HK1194301B - Radiolabelled glutaminyl cyclase inhibitors - Google Patents

Description

Radiolabeled glutaminyl cyclase inhibitors

Technical Field

The present invention relates to the use of radiolabeled glutaminyl cyclase (QC) inhibitors as imaging agents, in particular, but not exclusively, as medical imaging agents for the detection of neurological disorders. The invention also relates to pharmaceutical compositions comprising said radiolabeled inhibitors, and to methods and kits for detecting neurological disorders.

Background

Glutaminyl cyclase (QC, EC 2.3.2.5) catalyses the intramolecular cyclization of N-terminal glutamine residues to pyroglutamic acid (pGlu)^*) Ammonia is released. In 1963, the first time Messer isolated QC from emulsions of the tropical plant papaya (Carica papaya) (Messer, m.1963nature4874, 1299). After 24 years, the corresponding enzyme activity was found in the animal pituitary (Busby, W.H.J., et al, 1987J Biol Chem)262,8532-. For mammalian QC, for precursors of TRH and GnRH, the conversion of Gln to pGlu by QC can be shown (Busby, W.H.J. et al, 1987J Biol Chem262,8532-8536; Fischer, W.H. and Spiess, J.1987Proc Natl Acad Sci U S A84, 3628-3632). Furthermore, initial QC localization experiments showed co-localization with its putative catalytic product in the bovine pituitary, which further enhanced the proposed function in peptide hormone synthesis (Bockers, T.M. et al, 1995J neuroendicrinol 7, 445-453). In contrast, the physiological function of plant QC is less clear. As regards the enzymes derived from papaya, a role in the plant defense against pathogenic microorganisms has been proposed (El Moussaoui, A. et al, 2001Cell Mol Life Sci58, 556-570). Recently, putative QCs from other plants have been identified by sequence comparison (Dahl, S.W. et al, 2000Protein Expr Purif20, 27-36). However, the physiological function of these enzymes is not yet clear.

QCs known from animals and plants show a strict specificity for L-glutamine located on the N-terminus of the substrate and their kinetic behaviour has been found to follow the Mimmann equation (Pohl, T. et al, 1991Proc Natl Acad Sci US A88,10059-10063; Conssalvo, A.P. et al, 1988Anal biochem175,131-138; Gooloboov, M.Y. et al, 1996Biol Chem Hoppe Seyler 377, 395-398). However, alignment of the primary structure of QC from papaya with that of highly conserved QC from mammals does not show any sequence homology (Dahl, s.w. et al, 2000Protein Expr Purif20, 27-36). While plant QC appears to belong to a new enzyme family (Dahl, s.w. et al, 2000Protein Expr Purif20,27-36), mammalian QC was found to have significant sequence homology to the bacterial aminopeptidases (Bateman, r.c. et al, 2001biochemistry40,11246-11250), concluding that QC from plants and animals has different evolutionary origins.

It was recently demonstrated that recombinant human QC and QC activity from brain extracts catalyze both N-terminal glutaminyl and glutamate cyclisation. The most striking finding is cyclase-catalyzed Glu₁The transformation is advantageously around pH6.0, while Gln₁Conversion to pGlu-derivatives at around 8.0Since inhibition of the production of pGlu-A β -related peptide can be achieved by inhibition of QC activity of recombinant human QC and porcine pituitary extracts, the enzyme QC is a target in the development of drugs for the treatment of Alzheimer's disease.

Alzheimer's Disease (AD) is the most common form of dementia and is an incurable, degenerative disorder. In 2006, 2660 million patients were worldwide. Alzheimer's disease is predicted to occur in 1 of every 85 people worldwide by 2050. Alzheimer's disease is usually diagnosed clinically by patient history, additional relatives history, and clinical observations based on the appearance of characteristic neurological and neuropsychological characteristics, and the absence of other conditions. Intellectual functional assessments, including memory tests, can further characterize the condition of the disease.

Recently, imaging technology has become a valuable tool in the diagnosis of alzheimer's disease. For example, when used as a diagnostic tool, Single Photon Emission Computed Tomography (SPECT) and Positron Emission Tomography (PET) neuroimaging (neuroimaging) can be used to confirm the diagnosis of alzheimer's disease and assessment including mental condition examination. In individuals already suffering from dementia, SPECT appears to be more advantageous than conventional methods using psychiatric testing and medical history analysis in distinguishing alzheimer's disease from other possible causes.

A new technique, called PiB PET, has been developed for directly and clearly imaging β amyloid deposits in vivo (using a tracer that selectively binds to A β deposits.) the use of PiB-PET compounds¹¹C PET scan. Recent studies have shown that PiB-PET predicts, with 86% accuracy, which patients with mild cognitive impairment will develop alzheimer's disease within two years, excluding the possibility of developing alzheimer's disease with 92% accuracy.

A similar PET scanning radiopharmaceutical compound named (E) -4- (2- (6- (2- (2- ([ 2 ]))¹⁸F]-Fluoroethoxy) ethoxy) pyrimidin-3-yl) vinyl) -N-methylaniline (also known as¹⁸F AV-45、florbeta par-fluoro-18 or florbetapir), which contains the more persistent radionuclide fluoro-18, and is tested as a possible diagnostic tool in alzheimer's patients like PiB florbetapir binds to β -like amyloid, but in contrast to PiB's 20 minute radioactive half-life, it has a half-life of 110 minutes due to its use of fluoro-18, it has also been found that a longer half-life allows for significantly more accumulation of tracer in the brain of AD patients, especially in the area known to be associated with β -amyloid deposits.

Accordingly, there is a need for other imaging agents that are capable of diagnosing neurological disorders (e.g., alzheimer's disease).

Drawings

FIG. 1 shows the administration of Compound (I) to two rats^dThe subsequent PET cumulative image (summation image) (0-60 minutes).

FIG. 2 shows administration of Compound (I)^dAfter that, the time-specific activity curve (% dose administered per gram of brain) of the brains of both rats.

Detailed Description

In a first aspect the present invention provides a radiolabeled glutaminyl cyclase (QC) inhibitor for use as an imaging agent.

Reference herein to "radiolabeled" includes compounds wherein 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 usually found in nature (i.e., naturally occurring). A non-limiting example is¹⁹F, which can detect molecules comprising the element without enriching the element to a higher degree than naturally occurring. Thus, having substituents¹⁹Compounds of F may also be referred to as "labeled" and the like. The term "radiolabeled" may be used in conjunction with "isotopically labeled", "isotopically labelled", "detectable isotope" or"radioligand" is used interchangeably.

In one embodiment, the glutaminyl cyclase (QC) inhibitor comprises a single radiolabeled group.

