Related Application

This application claims the benefit of priority to U.S. Provisional Patent Application No. 62/609,514, filed December 22, 2017, which application is hereby incorporated by reference in its entirety.

Background of the Invention

Presented herein are processes for preparing cyclohexanol ring systems that are useful in the preparation of compounds for treating at least one condition associated with inhibitition of the deposition of amyloid b peptide (Ab) and portions thereof by inhibiting or the beta site APP Cleaving Enzyme (BACE).

The prime neuropathological event distinguishing Alzheimer's disease (AD) is deposition of the 40-42 residue Ab in brain parenchyma and cerebral vessels. A large body of genetic, biochemical and in vivo data supports a pivotal role for Ab in the pathological cascade that eventually leads to AD. Patients usually present early symptoms (commonly memory loss) in their sixth or seventh decades of life. The disease progresses with increasing dementia and elevated deposition of Ab. In parallel, a hyperphosphorylated form of the microtubule-associated protein tau accumulates within neurons, leading to a plethora of deleterious effects on neuronal function. The prevailing working hypothesis regarding the temporal relationship between Ab and tau pathologies states that Ab deposition precedes tau aggregation in humans and animal models of the disease. Within this context, it is worth noting that the exact molecular nature of Ab mediating this pathological function is presently an issue under intense study. Most likely, there is a continuum of toxic species ranging from lower order Ab oligomers to supramolecular assemblies such as Ab fibrils.

The Ab peptide is an integral fragment of the Type I protein APP (Ab amyloid precursor protein), a protein ubiquitously expressed in human tissues. Since soluble Ab can be found in both plasma and cerebrospinal fluid (CSF), and in the medium from cultured cells, APP has to undergo proteolysis. There are three main cleavages of APP that are relevant to the pathobiology of AD, the so-called a-, b-, and g-cleavages. The a-cleavage, which occurs roughly in the middle of the Ab domain in APP, is executed by the metalloproteases

ADAM110 or ADAM117 (the latter also known as TACE). The b-cleavage, occurring at the N terminus of Ab, is generated by the transmembrane aspartyl protease Beta site APP

Cleaving Enzymel (BACE1). The g-cleavage, generating the Ab C termini and subsequent release of the peptide, is carried out by a multi-subunit aspartyl protease named g-secretase. ADAM10/17 cleavage followed by g-secretase cleavage results in the release of the soluble p3 peptide, an N-terminally truncated Ab fragment that fails to form amyloid deposits in humans. This proteolytic route is commonly referred to as the nonamyloidogenic pathway. Consecutive cleavages by BACE1 and g-secretase generate the intact Ab peptides; hence this processing scheme has been termed the amyloidogenic pathway. With this knowledge at hand, it is possible to envision two possible avenues of lowering Ab production: stimulating non-amyloidogenic processing, or inhibiting or modulating amyloidogenic processing. This application focuses on the latter strategy, inhibition or modulation of amyloidogenic processing.

Amyloidogenic plaques and vascular amyloid angiopathy also characterize the brains of patients with Trisomy 21 (Down's Syndrome), Hereditary Cerebral Hemorrhage with Amyloidosis of the Dutch-type (HCHW A-D), and other neurodegenerative disorders.

Neurofibrillary tangles also occur in other neurodegenerative disorders including dementia- inducing disorders (Varghese, J., et al, Journal of Medicinal Chemistry, 2003, 46, 4625-4630). b-amyloid deposits are predominately an aggregate of Ab peptide, which in turn is a product of the proteolysis of amyloid precursor protein (APP). More specifically, Ab peptide results from the cleavage of APP at the C-terminus by one or more g-secretases, and at the N- terminus by b-secretase enzyme (BACE), also known as aspartyl protease or Asp2 or Beta site APP Cleaving Enzyme (BACE), as part of the b-amyloidogenic pathway.

BACE activity is correlated directly to the generation of Ab peptide from APP (Sinha, et al, Nature, 1999, 402, 537-540), and studies increasingly indicate that the inhibition of BACE inhibits the production of Ab peptide (Roberds, S. L., et al, Human Molecular

Genetics, 2001, 10, 1317-1324). BACE is a membrane-bound type 1 protein that is synthesized as a partially active proenzyme and is abundantly expressed in brain tissue. It is thought to represent the major b-secretase activity, and is considered to be the rate-limiting step in the production of amyloid^-peptide (Ab).

Drugs that reduce or block BACE activity should therefore reduce Ab levels and levels of fragments of Ab in the brain, or elsewhere where Ab or fragments thereof deposit, and thus slow the formation of amyloid plaques and the progression of AD or other maladies involving deposition of Ab or fragments thereof. BACE is therefore an important candidate for the development of drugs as a treatment and/or prophylaxis of Ab-related pathologies such as Down's syndrome; b-amyloid angiopathy, such as but not limited to cerebral amyloid angiopathy or hereditary cerebral hemorrhage; disorders associated with cognitive impairment such as but not limited to MCI ("mild cognitive impairment"); Alzheimer's Disease; memory loss; attention deficit symptoms associated with Alzheimer's disease; neurodegeneration associated with diseases, such as Alzheimer's disease or dementia, including dementia of mixed vascular and degenerative origin, pre-senile dementia, senile dementia, and dementia associated with Parkinson's disease; progressive supranuclear palsy or cortical basal degeneration. It would therefore be useful to inhibit the deposition of Ab and portions thereof by inhibiting BACE through inhibitors.

International PCT publications WO 2012/087237 and WO 2016/055858 describe the synthesis of a number of compounds with dihydroimidazole ring systems shown to be BACE inhibitors. Improved processes of making these compounds would be advantageous.

Described herein are stereo- and regioselective processes for preparing cyclohexanol ring systems that are useful in the preparation of such compounds.

Summary of the Application

The present application provides processes for preparing the compounds disclosed herein, wherein the compounds are BACE inhibitors, or salts thereof, and/or intermediates useful in the preparation of BACE inhibitors. In certain such embodiments, the BACE inhibitors are useful in the treatment or prevention of Ab-related pathologies, such as a b- amyloid angiopathy, cerebral amyloid angiopathy, hereditary cerebral hemorrhage, a disorder associated with cognitive impairment, MCI ("mild cognitive impairment"), Alzheimer's Disease, memory loss, attention deficit symptoms associated with Alzheimer's disease, neurodegeneration associated with Alzheimer's disease, dementia of mixed vascular origin, dementia of degenerative origin, pre-senile dementia, senile dementia, dementia associated with Parkinson's disease, progressive supranuclear palsy or cortical basal degeneration.

The present application provides a process comprising contacting a compound of o formula (I), , or a salt thereof, with a ketoreductase enzyme to form a O compound of formula (II), , or a salt thereof, wherein R ⁷ is selected from halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁸ is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; or R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring; and o is 1 , 2, or 3.

In certain embodiments, the ketoreductase enzyme provides both stereoselective and regioselective reduction of the compound of formula (I), or a salt thereof.

In certain embodiments, the compound of formula (II), or salt thereof, is produced in

OH excess of a compound of formula (III), or a salt thereof and in excess

OH of a compound of formula (IV), or a salt thereof. In certain such embodiments, the compound of formula (II), or salt thereof, and the compound of formula (III), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1,

60:1, 70:1, 80:1, 90:1, 95:1, or 100:1. In certain embodiments, the compound of formula (II), or salt thereof, and the compound of formula (IV), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 95:1, or 100:1.

In certain embodiments, the compound of formula (II), or salt thereof, is a compound

O

ΌH

(lla)

of formula (lla), or a salt thereof, or a compound of formula (lib), O , or a salt thereof, or a mixture of any of the foregoing. In certain such embodiments, the compound of formula (lla), or salt thereof, and the compound of formula (lib), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 95:1, or 100:1, such as at least 100:1. In certain embodiments of the foregoing processes, the percent yield of compound of formula (Ila), or salt thereof, is greater than 85%, 90%, 95%, or 99%.

The present application further provides a process comprising contacting a compound

of formula (G), , or a salt thereof, with a ketoreductase enzyme

to form a compound of formula (IG), , or a salt thereof, wherein R ² independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; o is 1, 2, or 3; and p is 0, 1, 2, 3, or 4.

In certain embodiments, the ketoreductase enzyme provides both stereoselective and regioselective reduction of the compound of formula (G), or a salt thereof.

In certain embodiments, the compound of formula (IG), or salt thereof, is produced in

OH excess of a compound of formula (IIG), or a salt thereof and

OH in excess of a compound of formula (IV’), or a salt thereof.

In certain such embodiments, the compound of formula (IG), or salt thereof, and the compound of formula (IIG), or salt thereof, are present in a ratio of at least 1: 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90:1, 95: 1, or 100: 1. In certain embodiments, the compound of formula (IG), or salt thereof, and the compound of formula (IV’), or salt thereof, are present in a ratio of at least 1 : 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90: 1, 95: 1, or 100: 1.

In certain embodiments, the compound of formula (IG), or salt thereof, is a compound (N’a)

of formula (H’a), , or a salt thereof, or a compound of

formula (H’b), , or a salt thereof, or a mixture of any of the foregoing. In certain such embodiments, the compound of formula (H’a), or salt thereof, and the compound of formula (H’b), or salt thereof, are present in a ratio of at least 1: 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90:1, 95: 1, or 100: 1, such as at least 100: 1.

In certain embodiments of the foregoing processes, the percent yield of compound of formula (H’a), or salt thereof, is greater than 85%, 90%, 95%, or 99%.

In certain embodiments, the compound of formula (G), or salt thereof, is compound 1, (1 ) , or a salt thereof.

