The present invention relates to radiolabeled ligands for 5-hydroxytryptamine-6 (5-HT ₆ ) receptors, useful for the labelling and diagnostic imaging of 5-HT ₆ receptor functionality.

Noninvasive, nuclear imaging techniques can be used to obtain basic and diagnostic information about the physiology and biochemistry of living subjects, including experimental animals, patients and volunteers. These techniques rely on the use of imaging instruments that can detect radiation emitted from radiotracers administered to living subjects. The information obtained can be reconstructed to provide planar and tomographic images which reveal the distribution and/or concentration of the radiotracer as a function of time.

Positron emission tomography (PET) is a noninvasive imaging technique that offers the highest spatial and temporal resolution of all nuclear medicine imaging modalities and has the added advantage that it can allow for true quantitation of tracer concentrations in tissues. The technique involves the use of radiotracers, labelled with positron emitting radionuclides, that are designed to have in vivo properties which permit measurement of parameters regarding the physiology or biochemistry of a variety of processes in living tissue.

Compounds can be labelled with positron or gamma emitting radionuclides. The most commonly used positron emitting radionuclides are ^O, 1 ^N, ^C and ^p, which are accelerator produced and have half lifes of 2, 10, 20 and 110 minutes respectively. The most widely used gamma emmitting radionuclides are ^p, 99mj _Cj 201 j\ _anc | 123|

In vitro studies using 5-HT ₆ receptor antagonists selectively binding with high affinity to recombinant and native 5-HT ₆ receptors have localised 5-HT ₆ receptors almost exclusively in the CNS (W. D. Hirst et al., Br. J. Pharmacol.. 130, 1597-1605 (2000), A. J. Sleight et al., Br. J. Pharmacol.. 124, 556-562 (1998)). As a therapeutic target the 5-HT ₆ receptor has been investigated for a variety of CNS disorders including anxiety, epilepsy, cognitive function, dementia psychosis and affective disorders.

WO 03/080580 discloses a series of quinoline compounds said to be 5-HT ₆ receptor antagonists and claimed to be useful in the treatment of various CNS disorders. When

Example 7 (GSK215083) of WO 03/080580 is radiolabeled it has been found to allow in vivo imaging of 5HT-6 receptors in the brain.

Accordingly the present invention provides a compound of formula (I):

(I)

for use as a 5-HT ₆ ligand, wherein:

Ri is a radiolabeled group incorporating or consisting of a radionuclide selected from ^H, ^{I I} _C , 13 _N , 15 _O 76 _B r, 18 _F> 123,, 125|_ 131 |_ 75 _Br , 76 _Br , 77 _Br , 82 _Br or 211 At and R ₂ is F; or Ri is C _1-4 alkyl or fluoroC _1-4 alkyl and R ₂ is a radiolabeled group incorporating or consisting of a radionuclide selected from the group consisiting of ^H, ¹¹ C, 13|M, ^O ₁ ^F, 123| 125I ₁ 1311, 75 _Bfi 76Br, 77 _Br , 82 _{Br or} 211 _At ; or a pharmaceutically acceptable salt thereof.

In one embodiment R ₁ is [ C] methyl and R ₂ is F or R ₁ is methyl and R ₂ is ¹ T .

In another embodiment a compound of formula (I) is [ ¹¹ C-Λ/-methyl]3-[(3- fluorophenyl)sulfonyl]-8-(4-methyl-1-piperazinyl)quinoline

The present invention also provides a radiopharmaceutical composition which comprises a compound of formula (I) and a pharmaceutically acceptable carrier or excipient.

The present invention also provides the use of a compound of formula (I) for the manufacture of a medicament for the treatment or prophylaxis of anxiety, depression, epilepsy, obsessive compulsive disorders, migraine, cognitive memory disorders (e.g. Alzheimers disease, age related cognitive decline and mild cognitive impairment), Parkinsons Disease, ADHD (Attention Deficit Disorder/Hyperactivity Syndrome), sleep

disorders (including disturbances of Circadian rhythm), feeding disorders such as anorexia and bulimia, panic attacks, withdrawal from drug abuse such as cocaine, ethanol, nicotine and benzodiazepines, schizophrenia (in particular cognitive deficits of schizophrenia), stroke, disorders associated with spinal trauma and/or head injury such as hydrocephalus, Gl (gastrointestinal) disorders such as IBS (Irritable Bowel Syndrome) and obesity.

The present invention further provides a method for labelling 5-HT ₆ receptors in a mammal which comprises administering to a mammal an effective amount of a compound of formula (I).

The present invention also provides a method for diagnostic imaging of 5-HT ₆ receptors in a mammal which comprises administering to a mammal an effective amount of a compound of formula (I).

