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WO1995020590A1 - Recepteurs enantioselectifs pour derives d'acide amine et autres composes - Google Patents

Recepteurs enantioselectifs pour derives d'acide amine et autres composes Download PDF

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Publication number
WO1995020590A1
WO1995020590A1 PCT/US1995/000948 US9500948W WO9520590A1 WO 1995020590 A1 WO1995020590 A1 WO 1995020590A1 US 9500948 W US9500948 W US 9500948W WO 9520590 A1 WO9520590 A1 WO 9520590A1
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Prior art keywords
composition
compound
mixture
purified
complex
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PCT/US1995/000948
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English (en)
Inventor
W. Clark Still
Julian A. Simon
Shawn D. Erickson
Seung Soo Yoon
Allen Borchardt
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The Trustees Of Columbia University In The City Of New York
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Priority claimed from US08/188,146 external-priority patent/US5599926A/en
Application filed by The Trustees Of Columbia University In The City Of New York filed Critical The Trustees Of Columbia University In The City Of New York
Priority to AU16894/95A priority Critical patent/AU1689495A/en
Publication of WO1995020590A1 publication Critical patent/WO1995020590A1/fr

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/22—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains four or more hetero rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B57/00—Separation of optically-active compounds
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07B—GENERAL METHODS OF ORGANIC CHEMISTRY; APPARATUS THEREFOR
    • C07B63/00—Purification; Separation; Stabilisation; Use of additives
    • C07B63/04—Use of additives
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D513/00—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00
    • C07D513/12—Heterocyclic compounds containing in the condensed system at least one hetero ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for in groups C07D463/00, C07D477/00 or C07D499/00 - C07D507/00 in which the condensed system contains three hetero rings
    • C07D513/18—Bridged systems

Definitions

  • This invention relates to the field of molecular recognition of small ligands. More particularly, the invention relates to compositions useful for the purification of enantiomers of amino acid derivatives and for the purification of certain compounds able to form hydrogen bonds, methods for preparing these compositions, and methods for using them.
  • Standard approaches to the optical resolution and purification of organic and biological molecules include crystallization, distillation, extraction, and chromatography (Eliel, Stereochemistry of Carbon Compounds, New York: McGraw-Hill, 1962).
  • Each methodology is based on a physical or chemical interaction of a molecule with an element of its environment, and may involve molecular sizing, electrostatics, hydrophobicity, sterics, or polarity.
  • the efficiency of purification increases as the differences in interaction energy for all the species present in the mixture increase.
  • the relevant interactions for cystallization are crystal lattice forces and solvation of the molecule; for distillation, the interaction is a liquid-gas phase transition; while for extraction and chromatography, the interaction is exchange between non-miscible phases.
  • the subject invention relates to a composition of matter having the structure:
  • the invention provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of isomers of such compounds which comprises contacting the mixture of isomers with the composition under conditions such that the enantiomeric isomer binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the enantiomeric isomer from the composition, and recovering the purified enantiomeric isomer.
  • the invention also provides a process of obtaining a purified organic compound of interest able to form hydrogen bonds from a mixture of organic compounds which comprises contacting the mixture with the composition under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate compound from the composition, and recovering the purified compound.
  • the invention further provides a process of preparing the composition which comprises: (a) reacting a chiral multifunctional reagent containing at least one protecting group with a compound having the structure:
  • step (c) treating the compound formed in step (b) with a condensing agent under conditions permitting multiple macrolactamization so as to thereby form the composition.
  • the subject invention further provides a composition of matter having the structure:
  • A has the structure:
  • the subject invention also provides a composition of matter having the structure:
  • A has the structure :
  • R 1 is H, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl) alkyl, or acylalkyl group, or an aryl group, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group.
  • the subject invention also provides a composition of matter having the structure:
  • composition of matter having the structure:
  • the invention provides a composition of matter having the structure:
  • A has the structure:
  • composition of matter having the structure:
  • the subject invention also provides a composition of matter having the structure:
  • R 1 and R 2 are each independently H, F, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl) alkyl, acylalkyl, aryl, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group; wherein Q is selected from a group consisting of H, linear or branched chain alkyl, acyl or aryl; and n is an integer from 0 to about 3.
  • Figure 1 shows a scheme for the practical synthesis host molecule 2: (a) Methanol/ammonia 4:1, rt, 2 days, 97%; (b) Boc 2 O, i-Pr 2 NEt, 4-DMAP (10 mol %), CH 2 Cl 2 , 1 h, 90%; (c) NaN(TMS) 2 , THF, -78°C, 3 min; add tetra-n-butylammonium iodide and methyl 3,5-bis(bromomethyl)benzoate; warm to 10°C, 2 h, 82%; (d) benzene-1, 3 , 5-trithiol, i-Pr 2 NEt, THF, 8 h, 78%; (e) TFA, anisole, CH 2 Cl 2 , rt, 16 h, quant; (f) Boc 2 O, i-Pr 2 NEt, K 2 CO 3 , CH 2 Cl 2 , rt, 24 h, 86%; (g) T
  • Figure 2 shows a diagram of a model for receptor-substrate binding.
  • Figure 3 shows composition 13a*.
  • Figure 4 shows a synthesis of 13a*.
  • Figure 5 shows compositions 11a* and 12*.
  • Figure 6(a) shows a synthesis of composition 11a*.
  • Figure 6(b) shows a synthesis of composition 12*.
  • Figure 7(a) shows compositions laa, lbb, lcc and 2aa.
  • Figure 7(b) shows a synthesis of composition 2aa.
  • Figure 8(a) shows compositions A 4 B 6 , precursors thereof (A, B), and precursors of A 4 B1 6 and A 4 B2 6 (B1 and B2, respectively).
  • Figure 8(b) shows a synthesis of labeled composition 2'.
  • Figure 9(a) shows compositions 1'' and 2''.
  • Figure 9(b) shows a comparison of models for interaction between compositions 1 and 1'' with a peptide substrate.
  • Figure 9(c) shows key intermediates 3'' and 4'' in the synthesis of composition 1".
  • Figure 10 shows gray-scale histograms of substrate sequence binding for 2".
  • composition of matter having the structure:
  • X, Y, and Z are each O; in another embodiment, they are each S.
  • R 1 , R 2 , and R 3 are each phenyl, or they are each 4-hydroxyphenyl.
  • n is desirably 1.
  • the invention further provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of isomers of such compounds which comprises contacting the mixture of isomers with the chiral host composition defined hereinabove under conditions such that the enantiomeric isomer binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the enantiomeric isomer from the composition, and recovering the purified enantiomeric isomer.
  • the process is used to purify enantiomers of amino acid derivatives, of which diamides are particularly effective.
  • the invention also provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the chiral host composition defined hereinabove under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the compound from the composition, and recovering the purified compound.
  • the process is used to purify derivatives of amino acids differing in side-chains. The process is particularly well suited to purify diamide derivatives of amino acids.
  • compositions of interest are bind it to a solid support such that a chromatographic adsorbent results which is specific for enantiomeric isomers of compounds of interest and other organic compounds of interest which differ only in side-chain substitution.
  • Effective use of the composition bound to a solid support is made to obtain the enantiomeric isomers of an amino acid derivative in a purified form and to obtain a purified organic compound of interest able to form hydrogen bonds from a mixture of compounds.
  • the compound to be purified by the composition is preferably a diamide.
  • the invention further provides a process of preparing the composition, which comprises:
  • step (b) treating the compound formed in step (a) under suitable conditions to cleave one protecting group to form a compound having the structure:
  • step (c) treating the compound formed in step (b) with a condensing agent under conditions permitting multiple macrolactamization, thereby forming the desired composition.
  • the preparation of the composition strategically exploits its C 3 symmetry.
  • the synthesis of the composition could proceed in a manner analogous to the detailed experimental examples given hereinbelow for embodiments in which X, Y, and Z are S, and R 1 , R 2 , R 3 are 4-hydroxy phenyl, except that if there is only one protecting group in the chiral multifunctional reagent of step (a), then none of the side-group protection reactions would pertain.
