HK40078470A - Apparatus and method for solid phase extraction - Google Patents

Description

Apparatus and method for solid phase extraction

Technical Field

The present invention relates to the field of Solid Phase Extraction (SPE). More particularly, the present invention relates to an apparatus and method for optimizing SPE conditions.

Background

Automated synthesis systems are important for the production of radiopharmaceuticals. Synthetic systems, such as FASTlab ® synthesizers (GE Healthcare), provide for the production of doses for clinical applications. The FASTlab synthesizer accepts and operates methods via equipment used to produce radiopharmaceuticals. Examples of such radiopharmaceuticals include¹⁸F-FLT ([¹⁸F]Fluoro-thymidine),¹⁸F-FDDNP (2-(1-{6-[(2-[¹⁸F]Fluoroethyl) (methyl) amino]2-naphthyl } ethylene) malononitrile), and,¹⁸F-FHBG (9-[4-[¹⁸F]Fluoro-3- (hydroxymethyl) butyl]Guanine or [2 ]¹⁸F]Penciclovir (I),¹⁸F-FESP ([¹⁸F]-fluoroethylspirone)¹⁸F-p-MPPF (4- (2-methoxyphenyl) -1- [2- (N-2-pyridyl) -p-, [2 ]¹⁸F]Fluorobenzamido]Ethylpiperazine) and¹⁸F-FDG ([¹⁸F]-2-deoxy-2-fluoro-D-glucose), and the like.

Such a synthesis system/device is used with a cartridge comprising: a flow path comprising a first end and a second end; and a plurality of valves oriented along the flow path, wherein each of the plurality of valves is selectively fluidly connected to one of a plurality of components, typically including reagent vials, reaction vessels in which one or more reactions of a process for producing a compound are conducted, cartridges, filters, syringes, tubing, and connectors for synthesizing a particular radiotracer. A dedicated cassette customized for the radiopharmaceutical is used to manufacture a different radiopharmaceutical. The apparatus is arranged to cooperatively engage the components so that each stopcock and syringe can be actuated to drive a source fluid having a radioisotope through the apparatus to effect a chemical synthesis process. The synthesis system may also include one or more (e.g., two) heating chambers/elements that receive the first reaction vessel in order to provide any heat required for the chemical reaction.

The synthesis system is programmed to operate the required pumps, syringes, valves, heating elements, and control the supply of motive gas (e.g., nitrogen) and application of vacuum in order to direct the source fluids to mix with the reagents, conduct chemical reactions through the appropriate purification cartridges, and selectively pump the output tracers and waste fluids into the appropriate vial containers for the final product, waste product, and the like. Although the fluid collected in the output vial is typically input into another system for purification and/or distribution, the synthesis system may also be connected to or include a separate purification system that returns the purified (or partially purified compound) to the system for further processing.

Such an automated synthesizer of the prior art is described in WO 2007/042781.

While existing synthesis systems (e.g., FASTlab synthesizers) may include some form of purification step, the systems are not designed to optimize the purification conditions, nor do they. Existing SPE manifold systems (where SPE is performed manually) are currently used to optimize purification methods, such as those used in the prior art (e.g., FASTlab systems). Determining an appropriate purification method in this manner is time consuming as it typically involves each manufacture of the product followed by one or more different purification parameters. Furthermore, once suitable purification conditions have been identified, the resulting purification process needs to be further tuned and optimized to be compatible with the synthesis system.

There are many examples of SPE robots available on the market. A key benefit of using the present invention is that it allows SPE purification methods to be developed quickly using existing production equipment. Effectively eliminating the time it takes to transfer the SPE method onto an existing device (e.g., FASTlab) because the backbone may be the same for both.

Existing SPE robots are designed to collect larger fractions than are typically used in the present invention. The ability to collect smaller fractions of the eluate for analysis is an improvement over current SPE robots. The larger the fraction collected, the less can be inferred about the position of the impurity/product elution.