The present invention also provides a method for diagnostic imaging of tissues expressing 5-HT ₆ receptors in a mammal which comprises administering to a mammal an effective amount of a compound of formula (I).

The present invention also provides a method for diagnostic imaging of the brain in a mammal which comprises administering to a mammal an effective amount of a compound of formula (I).

The present invention further provides a method for the detection or quantification of 5- HT ₆ receptor functionality in mammalian tissue which comprises administering to a mammal in which such detection or quantification is desired an effective amount of a compound of formula (I).

Preferably, in the methods of the present invention the mammal is human.

[ ¹¹ C-Λ/-methyl]3-[(3-fluorophenyl)sulfonyl]-8-(4-methyl- 1-piperazinyl)quinoline ([ ¹¹ C]GSK215083) is a compound of formula (I).

The present invention also relates to a process for the preparation of [ ¹¹ C-/V-methyl] 3-[(3- fluorophenyl)sulfonyl]-8-(4-methyl-1-piperazinyl)quinoline which comprises reacting 3-[(3- fluorophenyl)sulfonyl]-8-(1-piperazinyl)quinoline with [ ¹¹ C]methyl triflate.

Suitable radionuclides that may be incorporated in compounds of formula (I) include: ³ H, 13N ₁ 15O 76 _Br , 18 _F , 123,, 125,, 131 ,, 75 _Br , 76 _Br , 77 _Br , 82 _{Br and} 211 _At . The choice of radionuclide to be incorporated into compounds of formula (I) will depend on the specific analytical or pharmaceutical application. Therefore, for in vitro labelling of 5-HT ₆ receptors and for competition assays compounds that incorporate ³ H 125| _or 77 _{Br W} ould be preferred. For diagnostic and investigative imaging agents, compounds that incorporate a radionuclide selected from 1 ^" O, ^F, 123| _or 76 _{Br are} preferred. Incorporation of a chelating radionuclide may be useful in certain applications.

Radiolabeled analogues of compound (I) may be used in clinical studies to evaluate the role of 5-HT ₆ receptor ligands in a variety of disease areas where 5-HT ₆ receptor ligands are believed to be involved.

Scheme 1 represents a synthetic route towards compounds of formula (I) wherein R _r is a leaving group and R ₂ is F or R1 ' is methyl and R ₂ ' is a leaving group.

Scheme 1

One synthetic route for the synthesis of a compound of formula (II) is shown in Scheme 2.

Scheme 2.

Steps invoved in the preparation of compound (II) are i: Copper(l) iodide, potassium phosphate, ethylene glycol, 3-fluorobenzenethiol, 8O ⁰ C, 3.5 h; ii: iron powder, acetic acid, 60 ⁰ C, 5 h; iii: trifluoroacetic, n-butylnitrite then tetra-n-butylammonium iodide, r.t, 1 h; iv: monomagnesium peroxyphthalate hexahydrate, 40 ⁰ C, 12 h; and v: Pd ₂ (dba) ₃ , 1 ,1 '-bis- diphenylphosphenoferrocene, sodium terfbutoxide, piperazine, 4O ⁰ C, 16 % h.

The starting materials and other reagents are available commercially or can be synthesised by well-known and conventional methods.

Example 1

[ ¹¹ C-N-methyl]-3-(3-Fluoro-benzenesulfonyl)-8-(4-methyl-p iperazin-1-yl)-quinoline ([ ¹¹ C]GSK215083)

The production of [ ¹¹ C]CO ₂ via the ¹⁴ N(p,α) ¹¹ C reaction was carried out by irradiation of a nitrogen target (N ₂ , 99.99%) with 0.5% O ₂ (99.99%) at a 17 MeV cyclotron (General Electric PET-trace). [ ¹¹ C]CH ₃ I was prepared by catalytic reduction (Ni) of [ ¹¹ C]CO ₂ to [ ¹¹ C]CH ₄ followed by gas phase iodination with I ₂ using the PETtrace MeI MicroLab

system(General Electric). Subsequently [ ¹¹ C]CH ₃ I was passed through a quartz tube loaded with silver triflate heated at 195 ⁰ C for conversion to [ ¹¹ C]MeOTf.

The [ ¹¹ C]methyl triflate was delivered to reaction containing compound (II) and 2,2,6,6- tetramethylpiperidine in methanohacetonitrile at room temperature. Following that first step, the reaction mixture was heated at 85°C for 5 min and then injected onto a semi- preparative column for purification. For all radiosyntheses the radiochemical purity of the product was >99%. The radiochemical yield (decay-corrected, related to [ ¹¹ C]methyliodide) ranged between 50 and 60% and the specific activity was >200 GBq/μmol. The average time for synthesis was 40 min from the end of beam (EOB) including HPLC purification and formulation.

