EP4565570A1 - Agents de contraste à haute relaxivité et préparation stéréosélective - Google Patents

Agents de contraste à haute relaxivité et préparation stéréosélective

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Publication number
EP4565570A1
EP4565570A1 EP23755294.8A EP23755294A EP4565570A1 EP 4565570 A1 EP4565570 A1 EP 4565570A1 EP 23755294 A EP23755294 A EP 23755294A EP 4565570 A1 EP4565570 A1 EP 4565570A1
Authority
EP
European Patent Office
Prior art keywords
compound
formula
independently
chelate
aryl
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
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EP23755294.8A
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German (de)
English (en)
Inventor
Mark Woods
Karley B. MAIER
Lauren N. RUST
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Portland State University
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Portland State University
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Application filed by Portland State University filed Critical Portland State University
Publication of EP4565570A1 publication Critical patent/EP4565570A1/fr
Pending legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D257/00—Heterocyclic compounds containing rings having four nitrogen atoms as the only ring hetero atoms
    • C07D257/02—Heterocyclic compounds containing rings having four nitrogen atoms as the only ring hetero atoms not condensed with other rings

Definitions

  • This disclosure concerns contrast agents for magnetic resonance imaging, as well as methods of stereoselectively preparing the contrast agents.
  • Contrast agents for magnetic resonance imaging are disclosed, along with methods of making and using the contrast agents. Aspects of the disclosed contrast agents are compounds according to formula IB or IA, or salts or stereoisomers thereof:
  • Each R 1 independently is an aryl or aliphatic group having an sp 2 -hybridized or sp-hybridized carbon at its attachment point to the carbon atom alpha to nitrogen; and Ln is a lanthanide ion.
  • each R 1 independently is aryl, alkenyl, or alkynyl.
  • each R 1 independently may be aryl substituted with one or more substituents R 2 , where each R 2 independently is -(CH2) n C(0)0R a , -(CH2) n 0R a , -(CH2) n 0C(0)R a , -(CH 2 )nC(O)R a , -(CH 2 )nC(S)R a , -(CH 2 ) n COR a , -(CH 2 ) n CSR a , -(CH 2 ) n SR a , -(CH 2 )nNO 2 , -(CH 2 ) n N(R b )R c , -(CH 2 )nC(O)N(R b )R c , -(CH 2 ) n C
  • a pharmaceutical composition include a compound according to formula IB and a pharmaceutically acceptable carrier.
  • the compound is a diastereomer and at least 70% of molecules of the compound independently are RRRR- or SSSS- enantiomers.
  • the compound is: or a combination thereof.
  • FIG. 1 is a X H NMR spectrum, focused on the most highly shifted ax s resonances, of ⁇ EuDOTMA recorded at 600 MHz in D2O at pD 6.
  • FIG. 2 shows the 1 H resonance of the chiral proton of R-m ethyl a -bromo phenylacetate R-2 prepared from J?rRhenylglycine in the absence (bottom, blue) and presence (top, red) of Eu(-)hfc3 at 400 MHz in CDCh.
  • FIG. 3 is a diagram showing all possible sequences of substitution in a racemic synthesis of a tetra-a-substituted DOTA derivative after the first alkylation step has afforded an R- configuration.
  • FIGS. 4A-4D show an analysis of chelates produced in the synthesis of EuDOTFA.
  • FIG. 4A is the chromatogram from the RP-HPLC separation of crude EuDOTFA (Cl 8 column, X - 254 nm).
  • FIG. 4B is the 3 H NMR spectrum (focused on the most shifted ax s protons) of the first peak to elute, which appears to be the RSRS- isomer.
  • FIG. 4C is the 3 H NMR spectrum (focused on the most shifted ax s protons) of the second peak to elute, which appears to be the RRRS-/SSSR- isomer.
  • FIG. 4D is the 3 H NMR spectrum (focused on the most shifted ax s protons) of the third peak to elute, which is the RRRR-/SSSS- isomer. NMR spectra were recorded at 400 MHz in D2O.
  • FIG. 5 shows the carbonyl region of the 13 C NMR spectrum (600 MHz, D2O) of HsDOTBA.
  • FIG. 6 shows 1 H NMR spectra (focused on the most shifted ax s region of the spectrum) of samples of EuDOTFA (left) and EuDOTBA (right) both prepared from ligand samples that contained a mixture of diastereoisomers.
  • FIG. 7 shows partial 'H NMR spectra (600 MHz) of EuDOTBA after incubation in D2O at pD 11, 60 °C focusing on the most shifted resonances of the SAP isomer: left, axial proton of the side carbon (ax s ) which serves as a control; right, the chiral methyne proton which was the anticipated point of alkylation.
  • FIG. 8 shows the up-field (right) and down-field (left) regions of the 1 H NMR spectra recorded during the chelation of Eu 3+ with ⁇ DOTBA in D2O at pD 13, recorded at 600 MHz.
  • a reference spectrum of the RRRR-/SSSS- isomer is shown (top).
  • FIG. 9 is a graph showing proportions of each stereoisomer produced as a function of the pH at which the chelation reaction was performed to provide EuDOTFA.
  • FIGS. 10A-10B show the temperature dependence of the 17 O reduced transverse relaxation rate constant of solvent water of solutions of GdDOTFA (FIG. 10 A) and GdDOTBA (FIG. 10B) at pH 6.6 and 11.7 T.