Furthermore, existing SPE robots occupy a large footprint in the laboratory. The present invention provides the advantage that no additional piece of equipment dedicated to SPE optimization is required in the case where the laboratory already has an automated synthesis system.

For example, the cassettes of the invention may be used in conjunction with the FASTlab system which has been programmed to evaluate up to three SPE cartridges having up to 6 different mobile phases in one sequence. Furthermore, this provides the advantage that the cartridges can be reused, allowing them to be tested repeatedly, which saves both time and cartridge costs.

It would be beneficial if methods could be developed to optimize product purification to reduce the timescale described above, and to be compatible with existing synthesizer systems.

Disclosure of Invention

The present invention provides apparatus for optimizing SPE purification conditions for separating a compound from a composition containing the compound, preferably from a crude reaction mixture. The compound to be isolated from the composition may be any radioactive or non-radioactive compound, preferably not a radioactive compound. The device of the first aspect of the invention is described as a cartridge. Such a cartridge may be a separate piece of equipment and/or may be a piece of equipment that fits into already existing equipment in place of some of the original components.

In a first aspect, the present invention provides a cartridge for determining optimized Solid Phase Extraction (SPE) purification conditions, wherein the cartridge comprises:

(i) a flow path comprising a first end and a second end; and

(ii) a plurality of valves oriented along the flow path, wherein each of the plurality of valves is selectively fluidly connected to one of a plurality of components,

wherein the component comprises:

(a)1-5 vials of the composition;

(b)1-3 SPE cartridges;

(c)4-10 solvent vials;

(d) a water vial; and

(e) a transfer line.

The cartridge of the present invention has a flow path comprising a first end and a second end. The flow path is a channel suitable for transporting materials (in particular fluids, such as solvents) and compositions (such as crude reaction mixtures). The composition vial may be a crude reaction mixture vial, a single reference standard vial, or a reference standard mixture vial. The "crude reaction mixture" may include the desired product in a mixture with one or more impurities. A "single reference standard" may be a single impurity, such as an impurity that is typically produced with the desired product. Similarly, a 'reference standard mixture' may be a mixture of impurities, such as those typically produced with the desired product.

By "fluidly connected" is meant that fluid can pass into and out of the vial and (optionally) through the valve to other portions of the cartridge. A suitable valve may be a 3-way valve having three ports and means for placing any two of the three associated ports in fluid communication with each other while fluidly isolating the third port. A suitable valve may also be a stopcock valve comprising a rotatable stopcock.

The cassette may be linked to or compatible with a system or device for synthesizing the compounds provided by the cassettes of the invention, such as the known FASTlab system. The cassette of the present invention is compatible with synthesizer systems for radiopharmaceutical synthesis, as described above, and is specifically set up to optimize SPE purification conditions. Thus, the present invention allows the determination of an optimized SPE purification method for a product on the same system and/or compatible system used to produce the product. This provides a significant advantage over existing methods. For example, because the purification process is optimized on the same or compatible system used to produce the desired product, the purification process does not require further adaptation to work on the system as compared to existing processes.

The cartridge is designed such that on the one hand it provides a combination of SPE cartridges and solvent vials providing different solvents (different chemical compositions or different concentrations) and on the other hand provides a device where alternative parameters of SPE and solvent can be used and combined to determine the optimal purification method.

Preferably, the cartridge comprises:

(i) 3 SPE cartridges and 6 solvent vials; or

(ii) 2 SPE cartridges and 8 solvent vials; or

(iii) 1 SPE cartridge and 10 solvent vials.

One or more SPE cartridges may be conditioned using one of the solvent vials. Conditioning may be performed by passing 100% solvent through the SPE cartridge, then passing water through the SPE cartridge. In this case, the number of solvent vials used for purification can be viewed as 5 solvent vials for 3 SPE cartridges, 7 solvent vials for 2 SPE cartridges, and 9 solvent vials for 1 SPE cartridge.