Suitable, non-limiting radiolabel groups include²H (D or deuterium),³H (T or tritium),¹¹C、¹³C、¹⁴C、¹³N、¹⁵N、¹⁵O、¹⁷O、¹⁸O、¹⁸F、³⁵S、³⁶Cl、⁸²Br、⁷⁵Br、⁷⁶Br、⁷⁷Br、¹²³I、¹²⁴I、¹²⁵I and¹³¹I. it is to be understood that isotopically labeled compounds need only enrich the detectable isotope to or above a level detectable by the technique applicable to the particular application, e.g., in use¹¹In the C-labeled detectable compound, the carbon atom in the labeling group of the labeled compound may comprise a proportion of the molecule¹²C or other carbon isotopes. The radionuclide that is added to the radiolabeled compound will depend on the particular application of the radiolabeled compound. For example, for in vitro plaque or receptor labeling and in competitive assays, it is common to add³H、¹⁴C or¹²⁵The compounds of formula I are most useful. For in vivo imaging applications, in general¹¹C、¹³C、¹⁸F、¹⁹F、¹²⁰I、¹²³I、¹³¹I、⁷⁵Br or⁷⁶Br is most useful. In one embodiment, the radiolabel is¹¹C. In another embodiment, the radiolabel is¹⁴C. In another alternative embodiment, the radiolabel is¹³C。

In one embodiment, the glutaminyl cyclase (QC) inhibitor is a compound of formula (I):

or a pharmaceutically acceptable salt, solvate or polymorph thereof, including all tautomers and stereoisomers thereof, wherein:

R¹represents heteroaryl, -carbocyclyl-heteroaryl, -C_2-6Alkenyl heteroaryl, -C_1-6Alkyl heteroaryl or (CH)₂)_aCR⁵R⁶(CH₂)_bHeteroaryl, wherein a and b independently represent an integer from 0 to 5, with the proviso that a + b =0 to 5, and R⁵And R⁶Are alkylene groups, which together with the carbon atom to which they are attached form C₃-C₅A cycloalkyl group;

wherein any of the above heteroaryl groups may be optionally substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

and wherein any of the above carbocyclic groups may optionally be substituted with one or more substituents selected from C_1-4Alkyl, oxo, halogen and C_1-4A radical substitution of alkoxy;

R²representation H, C_1-8Alkyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, -C_1-4Alkylaryl, -C_1-4Alkyl heteroaryl, -C_1-4Alkyl carbocyclylor-C_1-4An alkyl heterocyclic group;

wherein any of the above aryl and heteroaryl groups may be optionally substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, C_1-6alkoxy-C_1-6Alkoxy-, nitro-, halogen, halogeno-C_1-6Alkyl, halo C_1-6Alkoxy, cyano, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -N (C)_1-4Alkyl) (C_1-4Alkyl) -N (C)_1-4Alkyl) (C_1-4Alkyl), -C_1-4alkyl-N (C)_1-4Alkyl) (C_1-4Alkyl), -C_1-4alkoxy-N (C)_1-4Alkyl) (C_1-4Alkyl), -N (C)_3-8Cycloalkyl) (C)_3-8Cycloalkyl), -N (-C)_1-6alkyl-C_1-6Alkoxy) (-C_1-6alkyl-C_1-6Alkoxy), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

and wherein any of the above carbocyclyl and heterocyclyl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, oxo, halogen, -C (O) C_1-6Alkyl and C_1-4Alkoxy groups.

Or R²Represents phenyl substituted by phenyl, phenyl substituted by monocyclic heteroaryl, phenyl substituted by phenoxy, phenyl substituted by heterocyclyl (wherein the heterocyclyl is substituted by phenyl), phenyl substituted by-O-C_1-4Alkyl-heterocyclyl substituted phenyl, benzyloxy substituted phenyl, carbocyclyl substituted phenyl (wherein said carbocyclyl is substituted by heterocyclyl), O-carbocyclyl substituted phenyl, phenyl substituted heterocyclyl, phenyl substituted carbocyclyl, carbocyclyl fused to carbocyclyl, phenyl fused to heterocyclyl, -C_1-4Alkyl (phenyl substituted by phenyl), -C_1-4Alkyl (phenyl substituted by monocyclic heteroaryl), -C_1-4Alkyl (phenyl substituted by monocyclic heterocyclyl), -C_1-4Alkyl (phenyl substituted by-O-carbocyclyl), -C_1-4Alkyl (phenyl substituted by benzyloxy), -C_1-4Alkyl (optionally substituted phenyl fused to an optionally substituted carbocyclyl) or-C_1-4An alkyl group (an optionally substituted phenyl group fused to an optionally substituted heterocyclic group);

wherein any of the above phenyl, benzyloxy, and heteroaryl groups can be optionally substituted with one or more groups selected from C_1-4Alkyl, halogen and C_1-4The substituent of the alkoxy group is replaced by the group,

and wherein any of the above carbocyclyl and heterocyclyl may be optionally substituted with one or more substituents selected from methyl, phenyl, oxo, halogen, hydroxy and C_1-4A radical substitution of alkoxy;

R³represents H, -C_1-4An alkyl or aryl group;

wherein the above aryl groups may optionally be substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

or R²And R³Are linked to form a carbocyclic ring, optionally substituted with one or more C_1-2Alkyl substitution;

or R²And R³Are linked to form a carbocyclic ring fused to a phenyl, wherein the carbocyclic group and/or the phenyl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, halogen and C_1-4A radical substitution of alkoxy;

or R²And R³Are linked to form a carbocyclic ring fused to a monocyclic heteroaryl, wherein the carbocyclic group and/or heteroaryl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, halogen and C_1-4A radical substitution of alkoxy;

x represents C = O, O, S, CR⁷R⁸、-O-CH₂-or-CH₂-CH₂-；

Y represents CHR⁹C = O or C = S;

z represents-N-R⁴O or CHR¹⁰Provided that when X represents O or S, Z must represent CHR¹⁰；

Or X and Z represent two adjacent carbon atoms of a phenyl ring fused at that position and optionally substituted by one or more halogens or C_1-2Alkyl substitution;

R⁴represents H, -C_1-8Alkyl, -C (O) C_1-6Alkyl or-NH₂；

R⁷And R⁸Independently represent H, -C_1-4An alkyl or aryl group;

wherein the above aryl group may be optionally substituted by C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) substituted;

R⁹and R¹⁰Independently represents H or methyl;

with the proviso that the radical-Y-Z-X-represents other than-C (= O) -N (-R)⁴) -C (= O) -or-C (= S) -N (-R)⁴) -a group of-C (= O) -;

the compounds of formula (I) are described in WO 2010/026212A1(Probiodrug AG).

In another embodiment, the compound of formula (I) is 1- (1H-benzo [ d ] imidazol-5-yl) -5- (4-propoxyphenyl) imidazolidin-2-one:

formula (I)^aThe compound is described in WO 2010/026212A1(Probiodrug AG) as example 12.

In another embodiment, the compound of formula (I) is (S) -1- (1H-benzo [ d ] imidazol-5-yl) -5- (4-propoxyphenyl) imidazolidin-2-one:

formula (I)^bThe compound is described in WO 2010/026212A1(Probiodrug AG) as example 14.

In one embodiment, the radiolabeled compound is of formula (I)^cA compound:

in one embodiment, the radiolabeled compound is of formula (I)^dA compound:

in one embodiment, the radiolabeled compound is of formula (I)^eA compound:

in one embodiment, the radiolabeled compound is of formula (I)^fA compound:

in one embodiment, the glutaminyl cyclase (QC) inhibitor is a compound of formula (II):

or a pharmaceutically acceptable salt, solvate or polymorph thereof, including all tautomers and stereoisomers thereof, wherein:

R¹represents-C_1-6Alkyl, -aryl, -C_1-6Alkylaryl, -cycloalkyl, -C_1-6Alkylcycloalkyl, -heteroaryl, -C_1-6Alkylheteroaryl, -heterocyclyl, -C_1-6Alkylheterocyclyl, -cycloalkyl substituted by phenyl, -cycloalkyl substituted by phenoxy, -phenyl substituted by cycloalkyl, -phenyl substituted by phenoxy, -phenyl substituted by phenyl, heterocyclyl substituted by phenyl, heteroaryl substituted by phenyl, phenyl substituted by heterocyclyl, phenyl substituted by heteroaryl, phenyl substituted by-O-cycloalkyl or phenyl substituted by-cycloalkyl-heterocyclyl;

and wherein any of the above aryl, cycloalkyl, heterocyclyl, heteroaryl, phenyl or phenoxy groups may optionally be substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

R²represents-C_1-6Alkyl radical, C_1-6Haloalkyl, -aryl, -C_1-6Alkylaryl, -cycloalkyl, -C_1-6Alkylcycloalkyl, -heteroaryl, -C_1-6Alkylheteroaryl, -heterocyclyl or-C_1-6An alkyl heterocyclic group;

and wherein any of the above aryl, heteroaryl or heterocyclyl groups may be optionally substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

R³is represented by C_1-6Alkyl or C_1-6A haloalkyl group;

n represents an integer selected from 0 to 3; and is

R^aIs represented by C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl groups).

