HK1071131B - Small molecule inhibitors of rotamase enzyme activity - Google Patents

Description

Small molecule inhibitors of rotamase enzyme activity

The application is a divisional application with application number 96194554.0, and the application date of application number 96194554.0 is 1996, 6 and 5.

Application to

This application is a partially-filed continuation-in-part application of U.S. patent application serial No. 08/479,436, filed on 7.6.1995.

Background

1. Scope of the invention

The present invention relates to neurotrophic compounds having affinity for FKBP-type immunophilins, their preparation and use as inhibitors of immunophilin-related enzyme activity, particularly inhibitors of peptidyl-proline isomerase or rotamase enzyme activity.

2. Description of the prior art

The term "immunophilins" refers to a wide variety of proteins that act as receptors for the major immunosuppressive drugs, the cyclosporin a (csa), FK506, and rapamycin. Several immunophilins are known as cyclophilins, and FK506 binding proteins, such as FKBP. The cyclosporin A binds to cyclophilin while FK506 and rapamycin bind to FKBP. These immunophilin drug complexes are compatible with a variety of intracellular signal transduction systems, particularly in the immune and nervous systems.

Immunophilins are known to have peptidyl-proline isomerase (PP1 enzyme) or rotamase activity. It has been determined that rotamase enzyme activity is catalytic during the interconversion of cis and trans isomers of immunophilins.

Immunophilins were originally discovered and studied in immune tissues. It was initially hypothesized by those skilled in the art that inhibition of immunophilin rotamase activity results in inhibition of T cell proliferation, thereby eliciting the immunosuppressive effects exhibited by immunosuppressive drugs such as cyclosporin A, FK506 and rapamycin. Further studies have shown that rotamase activity is, in essence, and naturally, inadequate for immunosuppressive activity. Schreiber et al (Schreiber) in Science 1990, vol.250, p556-559 show that immunophilin drug complexes interact with ternary protein targets as their mode of activity. Schleber et al, Cell (Cell)1991, vol.66, p807-815, believe that in the case of FKBP-FK506 and FKBP-CsA, the drug immunophilin complex binds calcineurin, an inhibitory T Cell receptor, which signals T Cell proliferation. Similarly, the complex of mycophenolate mofetil and FKBP interacts with RAFT1/FRAP protein, inhibiting signaling by IL-2 receptors.

It has now been found that immunophilins exist in high concentrations in the central nervous system. The immunophilin in the central nervous system is 10-50 times more abundant than in the immune system. In neural tissue, immunophilins appear to affect neuronal progression, nitric oxide synthesis, and neurotransmission release processes.

It has been found that picomolar concentrations of immunosuppressants such as FK506 and rapamycin stimulate neurite outgrowth in PC12 cells and sensory nerves, i.e., dorsal root ganglion cells (DRGs). Lyons et al, Natl.Acad.Sci, 1994, vol.91, p.3191-3195. In all animal experiments, FK506 was shown to stimulate nerve regeneration after surface nerve injury and to result in functional recovery in sciatic nerve injured animals.

It has been unexpectedly found that drugs with high affinity for FKBP are potent rotamase inhibitors that can elicit neurotrophic effects. Lyon et al believe that this finding suggests that immunosuppressive agents may be used to treat a variety of peripheral neuropathies and enhance neuronal regeneration in the Central Nervous System (CNS). Studies have shown that neurodegenerative diseases such as alzheimer's disease, parkinson's disease and Amyotrophic Lateral Sclerosis (ALS) can develop as a result of the loss or diminished access to specific neurotrophic substances by certain populations whose neurons are abnormally affected.

Neurotrophic factors have now been identified that affect specific neuronal populations in the central nervous system. For example, Alzheimer's disease has been hypothesized to be caused by a reduction or loss of Nerve Growth Factor (NGF). Thus, it has been suggested that Alzheimer's disease may be treated by increasing the activity of degenerated neuronal populations using extrinsic nerve growth factors or other neurotrophic proteins, such as glial derived neurotrophic factor (BDNF), ciliary neurotrophic factor, and neurotrophic factor-3.