In one embodiment, the cassette of the invention does not comprise a means for processing a radioisotope (e.g., [2 ]¹⁸F]Fluoride) to be used. For example, the cartridge does not comprise an 'ion exchange cartridge', e.g. an SPE cartridge, when coming from a nuclear reaction¹⁸O(p,n)¹⁸The aqueous solution of F is retained when it passes through¹⁸F and allow¹⁸And (4) passing the oxygen. Such ion exchange cartridges include anion exchange cartridges, for example, methyl quaternary ammonium (QMA) cartridges, and these are typically found in known cassettes designed for the radiosynthesis of radiolabeled compounds. In one embodiment, the cassette also does not include a 'cationic counterion', where the cationic counterion may be a positively charged counterion, such as a cryptand or a metal complex of a tetraalkylammonium salt or a large but soft metal ion, such as rubidium or cesium。

In one embodiment, the cartridge does not include a reaction vessel.

The cartridge may have 25 valves arranged linearly. When the cartridge has 25 valves arranged linearly and the cartridge contains 3 SPE cartridges and 6 solvent vials,

(i) 1-5 composition vials fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 3 composition vials are used;

(ii) the 3 SPE cartridges are preferably fluidly connected to the 18 th, 20 th and 22 th valves;

(iii) the 6 solvent vials were fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th and 10 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

When the cartridge has 25 valves arranged linearly and the cartridge contains 2 SPE cartridges and 8 solvent vials,

(i) 1-5 composition vials fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 2 composition vials are used;

(ii) the 2 SPE cartridges are preferably fluidly connected to the 20 th and 22 th valves;

(iii) the 8 solvent vials were fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th, 10 th, 17 th and 18 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

When the cartridge has 25 valves arranged linearly and the cartridge contains 1 SPE cartridge and 10 solvent vials,

(i) 1-5 composition vials fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 1 composition vial is used;

(ii) the 1 SPE cartridge is preferably fluidly connected to the 22 nd valve;

(iii) the 10 solvent vials are fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th, 10 th, 17 th, 18 th, 19 th, and 20 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

The above cartridge may further comprise:

(v) a syringe fluidly connected to the 3 rd valve, preferably wherein the syringe is a 1 mL syringe; and

(vi) a syringe fluidly connected to the 11 th valve, preferably wherein the syringe is a 7 mL syringe; and

(vii) a syringe fluidly connected to the 24 th valve, preferably wherein the syringe is a 7 mL syringe.

The above cartridge may further comprise:

(viii) a tube connecting the 1 st and 25 th valves.

The above cartridge may further comprise:

(ix) an output of eluent from one or more SPE cartridges fluidly connected to the 23 th valve.

In a second aspect of the invention, the invention provides a method for determining optimized SPE purification conditions for a compound from a composition, the method comprising:

(i) providing a cartridge as defined according to the first aspect of the invention;

(ii) said cartridge containing a composition of said compound in each of said 1-5 composition vials or adding such a composition to each of said 1-5 composition vials;

(iii) passing an aliquot of the composition into each of the 1-3 SPE cartridges;

(iv) passing a specific combination of aliquots of solvent from at least 4 of the 4-10 solvent vials into one or more of the SPE cartridges, wherein the solvent in each of the 4-10 solvent vials is a different solvent or the same solvent at a different concentration;

(v) eluting the compound to be purified from the or each SPE cartridge;

(vi) (vi) evaluating the eluted product of step (v); and

(vii) (vi) determining the optimal purification conditions by comparing the products eluted from each cartridge and each solvent in step (v).

The at least one solvent is selected from: (i) ethanol, (ii) methanol, (iii) acetonitrile, or any alternative organic solvent known in the art, or combinations thereof. Preferably, the product is eluted with ethanol or aqueous ethanol, since if a different solvent is used, an aqueous solvent exchange step needs to be included.

