Patent Document

CROSS REFERENCE TO RELATED APPLICATIONS 
     This application is a divisional application of U.S. application Ser. No. 13/517,276, filed on Jun. 20, 2012, now allowed, which is a filing under 35 U.S.C. 371 of international application number PCT/US2010/051891, file Oct. 8, 2010, which claims priority to application No. 61/249,656 filed Oct. 8, 2009, 61/285,239 filed Dec. 10, 2009 and 61/315,507 filed Mar. 19, 2010, the entire disclosure of each of which is hereby incorporated by reference. 
    
    
     FIELD OF THE INVENTION 
     The present invention relates to a diagnostic imaging agent useful for positron emission tomography (PET) imaging as well as to improved hardware for producing such imaging agents. More specifically, the present invention is directed to method of purifying crude [ 18 F]flutemetamol which in turn can then be formulated into Flutemetamol [ 18 F] Injection for the imaging of β-amyloid plaques in the brain and methods and devices for preparing same. More specifically, the present invention is directed to the automated synthesis and purification of [ 18 F]flutemetamol by means of solid phase extraction (SPE). 
     BACKGROUND OF THE INVENTION 
     Flutemetamol [ 18 F] Injection is a diagnostic positron emission tomography (PET) agent for the imaging of β-amyloid plaques in the brain. The synthesis of the agent can be performed using automated synthesis platforms with or without using specially-tailored cassettes. For example, the synthesis can be performed using either the TRACERlab FX F-N platform or the FASTlab™ platform, commercially available from GE Healthcare a division of General Electric Company in conjunction with auxiliary preparative high pressure liquid chromatography equipment. After synthesis, the bulk agent is transferred to high pressure liquid chromatography (HPLC) equipment to separate the physico-chemically similar compounds [ 18 F]flutemetamol from its deprotected precursor, AH111832 (6-hydroxy-2-(4′-(N-methyl)amino-3′-nitro)phenylbenzothiazole) and hence obtain purified [ 18 F]flutemetamol. 
     However there still exists a need in the art for alternative purification methods for the preparation of [ 18 F]flutemetamol. The invention as described below answers such a need. Specifically, Applicants have now found a process that eliminates the use of preparative HPLC equipment. 
     SUMMARY OF THE INVENTION 
     As [ 18 F]flutemetamol and its deprotected precursor, AH111832 (6-hydroxy-2-(4′-(N-methyl)amino-3′-nitro)phenylbenzothiazole) are physico-chemically very similar, preparative HPLC is required to separate them. However, Applicants have now found that it is possible to replace the preparative HPLC equipment in previous purification processes with low cost, single-use solid phase extraction (SPE) cartridges for purification of [ 18 F]flutemetamol. 
     Accordingly, the present invention provides a purification process comprising the following steps: 
     (a) passing a diluted crude product reaction mixture comprising flutemetamol through a first reverse phase SPE cartridge; 
     (b) washing said first reverse phase SPE cartridge with a water/acetonitrile, tetrahydrofuran (THF)/water, methanol (MeOH)/water or isopropanol/water mixture; preferably, a water/acetonitrile mixture; 
     (c) rinsing said first reverse phase SPE cartridge with water once step (b) is completed; 
     (d) eluting said first reverse phase SPE cartridge with acetonitrile or tetrahydrofuran; preferably, acetonitrile; 
     (e) directly passing the mixture from said eluting step (d) through a normal phase SPE cartridge to give an acetonitrile or tetrahydrofuran solution; preferably, an acetonitrile solution, comprising purified flutemetamol; 
     (f) diluting said acetonitrile or tetrahydrofuran solution; preferably, an acetonitrile solution, comprising purified flutemetamol, with water to form a diluted water/acetonitrile or a diluted water/tetrahydrofuran solution; preferably, a diluted water/acetonitrile solution, comprising purified flutemetamol, wherein said water/acetonitrile solution contains about 40-70% (v/v) water; preferably at least about 40% (v/v) water; more preferably at least about 50% (v/v) water; 
     (g) passing the diluted water/acetonitrile or diluted water/tetrahydrofuran solution; preferably, diluted water/acetonitrile solution, comprising purified flutemetamol of step (f) through a second reverse phase SPE cartridge and trapping the flutemetamol on said cartridge second reverse phase SPE cartridge; 
     (h) rinsing said second reverse phase SPE cartridge with water; and 
     (i) eluting the trapped purified flutemetamol from second reverse phase SPE cartridge with an injectable organic solvent; preferably, ethanol or DMSO; preferably with ethanol. 