In certain embodiments, the compound of formula (IG), or salt thereof, is compound salt thereof

In certain embodiments, the compound of formula (H’a), or salt thereof, is compound

(2a)

, or a salt thereof, and the compound of formula (H’b), (2b)

or salt thereof, is compound (2b), or a salt thereof

In certain embodiments, the ketoreductase enzyme is selected from the group consisting of KRED-P1-A04, KRED-P1-B02, KRED-P1-B05, KRED-P1-B10, KRED-P1-

B12, KRED-P1-C01, KRED-P1-H08, KRED-P1-H10, KRED-P2-B02, KRED-P2-C02, KRED-P2-C11, KRED-P2-D03, KRED-P2-D11, KRED-P2-D12, KRED-P2-G03, KRED- P2-H07, KRED-P3-B03, KRED-P3-G09, KRED-P3-H12, ADH-101, ADH-105, and ADH-

112, such as KRED-P2-D12, KRED-P1-H10, KRED-P1-B05, KRED-P1-B04, and KRED-

P1-H08, e.g., KRED-P1-H08.

In certain embodiments, the ketoreductase enzyme is present in an amount selected from 1-100% w/w based on the weight of the compound of formula (I), or salt thereof, such as 5% w/w based on the weight of the compound of formula (I), or salt thereof.

In certain embodiments, the compound of formula (I), or salt thereof, is present in an amount selected from 1-200 g/L, such as 100 g/L.

In certain embodiments of any of the processes disclosed herein, the process further comprises contacting the compound of formula (I), or salt thereof, with a cofactor. In certain such embodiments, the cofactor is selected from NAD or ADP, e.g., NADP. In certain embodiments, the compound of formula (G), or salt thereof, is contacted with the ketoreductase enzyme in the presence of isopropanol (IPA). In certain such embodiments, the aqueous IPA is present in an amount selected from 5-95% v/v, such as 50% v/v.

In certain embodiments of any of the foregoing processes, the process is performed at a pH between about 5 and about 9, such as at a pH of about 5.5.

In certain embodiments of any of the foregoing processes, the process is performed at a temperature of between about 30 to 50 °C, such as at a temperature of about 40 °C.

The present application further provides a process for preparing a compound of

(V)

formula ( , or a salt thereof, comprising any one of the foregoing processes,

wherein R ³ independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁷ is selected from halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁸ is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; or R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring; and o is 1, 2, or 3.

The present application further provides a process for preparing a compound of

)

formula ( , or a salt thereof, comprising any one of the foregoing processes, wherein R ² independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ³ independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; p is 0, 1, 2, 3, or 4; and o is 1, 2, or 3. Additional embodiments, features, and advantages of the application will be apparent from the following detailed description and through practice of the embodiments described in this application.

Detailed Description

The present application provides a process comprising contacting a compound of

O

(I)

formula (I), or a salt thereof, with a ketoreductase enzyme to form a

O compound of formula (II), , or a salt thereof, wherein R ⁷ is selected from halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁸ is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; or R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring; and o is 1 , 2, or 3.

In certain embodiments, R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring. In certain such embodiments, R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted aryl ring, such as an optionally substituted benzene ring. In certain embodiments of the foregoing, the optionally substituted aryl ring, such as an optionally substituted benzene ring, is substituted with halogen, such as bromine.

In certain embodiments, o is 1.

In certain embodiments, the ketoreductase enzyme provides both stereoselective and regioselective reduction of the compound of formula (I), or a salt thereof.

In certain embodiments, the compound of formula (II), or salt thereof, is produced in

OH excess of a compound of formula (III), , or a salt thereof. In certain embodiments, the compound of formula (II), or salt thereof, is produced in excess of a OH compound of formula (IV), or a salt thereof. In certain

embodiments, the compound of formula (II), or salt thereof, is produced in excess of a

OH

(III)

compound of formula (III), or a salt thereof, and in excess of a

compound of formula (IV), , or a salt thereof. In certain such embodiments, the compound of formula (II), or salt thereof, and the compound of formula (III), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1,

60:1, 70:1, 80:1, 90:1, 95:1, or 100:1. In certain embodiments, the compound of formula (II), or salt thereof, and the compound of formula (IV), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 95:1, or 100:1.

In certain embodiments, the compound of formula (II), or salt thereof, is a compound

O of formula (Ila), , or a salt thereof, or a compound of formula (lib),

O

OH

(lib)

or a salt thereof, or a mixture of any of the foregoing In certain such embodiments, the compound of formula (Ila), or salt thereof, and the compound of formula (lib), or salt thereof, are present in a ratio of at least 1:1, 5:1, 10:1, 20:1, 30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, 95:1, or 100:1, such as at least 100:1.

In certain embodiments of the foregoing processes, the percent yield of compound of formula (Ila), or salt thereof, is greater than 85%, 90%, 95%, or 99%.

The present application further provides a process comprising contacting a compound

of formula (G), , or a salt thereof, with a ketoreductase enzyme

to form a compound of formula (IG), , or a salt thereof, wherein R ² independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; o is 1, 2, or 3; and p is 0, 1, 2, 3, or 4.

In certain embodiments, R ² is halogen, such as bromine.

In certain embodiments, o is 1.

In certain embodiments, p is 1.

In certain embodiments, the ketoreductase enzyme provides both stereoselective and regioselective reduction of the compound of formula (G), or a salt thereof.

In certain embodiments, the compound of formula (IG), or salt thereof, is produced in

OH excess of a compound of formula (IIG), (in') , or a salt thereof. In certain embodiments, the compound of formula (IG), or salt thereof, is produced in excess of

OH (IV)

a compound of formula (IV’), , or a salt thereof In certain embodiments, the compound of formula (IG), or salt thereof, is produced in excess of a

OH (HI')

compound of formula (IIG), or a salt thereof, and in excess

of a compound of formula (IV’), or a salt thereof In certain such embodiments, the compound of formula (IG), or salt thereof, and the compound of formula (IIG), or salt thereof, are present in a ratio of at least 1 : 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90: 1, 95: 1, or 100: 1. In certain embodiments, the compound of formula (IG), or salt thereof, and the compound of formula (IV’), or salt thereof, are present in a ratio of at least 1 : 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90: 1, 95: 1, or 100: 1.

In certain embodiments, the compound of formula (IG), or salt thereof, is a compound a)

of formula ( , or a salt thereof, or a compound of formula ( salt thereof, or a mixture of any of the foregoing. In certain such embodiments, the compound of formula (H’a), or salt thereof, and the compound of formula (H’b), or salt thereof, are present in a ratio of at least 1: 1, 5: 1, 10: 1, 20: 1, 30: 1, 40: 1, 50: 1, 60: 1, 70: 1, 80: 1, 90:1, 95: 1, or 100: 1, such as at least 100: 1.

In certain embodiments of the foregoing processes, the percent yield of compound of formula (H’a), or salt thereof, is greater than 85%, 90%, 95%, or 99%.

In certain embodiments, the compound of formula (G), or salt thereof, is compound 1, il) or a salt thereof.

In certain embodiments, the compound of formula (IG), or salt thereof, is compound salt thereof

In certain embodiments, the compound of formula (H’a), or salt thereof, is compound

O , or a salt thereof, and the compound of formula (H’b),

O or salt thereof, is compound (2b), , or a salt thereof.

In certain embodiments, the ketoreductase enzyme may be a natural or genetically engineered enzyme. In certain embodiments, the ketoreductase enzyme may be used with any suitable cofactor, such as NADH or NADPH. In certain embodiments, the ketoreductase enzyme is a NADPH-dependent ketoreductase enzyme that is used in conjunction with NADPH as a co-factor. In other embodiments, the ketoreductase enzyme is a NADPH- dependent ketoreductase enzyme that is used in conjunction with with cofactors other than NADPH, such as NADH. In other embodiments, the ketoreductase enzyme is a NADH- dependent ketoreductase enzyme that is used in conjunction with NADPH as a co-factor. In yet other embodiments, the ketoreductase enzyme is a NADH-dependent ketoreductase enzyme that is used in conjunction in conjunction with cofactors other than NADH, such as NADPH. In certain embodiments, the ketoreductase enzyme is selected from the group consisting of KRED-P 1 -A04, KRED-P1-B02, KRED-P1-B05, KRED-P1-B10, KRED-P1- B12, KRED-P1-C01, KRED-P 1-H08, KRED-P l-H 10, KRED-P2-B02, KRED-P2-C02, KRED-P2-C11, KRED-P2-D03, KRED-P2-D11, KRED-P2-D12, KRED-P2-G03, KRED- P2-H07, KRED-P3-B03, KRED-P3-G09, KRED-P3-H12, ADH-101, ADH-105, and ADH- 112, all available from Codexis, Inc. in Redwood City, California, USA. In certain such embodiments, the ketoreductase enzyme is selected from the group consisting of KRED-P2- D12, KRED-P l-H 10, KRED-P 1-B05, KRED-P 1-B04, and KRED-P 1-H08, such as KRED- P1-H08.

In certain embodiments, the ketoreductase enzyme is present in an amount selected from 1-100% w/w based on the weight of the compound of formula (I), or salt thereof. In certain embodiments, the ketoreductase enzyme is present in an amount selected from 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, or 10% w/w based on the weight of the compound of formula (I), or salt thereof, such as 5% w/w based on the weight of the compound of formula (I), or salt thereof.

In certain embodiments, the compound of formula (I), or salt thereof, is present in an amount selected from 1-200 g/L. In certain embodiments, the compound of formula (I), or salt thereof, is present in an amount selected from 1-150 g/L, 25-150 g/L, 50-150 g/L, 75-150 g/L, 75-125 g/L, or 75-100 g/L, such as 100 g/L.

In certain embodiments, the compound of formula (G), or salt thereof, is contacted with the ketoreductase enzyme in the presence of an alcohol, such as methanol, ethanol, or isopropanol (IP A). In certain embodiments, the compound of formula (G), or salt thereof, is contacted with the ketoreductase enzyme in the presence of isopropanol (IP A). In certain such embodiments, the alcohol, such as aqueous IPA, is present in an amount selected from 5-95% v/v. In certain embodiments of the foregoing, the alcohol, such as aqueous IPA, is present in an amount selected from 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, or 70% v/v, such as 50% v/v.

In certain embodiments of any of the foregoing processes, the process is performed at a pH between about 5 and about 9. In certain such embodiments, the process is performed at a pH between about 4.5 and about 6.5, such as at a pH of between about 5 and about 6. In certain embodiments, the process is performed at a pH of about 5.5.

