Classifications

• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
  • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
  • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/041—Heterocyclic compounds
  • A61K51/044—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins
  • A61K51/0455—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine, rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
• • A—HUMAN NECESSITIES
  • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
  • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
  • A61K51/00—Preparations containing radioactive substances for use in therapy or testing in vivo
  • A61K51/02—Preparations containing radioactive substances for use in therapy or testing in vivo characterised by the carrier, i.e. characterised by the agent or material covalently linked or complexing the radioactive nucleus
  • A61K51/04—Organic compounds
  • A61K51/08—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins
  • A61K51/088—Peptides, e.g. proteins, carriers being peptides, polyamino acids, proteins conjugates with carriers being peptides, polyamino acids or proteins
• • C—CHEMISTRY; METALLURGY
  • C07—ORGANIC CHEMISTRY
  • C07D—HETEROCYCLIC COMPOUNDS
  • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
  • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
  • C07D401/06—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a carbon chain containing only aliphatic carbon atoms

Definitions

• the present invention relates to maleimide compounds labeled with fluorine-18.
• the invention also relates to a process for the preparation of these compounds.
• the invention finally relates to the use of these maleimide compounds, in particular labeled with fluorine-18, for the labeling of. macromolecules, such as oligonucleotides, proteins, antibodies and peptides.
• the technical field of the invention can be defined, in general, as that of radioactive labeling of macromolecules and, in particular, of proteins and peptides.
• macromolecules such as proteins or even peptides can be coupled to a labeling molecule allowing their detection, this labeling molecule can be, for example, a fluorescent molecule, gold particles, a paramagnetic compound or a molecule carrying a radioelement.
• Proteins have been radioactively labeled with radioisotopes, iodine and various metal radioisotopes, such as technetium, indium and gallium. More recently, proteins have been labeled with fluorine-18.
• peptides coupled to radioelements allow "in vivo" detection of the localization of thrombotic zones during vascular accidents of all kinds, in particular apoptotic and inflammatory foci, using systems of imaging.
• radioactive atoms emitting positrons with a short lifespan and in particular 18 F can, in particular, be detected by positron emission tomography (PET) devices (PET or “Positon Emission Tomography”).
• PET positron emission tomography
• Radioactive labeling with fluorine-18 poses, in particular due to the very short period of fluorine-18 (close to 109.8 minutes), specific problems which make labeling with fluorine-18 fundamentally different from that with other halogens, such as iodine.
• the above coupling can be carried out by all the conventional techniques of organic chemistry known to those skilled in the art, and by the synthesis of protein and peptide markers carrying one or more radioactive atoms with a short lifespan, in particular 18 F.
• This marker generally consists, on the one hand, of a part capable of receiving, for example, an 18 F atom and, on the other hand, of a part comprising any conventional function of binding to the macromolecule, by for example, protein.
• These markers must meet the requirement for rapid and easy synthesis, because due to the short lifespan of radioisotopes such as 18 F, the duration of synthesis should not generally exceed a few hours.
• the methods for labeling proteins or peptides with fluorine-18 make use of markers also called “conjugated” or “synthon” labeled, which are classified into three main families, depending on whether they react with the amino groups, sulfhydryl groups, or carbohydrate groups of macromolecules, such as proteins and peptides.
• imidates such as 3- [ 18 F] fluoro-5-nitrobenzoimidate, which react, for example, with the ⁇ -NH 2 group of lysine to bind to a protein
• activated esters such as
• N-succinimidyl- [ 18 F] fluorobenzoate N-succinimidyl- [ 18 F] fluorobenzoate
• carboxylic acids such as N- (4- [ 18 F] fluorobenzoic acid
• aldehydes such as 4- [ 18 F] pentafluorobenzaldehyde
• isothiocyanates such as 4- ([ 18 F] fluoromethyl phenylisothiocyanate).
• Activated halides such as (4- [ 18 F] fluorophenacyl bromide), react with groups amino, such as the ⁇ -NH 2 group of lysine or the -SH group of cysteine.
• Amines such as 1- (4- ([ 18 F] fluoromethyl) benzoyl) -aminobutane-4-amine react with the CO 2 H groups, for example glutamic acid or aspartic acid or with CHO groups of glycoproteins.