The title compound was obtained by methylation of the corresponding desmethyl precursor compound (II) with [ ¹¹ C]MeoTf. The [ ¹¹ C]MeOTf produced was trapped at room temperature into a 1 ml glass container loaded with compound (II) (1 mg) 2,2,6,6- tetramethylpiperidine (10 μl) in MeOH:acetonitrile, 2:1 (300 μl). After trapping the reaction mixture mixture was heated 5 min at 80 ⁰ C and injected onto a semi-preparative column for purification. The title compound was purified on a C18 column (Sphereclone ODS(2) C-18 250 x 7.4 mm); using ACN:70 mM NaH2PO4 (60:40) as mobile phase at a flow rate of 9 ml/min, the title compound eluted at 10.5 min with baseline separation between the product and the starting material. The product fraction collected after 10min was evaporated to dryness, and reformulated in 10 ml of 0.9% NaCI. Quality controls were performed on a Sphereclone ODS(2) C-18 250 x 4.6 mm using ACN:70 mM NaH2PO4 (60:40) as mobile phase at a flow rate of 2 ml/min, the title compound eluted at 3.5 min .

Biological Data

1. In vitro activity

GSK215083 has very high affinity for the human recombinant 5-HT ₆ receptor stably expressed in the HeLa cell line, with a pKj value of 9.82 (competition studies with [ ³ H]LSD as a radioligand) and is selective for the human 5-HT ₆ receptor over other receptors screened to date.

Table 1. Binding Profile (pKi values) of GSK215083

2. 5-HT ₆ localisation in the brain

Yorkshire/Danish Landrace porcines (~ 40 Kg; n=4) were housed singly in thermostatically controlled (20 ⁰ C) and naturally illuminated stalls. The pigs were provided with Soavl® chow (DLG, Denmark) and water ad libitum by trough. Animals were fasted for 24 hours prior to PET examination, with free access to water during this interval. Animals were anaesthetised by induction with ketamine and midazolam (both intramuscular and intravenous (i.v.)) and maintained in deep anaesthesia using isoflurane (1 - 2 %; Abbott, Denmark). The left femoral artery and vein of each animal were surgically cannulated using catheters (Avanti® size 4F-7F). The femoral artery for blood sampling and blood pressure (BP) recordings and the femoral vein for administration of radiolabeled and non- labelled agents. Animals were placed supine in a Siemens ECAT EXACT HR tomograph, with the head immobilised in a custom-made holding device. During the study, blood pH, pCO ₂ and pθ ₂ levels were monitored and maintained within the normal physiological range. In addition, BP and heart rate were recorded throughout the study. At the end of each study day, the animal was terminated via an intravenous dose of sodium pentobarbital (20 ml). [ ¹¹ C]GSK215083 was administered intravenously in the femoral vein as a 1 minute bolus injection. PET scanning and arterial blood sampling was commenced upon start of the radioligand administration.

[ ¹¹ C]GSK215083 readily enters the brain reaching peak regional tissue concentrations at approximately 20min post injection followed by a slow washout from brain regions known to be rich in 5-HT ₆ receptors with highest uptake and retention observed in striatum. The observed rank order of regional brain concentrations was striatum>cortical regions>cerebellum, consistent with reported 5-HT ₆ receptor densities and localisation determined by tissue section autoradiograpgy in animals and man. [ ¹¹ C]GSK215083 concentation was low in cerebellum, a brain region known to possess very low level of 5- HT ₆ receptors.

3. Saturation of 5-HT ₆ receptors in the brain

Four sequential high specific activity iv radioligand ([ ¹¹ C]GSK215083) administrations were performed in same animal, same day. Following a baseline scan, [ ¹¹ C] GSK215083 was coadministered with escalating dose of unlabelled GSK215083 (0.005, 0.05 and 0.5mg/kg). [ ¹⁵ O]CO and [ ¹⁵ O]H2O were administered pre and post administration of GSK215083 to monitor and to correct for changes in cerebral blood volume and changes in cerebral blood flow. Upon injection of [ ¹¹ C]GSK215083, striatum to cerebellum and cortex to cerebellum ratios of 2 to1 and 1.5 to 1 , respectively were reached at 60 min post injection. In contrast, the same ratios decreased to 1.5 to 1 and 1.2 to 1 at 60 min post coinjection with 0.005mg/kg of GSK215083. When 0.05mg/kg of GSK215083 was administered striatum to cerebellum and cortex to cerebellum ratios further decreased to 1.3 to 1 and 1.1 to 1 , respectively. Finally, ratios of 1.1 to 1 and 1 to 1 , respectively were measured 60min post administration of 0.5mg/kg. The dose dependent decrease in [ ¹¹ C]GSK215083 signal in brain regions known to be rich in 5-HT ₆ .