In certain embodiments of any of the foregoing processes, the process is performed at a temperature of between about 30 to 50 °C, such as at a temperature of between about 35-45 °C. In certain such embodiments, the process is performed at a temperature of about 40 °C. The present application further provides a process for preparing a compound of

(V)

formula ( , or a salt thereof, comprising any one of the foregoing processes,

wherein R ³ independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁷ is selected from halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ⁸ is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; or R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring; and o is 1, 2, or 3.

In certain embodiments, R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused cycloalkyl, heterocycloalkyl, cycloalkenyl, heterocycloalkenyl, aryl, or heteroaryl ring. In certain such embodiments, R ⁷ and R ⁸ taken together with with the carbon(s) to which they are attached, form an optionally substituted fused aryl ring, such as an optionaly substituted fused benzene ring. In certain such embodiments, the fused aryl ring, such as a fused benzene ring, is substituted with halogen, such as bromine.

In certain embodiments, o is 1.

The present application further provides a process for preparing a compound of

)

formula ( , or a salt thereof, comprising any one of the foregoing processes, wherein R ² independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; R ³ independently for each occurrence, is selected from hydrogen, halogen, CN, and optionally substituted alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, or oxime; p is 0, 1, 2, 3, or 4; and o is 1, 2, or 3.

In certain embodiments, R ² is halogen, such as bromine. In certain embodiments, p is 1.

In certain embodiments, o is 1.

In certain embodiments wherein alkyl, alkenyl, alkynyl, alkoxy, aryl, heteroaryl, cycloalkyl, heterocycloalkyl, or oxime are substituted, they are substituted, valency permitting, with one or more substituents selected from substituted or unsubstituted alkyl, such as perfluoroalkyl (e.g., trifluoromethyl), alkenyl, alkoxy, alkoxyalkyl, aryl, aralkyl, arylalkoxy, aryloxy, aryloxyalkyl, hydroxyl, halo, alkoxy, such as perfluoroalkoxy (e.g., trifluoromethoxy), alkoxyalkoxy, hydroxyalkyl, hydroxyalkylamino, hydroxyalkoxy, amino, aminoalkyl, alkylamino, aminoalkylalkoxy, aminoalkoxy, acylamino, acylaminoalkyl, such as perfluoro acylaminoalkyl (e.g., trifluoromethylacylaminoalkyl), acyloxy, cycloalkyl, cycloalkylalkyl, cycloalkylalkoxy, heterocyclyl, heterocyclylalkyl, heterocyclyloxy, heterocyclylalkoxy, heteroaryl, heteroarylalkyl, heteroarylalkoxy, heteroaryloxy,

heteroaryloxyalkyl, heterocyclylaminoalkyl, heterocyclylaminoalkoxy, amido, amidoalkyl, amidine, imine, oxo, carbonyl (such as carboxyl, alkoxycarbonyl, formyl, or acyl, including perfluoroacyl (e.g., C(0)CF3)), carbonylalkyl (such as carboxyalkyl, alkoxycarbonylalkyl, formylalkyl, or acylalkyl, including perfluoroacylalkyl (e.g., -alkylC(0)CF3)), carbamate, carbamatealkyl, urea, ureaalkyl, sulfate, sulfonate, sulfamoyl, sulfone, sulfonamide, sulfonamidealkyl, cyano, nitro, azido, sulfhydryl, alkylthio, thiocarbonyl (such as thioester, thioacetate, or thioformate), phosphoryl, phosphate, phosphonate or phosphinate.

When a bond to a substituent is shown to cross a bond connecting two atoms in a ring, then such substituent may be bonded to any atom on the ring. When a substituent is listed without indicating the atom via which such substituent is bonded to the rest of the compound of a given formula, then such substituent may be bonded via any atom in such substituent. Combinations of substituents, positions of substituents and/or variables are permissible only if such combinations result in stable compounds.

Those skilled in the art will recognize that the species listed or illustrated herein are not exhaustive, and that additional species within the scope of these defined terms may also be selected.

Throughout this disclosure it is to be understood that, where appropriate, suitable protecting groups may be added to, and subsequently removed from, the various reactants and intermediates in a manner that will be readily understood by one skilled in the art of organic synthesis. Conventional procedures for using such protecting groups as well as examples of suitable protecting groups are described, for example, in“Protective Groups in Organic Synthesis,” T. W. Green, P. G. M. Wuts, Wiley-Interscience, New York, (1999).

A transformation of a group or substituent into another group or substituent by chemical manipulation can be conducted on any intermediate or final product on the synthetic path toward the final product; the type of transformation is limited only by the inherent incompatibility of other functional groups contained in the molecule to the conditions or reagents employed in the transformation. Such inherent incompatibilities, and ways to circumvent them by carrying out appropriate transformations and synthetic steps in a suitable order, will be readily understood by one skilled in the art of organic synthesis.

Examples of transformations are given throughout this disclosure, and it is understood that the described transformations are not limited only to the generic groups or substituents for which the transformations are exemplified. References and descriptions of other suitable transformations are given in“Comprehensive Organic Transformations— A Guide to Functional Group Preparations” R. C. Larock, Wiley VCH, 2 ^nd Edition (1999).

Examplary reaction conditions are given throughout this disclosure, and it is understood that the described reaction conditions are not limited only to the described reaction conditions. References and descriptions of other suitable reaction conditions are described in textbooks of organic chemistry, such as, for example,“Advanced Organic Chemistry”, March 6 ^th Edition, Wiley Interscience (2007), and“Organic Synthesis”, Smith, 2 ^nd Edition, McGraw Hill, (2001).

In certain embodiments, the present application provides processes for the preparation of BACE inhibitors that are useful in the treatment or prevention of Ab-related pathologies.

In certain such embodiments, the Ab-related pathology is Down's syndrome, a b-amyloid angiopathy, cerebral amyloid angiopathy, hereditary cerebral hemorrhage, a disorder associated with cognitive impairment, MCI ("mild cognitive impairment"), Alzheimer's Disease, memory loss, attention deficit symptoms associated with Alzheimer's disease, neurodegeneration associated with Alzheimer's disease, dementia of mixed vascular origin, dementia of degenerative origin, pre-senile dementia, senile dementia, dementia associated with Parkinson's disease, progressive supranuclear palsy or cortical basal degeneration.

Definitions

The definitions set forth in this application are intended to clarify terms used throughout this application. Unless defined otherwise, all technical and scientific terms used herein have the same meaning as is commonly understood by one of ordinary skill in the art to which this application belongs. All patents, applications, published applications and other publications referred to herein are incorporated by reference in their entireties to disclose and describe the methods and/or materials in connection with which the publications are cited. If a definition set forth in this section is contrary to or otherwise inconsistent with a definition set forth in a patent, application, or other publication that is herein incorporated by reference, the definition set forth in this section prevails over the definition incorporated herein by reference.

Although any methods and materials similar or equivalent to those described herein can also be used in the practice or testing of the embodiments in present application, the preferred methods and materials are now described.

To provide a more concise description, some of the quantitative expressions given herein are not qualified with the term“about.” It is understood that, whether the term“about” is used explicitly or not, every quantity given herein is meant to refer to the actual given value, and it is also meant to refer to the approximation to such given value that would reasonably be inferred based on the ordinary skill in the art, including equivalents and approximations due to the experimental and/or measurement conditions for such given value. Whenever a yield is given as a percentage, such yield refers to a mass of the entity for which the yield is given with respect to the maximum amount of the same entity that could be obtained under the particular stoichiometric conditions. Concentrations that are given as percentages refer to mass ratios, unless indicated differently.

Except as otherwise noted, the methods and techniques of the present embodiments are generally performed according to conventional methods well known in the art and as described in various general and more specific references that are cited and discussed throughout the present specification. See, e.g., Loudon, Organic Chemistry, Fourth Edition, New York: Oxford University Press, 2002, pp. 360-361, 1084-1085; Smith and March, March’s Advanced Organic Chemistry: Reactions, Mechanisms, and Structure, Fifth Edition, Wiley-Interscience, 2001.

The nomenclature used herein to name the subject compounds is illustrated in the Examples herein. This nomenclature has generally been derived using the commercially - available ChemBioDraw Ultra software (Cambridgesoft/Perkin Elmer), Version 12.0.

It is to be understood that the present description is not limited to particular embodiments described, as such may, of course, vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to be limiting, since the scope of the present application will be limited only by the appended claims.

It is appreciated that certain features of the application, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the application, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination. All combinations of the embodiments pertaining to the chemical groups represented by the variables are specifically embraced by the present application and are disclosed herein just as if each and every combination was individually and explicitly disclosed, to the extent that such combinations embrace compounds that are stable compounds (i.e., compounds that can be isolated, characterized, and tested for biological activity). In addition, all subcombinations of the chemical groups listed in the embodiments describing such variables are also specifically embraced by the present application and are disclosed herein just as if each and every such sub-combination of chemical groups was individually and explicitly disclosed herein.

Any formula depicted herein is intended to represent a compound of that structural formula as well as certain variations or forms. E.g., a formula given herein is intended to include a racemic form, or one or more enantiomeric, diastereomeric, or geometric isomers, or tautomeric forms, or a mixture thereof. Additionally, any formula given herein is intended to refer also to a solvate, such as a hydrate, solvate, or polymorph of such a compound, or a mixture thereof. Any formula given herein is intended to refer to amorphous and/or crystalline physical forms of the compound. The compounds described herein may be analytically pure, or a mixture in which the compound comprises at least 50%, at least 70%, at least 80%, at least 90%, at least 95%, or at least 98% by weight of the mixture.

In addition, where features or aspects of the embodiments of this application are described in terms of Markush groups, those skilled in the art will recognize that

embodiments described herein is also thereby described in terms of any individual member or subgroup of members of the Markush group. E.g., if X is described as selected from the group consisting of bromine, chlorine, and iodine, claims for X being bromine and claims for X being bromine and chlorine are fully described.