  • step (a) above can be carried out by several alternative methods of forming amide bonds.
  • One approach is to contact the achiral tetraaromatic triamino triester above shown with the p-nitrophenyl active ester of the chiral multifunctional reagent, made from p-nitrophenol, N-hydroxybenzotriazole, and N,N-dicyclohexylcarbodiimide.
  • the reaction may be performed in the presence of aprotic dipolar solvents, such as N,N-dimethylformamide, tetrahydrofuran, or dimethylsulfoxide, diluted with a miscible cosolvent, such as dichloromethane, to the extent required to achieve solubility of all reactants, at temperatures from about 0 to 100°C, preferably from 0 to 30°C.
  • aprotic dipolar solvents such as N,N-dimethylformamide, tetrahydrofuran, or dimethylsulfoxide
  • a miscible cosolvent such as dichloromethane
  • the chiral multifunctional reagent containing at least one protecting group in step (a) is an amino acid containing an N-protecting group.
  • the amino acid is L-phenylalanine or L-tyrosine.
  • the N-protecting group is preferably chosen such that it may be removed in process step (b) by an acid, for example, trifluoroacetic acid.
  • process step (b) involves the removal of three protecting groups on the tetraaromatic intermediate.
  • This reaction could be effected by any method corresponding to the lability of the protecting group.
  • a large variety of protecting groups are available for the purpose, including t-butyloxycarbonyl (BOC), benzyloxycarbonyl, 2-bromobenzyloxycarbonyl, and p-toluensulfonyl. While a preferred method is to use acid-sensitive BOC groups, other effective protecting groups also removable by acid include biphenylisopropyloxycarbonyl (Bpoc) and adamantyloxycarbonyl (Adoc). Still other protecting groups may be selected such that alternative methods of removal are feasible according to the invention, including photolytic, reductive, electrochemical, and mild base conditions. This flexibility allows a wide range of chiral multifunctional reagents to be used to prepare the composition.
  • the protecting ester group for example, methyl
  • the protecting ester group for example, methyl
  • the carboxylic acid by (i) transesterification with trimethylsilylethanol, followed by (ii) fluoride-induced silane elimination.
  • the condensing agent in step (c) could comprise a reagent generated (i) from an agent selected from a group comprising pentafluorophenol, hydroxybenzotriazole, 4-nitrophenol, 2-nitrophenol, pentachlorophenol, hydroxysuccinimide, and hydroxypiperidine and (ii) from an agent selected from a group consisting of N,N-dicyclohexyldiimide, diisopropylcarbodiimide, and carbonyldiimidazole.
  • condensing methods may also serve the purpose, including Woodward's reagent K, mixed anhydrides, triphenylphosphine/2,2'-dipyridyl sulfide, ketenimines, and acyloxyphosphonium salts.
  • the condensing agent is the combination of N,N-dicyclohexylcarbodiimide and pentafluorophenol. If the multifunctional chiral reagent of step (a) contains an alcohol function, the process of steps (b) and (c) could be simply adapted to generate three ester linkages after multiple macrolactonization. Other modifications in the multifunctional chiral reagent of step (a) could be readily envisioned to form such alternative linkages as thioesters, thionoesters, and phosphoramides.
  • R 1 , R 2 , and R 3 are 4-hydroxyphenyl which should be made by coupling with the suitably protected multifunctional chiral reagent Boc-L-tyrosine (Tyr).
  • the protecting group on the Tyr is preferably an allyl ether.
  • composition of matter hereinafter denoted 9 having the structure:
  • A has the structure:
  • R 1 and R 2 are independently the same or different and are H, F, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl)alkyl, or acylalkyl group, or an aryl group, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group;
  • X is CH 2 or NH;
  • n is 0 to about 3.
  • the subject invention provides a composition wherein X is NH.
  • the invention provides a composition wherein R 1 and R 2 are H.
  • the subject invention also provides a composition of matter (hereinafter referred to as 10) having the structure:
  • A has the structure:
  • R 1 is H, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl) alkyl, or acylalkyl group, or an aryl group, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group.
  • the invention provides a composition wherein R 1 is a phenyl group.
  • the invention provides a composition wherein R 1 is a benzyloxymethyl group.
  • composition of matter (hereinafter referred to as 2A) having the structure:
  • the subject invention also provides a compound which comprises the compositions of matter 9, 10, or 2A, bound to a solid support.
  • the subject invention further provides a complex which comprises the compositions 9, 10, or 2A, bound to a derivative of an amino acid.
  • the invention provides a composition wherein the derivative is an amide.
  • the subject invention provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of isomers of such compounds which comprises contacting the mixture of isomers with the compositions 9, 10, or 2A, under conditions such that the enantiomeric isomer binds to the compositon to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the enantiomeric isomer from the composition, and recovering the purified enantiomeric isomer.
  • the subject invention further provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the compositions 9, 10, or 2A, under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the organic compound from the composition, and recovering the purified compound.
  • the invention provides a process wherein the purified organic compound is an amino acid derivative.
  • the subject invention also provides a process of preparing the composition having the structure:
  • A has the structure:
  • R 1 and R 2 are H and n is 1 which comprises:
  • step (b) hydrolyzing the compound formed by step (a) under suitable conditions to form an acid compound having the structure:
  • step (c) treating the compound formed in step (b) under suitable conditions so as to activate the acid compound to form a compound having the structure:
  • step (d) reacting the compound formed in step (c) under suitable conditions with a compound having the structure:
  • step (e) saponifying the compound formed by step (d) under suitable conditions to form a diacid having the structure:
  • step (f) activating the diacid formed in step (e) under suitable conditions to form a compound having the structure:
  • step (g) deprotecting the compound formed in step (f) under suitable conditions to form a diamino diacid having the structure:
  • step (h) dimerizing the diamino diacid formed in step (g) under suitable conditions to form the composition having the structure:
  • A has the structure:
  • R 1 and R 2 are H and n is l.
  • esters other than methyl esters may be used in an equivalent manner for the purposes of the process.
  • Other useful esters include ethyl, propyl, phenol, and benzyl esters.
  • the condensing agent in step (a) could comprise a reagent generated (i) from an agent selected from a group comprising pentafluorophenol, hydroxybenzotriazole, 4-nitrophenol, 2-nitrophenol, pentachlorophenol, hydroxysuccinimide, and hydroxypiperidine and (ii) from an agent selected from a group consisting of N,N-dicyclohexyldiimide, diisopropylcarbodiimide, and carbonyldiimidazole.
  • Condensing methods may also serve the purpose, including Woodward's reagent K, mixed anhydrides, triphenylphosphine/2, 2'-dipyridyl sulfide, ketenimines, and acyloxyphosphonium salts.
  • Hydrolyzing step (b) may be performed using base or acid catalysis, though preferably base catalysis.
  • Treating step (c) is effected by a wide variety of procedures, including reaction of pentafluorophenol with DCC or 1-(3-(dimethylamino)propyl)-3-ethylcarbodiimide hydrochloride (EDC).
  • Reacting step (d) is performed in the presence of a nonnucleophilic base such as triethylamine.
  • a nonnucleophilic base such as triethylamine.
  • Good solvents for the purpose include dimethyl acetamide or dimethyl formamide.
  • Saponifying step (e) is carried out using a base, such as sodium hydroxide.
  • bases which effect the step include lithium hydroxide, potassium hydroxide, and tetramethylammonium hydroxide.
  • Activating step (f) is effectively performed using an activating agent such as
  • step (g) is carried out preferrably under mildly acidic conditions.
  • Useful acids include trifluoroacetic, trichloroacetic acid and hydrochloric acid in dioxane solution. Scavengers such as anisole help prevent untoward alkylation reactions.
  • Dimerizing step (h) may be effectively performed in the presence of a mild nonnucleophilic base, such as diisopropylethylamine or triethylamine in a dipolar nonaqueous solvent, such as tetrahydrofuran.