• Nitrenes with photochemical active centers such as azidophenacyl fluoride [ 18 F] also react with amino groups, for example the ⁇ -NH 2 group of lysine.
• Two more specific methods for labeling peptides and nucleotides have good specificity with respect to sulfur atoms, for example cysteine for the peptides and a phosphoro-thioate function for the nucleotides.
• the first of these compounds is not easy to label with fluorine-18 with high specific activity.
• fluorine F 2 would allow easy labeling of (as with iodine) and it is precisely that F 2 is generally a product with low specific activity.
• F 2 is not suitable for the manufacture of so-called “radiotracers” compounds which are preferably targeted according to the invention, simply because the injected mass of labeled molecule becomes large and that, then, the basic principle governing this “tracer”, ie the extremely low occupation (for example, less than 5%) of the receiving sites, is not respected.
• the second of the compounds cited in the document by SHIUE et al. has an amide chain which is not chemically very solid and which is easily cleaved, broken, in vivo.
• X is a radioactive halogen chosen from bromine-75, bromine-76, bromine -82, iodine-123, iodine-125, iodine-131 and fluorine-18.
• the chemistry used to manufacture the fluorinated compound of document US Pat. No. 4,735,792 is a complex and long-term chemistry.
• the object of the present invention is to provide a fluorine-18 labeled maleimide compound which meets, among other things, these needs.
• the aim of the present invention is also to provide a fluorine-18 labeled maleimide compound which does not have the drawbacks, defects, limitations and disadvantages of the compounds of the prior art and which solves the problems of the prior art.
• n represents an integer from 1 to 10, such as 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10;
• Y represents a group chosen from monocyclic or bicyclic heterocyclic groups chosen from imidazolyl, pyrazolyl, benzimidazolyl, pyridinyl, piridazinyl, pyrimidinyl, pyrazinyl, triazinyl, quinolinyl, isoquinolinyl, cinnolinyl, quinazolinyl, quinoxalinyl, possibly quinoxalinyl substituted by one or more substituents each of said substituents being independently selected from hydrogen, halogen (non-radioactive), phenyl, alkyl C ⁇ - 6 alkoxy C ⁇ - 6, aryloxy, amino, mono- or di (alkyl C ⁇ _ 6) alkylamino, mono- or di (aryl) amino, thio, alkyl C ⁇ - 6 alkylthio, arylthio, formyl, alkyl C ⁇ - 6 alkylcarbonyl, arylcarbonyl,
• - a, b, c, d, e, f, g each independently represent an integer of 0 to 10, such as 0, 1, 2, 3, 4, 5, 6, 7, 8, 9;
• halogen means fluorine, chlorine, bromine or iodine.
• C ⁇ _ 6 alkyl corresponds to saturated hydrocarbon radicals with linear and branched chains having from 1 to 6 carbon atoms, such as methyl, ethyl, propyl, butyl, pentyl and hexyl.
• the attachment and substitution of heterocycles, aryl group, etc., can be done at any position.
• the attachment of 18 F to Y or X can be done at any position, in particular at any position on a heterocycle.
• n 1, and Y is a 3-pyridinyl group.
• the compounds of formula (I) can belong to various families, a first family can be defined as that of "alkyl ethers", which correspond to the following formula (II):
• a second family of compounds of formula (I) can be defined as those of “phenylalkyl ethers”, which correspond to the following formula (III):
• q and r independently represent an integer from 0 to 10, such as 0, 1, 2, 3, 4, 5, 6, 7, 8, 9.
• a third family is that of the compounds which correspond to the following formula (IV):
• a preferred compound of formula (IV) is the following compound:
• a fourth family is that of the compounds which correspond to the following formula (V):
• Preferred compounds of formula (V) are the following compounds:
• the compounds according to the invention differ fundamentally from the compounds of the prior art, because of their specific structure in which the part carrying the atom of fluorine-18 is constituted, according to the invention, by a specific group Y which is in particular a pyridinyl group, the linking part, of coupling to a macromolecule, such as a protein or a peptide, is constituted, according to the invention by a specific function, namely a maleimido function, and, finally, the part of bond to a macromolecule and the part carrying the fluorine-18 atom are linked according to the invention by a new specific chain or spacer arm, for example of the alkyl type (generally from 2 to 6C), alkyl ether, ethers phenylalkyl, alkenyl, which are not fragile and are not susceptible to ruptures "in vivo".
