-amine

Benzylamine (0.37 g, 3.49 mmol) and TEA (1.06 g, 10.5 mmol) were added to a stirred solution of 2,4-dichloro-8-methoxyquinazoline (0.8 g, 3.49 mmol) in MeCN (5 mL) under a nitrogen atmosphere. The mixture was stirred at room temperature for 2 hours then the resulting precipitate collected by filtration. The collected solid was washed with MeCN (20 mL) and dried under reduced pressure to give the title compound (0.72 g, 68%) as an off-white solid.

A-B: 1-f4-(benzylamino)-8-methoxy-quinazolin-2-yl1-2-methyl-indol e-4-carbonitrile A stirred mixture of N-benzyl-2-chloro-8-methoxy-quinazolin-4-amine (0.50 g, 1.67 mmol), 2-methyl-1 H-indole-4-carbonitrile (0.37 g, 2.34 mmol) and CS2CO3 (1.1 g, 3.34 mmol) in 1 ,4-dioxane (10 mL) was degassed with nitrogen for 15 mins. Pd2(dba)s (230 mg, 0.25 mmol) and XPhos (119 mg, 0.25 mmol) were added to the reaction mixture which was heated to 100°C for 16 hours under a nitrogen atmosphere. The cooled mixture was partitioned between water (30 mL) and EtOAc (30 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (0.58 g, 76%) as a pale yellow solid.

A-C: 1-r4-(benzylamino)-8-methoxy-quinazolin-2-yl1-2-methyl-indol e-4-carboxamide

A stirred mixture of 1-[4-(benzylamino)-8-methoxy-quinazolin-2-yl]-2-methyl-indol e-4- carbonitrile (0.58 g, 1.38 mmol), acetaldehyde oxime (163 mg, 2.76 mmol), Pd(OAc)2 (31 mg, 0.48 mmol) and polystyrene supported PPhs (127 mg, 0.48 mmol) in EtOH (15 mL) and water (1 mL) was heated to 100°C for 16 hours then cooled to room temperature and the solvents removed under reduced pressure. The resulting residue was partitioned between EtOAc (30 mL) and water (30 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (75 mg, 12%) as a pale yellow solid.

Synthetic Route G

(Illustrated with reference to Example B: 1-[4-(benzylamino)-5,6,7,8-tetrahydroquinazolin- 2-yl]-2-methyl-indole-4-carboxamide)

EXAMPLE B

B-A: N-benzyl-2-chloro-5,6,7,8-tetrahydroquinazolin-4-amine Benzylamine (0.52 g, 4.90 mmol) and TEA (1 g, 9.8 mmol) were added to a stirred solution of 2,4-dichloro-5,6,7,8-tetrahydroquinazoline (1.0 g, 4.90 mmol) in MeCN (10 mL) under a nitrogen atmosphere. The stirred reaction mixture was heated to 40°C for 16 hours then the solvents were removed under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (0.90 g, 67%) as a white solid.

B-B: 1-r4-(benzylamino)-5,6,7,8-tetrahydroquinazolin-2-yl1-2-meth yl-indole-4-carbonitrile

A stirred mixture of N-benzyl-2-chloro-5,6,7,8-tetrahydroquinazolin-4-amine (0.55 g, 2.01 mmol), 2-methyl-1 H-indole-4-carbonitrile (0.35 g, 2.21 mmol) and CS2CO3 (1.31 g, 4.02 mmol) in 1 ,4-dioxane (10 mL) was degassed with nitrogen for 15 mins. Pd2(dba)s (0.28 g, 0.30 mmol) and XPhos (143 mg, 0.30 mmol) were added to the reaction mixture which was heated to 100°C for 16 hours under a nitrogen atmosphere. The cooled mixture was partitioned between water (30 mL) and EtOAc (30 mL) and the separated aqueous phase extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (0.68 g, 81%) as a pale yellow solid.