  • FIG. 11 shows 'H nuclear magnetic relaxation dispersion (NMRD) profiles, recorded at 298K of GdDOTFA (open circles) and GdDOTBA (closed diamonds). For reference, the fit of the 'H NMRD profile of GdDOTA is shown (dashed line).
  • FIGS. 12A and 12B are graphs showing relaxometric titrations at 298 K and 0.47 T of human serum albumin into 0.09 mM and 0.10 mM solutions of GdDOTFA (FIG. 12A) and GdDOBTA (FIG. 12B).
  • TR 100 ns
  • TR 2 ns
  • TM water exchange lifetime
  • FIGS. 15A and 15B show calculated relaxivity as a function of time for GdDOTA (FIG. 15 A) and GdDOTFA (FIG. 15B) incubated in 1 M HC1.
  • FIG. 16 shows chemical structures of Gd 3+ chelates used as contrast agents that are either in clinical trials or already in clinical use.
  • FIG. 17 shows the relaxivity (expressed in terms of the total relaxivity per molecule) of GdDOTBA at 1.5 T (top) and 3.0 T (bottom) at 310 K.
  • the relaxivities of clinically available contrast agents and another currently in trials under the same conditions are shown for comparative purposes.
  • FIG. 18 shows relaxivity (expressed in terms of the relaxivity per water molecule bound to Gd 3+ ) of GdDOTBA at 1.5 T (top) and 3.0 T (bottom) at 310 K.
  • the relaxivities of clinically available contrast agents and another currently in trials under the same conditions are shown for comparative purposes.
  • FIG. 19 shows NMR spectra of crude samples of four aryl -substituted EuDOTA chelates, focused on the most shifted axial proton resonance.
  • This disclosure concerns compounds having a structure according to formula IA or IB, or a salt or stereoisomer thereof: independently is an aryl or aliphatic group having an sp 2 -hybridized or sp-hybridized carbon at its attachment point to the carbon atom alpha to nitrogen, and Ln is a lanthanide ion.
  • the disclosed compounds may be useful as contrast agents, such as magnetic resonance imaging contrast agents.
  • Some stereoisomers of the disclosed chelated compounds according to formula IB have a high relaxivity. In certain instances, the relaxivity is close to a theoretical maximum value for the chelate.
  • Embodiments of a stereoselective synthesis for preparing the compounds also are disclosed.
  • Adduct A product resulting from an addition reaction between two or more molecules. For example, molecules A and B react to form the product AB.
  • Aliphatic A substantially hydrocarbon-based compound, or a radical thereof (e.g., Cr.His, for a hexane radical), including alkanes, alkenes, alkynes, including cyclic versions thereof (also referred to as cycloaliphatic), and further including straight- and branched-chain arrangements, and all stereo and position isomers as well.
  • an aliphatic group contains from one to twenty-five carbon atoms; for example, from one to fifteen, from one to ten, from one to six, or from one to four carbon atoms.
  • the term "lower aliphatic" refers to an aliphatic group containing from one to ten carbon atoms.
  • An alkyl group is a hydrocarbon group having a saturated carbon chain.
  • the chain may be cyclic, branched or unbranched. When the chain is cyclic, the group may be referred to as cycloalkyl. Examples, without limitation, of alkyl groups include methyl, ethyl, propyl, butyl, pentyl, hexyl, heptyl, octyl, nonyl and decyl.
  • the term lower alkyl means the chain includes 1-10 carbon atoms.
  • alkenyl and alkynyl refer to hydrocarbon groups having carbon chains containing one or more double or triple bonds, respectively; cyclic versions may be referred to a cycloalkenyl or cycloalkynyl, respectively.
  • An aliphatic chain may be substituted or unsubstituted. Unless expressly referred to as an “unsubstituted aliphatic,” an aliphatic group can either be unsubstituted or substituted.
  • substituents include, but are not limited to, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, alkylthio, acyl, aldehyde, amide, amino, aminoalkyl, aryl, arylalkyl, carboxyl, cyano, cycloalkyl, dialkylamino, halo, haloaliphatic, heteroaliphatic, heteroaryl, heterocycloaliphatic, hydroxyl, oxo, sulfonamide, sulfhydryl, thioalkoxy, or other functionality.
  • a substituted aliphatic group includes at least one sp 3 -hybridized carbon or two sp 2 -hybridized carbons bonded with a double bond or at least two sp-hybridized carbons bonded with a triple bond.
  • Aromatic Unsaturated, cyclic hydrocarbons having alternate single and double bonds. Benzene, a 6-carbon ring containing three double bonds, is a typical aromatic compound.
  • Aryl A monovalent aromatic carbocyclic group of, unless specified otherwise, from 6 to 15 carbon atoms having a single ring (e.g., phenyl) or multiple condensed rings in which at least one ring is aromatic (e.g., quinoline, indole, benzodioxole, and the like), provided that the point of attachment is through an atom of an aromatic portion of the aryl group and the aromatic portion at the point of attachment contains only carbons in the aromatic ring. If any aromatic ring portion contains a heteroatom, the group is a heteroaryl and not an aryl. Unless expressly referred to as an “unsubstituted aryl,” an aryl group can either be unsubstituted or substituted.
  • Chelate A coordination compound in which a central metal ion is attached by coordinate bonds to two or more nonmetal atoms of a single molecule or ligand.
  • the chelate may have a cyclic structure. As a verb, to form a chelate.
  • Coordination compound A compound formed by a metal ion bonded to a non-metallic ion or molecule called a ligand.