     According to the invention, the purified flutemetamol can be collected after step (i). 
     The present invention also provides a purification process of the present invention, wherein the process is automated. 
     The present invention also provides a cassette on which a purification process of the present invention can be performed. 
    
    
     
       BRIEF DESCRIPTION OF THE FIGURES 
         FIG. 1  depicts hydrophilic precursor derivatives. 
         FIG. 2  depicts the effect of ambient temperature and acetonitrile concentration on yield. 
         FIG. 3  depicts the effect of ambient temperature and acetonitrile concentration on level of chemical impurities. 
         FIG. 4  depicts a flow chart describing the production and formulation of Flutemetamol ( 18 F) Injection on a cassette of the present invention. 
         FIG. 5  is a picture of a fully assembled cassette of the present invention for the production of Flutemetamol ( 18 F) Injection, showing all tubing and prefilled reagent vials and the SPE cartridge. 
         FIG. 6  shows the numbering of each position of the cassette manifold of the present invention. 
         FIGS. 7 and 8  list the raw materials required and the location of each of the main components on the cassette of the present invention. 
         FIG. 9  depicts an SPE cartridge of the present invention. 
         FIG. 10  depicts deprotected precursor AH111832 (6-hydroxy-2-(4′-(N-methyl)amino-3′-nitro)phenylbenzothiazole). 
         FIG. 11  is an alternate view of the cassette of the present invention. 
     
    
    
     DETAILED DESCRIPTION 
     [ 18 F]Flutemetamol may be prepared by nucleophilic substitution of a nitro group in the precursor AH111907 (6-ethoxymethoxy-2-(4′-(N-formyl-N-methyl)amino-3′-nitro)phenylbenzothiazole) by [ 18 F]fluoride followed by deprotection as illustrated in Scheme 1: 
     
       
                 
         
             
             
         
      
     
     Initial studies carried out on AH111907 (6-ethoxymethoxy-2-(4′-(N-formyl-N-methyl)amino-3′-nitro)phenylbenzothiazole) and (non-radioactive) flutemetamol demonstrated that the former reacted with strong bases to produce less lipophilic species (e.g., hydrophilic precursor derivatives of  FIG. 1 ) while leaving the latter unaffected. Any suitable base may be used. In one embodiment, alkoxide, alkali metal hydroxides, or thiooxide bases can be used. In a further embodiment, the base is selected from the group consisting of sodium hydroxide, potassium hydroxide, sodium hydride, sodium thiomethoxide, sodium ethoxide, and sodium methoxide. In a further embodiment, the base is sodium ethoxide or sodium methoxide. In a further embodiment, the base is sodium methoxide. 
     In one embodiment of the invention, the “crude product reaction mixture comprising flutemetamol” of step (a) is the crude [ 18 F]fluoride substitution reaction mixture comprising (i) flutemetamol, (ii) the hydroxyl- and amino-protected flutemetamol having the following structure: 
                                
and (iii) AH111907 (6-ethoxymethoxy-2-(4′-(N-formyl-N-methyl)amino-3′-nitro)phenylbenzothiazole), each as described herein, and is treated with base at a temperature of about &gt;100° C. followed by treatment with acid. In one embodiment of the invention, base treatment of the “crude product reaction mixture comprising flutemetamol”; preferably, the crude [ 18 F]fluoride substitution reaction mixture is performed at a temperature ranging between about 120-140° C.; more preferably at about 130° C. According to the invention, for the subsequent acid treatment, any mineral acid can be used. Examples of suitable acids include, but are not limited to, sulphuric acid, nitric acid, hydrochloric acid, phosphoric acid, hydrobromic acid (HBr); preferably the acid used is hydrochloric acid. The resulting less lipophilic species are then separable from [ 18 F]flutemetamol using solid phase extraction (SPE) cartridge.
 
     According to the present invention, flutemetamol as used herein can be either unlabelled or radiolabeled flutemetamol. In a preferred embodiment, flutemetamol will be [ 18 F]flutemetamol. [ 18 F]flutemetamol may be prepared by any means known in the art including, but not limited to, the synthesis set forth in Scheme 1 as described herein, to give the “crude product reaction mixture comprising flutemetamol” of step (a). 