B-C: 1-r4-(benzylamino)-5,6,7,8-tetrahydroquinazolin-2-yl1-2-meth yl-indole-4-carboxamide

A stirred mixture of 1-[4-(benzylamino)-5,6,7,8-tetrahydroquinazolin-2-yl]-2-meth yl-indole- 4-carbonitrile (0.17 g, 0.43 mmol), acetaldehyde oxime (51 mg, 0.86 mmol), Pd(OAc)2 (10 mg, 0.04 mmol) and polystyrene supported PPhs (23 mg, 0.08 mmol) in EtOH (9 mL) and water (1 mL) was heated to 100°C for 10 hours then cooled to room temperature and the solvents removed under reduced pressure. The resulting residue was partitioned between EtOAc (20 mL) and water (20 mL) and the separated aqueous phase was extracted with EtOAc (3 x 30 mL). The combined organic extracts were dried over Na2SO4 and concentrated under reduced pressure. The resulting residue was purified by flash column chromatography (silica gel, 0-100% EtOAc in Hexane) to give the title compound (52 mg, 28%) as a pale yellow solid.

BIOLOGICAL ACTIVITY

EXAMPLE 54 - VCP ATPase Activity The compounds of the invention were tested for activity against VCP using the materials and protocols set out below.

ATPase assay was assembled in a low bind, low volume, white 384 well plates (Corning 3824) and read using a BMG Clariostar plate reader.

Small molecule inhibitors were diluted in 100% DMSO, followed by intermediate dilution into assay buffer (20 mM HEPES pH 7.4, 250 mM KCI, 1 mM MgCh, 5% glycerol, 1 mM DTT) with DMSO adjusted to 5% final concentration. From this intermediate dilution, 4 pL were added to respective wells in assay plate, followed by 3 pL of VCP enzyme solution at 666 nM in assay buffer. Assay was incubated for 15min at room temperature, followed by addition of 3 pL of ATP at 333 pM to wells containing VCP/inhibitors. At this point, ATP/ADP standard curve was prepared in assay buffer, with indicated concentrations below, and 10 pL of the mixture was added to control wells in assay plate.

Reaction was incubated at 37°C for 45 minutes followed by equilibrating at room temperature for 15 minutes, at which point 10 pL of ADP-Glo reagent (Promega) was added to quench the reactions. Assay was incubated for 60 minutes, after which 20 pL of ADP-Glo detection reagent (Promega) was added to wells. Luminescence was then measured after incubating for 20 minutes at room temperature, using a Clariostar plate reader (BMG).

Data was analysed using GraphPad Prism and normalized dose responses plotted, by using the buffer only sample with VCP as 100% activity, and ATP alone as the 0% activity.

ATPase information:

VCP - Uniprot ID P55072

His tagged full length VCP protein (2-806) was expressed in E. coli BL21 (DE3) and purified following procedure in Chou et al, 2011 (PMID: 21383145). VCP was purified using Ni-NTA affinity purification and gel size exclusion chromatography to a final purity greater than 90%. Purified VCP was subsequently concentrated and protein aliquots stored in VCP storage buffer (20 mM HEPES pH 7.4, 250 mM KCI, 1 mM MgCI ₂ , 5% glycerol, 1 mM DTT).

Final concentration in assay = 200 nM From the results obtained by following the above protocol, the IC50 values against VCP ATPase activity of each of the compounds of Examples 1 to 53, A, B and C were determined.

The compounds of Examples 1 to 53, A, B and C all had IC50 values of less than 1 pM. Of these, the compounds of Examples 1 , 2, 3, 4, 5, 6, 7, 8, 11 , 17, 18, 19, 20, 21 , 23, 24, 26,

28, 29, 30, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 44, 45, 46, 47, 48, 49, 50, 51 , 52, A,

B and C all had IC50 values of less than 0.1 pM. The compounds of Examples 1 , 2, 3, 6, 7, 8, 11 , 17, 18, 19, 21 , 26, 29, 30, 33, 37, 38, 40, 41 , 42, 44, 48, 49 and 52 all had IC50 values in the range from 0.02 to 0.1 pM, and the compounds of Examples 4, 5, 20, 23, 24, 28, 32, 34, 35, 36, 39, 45, 46, 47, 50, 51 , , A, B and C all had IC50 values in the range from 0.001 to 0.02 pM.