  • a coordinate bond is a covalent bond consisting of a pair of shared electrons donated by the ligand.
  • Effective amount with respect to a compound or composition refers to an amount of the compound or composition sufficient to achieve a particular desired result, e.g., detection during an imaging process.
  • Isomer One of two or more molecules having the same number and kind of atoms, but differing in the arrangement or configuration of the atoms. Isomers that differ in the arrangement of their atoms in space are termed “stereoisomers”. Stereoisomers that are not mirror images of one another are termed “diastereomers” and those that are non-superimposable mirror images of each other are termed “enantiomers.” When a compound has an asymmetric center, for example, if a carbon atom is bonded to four different groups, a pair of enantiomers is possible.
  • An enantiomer can be characterized by the absolute configuration of its asymmetric center and is described by the R- and S-sequencing rules of Cahn and Prelog, or by the manner in which the molecule rotates the plane of polarized light and designated as dextrorotatory or levorotatory (z.e., as (+) or (-) isomers respectively).
  • a chiral compound can exist as either individual enantiomer or as a mixture thereof.
  • a mixture containing equal proportions of the enantiomers is called a “racemic mixture.”
  • the term “coordination isomers” refers to isomers that differ in the geometry around a central coordinated atom.
  • embodiments of the disclosed chelates may be coordination isomers having a square antiprism (SAP) or twisted square antiprism (TSAP) geometry around a coordinated lanthanide ion.
  • Macromolecule refers to a large molecule having a largest dimension (e.g., diameter) greater than 0.5 nm.
  • compositions that can be taken into a subject without significant adverse toxicological effects on the subject.
  • compositions and formulations suitable for pharmaceutical delivery of one or more compositions are conventional.
  • Remington The Science and Practice of Pharmacy, The University of the Sciences in Philadelphia, Editor, Lippincott, Williams, & Wilkins, Philadelphia, PA, 21 st Edition (2005), describes compositions and formulations suitable for pharmaceutical delivery of one or more compositions.
  • the nature of the carrier will depend on the particular mode of administration being employed.
  • parenteral formulations usually comprise injectable fluids that include pharmaceutically and physiologically acceptable fluids such as water, physiological saline, balanced salt solutions, aqueous dextrose, glycerol or the like as a vehicle.
  • the pharmaceutically acceptable carrier may be sterile to be suitable for administration to a subject (for example, by parenteral, intramuscular, or subcutaneous injection).
  • pharmaceutical compositions to be administered can contain minor amounts of non-toxic auxiliary substances, such as wetting or emulsifying agents, preservatives, and pH buffering agents and the like, for example sodium acetate or sorbitan monolaurate.
  • the pharmaceutically acceptable carrier is a non-naturally occurring or synthetic carrier.
  • the carrier also can be formulated in a unit-dosage form that carries a preselected therapeutic dosage of the active agent, for example in a pill, vial, bottle, or syringe.
  • Substituent An atom or group of atoms that replaces another atom in a molecule as the result of a reaction.
  • the term “substituent” typically refers to an atom or group of atoms that replaces a hydrogen atom, or two hydrogen atoms if the substituent is attached via a double bond, on a parent hydrocarbon chain or ring.
  • the term “substituent” may also cover groups of atoms having multiple points of attachment to the molecule, e.g., the substituent replaces two or more hydrogen atoms on a parent hydrocarbon chain or ring. In such instances, the substituent, unless otherwise specified, may be attached in any spatial orientation to the parent hydrocarbon chain or ring.
  • substituents include, for instance, alkyl, alkenyl, alkynyl, alkoxy, alkylamino, alkylthio, acyl, aldehyde, amido, amino, aminoalkyl, aryl, arylalkyl, arylamino, carbonate, carboxyl, cyano, cycloalkyl, dialkylamino, halo, haloaliphatic (e.g., haloalkyl), haloalkoxy, heteroaliphatic, heteroaryl, heterocycloaliphatic, hydroxyl, oxo, sulfonamide, sulfhydryl, thio, and thioalkoxy groups.
  • alkyl alkenyl, alkynyl, alkoxy, alkylamino, alkylthio, acyl, aldehyde, amido, amino, aminoalkyl, aryl, arylalkyl, arylamino, carbonate
  • a fundamental compound such as an aryl or aliphatic compound, or a radical thereof, having coupled thereto one or more substituents, each substituent typically replacing a hydrogen atom on the fundamental compound.
  • a person of ordinary skill in the art will recognize that compounds disclosed herein may be described with reference to particular structures and substituents coupled to such structures, and that such structures and/or substituents also can be further substituted, unless expressly stated otherwise or context dictates otherwise.
  • a substituted aryl compound may have an aliphatic group coupled to the closed ring of the aryl base, such as with toluene.
  • a long-chain hydrocarbon may have a hydroxyl group bonded thereto.
  • the contrast agents are useful for magnetic resonance imaging.
  • the contrast agents may exhibit high relaxivity, e.g., > 6 rnNf's’ 1 at a nuclear magnetic resonance field strength of 0.5 T to 7 T.
  • the disclosed compounds have a structure according to formula IA or IB, or a salt or stereoisomer thereof: where each R 1 independently is an aryl or aliphatic group having an sp 2 -hybridized or sp-hybridized carbon at its attachment point to the carbon atom alpha to nitrogen; and Ln is a lanthanide ion.
  • the compound is optically active.