The term "herein" refers to the entire application. As used herein, the singular forms“a,”“an,” and“the” include plural referents unless the context clearly dictates otherwise. It is further noted that the claims may be drafted to exclude any optional element. As such, this statement is intended to serve as antecedent basis for use of such exclusive terminology as“solely,”“only” and the like in connection with the recitation of claim elements, or use of a“negative” limitation.

As used herein, the terms“including,”“containing,” and“comprising” are used in their open, non-limiting sense.

As used herein,“subject” (as in the subject of the treatment) refers to both mammals and non-mammals. Mammals include, e.g., humans; non-human primates, e.g. apes and monkeys; and non-primates, e.g. mice, rats, rabbits, dogs, cats, cattle, horses, sheep, and goats. Non-mammals include, e.g., worms, fish and birds. In some embodiments, the subject is a human.

"Substantially" as the term is used herein refers to being completely or almost completely; e.g., a composition that is "substantially free" of a component either has none of the component or contains such a trace amount that any relevant functional property of the composition is unaffected by the presence of the trace amount, or a compound is

"substantially pure" is there are only negligible traces of impurities present.

The term“acyl” is art-recognized and refers to a group represented by the general formula hydrocarbylC(O)-, preferably alkylC(O)-.

The term“acylamino” is art-recognized and refers to an amino group substituted with an acyl group and may be represented, e.g., by the formula hydrocarbylC(0)NH-.

The term“acyloxy” is art-recognized and refers to a group represented by the general formula hydrocarbylC(0)0-, preferably alkylC(0)0-.

The term“alkoxy” refers to an alkyl group, preferably a lower alkyl group, having an oxygen attached thereto. Representative alkoxy groups include methoxy, ethoxy, propoxy, tert-butoxy and the like.

The term“alkoxyalkyl” refers to an alkyl group substituted with an alkoxy group and may be represented by the general formula alkyl-O-alkyl.

The term“alkenyl”, as used herein, refers to an aliphatic group containing at least one double bond and is intended to include both "unsubstituted alkenyls" and "substituted alkenyls", the latter of which refers to alkenyl moieties having substituents replacing a hydrogen on one or more carbons of the alkenyl group. Such substituents may occur on one or more carbons that are included or not included in one or more double bonds. Moreover, such substituents include all those contemplated for alkyl groups, as discussed below, except where stability is prohibitive. E.g., substitution of alkenyl groups by one or more alkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups is contemplated.

The term“alkynyl”, as used herein, refers to an aliphatic group containing at least one triple bond and is intended to include both "unsubstituted alkynyls" and "substituted alkynyls", the latter of which refers to alkynyl moieties having substituents replacing one or more hydrogens on one or more carbons of the alkynyl group. Such substituents may occur on one or more carbons that are included or not included in one or more triple bonds.

Moreover, such substituents include all those contemplated for alkyl groups, as discussed above, except where stability is prohibitive. E.g., substitution of alkynyl groups by one or more alkyl, carbocyclyl, aryl, heterocyclyl, or heteroaryl groups is contemplated.

An“alkyl” group or“alkane” is a straight chained or branched non-aromatic hydrocarbon which is completely saturated. Typically, a straight chained or branched alkyl group has from 1 to about 20 carbon atoms, such as from 1 to 12 carbon atoms, preferably from 1 to about 10, more preferably from 1 to 4, unless otherwise defined. Examples of straight chained and branched alkyl groups include methyl, ethyl, n-propyl, isopropyl, n- butyl, isobutyl, sec -butyl, tert-butyl, pentyl, isopentyl, tert-pentyl, hexyl, isohexyl, pentyl and octyl. A Ci-Ce straight chained or branched alkyl group is also referred to as a "lower alkyl" group.

Moreover, the term "alkyl" (or "lower alkyl") as used throughout the specification, examples, and claims is intended to include both "unsubstituted alkyls" and "substituted alkyls", the latter of which refers to alkyl moieties having substituents replacing a hydrogen or more hydrogens on one or more carbons of the hydrocarbon backbone. Such substituents, if not otherwise specified, can include, e.g., a halogen, a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbonyl, a formyl, or an acyl), a thiocarbonyl (such as a thioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, a phosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine, an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, a sulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, a heterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. It will be understood by those skilled in the art that the moieties substituted on the hydrocarbon chain can themselves be substituted, if appropriate. For instance, the substituents of a substituted alkyl may include substituted and unsubstituted forms of amino, azido, imino, amido, phosphoryl (including phosphonate and phosphinate), sulfonyl (including sulfate, sulfonamido, sulfamoyl and sulfonate), and silyl groups, as well as ethers, alkylthios, carbonyls (including ketones, aldehydes, carboxylates, and esters), -CF ₃ , -CN and the like. Exemplary substituted alkyls are described below. Cycloalkyls can be further substituted with alkyls, alkenyls, alkoxys, alkylthios, aminoalkyls, carbonyl-substituted alkyls, -CF ₃ , -CN, and the like.

The term“(ATOM)i-j” with j > i, when used in conjunction with a chemical moiety, such as, acyl, acyloxy, alkyl, alkenyl, alkynyl, or alkoxy is meant to include groups that contain from i to j (including i and j) atoms. E.g., the term“C _x-y alkyl” refers to substituted or unsubstituted saturated hydrocarbon groups, including straight-chain alkyl and branched- chain alkyl groups that contain from x to y carbons in the chain, including haloalkyl groups such as trifluoromethyl and 2,2,2-tirfluoroethyl, etc. Co alkyl refers to a hydrogen atom where the group is in a terminal position, a bond if internal. Similarly, e.g., C _3-6 cycloalkyl refers to a cycloalkyl as defined herein that has 3 to 6 carbon ring atoms. The terms“C2- _y alkenyl” and “C2- _y alkynyl” refer to substituted or unsubstituted unsaturated aliphatic groups analogous in length and possible substitution to the alkyls described above, but that contain at least one double or triple bond respectively.

The term“alkylamino”, as used herein, refers to an amino group substituted with at least one alkyl group.

The term“alkylthio”, as used herein, refers to a thiol group substituted with an alkyl group and may be represented by the general formula alkylS-.

The term“hydrocarbyl”, as used herein, refers to a group that is bonded through a carbon atom that does not have a =0 or =S substituent, and typically has at least one carbon- hydrogen bond and a primarily carbon backbone, but may optionally include heteroatoms. Thus, groups like methyl, ethoxyethyl, 2-pyridyl, and trifluoromethyl are considered to be hydrocarbyl for the purposes of this application, but substituents such as acetyl (which has a =0 substituent on the linking carbon) and ethoxy (which is linked through oxygen, not carbon) are not. Hydrocarbyl groups include, but are not limited to aryl, heteroaryl, carbocycle, heterocyclyl, alkyl, alkenyl, alkynyl, and combinations thereof.

The terms“amine” and“amino” are art-recognized and refer to both unsubstituted and substituted amines and salts thereof, e.g., a moiety that can be represented by , wherein each R ³⁰ independently represents a hydrogen or a hydrocarbyl group, or two R ³⁰ are taken together with the N atom to which they are attached complete a heterocycle having from 4 to 8 atoms in the ring structure. The term“aminoalkyl”, as used herein, refers to an alkyl group substituted with an amino group.

o

I I R

The term“amide”, as used herein, refers to a group: R ³⁰ , wherein each R ³⁰ independently represent a hydrogen or hydrocarbyl group, or two R ³⁰ are taken together with the N atom to which they are attached complete a heterocycle having from 4 to 8 atoms in the ring structure.

The term“carbamate” is art-recognized and refers to a group

wherein R ²⁹ and R ³⁰ independently represent hydrogen or a hydrocarbyl group, such as an alkyl

group, or R ²⁹ and R ³⁰ taken together with the intervening atom(s) complete a heterocycle having

from 4 to 8 atoms in the ring structure.

The term“halogen,” or“halide” represents chlorine, fluorine, bromine, or iodine. The term“halo” represents fluoro, chloro, bromo, or iodo.

The term“haloalkyl”, as used herein, refers to an alkyl group with one or more halo substituents, or one, two, or three halo substituents. Examples of haloalkyl groups include - CFs, -CH ₂ F, -CHF2, -CH ₂ Br, -CfFCFs, and -CH ₂ CH ₂ F.

The term“heteroatom”, as used herein, refers to an atom of any element other than carbon or hydrogen. Exemplary heteroatoms include but are not limited to nitrogen, oxygen, and sulfur.

The term“heteroalkyl”, as used herein, refers to a saturated or unsaturated chain of carbon atoms and at least one heteroatom, wherein no two heteroatoms are adjacent.

The term“aryl”, as used herein, includes substituted or unsubstituted monocyclic aromatic rings in which each atom of the ring is carbon. Preferably the ring is a 5- to 7- membered ring, more preferably a 6-membered ring. The term“aryl” also includes polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is aromatic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls.

Aryl groups include benzene, naphthalene, phenanthrene, phenol, aniline, and the like. The term“aralkyl”, as used herein, refers to an alkyl group substituted with an aryl group.

An“aroyl” group, as the term is used herein, refers to an aryl group bonded via an exocyclic carbonyl group, such as a benzoyl group.

The term“heteroaryl” , as used herein, includes substituted or unsubstituted monocyclic aromatic ring system, preferably 5- to 7-membered aromatic rings, more preferably 5- to 6-membered rings, whose ring structures include at least one heteroatom, preferably one to four heteroatoms, more preferably one to two heteroatoms. E.g., a 5- membered heteroaryl is furan, thiophene, pyrrole, oxazole, isoxazole, thiazole, isothiazole, pyrazole, imidazole, oxadiazole, thiadiazole, triazole, or tetrazole. In another example, a 6- membered heteroaryl is pyridine, pyrazine, pyrimidine, pyridazine, or triazine. The term “heteroaryl” also include substituted or unsubstituted“polycyclic” ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is heteroaromatic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls.