The compounds of each of Examples 1 to 53, A, B and C have the IC50 values shown in Table 3. EXAMPLE 55 - PDE6 Assay

PDE6 phosphodiesterase enzymatic activity assay was performed by Eurofins Panlabs Discovery Services (#156100), following the procedure first described in Baehr et al (J Biol Chem, 1979) and Gillespie and Beavo (Mol Pharma, 1989). Alternatively:

Bovine retinal rod phosphodiesterase PDE6 activated by trypsin was used. Test compound and/or vehicle was preincubated with 0.2 pg/ml enzyme in Tris-HCI buffer pH 7.5 for 15 minutes at 25°C. The reaction was initiated by addition of 100 pM cGMP and 0.03 pM [3H]cGMP followed by 20 minute incubation period and terminated by heating to 100°C. The resulting [3H] cGMP was converted to [3H]Guanosine by addition of snake venom nucleotidase and separated by AG1-X2 resin. An aliquot was removed and counted to determine the amount of [3H]Guanosine formed. IC50 of compounds were determined alongside the control PDE6 inhibitor Zaprinast.

The compounds of a selection of Examples 1 to 48, A and B have the IC50 values shown in Table 4.

EXAMPLE 56 - FRAME-SHIFT REPORTER ASSAY AND CYTOTOXICITY In order to read out mutagenesis in mammalian cells, a Frameshift Reporter (FSR) assay was used. The assay specifically focuses on frameshifts because these are the ones that most abundantly generate neoantigens (see, for example, Turajlic et al. Lancet Oncol. 2017 (18; 1009-21)). The assay based on the activity of the NanoLuciferase (NanoLuc®, Promega) reporter enzyme was developed (the “CA( _n )-NanoLuc assay”). Copies of the CA dinucleotide repeat (referred to as “CA(n)-) were cloned upstream of the NanoLuc coding sequence. Typically 18 copies of the CA dinucleotide repeat (CA(18>) were used. Alternatively, the assay used 20 copies (CA(2o>). This CA( _n > tract renders the NanoLuc coding sequence out of frame, and therefore there is no reporter enzyme expression and reporter activity is low. The CA( _n > tract is, however, a sequence which is subject to frequent DNA replication errors, and is reliant on the MM R pathway to repair any post- replicative DNA mismatches. If, a small molecule inhibits VCP activity frameshift mutations may occur, and some cells in a population will now express a functional NanoLuc protein. This is depicted in Figure 1.

Mutagenic insult can therefore be reported as an increase in NanoLuc® activity when a plasmid containing a CA repeat, such as the CA(is) NanoLuc construct, is transfected into cells. Since NanoLuc® is a highly processive enzyme with a large and linear dynamic range, it was predicted that only a small number of frameshift mutations would be required to generate a positive signal, making for a sensitive assay with a large signal to noise ratio.

Detailed method as follows:

Preparation of cell culture

HEK293FT cells were cultured in DMEM supplemented with 10% Fetal Bovine Serum (FBS), 1 % Penicillin and Streptomycin (PenStrep) and L-Glutamine at 37°C (5% CO2, > 90% humidity).

In brief, cells were washed in 10 mL PBS and trypsinised with 3 mL TrypLE Express. After cell detachment, TrypLE Express was blocked with 7 ml fresh media and cell suspension moved to a 50 mL Falcon tube. Cells were centrifuged at 300 x g for 5 mins, supernatant aspirated and cells resuspended in 10 mL cell specific growth medium. Cell density was calculated using a ThermoScientific Countess II automated cell counter: 10 pL cell culture added to 10 pL 0.4% Trypan Blue. Cells were diluted in a 50 mL falcon to 100000 cells/ml in DMEM and distributed in 6 well plates. Cells incubated overnight at 37°C 5% CO2 in a humidified atmosphere to allow adherence.