  • each R 1 independently may be aryl, alkenyl, or alkynyl. Each R 1 may be substituted or unsubstituted. R 1 has an sp 2 -hybridized or sp-hybridized carbon at its attachment point to the carbon atom alpha to nitrogen. For example, if
  • R 1 is butenyl, it may have a formula propynyl, it has a formula
  • R 1 is substituted or unsubstituted aryl, such as substituted or unsubstituted phenyl, or substituted or unsubstituted naphthalenyl.
  • R 1 is aryl substituted with one or more substituents R 2 , where each R 2 independently is -(CH 2 ) n C(O)OR a , -(CH 2 )nOR a , -(CH 2 )nOC(O)R a , -(CH 2 ) n C(O)R a , -(CH 2 ) n C(S)R a , -(CH 2 ) n COR a , -(CH 2 ) n CSR a , -(CH 2 ) n SR a , -(CH 2 ) n NO 2 , -(CH 2 ) n N(R b )R c , -(CH 2 )
  • n is 0, and R 2 is -C(O)OR a , -OR a , -OC(O)R a , -C(O)R a , -C(S)R a , -COR a , -CSR a , -SR a , -NO 2 , -N(R b )R c , -C(O)N(R b )R c , -C(S)N(R b )R c , -CN, alkyl, alkenyl, alkynyl, or halo.
  • each R 2 independently is -C(O)OR a , OR a , alkyl, -SR a , -NO 2 , or halo.
  • each R a , R b , and R c independently is H or C1-C3 alkyl.
  • an alkyl, alkenyl, or alkynyl group may be substituted or unsubstituted unless otherwise specified.
  • R 1 is unsubstituted phenyl, phenyl substituted with one or more R 2 groups, or unsubstituted naphthalenyl.
  • R 2 groups include, but are not limited to, -C(O)OR a -OR a , and alkyl, where R a is H or C 1 -C 3 alkyl.
  • R 2 is -COOH, -OCH 3 , -CF 3 , -COOCH 3 ,
  • R 1 is unsubstituted aryl or aryl substituted with one R 2 group.
  • the aryl group is a phenyl group or a naphthalenyl group.
  • R 1 is or unsubstituted phenyl. In particular examples, R 1 is In some embodiments, each R 1 is the same.
  • Ln is a lanthanide ion, such as Ln 3+ .
  • the lanthanides include lanthanum and the elements with atomic numbers 58-71.
  • Ln is Gd or Eu.
  • Ln is Gd, and the compound according to formula IB is a Gd 3+ chelate.
  • Ln is Gd or Eu.
  • Embodiments of the disclosed compounds are stereoisomers with stereochemistry at the carbon atoms bonded to R 1 (i.e., stereochemistry at the carbon atoms alpha to the nitrogen atoms). With four stereocenters, RRRR-, SSSS-, RRRS-, SSSR-, RSRS-, and RRSS- isomers are possible.
  • the compound is a diastereomer, and molecules of the diastereomer comprise a mixture of RRRR- and SSSS- enantiomers.
  • the compound may be:
  • a solution of the compound in water may have a relaxivity n > 6 mM ⁇ s' 1 at a nuclear magnetic resonance field strength of 0.5 T to 7 T and 310K.
  • a solution of the compound in water has a relaxivity ri of > 6 mM’ 1 s’ 1 , > 7 mM 4 s 4 , or > 8 mM 4 s 4 at 0.5 T to 7 T and 310K, such as a relaxivity of 6 mM 4 s 4 to 15 mM 4 s 4 , 7 mM 4 s 4 to 12 mM 4 s 4 , or 8 mM 4 s 4 to 10 mM 4 s 4 at 0.5 T to 7 T and 310K.
  • a pharmaceutical composition comprises a compound according to formula IB and a pharmaceutically acceptable carrier.
  • the compound in the pharmaceutical composition may be a diastereoisomer with at least 70% of the compound molecules independently being RRRR- or SSSS- enantiomers. In some implementations, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% of the molecules independently are RRRR- or SSSS- enantiomers.
  • the pharmaceutically acceptable carrier may be any conventional carrier.
  • the pharmaceutical composition is formulated for use as a magnetic resonance imaging contrast agent.
  • the pharmaceutical composition may be formulated for parenteral administration.
  • the pharmaceutical composition is an injectable composition, and the compound according to formula IB is dissolved or suspended in the pharmaceutically acceptable carrier.
  • Embodiments of the disclosed contrast agents may be suitable for diagnostic imaging, such as magnetic resonance imaging.
  • a method for magnetic resonance imaging may include administering to a subject an effective amount of a pharmaceutical composition comprising a compound according to formula IB, and performing a magnetic resonance imaging procedure on the subject.
  • GdDOTFA shown below
  • GdDOTFA when bound to human serum albumin, achieved theoretical maximum relaxivity of 110 mM’ 1 s’ 1 , and is believed to be the first chelate to
  • Embodiments of compounds according to formula IB may be stereoselectively synthesized.
  • R 1 may be aryl, alkenyl, or alkynyl, and may be substituted or unsubstituted.
  • R is a C 1 -C 4 alkyl. In some implementations, R is methyl, ethyl, or isopropyl.
  • X is halo, e.g., Br, Cl, or I. In certain implementations, X is Br.
  • R 1 is alkenyl
  • an alkylating agent e.g., where R' is a tritiate, tosylate, or mesylate group, may be utilized in place of the halogenated starting material, with the remainder of synthesis proceeding as shown above.