Illustrative examples of heteroaryl groups include but are not limited to the following entities, in the form of properly bonded moieties:

The term“heteroaralkyl” or“hetaralkyl”, as used herein, refers to an alkyl group substituted with a heteroaryl group.

A“heteroaroyl” group, as the term is used herein, refers to a heteroaryl group bonded via an exocyclic carbonyl group, analogous to a benzoyl group but wherein the phenyl ring of the benzoyl group is replaced by a heteroaryl group.

The terms“heterocyclyl”,“heterocycle”, and“heterocyclic”, as used herein, refer to substituted or unsubstituted non-aromatic ring structures, preferably 3- to lO-membered rings, more preferably 3- to 7-membered rings, whose ring structures include at least one heteroatom, preferably one to four heteroatoms, more preferably one or two heteroatoms. The terms“heterocyclyl” and“heterocyclic” also include substituted or unsubstituted polycyclic ring systems having two or more cyclic rings in which two or more carbons are common to two adjoining rings wherein at least one of the rings is heterocyclic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or

heterocyclyls. Heterocyclyl groups include, e.g., piperidine, piperazine, pyrrolidine, morpholine, lactones, lactams, and the like.

The term“heterocyclylalkyl”, as used herein, refers to an alkyl group substituted with a heterocycle group which is optionally substituted.

The terms“carbocycle”, and“carbocyclic”, as used herein, refers to a saturated or unsaturated ring in which each atom of the ring is carbon. The term carbocycle includes both aromatic carbocycles and non-aromatic carbocycles. Non-aromatic carbocycles include both cycloalkane rings, in which all carbon atoms are saturated, and cycloalkene rings, which contain at least one double bond.“Carbocycle” includes 5-7 membered monocyclic and 8-12 membered bicyclic rings. Each ring of a bicyclic carbocycle may be selected from saturated, unsaturated and aromatic rings. Carbocycle includes bicyclic molecules in which one, two or three or more atoms are shared between the two rings. The term“fused carbocycle” refers to a bicyclic carbocycle in which each of the rings shares two adjacent atoms with the other ring. Each ring of a fused carbocycle may be selected from saturated, unsaturated and aromatic rings. In an exemplary embodiment, an aromatic ring, e.g., phenyl, may be fused to a saturated or unsaturated ring, e.g., cyclohexane, cyclopentane, or cyclohexene. Any combination of saturated, unsaturated and aromatic bicyclic rings, as valence permits, is included in the definition of carbocyclic. Exemplary“carbocycles” include cyclopentane, cyclohexane, bicyclo[2.2. l]heptane, l,5-cyclooctadiene, l,2,3,4-tetrahydronaphthalene, bicyclo[4.2.0]oct-3-ene, naphthalene and adamantane. Exemplary fused carbocycles include decalin, naphthalene, l,2,3,4-tetrahydronaphthalene, bicyclo[4.2.0]octane, 4,5,6,7-tetrahydro- lH-indene and bicyclo[4. l.0]hept-3-ene.“Carbocycles” may be susbstituted at any one or more positions capable of bearing a hydrogen atom.

A“cycloalkyl” group, as used herein, refers to a substituted or unsubstituted cyclic hydrocarbon which is completely saturated.“Cycloalkyl” includes substituted or

unsubstituted monocyclic and bicyclic rings. Typically, a monocyclic cycloalkyl group has from 3 to about 10 carbon atoms, more typically 3 to 8 carbon atoms unless otherwise defined. Such a monocyclic cycloalkyl group may be substituted or unsubstituted. The second ring of a bicyclic cycloalkyl may be selected from saturated, unsaturated and aromatic rings that are substituted or unsubstituted. Cycloalkyl includes substituted or unsubstituted bicyclic molecules in which one, two or three or more atoms are shared between the two rings. The term“fused cycloalkyl” refers to a substituted or unsubstituted bicyclic cycloalkyl in which each of the rings shares two adjacent atoms with the other ring. The second ring of a fused bicyclic cycloalkyl may be selected from saturated, unsaturated and aromatic rings.

The term“carbocyclylalkyl”, as used herein, refers to an alkyl group substituted with a carbocycle group.

A“cycloalkenyl” group, as used herein, refers to a cyclic hydrocarbon containing one or more double bonds. A“cycloalkynyl” group is a cyclic hydrocarbon containing one or more triple bonds.

The terms“polycyclyl”,“polycycle”, and“polycyclic”, as used herein, refer to two or more rings (e.g., cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls) in which two or more atoms are common to two adjoining rings, e.g., the rings are“fused rings”. Each of the rings of the polycycle can be substituted or unsubstituted. In certain embodiments, each ring of the polycycle contains from 3 to 10 atoms in the ring, preferably from 5 to 7.

The term“carbonate” is art-recognized and refers to a group -OCO2-R ³⁰ , wherein R ³⁰ represents a hydrocarbyl group.

The term“carboxy”, as used herein, refers to a group represented by the

formula -CO2H.

The term“ester”, as used herein, refers to a group -C(0)0R ³ ° wherein R ³⁰ represents a hydrocarbyl group.

The term“ether,” as used herein, refers to a hydrocarbyl group linked through an oxygen to another hydrocarbyl group. Accordingly, an ether substituent of a hydrocarbyl group may be hydrocarbyl-O-. Ethers may be either symmetrical or unsymmetrical.

Examples of ethers include, but are not limited to, heterocycle-O-heterocycle and aryl-O- heterocycle. Ethers include“alkoxyalkyl” groups, which may be represented by the general formula alkyl-O-alkyl.

The term“sulfate” is art-recognized and refers to the group -OSO3H, or a

pharmaceutically acceptable salt thereof. The term“sulfonamide” is art-recognized and refers to the group represented by the

general formulae

wherein R ²⁹ and R ³⁰ independently represents hydrogen or hydrocarbyl, such as alkyl, or R ²⁹ and R ³⁰ taken together with the intervening atom(s) complete a heterocycle having from 4 to 8 atoms in the ring structure.

The term“sulfoxide” is art-recognized and refers to the group -S(0)-R ³ °, wherein R ³⁰ represents a hydrocarbyl.

The term“sulfonate” is art-recognized and refers to the group SO3H, or a

pharmaceutically acceptable salt thereof.

The term“sulfone” is art-recognized and refers to the group -S(0)2-R ³ °, wherein R ³⁰ represents a hydrocarbyl.

The term“thioalkyl”, as used herein, refers to an alkyl group substituted with a thiol group.

The term“thioester”, as used herein, refers to a group -C(0)SR ³ ° or -SC(0)R ³ ° wherein R ³⁰ represents a hydrocarbyl.

The term“thioether”, as used herein, is equivalent to an ether, wherein the oxygen is replaced with asulfur.

The term“urea” is art-recognized and may be represented by the general formula

^L IAN' ^r30

329 ,29

wherein R ²⁹ and R ³⁰ independently represent hydrogen or a hydrocarbyl, such as alkyl, or either occurrence of R ²⁹ taken together with R ³⁰ and the intervening atom(s) complete a heterocycle having from 4 to 8 atoms in the ring structure.

The term“substituted”, as used herein, refers to moieties having substituents replacing one or more hydrogens on one or more carbons of the backbone. It will be understood that“substitution” or“substituted with” includes the implicit proviso that such substitution is in accordance with permitted valence of the substituted atom and the substituent, and that the substitution results in a stable compound, e.g., which does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc. As used herein, the term“substituted” is contemplated to include all permissible substituents of organic compounds. In a broad aspect, the permissible substituents include acyclic and cyclic, branched and unbranched, carbocyclic and heterocyclic, aromatic and non-aromatic substituents of organic compounds. The permissible substituents can be one or more and the same or different for appropriate organic compounds. For purposes of this application, the heteroatoms such as nitrogen may have hydrogen substituents and/or any permissible substituents of organic compounds described herein which satisfy the valences of the heteroatoms. In some embodiments,“substituted” means that the specified group or moiety bears one, two, or three substituents. In other embodiments,“substituted” means that the specified group or moiety bears one or two substituents. In still other embodiments, “substituted” refers to the specified group or moiety bears one substituent.

Substituents can include any substituents described herein, e.g., a lower alkyl (such as Ci-6 alkyl, e.g., -methyl, -ethyl, and -propyl), a halogen, a hydroxyl, a carbonyl (such as a carboxyl, an alkoxycarbonyl, a formyl, or an acyl), a thiocarbonyl (such as a thioester, a thioacetate, or a thioformate), an alkoxyl, a phosphoryl, a phosphate, a phosphonate, a phosphinate, an amino, an amido, an amidine, an imine, a cyano, a nitro, an azido, a sulfhydryl, an alkylthio, a sulfate, a sulfonate, a sulfamoyl, a sulfonamido, a sulfonyl, a heterocyclyl, an aralkyl, or an aromatic or heteroaromatic moiety. It will be understood by those skilled in the art that substituents can themselves be substituted, if appropriate.

Unless specifically stated as“unsubstituted,” references to chemical moieties herein are understood to include substituted variants. E.g., reference to an“aryl” group or moiety implicitly includes both substituted and unsubstituted variants. The term“unsubstituted” refers to that the specified group bears no substituents.

The term“optionally substituted”, as used herein, means that substitution is optional and therefore it is possible for the designated atom or moiety to be unsubstituted.

Any disubstituent referred to herein is meant to encompass the various attachment possibilities when more than one of such possibilities are allowed. E.g., reference to disubstituent -A-B-, where A ¹ B, refers herein to such disubstituent with A attached to a first substituted member and B attached to a second substituted member, and it also refers to such disubstituent with A attached to the second substituted member and B attached to the first substituted member.