The clinical utility of these proteins in various neurological disorders has been hampered by the difficulty in delivering large protein molecules to target tissues in the nervous system, and the bioavailability of such large proteins that are difficult to utilize by target tissues in the nervous system. In contrast, immunosuppressant drugs with neurotrophic activity are relatively small in size and exhibit excellent bioavailability and specificity. However, immunosuppressive agents exhibit a number of potentially serious side effects when administered chronically, including nephrotoxicity, such as glomerular filtration damage and irreversible interstitial fibrosis [ see kupa (Kopp) et al journal of the american society for renal disease (j.am.soc.nephrol.)1991, 1: 162 ], nerve damage such as occasional tremors, or non-specific cerebral colic such as fixed-position-free headache (see De glycone (De Groen) et al in new england journal of medicine (n.engl.j.med.)1987, 317: 861 and the resulting vascular hypertension induced by these [ see Kahan et al in New England journal of medicine (N.Engl. J.Med.) ] 1989, 321: 1725 ].

To prevent the side effects associated with the use of these immunosuppressive compounds, the present invention proposes a non-immunosuppressive compound containing a small molecule FKBP rotamase inhibitor for promoting nerve growth and regeneration in various neurological disease states. Neuronal repair can now be readily performed, including peripheral nerve damage caused by mechanical injury or disease (such as diabetes), mechanical damage to the central nervous system (spinal cord and brain), brain damage associated with shock, and also for the treatment of neurological disorders associated with neurodegeneration, including Parkinson's disease, Alzheimer's disease, and amyotrophic lateral sclerosis.

Summary of The Invention

The present invention relates to a novel class of neurotrophic compounds having affinity for FKBP-type immunophilins. Once bound to the protein, the neurotrophic compounds are potent inhibitors of immunophilin-related enzyme activity, particularly rotamase activity, thereby stimulating neuronal regeneration and outgrowth. The key feature of the compounds of the invention is that they do not produce any significant immunosuppressive activity other than their neurotrophic activity.

A preferred embodiment of the present invention is a neurotrophic compound of the formula:

in the formula:

R₁is selected from optionally substituted C₃-C₈Cycloalkyl-substituted C₁-C₉Straight or branched alkyl or alkenyl, C₃Or C₅Cycloalkyl radical, C₅-C₇Cycloalkenyl radical, Ar₁The alkyl, alkenyl, cycloalkyl or cycloalkenyl groups mentioned herein may optionally carry C₁-C₄Alkyl radical, C₁-C₄An alkenyl or hydroxy substituent; and Ar₁Selected from the group consisting of 1-naphthyl, 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-, 4-pyridyl and phenyl, each of which may have 1 to 3 substituents each independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

x is selected from oxygen, sulfur, methylene (CH)₂) Or H₂；

Y is selected from oxygen or NR₂Where R is₂Is hydrogen or C₁-C₆An alkyl group; and

z is selected from C₂-C₆A linear or branched alkyl or alkenyl group,

wherein the alkyl chain is substituted at one or more positions by Ar as defined above₁Substitution, C₃-C₈Cycloalkyl radical, of₁-C₆Cycloalkyl linked by a straight or unbranched alkyl or alkenyl chain and Ar₂Ar here₂Selected from the group consisting of 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-or 4-pyridyl and phenyl, each of which may have 1 to 3 substituents independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

z may also be a moiety of the formula:

in the formula R₃Is selected from optionally substituted C₃-C₈Cycloalkyl-substituted C₁-C₈Straight or branched alkyl, or Ar as defined above₁And unsubstituted Ar₁；

X₂Is O or NR₅Where R is₅Selected from hydrogen, C₁-C₆Straight or branched alkyl and alkenyl;

R₄selected from phenyl, benzyl, C₁-C₅Straight or branched alkyl or alkenyl and C substituted by phenyl₁-C₅Straight or branched alkyl or alkenyl.