Four hours prior to transfection media was aspirated and 2 mL fresh DMEM added to each well.

Plasmid stock 003: pcDNA3.1 hygro CA18-nanoluc (1.1 pg/mL) and JetPrime Buffer (Polypus Transfection: Cat# 114-75) were equilibrated to room temp. 24 pg (21 pL stock) was added to 9.6 mL JetPrime Buffer in a 15 mL Falcon tube and mixed by vortex for 10 secs. 192 pL JetPrime transfection reagent was added and mixed by vortex for 10 secs. Tube was centrifuged (54 x g, 30 secs) and incubated for 10 minutes at room temperature. 200 pL of JetPrime/DNA complex was added dropwise to each well of 6 well plates containing cultured HEK293FT cells. Plates were incubated for 4 hours before aspiration of media and replacement with 2 mL/well fresh DMEM followed by overnight incubation.

One day before the compound treatment, media was aspirated, and wells were washed with 2 ml PBS. 500 pL TrypLIE Express was added to each well of 6 well plates and incubated for 5 minutes before cells were dissociated, blocked in 2 mL DMEM and all cells pooled in 50 mL Falcon tubes. Cells were pelleted by centrifugation at 1000 x g for 5 mins and washed in 20 mL PBS followed by centrifugation (1000 x g, 5 mins). PBS was aspirated and cells were resuspended in 20 mL DMEM. Cell density was calculated as described above and cells were resuspended to 1.0 x 10 ⁵ cells/mL in 180 mL DMEM. 50 pL was added to the inner 60 wells of a white, clear bottom 96 well plate. In 6 wells of column A, 100 pL of media only was added, and to the rest of the wells in the plate, 200 pL of PBS. One plate per compound was prepared and incubated overnight.

VCP inhibitors were made to 4 mM in DMSO, and 9 x two-fold serial dilutions were made in DMSO, keeping the DMSO concentration constant. A 2 x solution of VCP inhibitor was made up in DMEM, with a top concentration of 20 pM, keeping the DMSO constant at 0.5%. 50 pL of each VCP inhibitor was added to all wells of 96 well plates (6 replicates per concentration). Plates were incubated for 72 hours at 37°C 5% CO2 in a humidified atmosphere.

To 3 wells per concentration a 1:1 ratio of CellTiter Gio (Promega) was added. To remaining wells, an equal volume of NanoGio (Promega) reagent was added, and plates were placed on a plate shaker for 2 minutes. Ten minutes later luminescence was read using CLARIOStar plate reader. Percentage viability was calculated for all conditions in comparison to a DMSO treated control. Percentage viability was calculated for all conditions in comparison to an DMSO treated control and nanoluciferase reading were normalised to viability.

On occasions where FSR was not measured, non-transfected cells were plated and treated as detailed above.

The compounds of a selection of Examples 1 to 53, A and B have the GI50 and FSR response values shown in Table 5.

EXAMPLE 57 - KRAS Cell Lines

Studies were carried out to test the sensitising effect of compounds of the invention on the ability to inhibit the growth of KRAS wild type (WT) and KRAS G13D mutant DLD1 cancer cells in 3D tissue culture.

The following protocol was used:

DLD-1 KRAS +/- and DLD-1 G13D/- were cultured in RPMI supplemented with 10% Fetal Bovine Serum (FBS), at 37°C (5% CO2, > 90% humidity).