• the invention relates to the use of a compound, as described above for labeling macromolecules.
• This macromolecule can be any macromolecule, in particular known biological, but it can be chosen, for example, from the oligonucleotides, proteins, antibodies and peptides. Said macromolecule is advantageously a macromolecule for recognizing a specific site chosen, preferably, from sites presenting target molecules specific for a pathology, such as sites of apoptosis, necrosis or tumor area.
• the invention also relates to a complex comprising a macromolecule coupled to a compound according to the invention, as described above.
• Said macromolecule is preferably chosen from oligonucleotides, proteins, antibodies and peptides.
• Said coupling is carried out by reaction of the double bond of the maleimido group of the compound according to the invention with specifically an -SH (thiol) function of cysteine in the case of a peptide, or a phosphoro-thioate function in the case of an oligonucleotide.
• cysteines are due to the presence in the molecule of the invention of a “dedicated” function, namely the maleimido function, which is a dedicated function for chemoselectivity towards the thiols of cysteines , or in a similar manner towards the phosphoro-thioate functions.
• a “dedicated” function namely the maleimido function, which is a dedicated function for chemoselectivity towards the thiols of cysteines , or in a similar manner towards the phosphoro-thioate functions.
• the labeling or coupling, via cysteine can be a direct labeling or coupling, that is to say that the cysteine already exists in the macromolecule which it is desired to couple to the compound according to the invention, either cysteine or a molecule (peptide) comprising it, can be introduced (coupled beforehand or not with the compounds of the invention) in the macromolecule which did not contain cysteine and coupling is then carried out if this has not been carried out beforehand cysteine or the molecule comprising it.
• the cysteine or molecule comprising it can, for example, be introduced "on order" into the macromolecule by engineering proteins / peptides in a position which does not compete with, or does not disturb the biological function.
• Said macromolecule is advantageously a macromolecule for recognition of a specific site, as described above.
• the coupling that is to say the labeling, is preferably such that it does not affect the recognition activity of the target, of the site, by the macromolecule.
• the invention also relates to an analysis and detection kit, for example for imaging medical comprising a compound according to the invention and a macromolecule.
• the invention also relates to an analysis and detection kit, for example for medical imaging, comprising a compound according to the invention coupled to a macromolecule, that is to say a complex according to the invention.
• the invention also relates to a diagnostic kit comprising a compound according to the invention and a macromolecule.
• the invention further relates to a diagnostic kit comprising a complex, as described above.
• the invention relates to the use of the complex or the compound, as described above, in a medical imaging method, such as positron emission tomography (PET) and the use of a complex or a compound according to the invention for manufacturing a product intended for medical imaging, for example for positron emission tomography (PET).
• a medical imaging method such as positron emission tomography (PET)
• PET positron emission tomography
• PET positron emission tomography
• the invention relates to a product for medical imaging, in particular positron emission tomography (PET) comprising a complex or a compound as described above and a pharmaceutically acceptable vehicle.
• PET positron emission tomography
• the compounds and complexes, according to the invention, comprising an atom of fluorine-18 show numerous advantages over the compounds with another radioactive halogen, for example iodine. Indeed, the only isotope of the positron emitting iodine is iodine-124, which could allow PET.
• iodine-124 is not a pure positron emitter (unlike fluorine -18.97%) and decreases by beta + emission at only 25% and by electronic capture at 75%; it has a large number of gamma lines ranging from 0.603 MeV (62%) to 2.75 MeV (1%).