  • a glycine is halogenated to provide the corresponding halogenated glycine, where X is Br, Cl, or I.
  • the halogenated glycine is esterified to provide the alkylating agent, .
  • the glycine is combined with a halide salt (e.g., NaBr, NaCl, or Nal) in a hydracid (HBr, HC1, or HI) and cooled to 0 °C. Sodium nitrite is gradually added, the solution is warmed to room temperature (e.g., 20-25 °C), and stirred until halogenation is complete.
  • a halide salt e.g., NaBr, NaCl, or Nal
  • a base e.g., KOH
  • a base e.g., KOH
  • the reaction mixture is extracted with diethyl ether.
  • the aqueous layer is acidified and extraction is repeated.
  • the combined extracts are dried and the solvents removed to provide the halogenated glycine.
  • a Fisher esterification is used to esterify the halogenated glycine.
  • the halogenated glycine may be dissolved in acidified methanol and heated with stirring until the halogenated glycine is esterified to form the alkylating agent.
  • a base e.g., K 2 CO 3
  • the extracts are dried and the solvents removed to provide the alkylating agent.
  • a carboxylic acid is esterified to provide
  • esterified compound is then halogenated to provide the alkylating agent, .
  • esterification may provide a diester .
  • the carboxymethyl compound is dissolved in a C 1 -C 4 alkanol, acidified, and reacted under effective conditions to provide an esterified compound .
  • the reaction proceeds under reflux with stirring for 12-24 hours, such as for 18 hours.
  • the solvents are removed, and the residue is purified by extraction with water and diethyl ether.
  • the esterified compound is recovered from the organic extract.
  • esterified compound is brominated by reaction with A-bromosuccinimide in acetonitrile to provide the alkylating agent, Alternatively, the esterified compound may be reacted with A-chlorosuccinimide or
  • the reaction proceeds under ultraviolet irradiation (72-watt UV, 365-395 nm) with heating at 50 °C to 70 °C for 3-5 days, such as at 60 °C for four days.
  • the solvent is removed.
  • the residue is dissolved (e.g., in diethyl ether), filtered, washed, and purified, such as by column chromatography.
  • the alkylating agent is prepared by hydroxylating an esterified alkene (e.g., with monoperoxyphthalate, MMPP) to introduce a hydroxyl group and then converting the hydroxyl group to a triflate:
  • an esterified alkene e.g., with monoperoxyphthalate, MMPP
  • the esterified alkene is reacted to introduce a hydroxyl group as shown, and the hydroxyl group is then converted to a tosylate or mesylate group.
  • the alkylating agent (e.g., when R 1 is alkenyl), is reacted with embodiments, the alkylating agent is reacted with the cyclen in acetonitrile and cesium chloride under effective conditions to provide the alkylated cycle.
  • the effective conditions may include a temperature of from 50 °C to 70 °C and a reaction time, with stirring, of from 18-30 hours, such as 20-25 hours. Following the reaction, the solvents are removed.
  • the residue is dissolved (e.g., in dichloromethane), and washed with water or a carbonate solution (e.g., K2CO3, pH 12-14).
  • the aqueous layer is extracted (e.g., with dichloromethane).
  • the organic layers are combined and dried, and the solvents are removed.
  • the residue is purified, e.g., by column chromatography) to provide an alkylated cy cl en.
  • the alkylated cyclen is saponified to remove R and provide a compound according to Saponification may be performed by combining the alkylated cyclen in an organic solvent (e.g., tetrahydrofuran) with an aqueous base, such as NaOH or KOH, under conditions effective to remove R.
  • saponification is performed at 50 °C to 70 °C with stirring for 12-60 hours, such as 16-20 hours or 45-50 hours.
  • the organic solvent is removed by evaporation, and the water is removed (e.g., by lyophilization) to yield the compound according to formula IA.
  • the reaction mixture may be acidified (e.g., to pH 3) prior to water removal.
  • a lanthanide (Ln) salt, oxide, or tritiate is chelated with the compound according to formula IA to provide a chelated compound according to Formula IB,
  • Ln is Gd, Eu, or a combination thereof. In particular examples, Ln is Gd.
  • Chelation is performed by combining the compound according to formula IA and the lanthanide salt, oxide, or triflate in aqueous solution. The reaction proceeds under effective conditions to provide the chelated compound according to Formula IB. In some examples, the effective conditions include a temperature 65 °C to 75 °C, with stirring for 36-96 hours.
  • the lanthanide salt is a lanthanide chloride
  • the pH is adjusted to be slightly acidic (e.g., pH 5-6)
  • the effective conditions include a temperature of 70 °C with stirring for 48 hours, followed by cooling and filtration.
  • the compound according to formula IA is combined with a lanthanide oxide, and the reaction is heated at 70 °C with stirring for 72 hours, followed by neutralization and filtration.
  • the compound according to formula IA is combined with a lanthanide triflate, the pH is adjusted to be slightly acidic (e.g., pH 5-6), and the effective conditions include a temperature of 70 °C with stirring for 48 hours, followed by cooling and filtration.
  • the chelate according to formula IB comprises a diastereomer, and molecules of the diastereomer comprise a mixture of RRRR- and SSSS- enantiomers. In some implementations, at least 75%, at least 80%, at least 85%, at least 90%, or at least 95% of the compound molecules independently are RRRR- or SSSS- enantiomers.
  • DOTA DOTA itself is one such acronym: L4.7.10-TetraazacycloDOdecane- L4.7.10-TetraAcetate.