In brief, cells were washed in 10 mL PBS and trypsinised with 3 mL TrypLE Express. After cell detachment, TrypLE Express was blocked with 7 ml fresh media and cell culture was moved to a 50 mL Falcon tube. Cells were centrifuged at 300 x g for 5 mins and resuspended in 10 mL RPMI growth medium. Cell density was calculated using a ThermoScientific Countess II automated cell counter: 10 pL cell culture added to 10 pL 0.4% Trypan Blue.

Cells were diluted in a 50 mL falcon to 520000 cells/mL in appropriate media and 50 pL of cell suspension was added to the inner 60 wells (3 rows contained DLD-1 KRAS +/- WT and the remaining 3 rows contained the DLD-1 KRAS G13D/-) of a low adherence round bottom plate, to stimulate the formation of spheres. In 6 wells of column A, 100 pL of media only was added, and to the rest of the wells in the plate, 200 pL of PBS. One plate per test compound was prepared and incubated for 3 days at 37°C 5% CO2 in a humidified atmosphere to allow the formation of spheres.

VCP inhibitors were made to either 0.8, 2 or 8 mM in DMSO and 9 x two-fold serial dilutions were made in DMSO, keeping the DMSO concentration constant. A 2 X solution of VCP inhibitor was made up in 400 pL of RPMI, with the top concentrations of 8, 20 or 80 pM, keeping the DMSO content at 0.5%. 50 pL of each VCP inhibitor was added to all wells of one 96 well plate (3 replicates per concentration for each cell line). Plates were incubated for 72 hours at 37°C 5% CO2 in a humidified atmosphere. CellTiter Glo-3D (Promega) was added at a ratio of 1 :1 to all plates and luminescence read using CLARIOStar plate reader. Percentage viability was calculated for all conditions in comparison to a DMSO treated control.

The compounds of a selection of Examples 1 to 23 have the GI50 values shown in Table 6.

EXAMPLE 58 - UNFOLDED PROTEIN RESPONSE VCP inhibition causes accumulation of misfolded proteins, a cellular insult that activates the unfolded protein response (UPR) to either resolve the insult or activate cell death pathways (Szcz^sniak PP et al (2022) VCP inhibition induces an unfolded protein response and apoptosis in human acute myeloid leukemia cells; Doultsinos D, et al. (2017) Control of the Unfolded Protein Response in Health and Disease).

A direct consequence of VCP or proteosome inhibition is the accumulation of K-48 polyubiquitinated protein chains and immediate dissociation of BiP from its sensors - PERK, IRE-1 , and ATF6 - to bind misfolded proteins, triggering prosurvival or, in the case of overwhelming stress, prodeath mechanisms. Some of the most common markers of UPR activation are the phosphorylation of the eukaryotic translation initiation factor 2a (elF2a), which promotes the translation and transcription of CHOP; the nonconventional splicing of XBP1 mRNA, and the release of the ATF6 cytosolic domain from the golgi, which increases the translation and transcription of endoplasmic reticulum (ER) chaperones such as BiP, amongst other proteins. Accumulation of misfolded proteins is also marked by a characteristic translocation of the chaperone calreticulin (CRT) from the ER lumen to the cell membrane surface, a signal capable of eliciting activation of the immune system.

The concept of immunogenic cell death (ICD) represents a unique cell response pattern that comprises the induction of signal cascades aiming to activate both the innate and adaptive immune response. Therefore, evidence of UPR activation gives an indication of the cellular activity upon VCP inhibition, which could in principle lead to the activation of an adaptive immune response through ICD.

The following protocol was used to assess the UPR response:

HCT116 cells were cultured in RPMI supplemented with 10% FBS and MC38 cells were cultured in DM EM supplemented with 10% FBS, both at 37°C (5% CO2, > 90% humidity)

Cells were harvested as previously described and diluted in 50 mL falcons to 750000 cells/mL in appropriate media and 1 mL of cell suspension was seeded into 6 well plates, one plate per compound for each cell line, and incubated overnight.