“Protecting group”, as used herein, refers to a group of atoms that, when attached to a reactive functional group in a molecule, mask, reduce or prevent the reactivity of the iunctional group. Typically, a protecting group may be selectively removed as desired during the course of a synthesis. Examples of protecting groups can be found in Greene and Wuts, Protective Groups in Organic Chemistry, 3 ^rd Ed., 1999, John Wiley & Sons, NY and Harrison et al, Compendium of Synthetic Organic Methods , Vols. 1-8, 1971-1996, John Wiley & Sons, NY. Representative nitrogen protecting groups include, but are not limited to, formyl, acetyl, trifluoroacetyl, benzyl, benzyloxycarbonyl (“CBZ”), tert-butoxycarbonyl (“Boc”), trimethylsilyl (“TMS”), 2-trimethylsilyl-ethanesulfonyl (“TES”), trityl and substituted trityl groups, allyloxycarbonyl, 9-fluorenylmethyloxycarbonyl (“FMOC”), nitro- veratryloxycarbonyl (“NVOC”) and the like. Representative hydroxy lprotecting groups include, but are not limited to, those where the hydroxyl group is either acylated (esterified) or alkylated such as benzyl and trityl ethers, as well as alkyl ethers, tetrahydropyranyl ethers, trialkylsilyl ethers (e.g., TMS or TIPS groups), glycol ethers, such as ethylene glycol and propylene glycol derivatives and allyl ethers.

The term "pharmaceutically acceptable" is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

A“pharmaceutically acceptable salt” is intended to mean a salt of a free acid or base of a compound represented herein that is non-toxic, biologically tolerable, or otherwise biologically suitable for administration to the subject. See, generally, S.M. Berge, et al, “Pharmaceutical Salts,” J. Pharm. Sci., 1977, 66, 1-19. Preferred pharmaceutically acceptable salts are those that are pharmacologically effective and suitable for contact with the tissues of subjects without undue toxicity, irritation, or allergic response. A compound described herein may possess a sufficiently acidic group, a sufficiently basic group, both types of functional groups, or more than one of each type, and accordingly react with a number of inorganic or organic bases, and inorganic and organic acids, to form a pharmaceutically acceptable salt.

For a compound described herein that contains a basic group, such as an amine, a pharmaceutically acceptable salt may be prepared by any suitable method available in the art, e.g., treatment of the free base with an inorganic acid, such as hydrochloric acid,

hydrobromic acid, sulfuric acid, sulfamic acid, nitric acid, boric acid, phosphoric acid, and the like, or with an organic acid, such as acetic acid, phenylacetic acid, propionic acid, stearic acid, lactic acid, ascorbic acid, maleic acid, hydroxymaleic acid, isethionic acid, succinic acid, valeric acid, fumaric acid, malonic acid, pyruvic acid, oxalic acid, glycolic acid, salicylic acid, oleic acid, palmitic acid, lauric acid, a pyranosidyl acid, such as glucuronic acid or galacturonic acid, an alpha-hydroxy acid, such as mandelic acid, citric acid, or tartaric acid, an amino acid, such as aspartic acid or glutamic acid, an aromatic acid, such as benzoic acid, 2-acetoxybenzoic acid, naphthoic acid, or cinnamic acid, a sulfonic acid, such as laurylsulfonic acid, p-toluenesulfonic acid, methanesulfonic acid, or ethanesulfonic acid, or any compatible mixture of acids such as those given as examples herein, and any other acid and mixture thereof that are regarded as equivalents or acceptable substitutes in light of the ordinary level of skill in this technology.

For a compound described herein that contains an acidic group, such as a carboxylic acid group, base addition salts can be prepared by any suitable method available in the art, e.g., treatment of such compound with a sufficient amount of the desired the desired base, either neat or in a suitable inert solvent. Examples of pharmaceutically acceptable base addition salts include, but are not limited to, lithium, sodium, potassium, calcium, ammonium, zinc, or magnesium salt, or other metal salts; organic amino salts, such as, alkyl, dialkyl, trialkyl, or tetra-alkyl ammonium salts.

Other examples of pharmaceutically acceptable salts include, but are not limited to, camsylate, sulfates, pyrosulfates, bisulfates, sulfites, bisulfites, phosphates, monohydrogen- phosphates, dihydrogenphosphates, metaphosphates, pyrophosphates, chlorides, bromides, iodides, acetates, propionates, decanoates, caprylates, acrylates, formates, isobutyrates, caproates, heptanoates, propiolates, oxalates, malonates, succinates, suberates, sebacates, fumarates, maleates, butyne-l,4-dioates, hexyne-l,6-dioates, benzoates, chlorobenzoates, methylbenzoates, dinitrobenzoates, hydroxybenzoates, methoxybenzoates, phthalates, sulfonates, methylsulfonates, propylsulfonates, besylates, xylenesulfonates, naphthalene- 1 - sulfonates, naphthalene-2-sulfonates, phenylacetates, phenylpropionates, phenylbutyrates, citrates, lactates, g-hydroxybutyrates, glycolates, tartrates, and mandelates. Lists of other suitable pharmaceutically acceptable salts are found in Remington's Pharmaceutical Sciences, 17* Edition, Mack Publishing Company, Easton, Pa., 1985.

The neutral forms of the compounds are preferably regenerated by contacting the salt with a base or acid and isolating the parent compound in the conventional manner. The parent form of the compound differs from the various salt forms in certain physical properties, such as solubility in polar solvents, but otherwise the salts are equivalent to the parent form of the compound for the purposes of the present application.

Compounds of the present application (such as compounds of formula II, such as formula Ila; formula IT, such as formula IT a (e.g., compound 2a); or formula V, such as formula V’) can also exist as various“solvates” or“hydrates.” A“hydrate” is a compound that exists in a composition with water molecules. The composition can include water in stoichiometic quantities, such as a monohydrate or a dihydrate, or can include water in random amounts. A“solvate” is a similar composition except that a solvent other that water, such as with methanol, ethanol, dimethylformamide, diethyl ether and the like replaces the water. E.g., methanol or ethanol can form an“alcoholate,"” which can again be stoichiometic or non-stoichiometric. Mixtures of such solvates or hydrates can also be prepared. The source of such solvate or hydrate can be from the solvent of crystallization, inherent in the solvent of preparation or crystallization, or adventitious to such solvent.

The compounds of the application, including their pharmaceutically acceptable salts and prodrugs, can exist as various polymorphs, pseudo-polymorphs, or in amorphous state. The term“polymorph”, as used herein, refers to different crystalline forms of the same compound and other solid state molecular forms including pseudo-polymorphs, such as hydrates, solvates, or salts of the same compound. Different crystalline polymorphs have different crystal structures due to a different packing of molecules in the lattice, as a result of changes in temperature, pressure, or variations in the crystallization process. Polymorphs differ from each other in their physical properties, such as x-ray diffraction characteristics, stability, melting points, solubility, or rates of dissolution in certain solvents. Thus crystalline polymorphic forms are important aspects in the development of suitable dosage forms in pharmaceutical industry.

The present application further embraces isolated compounds according to formula II, such as formula Ila, formula IT, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’. The term“isolated compound” refers to a preparation of a compound of formula II, such as formula Ila, formula IT, such as formula IT a (e.g., compound 2a), or formula V, such as formula V’, or a mixture of compounds according to formula II, such as formula Ila, formula IT, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’, wherein the isolated compound has been separated from the reagents used, and/or byproducts formed, in the synthesis of the compound or compounds.“Isolated” does not mean that the preparation is technically pure (homogeneous), but it is sufficiently pure to compound in a form in which it can be used therapeutically. Preferably an“isolated compound” refers to a preparation of a compound of formula II, such as formula Ila, formula IT, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’, or a mixture of compounds according to formula II, such as formula Ila, formula IT, such as formula IT a (e.g., compound 2a), or formula V, such as formula V’, which contains the named compound or mixture of compounds according to formula II, such as formula Ila, formula IG, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’, in an amount of at least 10 percent by weight of the total weight. Preferably, the preparation contains the named compound or mixture of compounds in an amount of at least 50% by weight of the total weight; more preferably at least 80% by weight of the total weight; and most preferably at least 90%, at least 95% or at least 98% by weight of the total weight of the preparation.

The compounds of the application and intermediates may be isolated from their reaction mixtures and purified by standard techniques such as filtration, liquid-liquid extraction, solid phase extraction, distillation, recrystallization or chromatography, including flash column chromatography, or HPLC.

Isomerism and Tautomerism in Described Compounds

Tautomerism

Within the present application it is to be understood that a compound described herein or a salt thereof may exhibit the phenomenon of tautomerism whereby two chemical compounds that are capable of facile interconversion by exchanging a hydrogen atom between two atoms, to either of which it forms a covalent bond. Since the tautomeric compounds exist in mobile equilibrium with each other they may be regarded as different isomeric forms of the same compound. It is to be understood that the formulae drawings within this specification can represent only one of the possible tautomeric forms. However, it is also to be understood that the application encompasses any tautomeric form, and is not to be limited merely to any one tautomeric form utilized within the formulae drawings. The formulae drawings within this specification can represent only one of the possible tautomeric forms and it is to be understood that the specification encompasses all possible tautomeric forms of the compounds drawn not just those forms which it has been convenient to show graphically herein. E.g., tautomerism may be exhibited by a pyrazolyl group bonded as indicated by the wavy line. While both substituents would be termed a 4-pyrazolyl group, it is evident that a different nitrogen atom bears the hydrogen atom in each structure.

Such tautomerism can also occur with substituted pyrazoles such as 3 -methyl, 5- methyl, or 3,5-dimethylpyrazoles, and the like. Another example of tautomerism is amido- imido (lactam-lactim when cyclic) tautomerism, such as is seen in heterocyclic compounds bearing a ring oxygen atom adjacent to a ring nitrogen atom. E.g., the equilibrium: o OH

is an example of tautomerism. Accordingly, a structure depicted herein as one tautomer is intended to also include the other tautomer.

Optical Isomerism

It will be understood that when compounds of the present application contain one or more chiral centers, the compounds may exist in, and may be isolated as pure enantiomeric or diastereomeric forms or as racemic mixtures. The present application therefore includes any possible enantiomers, diastereomers, racemates in their pure forms or mixtures thereof, and salts thereof, of the compounds of the application.