     The suitable source of [ 18 F]-fluoride ion ( 18 F − ) can be obtained as an aqueous solution from the nuclear reaction  18 O (p,n) 18 F and is made reactive by the addition of a cationic counterion and the subsequent removal of water. Suitable cationic counterions should possess sufficient solubility within the anhydrous reaction solvent to maintain the solubility of  18 F − . Therefore, counterions that have been used include large but soft metal ions such as rubidium or caesium, potassium complexed with a cryptand such as Kryptofix™, or any tetraalkylammonium salt known in the art. A preferred counterion is tetrabutyl ammonium salt. A more detailed discussion of well-known  18 F labelling techniques can be found in Chapter 6 of the “Handbook of Radiopharmaceuticals” (2003; John Wiley and Sons: M. J. Welch and C. S. Redvanly, Eds.). According to the present invention, the washing step (b) removes hydrophilic compounds including the hydrophilic precursor derivatives produced during the sodium methoxide reaction (see  FIG. 1 ) from the cartridge to waste such that flutemetamol and species of similar hydrophobicity are retained on the first reverse phase SPE cartridge. As would be understood by one of skill in the art, the specific composition of the solvent mixture will depend on the SPE cartridge used. 
     According to the present invention, the normal phase SPE cartridge of step (e) of a process of the invention, each as described herein, serves to retain many of the remaining hydrophilic impurities. Flutemetamol and other hydrophobic compounds pass through the normal phase SPE cartridge with minimal retention. 
     According to the present invention, the rinsing step (h) is performed until residual acetonitrile is present at acceptable levels for injection. 
     According to the invention, the eluting step (i) elutes flutemetamol and compounds of similar hydrophobicity such as residual quantities of its deprotected precursor AH111832 (6-hydroxy-2-(4′-(N-methyl)amino-3′-nitro)phenylbenzothiazole). The purified flutemetamol of step (i) is suitable for formulation. 
     According to the invention, the purified flutemetamol can be collected into any suitable collection vial as would be understood by one of skill in the art. A process of the present invention optionally further comprises the step of further/additional rinsing of said second reverse phase SPE cartridge with water to completely remove any flutemetamol and ethanol on the second reverse phase SPE cartridge for transfer to a collection vial, each as described herein. 
     According to the invention, the reverse phase SPE cartridge can be any reverse phase SPE cartridge known in the art having a chain length longer than C8; preferably longer than C18; most preferably, a C30 SPE cartridge.  FIG. 9  depicts an example of reverse phase SPE cartridge for use in a process of the present invention. The reverse phase SPE cartridge includes a commercially available sorbent packed between two porous media layers within an elongate cartridge body. The cartridge body includes luer fittings for simplified connection. Suitable assembled reverse phase SPE cartridges for use in the present invention can be any assembled reverse phase SPE cartridge known in the art including, but not limited to, those commercially available from Macherey-Nagel GmbH &amp; Co. KG, Neumann-Neander-Strasse 6-8, D-52355 Dueren, Germany. Suitable sorbents for use in a reverse phase SPE cartridge can be any sorbent know in the art including, but not limited to those, commercially available from Princeton Chromatography Inc., Cranbury, N.J. 08512 USA. An example of suitable sorbent is a C30 sorbent. According to the invention, a C30 cartridge is preferred as it provides higher retention capability compared to shorter-chain reverse phase cartridges (C8, C18) and can be used for the separation of flutemetamol from its hydrophilic precursor derivatives. 
     Primary Purification—Reverse Phase SPE 
     In one embodiment of the invention, the first reverse phase SPE cartridge may be a reverse phase SPE cartridge as described herein. In a preferred embodiment of the invention the first reverse phase SPE cartridge is a C30 cartridge. 
     In one embodiment of the invention, the first reverse phase SPE cartridge may optionally be conditioned with acetonitrile followed by water prior to step (a) as described above. 
     In one embodiment of the invention, after step (b), the first reverse phase SPE cartridge may optionally be flushed with nitrogen and/or vacuum. 
     In one embodiment of the invention, in step (b) the sorbent of the first reverse phase SPE cartridge is washed with 40% acetonitrile:60% water (v/v) and then the first reverse phase SPE cartridge is flushed with nitrogen and/or vacuum. 