The compounds of formula (II) are described in WO 2011/110613A1(Probiodrug AG).

In another embodiment, the glutaminyl cyclase (QC) inhibitor is a compound of formula (I) or a compound of formula (II) as defined above.

In another embodiment, the compound of formula (II) is 1- (1H-benzo [ d ] imidazol-6-yl) -5- (2, 3-difluorophenyl) -3-methoxy-4-methyl-1H-pyrrol-2 (5H) -one:

formula (II)^aThe compound is described in WO 2011/110613A1(Probiodrug AG) as example 8.

In another embodiment, the compound of formula (II) is (R) -1- (1H-benzo [ d ] imidazol-6-yl) -5- (2, 3-difluorophenyl) -3-methoxy-4-methyl-1H-pyrrol-2 (5H) -one:

formula (II)^bThe compound is described in WO 2011/110613A1(Probiodrug AG) as example 9.

In one embodiment, the radiolabeled compound is of formula (II)^c：

In another embodiment, the radiolabeled compound is of formula (II)^d：

In one embodiment, the glutaminyl cyclase (QC) inhibitor is a compound of formula (III):

or a pharmaceutically acceptable salt, solvate or polymorph thereof, including all tautomers and stereoisomers thereof, wherein:

R¹represents-C_3-8Carbocyclyl-heteroaryl, -C_2-6Alkenyl heteroaryl, -C_1-6Alkyl heteroaryl or (CH)₂)_aCR⁵R⁶(CH₂)_bHeteroaryl, wherein a and b independently represent an integer from 0 to 5, with the proviso that a + b =0 to 5, and R⁵And R⁶Are alkylene groups which, together with the carbon to which they are attached, form C₃-C₅Cycloalkyl or bicyclic heteroaryl;

wherein any of the above heteroaryl groups may be optionally substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

and wherein any of the above carbocyclic groupsOptionally substituted by one or more groups selected from C_1-4Alkyl, oxo, halogen and C_1-4A radical substitution of alkoxy;

R²is represented by C_1-8Alkyl, aryl, heteroaryl, carbocyclyl, heterocyclyl, -C_1-4Alkylaryl, -C_1-4Alkyl heteroaryl, -C_1-4Alkyl carbocyclyl or-C_1-4An alkyl heterocyclic group;

wherein any of the above aryl and heteroaryl groups may be optionally substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

and wherein any of the above carbocyclyl and heterocyclyl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, oxo, halogen and C_1-4A radical substitution of alkoxy;

or R²Represents phenyl substituted by phenyl, phenyl substituted by monocyclic heteroaryl, phenyl substituted by benzyloxy, phenyl fused to carbocyclyl, phenyl fused to heterocyclyl, -C_1-4Alkyl (phenyl substituted by phenyl), -C_1-4Alkyl (phenyl substituted by monocyclic heteroaryl), -C_1-4Alkyl (phenyl substituted by benzyloxy), -C_1-4Alkyl (optionally condensed to an optionally substituted carbocyclic group)Substituted phenyl) or-C_1-4An alkyl group (an optionally substituted phenyl group fused to an optionally substituted heterocyclic group);

wherein any of the above phenyl, benzyloxy, and heteroaryl groups can be optionally substituted with one or more groups selected from C_1-4Alkyl, halogen and C_1-4The substituent of the alkoxy group is replaced by the group,

and wherein any of the above carbocyclyl and heterocyclyl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, oxo, halogen and C_1-4A radical substitution of alkoxy;

R³represents H, -C_1-4An alkyl or aryl group;

wherein the above aryl groups may optionally be substituted by one or more groups selected from C_1-6Alkyl radical, C_2-6Alkenyl radical, C_2-6Alkynyl, C_1-6Haloalkyl, -C_1-6Sulfanyl, -SOC_1-4Alkyl, -SO₂C_1-4Alkyl radical, C_1-6Alkoxy-, -O-C_3-8Cycloalkyl radical, C_3-8Cycloalkyl, -SO₂C_3-8Cycloalkyl, -SOC_3-6Cycloalkyl radical, C_3-6alkenyloxy-C_3-6Alkynyloxy-, -C (O) C_1-6Alkyl, -C (O) OC_1-6Alkyl radical, C_1-6alkoxy-C_1-6Alkyl-, nitro-, halogen-, cyano-, hydroxy, -C (O) OH, -NH₂、-NHC_1-4Alkyl, -N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) N (C)_1-4Alkyl) (C_1-4Alkyl), -C (O) NH₂、-C(O)NH(C_1-4Alkyl) and-C (O) NH (C)_3-10Cycloalkyl) groups;

or R²And R³Are linked to form a carbocyclic ring, optionally substituted with one or more C_1-2Alkyl substitution;

or R²And R³Are linked to form a carbocyclic ring fused to a phenyl, wherein the carbocyclic group and/or the phenyl may optionally be substituted with one or more substituents selected from C_1-4Alkyl, halogen and C_1-4A radical substitution of alkoxy;

or R²And R³Are linked to form a carbocyclic ring fused to a monocyclic heteroaryl, wherein the carbocyclic and/or heteroaryl ring(s) described above may optionally be substituted with one or more substituents selected from C_1-4Alkyl, halogen and C_1-4A radical substitution of alkoxy;

R⁴represents H, -C_1-8Alkyl, -C (O) C_1-6Alkyl or-NH₂；

X represents O or S; and is

Y represents O or S.

The compounds of formula (III) are described in GB patent application No. 1003936.0 (Probiodrug AG).

In one embodiment, the radiolabeled glutaminyl cyclase (QC) inhibitor is a compound of formula (IV):

in one embodiment, the radiolabeled glutaminyl cyclase (QC) inhibitor is a compound of formula (V):

the method of adding the radiolabel to the glutaminyl cyclase (QC) inhibitor may be in accordance with known labelling methods. For example, WO 2010/111303 describes a method of labelling compounds with 18-fluoro isotopes.

For example, compounds of formula (IV) may be prepared according to the method shown in scheme A:

alternatively, the compounds of formula (V) may be prepared according to the method shown in scheme B:

in one embodiment, the inhibitor as defined herein is used as a medical imaging agent. In another embodiment, the inhibitors defined herein are used as medical imaging agents for the detection of neurological disorders.

According to another aspect of the present invention there is provided a pharmaceutical composition comprising a radiolabeled compound as defined herein, including all tautomers and stereoisomers thereof, or a pharmaceutically acceptable salt, solvate or polymorph thereof, and one or more pharmaceutically acceptable excipients.

Pharmaceutically acceptable salts:

in view of the close association between the free compound and the salt or solvate form of said compound, whenever a compound is referred to herein, it is also intended to mean its corresponding salt, solvate or polymorph, provided that this is possible or appropriate.

Salts and solvates and physiologically functional derivatives of medically useful glutaminyl cyclase (QC) inhibitors are those in which the counterion or bound solvent (association solvent) is pharmaceutically acceptable. However, salts and solvates containing non-pharmaceutically acceptable counterions or bound solvents are within the scope of the invention, e.g., as intermediates in the preparation of other compounds and pharmaceutically acceptable salts and solvates thereof.