Examples 2 and 4, alongside Bortezomib - a proteossome inhibitor - as well as Thapsigargin - a non-competitive inhibitor of the sarco/endoplasmic reticulum Ca ATPase (SERCA) - were prepared in DMSO and directly dissolved into the media of previously plated cells, keeping the DMSO constant at 0.5% at final concentrations of 0.3, 1 , 3 and 10 pM. A DMSO control was also included.

After 24 hours of treatment, all cells from the wells were collected with TrypLE Express. Cell pellets were washed with cold PBS and cells lysed in RIPA buffer supplemented with protease and phosphatase inhibitors. Supernatant was quantified with the Pierce BCA protein assay kit.

25 pg of samples were loaded in NuPAGE™ 4 to 12%, Bis-Tris, 1.0 mm, Midi Protein Gels and transferred to Trans-Blot Turbo Midi 0.2 pm PVDF membranes. Membranes were blocked with 5% Bovine Serum Albumin (BSA) and incubated with the primary antibodies listed in Table 7 overnight at 4°C. Membranes were then washed 3 times for 5 minutes with TBST buffer, incubated with secondary antibodies for 1 hour at RT and washed again as previously. Membranes were incubated with SuperSignal West Pico PLUS and signal was acquired with Licor using the chemiluminescent and RGB channels for specific times.

Western blot Figures 2 and 3 show an increase in Bip and CHOP protein levels and increase in K48-linked poly ubiquitinated protein products, indicating unfolded protein responses upon inhibition of VCP. This was observed both in human HCT116 cells and mouse MC38 mouse cell line.

EXAMPLE 59 - CALRETICULIN EXPOSURE

As described previously, cellular insults capable of activating the UPR can initiate immunogeninc cell death. In response to inducers of ICD, malignant cells expose Calreticulin (CRT) and other ER chaperones on their surface, secrete ATP, initiate a cell-intrinsic type I interferon (IFN) response culminating in the production of CXC-chemokine ligand 10 (CXCL10), and release of high-mobility group box 1 (HMGB1) and annexin A1 (ANXA1). (Galluzzi, L., Buque, A., Kepp, O. et al. Immunogenic cell death in cancer and infectious disease. Nat Rev Immunol 17, 97-111 , 2017; Obeid, M. et al. Calreticulin exposure dictates the immunogenicity of cancer cell death. Nat Med 13, 54-61 , 2007). Compounds that activate ICD might offer a distinct advantage in cancer treatment as they increase antigenicity and adjuvanticity.

The following protocol was used to assess the exposure of CRT in the cell surface membrane:

HCT 116 and MC38 cells were cultured and harvested as previously. Cells were diluted in a 50 mL falcon to 750000 cells/mL in appropriate media and 1 mL of cell suspension was added into all wells of a 6 well plates. One plate per compound for each cell line was seeded and incubated overnight.

VCP inhibitors, alongside Bortezomib - a proteossome inhibitor - as well as Cisplatin - a DNA crosslinker as negative control for ICD - were directly dissolved into the media of previously plated cells, keeping the DMSO constant at 0.5% at final concentrations of 0.5, 1 , 5 and 10 pM. A DMSO control was also included. Experimental samples were only collected where >50% of cells were attached to the plate.

After 24 hours, all cells from the wells were collected with cell dissociation buffer (Gibco), centrifuged at 600 x g for 5 minutes, washed with 2mL of cold PBS and diluted to 1000000 cells/mL in falcon tubes. 100 pL of cell suspension of each treatment and control were distributed in a round-bottom polypropylene 96 well plate and centrifuged at 600 x g for 5 minutes. Cells were incubated with 25 pL of cold blocking buffer (0.1% BSA in PBS) with the Alexa Fluor® 647 Anti-Calreticulin antibody [EPR3924] (Abeam) at a final concentration of 0.04 mg/mL for 30 minutes at 4°C. Cells were then washed with 200 pL of cold blocking buffer three times and resuspended in 100 pL of PBS. Cells were gated for single cells and the mean fluorescence values extracted from that population.