     In one embodiment of the invention, in step (b) the sorbent of the first reverse phase SPE cartridge is washed with water and then the first reverse phase SPE cartridge is flushed with nitrogen and/or vacuum. 
     In one embodiment of the invention, in step (d) flutemetamol is eluted from the first reverse phase SPE cartridge with 35-45% acetonitrile:water (v/v). 
     In an embodiment of the invention, acceptable yield and purity is obtained by performing these steps in the first reverse phase SPE cartridge at a temperature between about 19° C. and about 34° C.; preferably between about 20° C.-30° C. (i.e., the hot cell ambient temperature in which the first reverse phase SPE cartridge is located) and using an acetonitrile/water mixture where the water concentration is about 35-45% of the total (v/v) (e.g., 40% water+60% acetonitrile); preferably, about 39.5-40.5% of the total (v/v). (See  FIGS. 2 and 3 ). At lower temperatures and lower concentrations of acetonitrile, flutemetamol-related compounds are bound more tightly to the solid phase and are therefore less susceptible to being lost to waste during the acetonitrile/water washing. The result is a high yield of flutemetamol but with a greater level of impurities. The opposite effect is seen at higher temperatures and higher concentrations of acetonitrile. This combination gives higher purity but much lower yield. 
     Secondary Purification—Normal Phase SPE 
     According to the invention, the normal phase SPE cartridge can be any normal phase SPE cartridge known in the art. Examples of suitable normal phase SPE cartridges include, but are not limited to, amino, cyano, diol, alumina, and silica normal phase SPE cartridges. 
     In one embodiment of the invention, a normal phase SPE cartridge will contain normal phase materials such as silica-based amino stationary phase to selectively trap hydrophilic impurities from an acetonitrile solution without also retaining flutemetamol. Any silica-based amino stationary phase (i.e., amino sorbent) known in the art can be used. Examples of suitable “silica-based amino stationary phase” include, but are not limited to, those commercially available from Waters (Milford, Mass., USA). In one embodiment of the invention, during the FASTlab™ process, as described herein, amino cartridges, or normal phase SPE cartridges, (e.g., Varian Bond Elut Jr NH 2  cartridge) can be used on FASTlab™. 
     In one embodiment of the invention, the amino sorbent of the normal phase SPE cartridge of step (e) is first conditioned by passing acetonitrile through the normal phase SPE cartridge and then drying the cartridge under a flow of nitrogen, prior to the flutemetamol/acetonitrile fraction from the first reverse phase SPE cartridge being passed through it. According to the present invention, the normal phase SPE cartridge of step (e) may optionally be further rinsed with acetonitrile in order to maximise flutemetamol recovery prior to step (f). 
     In one embodiment of the invention, the amino sorbent is first conditioned by passing acetonitrile through the normal phase SPE cartridge and then dried under a flow of nitrogen. The flutemetamol/acetonitrile fraction from the first reverse phase SPE cartridge is passed through the amino cartridge and into a FASTlab™ syringe. The amino cartridge is then rinsed with a further acetonitrile to maximise flutemetamol recovery. 
     Solvent Exchange—Second Reverse Phase SPE 
     After the secondary purification by means of the normal phase SPE cartridge, acetonitrile (and any residual methanol) may be removed before the purified drug substance (i.e. flutemetamol) is transferred to the product collection vial. This can be achieved by performing solvent exchange on a second reverse phase SPE cartridge. According to the present invention, the second reverse phase SPE cartridge may be a reverse phase SPE cartridge as described herein. In a preferred embodiment of the invention, the second reverse phase SPE cartridge is a C30 cartridge. 
     In one embodiment of the invention, the second reverse phase SPE cartridge can optionally be pre-conditioned with acetonitrile and water. 
     In one embodiment of the invention, prior to passing through the second reverse phase SPE cartridge, the acetonitrile/flutemetamol product solution from the normal phase SPE/amino cartridge is diluted with water such that the loading solution is below about 50% acetonitrile in order to trap flutemetamol onto the sorbent of the second reverse phase SPE cartridge. 
     In one embodiment of the invention, the second reverse phase SPE cartridge is subsequently rinsed with water to remove residual solvents, before the flutemetamol is eluted from the cartridge into a product collection vial using first ethanol then water. 
     A purification process of the invention can be performed manually. A purification process of the invention can be automated. In a preferred embodiment, a purification process of the invention is performed on an automated system/platform. 