The isomers resulting from the presence of a chiral center comprise a pair of non-superimposable isomers that are called“enantiomers.” Single enantiomers of a pure compound are optically active, i.e., they are capable of rotating the plane of plane polarized light. Single enantiomers are designated according to the Cahn-Ingold-Prelog system. The priority of substituents is ranked based on atomic weights, a higher atomic weight, as determined by the systematic procedure, having a higher priority ranking. Once the priority ranking of the four groups is determined, the molecule is oriented so that the lowest ranking group is pointed away from the viewer. Then, if the descending rank order of the other groups proceeds clockwise, the molecule is designated ( R ) and if the descending rank of the other groups proceeds counterclockwise, the molecule is designated (S). In the example in Scheme 14, the Cahn-Ingold-Prelog ranking is A > B > C > D. The lowest ranking atom, D is oriented away from the viewer.

(R) configuration (S) configuration

Isolated optical isomers may be purified from racemic mixtures by well-known chiral separation techniques, such as but not limited to, normal and reverse phase chromatography, and crystallization. According to one such method, a racemic mixture of a compound of the application, or a chiral intermediate thereof, is separated using a chiral salt or carried out on a Chiralcell OD column. The column is operated according to the manufacturer’s instructions.

Isolated optical isomers (enantiomerically pure compounds) can also be prepared by the use of chiral intermediates or catalysts in synthesis. When a chiral synthetic intermediate is used, the optical center (chiral center) can be preserved without racemization throughout the remainder of the preparative procedure, as is well known in the art. Chiral catalyst can be used to impart at least some degree of enantiomeric purity to products of reactions catalyzed by the chiral catalyst. And, in some cases, compounds having at least some degree of enantiomeric enrichment can be obtained by physical processes such as selective

crystallization of salts or complexes formed with chiral adjuvants.

In certain embodiments, compounds of the application may be racemic. In certain embodiments, compounds of the application may be enriched in one enantiomer. For example, a compound of the application may have greater than 30% ee, 40% ee, 50% ee,

60% ee, 70% ee, 80% ee, 90% ee, or even 95% or greater ee.

In certain embodiments, the compound prepared according to any one of the foregoing processes may be enriched to include predominantly one enantiomer (e.g., of formula II, such as formula Ila; formula IF, such as formula IFa (e.g., compound 2a); or formula V, such as formula V’). An enantiomerically enriched mixture may comprise, for example, at least 60 mol percent of one enantiomer, or more preferably at least 75, 90, 95, or even 99 mol percent. In certain embodiments, the compound enriched in one enantiomer is substantially free of the other enantiomer, wherein substantially free means that the substance in question makes up less than 10%, or less than 5%, or less than 4%, or less than 3%, or less than 2%, or less than 1% as compared to the amount of the other enantiomer, e.g., in the composition or compound mixture. For example, if a composition or compound mixture contains 98 grams of a first enantiomer and 2 grams of a second enantiomer, it would be said to contain 98 mol percent of the first enantiomer and only 2% of the second enantiomer.

In certain embodiments, compounds of the application may have more than one stereocenter. In certain such embodiments, compounds of the application may be enriched in one or more diastereomer. For example, a compound of the application may have greater than 30% de, 40% de, 50% de, 60% de, 70% de, 80% de, 90% de, or even 95% or greater de.

In certain embodiments, the compound prepared according to any one of the foregoing processes may be enriched to provide predominantly one diastereomer of a compound (e.g., of formula II, such as formula Ila; formula IF, such as formula IFa (e.g., compound 2a); or formula V, such as formula V’). A diastereomerically enriched mixture may comprise, for example, at least 60 mol percent of one diastereomer, or more preferably at least 75, 90, 95, or even 99 mol percent.

A variety of compounds in the present application may exist in particular geometric or stereoisomeric forms. The present application takes into account all such compounds, including tautomers, cis- and trans-isomers, R- and S-enantiomers, diastereomers, (D)- isomers, (L)-isomers, the racemic mixtures thereof, and other mixtures thereof, as being covered within the scope of this application. All tautomeric forms are encompassed in the present application. Additional asymmetric carbon atoms may be present in a substituent such as an alkyl group. All such isomers, as well as mixtures thereof, are intended to be included in this application, unless the stereochemistry or isomeric form is specifically indicated.

Rotational Isomerism

It is understood that due to chemical properties (i.e., resonance lending some double bond character to the C-N bond) of restricted rotation about the amide bond linkage (as illustrated below) it is possible to observe separate rotamer species and even, under some circumstances, to isolate such species (see below). It is further understood that certain structural elements, including steric bulk or substituents on the amide nitrogen, may enhance the stability of a rotamer to the extent that a compound may be isolated as, and exist indefinitely, as a single stable rotamer. The present application therefore includes any possible stable rotamers of formula (I) which are biologically active in the treatment of cancer or other proliferative disease states.

Regioisomerism

The preferred compounds of the present application have a particular spatial arrangement of substituents on the aromatic rings, which are related to the structure activity relationship demonstrated by the compound class. Often such substitution arrangement is denoted by a numbering system; however, numbering systems are often not consistent between different ring systems. In six-membered aromatic systems, the spatial arrangements are specified by the common nomenclature“para” for 1, 4-substitution,“meta” for

1, 3-substitution and“ortho” for 1, 2-substitution as shown below.

Isotopical Labeline in Described Compounds

The present application further includes all pharmaceutically acceptable isotopically labeled compound [e.g., of formula II, such as formula Ila, formula IT, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’]. An "isotopically" or "radio-labeled" compound is a compound where one or more atoms are replaced or substituted by an atom having an atomic mass or mass number different from the atomic mass or mass number typically found in nature (i.e., naturally occurring). E.g., in certain embodiments, in compounds [e.g., of formula II, such as formula Ila, formula IG, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’], hydrogen atoms are replaced or substituted by one or more deuterium or tritium (e.g., hydrogen atoms on a Ci-6 alkyl or a Ci-6 alkoxy are replaced with deuterium, such as ί/3-methoxy or l,l,2,2-i/ ₄ -3-methylbutyl).

Certain isotopically labeled compounds [e.g., compounds of formula II, such as formula Ila, formula IG, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’,], e.g., those incorporating a radioactive isotope, are useful in drug and/or substrate tissue distribution studies. The radioactive isotopes tritium, i.e., ³ H, and carbon 14, i.e., ¹⁴ C, are particularly useful for this purpose in view of their ease of incorporation and ready means of detection.

Such isotopically labeled compounds are useful in metabolic studies (preferably with ¹⁴ C), reaction kinetic studies (with, for example ² H or ³ H), detection or imaging techniques [such as positron emission tomography (PET) or single-photon emission computed tomography (SPECT)] including drug or substrate tissue distribution assays, or in radioactive treatment of patients. Further, substitution with heavier isotopes such as deuterium (i.e., ² H) may afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life or reduced dosage requirements.

Substitution with positron emitting isotopes, such as ⁿ C, ¹⁸ F, ¹⁵ 0, and ¹³ N, can be useful in Positron Emission Topography (PET) studies for examining substrate receptor occupancy.

Isotopically labeled compounds [e.g., of formula II, such as formula Ila, formula IT, such as formula H’a (e.g., compound 2a), or formula V, such as formula V’] or their corresponding prodrugs can generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying examples using an appropriate isotopically labeled reagent in place of the non-labeled reagent previously employed. Suitable isotopes that may be incorporated in compounds of the present application include but are not limited to isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine, chlorine, and iodine, such as ² H (also written as D for deuterium), ³ H (also written as T for tritium), ⁿ C, ¹³ C, ¹⁴ C, ¹³ N, ¹⁵ N, ¹⁵ 0, ¹⁷ 0, ¹⁸ 0, ¹⁸ F, ³⁵ S, ³⁶ Cl , ⁸² B r, ⁷⁵ Br, ⁷⁶ B r, ⁷⁷ Br, ¹²³ I, ¹²⁴ I, ¹²⁵ 1, ¹³¹ 1, ³¹ P, and ³² P. Isotopically labeled compounds of this application and prodrugs thereof can generally be prepared by carrying out the procedures disclosed in the schemes or in the examples and preparations described below by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent.

Provisos may apply to any of the disclosed categories or embodiments such that specific embodiments or species may be excluded from such categories or embodiments.

In various embodiments, the compound or set of compounds, such as are used in the inventive methods, can be any one of any of the combinations and/or sub-combinations of the above-listed embodiments.

Exemplification

The invention is further defined in the following Examples. It should be understood that the Examples are given by way of illustration only. From the above discussion and the Examples, one skilled in the art can ascertain the essential characteristics of the disclosure, and without departing from the spirit and scope thereof, can make various changes and modifications to adapt the disclosure to various uses and conditions. As a result, the disclosure is not limited by the illustrative examples set forth hereinbelow.

All temperatures are in degrees Celsius (°C) and are uncorrected.

Unless otherwise noted, commercial reagents used in preparing the example compounds were used as received without additional purification.

Unless otherwise noted, the solvents used in preparing the example compounds were commercial anhydrous grades and were used without further drying or purification.

All starting materials are commercially available, unless stated otherwise.