The method may further comprise the step of conditioning 1-3 SPE cartridges as a first step of the method. The cartridge can be conditioned with 100% organic solvent (methanol, ethanol or acetonitrile) and then with water. The volumes of organic solvent and water used for adjustment can be varied to suit different cartridges. For example, 7 mL of organic solvent and 7 mL of water may be suitable. Or a smaller volume of organic solvent may be suitable, for example 2 mL, and in this case, multiple cartridges may be adjusted using solvent from only one syringe. In one embodiment, a syringe filled with water is used to ensure that the solvent is completely removed. After conditioning, the crude product was loaded onto one or more SPE cartridges.

Each cartridge was then washed with mobile phase eluent in variable volume fractions, where the volume is a compromise between "more detail" (i.e. smaller volume) and "analysis time", i.e. the smaller the volume collected, the more sample that needs to be analysed. The volume of the fraction is preferably 1 mL. The eluate fractions are collected and analyzed, for example by analytical HPLC. During this process, the cartridge can be cleaned with 100% organic solvent and reconditioned for reuse with a different mobile phase composition. To minimize operator handling time, samples can be collected in 96-well plates and analyzed on an HPLC system using an autosampler injection system. The collected samples can be analyzed overnight and then the results interpreted the next morning. The evaluation in step (vi) may be performed using any suitable method, such as HPLC, LC-MS or TLC.

1, 2, 3, 4 or 5 vials of the composition may be used. In particular, 1, 2 or 3 vials of the composition may be used. Preferably, the number of composition vials may be the same as the number of SPE cartridges.

The term "elution" refers to passing a solution through the SPE cartridge with the purpose of releasing one or more compounds of interest that have bound to the solid phase. Elution can be carried out by passing a suitable solvent through the SPE cartridge and through the transfer line for collection, preferably wherein collection is carried out in a 96-well plate. Suitable solvents are solvents for releasing the compound of interest from the SPE cartridge and may be, for example, organic solvents, acidic solvents or basic solvents, depending on the chemistry of the compound of interest and the nature of the SPE column chemistry.

The above method may further comprise the step of eluting impurities. Elution of the impurities may be carried out before and/or after step (v) above where the desired product is eluted. Preferably, elution of the impurities may be performed before and after step (v) above where the desired product is eluted.

The purification process is ideally effective if the product obtained from the composition is obtained in a purity of at least 95%, preferably at least 97%, more preferably at least 99%.

The invention also provides a compound purified according to the second aspect of the invention.

In a third aspect, the present invention provides a kit comprising:

(i) according to the cartridge of the first aspect,

(ii) 1-5 vials of the composition;

(iii) 1-3 SPE cartridges;

(iv) 4-10 solvent vials;

(v) a vial of water; and

(vi) a transfer line.

Drawings

Fig. 1 to 6 are provided to illustrate the invention in a non-limiting manner:

figure 1 provides an example of an embodiment of a cartridge of the present invention comprising 3 SPE cartridges and 6 solvent vials (labeled mobile phase).

Figure 2 provides an example of an embodiment of a cartridge of the present invention comprising 2 SPE cartridges and 8 solvent vials (labeled mobile phase).

Figure 3 provides an example of an embodiment of a cartridge of the present invention comprising 1 SPE cartridge and 10 solvent vials (labeled mobile phase).

Fig. 4 provides an example of a cassette typically used with the FASTlab system for comparison with the present invention.

Figure 5 shows the results of an embodiment of the process of the present invention applied to the hydroxyl impurities in the crude Flurpiridaz product.

Figure 6 shows the results of an embodiment of the process of the present invention applied to a crude reaction mixture of Flurpiridaz. In FIG. 6, the x-axis is volume (1-41 ml) and the y-axis is ug (0-25).