• the invention also relates to a process for the preparation of a compound of formula (I), as described above, in which: a) a precursor compound of formula (la) is brought into contact:
• PRi and PR 2 independently represent a hydrogen atom or a group protecting the amino function, provided that PRi and PR 2 are not both (simultaneously) a hydrogen atom, or PRi and PR 2 together with the nitrogen atom form a cyclic protecting group for the amino function
• Gp represents a leaving group capable of being replaced by a fluorine atom
• 18 and X, Y, m and n have the meaning already given more high ; with a source of fluoride ions F " marked with [ 18 F], to give a compound of formula (Ib):
• the method according to the invention is simple, reliable, easy to implement and can be easily robotized. It only has three steps, one of which is an extremely simple deprotection step.
• the overall duration of the process is short: for example, it is generally from 60 to 120 minutes, preferably from 75 to 85 minutes.
• the incorporation of the halogen fluor-18 is carried out extremely efficiently with a high yield, for example 70 to 100%, because, in particular, it is carried out on a heterocyclic group, such as pyridine.
• the final yield of the whole process for a purified product is extremely high, for example from 15% to 25% and the potential quantities of “synthon” compound, at the end of the synthesis, are also very large.
• the groups PR 1 and PR 2 when they are protective groups can be any protective group known in organic chemistry. They are preferably chosen from the tert-butoxycarbonyl (BOC) and fluorenylmethoxy carbonyl (FMOC) groups.
• this protective group can be, for example, a pthalimido group.
• the group Gp may be any leaving group capable of being replaced by a fluorine-18 atom; Gp is preferably chosen from halogens, such as F, Cl, Br, I, mesyl, tosyl and triflate groups, when Y is an alkyl group; and Gp is preferably chosen from halogens, ammonium salts, such as trimethylammonium-trifluoromethane sulfonate, and the nitro group, when Y is an aromatic or heterocyclic group.
• halogens such as F, Cl, Br, I, mesyl, tosyl and triflate groups
• Y is an alkyl group
• Gp is preferably chosen from halogens, ammonium salts, such as trimethylammonium-trifluoromethane sulfonate, and the nitro group, when Y is an aromatic or heterocyclic group.
• the source of fluoride ions marked with 18 F comprises said fluoride ions and a counterion, chosen from large cations, such as rubidium, and tetrabutylammonium, and small cations , such as potassium, sodium and lithium, said small cations being trapped, stabilized, for example, by a cryptand or a crown ether, etc., said cryptand or crown ether being adapted to the small cation used.
• a counterion chosen from large cations, such as rubidium, and tetrabutylammonium
• small cations such as potassium, sodium and lithium
• KRYPTOFIX ® K 222 (4, 7, 13, 16, 21, 24-hexaoxa-1,10-diazabicyclo [8.8.8] hexacosane) which traps, for example, potassium ion.
• the counterion or cation can be brought in the form of any salt, for example, it can be K 2 C0 3 , in the case of potassium.
• Step a) is generally carried out in a solvent, which can be any suitable solvent, such as DMSO.
• Step a) can be carried out under conditions known to a person skilled in the art, with heating generally at a temperature of 50 to 200 ° C., for example, 145 ° C., for a duration generally of 1 to 30 minutes, for example from 4 to 6 minutes.
• Step b) of removal of the protective group from the amino function, of deprotection, to give the compound of formula (Ic), where the amino group is free can be carried out by any known deprotection method. It is possible, for example, to put the compound (Ib) in contact with TFA in CH 2 C1 2 for a duration generally of 1 to 5, for example of 2 minutes.
• the reagent capable of giving a maleimido group from an amido group can be any known compound. It can thus be chosen from N-methoxycarbonylmaleimide and succinimide.
• Step c) can be carried out under conditions known to a person skilled in the art, for example in a solvent, such as xylene, THF, with heating generally at a temperature of 100 to 200 ° C., for example 190 ° C, for a period of 1 to 20 minutes, for example 5 minutes.
• a solvent such as xylene, THF
• Step c) can in another embodiment also be carried out in a two-phase mixture for example of dioxane and aqueous sodium bicarbonate, at room temperature for a period of 3 to 15 minutes, for example 10 minutes;
• This mode of implementation of step c) offers the advantage of better performance and be implemented at room temperature, without the need to heat the mixture.