     In a preferred embodiment, a process of the present invention is automated. [ 18 F]flutemetamol may be conveniently prepared in an automated fashion by means of an automated radiosynthesis apparatus. There are several commercially-available examples of such apparatus, including TRACERlab™ and FASTlab™ (both commercially available from GE Healthcare a division of General Electric Company). In a preferred embodiment of the invention, the automated radiosynthesis apparatus is FASTlab™. Automated radiosynthesis apparatus commonly comprises a “cassette”, often disposable, in which the radiochemistry is performed, which is fitted to the apparatus in order to perform a radiosynthesis. The cassette normally includes fluid pathways, a reaction vessel, and ports for receiving reagent vials as well as any solid-phase extraction cartridges used in post-radiosynthetic clean up steps. 
     The present invention therefore provides in another aspect of the present invention, a cassette for the automated synthesis and purification of [ 18 F]flutemetamol each as defined herein comprising: 
     (i) a vessel containing crude product reaction mixture comprising flutemetamol; 
     (ii) a first reverse phase SPE cartridge; 
     (iii) means for washing and eluting the first reverse phase SPE cartridge; 
     (iv) a normal phase SPE cartridge; 
     (v) a second reverse phase SPE cartridge; and 
     (vi) means for rinsing and eluting the second reverse phase SPE cartridge; 
     wherein each component is as described herein. 
     Advantages to such an on-cassette SPE cartridge-based process includes reduction of overall synthesis time and cost as well as improved process reproducibility. 
     Reference is now made to  FIGS. 5-9 and 11 , which depict a disposable synthesis cassette  110  and its components which are useful for performing the method of the present invention.  FIG. 6  depicts the numbering of each position  1 - 25  of the cassette manifold of the present invention, each position also referring to the manifold valve of manifold  112 .  FIG. 7  lists the raw materials required for the cassette of the present invention.  FIG. 8  lists the location of each of the main components on the cassette of the present invention. 
     Cassette  110  includes, a manifold  112  including twenty-five 3 way/3 position stopcocks valves  1 - 25 , respectively. Manifold valves  1 - 25  are also referred to as their manifold positions  1 - 25  respectively. Manifold valves  1 ,  4 - 5 ,  7 - 10 ,  17 - 23 , and  25  have female luer connectors projecting up therefrom. Valves  2 ,  6 , and  12 - 16  have an elongate open vial housing upstanding therefrom and support an upstanding cannula therein for piercing a reagent vial inserted in the respective vial housing. Movement of the reagent vial to be pierced by the respective cannula is performed under actuation by the synthesizer device. Valves  3 ,  11 , and  24  support an elongate open syringe barrel upstanding therefrom. Valves  1 - 25  include three open ports opening to adjacent manifold valves and to their respective luer connectors, cannulas, and syringe barrels. Each valve includes a rotatable stopcock which puts any two of the three associated ports in fluid communication with each other while fluidically isolating the third port. Manifold  112  further includes, at opposing ends thereof, first and second socket connectors  121  and  123 , each defining ports  121   a  and  123   a , respectively. Manifold  112  and the stopcocks of valves  1 - 25  are desirably formed from a polymeric material, e.g. PP, PE, Polysulfone, Ultem, or Peek. 
     Cassette  110  is a variant of a pre-assembled cartridge designed to be adaptable for synthesizing clinical batches of different radiopharmaceuticals with minimal customer installation and connections. Cassette  110  includes reaction vessel, reagent vials, cartridges, filters, syringes, tubings, and connectors for synthesizing a radiotracer according to the present invention. Connections are desirably automatically made to the reagent vials by driving the septums thereof onto penetrating spikes to allow the synthesizer access to use the reagents. 
     Cassette  110  is attachable to a synthesis device, such as FASTLab, which cooperatively engages the cassette so as to be able to actuate each of the stopcocks and syringes to drive a source fluid with a radioisotope through the cassette for performance of a chemical synthesis process. Additionally, the synthesis device can provide heat to the reaction vessel of cassette  110  as required for chemical reactions. The synthesizer is programmed to operate pumps, syringes, valves, heating element, and controls the provision of nitrogen and application of vacuum to the cassette so as to direct the source fluid into mixing with the reagents, performing the chemical reactions, through the appropriate purification cartridges, and selectively pumping the output tracer and waste fluids into appropriate vial receptacles outside the cassette. The fluid collected in the output vial is typically input into another system for either purification and/or dispensement. After product dispensement, the internal components of cassette  110  are typically flushed to remove latent radioactivity from the cassette, although some activity will remain. Cassette  110  thus can be operated to perform a two-step radiosynthesis process. By incorporating SPE cartridges on the manifold, cassette  110  is further able to provide simple purification so as to obviate the need for HPLC. 