Detailed Description

Examples

The following examples describe the invention in a non-limiting manner:

the first experiment was performed using a crude sample containing compound GE-179, with mobile phases consisting of 20, 30 and 40% ethanol in 0.1% aqueous formic acid solution. The structure of GE-179 is as follows:

the FASTlab cassette was set up as shown in figure 1. All 3 cartridges were conditioned with 100% ethanol (2 mL) followed by 100% water (7 mL). The crude product (dissolved in 10% ethanol 90% water) was loaded onto a 3 xtc 18 cartridge. The first cartridge was washed with 20% ethanol, the second cartridge with 30% ethanol, and the third cartridge with 40% ethanol (each through 18 mL, collecting 1 mL fractions). 54 samples were collected in 96-well plates and analyzed using an analytical HPLC system with an autosampler. The resulting data show that with 40% EtOH, all material was eluted from the cartridge in the first 6 mL, while with 20% EtOH, the desired product and all subsequent elution peaks were captured on the cartridge after 18 mL washing. These results reveal that the optimum conditions for washing the cartridge will be greater than 20% EtOH but less than 40% EtOH. Further experimentation using the same technique was required to determine the optimal selective elution of the product.

Experiments involving purification of Flurpiridaz were also performed, which has the following structure:

the major impurity in the crude Flurpiridaz product is a hydroxyl impurity, which has the following structure:

the FASTlab cassette was set up as shown in figure 2. Both cartridges were conditioned with 100% ethanol (7 mL) followed by 100% water (7 mL). The hydroxyl impurities (dissolved in 1:10 ethanol: water) were loaded onto a 2 XTC 18 cartridge. The first cartridge was washed with 40% acetonitrile and the second cartridge was washed with 35% ethanol (each by 41 mL, collecting 1 mL fractions). 82 samples were collected in 96-well plates and analyzed using an analytical HPLC system with an autosampler.

Figure 5 shows that the bands for the hydroxyl impurity eluted with 40% acetonitrile (dashed line) are tighter compared to 35% ethanol (solid line). These results demonstrate that 40% acetonitrile is superior to 35% ethanol and that a wash volume of 14-21 mL is sufficient to remove the major impurities from the crude product. Note that the line corresponding to 35% ethanol elution in fig. 5 includes a sharp drop due to the wrong data point.

The experiment was repeated with a crude reaction mixture of Flurpiridaz:

the FASTlab cassette was set up as shown in figure 3. The cartridges were conditioned with 100% ethanol (7 mL) followed by 100% water (7 mL). The crude product (dissolved in about 20% acetonitrile 80% aqueous solution) was loaded onto a tC18 cartridge. The cartridge was washed with 40% acetonitrile (41 mL, 1 mL fractions were collected). 41 samples were collected in 96-well plates and analyzed using an analytical HPLC system with an autosampler. When combined with information from the hydroxyl standard experiment described above, these results shown in figure 6 confirm that a wash volume of about 16 mL is sufficient to remove the hydroxyl impurities without eluting the product. Other impurities present in very small amounts are cyano impurities and chloro impurities.

Those skilled in the art will readily appreciate that the embodiments of the invention described herein are capable of broad use and application. Thus, while the present invention is described herein in detail with respect to exemplary embodiments, it is to be understood that the present disclosure illustrates and exemplifies embodiments and is intended to provide an enabling disclosure of exemplary embodiments. The present disclosure is not intended to be construed as limiting embodiments of the invention or to otherwise exclude any other such embodiments, adaptations, variations, modifications and equivalent arrangements. The scope of the invention is defined by the appended claims.

Claims (15)

1. A cartridge for determining optimized Solid Phase Extraction (SPE) purification conditions, wherein the cartridge comprises:

(i) a flow path comprising a first end and a second end; and

(ii) a plurality of valves oriented along the flow path, wherein each of the plurality of valves is selectively fluidly connected to one of a plurality of components,

wherein the component comprises:

(a)1-5 vials of the composition;

(b)1-3 SPE cartridges;

(c)4-10 solvent vials;

(d) a water vial; and

(e) a transfer line.

2. The cassette of claim 1, wherein the cassette does not comprise for processing [2 ]¹⁸F]Any device for fluoride.

3. The cartridge of claim 1 or claim 2, wherein the cartridge comprises:

(i) 3 SPE cartridges and 6 solvent vials; or

(ii) 2 SPE cartridges and 8 solvent vials; or

(iii) 1 SPE cartridge and 10 solvent vials.