Suitable salts of the present invention include those formed with organic acids or bases as well as inorganic acids or bases. Pharmaceutically acceptable acid addition salts include those formed with the following acids: hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, citric acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, trifluoroacetic acid, triphenylacetic acid, sulfamic acid, sulfanilic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, malic acid, mandelic acid, glutamic acid, aspartic acid, oxaloacetic acid, methanesulfonic acid, ethanesulfonic acid, arylsulfonic acids (e.g., p-toluenesulfonic acid, benzenesulfonic acid, naphthalenesulfonic acid, or naphthalenedisulfonic acid), salicylic acid, glutaric acid, gluconic acid, tricarballylic acid, cinnamic acid, substituted cinnamic acids (e.g., cinnamic acid substituted with phenyl, methyl, methoxy, or halogen, including 4-methylcinnamic acid and 4-methoxycinnamic acid), ascorbic acid, oleic acid, naphthoic acid, hydroxynaphthoic acid (e.g., 1-or 3-hydroxy-2-naphthoic acid), naphthoic acid (e.g., naphthalene-2-acrylic acid), Benzoic acid, 4-methoxybenzoic acid, 2-or 4-hydroxybenzoic acid, 4-chlorobenzoic acid, 4-phenylbenzoic acid, phenylacrylic acid (e.g. 1, 4-benzenediacrylic acid), isethionic acid, perchloric acid, propionic acid, glycolic acid, isethionic acid, pamoic acid, cyclamic acid, salicylic acid, saccharinic acid and trifluoroacetic acid. Pharmaceutically acceptable base salts include ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, and salts with organic bases such as dicyclohexylamine and N-methyl-D-glucamine.

The scope of the present invention is intended to include all pharmaceutically acceptable acid addition salt forms of the compounds of the present invention.

Polymorphic crystalline form:

furthermore, certain crystalline forms of the compounds may exist as polymorphs and as such are intended to be included in the present invention. In addition, certain compounds may form solvates with water (i.e., hydrates) or common organic solvents, and such solvates are also intended to be included within the scope of the present invention. The compounds, including their salts, may also be obtained in the form of their hydrates or include other solvents used to crystallize them.

Pharmaceutically acceptable excipients:

thus, for liquid oral preparations such as suspensions, elixirs and solutions, suitable carriers and additives may advantageously include water, glycols, oils, alcohols, flavoring agents, preservatives, coloring agents and the like; for solid oral formulations such as powders, capsules, soft capsules and tablets, suitable carriers and additives include starches, sugars, diluents, granulating agents, lubricants, binders, disintegrating agents and the like.

Carriers that may be added to the mixture include necessary and inert pharmaceutical excipients including, but not limited to, suitable binders, suspending agents, lubricants, flavorants (flavanants), sweeteners, preservatives, coating agents, disintegrating agents, dyes and coloring agents.

Soluble polymers as targeted drug carriers may include polyvinylpyrrolidone, pyran copolymer, polyhydroxypropylmethacrylamide phenol (polyhydroxypropylmethacrylamide), polyhydroxyethylaspartamide phenol (polyhydroxyethylaspartamide-phenol), or polyethyleneoxide polylysine substituted with palmitoyl residues. Furthermore, the compounds of the present invention may be coupled to biodegradable polymer types used to achieve controlled release of drugs, such as polylactic acid, poly-caprolactone, polyhydroxybutyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates and crosslinked or amphipathic block copolymers of hydrogels.

Suitable binders include, but are not limited to, starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, and the like.

Disintegrants include, but are not limited to, starch, methylcellulose, agar, bentonite, xanthan gum, and the like.

According to another aspect of the present invention there is provided a pharmaceutical composition as defined herein for use as an imaging agent in the detection of a neurological condition.

Suitable and non-limiting examples of neurological disorders include: mild cognitive impairment, alzheimer's disease, familial dementia of the british type, familial dementia of the danish type, down's syndrome and neurodegeneration in huntington's disease. In a particular embodiment, the neurological disorder is alzheimer's disease.

In one embodiment, the inhibitor or composition of the invention is used for detecting amyloid peptides.

In one embodiment, the inhibitor or composition of the invention is used to detect tau protein of neurofibrillary tangles.

The detection of such amyloid peptides is useful for the detection and quantification of amyloid deposits and/or neurofibrillary tangles in diseases including, but not limited to, mediterranean fever, MuckleWells syndrome, idiopathic myeloma, amyloid polyneuropathy, amyloid cardiomyopathy, systemic senile amyloidosis, amyloid polyneuropathy, hereditary cerebral hemorrhage with amyloidosis, down's syndrome, scrapie, Creutzfeldt-Jacob disease, kuru, Gerstamnn-Straussler-Scheinker syndrome, medullary thyroid carcinoma, isolated atrial amyloidosis, dialysis patients [ beta ] 2-microglobulin amyloidosis, inclusion body myositis, beta 2-amyloid deposits in muscular dystrophy, chronic traumatic encephalopathy (CET), and islet type II diabetes.

The radiolabeled compounds of the invention may be administered by any route known to those skilled in the art. For example, administration can be topical or systemic and accomplished by oral, parenteral, inhalation spray, topical, rectal, inhalation, nasal, buccal, vaginal, or by an implantable reservoir. The term "parenteral" as used herein includes subcutaneous, intravenous, intraarterial, intramuscular, intraperitoneal, intrathecal, intraventricular, intrasternal, intracranial and intraosseous injection and infusion techniques.

Dosage levels may range from about 0.001 μ g/kg/day to about 10,000 mg/kg/day. In one embodiment, the dosage level is from about 0.001 μ g/kg/day to about 10 g/kg/day. In another embodiment, the dosage level is from about 0.01 μ g/kg/day to about 1.0 g/kg/day. In another embodiment, the dosage level is from about 0.1 mg/kg/day to about 100 mg/kg/day.

The exact dosage regimen and dosage level will depend upon a variety of factors including the age, weight, general health, sex, and diet of the patient; determination of specific dosing regimens is well known to those of ordinary skill in the art. The regimen may include pretreatment and/or simultaneous administration with other compounds (e.g., therapeutic agents).

Another aspect of the invention provides a method for imaging and detecting senile plaques and/or neurofibrillary tangles in brain tissue, the method comprising treating the tissue with an inhibitor as defined herein to detect a neurological disorder.

In one embodiment, the neurological disorder is detected by measuring the affinity of an inhibitor as defined herein for age spots.

In one embodiment, the neurological disorder is detected by measuring the affinity of an inhibitor as defined herein for tau aggregates.

In another aspect of the invention there is provided a method of detecting amyloid deposits in brain tissue ex vivo or in vitro comprising treating the tissue with an inhibitor as defined herein to detect said amyloid deposits.

In another aspect, the invention provides a method of detecting amyloid deposits in a patient in vivo, said method comprising administering to said patient an effective amount of an inhibitor as defined herein, and detecting the level of binding of the compound to amyloid deposits in said patient.

Another aspect of the invention provides a method of detecting tau protein in brain tissue ex vivo or in vitro, the method comprising treating the tissue with an inhibitor as defined herein to detect neurofibrillary tangles.

In a further aspect the invention provides a method of detecting neurofibrillary tangles in a patient in vivo, said method comprising administering to said patient an effective amount of an inhibitor as defined herein and detecting the level of binding of the compound to tau protein.

In one embodiment, the method involves detecting senile plaques and neurofibrillary tangles characteristic of a neurological disorder.

In one embodiment, the detection is performed using gamma imaging, magnetic resonance spectroscopy, or fluorescence spectroscopy.