       FIGS. 5 and 11  depicts a fully assembled cassette  110  of the present invention for the production of Flutemetamol ( 18 F) Injection, showing all tubing and prefilled reagent vials. Cassette  110  includes a polymeric housing  111  having a planar major front surface  113  and defining a housing cavity  115  in which manifold  112  is supported. A first reverse phase SPE Cartridge  114  is positioned at manifold position  18  while a second reverse phase SPE cartridge  116  is positioned at manifold position  22 . A normal phase (or amino) SPE cartridge  120  is located at manifold position  21 . First SPE Cartridge  114  is used for primary purification. The amino cartridge  120  is used for secondary purification. The second SPE Cartridge  116  is used for solvent exchange. A 50 cm to over-2 m length of Tygon tubing  118  is connected between cassette position  19  and a product collection vial  129  in which occurs the formulation of the drug substance. Tubing  118  is shown in partial phantom line to indicate where is passing behing front surface  113  on the far side of manifold  112  in the view. While some of the tubings of the cassette are, or will be, indentified as being made from a specific material, the present invention contemplates that the tubings employed in cassette  110  may be formed from any suitable polymer and may be of any length as required. Surface  113  of housing  111  defines an aperture  119  through which tubing  118  transits between valve  19  and the product collection vial  139 .  FIG. 11  depicts the same assembled manifold of the cassette and shows the connections to a vial containing a mixture of 40% MeCN and 60% water at manifold position  9 , a vial of 100% MeCN at manifold position  10 , a water vial connected at the spike of manifold position  14 , and a product collection vial connected at manifold position  19 .  FIG. 11  depicts manifold  112  from the opposite face, such that the rotatable stopcocks and the ports  121   a  and  123   a  are hidden from view. 
     A 14 cm length of a tubing  122  extends between the free end of cartridge  114  and the luer connector of manifold valve  17 . An 8 cm length of tubing  124  extends between the free end of cartridge  116  and the luer connector of manifold valve  23 . A 14 cm length of tubing  126  extends between the free end of cartridge  120  and the luer connector of manifold valve  20 . Additionally, tubing  128  extends from the luer connector of manifold valve  1  to a target recovery vessel  129  (shown in  FIG. 11 ) which recovers the waste enriched water after the fluoride has been removed by the QMA cartridge. The free end of tubing  128  supports a connector  131 , such as a luer fitting or an elongate needle and associated tubing, for connecting the cavity to the target recovery vessel  129 . In the method of the present invention, the radioisotope is [ 18 F]fluoride provided in solution with H 2 [ 18 O] target water and is introduced at manifold valve  6 . 
     A tetrabutylammonium bicarbonate eluent vial  130  is positioned within the vial housing at manifold valve  2  and is to be impaled on the spike therein. An elongate 1 mL syringe pump  132  is positioned at manifold valve  3 . Syringe pump  132  includes an elongate piston rod  134  which is reciprocally moveable by the synthesis device to draw and pump fluid through manifold  112  and the attached components. QMA cartridge  136  is supported on the luer connector of manifold valve  4  and is connected via a 14 cm length of silicone tubing  138  to the luer connector of manifold position  5 . Cartridge  136  is desirably a QMA light carbonate cartridge sold by Waters, a division of Millipore. The tetrabutylammonium bicarbonate in an 80% acetonitrile; 20% water (v/v) solution provides elution of [ 18 F]fluoride from QMA and phase transfer catalyst. A fluoride inlet reservoir  140  is supported at manifold valve  6 . 