The gating strategy is shown in Figures 4 and 5 and CRT levels are shown in Figures 6 and 7.

EXAMPLE 60 - ATP RELEASE As described previously, ATP secretion is a characteristic of ICD initiation and it can be monitored over time with the RealTime-Glo™ Extracellular ATP Assay kit from Promega, following manufacture’s guidelines.

The following protocol was used to measure extracellular ATP:

HCT 116 cells were cultured and harvested as previously, and cells were diluted in a 50 mL falcon 325000 cells/mL in cell specific media. 50 pL was added to the inner 60 wells of a white, clear bottom 96 well plate. 200 pL of PBS were added to the remaining wells.

The compound of Example 2, alongside Bortezomib, were made to 8 mM, as well as 4 x 4-fold dilutions, in DMSO. Compounds were dissolved into the media to a top concentration of 160 pM (4X). 25 pL of each media dilution was added to the plated cells, as well as 25 pL of the RealTime-Glo™ Extracellular ATP Assay solution (4X). A DMSO control was also added. Luminescence measurements were taken at specific times up to 24 hours.

The results are shown in Figures 8 and 9.

EXAMPLE 61 - PHARMACEUTICAL FORMULATIONS

(i) Tablet Formulation

A tablet composition containing a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 is prepared by mixing 50 mg of the compound with 197 mg of lactose (BP) as diluent, and 3 mg magnesium stearate as a lubricant and compressing to form a tablet in known manner.

(ii) Capsule Formulation

A capsule formulation is prepared by mixing 100 mg of a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 with 100 mg lactose and filling the resulting mixture into standard opaque hard gelatin capsules.

(iii) Injectable Formulation I

A parenteral composition for administration by injection can be prepared by dissolving a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 (e.g. in a salt form) in water containing 10% propylene glycol to give a concentration of active compound of 1.5 % by weight. The solution is then sterilised by filtration, filled into an ampoule and sealed.

(iv) Injectable Formulation

A parenteral composition for injection is prepared by dissolving in water a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 (e.g. in salt form) (2 mg/ml) and mannitol (50 mg/ml), sterile filtering the solution and filling into sealable 1 ml vials or ampoules. v) Injectable formulation III

A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of formula (1) as defined in any one of Embodiments 1.1 to 1.85 (e.g. in a salt form) in water at 20 mg/ml. The vial is then sealed and sterilised by autoclaving. vi) Injectable formulation IV

A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of formula (1) as defined in any one of Embodiments 1.1 to 1.85 (e.g. in a salt form) in water containing a buffer (e.g. 0.2 M acetate pH 4.6) at 20 mg/ml. The vial is then sealed and sterilised by autoclaving.

(vii) Subcutaneous Injection Formulation

A composition for sub-cutaneous administration is prepared by mixing a compound of the formula (1) as defined in any one of Embodiments 1.1 to 1.85 with pharmaceutical grade corn oil to give a concentration of 5 mg/ml. The composition is sterilised and filled into a suitable container. viii) Lyophilised formulation

Aliquots of formulated compound of formula (1) as defined in any one of Embodiments 1.1 to 1.85 are put into 50 ml vials and lyophilized. During lyophilisation, the compositions are frozen using a one-step freezing protocol at (-45 °C). The temperature is raised to - 10 °C for annealing, then lowered to freezing at -45 °C, followed by primary drying at +25 °C for approximately 3400 minutes, followed by a secondary drying with increased steps if temperature to 50 °C. The pressure during primary and secondary drying is set at 80 millitor. Equivalents

The foregoing examples are presented for the purpose of illustrating the invention and should not be construed as imposing any limitation on the scope of the invention. It will readily be apparent that numerous modifications and alterations may be made to the specific embodiments of the invention described above and illustrated in the examples without departing from the principles underlying the invention. All such modifications and alterations are intended to be embraced by this application.