4. The cartridge of claim 3 part (i), wherein the cartridge has 25 valves arranged linearly, an

(i) The 1-5 composition vials are fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 3 composition vials are used;

(ii) the 3 SPE cartridges are fluidly connected to the 18 th, 20 th and 22 th valves;

(iii) the 6 solvent vials are fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th and 10 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

5. The cartridge of claim 3 part (ii), wherein the cartridge has 25 valves arranged linearly, an

(i) The 1-5 composition vials are fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 2 composition vials are used;

(ii) the 2 SPE cartridges are preferably fluidly connected to the 20 th and 22 th valves;

(iii) the 8 solvent vials are fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th, 10 th, 17 th and 18 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

6. The cartridge of claim 3 part (iii), wherein the cartridge has 25 valves arranged linearly, an

(i) The 1-5 composition vials are fluidly connected to the 2 nd, 12 th, 13 th, 14 th and/or 16 th valves, preferably wherein up to 1 composition vial is used;

(ii) the 1 SPE cartridge is preferably fluidly connected to the 22 th valve;

(iii) the 10 solvent vials are fluidly connected to the 4 th, 5 th, 7 th, 8 th, 9 th, 10 th, 17 th, 18 th, 19 th, and 20 th valves; and

(iv) the water vial is fluidly connected to the 15 th valve.

7. The cartridge of any one of claims 4-6, wherein the cartridge further comprises:

(i) a syringe fluidly connected to the 3 rd valve, preferably wherein the syringe is a 1 mL syringe; and

(ii) a syringe fluidly connected to the 11 th valve, preferably wherein the syringe is a 7 mL syringe; and

(iii) a syringe fluidly connected to the 24 th valve, preferably wherein the syringe is a 7 mL syringe.

8. A method for determining optimized SPE purification conditions for isolating a compound from a composition, the method comprising:

(i) providing a cartridge as defined in any one of claims 1-7;

(ii) said cartridge containing a composition of said compound in each of said 1-5 composition vials or adding such a composition to each of said 1-5 composition vials;

(iii) passing an aliquot of the composition into each of the 1-3 SPE cartridges;

(iv) passing a specific combination of aliquots of solvent from at least 4 of the 4-10 solvent vials into one or more of the SPE cartridges, wherein the solvent in each of the 4-10 solvent vials is a different solvent or the same solvent at different concentrations;

(v) eluting the compound to be purified from the or each SPE cartridge;

(vi) (vi) evaluating the eluted product of step (v); and

(vii) (vi) determining the optimal purification conditions by comparing the products eluted from each cartridge and each solvent in step (v).

9. The method of claim 8, wherein the method further comprises a step of eluting impurities, preferably wherein the step of eluting impurities is performed before and/or after step (v).

10. The method of claim 8 or claim 9, wherein at least one solvent is selected from the group consisting of: (i) ethanol, (ii) methanol, (iii) acetonitrile, or a combination thereof.

11. The method of any one of claims 8-10, wherein said method further comprises the step of conditioning said 1-3 SPE cartridges prior to step (ii).

12. The method of any one of claims 8-11, wherein the evaluating in step (vi) is performed using HPLC, LC-MS, or TLC.

13. The method of any one of claims 8-12, wherein elution is performed by passing an organic solvent through the SPE cartridge and through the transfer line for collection, preferably wherein the collection is performed in a 96-well plate.

14. The cartridge of any one of claims 1-7 or the method of any one of claims 8-13, wherein the 1-5 composition vials are:

(i) 1-5 vials of crude reaction mixture;

(ii) 1-5 single reference standard vials; or

(iii) 1-5 vials of the reference standard mixture,

preferably wherein 1, 2, 3, 4 or 5 vials of the composition are used, more preferably wherein 1, 2 or 3 vials of the composition are used.

15. A kit, comprising:

(i) the cartridge as defined in any one of claims 1 to 7,

(ii) 1-5 vials of the composition;

(iii) 1-3 SPE cartridges;

(iv) 4-10 solvent vials;

(v) a water vial; and

(vi) a transfer line.