  • the subject invention also provides a process of preparing the composition having the structure:
  • A is a 1,3,5-trisubstituted phenyl moiety and R 1 and R 2 are H and n is 1 which comprises:
  • R 1 and R 2 are H and n is 1.
  • the reacting step may be effectively performed as a onepot procedure in the presence of a mild nonnucleophilic base such as diisopropylethylamine.
  • a mild nonnucleophilic base such as diisopropylethylamine.
  • Useful solvents include dipolar nonaqueous solvents such as dimethyl formamide and tetrahydrofuran.
  • the reaction may be carried out over a range of temperatures from -25°C to 60°C, but preferably at 0-10°C.
  • the subject invention also provides a process of preparing the composition having the structure:
  • step (b) reacting the compound formed by step (a) with an acylating agent under suitable conditions to form a plurally acylated compound having the structure:
  • step (c) reacting the plurally acylated compound formed by step (b) with a compound having the structure:
  • step (d) reacting the alkylated amide formed by step (c) with benzene-1,3,5-trithiol under suitable conditions to form a sulfide having the structure:
  • step (e) deprotecting the sulfide formed by step (d) under suitable conditions to form a free amine ester having thh structure:
  • step (f) re-acylating the free amine ester formed by step (e) under suitable conditions to form an acylamine ester having the structure:
  • step (g) saponifying the acylamine ester formed by step (f) under suitable conditions to form an acylamine acid having the structure:
  • step (h) activating the acylamine acid formed by step (g) under suitable conditions to form an acylamine activated ester having the structure:
  • step (i) de-protecting the acylamine activated ester formed by step (h) under suitable conditions to form a free amine activated ester having the structure:
  • Reacting step (a) may be carried out in the presence of a miscible co-solvent such as methanol, and occurs in high yield when performed at ambient temperatures.
  • Reacting step (b) may be carried out using a variety of acylating agents in the presence of nonnucleophilic base and 4-dimethylaminopyridine catalyst. Common agents include t-Boc-Cl and Amyloxycarbonyl chloride.
  • Reacting step (c) is efficiently performed using sodium hexamethyldisilylazide in tetrahydrofuran solution. Preferred temperatures range from -80°C to -70°C.
  • Reacting step (d) is readily effected in the presence of a nonnucleophilic base such as diisopropylethylamine in a dipolar nonaqueous solvent such as tetrahydrofuran.
  • Deprotecting step (e) occurs well by using a mild acid, such as trifluoacetic acid in the presence of a scavenger such as anisole.
  • Reacylating step (f) is carried out using a variety of acylating agents.
  • t-Boc 2 O is a preferred acylating agent for the purposes of the synthesis.
  • Saponifying step (g) may be carried out using such bases as lithium hydroxide and sodium hydroxide.
  • Lithium hydroxide is a preferred base.
  • Activating step (h) is carried out using pentafluorophenol in the presence of various condensing agents, including DCC and EDC.
  • De-protecting step (i) may be performed using a mild acid such as trifluoroacetic acid and a scavenger.
  • Cyclizing step (j) is performed using a dropwise addition technique and a nonnucleophilic base such as diisopropylethylamine in a dipolar nonaqueous solvent such as dimethyl acetamide or dimethyl formamide.
  • Receptors 1 and 2 are capable of high binding selectivity among simple amino acid derivatives (Table I).
  • the chiral host compounds may be utilized in any manner suitable for the intended purpose.
  • the host may be covalently bound to a polymer by modification of the synthetic method described above by replacing phloroglucinol or a similar starting material with one which has the additional substitution of an alkyl, aryl, or aralkyl, linker containing a reactive moiety at its terminus, comprising a halide, amine, carboxylate, alcohol, or thiol, if necessary in suitably protected form.
  • the resulting chiral polymer may serve as an adsorbent for use as a convenient extractive reagent, in which the polymer may be combined with a mixture of racemic amino acid derivatives or a mixture of compounds related by differing side-chain substitution in a range of polar or nonpolar solvents.
  • the polymeric complex is then separated by gravity or suction filtration, centrifugation, or sedimentation and decanting.
  • the desired enantiomeric derivative or related compound may be obtained by washing the polymer with a suitable buffer, solvent, or mixture of solvents at a temperature suitable for releasing the derivative from the polymeric host.
  • the chiral polymer may also serve as an adsorbent in a chromatographic column, in which the mixture of enantiomers or related compounds may bind with different affinities, and then be eluted after washing with a suitable buffer, solvent, or mixture of solvents.
  • the adsorbent is preferably prepared using finer meshes (>400 U.S.
  • Any polymeric resin selected from the group consisting of polyacrylamide, phenolformaldehyde polymers, polymethacrylate, carbohydrates, aluminates, and silicates may serve as the solid medium.
  • the chiral hosts of the subject invention bind diamides of certain amino acids with high selectivity which is dependent upon the nature of the amino acid side chain (2-kcal/mol for serine vs alanine) and the identity of the N-alkyl substituent (>3 kcal/mol for methyl vs tert- butyl).
  • These synthetic hosts are among the most enantioselective known, and bind certain derivatives of L-amino acids with selectivities as high as 3 kcal/mol. No other composition has been available to the art which achieves binding energy differentials of the magnitude herein disclosed for diastereoselective complexation of amino acid derivatives.
  • composition of matter having the structure:
  • the invention provides the composition wherein A, B and C are O.
  • the invention provides the composition wherein A, B and C are S.
  • the invention also provides the composition wherein R 1 , R 2 , and R 3 are each independently phenyl, 4-hydroxyphenyl, pyridyl, pyrrolyl, indolyl or naphthyl.
  • the invention further provides the composition wherein R 1 , R 2 , and R 3 are 4-hydroxymethylphenyl.
  • the invention also provides the composition wherein R 1 , R 2 , and R 3 are each independently 4-allyloxyphenyl, 4-alkoxyphenyl, 4- acyloxyphenyl or 4-(dye-substituted-acyloxy)phehnyl.
  • the invention provides the composition 43, wherein n is 1.
  • the invention provides a compound which comprises the composition having the structure:
  • the invention provides a compound which comprises the above composition bound to a derivative of an amino acid.
  • the invention provides the compound above wherein the derivative is an oligopeptide.
  • the subject invention provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of optical isomers of such compound which comprises contacting the mixture of isomers with the composition having the structure:
  • the invention also provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the composition shown above under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the organic compound from the composition, and recovering the purified compound.
  • the invention provides the above process wherein the purified organic compound is an amino acid derivative.
  • the invention provides the above process wherein the amino acid derivative is an oligopeptide.
  • the invention provides the above process wherein the purified organic compound is a biopolymer.
  • the invention provides the above process wherein the biopolymer is an enzyme.
  • the invention provides the above process wherein the purified organic compound is a monosaccharide or a polysaccharide.
  • the invention provides the above processed directed to optical isomers and organic compounds wherein the composition shown above is bound to a permeable membrane.
  • the invention provides a composition of matter having the structure:
  • A has the structure:
  • the invention provides the above composition wherein R 1 and R 2 are H. In another embodiment, the invention provides the composition wherein Q is an acyl group. In a certain embodiment, the invention provides the above composition wherein Q is an acyl moiety sustituted by a dye molecule. In a particular embodiment, the invention provides the above composition wherein the acyl moiety is:
  • the invention also provides a compound which comprises the above shown composition of matter bound to a solid support.
  • the invention also provides a compound which comprises the above shown composition bound to a derivative of an amino acid.
  • the invention provides the above compound wherein the derivative is an oligopeptide.
  • the subject invention provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of optical isomers of such compound which
  • A has the structure:
  • the invention also provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the above shown composition under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the organic compound from the composition, and recovering the purified compound.
  • the invention provides the above process wherein the purified organic compound is an amino acid derivative.
  • the invention provides the above process wherein the amino acid derivative is an oligopeptide.
  • the purified organic compound is a biopolymer.
  • the invention provides the above process wherein the biopolymer is an enzyme.
  • the invention provides the above process wherein the purified organic compound is a monosaccharide or a polysaccharide.