  • DOTMA was one of the first derivatives of DOTA to be reported. Its acronym denotes four acetate arms by the presence of the T (denoting tetra) and the A (denoting acetate. Note: A does not denote acetic acid: in the ligand the carboxylate is deprotonated). That these acetates are substituted is indicated by the M that separates the T and the A.
  • DOTF 3 A has four acetate arms, three of which are substituted with phenyl groups. In this way multiple different substituents may be noted.
  • DOTM 3 B 1 A would have four acetate arms, three of which are substituted with methyl groups with the fourth substituted by a benzoate.
  • substituents can be denoted as laid out about, e.g., D03MA (three acetate arms each with a methyl substituent), D02F 1 A (two acetate arms, one of which has a phenyl substituent).
  • substituents could be (and in some cases have been) denoted prior to the “DOTA” acronym. Examples include TCE- DOTA and NB-DOTA, the former denoting four a-carboxyethyl groups and the latter denoting a single nitrobenzyl substituent on a carbon of the macrocycle.
  • Chelates of both DOTFA and DOTBA were purified by eluting with water (0.037 % w/w HC1) for 5 minutes followed by a linear gradient to 80 % acetonitrile at 15 minutes with a flow rate of 50 mLmin' 1 .
  • the eluent was maintained at 20 % water and 80 % acetonitrile for a further 7 minutes. In all cases absorbance was monitored at 205 and 254 nm.
  • T H and 13 C NMR spectra were acquired on a Bruker® Avance® Ila spectrometer (Bruker Corporation, Billerica, MA) operating at either 400.13 and 100.61 MHz, respectively, or a Bruker® Avance® III NMR spectrometer operating at 600.17 MHz and 150.93 MHz, respectively. Melting points were determined on a Bibby Scientific SMP10 (Bibby Scientific Ltd, Stone, UK). The rotation of plane polarized light was measured on a Rudolph Research Analytical Autopol® 1 (Rudolph Research Analytical, Hackettstown, NJ).
  • Infrared spectra we measured on a ThermoScientific Nicolet® iSlO FTIR spectrometer (Thermo Fisher Scientific, Inc., Waltham, MA) equipped with a total attenuated reflectance sample holder. Mass spectra were measured on a ThermoScientific LTQ-Orbitrap® Discovery mass spectrometer equipped with an Accula® autosampler (Thermo Fisher Scientific, Inc., Waltham, MA).
  • DOTMA refers to the RRRR- enantiomer (Aime et al., Inorg. Chem. 2011, 50:7955-7965).
  • the stereospecificity of each synthetic approach to DOTMA has not been tested - some purification steps (especially crystallization) may remove traces of unwanted diastereoisomers. Nonetheless, diastereoisomerically pure chelates can be readily obtained.
  • ⁇ DOTMA was prepared using racemic 2-bromo propionic acid as shown in Scheme 1. Reagents and Conditions: i. /-butanol / cat. H2SO4 / MgSO4; ii. cyclen / CS2CO3 / MeCN; iii. TFA / 40 °C; iv. EuCL / H2O / pH 5.5.
  • Carboxylates were protected as t-butyl esters and although the tetra-t-butyl protected ligand was subjected to column purification, this technique was not thought to have removed of one or more diastereoisomer from the mixture.
  • the t-butyl esters were removed in trifluoroacetic acid and Eu 3+ introduced under standard conditions (60 °C, pH 5.5).
  • FIG. 3 shows the substitution sequences that lead to each isomer when the first alkylation step has afforded an A’-configuration.
  • the mirror image of these pathways exists for when the first alkylation yields an S- configuration.
  • R and S are used to denote the absolute stereochemistry at the a-position of a pendant arm substituent. Substitutions resulting in an Rconfiguration are shown proceeding to the left, those resulting in an S- configuration are shown proceeding to the right.
  • RRRS-/SSSR- isomer obtained is only slightly higher than the statistically predicted amount. The difference is made up by a larger than expected amount of the RSRS- isomer. But only two pathways afford the RSRS- isomer. If over-production of RSRS- arises primarily through a sequence involving trcms-R,R- (upper left pathway), a concomitant decrease the production of both RRRR-/SSSS- and RRRS-/SSSR- would be expected. But this is not observed. This implies that the sequences through cis- substitution (lower right pathway) that directly competes with RRSS- production must play a significant role in the distribution of isomers.
  • Scheme 2 shows the synthetic route for the preparation of an enantiomerically pure alkylating agent to introduce phenyl groups into the a-position of DOTA (top).
  • Reagents and conditions' i. NaNO2/HBr/KBr; ii. MeOH/FLSO ⁇ iii. R-2, ⁇ 2 or 4/Cs2CO3/MeCN/60 °C; iv. KOH followed by HC1 (pH 3).
  • R-2 was used to tetra-alkylate cyclen in acetonitrile with caesium carbonate at 60 °C. To permit the evaluation of the stereochemical selectivity of this synthesis the esters were removed by saponification without performing extensive purification.
  • the rare earth chelates of DOTFA were prepared in aqueous solution at pH 5.5 at 70 °C from the corresponding chloride or from oxide without adjustment of the pH.
  • Peak 1 is found to possess C2-symmetry and was assigned as the RSRS- isomer
  • peak 2 exhibits no symmetry and, based on the isomeric distribution observed for ⁇ EuDOTMA, was assigned to the SSSR- isomer.