Another preferred embodiment of the invention is a neurotrophic compound of the formula:

in the formula:

R₁is optionally covered with C₃-C₈Cycloalkyl-substituted C₁-C₉Straight or branched alkyl or alkenyl, C₃Or C₅Cycloalkyl radical, C₅-C₇Cycloalkenyl group, or Ar₁The alkyl, alkenyl, cycloalkyl or cycloalkenyl groups described herein may optionally carry C₁-C₄Alkyl radical, C₁-C₄Alkenyl or hydroxy substituents, and Ar₁Selected from 1-naphthyl, 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-, or 4-pyridyl or phenyl, each of which may have 1 to 3 substituents independently selected from hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino substituents;

z is C₂-C₆Wherein the alkyl chain is substituted at one or more positions by Ar as defined above₁Substituted, C₃-C₈Cycloalkyl radical, of₁-C₆Cycloalkyl or Ar linked by a straight or unbranched alkyl or alkenyl chain₂Ar here₂Selected from 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-, or 4-pyridyl, or phenyl, each of which has 1 to 3 substituents independently selected from hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino.

Another preferred embodiment of the present invention is to provide a neurotrophic compound having an affinity for FKBP-type immunophilins, which inhibits the rotamase activity of the immunophilin.

Another preferred embodiment of the present invention relates to a method for treating a neurological disease in an animal comprising administering a therapeutically effective amount of a compound having affinity for FKBP-type immunophilins, which inhibits rotamase activity of the immunophilin.

Another preferred embodiment of the present invention is a method of promoting neuronal regeneration and growth in a mammal which comprises administering to the mammal an effective amount of a neurotrophic compound having affinity for FKBP-type immunophilins, which inhibits rotamase activity of the immunophilin.

Yet another preferred embodiment of the present invention is a method for preventing neurodegeneration in an animal comprising administering to the animal an effective amount of a neurotrophic compound having an affinity for FKBP-type immunophilins, which inhibits rotamase activity of the immunophilin.

Another preferred embodiment are neurotrophic N-glyoxyl-alanyl esters of the formula:

in the formula:

R₁is optionally provided with C₃-C₆C of substituents of cycloalkyl₁-C₅Straight or branched alkyl or alkenyl or Ar₁Ar here₁Selected from 2-furyl, 2-thienyl or phenyl;

x is selected from oxygen and sulfur;

y is oxygen; and

z is a straight or branched alkyl or alkenyl group in which the alkyl chain is Ar as defined above at 1 or more positions₁Substitution, C₃-C₆Cycloalkyl radical, Ar₂Ar here₂Is selected from 2-,3-or 4-pyridyl, or phenyl, each of which has 1 to 3 substituents independently selected from hydrogen and C₁-C₄A substituent of an alkoxy group.

Particularly preferred neurotrophic N-glyoxyl-alanyl ester compounds according to the above formula are selected from:

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid-3- (2, 5-dimethoxyphenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid-2- (3, 4, 5-trimethoxyphenyl) -1-ethyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4-pyridyl) -1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -N- ([ 2-thienyl ] glyoxyl) pyrrolidine carboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxobutyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1-Cyclohexylglyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, and

(2S) -1- (2-thienyl) glyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester.

Brief Description of Drawings

Figure 1 is a photomicrograph of a dorsal root ganglion of a chicken treated with the compound of example 17 at the various concentrations indicated. FIG. 1 shows that the compound of example 17 of the present invention strongly promotes the outgrowth of axons in sensory neuron cultures. Graft cultures isolated from 9-10 day old chick embryo dorsal root ganglia were treated with the compound of example 17 at various concentrations as specified. After 48 hours, the number of axons greater than 1 dorsal root ganglion explanted were counted. The number of axons represented by the dorsal root ganglion of the untreated group was subtracted from the number of axons of the specimen treated with the compound of example 17 to obtain the number of specific neurite outgrowths depending on the compound of example 17. The figure shows microgram of the compound of example 17 treated dorsal root ganglion and the dose-related extraaxonal growth condition caused by the compound of example 17.