  • the invention provides the above processes directed to optical isomers and organic compounds wherein the composition shown above is bound to a permeable membrane.
  • the subject invention provides a composition of matter having the structure:
  • the invention provides the above composition wherein A, B and C are 0.
  • the invention provides the above composition wherein A, B and C are NH.
  • the invention provides the above composition wherein Q is an acyl group.
  • the invention provides the above composition wherein Q is an acyl moiety sustituted by a dye molecule.
  • the invention provides6 the above composition wherein the acyl moiety sustituted by a dye molecule is:
  • the invention provides the above composition wherein n is 1 or 2.
  • the invention provides a compound which comprises the above shown composition of matter bound to a solid support.
  • the invention provides a compound which comprises the above shown composition bound to a derivative of an amino acid.
  • the invention provides the compound above wherein the derivative is an oligopeptide.
  • the invention also provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of optical isomers of such compound which
  • the invention also provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the above shown composition under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the organic compound from the composition, and recovering the purified compound.
  • the invention provides the above process wherein the purified organic compound is an amino acid derivative.
  • the invention provides the above process wherein the amino acid derivative is an oligopeptide.
  • the invention provides the above process wherein the purified organic compound is a biopolymer.
  • the invention provides the above process wherein the biopolymer is an enzyme.
  • the invention provides the above process wherein the purified organic compound is a monosaccharide or a polysaccharide.
  • the invention provides the above processes directed to optical isomers and organic compounds wherein the composition shown above is bound to a permeable membrane.
  • the permeable membrane is one of the type used to filter and separate molecules according to selected physical parameters, such as size, molecular weight, etc.
  • a variety of synthetic and natural memranes are suitable for the purpose, including polyamide, nylon, perfluoroethylene, cellulose acetate, etc.
  • the subject invention also provides a composition of matter having the structure:
  • R 1 and R 2 are each independently H, F, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl) alkyl, acylalkyl, aryl, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group; wherein Q is selected from a group consisting of H, linear or branched chain alkyl, acyl or aryl; and n is an integer from 0 to about 3.
  • the invention provides the above composition wherein R 1 and R 2 are H.
  • the invention provides the above composition wherein Q is an acyl group. In a particular embodiment, the invention provides the above composition wherein Q is an acyl moiety sustituted by a dye molecule. In another particular embodiment, the invention provides the above composition wherein the acyl moiety sustituted by a dye molecule is:
  • the invention also provides a compound which comprises the composition of matter having the above structure bound to a solid support.
  • the invention also provides a compound which comprises the above shown composition bound to a derivative of an amino acid.
  • the invention provides the above compound wherein the derivative is an oligopeptide.
  • the invention also provides a process of obtaining a purified enantiomeric isomer of a compound of interest from a mixture of optical isomers of such compound which
  • R 1 and R 2 are each independently H, F, a linear or branched chain alkyl, arylalkyl, alkoxyalkyl, aminoalkyl, alkylaminoalkyl, hydroxyalkyl, (cycloalkyl)alkyl, acylalkyl, aryl, a linear or branched chain alkylaryl, pyridyl, thiophene, pyrrolyl, indolyl or naphthyl group; wherein Q is selected from a group consisting of H, linear or branched chain alkyl, acyl or aryl; and n is an integer from 0 to about 3; under conditions such that the enantiomeric isomer binds to the compositon to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the enantiomeric isomer from the composition, and recovering the purified enantiomeric isomer.
  • the invention also provides a process of obtaining a purified organic compound of interest from a mixture of organic compounds able to form hydrogen bonds, which comprises contacting the mixture with the above shown composition under conditions such that the organic compound binds to the composition to form a complex, separating the resulting complex from the mixture, treating the complex so as to separate the organic compound from the composition, and recovering the purified compound.
  • the invention provides the above process wherein the purified organic compound is an amino acid derivative.
  • the invention provides the above process wherein the amino acid derivative is an oligopeptide.
  • the invention provides the above process wherein the purified organic compound is a biopolymer.
  • the invention provides the above process wherein the biopolymer is an enzyme.
  • the invention provides the above process wherein the purified organic compound is a monosaccharide or a polysaccharide.
  • the invention provides the above processes directed to optical isomers and organic compounds wherein the composition shown above is bound to a permeable membrane.
  • N- ⁇ -BOC-L-tyrosine methyl ester (12.5 g, 42.3 mmol) in N,N-dimethylformamide (100 ml) was treated with allyl bromide (4.5 ml, 51.8 mmol), tetra-n-butylammonium iodide (1.5 g, 4.3 mmol) and potassium carbonate (12 g, 86.4 mmol) and allowed to stir overnight.
  • reaction mixture was diluted with an equal volume of ethyl acetate, extracted with two 200 ml portions of 5% aqueous hydrochloric acid, two 200 ml portions of saturated aqueous sodium bicarbonate, and 100 ml saturated aqueous sodium chloride.
  • the organic phase was dried over magnesium sulfate, concentrated under reduced pressure and chromatographed using a gradient of 100% chloroform - 5% methanol/chloroform to yield the product (892 mg, 65% yield) as a pale yellow powder.
  • a solid phase peptide reaction vessel was charged with Merrifield resin (chloromethylated polystyrene cross-linked with 2% divinylbenzene ; 100 mg, 0. 100 meq), the macrocyclic tris-phenol made according to Example 16 (110.0 mg, 0.100 mmol), potassium carbonate (14 mg, 0.100 mmol), and N,N-dimethylformamide (2 ml).
  • the mixture was placed on a rotary agitator for four days.
  • the reaction mixture was washed successively with 5 ⁇ 5 ml portions of methylene chloride, methanol, deionized water, methanol, and methylene chloride.
  • the resulting solid was dried under high vacuum and weighed to dermine the amount of alkylation.
  • the coupled resin weighed 116.3 mg (approximately 15% based on chloromethyl groups).
  • the organic washes were diluted with 100 ml ethyl acetate and extracted with 50 ml portions of 1 M aqueous potassium hydrogen sulfate, saturated aqueous sodium bicarbonate, and saturated aqueous sodium chloride.
  • the organic phase was dried over magnesium sulfate, concentrated under reduced pressure, and chromatographed using 10% methanol/chloroform to recover unreacted tris-phenol (40.1 mg).
  • the infrared spectrum shows type I, II, and III amide bands (1650, 1510, and 1230 cm -1 ).
  • N- ⁇ -BOC-L-alanine p-nitrophenyl ester On neutralization with triethylamine, the resulting amine was reacted with N- ⁇ -BOC-L-alanine p-nitrophenyl ester to give N- ⁇ -BOC-L-alanylvaline methylamide (19.0 mg, 97.2%) after chromatography.
  • NMR integration and comparison with authentic DL diastereomeric compounds revealed an 85:15 mixture of diastereomers, i.e., 70% enantiomeric enrichment.
  • the resin could be regenerated by washing five times with 50 ml portions of methanol, dried under a stream of argon, and re-swelled with chloroform.
  • Ammonia (20 mL) was condensed into a solution of N-Boc-O-allyl-L-tyrosine methyl ester in CH 3 OH (60 mL) at -78 °C in a high pressure glass reaction vessel. The vessel was sealed and slowly warmed to rt. After 2 days, the vessel was cooled to -78 °C and opened. Argon was bubbled through the solution while it was allowed to warm slowly to rt. After 1 h, the solution was transferred to a round-bottom flask and all volatiles were removed.
  • i-Pr 2 -NEt 6.52 mL, 37.5 mmol
  • DMAP 192 mg, 1.56 mmol
  • di-tert-butyl dicarbonate 5.12 g, 23.5 mmol
  • Nona-Boc Trisulfide 6 Compound 5 (2.0 g, 2.63 mmol) was added to a suspension of benzene-1, 3 , 5-trithiol 6 (140 mg, 0.80 mmol) and i-Pr 2 NEt (610 ⁇ L, 35.1 mmol) in THF (20 mL) at rt. The reaction mixture was quenched with aq NH 4 C1 after 6 h and extracted with ether (2X).