In one embodiment, the gamma imaging used for detection is PET or SPECT. Positron Emission Tomography (PET) is an accurate and complex technique using isotopes produced in a cyclotron. Positron emitting radionuclides are typically introduced by injection and accumulate in the target tissue. As it decays, it emits a positron that rapidly combines with a nearby electron, resulting in the simultaneous emission of two identifiable gamma rays in opposite directions. These are detected by the PET camera and their origin is very accurately indicated. Since the demonstration that PET is the most accurate non-invasive method for detecting and evaluating most cancers, the most important clinical use of PET is in oncology (fluorine-18 as a tracer). It is also used for cardiac and brain imaging.

Many medical diagnostic methods, including PET and SPECT with radiolabeled compounds, are well known in the art. PET and SPECT are very sensitive techniques and require small amounts of radiolabeled compounds (known as tracers). The transport, accumulation and transformation of the labeled compound in vivo is the same as for the corresponding nonradioactive compound. The tracer or probe may be a radionuclide for PET imaging (e.g. as described above)¹¹C、¹³N、¹⁵O、¹⁸F、⁶⁴Cu and¹²⁴I) radiolabels, or radionuclides suitable for use in SPECT imaging (e.g. using⁹⁹Tc、⁷⁷Br、⁶¹Cu、¹⁵³Gd、¹²³I、¹²⁵I、¹³¹I and³²p) radiolabelling.

PET produces images based on the distribution of molecular imaging tracers with positron emitting isotopes within the tissues of a patient. The PET method has the potential to detect malfunctions of the investigated tissue or organ at the cellular level. PET has been used in clinical oncology, such as imaging of tumors and metastases, and has been used in the diagnosis of certain brain diseases, as well as in the study of brain and heart function. Similarly, SPECT can be used to supplement any gamma imaging study when true 3D characterization (3D representation) can help, such as in imaging of tumors, infections (leukocytes), thyroid, or bone.

The person skilled in the art is familiar with various methods for detecting labeled compounds for imaging purposes. For example, a radiolabeled compound may be detected using Positron Emission Tomography (PET) or Single Photon Emission Computed Tomography (SPECT). The label incorporated into the compound may depend on the desired detection method. PET is well known to those skilled in the art for detecting positron-emitting atoms, such as F. The invention also relates to specific compounds described herein wherein the F atom is replaced with a non-radiolabeled fluorine atom. SPECT is well known to those skilled in the art for detecting photon-emitting atoms, e.g.¹²³I or⁹⁹Tc。

The radiolabeled glutaminyl cyclase inhibitors of the invention should generally have sufficient radioactivity and radioactive concentration to ensure diagnostic reliability. Imaging of amyloid deposits and imaging of neurofibrillary tangles can also be performed quantitatively, such that the amount of amyloid deposits and neurofibrillary tangles can be determined.

An important prerequisite for in vivo imaging agents in the brain is the ability to cross the intact blood brain barrier following rapid intravenous injection. In the first step of the imaging method, a radiolabeled glutaminyl cyclase inhibitor of the invention is introduced into a tissue or a patient in a detectable amount. Typically, the compound is part of a pharmaceutical composition and is administered to the tissue or the patient by methods well known to those skilled in the art.

In another embodiment, a radiolabeled glutaminyl cyclase inhibitor of the invention is introduced into a patient in a detectable amount and after a time sufficient for the compound to bind to amyloid deposits and/or tau protein, the labeled compound is detected non-invasively. In another embodiment of the invention, a radiolabeled glutaminyl cyclase inhibitor of the invention is introduced into a patient, after a time sufficient for the compound to bind to amyloid deposits, a tissue sample is taken from the patient, and the radiolabeled compound is detected in the tissue outside the patient. In another embodiment of the invention, a tissue sample is taken from a patient and a radiolabeled glutaminyl cyclase inhibitor of the invention is introduced into the tissue sample. After a time sufficient for the compound to bind to amyloid deposits and/or tau protein, the compound is detected.

The detectable amount is the amount of labeled compound required to be detected by the detection method chosen. The amount of radiolabeled glutaminyl cyclase inhibitor of the invention to be introduced into a patient for detection can be readily determined by one skilled in the art. For example, an increased amount of a radiolabeled compound may be administered to a patient until the compound is detected by the selected detection method. A label is introduced to a compound to render the compound detectable.

The time required can be readily determined by introducing a detectable amount of a radiolabeled glutaminyl cyclase inhibitor of the invention to a patient and then detecting the radiolabeled compound at various times after administration.

Another aspect of the invention provides a kit for diagnosing a neurological disorder comprising a pharmaceutical composition as defined herein and instructions for using said kit in accordance with the methods described herein.

Examples

Example 1

[ benzimidazole-2- ¹⁴ C]Formula (I) ^b Compound (I) ^c ) Preparation of

Intermediate 1

To 5-amino 2-¹⁴C]Benzimidazole dihydrochloride (1.30g,6.27mmol,375mCi) was added water (10ml) followed by 2M sodium hydroxide solution (6.3ml,12.60 mmol). The mixture was stirred at room temperature for 5 minutes, and then the solvent was removed under reduced pressure. Acetic acid (6.2ml) was added to the residue, and the slurry was stirred at room temperature. 4-Propoxybenzaldehyde (935mg,5.69mmol) was then added dropwise over 15 minutes. And trimethylsilyl cyanide (846mg,8.52mmol) was added dropwise over 15 minutes, and the reaction mixture was stirred at room temperature under a nitrogen atmosphere for 3 hours.

The reaction mixture was added to an ice-cold 28% ammonium hydroxide solution (15ml) with stirring. The product was extracted into ethyl acetate (3 × 20ml) and the extracts combined. Dry over sodium sulfate, then filter the slurry and remove the solvent under reduced pressure. The product was purified by flash chromatography and the desired fractions were combined. The solvent was removed under reduced pressure and the residual solid was pumped dry under vacuum to constant weight to give the title compound (1.67g,5.22mmol,312 mCi).

Intermediate 2

To intermediate 1(267mg,0.84mmol,50.0mCi) was added a slurry of 10% palladium on carbon (Degussa E101R/W form, 51mg) in acetic acid (3ml) under a nitrogen atmosphere. The mixture was stirred at room temperature under hydrogen for 18 hours.

The catalyst was removed by filtration through a pad of Celite (Celite), then washed with acetic acid (10 ml). The filtrate was evaporated to dryness under reduced pressure, and toluene (20ml) was added to the residue. The solvent was removed under reduced pressure to give the title compound (0.75mmol, equivalent to 45 mCi).

[ benzimidazole-2-¹⁴C]Formula (I)^aCompound (I)

To intermediate 2(0.75mmol,45mCi) was added tetrahydrofuran (2.8ml), triethylamine (227mg,2.25mmol) and 1, 1-carbonyldiimidazole (146mg,0.90 mmol). The reaction mixture was stirred at 85 ℃ for 2 hours.

It was cooled to room temperature then water (15ml) was added and the product extracted into ethyl acetate (3 × 20 ml). The extracts were combined, washed with saturated sodium chloride solution (10ml) and then dried over sodium sulfate. The slurry was filtered and the solvent was removed under reduced pressure.

The product was purified by reverse phase high performance liquid chromatography. The desired fractions were combined and the organic solvent was removed under reduced pressure. To the remaining aqueous phase was added saturated sodium chloride solution (15ml) and the product was extracted into ethyl acetate (2 × 15 ml). The extracts were combined and the solvent was removed under reduced pressure. The title compound (0.098mmol, corresponding to 5.9mCi) was obtained.