FIG. 2 shows the quantitative relationship of neurite outgrowth at dorsal root ganglion of chicks treated with the compound of example 17 at the various concentrations indicated. FIG. 2 shows that the compound of example 17 of the present invention effectively promotes the outgrowth of axons in sensory neuron cultures. Graft cultures isolated from 9-10 day old chick embryo dorsal root ganglion were treated with the indicated concentrations of the compound of example 17. After 48 hours, the number of axons greater than 1 dorsal root ganglion graft in length was counted. The number of axons expressed in the dorsal root ganglion of the untreated group was subtracted from the number of axons in the sample treated with the compound of example 17 to obtain the specific neurite outgrowth amount depending on the compound of example 17, which shows the quantitative dependence of the neurite outgrowth on the dose of the compound of example 17.

FIG. 3 is a photomicrograph of a section of a rat sciatic nerve. FIG. 3 shows that example 1 of the present invention promotes neuronal regeneration after sciatic nerve damage. Sciatic nerves from 150 grams of Spraque-Dawley males were crushed near the hip. Within 21 days thereafter, the compound of example 1(30mg/kg, subcutaneously), the inactive substance (30mg/kg, subcutaneously) or the lipid emulsion vehicle was administered once daily. The animals were sacrificed, the sciatic nerve was isolated, and a section of the nerve 2mm distal to the crush point was sectioned and stained with a Holmes (Holmes) silver stain (to estimate the number of axons) and ruxol (Luxol) fast blue stain (to estimate the number of remyelination). Micrographs show 630-value magnification micrographs of Sham (Sham) operated mice, vehicle-treated compromised animals, compound-treated example 1, and non-active treated sciatic nerve sections, 4 animals per group.

FIG. 4 is the binding of striatal membrane protein per μ g³H]-CFT map. FIG. 4 shows that the neuroimmunophilin ligands of the invention promote dopamine neuron recovery after treatment of mice with MPTP. 25g of CD1 mice were treated daily with 30mg/kg MPTP (i.p.) for 5 days. Animals were also dosed daily with either lipid emulsion vehicle, the compound of example 1 (100mg/kg subcutaneously) or the compound of example 17 (40, 20, 10mg/kg subcutaneously as specified) simultaneously with MPTP for an additional 5 days. After 18 days, the mice were sacrificed and the striatum harvested from 5 mice per group was pooled and processed into washed membrane preparations. Measurement of [3H]-the number of CFTs bound to these different groups of striatal membrane preparations to determine the content of dopamine-delivering factors at the viable nerve terminals. The amount of binding in the presence of 10. mu.M unlabeled CFT was provided based on an estimate of non-specific binding, i.e.the quantitative specificity [3H ] was subtracted from the total amount of binding]-the amount of CFT binding. The amount of binding for each experimental group was normalized to the protein content of the striatal membranes.Coronal and sagittal sections of brain from MPTP and drug-treated animals were stained with anti-Tyrosine Hydroxylase (TH) Ig to determine the TH level of striatum, mid-proximal forebrain tract axons and stroma, which are indicative of functional dopaminergic neurons.

FIG. 5 is [ 2 ]³H]Histogram of CFT vs 200. mu.g membrane protein. FIG. 5 shows that the neuroimmunophilin ligand of the invention promotes dopaminergic neuron recovery after treatment of mice with MPTP according to the procedure described in FIG. 4.

Figure 6 is a photomicrograph of coronal and sagittal sections of the brain at 630 x magnification. FIG. 6 shows that staining of brain sections of MPTP and drug-treated animals with anti-Tyrosine Hydroxylase (TH) Ig allows determination of the TH level of the striatum, an indication of functional dopaminergic neurons.

Figure 7 is a photomicrograph of coronal and sagittal sections of the brain at 50 x magnification. FIG. 7 shows that staining of brain sections of MPTP and drug-treated animals with anti-Tyrosine Hydroxylase (TH) Ig allows determination of the TH level of the stroma, which is indicative of functional dopaminergic neurons.

Fig. 8 is a photomicrograph of coronal and sagittal sections of the brain at 400 x magnification. FIG. 8 shows that staining MPTP with anti-Tyrosine Hydroxylase (TH) Ig and treatment of brain sections of animals with drugs can determine TH levels of the proximal forebrain bundle axons, which are indicative of functional dopaminergic neurons.

Detailed Description

The molecules of the novel neurotrophic compounds of the present invention are small compared to other known compounds that bind FKBP-type immunophilins, such as rapamycin, FK506, and cyclosporin.