  • Pentafluorophenyl Ester of 7. A solution of 1 M aq LiOH (15 mL, 15 mmol) was added to 7 (500 mg, 0.309 mmol) in THF/EtOH/H 2 O (6:3:2, 100 mL). The reaction mixture was poured into 1. M aq KHSO 4 after 8 h and extracted with ethyl acetate (3X). After the extracts were washed with brine and dried, solvent removal afforded the crude acid as a light brown powder which was washed with ether.
  • Pentafluorophenol 600 mg, 3.26 mmol
  • 1-(3-(dimethylamino) propyl)-3-ethylcarbodiimide hydrochloride 320 mg, 1.69 mmol
  • THF 7.0 mL
  • Tyrosine Macrocycle 2A Anisole (12 mL) and trifluoroacetic acid (60 mL) were added via syringe to a stirring solution of the above tris (pentafluorophenyl ester) (3.16 g, 1.52 mmol) in CH 2 -Cl 2 (125 mL). After 6 h, the reaction mixture was concentrated. The resulting pink oil was triturated with ether to yield the tris-TFA amine salt as a white powder (3.20 g).
  • O-Allyltyrosine methyl ester (20 mg, 0.084 mmol) was added to a stirred solution of (S)-(-)-methoxy(trifluoromethyl) phenyl-acetic acid (28.0 mg, 0.120 mmol) and DCC (40 mg, 0.20 mmol) in CH 2 Cl 2 (0.50 mL). After 3 h the reaction mixture was diluted with CH 2 Cl 2 (10.0 mL), filtered, and washed with 0.5 M aq NaOH.
  • EXAMPLE 21 One-step synthesis of 9. To an ice cold solution of (-)-(1R, 2R)-diaminocyclohexane (24 mg, 0.211 mmol) and iPrNEt 2 (0.11 mL, 0.417 mmol) in THF (100 mL) and dimethylacetamide (10 mL) was added 1,3,5-benzenetricarbonyl trichloride (36 mg, 0.139 mmol) as a single portion with stirring. After 2 hours at 0 °C, the mixture was allowed to warm to room temperature and stirred for an additional 12 hours.
  • 3aa To a solution of trimesic acid pentafluorophenyl dimethyl ester (0.42 g, 1.04 mmol) and tri (aminomethyl) benzene triHCl salt (86 mg, 0.31 mmol) in 10 mL of dry N,N-dimethylacetamide (DMA) was added 0.36 mL of iPr 2 NEt. After stirring 8 hr, the mixture was concentrated at reduced pressure and purified by flash chromatography (silica gel, 5% MeOH in CH 2 Cl 2 ) to give 3aa as an amorphous white solid (0.20 g, 78%).
  • DMA dry N,N-dimethylacetamide
  • EXAMPLE 25 A 2 B 2 dimethyl ester (3').
  • DMA dimethylacetamide
  • N-Succinyl dye-3R,4R-pyrrolidine diamine diTFA salt (5').
  • Dyed receptor (2') A solution of 0.12 g of Ms (pentafluorophenyl) ester 4' (0.131 mmol) and 0.11 g of 5' TFA 2 (0.146 mmol, 1.1 eq.) in 10 mL of DMA was added via syringe pump over 20 h to a solution of 0.23 mL of iPr 2 NEt (1.31 mmol) in 200 ml of THF at 28°C. After stirring for additional 8 hr, all volatiles were removed at reduced pressure.
  • Solid Phase Binding Assay The solid phase substrate library was prepared by the encoded split synthesis as described previously and included 50,625 different acylated tripeptide sequences corresponding to all possible combinations of the 15 acylating agents and 15 amino acids (used three times) listed in the text.
  • a 10 mg sample of the library ( ⁇ 10 5 beads) was mixed in a 1.5 mL Eppendorf tube with 0.3 mL of ⁇ 50 ⁇ M 2 ' in CHCl 3 . After agitation on a wrist-action shaker for 48 h, ⁇ 1% of the beads were found to be stained deep red. Fifty-five of these deep red beads were picked by hand under a 4X wide-field microscope and photolyzed (350 nm, 4 hrs) in 1-2 ⁇ L of DMF to release the tag molecules. After silylation (CH 3 C(OTMS)NTMS, ⁇ 0.1 ⁇ L), electron capture GC was used to analyze the tag complement of each picked bead.
  • the C 3 -symmetric receptor 1A (Hong, J.-LI; Namgoong, S.K.; Bernardi, A.; Still, W.C. J. Am. Chem. Soc. 1991, 113, 5111) described hereinabove is one of the most enantioselective synthetic receptors yet reported and binds N-Boc-N'-methylamide derivatives of simple amino acids with enantioselectivity ranging from 2 to 3 kcal/mol (90-99% ee) (Other enantioselective hosts for neutral molecules: Canceill, J.; Lacombe, L.; Collet, A.; J. Am. Chem. Soc. 1985, 107. 6993.
  • the O-allyl derivative 2A is appropriate.
  • Such otherwise stable ethers can be deprotected (Kunz, H.; Unverzagt, C. Angew. Chem., Int. Ed. Engl. 1984, 23, 436) with transition metals to free phenols or attached (Tambute, A.; Begos, A.; Lienne, M.; Macaudiere, P.; Caude, M.; Rosset, R.; New J. Chem. 1989, 13, 625) directly to a support using free radical chemistry.
  • the present synthesis avoids the problematic di-tert-butyl iminodicarboxylate anion coupling and addition of nitrogen and amino acid in separate steps. Instead, a more convergent route is provided in which an N-anionic amino acid fragment would be added to bis (bromomethyl) benzoate in a single step.
  • Use of a Boc-stabilized amide ion made from N-Boc-O-allyltyrosine amide is summarized in Figure 1, and proved more reactive to acylation than was the primary amide.
  • the major product with 1 equiv of Boc 2 O/DMAP the tri-Boc material could be isolated in 95% yield.
  • 5 might be acidic enough to have racemized under the basic conditions of the alkylation.
  • a sample of 5 was treated with K 2 CO 3 in methanol and then HCl in methanol.
  • the first treatment converted (Flynn, D.A.; Zelle, R.E.; Grieco, P.A. J. Org. Chem. 1983, 48, 2824) the C-terminal Boc-amide to methyl ester while the second removed the two N-terminal Boc groups, yielding O-allyltyrosine methyl ester.
  • the benzylic bromide 5 was then used to triply alkylate sym-trimercaptobenzene (Bellavita, V. Chim. Ital. 1932, 62, 655) using Hunig's base (i-Pr 2 NEt) providing C 3 -symmetric 6 in 78% yield.
  • the remainder of the synthesis involved a triple macrolactamization via an activated benzoic acid ester.
  • the Boc-substituted amide was quite labile toward acid and base, and conversion of the methyl ester to acid was difficult in its presence.
  • the problematic Boc could not be removed from the C-terminal amide without simultaneously deprotecting the tyrosyl amine.
  • An effective solution to the problem was to remove all Boc protecting groups with TFA and then restore Boc protection of the free amines with Boc 2 O to obtain 7 in 86% yield over both steps.
  • This triple macrolactamization was suitable for reactions of this type and provided 2A in 78% yield after silica gel chromatography.
  • 9 was made by first preparing an amide-linked Boc-B-A-B-A-B-Boc oligomer having the two internal carboxylates activated as pentafluorophenyl esters. When this material was deprotected (TFA, anisole) and slowly added to iPr 2 NEt/THF, it dimerized to 9 in 39% yield.
  • 9 could be prepared in a single step (13% yield) by simply mixing commercially available A acid trichloride and B at 3 mM concentration with iPr 2 NEt in dry THF.
  • a related pair of intramolecular hydrogen bonds closes the unbound end of 9 to produce a deep cavity which fully encapsulates the side chain (R) of a bound L-peptide.
  • L-valine With L-valine, this structure places the sidechain isopropyl near the face of the four aromatic rings (A) of 9. It is incompatible with the 1 H NMR of the corresponding L-valine methylamide complex, which shows a 2.5 ppm upfield shift for the side-chain methyls and an ⁇ 1ppm downfield shift of only one of the three different types of host NH's.