The following abbreviations may be employed herein: ¹³ C NMR: carbon nuclear magnetic resonance; d: doublet; DMF: A,A-dimcthyl formamide; DMSO: dimethyl sulfoxide; EDCI x HC1: 1 -ethyl-3 -(3 -dimethylaminopropyl) carbodiimide hydrochloride; ES:

electrospray; g: gram; h: hour(s); ^X H NMR: proton nuclear magnetic resonance; HPLC: high pressure liquid chromatography; kg: kilogram; L: liter; m: multiplet; M: molar; mL: milliliter; MHz: megahertz; min: minute(s); mmol: millimole; mol: mole; MS: mass spectrometry; NMM: N-methyl-morpholine; ppm: parts per million; s: singlet; 2-MeTHF: 2-methyl- tetrahydrofiiran; br.: broad; Bu: butyl; calcd: calculated; Celite® : brand of diatomaceous earth filtering agent, registered trader of Celite Corporation; d: doublet; dd: doublet of doublet; ddd: doublet of doublet of doublet; dddd: doublet of doublet of doublet of doublet; DABCO: l,4-diazabicyclo[2.2.2]octane; DCE: dichloroethane; DCM: dichloromethane; DIPEA: N-ethyl-N-isopropylpropan-2-amine; DME: dimethyl ether; DMEA: dimethyl ethylamine; dq: doublet of quartet; dt: doublet of triplet; EDC: 1 -ethyl-3 -(3- dimethylaminopropyl) carbodiimide hydrochloride; ESI: electrospray ion source; EtOAc: ethyl acetate; EtOH: ethanol; g: gram; h: hour(s); HBTU: 0-Benzotriazole-N,N,N’,N’- tetramethyl-uronium-hexafluoro-phosphate; HOBT: N-Hydroxybenzotriazole; HRMS: high resolution mass spectrometry; iPrOH: iso-propanol; MeOH: methanol; mg: milligram;

MgS0 ₄ : anhydrous magnesium sulfate (drying agent); MPLC: medium pressure liquid chromatography; MTBE: methyl /er/-butyl ether; NaHCC : sodium bicarbonate; NH ₄ Q: ammonium chloride; q: quartet; quin: quintet; rt: room temperature; sat: saturated; t: triplet; TEA: triethylamine; tBuOH: /er/-butanol; td: triplet of doublet; TFA: trifluoroacetic acid; and THF : tetrahydrofuran.

The 1H NMR spectra were recorded on a Bruker Ultrashield AV3 500 MHz spectrometer fitted with a QCI cryoprobe and operating with Topspin3.5pl5 software or on a Bruker Ultrashield AV3 400 MHz spectrometer fitted with a BBFO probe and operating with Topspin3.5pl5 software. NMR data were processed using either ACD Spectrus Processor 2015 Pack 2 or Mestrenova version 1 l.0.2.Bruker UltraShield Advance 400MHz / 54mm spectrometer and processed with XWIN-NMR version 2.6 software. The chemical shifts (d) are reported in parts-per-million from the deuterated solvent used.

The 13C NMR spectra were recorded on Bruker Ultrashield AV3 500 MHz spectrometer fitted with a QCI cryoprobe and operating with Topspin3.5pl5 software or on a Bruker Ultrashield AV3 400 MHz spectrometer fitted with a BBFO probe and operating with Topspin3.5pl5 software. NMR data were processed using either ACD Spectrus Processor 2015 Pack 2 or Mestrenova version 11.0.2. a Bruker UltraShield Advance l25MHz / 54mm spectrometer and processed with XWIN-NMR version 2.6 software. The chemical shifts (d) are reported in parts-per-million from the deuterated solvent used.

Example 1 : Compound Synthesis

A solution of dibasic Potassium Phosphate (2.18g, 0.125 mol) in water (100 mL) was prepared-Solution A. A solution of monobasic Potassium Phosphate (l.70g, 0.125 mol) in water (100 mL) was prepared-Solution B. 8 mL of Solution A was mixed with 92 mL of Solution B to give 0.125M Potassium Phosphate Buffer. The pH of this solution was adjusted to pH 5.5 with the addition of 5M NaOH dropwise to give Solution C. Magnesium Sulphate (0.0l5g, 1.2 x 10 ⁴ mol) and Beta-Nicotinamide Adenine Dinucleotide Phosphate Disodium Salt (0.045g, 5.7 x 10 ⁵ mol) were added to Solution C to give Solution D

A 250 mL Jacketed Vessel was charged with KRED-P1-H08 (l.OOg), followed by Solution D (lOOmL). The mixture was stirred at 25oC until all the enzyme powder had dissolved (~5 minutes). The resulting solution was warmed to 40 °C. Compound 1 (20.00g, 0.068 mol) was added, followed by Isopropanol (100 mL).

The resulting suspension was stirred at 40 °C under a slow stream of nitrogen. After 24h, HPLC analysis showed that less than 0.5% by area of compound 1 was present. The reaction mixture was heated to 70 °C for l.5h, before being cooled to 25 °C. The cloudy solution produced was fdtered. The jacketed vessel was washed out with a mixture of Isopropanol (60 mL) and Water (60 mL). The vessel washings were then used to wash the fdter pad. The combined filtrate was then concentrated under vacuum on the Rotary Evaporator (to remove isopropanol) to give an oily suspension.

The oily suspension was extracted with Toluene (100 mL), twice. The combined Toluene extracts were dried (Na ₂ S04) and evaporated under reduced pressure to give compound 2a as pale brown oil that crystallised on standing (l8.74g, 93%). 1H NMR (CD3SOCD3), 7.86 (dd, J=2.0, 8.1 Hz, 1H), 7.76 (d, J=2.0 Hz, 1H), 7.56 (d, J=8.l Hz, 1H), 4.65 (d, J=4.3 Hz), 3.48 (m, 1H), 3.00 (s, 2H), 1.85 (m, 2H), 1.56 (m, 2H), 1.35 (m, 4H).

Example 2: Biological Activity

Assays

The level of activity of compounds prepared according to the processes disclosed here can be tested using the following methods:

TR-FRET Assay

The b-secretase enzyme used in the TR-LRET is prepared as follows:

Human BACE1: the cDNA for the soluble part of the human b-Secretasel (AA1-AA460) is cloned using the BACE 1(1 -460)-(AVT)-E c-pGEN-IRES-neo mammalian expression vector. The gene is fused to the Fc domain of IgGl (affinity tag) and stably cloned into HEK 293 cells. Purified sBACE-Fc is stored in -80°C in Tris buffer, pH 9.2 and has a purity of -40%. Human BACE2: the cDNA for the soluble part of the human b-8eoGeίh8e2 (AA1-AA473) is cloned using BACE2(l-473)-(AVT)-Fc-pDESTl2.2 mammalian expression vector. The gene is fused to the Fc domain of IgGl (affinity tag) and stably cloned into HEK 293 cells. Purified sBACE-Fc is stored in -80°C in 50 mM Glycine, 10 mM Tris-HCl, pH 7-8, and has a purity of -70%.

The enzyme (truncated form) is diluted to 6 pg/mL (stock hBacel: l.3mg/mL, hBace2: l.6mg/ml) and the TruPoint BACE1 Substrate to 200 nM (stock 120 uM) in reaction buffer (NaAcetate, chaps, triton c-100, EDTA pH4.5). A multidrop Combi is used for the liquid handling. Enzyme (7 pF) is added to the compound plate (containing 0.8 pL of compound in dimethy lsulphoxide) . The plate is incubated for 10 minutes. Substrate (8 pL) is then added, and the reaction proceeds for 17 minutes at r.t. The reaction is stopped with the addition of Stop solution (5.5 pL, NaOAc, pH 9). Fluorescence is measured on a Pherastar plate reader using HTRF module. The assay is performed in a 384 well polystyrene, black, round bottom, small volume plate (Greiner 784076). The final concentration of the enzyme is 2.7 pg/mL; the final concentration of substrate is 100 nM (Km hBACEl: 250 nM, hBACE2: 350 nM).

The dimethylsulphoxide control, instead of test compound, defines the 100% activity level and 0% activity is defined by a control inhibitor compound (2-amino-6-(3'-methoxybiphenyl- 3 -yl)-3 ,6-dimethyl-5 ,6-dihydropyrimidin-4(3 H)-one, at a final concentration of 50 pM). 5 reference inhibitors with different affinities are used at all screen occasions in dose response. Diluted TR-FRET Assay

Compounds with a high affinity are further tested in a diluted TR-FRET assay, conditions as described above for the TR-FRET assay, but with 50 times less enzyme and a 6.5 h reaction time at r.t. in the dark.

sAPPB Release Assay

SH-SY5Y cells are cultured in DMEM/F-12 with Glutamax, 10% FCS and 1% non- essential amino acids and cryopreserved and stored at -140 °C at a concentration of 7.5- 9.5xl0 ⁶ cells per vial.

Cells are thawed and seeded at a concentration of around 10000 cells/well in

DMEM/F- 12 with Glutamax, 10% FCS and 1 % non-essential amino acids to a 384-well tissue culture treated plate, 30 pL cell susp/well. The cell plates are then incubated for 7-24 h at 37 °C, 5% C0 ₂ .

The cell medium is removed, followed by addition of 50 pL compound diluted in DMEM/F- 12 with Glutamax, 10% FCS, 1 % non-essential amino acids to a final cone of 0.5% DMSO. The compounds are incubated with the cells for 16-17 h (overnight) at 37 °C, 5%

CO2.

Meso Scale Discovery (MSD) plates are used for the detection of sAPP release. MSD bARRb plates are blocked in 1% BSA in Tris wash buffer for 1 h on shake at r.t. and washed 1 time in Tris wash buffer. 20 pL of medium is transferred to the pre-blocked and washed MSD sAPP microplates, and the cell plates are further used in an ATP assay to measure cytotoxicity. The MSD plates are incubated with shaking at r.t. for 2 h and the media discarded. 10 pL detection antibody is added (1 nM) per well followed by incubation with shaking at r.t. for 2 h and then discarded. 35 pL Read Buffer is added per well and the plates are read in a Meso Scale Discovery SECTOR6000 Imager.

ATP assay

As indicated in the bARRb release assay, after transferring 20 pL medium from the cell plates for bARRb detection, the plates are used to analyse cytotoxicity using the

ViaLightTM Plus cell proliferation/cytotoxicity kit from Cambrex Bioscience that measures total cellular ATP. The assay is performed according to the manufacture's protocol. Briefly, 10 uL cell lysis reagent is added per well. The plates are incubated at r.t. for 10 min. Two min after addition of 25 pL reconstituted ViaLightTM Plus ATP reagent, the luminescence is measured in an Envision reader. Tox threshold is a signal below 70% of the control.

Equivalents

Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the compounds and methods of use thereof described herein. Such equivalents are considered to be within the scope of this invention and are covered by the following claims.

The contents of all references, patents and published patent applications cited throughout this Application, as well as their associated figures are hereby incorporated by reference in entirety.