  • the alkylating agent R-2 was confirmed to be enantiopure (FIG. 2) and so the production of unintended diastereoisomers may be attributable to a small degree of racemization during the alkylation reaction.
  • the synthesis of DOTFA chelates employed enantio-pure starting materials in procedures comparable to those used to obtain single enantiomers of other similar chelates (Kang et al., Inorg. Chem.
  • DOTFA is distinguished from other tetra-a-substituted chelates by its aryl substituents, which can potentially stabilize enolate intermediate facilitating deracemization under mild conditions.
  • aryl substituents can potentially stabilize enolate intermediate facilitating deracemization under mild conditions.
  • bulky a-substituents will tend to drive the orientation of the pendant arms into the lowest energy diastereoisomer: RRRR-/SSSS-. Because this is a thermodynamically driven process the preferred isomer is not expected to be the same as that in the study of ⁇ EuDOTMA.
  • the ligands of DOTFA and DOTBA were prepared in racemic form.
  • a suitable alkylating agent to produce DOTBA was prepared from an achiral source. 4-(Carboxymethyl) benzoic acid was protected as a diethyl ester also by Fisher esterification to afford 3 (Scheme 3).
  • the benzylic carbon was then brominated with ABS in MeCN at 60 °C with constant irradiation at 365 nm - 395 nm to afford ⁇ 4 after column chromatography. Cyclen was then exhaustively alkylated using either ⁇ 2 or ⁇ 4 in acetonitrile with caesium carbonate at 60 °C (Scheme 2).
  • RRRR-/SSSS- is the most thermodynamically stable isomer since this is the only isomer with congruence on all four pendant arms between the configuration at carbon and the helicity of the pendant arms. Racemization at the a-carbon leads to the conversion of the other isomeric chelates to the most thermodynamically stable isomer. What is more difficult to understand is why SSSS- H4DOTFA should convert into A’A’A’A’-EuDOTFA. To do so the chelate must pass through two other isomers both of which are higher in energy than the starting chelate.
  • the mechanism of this H/D exchange - formation of an enolate at the a-carbon - is the same as that expected in deracemization.
  • the metal ion interacts only with the pendant arms and the cyclen ring is diprotonated.
  • the rate determining step the protons are removed from the cyclen ring, allowing the metal ion to drop down into the coordination cage of the ligand forming the final chelate.
  • the second step occurs easily at moderate pH, but occasionally unusually high pHs are required to deprotonate the cyclen ring - this is most common when ligands contain potential ligating groups in peripheral positions (Kalman et al., Inorg. Chem. 2007, 46:5260-5270). The requirement for a high pH when chelating DOTBA is perhaps not surprising in this context.
  • Type I complexes appear to bind cooperatively (Stenson et al., Dalton Trans 2006, 3291-3293), and so the same relationship between helicity and stereochemical configuration is expected. This suggests that deracemization could occur in the Type I intermediate.
  • a solution of the DOTBA and EuCE in D 2 O at pD 13 (NaOD) was incubated in a sealed NMR tube at 60 °C for 72 hours followed by 120 hours at 100 °C. Reaction progress was monitored periodically by 'H NMR spectroscopy.
  • each sample was lyophilized and redissolved in D2O.
  • the 'H NMR spectrum of each was recorded with pre- saturation suppression of the residual acetate buffer peak.
  • the amount of each isomer present was determined by the line-fitting procedure described above.
  • the desired RRRR-/SSSS- isomer was the predominant reaction product.
  • this isomer made up 80 % of the produced EuDOTFA.
  • the proportion of the RRRR-/SSSS- isomer increased, eventually rising to 91 % when the reaction was performed at pH 4 (FIG. 9).
  • RSRS- and RRRS-/SSSR- were produced in measurable quantities. Only 1 - 2 % of the RSRS- isomer was produced, the quantity of this isomers seemingly largely unaffected by pH. In contrast the amount of RRRS-/SSSR- isomer produced exhibited a clear decrease as the pH was increased.
  • the rate determining step for inversion of the a-stereocenter in the Type I complex is the formation of the enolate (designated by the rate constant ki).
  • Conversion of the Type I complex to the Type II complex is the rate determining step of the chelation reaction (designated by the rate constant ki) (Toth et al.. Inorg. Chem. 1994, 33:4070-4076). This process involves removing two protons from the cyclen ring and is intrinsically pH dependent.
  • the difference in isomeric distribution between EuDOTFA and EuDOTBA is larger than any change affected by change in conditions.
  • the benzoate carboxylate is the only feature that distinguishes DOTBA from DOTFA.
  • the NMR spectrum of EuDOTBA has peaks that were attributed to the residual Type I complex, this implies that the peripheral carboxylate may cause a decrease in ki by stabilizing the Type I intermediate (Kalman et al., Inorg. Chem. 2007, 46:5260-5270). This would increase the lifetime of the intermediate, affording more time for the inversion of stereochemistry to occur and possibly accounting for the almost complete deracemization of EuDOTBA.
  • the Gd 3+ chelates used as MRI contrast agents are notoriously inefficient, requiring high doses: typically 1.0 - 1.5 g per dose.
  • the relaxivity of a chelate depends upon the number of solvent water molecules that can coordinated directly to the Gd 3+ (q), the distance of their protons from the metal (rcdu), the rate at which they exchange with the bulk solvent (1/TM) as well as the rate at which the chelate tumbles (1/TR) and the electronic relaxation parameters zl 2 and TV.