  • each binding energy is the average of two to five independent measurements on different protons, and the average of two to five independent measurements on different protons, and the largest deviation from the average is ⁇ 0.2 kcal/mol).
  • b Enantioselectivity favoring L.
  • c PGly phenylglycine.
  • NC no complex observed.
  • e Oc octanoyl.
  • f HSer homoserine.
  • peptide derivatives are bound with high selectivity for the L-configuration except when side chains are large (entries 7 and 8).
  • Valine and phenylglycine side chains appear to fit the binding cavity quite well, but substantial reductions in binding occur when even single methylenes are added (entries 3 vs 4 and 5 and 6 vs 7 and 8). Removal of side-chain bulk from a near-optimal side chain (iPR) also diminishes binding. Thus stepwise truncation of side-chain iPR to Me to H costs 1.5 kcal/mol per step with L-amino acids. The effect is less significant with D-amino acids, which the model suggests to have side chains projecting away from the binding site and into solvent. Finally, the large binding energies in entries 14 and 15 suggest that 9 can interact associatively with as many as three residues, a feat that appears unique among synthetic receptors. Presumably, the terminal residues of such peptides are able to form additional hydrogen bonds to the outlying amides of the host (NHCO and CONH in the schematic).
  • a 4 B 6 macrotricycle described herein is remarkable for several reasons.
  • Second, A 4 B 6 is a highly selective receptor for neutral peptides. For example, it binds derivatives of L amino acids with enantioselectivities as high as 99% ee and can also distinguish between peptides based on the steric requirements of their sidechains.
  • this sidechain selectivity can be quite large and exceed 3 kcal/mol even when the peptides being compared differ only by a single methylene (e.g. phenylglycine vs phenylalanine).
  • the conformationally rigid building blocks used minimize its flexibility.
  • the synthesis and properties of two related A 4 B 6 cyclooligomers which are constructed from more conformationally flexible acyclic diamines (1R,2R)-1,2-diphenylethylenediamine (hereinafter B1) and (2R,3R)-2,3-diaminobutane-1,4-diol (hereinafter B2).
  • the binding properties in this series of receptors are sensitive to the structure of the components used to assemble them, but rigid cyclic building blocks need not be used to obtain high binding selectivity.
  • the receptors To prepare the receptors, a simple one-step coupling was performed on the amines and the triacid chloride as described for A 4 B 6 . With B1, the A 4 B1 6 receptor was obtained in 10% yield when the coupling was carried out at a concentration corresponding to 6 nM in receptor.
  • Binding energies were measured by titrating 0.5 mM solutions of receptor in CDCl 3 with various N-acetyl amino acid methylamides and monitoring the receptor protons by 400 MHz NMR. In general, signals which showed the largest shifts upon binding were certain aromatic (H-C) and amide (H-N) protons. The binding energies found are given in Table III and all represent averages of at least two different binding measurements. Scatchard treatment of binding data indicated 1:1 complexes in all cases.
  • Both A 4 B 6 and A 4 B1 6 show surprisingly high selectivity among L amino acids which are distinguished only by the size and shape of their unfunctionalized, hydrocarbon sidechains.
  • Amino acids having branched sidechains bind well only when the branch occurs at the substrate ⁇ -carbon.
  • the receptors also distinguish substrates by sidechain length.
  • receptor A 4 B2 6 Like A 4 B 6 and A 4 B1 6 which bind L-peptides based on the steric reguirements of their sidechains, receptor A 4 B2 6 also distinguishes peptide sidechains sterically but with different selectivity. In particular, A 4 B2 6 selects for L-peptides whose sidechains are small and compact; thus alanine, valine and ethylglycine are well-bound while isoleucine, leucine, phenylglycine, propylglycine and butylglycine are more weakly bound relative to the other receptors. Thus A 4 B2 6 appears to have a smaller binding cavity, a property which may follow from cavity occupancy by benzyloxymethyl substituents or from partial cavity collapse due to the flexible nature of the B2 fragment.
  • a 4 B 6 receptor may be general to cyclooligomeric molecules of this class and that binding selectivity can be altered by starting with different amine and acid chloride fragments. It may be noted that these receptors incorporate diamine fragments in two different structural environments: the upper and lower macrocycles include four equivalent B amines while two other B's link those macrocycles together. By varying these distinct B fragments independently, even more receptor diversity can be generated.
  • a new tetrahedrally symmetric cagelike receptor 13*(A A B 6 ), an isomer of 7* (A 4 B 6 ) where A and B's are combined in a different way, is described.
  • the global minimum conformation of 13 by molecular mechanics calculation reveals several interesting structural features. First, it is conformationally homogeneous. Within 3 kcal/mol of the global minimum conformation, 13 exits in a single family of closely related conformations. Second, it has a well-defined cavity with hydrogen bond donor/acceptors on symmetrically positioned about the periphery. Each hydrogen bond donor/acceptors may interact with a peptide substrate bound in the central cavity.
  • MOM s-n-P 52 MOM s-S-S
  • the binding energy between a C 2 symmetric receptor and imidazole can be increased up to 3.3 kcal/mol by changing the solvent from CHCl 3 (3.7 kcal/mol) to CHCl 2 CHCl 2 (7.0 kcal/mol).
  • receptors with preorganized three-dimensional binding cavities should exhibit increases in binding energies in media composed of increasingly bulky solvent molecules.
  • Molecular mechanics calculations and CPK modeling studies show that receptor 13a* has a big cavity enough to accommodate a CHCl 3 solvent molecule, but too small to accommodate a CHCl 2 CHCl 2 .
  • changing the solid phase assay solvent from CHCl 3 to CHCl 2 CHCl 2 would increase binding energies and lead to different selectivities.
  • a solid phase color assay of 13a* was employed with th. side-chain protected substrate library in CHCl 2 CHCl 2 . The results are summarized in Table 15.
  • structure 11* (Fig. 5) is appealing because of its welldefined binding cavity and appropriately positioned hydrogen bonding groups.
  • the central benzene ring presumably reduces conformational flexibility and provides a hydrophobic region for nonbonded interactions with peptides, and is likely crucial for the high observed binding energies and selectivities.
  • the protected substrate library was screened for binding by treatment with 50 ⁇ M solution of the red receptor 11a* in CHCl 3 . After 24 hr of equilibration with the library, ca. 10% of the beads had become colored with ca. 1% being very deep red. The most deeply stained beads were
  • the binding data in Table 7 show that extraordinary selectivity was observed for the terminal acylating groups.
  • the terminal R was composed of three non-hydrogen atoms (31 for MeOCH 2 and 13 Me 2 N) . High selectivity was observed for the AA 3 position.
  • the residue in AA 3 was composed of D-amino acids with an amide group in the side chain (31 for Gln, 7 for Asn and 5 for Lys).
  • the data indicate that receptor 11a* discriminates between substrates most effectively when structural differences occur near the free end of the substrate chain.
  • the number of accepted residues is minimal in the case of the terminal acylating group (R) and increases with distance from the terminus.
  • receptor 12* does not show any binding
  • highly selective complexation of 11a* was also found by a related binding assay using the side-chain deprotected
  • 11a* is a readily accessible heterooligomeric assembly from trimesic acid (A) and diamine (B) linked through a tris(aminomethyl)benzene. The results described here not
  • 11a* is a highly selective receptor for peptide substrates but also demonstrate the power of directed screening of large chemical libraries as a method to find novel molecules having sought-after properties.
  • the remarkable differences in peptide substrate binding properties between lla* and 12* suggest that conformational homogeneity may be a key to the designing the highly selective receptors .
  • receptor 2aa and its relatives are attractive candidates.
  • laa and 2aa are closely related in that their cup-shaped binding cavities have both similar dimensions and analogous patterns of unassociated hydrogen bond donors and acceptors on their peripheries. Described herein is a simple synthesis of 2aa and its binding properties as revealed using an encoded combinatorial library of ⁇ 50,000 acylated tripeptide substrates. 2aa is a selective receptor for peptides.