[ benzimidazole-2-¹⁴C]Formula (I)^bCompound (I)

Reacting [ benzimidazole-2-¹⁴C]Formula (I)^aThe compound (0.098mmol, corresponding to 5.9mCi) was dissolved in n-heptane ethanol methanol diethylamine (500:250:250:5;5ml) and the isomers were resolved by chiral high performance liquid chromatography using a Pirkle Whelk column.

The desired fractions were combined and the solvent was removed under reduced pressure. The remaining residue was dissolved in acetonitrile: water (33:66;5ml) and then lyophilized to give a solid which was vacuum dried to constant weight. The title compound (14.0mg,0.0415mmol,2.49mCi) was obtained.

The technical parameters are as follows:

specific activity

Determined by:

mass spectrometry 61mCi/mmol 2.26GBq/mmol

Gravimetric 178. mu. Ci/mg 6.59MBq/mg

Corresponding to 60mCi/mmol 2.22GBq/mmol

Molecular weight (for the specific activity) 338.3

Radiochemical purity (HPLC) 99.9%

Column Phenomenex Luna C18(2)150x4.6mm

Temperature of room temperature

Solvent A0.05% trifluoroacetic acid in water

Solvent B0.05% trifluoroacetic acid in acetonitrile

Gradient time (min) 015202130

%B 0 100 100 0 0

Flow rate 1.0 ml/min

Ultraviolet detection at 254nm

Chemical purity (HPLC) 99.0%

Column Phenomenex Luna C18(2)150x4.6mm

Temperature of room temperature

Solvent A0.05% trifluoroacetic acid in water

Solvent B0.05% trifluoroacetic acid in acetonitrile

Gradient time (min) 015202130

%B 0 100 100 0 0

Flow rate 1.0 ml/min

Ultraviolet detection at 254nm

Chiral purity (HPLC) >99.9%

Column, Regis Pirkle Whelk02(R, R)250x4.6mm10 μm

Temperature of room temperature

Solvent n-heptane ethanol methanol diethylamine (50:25:25:0.5)

Gradient of equal degree for 30 min

Flow rate 1.0 ml/min

Example 2

[ benzimidazole-2- ¹¹ C]Formula (I) ^b Compound (I) ^d ) Preparation of

At-20 deg.C, mixing¹¹C]CO₂Mu.l THF and 50. mu.l LiEt were added to the reaction vessel₃In BH. After 40 seconds of reaction, 500. mu. l H were added₂And O is hydrolyzed. To obtain [11 ] as a reaction product^C]HCOOH。

Then (S) -1- (3, 4-diaminophenyl) -5- (4-propoxyphenyl) imidazolidin-2-one (1mg in 300. mu.l of 2N aqueous HCl) was added. After 10 minutes of reaction at 140 ℃, the reaction mixture was cooled and the product was purified by HPLC:

column Chromolith Performance RP-18end clamped 100-4,6mm monolithic HPLC-column (MERCK)

Solvent 16% acetonitrile in water (0.1% TFA)

Flow rate 6 ml/min

Retention time (S) -1- (3, 4-diaminophenyl) -5- (4-propoxyphenyl) imidazolidin-2-one 3-7 minutes; compound (I)^d8-9.5 minutes

Will comprise compound (I)^dThe product peak of (2) was collected into 100ml of H₂O, and for further purification it was loaded onto SepPak tc18 column. The SepPak tc18 column was washed with 10ml of water. The compound (I) was then eluted with 3ml of ethanol^d. The product was then dried at 96 ℃ under an argon atmosphere.

Adding NaCl, compound (I)^dDissolve in 100 μ l ethanol (10% ethanol maximum final concentration) to give the final tracer solution.

Specific activity 35,7 GBq/. mu.mol

Stability of the final tracer solution after 1.5 hours at room temperature >98% (n =6)

The technical parameters are as follows:

analytical HPLC

HPLC:Agilent HP1200DAD incl.Autosampler and Raytest RA detector(BGOcell)

Column Chromolith Performance RP-18 endclamped 100-4,6mm monolithic HPLC-column (MERCK)

Solvent A0.1% aqueous TFA

B is acetonitrile

Flow rate 1 ml/min

Gradient 0-10 min 15-20% B

20-50% of B in 10-24 min

50-95% of B in 24-26 min

26-27 minutes, 95% B

27-28 min 15% B

28-30 min 15% B

Balancing 8 min 15% B (before sample introduction)

Ultraviolet detection at 225nm

Analytical HPLC-chiral method

HPLC:Agilent HP1100DAD incl.Raytest RA Detector(PET)

Column Chiralcel OD-H (ODH0CE-PA130)4,6X250mm +4,5X10mm incl precolumn

Solvent n-hexane/ethanol 80/20

Flow rate 1 ml/min

Ultraviolet detection at 225nm

Example 3

1- (1H-benzimidazol-5-yl) -5- (4-propoxyphenyl-, [ solution of ] A ¹³ C ₆ ]Imidazolidin-2-one (formula (I) ^e Compound (I) Preparation of

The intermediate 1 is propoxyphenyl-, [2 ]¹³C₆]

Phenol-, [2 ]¹³C₆](1.20g,12.0mmol) was dissolved in DMSO (12 ml). Finely powdered sodium hydroxide (1.9g,48mmol) was added and stirred rapidly at room temperature for 15 minutes. Iodopropane (4.08g,24.0mmol) was then added dropwise over 3 minutes and the reaction mixture was stirred for 30 minutes. The reaction was sampled for minor work-up and analyzed by GC-MS. A single peak at 6.3 min (m/z142) indicated complete reaction, which was worked up by addition of cold water (100 ml). The quenched reaction was extracted with hexane (4 × 25ml), combined, and then washed with waterDilute sodium hydroxide solution and brine wash. The organic extracts were dried over sodium sulfate, filtered, and the solvent removed in vacuo to give the product as a slurry (1.4g,9.9mmol, 82%). In a similar manner, with 1.6g of phenol-, sodium alginate¹³C₆](16mmol) the reaction was repeated to give 1.50g (10.6mmol,66%) of product, which was combined with the preparation described above. The combined title compound was used in the subsequent step without further purification.

The intermediate 2 is 1-bromo-4-propoxybenzene-, [2 ] or a salt thereof¹³C₆]

To intermediate 1(2.76g,19.4mmol) dissolved in acetonitrile (15ml) was added ammonium nitrate (0.15g,1.9mmol,0.1 eq, ACS grade) and stirred for 10 min. N-bromosuccinimide (3.42g,19.2mmol,0.99 eq., recrystallized in water) was added and stirred at room temperature for 30 min. The consumption of starting material was confirmed by GC-MS analysis and showed a new product peak containing bromine at 10.8 minutes (m/z220+ 222). The reaction was quenched with 50ml and 50ml hexane. The aqueous layer was extracted with additional ethyl acetate-hexanes (1:1,4 × 25ml) and the combined organic layers were washed successively with water, brine and then dried over sodium sulfate. Filtration and evaporation of the solvent gave the title compound (4.2g,19mmol,98%) which was used in the subsequent step without further purification.

Intermediate 3: 4-propoxybenzaldehyde-, [ solution of ]¹³C₆]

To intermediate 2(4.0g,18mmol) in dry THF (16ml) was added a solution of n-butyllithium (2.5M in hexane, 10.9ml,27.1mmol,1.5 equiv) at-78 deg.C over 5 minutes under an inert atmosphere. The cold mixture was stirred for an additional 75 minutes. A solution of dry DMF (2.6g,36mmol,2 equiv.) in dry THF (16ml) was slowly added and the reaction stirred for 2.5 h while the cooling bath warmed to 0 ℃. GC-MS analysis then indicated complete reaction and the product was at 11.5 minutes (m/z 170). The reaction was quenched in cold dilute citric acid and extracted with methyl tert-butyl ether-hexane (1:1,4 × 25 ml). The combined extracts were washed with water (2 × 25ml) then brine (25ml) and dried over sodium sulfate. Filtration and evaporation of the solvent gave 3.5g of crude product. The crude product was purified on silica with ethyl acetate-hexane (7.5:92.5) to give 2.83g of the title compound (16.6mmol, 92%).