  • the contrast agents currently in use are sub-optimal: exchanging water too slowly and tumbling too rapidly (Sherry et al., Curr. Opin. Chem. Biol. 2013, 17: 167-174).
  • GdDOTFA and GdDOTBA were prepared as described above.
  • the relaxometric analyses described herein were performed using previously described instrumentation and methods (Webber et al., Inorg. Chem. 2020, 59:9037-9046).
  • GdDOTA itself has sub-optimal water exchange kinetics (Aime et al., Inorg. Chem. 2011, 50:7955-7965).
  • DOTA chelates are found to adopt both square antiprismatic (SAP) and twisted square antiprismatic (TSAP) coordination geometries. This is of significance because water exchange in TSAP isomers is found to be up to 100* faster than the SAP isomer (Woods et al., Angew. Chem. In. Ed. 2003, 5889-5892; Aime et al. , Angew. Chem. Int. Ed. 1998, 37:2673-2675; Woods et al., J. Am. Chem. Soc.
  • GdDOTA predominates as the SAP isomer, increasing the proportion of TSAP isomer is a strategy for improving water exchange kinetics (Woods et al., ANgew. Chem. In. Ed. 2003, 5889-5892; Dai et al., Nat. Commun. 2018, 9:857).
  • Substituting the a -position of the pendant arms of a DOTA chelate as shown above was found to increase the TSAP/SAP ratio (298 K) from 1 :4 (EuDOTA) to 7: 1 (EuDOTFA) and 10: 1 (EuDOTBA).
  • GdDOTFA and GdDOTBA can be obtained from a quantitative analysis of the nuclear magnetic relaxation dispersion (NMRD) profiles of these chelates which measure relaxivity as a function of the applied magnetic field (Bo).
  • FIG. 11 shows the NMRD profiles at 298 K of both GdDOTFA (open circles) and GdDOTBA (closed diamonds).
  • the fit of the NMRD profile of GdDOTA is shown (dashed line).
  • the profiles are notable for several reasons.
  • the relaxivity at all fields is significantly higher than that of GdDOTA, this can be attributed to the larger size of these chelates - slower rotational tumbling (longer TR). At low fields this difference is especially pronounced: indicative of more favourable electronic relaxation properties.
  • HSA Human serum albumin
  • HSA was titrated into solutions of GdDOTFA (0.09 mM) and GdDOTBA (0.1 mM), and binding was monitored by relaxometry at 0.5 T and 298 K (FIGS. 12A and 12B). Fitting these data using a simple 1 : 1 binding model shows that the binding of both chelates to HSA was quite weak.
  • the two chelates differ primarily in the peripheral carboxylates of GdDOTBA; presumably these groups increase chelate hydrophilicity thereby decreasing the hydrophobic interactions between chelate and protein.
  • GdDOTBA The relaxivity of GdDOTBA compares very favourably with established clinical contrast agents (Laurent et al., Contrast Media Mol Imaging 2006, 1 : 128-137), between 2 and 3 times higher (FIGS. 16-18).
  • gadopiclenol and gadoquatrane - have relaxivities (Robert et al., Radiology 2020, 294: 117-126; Lohrke et al., Invest. Radiol.
  • Gadopiclenol employs a heptadenate ligand opening a second coordination site to water, a strategy that was once considered risky for a chelate that is to be used in vivo because of the risk of compromising chelate robustness.
  • Gadoquatrane increases the number of Gd 3+ ions from 1 to 4, effectively quadrupling the dose. Neither of these chelate meet the criteria of a Gd 3+ chelate that has one site open for coordination by water.
  • GdDOTBA as a discrete chelate, exhibits unprecedentedly high relaxivity even at the higher fields (1.5 T and 3.0 T) typically used in clinical MRI. These relaxivity gains are all achieved while preserving octadentate coordination of the DOTA framework - these chelates are expected to retain the robustness (and thus safety) associated with DOTA chelates.
  • some compounds are prepared by a modified procedure using the chemical initiator AIBN in place of UV-irradiation.
  • This alternative procedure may be useful when Ar is alkoxy-substituted phenyl, resulting in selective bromination primarily at the benzylic position as desired.
  • R 1 when R 1 is an alkenyl group, the compound may be synthesized by functionalizing with a hydroxide instead of a halide.
  • MMPP introduces an a-hydroxyl, is then converted to a triflate, tosylate, or mesylate.
  • MMPP is magnesium monoperoxyphthalate.

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Abstract

La divulgation concerne un composé ayant une structure selon la formule IA ou IB, ou un sel ou un stéréoisomère de celui-ci : (IA) ou (IB), chaque R1 étant indépendamment un groupe aryle ou aliphatique ayant un carbone sp2- hybridé ou sp-hybridé à son point de liaison à l'atome de carbone alpha par rapport à l'azote et Ln est un ion lanthanide. Le groupe aryle ou aliphatique peut être substitué ou non substitué. Les composés divulgués peuvent être utiles en tant qu'agents de contraste, tels que des agents de contraste d'imagerie par résonance magnétique. Certains stéréoisomères des composés chélatés selon la formule IB ont une relaxivité élevée. Dans certains cas, la relaxivité est proche d'une valeur maximale théorique pour le chélate. La divulgation concerne également une synthèse stéréosélective pour la préparation des composés.
EP23755294.8A 2022-08-01 2023-07-24 Agents de contraste à haute relaxivité et préparation stéréosélective Pending EP4565570A1 (fr)

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