  • AAn represents any one of the following fifteen side-chain-protected [(N-trityl)Asn, (N-trityl)Gln, (N-Boc)Lys, (O-tBu)Ser] amino acids:
  • R represents any one of the following fifteen groups: methyl (Me), ethyl (Et), i-propyl (iPr), t-butyl (tBu), i-butyl (iBu), neopentyl (neoPe), trifluoro-methyl (TFM), methoxymethyl (MOM), cyclopropyl (cPr), cyclobutyl (cBu), cyclopentyl (cPe), acetoxymethyl (AcOM), phenyl (Ph), dimethylamino (Me 2 N), morpholino (Mor)
  • the library was prepared using split synthesis . (A. Furka, M. Sebestyen, M. Asgedom and G. Dibo, Abstr. 14th
  • the binding assay was carried out by equilibrating a 10 mg sample ( ⁇ 10 5 beads) of the above peptide library with ⁇ 0.3 mL of 50 ⁇ M 2aa in CHCl 3 . After 24 hrs of agitation, ⁇ 1% of the beads had developed deep red-orange coloration. These beads carried peptides that bound red 2aa most tightly. (Previous studies have shown the color assay readily distinguishes substrates differing in binding energy by as little as 1.0 kcal/mol. Control
  • receptor 2aa bound a different set of peptides with the deprotected library. It was most discriminating at the internal AA1 site where Pro was strongly preferred.
  • 2aa preferentially bound the two partial sequences XCO-(D)Gln- (D)X-(L) Pro (40% of beads) and XCO-X-(L)Gln-(D)Pro (40% of beads) [X indicates no significant with both the configuration and position of the downstream Gln is particularly interesting. This novel Gln...
  • Pro selectivity may reflect hydrogen boning between the Gin sidechain and amides in the bottom of 2aa's binding cavity - a possibility not available to laa.
  • the ability of 2aa to selectively bind peptide spans having as many as three residues is remarkable for such a small host molecule.
  • the conformationally rigid building blocks A and B were chosen to minimize its flexibility.
  • the present inventors also describe herein the synthesis and properties of two related A 4 B 6 cyclooligomers which are constructed from more conformationally flexible acyclic diamines B1 and B2. Binding properties in this series of receptors are sensitive to structure of the components used to assemble them, but rigid cyclic building blocks need not be used to obtain high binding selectivity.
  • Binding energies were measured by titrating 0.5 mM solutions of receptor in CDCl 3 with various N-acetyl amino acid methylamides and monitoring the receptor protons by 400 MHz (H-C) and amide (H-N) protons.
  • the binding energies found are given in Table IA and all represent averages of at least two different binding measurements. Scatchard treatment of binding data indicated 1:1 complexes in all cases.
  • receptor A 4 B2 6 Like A 4 B 6 and A 4 B1 6 which bind L-peptides based on the steric requirements of their sidechains, receptor A 4 B2 6 also distinguishes peptide sidechains sterically but with different selectivity. In particular, A 4 B2 6 selects for L-peptides whose sidechains are small and compact. Thus, alanine, valine and ethylglycine are well-bound while isoleucine, leucine, phenylglycine, propylglycine and butylglycine are more weakly bound relative to the other receptors. A 4 B2 6 appears to have a smaller binding cavity, a property which may follow from cavity occupancy by benzyloxymethyl substituents or from partial cavity collapse due to the flexible nature of the B2 fragment.
  • This molecule is a cyclooligomer of trimesic acid and ( 1R, 2R) -diaminocyclohexane, and can be synthesized in one step from commercially available materials. Its most interesting property, however, is that it binds certain ⁇ -amino acid derivatives with high selectivity. In particular, 1' was found to bind L-amino acids enantioselectively (70-99% ee) and to select for amino acid side-chains having a particular size (e.g. phenyl>>benzyl, ethyl»methyl).
  • the approach to evaluating the binding properties of a synthetic receptor is closely related to methods developed for finding good ligands to biological receptors such as antibodies.
  • a receptor e. g. with a fluorescent dye or radioisotope
  • the general scheme involves labeling a receptor (e. g. with a fluorescent dye or radioisotope) so that it may be sensitively detected and then treating the labeled receptor with a large collection of potential substrates. If these substrates are spatially separated (e.g. on different solid particles or in different location on a plate), then those areas occupied by substrates which bind the receptor will themselves become labeled.
  • K a simplifies to 1/ [Receptor] free .
  • K a-min the minimum K a (here termed K a-min )of receptor for substrates on any fully labeled particles by measuring the equilibrium concentration of the labeled receptor in free solution over the particles and taking its reciprocal.
  • the particular solid phase assay used here generally involves labeling the receptor with a colored dye and mixing it in dilute solution with a library of peptide-like substrates attached to Merrifield synthesis beads. After 48 hours of agitation to equilibrate a dilute solution of the colored receptor and the initially colorless substrate bead library, a small percentage of the beads take on deep colorations. Then the concentration of free receptor remaining in solution is measured to determine K a-min , and then those beads having the deepest coloration are picked. By determining the structures of the substrates on those beads, one learns which substrates in the library bind the receptor with association constants of at least K a-min assuming the substrates on the deeply colored beads are at least 50% saturated by labeled receptor.
  • split synthesis prescribes a simple protocol for preparing the library of products resulting from all possible combinations of all alternative reagents used.
  • Split synthesis is carried out on small solid support particles (e.g. Merrifield beads) and yields a particle- supported library in which any particular particle carries the product from one particular set of reagents.
  • the split synthesis method was developed originally for oligopeptide synthesis and can yield very large libraries.
  • the final library will contain 3,200,000 (20 5 ) different pentapeptides.
  • any particular synthesis particle will bear only one type of pentapeptide (or at least have been submitted to only one particular, well-defined series of chemical steps).
  • Encoding entails attaching arrays of molecular tags to the solid support particles during each synthetic step to create unique, tag-encoded records of the particular reagents used in the synthesis of each library member.
  • a substrate library of 50,625 (15 4 ) terminally acylated tripeptides is used and is prepared by split synthesis on 50-80 ⁇ polystyrene (Merrifield) beads as described above.
  • the library is encoded using a set of sixteen highly electrophoric tagging molecules which can be detached from single synthesis beads and analyzed using electron capture capillary gas chromatography (ECGC).
  • ECGC electron capture capillary gas chromatography
  • a labeled variant of receptor 1' ( Figure 8(b)) was made that could be visualty detected by simple inspection. Since 1' itself has no appropriate label attachment site, a relative of 1 ' in which the two spanning transdiaminocyclohexane ⁇ (B in 1') were replaced by stereochemically similar trans-3 ,4-diaminopyrrolidine ⁇ was made. The pyrrolidine ring nitrogen could then serve as the label attachment point. For the label, an intense red dye, Disperse Red 1, was selected, and the final structure of the labeled receptor thus became 2'.
  • red-labeled receptor 2' is straight-forward and is outlined in Figure 8(c) (EXAMPLE 25). Because 2' is not significantly strained and has few rotatable bonds, it could be constructed by a few simple reactions which both couple fragments and macrocyclize in a single step. Thus, the macrocyclic tetramide 3' was prepared in >50% yield by a single reaction which linked and cyclized two molecules of trans-1,2-diaminocyclohexane and two molecules of diactivated trimesic ester.
  • the synthesis was completed via diactivation of 3' as a bis-pentafluorophenyl ester (4') followed by another single step coupling/macrocyclization ( ⁇ 30% yield) using labeled diamine 5' to give the desired dye-labeled receptor 2'.