     Manifold valve  7  supports a tubing  142  at its luer connector which extends to a first port  144  of a reaction vessel  146 . The luer connector of manifold valve  8  is connected via a 14 cm length of tubing  148  to a second port  150  of reaction vessel  146 . The luer connector of manifold valve  9  is connected via a 42 cm length of tubing  152  to a vial  154  containing a mixture of 40% MeCN and 60% water (v/v). The acetonitrile and water mixture is used to enable primary purification of flutemetamol at the first SPE cartridge  114 . The luer connector of manifold valve  10  is connected via a 42 cm length of tubing  156  to a vial  158  containing 100% MeCN used for conditioning of the cartridges and the elution of flutametamol from the first SPE cartridge  114 . Manifold valve  11  supports a barrel wall for a 5 ml syringe pump  160 . Syringe pump  160  includes an elongate piston rod  162  which is reciprocally moveable by the synthesis device so as to draw and pump fluid through manifold  112 . The vial housing at manifold valve  12  receives vial  164  containing 6-ethoxymethoxy-2-(4′-(N-formyl-N-methyl)amino-3′-nitro)phenylbenzothiazole). The vial housing at manifold valve  13  receives a vial  166  containing 4M hydrochloric acid. The hydrochloric acid provides deprotection of the radiolabelled intermediate. The vial housing at manifold valve  14  receives a vial  168  of a methanol solution of sodium methoxide. The vial housing at manifold valve  15  receives an elongate hollow spike extension  170  which is positioned over the cannula at manifold valve  15  and provides an elongate water bag spike  170   a  at the free end thereof. Spike  170  pierces a cap  172  of a water bottle  174  containing water for both diluting and rinsing the fluid flowpaths of cassette  110 . The vial housing at manifold valve  16  receives a vial  176  containing ethanol. Ethanol is used for the elution of the drug substance from the second SPE cartridge  116 . The luer connector of manifold valve  17  is connected to a 14 cm length of silicone tubing  122  to SPE cartridge  114  at position  18 . Manifold valve  24  supports the elongate barrel of a 5 ml syringe pump  180 . Syringe pump  180  includes an elongate syringe rod  182  which is reciprocally moveable by the synthesis device to draw and pump fluid through manifold  112  and the attached components. The luer connector of manifold valve  25  is connected to a 42 cm length of a tubing  184  to a third port  186  of reactor vessel  146 . 
     Cassette  110  is mated to an automated synthesizer having rotatable arms which engage each of the stopcocks of valves  1 - 25  and can position each in a desired orientation throughout cassette operation. The synthesizer also includes a pair of spigots, one of each of which insert into ports  121   a  and  123   a  of connectors  121  and  123  in fluid-tight connection. The two spigots respectively provide a source of nitrogen and a vacuum to manifold  112  so as to assist in fluid transfer therethrough and to operate cassette  110  in accordance with the present invention. The free ends of the syringe plungers are engaged by cooperating members from the synthesizer, which will then apply the reciprocating motion thereto within the syringes. A bottle containing water is fitted to the synthesizer then pressed onto spike  170  to provide access to a fluid for driving compounds under operation of the various-included syringes. The reaction vessel will be placed within the reaction well of the synthesizer and the product collection vial and waste vial are connected. The synthesizer includes a radioisotope delivery conduit which extends from a source of the radioisotope, typically either vial or the output line from a cyclotron, to a delivery plunger. The delivery plunger is moveable by the synthesizer from a first raised position allowing the cassette to be attached to the synthesizer, to a second lowered position where the plunger is inserted into the housing at manifold valve  6 . The plunger provides sealed engagement with the housing at manifold valve  6  so that the vacuum applied by the synthesizer to manifold  112  will draw the radioisotope through the radioisotope delivery conduit and into manifold  112  for processing. Additionally, prior to beginning the synthesis process, arms from the synthesizer will press the reagent vials onto the cannulas of manifold  112 . The synthesis process may then commence. 
       FIG. 9  depicts an SPE cartridge  210  of the present invention. The sorbent fill  212  of the reverse phase SPE cartridges will differ from the fill of the normal phase SPE cartridge. Cartridge  210  includes an elongate tubular body  214  defining a cylindrical cavity  216 . A first end  214   a  of body  214  includes a transverse annular wall  218  defining a exit aperture  220  in fluid communication with cavity  216 . Annular wall  218  also supports an elongate open tubular wall  222  forming a luer tip  224 . The opposing second end  214   b  of body  214  supports an end cap  226  having a cap body  228  defining an inlet aperture  230  in fluid communication with cavity  216 . Cap body  228  includes an outer annular rim  232  engaging the outer surface  234  of tubular body  214  at second end  214   b  and an inner annular wall  236  engaging the inner surface  238  of tubular body  214  at second end  214   b . Cartridge  210  also includes circular disc-shaped porous filter elements  240  and  242  spanning across cavity  216  with sorbent fill  212  therebetween. By way of illustration and not of limitation, cartridge  210  is generally about 48.6 mm in length, about 15.2 mm in diameter at second end  214   b , about 12.0 mm in diameter at first end  214   a  and cavity  216  is about 34.6 mm in length, although the size and shape of cartridge  210  may be selected as will be suitable for its intended purpose. 