Intermediate 4: [ (1H-benzimidazol-5-ylamino)]- (4-propoxyphenyl-, [2 ]¹³C₆]) Acetonitrile

Intermediate 3(2.0g,11.8mmol) was added to a solution of 5-aminobenzimidazole (1.73g,13.0mmol,1.1 equiv.) in acetic acid (14ml) and stirred for 15 min. Trimethylsilyl cyanide (2.3ml,1.8g,18mmol) was added dropwise over 15 min, and the resulting dark reaction solution was stirred at room temperature for 3 h. The progress of the reaction was monitored by TLC (methanol-chloroform, 10:90) and MS. The reaction mixture was quenched by the addition of cooled 25% ammonium hydroxide (35 ml). The resulting solid product was retained and dissolved in ethyl acetate and the aqueous mixture was further extracted with ethyl acetate (3 × 25 ml). The combined organic solutions were washed successively with water (2 × 25ml) and brine (25ml), dried over sodium sulfate, filtered and evaporated to give the crude product, which was used in the subsequent step without further purification.

Intermediate 5: N¹- (1H-benzimidazol-5-yl) -1- (4-propoxyphenyl-, [ solution of ] A¹³C₆]) Ethane-1, 2-diamine

The crude product of intermediate 4 is dissolved inHydrogenation in acetic acid (40ml) using Pd-carbon (10%,0.8g) and 40psi of hydrogen was continued for 24 hours. Filtration over celite (celite) and evaporation of the solvent gave 10g of product as a slurry. TLC (methanol-chloroform 10:90, R)_f=0) and MS (+) (m/z317) confirm completion of the reaction. The crude product was purified on a silica column using methanol-dichloromethane-triethylamine (2L,10:90:0.1) followed by methanol-dichloromethane-triethylamine (1.2L,20:80:0.1) to give 2.9g of the title compound (9.2mmol,78%) by two steps.

1- (1H-benzimidazol-5-yl) -5- (4-propoxyphenyl-, [2 ]¹³C₆]Imidazolidin-2-one (formula (I)^eCompound (I)

To a solution of triethylamine (1.28g,12.6mmol,4 equiv.) and 1, 1' -carbonyldiimidazole (CDI,0.77g,4.7mmol,1.5 equiv., previously recrystallized from dry THF) in dry THF (15ml) was added pure intermediate 5(1.00g,3.16mmol) over 5 min. The resulting mixture was heated at 73 ℃ overnight under an inert atmosphere. The reaction mixture was cooled, added to water (50ml) and extracted with ethyl acetate (4 × 25 ml). The combined organic layers were washed with water (2 × 25ml) and brine (25ml) and dried over sodium sulfate. After filtration and evaporation of the solvent, a product was obtained as a syrup (0.7g), which was purified on silica with methanol-dichloromethane (20: 80). After purification, 0.245g of the title compound (TLC: methanol-chloroform (20:80), R) is obtained_f=0.55, reference standard co-migration; MS (+) m/z344/345), and additionally 0.070g of a mixture comprising the title compound.

The reaction was repeated with another 1.7g of intermediate 5(5.4mmol) and using only 1.2 equivalents of CDI (6.5 mol). The second preparation was purified by applying a gradient of methanol-dichloromethane (7:93 to 20:80) over silica to give 0.376g of the title compound as a tan solid. As before, a mixture (0.301g) containing the desired product was obtained. In each purification step, HPLC (Eclipse XDB-C18,4.6x150mm,3.5 μm, a = water-acetonitrile-trifluoroacetic acid (90:10:0.1), B = water-acetonitrile-trifluoroacetic acid (10:90:0.1), 0% B-100% B in 20 minutes, retention time =9.2 minutes) was used to determine fractions containing the desired title compound of higher purity. At this stage of purification, the purity of the combined product was about 90%. The final purification of the title compound was performed by stepwise gradient elution with water-acetonitrile (85:15, 75:25, 67:33) on an Amberchrom CG161m (4x30cm) column. Fractions containing pure product were again determined using RP-HPLC. The pooled fractions were lyophilized overnight. The solid product was then redissolved in methanol-dichloromethane (5:95) and washed with half saturated sodium bicarbonate and brine, thoroughly backwashing all aqueous solutions. The organic layer was dried over sodium sulfate, and the solvent was distilled off with heptane azeotrope to yield 0.317g of the title compound (0.93 mmol).

The technical parameters are as follows:

purity (HPLC)

method-Waters Acquity with ELS detector

Phenomenex Polar RP4.6x150x4μm

A:H₂O

B:MeOH

Time (% of minutes)% A% B

0 95 5

5 5 95

9 5 95

Flow rate 0.6 ml/min

Results are >99%

Retention time 6.43 minutes

Isotope incorporation (mass spectrometry)

Method Agilent MSD1100

Condition ES-API ionization Pattern

Positive polarity

6mM ammonium formate in methanol: water 7:3

As a result, the molecular ion peak at 343 is consistent with the expected labeling and mass ionization methods.

Conclusion Compound (I)^eTotal isotope incorporation>99%M+6。

Example 4

1- (1H-benzo [ d ]]Imidazol-5-yl) -5- (4-hydroxyphenyl-, [2 ] ¹³ C ₆ ]Imidazolidin-2-one (formula (I) ^f Compound (I) Preparation of

Boron tribromide (0.17ml,0.44g,1.8mmol) was added dropwise to a solution of example 3(0.200g,0.58mmol) in dry dichloromethane at-20 ℃ under an inert atmosphere. The bath was then cooled with ice water (0 ℃) and the reaction was stirred for 1 hour with cooling. The reaction was stirred for an additional 1 hour using a room temperature water bath. The reaction was quenched by slow addition of water (18 ml). The organic layer was retained and re-extracted with more water. All clear colorless aqueous layers (pH 3) were combined, cooled to 5 ℃ and basified by addition of 1N sodium hydroxide. The aqueous phase was ice-cooled for 1 hour and centrifuged to give a white precipitate which was washed with cold water and dried with Drierite overnight to give 0.138g of product which required additional purification. Amberchrom CG161m column (2x30cm) using a gradient of water-acetonitrile (10:90 to 50: 50). Fractions were analyzed by RP-HPLC and combined to give two batches of the title compound (0.038g and 0.063 g).

The technical parameters are as follows:

purity (HPLC)

Method for preparing Zorbax Bonus RP4.6x150x5 μm

A:H₂O

B:MeOH

The flow rate is 1.0 ml/min; ultraviolet: 254nm

As a result, 97.4 percent

Retention time 2 batches: 9.88 minutes and 9.5 minutes

Isotope incorporation (mass spectrometry)

Method Agilent MSD1100

Condition ES-API ionization Pattern

Positive polarity

6mM ammonium formate in methanol: water 7:3

As a result, the molecular ion peak at 301 was consistent with the expected labeling and mass ionization methods.