  • the polymer-supported, encoded substrate library has been described and has the general structure:
  • AAn any one of the following fifteen sidechain-protected (side-chain protection: Asn (trityl), Lys (Boc), Ser (tBu)) amino acids (standard single letter codes for amino acids in parentheses):
  • R represents any one of the following fifteen groups: methyl (Me), ethyl (Et), i-propyl (iPr), t-butyl (tBu), i-butyl(iBu), neopentyl (neoPe), trifluoromethyl (TFM), methoxymethyl (MOM), cyclopropyl (cPr), cyclobutyl (cBu), cyclopentyl (cPe), acetoxymethyl (AcOM), phenyl (Ph)m dimethylamino (Me 2 N), morpholino (
  • the substrate library was made by split synthesis using fifteen different amino acids at each of the three AA sites and terminated the tripeptide chain with fifteen different acylating agents, the total number of different substrates in the library is 15 4 or 50,625.
  • sixteen GC-distinct electrophoric tagging molecules were used.
  • a 10 mg sample ( ⁇ 10 5 beads) of the substrate library was suspended in an Eppendorf tube containing -0.3 mL CHCl 3 to which was then added ⁇ 25 ⁇ L of a 600 ⁇ M CHCl 3 solution of red 2 ' .
  • the bead library immediately extracted most of the colored receptor from solution, but examination of the mixture through a low power microscope showed that all beads looked essentially the same. However, after 30 min of agitation on a wrist-action shaker, ⁇ 5% of the beads had turned light orange. After 48 hours of agitation, ⁇ 1% of the beads were stained deep red-orange along with more having various lighter orange colorations.
  • tags were attached by photolabile ortho-nitrobenzylic carbonate linkages, they could be released for ECGC analysis by long wavelength ultraviolet (350 nm) irradiation. After silylation (bistrimethylsilylacetamide) to increase tag volatility, the solution over each bead was injected into an electron capture capillary gas chromatograph for tag analysis.
  • V site following V
  • S L-(O-tBu) serine
  • G glycine
  • the binding energies measured by solid phase experiments are not directly comparable to those from solution phase experiments because the environment inside the bead is not quite the same as free solution.
  • the concentration of supported peptide in the bead is ⁇ 0.1M and this relatively high concentration favors peptide substrate aggregation which would diminish binding to receptor.
  • Comparison measurements of binding of several (L)Ala- and (L) Val- containing peptides in free solution and on polystyrene supports using receptor 1' showed that peptide binding is 2-3 kcal/mol stronger when binding is measured in dilute free solution.
  • receptor 2 binds peptide substrates having particular amino acid sequences with remarkable selectivity. This selectivity includes selection based on side-chain stereochemistry (favoring D-Asn/Gln, L-Val), size (Val»Ala) and functionality (favoring the carboxamides of Asn and Gln).
  • Receptor 2 ' binds certain side-chain-protected di- and tripeptides with remarkable selectivity and shows a large preference for n/q-V-containing substrates. Based on the number of replicate substrate sequences found in the binding assay, the total number of different substrate sequences which are bound by 2 ' at -4 kcal/mol or better are 500-1000 out of the entire library of 50,625 sequences.
  • Bio receptors use similar means but also distinguish substrates or fragments by size and seem to make such distinctions based in part upon a substrate's ability to precisely fill a binding cavity.
  • 1'' stereoselectively binds substrates having L-amino acids adjacent to L-Pro (90-99% de for L-Ala) and with binding constants (K a 2.5 ⁇ 10 5 for iPrCO-(L)Pro-(L)Ala) that are among the largest reported for binding a neutral guest by a synthetic host.
  • Receptor 1 has a cup-like shape with a ca. 6A diameter binding cavity surrounded by six unassociated hydrogen bond donors (D) and acceptors (A). According to molecular mechanics, the design of 1" has a very similar conformation except that the three naphthalenes substantially enlarged the binding cavity (ca. 8A diameter).
  • a 5,000-step Monte Carlo conformational search (Goodman, J.M.; Still, W.C. J. Comput . Chem. 1991, 12 , 1110) for 1 (L-Tyr modeled by L-Ala) using the MacroModel/AMBER * force field (McDonald, D.Q.; Still, W.C. Tetrahedron Lett.
  • R methyl (Me), ethyl (Et), isopropyl (iPr), t-butyl (tBu), neopentyl (neoPe), trifluoromethyl (CF 3 ), isobutyl (iBu), methoxymethyl (MOM) , acetoxymethyl (AcOM), cyclopropyl (cPr), cyclobutyl (cBu), cyclopentyl (cPe), phenyl (Ph), morpholino (Morph), dimethylamino (Me 2 N).
  • AA1-AA3 Gly
  • the library was supported on 50-80 ⁇ polystyrene beads and was prepared both with and without N-trityl/N-Boc sidechain protection. Each bead carried only one type of tripeptide substrate.
  • R Me, Et, . . .
  • AAn L-Ala, L-Ser, . . .
  • the selectivity for AA1-AA3 L-amino acids can be seen as a generally dark region in the upper half of the histograms just below AA1-AA3 .
  • the large number of gray areas below R indicate little receptor selectivity for the terminal acyl group.
  • the general similarity of the protected and unprotected histograms indicates that 2" binds both libraries with similar selectivity.
  • receptor 1 has an enlarged binding cavity but that is otherwise very similar to peptide receptor 1. 10
  • the structural similarities of these two receptors far outweigh the differences, their binding properties are very different. Whereas the previous receptor selects for small terminal residue substituents (as in O), 1" shows little selectivity for the terminal substituent but instead selects for internal L-Pro with high stereochemical and steric selectivity (as in N).

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  • Organic Chemistry (AREA)
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Abstract

Molécules réceptrices chirales utiles pour la purification d'énantiomères de dérivés d'acides aminés et d'autres composés. La présente invention concerne également des procédés de préparation desdites molécules réceptrices.
PCT/US1995/000948 1994-01-27 1995-01-27 Recepteurs enantioselectifs pour derives d'acide amine et autres composes WO1995020590A1 (fr)

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US08/188,146 US5599926A (en) 1992-06-19 1994-01-27 A4 B6 macrotricyclic enantioselective receptors for amino acid derivatives, and other compounds
US08/188,146 1994-01-27
US35766394A 1994-12-16 1994-12-16
US08/357,663 1994-12-16

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006043906A1 (fr) * 2004-10-20 2006-04-27 Agency For Science, Technology And Research Procede de dissolution de stereoisomeres d'un compose
JP2015081248A (ja) * 2013-10-24 2015-04-27 メルクパフォーマンスマテリアルズマニュファクチャリング合同会社 レジスト下層膜形成組成物

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4596819A (en) * 1984-01-23 1986-06-24 Warner-Lambert Company Modified tripeptides
US5326862A (en) * 1990-07-16 1994-07-05 Rhone-Poulenc Rorer S.A. Process for the preparation of sulphinylpristinamycin IIB
US5342935A (en) * 1990-06-25 1994-08-30 Merck & Co., Inc. Antagonists of immunosuppressive macrolides
US5344925A (en) * 1991-09-09 1994-09-06 Merck & Co., Inc. Imidazolidyl macrolides having immunosuppressive activity
US5384424A (en) * 1991-06-26 1995-01-24 Lucky Limited Process for the selective preparation of 4,4-methylene-bis-(N-phenylalkylcarbamate)

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4596819A (en) * 1984-01-23 1986-06-24 Warner-Lambert Company Modified tripeptides
US5342935A (en) * 1990-06-25 1994-08-30 Merck & Co., Inc. Antagonists of immunosuppressive macrolides
US5326862A (en) * 1990-07-16 1994-07-05 Rhone-Poulenc Rorer S.A. Process for the preparation of sulphinylpristinamycin IIB
US5384424A (en) * 1991-06-26 1995-01-24 Lucky Limited Process for the selective preparation of 4,4-methylene-bis-(N-phenylalkylcarbamate)
US5344925A (en) * 1991-09-09 1994-09-06 Merck & Co., Inc. Imidazolidyl macrolides having immunosuppressive activity

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2006043906A1 (fr) * 2004-10-20 2006-04-27 Agency For Science, Technology And Research Procede de dissolution de stereoisomeres d'un compose
JP2015081248A (ja) * 2013-10-24 2015-04-27 メルクパフォーマンスマテリアルズマニュファクチャリング合同会社 レジスト下層膜形成組成物

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