     EXAMPLES 
     Example 1 
     FASTlab™ Synthesis of [ 18 F]flutemetamol Injection Using SPE Purification 
     With reference to  FIGS. 4, 7 and 8 , the production and formulation of Flutemetamol ( 18 F) Injection on a cassette of the present invention is described. For this process, the cassette of  FIGS. 5 and 11  was constructed for operation by the FASTlab™ machine. First, [ 18 F]fluoride solution is transferred by vacuum to the FASTlab™ and is trapped on a QMA cartridge (commercially available from Waters (Milford, Mass. USA)) that had been preconditioned. The [ 18 O]enriched water is recovered and takes no further part in the synthesis. The [ 18 F]fluoride is then eluted directly from the QMA cartridge into the reaction vessel with tetrabutylammonium bicarbonate solution (350 μl, 0.15 M in 80:20 acetonitrile:water). 
     The reaction vessel is heated by the synthesis unit under a flow of nitrogen and a vacuum in order to dry the [ 18 F]fluoride and remove the QMA eluent solvents. For the radiolabelling reaction, the final intermediate, AH111907 (6-ethoxymethoxy-2-(4′-(N-formyl-N-methyl)amino-3′-nitro)phenylbenzothiazole) in anhydrous dimethyl sulfoxide (DMSO) is added to the reaction vessel. The vessel is subsequently sealed, by positioning manifold valves  7 ,  8 , and  25  to seal the vessel, and heated. Sodium methoxide in methanol is added to the reaction vessel, which is then heated. To effect deprotection, hydrochloric acid is added to the reaction vessel, which is then heated. The crude deprotected reaction mixture containing [ 18 F]flutemetamol is diluted with sterile purified water (about 2 ml) before being passed through the first reverse phase SPE cartridge  114 . 
     For the primary SPE purification, the first reverse phase SPE cartridge  114  is washed with 12 ml 40% acetonitrile:60% water (v/v), followed by 5 ml water in order to remove the majority of the impurities (including the hydrophilic precursor derivatives). The partially purified flutemetamol is eluted from the first reverse phase SPE cartridge  114  in 2 ml acetonitrile. 
     For the secondary SPE purification, the normal phase, the 2 ml acetonitrile solution from the first reverse phase SPE cartridge is passed back and forth through the amino cartridge (or normal phase SPE cartridge)  120  in order to trap many of the remaining hydrophilic impurities. The amino cartridge is then rinsed with a further 1 ml acetonitrile to maximise recovery of flutemetamol. 
     For then performing solvent exchange and formulation, the acetonitrile solution (about 3 mL) from the amino cartridge  120  is diluted with water (about 5.5 mL) and passed through the second C30 cartridge  116 . The C30 cartridge  116  is then rinsed three times with water (up to about 20 mL in total) to reduce levels of acetonitrile and methanol. The drug substance [ 18 F]flutemetamol is retained on the cartridge  116  and eluted with ethanol (about 3.5 ml) into the product collection vial prefilled with polysorbate 80, phosphate buffer and sodium chloride to give Flutemetamol [ 18 F] Injection. The C30 cartridge  116  is further eluted with water (about 9.3 mL) in order to flush out any remaining ethanol and this passes directly into the product collection vial. One of the FASTlab™ syringes then draws the contents of the product collection vial up and down in order to homogenise the drug product. 
     All patents, journal articles, publications and other documents discussed and/or cited above are hereby incorporated by reference. 
     While the particular embodiment of the present invention has been shown and described, it will be obvious to those skilled in the art that changes and modifications may be made without departing from the teachings of the invention. The matter set forth in the foregoing description and accompanying drawings is offered by way of illustration only and not as a limitation. The actual scope of the invention is intended to be defined in the following claims when viewed in their proper perspective based on the prior art.

Technology Category: 7