• the compound of formula (la) can correspond to the following formula (IIa):
• the compound (Ha) preferably corresponds to the following formula (11b):
• the compound of formula (la) can, in another embodiment, correspond to the following formula (Illa):
• the compound (Illa) preferably corresponds to the following formula (Illb):
• the compound of formula (la) can, in yet another embodiment, correspond to the following formula (IVa):
• the compound (IVa) preferably corresponds to the following formula (IVb):
• the compound of formula (la) can correspond to the following formula (Va):
• the compound (Va) preferably corresponds to the following formula (Vb):
• the invention also relates to the precursor compounds of formulas (la), (lia), (Ilb), (Illa),
• the precursor compounds can be chosen, in particular, from the final compounds defined, listed above, in which the [ 18 F] is replaced by a non-radioactive halogen, such as 19 F, Cl, Br, I, a salt d ammonium, such as trimethylammonium-trifluoromethanesulfonate or an NO 2 group and the group l-pyrrole-2,5-dione is replaced by a tert-butoxycarbonylamino group.
• a non-radioactive halogen such as 19 F, Cl, Br, I
• a salt d ammonium such as trimethylammonium-trifluoromethanesulfonate or an NO 2 group
• the group l-pyrrole-2,5-dione is replaced by a tert-butoxycarbonylamino group.
• Preferred precursor compounds are, for example [3- (3-tert-butoxycarbonylamino-propoxy) - pyridin-2-yl] -trimethyl-ammonium, trifluoromethane sulfonate; and [3- (2-nitro-pyridin-3-yloxy) -propyl] -carbamic acid tert-butyl ester.
• the chemicals were obtained from various suppliers (ALDRICH, FLUKA or SIGMA France) and they were used without further purification, except when mentioned.
• the TLCs are carried out on plates coated, beforehand, with silica gel 60F 2 5 from MERCK.
• the compounds were located (1) if possible at 254 nm, using a UV lamp and / or (2) by iodine staining and / or (3) by soaking the TLC plates in an ethanolic solution at 1 % ninhydrin (or a 1% aqueous solution of KMn0 4 ) and by heating on a hot plate. Radioactive points, spots or "spots" are detected using a BERTHOLD TRACE MASTER 20 automatic linear analysis device.
• the NMR spectra are recorded on a BRUKER AMX device (300 MHz), using the hydrogenated residue of deuterated solvents (DMS0-d 6 , ⁇ : 2.50 ppm; CD 2 C1 2 , ⁇ : 5.32 ppm; CD 3 OD , ⁇ : 4.78; CD 3 CN, ⁇ : 1.93 ppm) and / or TMS as internal standards for 1 H NMR, and deuterated solvents (DMS0-d 6; ⁇ : 39.5ppm; CD 2 C1 2 , ⁇ : 53.8 ppm; CD 3 OD, ⁇ : 49.3 ppm) and / or TMS, as internal standards for 13 C NMR.
• deuterated solvents DMS0-d 6; ⁇ : 39.5ppm; CD 2 C1 2 , ⁇ : 53.8 ppm; CD 3 OD, ⁇ : 49.3 ppm
• MS are measured on a Quadripolair Finnigan 4 600 (DCI / NH 4 + ).
• Aqueous fluoride [ 18 F] ions were prepared in a CGR-MeV 520 cyclotron by irradiation of a 2 mL water target, using a 20 MeV proton beam on water enriched in [ 18 0 ] to 95% by the nuclear reaction [ 18 0 (p, n) 18 F].
• the fluoride ions were transferred to the appropriate shielded cell.
• the radiosyntheses using fluor-18, including the purifications by semi-preparative HPLC were carried out in a 7.5 cm cell shielded with lead, using a ZYMATE robot system controlled by computer (from ZYMACK CORP., USA) .
• activation in the microwave is carried out with a MICRO ELL 10 oven (2.45 GHz), supplied by LAB ELL AB, Sweden.
• the specific radioactivity is determined as follows: the area of the UV absorbance peaks, corresponding to the radiolabelled product, is measured on the HPLC chromatogram and compared to a standard curve giving the mass, as a function of the UV absorbance.