Conclusion Compound (I)^fTotal isotope incorporation>99%M+6。

Example 5

[ benzimidazole-2- ¹⁴ C]Formula (II) ^a Compounds and formula (II) ^b Compound (formula (II) ^c And formula (II) ^d Compound) of Preparation of

Intermediate 1

To 5-amino 2-¹⁴C]Benzimidazole.2 HCl (supplier IOI; catalog number CC-544) (52.2mCi,60mCi/mmol,0.87mmol) in methanolTo the suspension in (2ml) were added potassium carbonate (468mg,3.388mmol) and triethylamine (236. mu.l, 1.694 mmol). The mixture was stirred at 0 ℃ for 1 hour, filtered and rotary evaporated to a brown solid. The brown solid was dissolved in methanol (1ml) and stirred at 0 ℃.2, 3-difluorobenzaldehyde (119mg,0.837mmol) was added thereto. The solution was warmed to room temperature and stirred for 2 hours. The solvent was removed by rotary evaporation to give an oil (52mCi,60mCi/mmol,0.867 mmol).

Intermediate 2

Intermediate 1(52mCi,60mCi/mmol,0.867mmol) was dissolved in 1, 2-dimethoxyethane (5 ml). Diethyl oxaloacetate (183. mu.l, 0.969mmol) was added thereto, and the solution was refluxed at 95 ℃ for 72 hours.

The product was purified by HPLC, eluted on a Gemini C18 column with a 20mM ammonium hydroxide methanol gradient system, and then rotary evaporated to a solid (21.2mCi,60mCi/mmol,0.353 mmol).

Intermediate 3

Intermediate 2(21.2mCi,60mCi/mmol,0.353mmol) was dissolved in concentrated hydrochloric acid (6ml) and refluxed at 110 ℃ for 16 h.

The solid was filtered, suspended in water (10ml) and basified to ph8.1 with saturated sodium bicarbonate. Stirring was continued for 30 min, then the mixture was filtered and rotary evaporated to a solid (16.2mCi,60mCi/mmol,0.27 mmol).

Racemic [ benzimidazole-2-¹⁴C]Formula (II)^aCompound (formula (II)^cCompound (I)

To a stirred solution of intermediate 3(16.2mCi,60mCi/mmol,0.27mmol) in methanol (4ml) was added diisopropylethylamine (53. mu.l, 0.303mmol) followed by (trimethylsilyl) diazomethane (2M in ether, 275. mu.l, 0.55 mmol). After 15 min, an additional portion of (trimethylsilyl) diazomethane (275 μ l,0.55mmol) was added and stirring was continued for 1 h. The solvent was removed by rotary evaporation to give a solid. The solid was then purified by HPLC, eluting on a Gemini C18 column with a 20mM ammonium hydroxide methanol gradient system, and then rotary evaporated to a solid.

Pure isomer [ benzimidazole-2-¹⁴C]Formula (II)^bCompound (formula (II)^dCompound (I)

Racemic compound (II)^cPurification by HPLC was performed on a Chirobiotic TAG column eluting with 40mM ammonium acetate methanol (4: 6). Pure isomer (II)^cFreeze-drying overnight afforded a white solid (1.94mCi,60mCi/mmol,0.032 mol).

The technical parameters are as follows:

specific activity:

mass spectrometry determination of 60mCi/mmol 2.22GBq/mmol

Molecular weight (at this specific activity) 357.3

Radiochemical purity (HPLC)

Column Zorbax Box RP3.5 μm (150x4.6mm)

Solvent A phosphate buffer pH6.0

Solvent B acetonitrile

The temperature is 25 DEG C

Flow rate 1.0 ml/min

Detection homogeneous (homogeneous) radiochemical Detector, DAD225nm

The result was 98.1%

Chiral purity (HPLC)

Column Chirobiotic Tag5 μm (250x4.6mm)

Solvent A40 mM ammonium acetate buffer pH4.0

Solvent B methanol

Gradient 60% B isocratic for 20 min

The temperature is 20 DEG C

Flow rate 1 ml/min

Detection by homogeneous radiochemical Detector, DAD220nm

The result is 98.8 percent

Biological examples

Small animal PET preliminary study in rats

With compounds (I)^dTwo female Sprague-Dawley rats were treated:

rat 1: 109.5MBq of compound (I) dissolved in 500. mu.l of 0.9% NaCl/EtOH (9/1, v/v)^dIntravenous injection into tail vein. Labeled Compound (I)^dThe specific activity of (A) is 23.7 GBq/. mu.mol. Administration of Compound (I) to rat 1^dThe final dose of (2) is 0.009 mg/kg.

Rat 2.29.5 MBq Compound (I)^dAnd 0.57mg of Compound (I) in an unlabeled form^dAdministration was intravenous in the tail vein. Administration of Compound (I) to rat 2^dThe final dose of (2) is 3.8 mg/kg.

PET scanning

The head regions of rat 1 and rat 2 were subjected to a 60 minute dynamic PET scan. Plasma samples were taken from the retroorbital region at the end of the PET scan. The PET cumulative image is shown in fig. 1.

1.5ml of plasma sample was thoroughly mixed with 3.0ml of acetonitrile. After centrifugation, the supernatant was evaporated at 100 ℃ under an argon atmosphere. The dried residue was dissolved in 2ml CH₃CN/0.1% trifluoroacetic acid in water (9/1) (containing 20. mu.l of unlabelled Compound (I)^d(2.3mg/kg)) and radioactivity was determined by HPLC:

chromatographic column Chromolith Performance RP-18end clamped 100-4,6mm monolithic HPLC-column (MERCK)

Solvent 13% acetonitrile in water (0.1% trifluoroacetic acid)

Flow rate 5 ml/min

Ultraviolet detection at 225nm

The time-specific activity graph is shown in fig. 2. After PET scanning, the specific activity concentrations (total radioactivity) in rat brain plasma were 0.27% ID/g (rat 1) and 0.19% ID/g (rat 2).

Claims (13)

1. Use of a radiolabelled compound of formula (I) for the manufacture of a medicament for use as an imaging agent in the detection of a neurological disorder, wherein the radiolabelled compound of formula (I) is of formula (I)^dA compound of the formula (I)^cA compound:

2. the use of claim 1, wherein the medicament further comprises one or more pharmaceutically acceptable excipients.

3. Use according to claim 1, wherein the neurological disorder is a neurodegenerative disorder in mild cognitive impairment, alzheimer's disease, familial dementia of the british type, familial dementia of the danish type, down's syndrome and huntington's disease.

4. The use of claim 1, wherein the medicament is for detecting amyloid peptide.

5. The use of claim 1, wherein the medicament is for detecting tau protein of neurofibrillary tangles.

6. A method of imaging and detecting senile plaques and/or neurofibrillary tangles in brain tissue, the method comprising treating the tissue with a medicament as defined in claim 1 to detect neurological disorders, the method being for non-diagnostic and non-therapeutic purposes.

7. The method of claim 6, wherein the neurological disorder is detected by measuring the affinity of a compound as defined in claim 1 for senile plaques.

8. The method of claim 6, wherein the neurological disorder is detected by measuring the affinity of a compound as defined in claim 1 for tau protein aggregates.

9. A method for ex vivo or in vitro detection of amyloid deposits in brain tissue, said method comprising treating said tissue with a medicament as defined in claim 1, said method being for non-diagnostic and non-therapeutic purposes.

10. A method for ex vivo or in vitro detection of tau protein in brain tissue, said method comprising treating said tissue with a medicament as defined in claim 1, said method being for non-diagnostic and non-therapeutic purposes.

11. A method of detecting neurofibrillary tangles in brain tissue, the method comprising treating the tissue with a drug as defined in claim 1 and detecting the level of binding of the compound to tau protein, the method being for non-diagnostic and non-therapeutic purposes.

12. The method of any one of claims 6-11, wherein the detecting is performed using gamma imaging, magnetic resonance spectroscopy, or fluorescence spectroscopy.

13. The method of claim 12, wherein the detection of gamma imaging is PET or SPECT.