• Example 2 The procedure described above in Example 1 is used with the 2-fluoro-3-hydroxypyridine prepared in Example 4 (1.29 g; 11.4 mmol) to give 1.9 g (62%) of tertiary butyl ester of [3- (2-fluoro-pyridin-3-yloxy) -propyl] -carbamic acid in the form of a yellow oil, after “flash” chromatography (eluent: CH 2 C1 2 pure, heptane / EtOAc: 70/30 to 50/50). Rf (CH 2 CL 2 / EtOAc: 95/5): 0.45.
• Example 2 The procedure described above in Example 2 is used with the tert-butyl ester of [3- (2-fluoro-pyridin-3-yloxy) -propyl] - carbamic acid prepared in a) (1, 0 g; molecular weight: 270.30; 3.7 mmol) to give 1.4 g / 95%) of 3- (2-fluoro-pyridin-3-yloxy) - propylamine. 2 TFA, in the form of a yellow oil.
• Example 2 The procedure described above in Example 1 is used with 2-dimethylamino-3-hydroxypyridine (0.250 g; molecular weight: 138.17; 1.8 mmol) to give 0.290 g of the tert-butyl ester of [3- (2-dimethylamino-pyridin-3-yloxy) -propyl] -carbamic acid (58%) as a yellow oil after “flash” chromatography (eluent: heptane / EtOAc: from 70/30 to 50/50).
• the tertiary butyl ester of [3- (2-dimethylamino-pyridin-3-yloxy) -propyl] -carbamic acid is then diluted in toluene (2 ml per 100 mg of ester) and the solution is cooled to 0 ° C (ice bath). To this solution is added methanetrifluoromethanesulfonate (50 microliters for 100 mg of ester) and the reaction medium is stirred for 1 hour at 0 ° C.
• DMSO freshly distilled (600 ⁇ L), containing 4.0 to 6.0 mg of the marker precursor “nitro” (tert-butyl ester of [3- (2-nitro-pyridin-3-yloxy) -propyl acid) ] -carbamic) is added directly to the tube containing the dried K [ 18 F] -K 222 complex.
• the tube (unsealed) is then placed in a heating block (at 145 ° C for 4 minutes). The tube is then cooled using an ice / water bath and the remaining radioactivity is measured.
• the reaction mixture obtained, dark in color, is then analyzed by radiochromatography.
• the incorporation yields are calculated from the TLC radiochromatogram and are defined by the area ratio of the tert-butyl ester derivative of [3- (2- [ 18 F] fluoro-pyridin-3-yloxy) -propyl acid. ] - carbamic on the total activity of 18 F fluor-18 (Si0 2 -CCM; eluent: EtOAc; Rf:: 0.75 and Rf: fluoride ion [ 18 F]: 0.0).
• the reaction mixture is diluted with 1 ml of water and transferred to a C18 Sep-pak (waters) cartridge.
• the tube is rinsed twice with 1 mL of water, which is also transferred and added to the diluted reaction mixture on the cartridge.
• the assembly is then passed through the cartridge.
• the cartridge is washed with 3 ml of water and partly dried for 0.5 minutes, sending a stream of nitrogen.
• the tert-butyl ester derivative of • [3- (2- [ 18 F] fluoro-pyridin-3-yloxy) -propyl] - carbamic acid is eluted from the cartridge with 3 mL of dichloromethane in a reaction flask containing 0.1 mL of TFA. 2 ml of dichloromethane are used twice to wash the cartridge and to completely transfer the derivative labeled with [ 18 F] mentioned above (5% of the amount of total radioactivity, involved in the fluorination process, remains on the cartridge) .
• the incorporation yield is also confirmed after the elution of Sep-pak by the ratio of the count values of CH 2 C1 2 to total eluted radioactivity (DMSO / H 2 0 + CH 2 C1 2 ).
• the resulting CH 2 C1 2 / TFA solution (50/1, V / V) is concentrated to dryness (at 65-75 ° C) under a moderate stream of nitrogen for 4 to 6 minutes).
• the deprotection yield is quantitative: No molecule, described above, protected by BOC can be detected by radiochromatography.
• the residue, above, is redissolved in 2 mL of CH 2 C1 and concentrated again to dryness to minimize the presence of TFA (at 65-75 ° C under a moderate stream of nitrogen for 4 to 6 minutes).
• the residue is then diluted with 0.5 mL of xylene containing 25 mg of N-methoxycarbonylmaleimide.
• the container is then hermetically closed, heated for 5 minutes at 190 ° C (strong reflux), then cooled for 2 minutes, using an ice / water bath.
• the reaction mixture is then injected onto a column of semi-preparative HPLC. Isocratic elution [eluent: heptane / EtOAc: 50/50; flow rate: 6.0 mL / minute] which gives 1- (3- (2- [ 18 F] fluor-pyridin-3-yloxy) -propyl] -pyrrole-2, 5- labeled dione, pure, retention time: 7.5 to 8.0 minutes.
• 60 to 70 mCi of 1- [3- (2- [ 18 F] fluor-pyridin-3-yloxy) -propyl] -pyrrole-2, 5-dione labeled, pure can be obtained in 75 to 85 minutes , from 550-650 mCi of a production batch
• the compound labeled with fluorine-18, 1- [3- (2- [ 18 F] fluoro-pyridin-3-yloxy) -propyl] -pyrrole-2, 5-dione can also be prepared by repeating steps in) and b) of the process described in Example 7, still using as labeling precursor, the compound “nitro” (tert-butyl ester of [3- (2-nitro-pyridin-3-yloxy) -propyl] acid] -carbamic), but by modifying the final part of the preparation (step c)) in the following way (variant according to which step c) is carried out in a biphasic mixture of dioxane and aqueous sodium bicarbonate).
• the cartridge is washed with 3 mL of water and partially dried for 0.5 minutes, sending a stream of nitrogen.
• the derivative labeled with fluorine-18 (1- [3- (2- [ 18 F] fluoro-pyridin-3-yloxy) -propyl] -pyrrole-2, 5-dione) is eluted from the cartridge with 3 mL of dichloromethane in a new empty vial. 1 ml of dichloromethane is used twice to wash the cartridge and to completely transfer the derivative labeled with [ 18 F] mentioned above.
• the solution containing the derivative labeled with [ 18 F] mentioned above is concentrated (at 65-75 ° C, under a moderate stream of nitrogen for 3 to 5 minutes) to a volume of approximately 1 mL and injected on a column of semi-preparative HPLC.
• the purification is identical to that described in Example 7.
• the fluorine-18 labeled compound, 1- [3- (2- [ 18 F] fluoro-pyridin-3-yloxy) -propyl] -pyrrole-2,5-dione can also be prepared by repeating steps a) and b) of the process described in Example 7 or 7a, but using as labeling precursor, the compound "trimethylammonium-trifluoromethane sulfonate” (trifluoromethanesulfonate of [3- (3-tert- butoxycarbonylamino-propoxy) -pyridin-2-yl] -trimethyl- ammonium).
• a peptide namely the peptide N-acetyl-Lys-Ala-Ala-Ala-Cys-amide
• a compound according to the invention which is 1- [3 - (2- [ 18 F] fluor-pyridin-3-yloxy) -propyl] -pyrrole-2, 5-dione.
• the labeled peptide is purified by HPLC on column C18 with a gradient of 0 to 34% acetonitrile / 0.1% TFA in

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• Nuclear Medicine (AREA)
• Pyridine Compounds (AREA)
• Saccharide Compounds (AREA)
• Plural Heterocyclic Compounds (AREA)
• Peptides Or Proteins (AREA)

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• 2002
  • 2002-07-01 FR FR0208203A patent/FR2841554B1/fr not_active Expired - Fee Related
• 2003
  • 2003-06-30 WO PCT/FR2003/002028 patent/WO2004002984A2/fr not_active Ceased
  • 2003-06-30 AU AU2003259312A patent/AU2003259312A1/en not_active Abandoned
  • 2003-06-30 US US10/517,612 patent/US7456202B2/en not_active Expired - Fee Related
  • 2003-06-30 JP JP2004516901A patent/JP2006504640A/ja active Pending
  • 2003-06-30 EP EP03761670A patent/EP1517903A2/de not_active Withdrawn
  • 2003-06-30 CA CA2491193A patent/CA2491193C/fr not_active Expired - Fee Related

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