The neurotrophic compounds of the present invention have affinity for FK506 binding proteins such as FKBP-12. When the neurotrophic compounds of the present invention are combined with FKBP, they have unexpectedly been found to inhibit prolyl-peptidyl cis-trans isomerase activity, i.e., rotamase activity of the binding protein, and to promote axonal growth without exhibiting immunosuppressive effects.

More particularly, the present invention relates to a novel class of neurotrophic compounds having the formula:

in the formula R₁Is an arbitrary quilt C₃-C₈Cycloalkyl-substituted C₁-C₉Straight or branched alkyl or alkenyl, C₃Or C₅Cycloalkyl radical, C₅-C₇Cycloalkenyl radical or Ar₁The alkyl, alkenyl, cycloalkyl or cycloalkenyl groups mentioned herein may optionally be substituted by C₁-C₄Alkyl radical, C₁-C₄Alkenyl or hydroxy substituted, and Ar₁Selected from 1-naphthyl, 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-or 4-pyridyl or phenyl, each of which has 1 to 3 substituents each independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

x is oxygen, sulfur, methylene (CH)₂) Or H₂；

Y is oxygen or NR₂Where R is₂Is hydrogen or C₁-C₆An alkyl group; and

z is C₂-C₆Straight or branched alkyl or alkenyl, where the alkyl chain is substituted at one or more positions with Ar as defined above₁Substitution, C₃-C₈Cycloalkyl radical, of₁-C₆Cycloalkyl or Ar linked by a straight or unbranched alkyl or alkenyl chain₂Ar here₂Selected from 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl2-thienyl, 3-thienyl, 2-, 3-or 4-pyridyl, or phenyl, each of which has 1 to 3 substituents independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

z can also be the following fragment:

in the formula

R₃Is selected from optionally substituted C₃-C₈Cycloalkyl or Ar as defined above₁Substituted C₁-C₈Straight or branched chain alkyl, and unsubstituted Ar₁；

X₂Is O or NR₅Where R is₅Selected from hydrogen, C₁-C₆Straight or branched alkyl and alkenyl;

R₄selected from phenyl, benzyl, C₁-C₅Straight or branched alkyl or alkenyl, and C substituted by phenyl₁-C₅Straight or branched alkyl or alkenyl.

Preferred compounds have the following structural formula:

in the formula:

R₁is an arbitrary quilt C₃-C₈Cycloalkyl-substituted C₁-C₉Straight or branched alkyl or alkenyl, C₃Or C₅Cycloalkyl radical, C₅-C₇Cycloalkenyl radical or Ar₁Alkyl, alkenyl, alkynyl, aryl, heteroaryl,cycloalkyl or cycloalkenyl radicals optionally substituted by C₁-C₄Alkyl radical, C₁-C₄Alkenyl or hydroxy substitution; and Ar₁Selected from 1-naphthyl, 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-or 4-pyridyl or phenyl, each bearing 1 to 3 substituents each independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

z is C₂-C₆Straight or branched alkyl or alkenyl in which the alkyl chain is substituted at one or more positions by Ar as defined above₁Substitution, C₃-C₈Cycloalkyl radical, of₁-C₆Cycloalkyl linked by a straight or unbranched alkyl or alkenyl group or Ar₂Ar herein₂Selected from 2-indolyl, 3-indolyl, 2-furyl, 3-furyl, 2-thiazolyl, 2-thienyl, 3-thienyl, 2-, 3-or 4-pyridyl or phenyl, each of which has 1 to 3 substituents independently selected from the group consisting of: hydrogen, halogen, hydroxy, nitro, trifluoromethyl, C₁-C₆Straight or branched alkyl or alkenyl, C₁-C₄Alkoxy or C₁-C₄Alkenyloxy, phenoxy, benzyloxy, and amino;

preferred compounds also include pharmaceutically acceptable salts or hydrates of the above compounds.

Preferred N-glyoxylprolyl esters have the following formula:

in the formula

R₁Is an arbitrary quilt C₃-C₆Cycloalkyl-substituted C₁-C₅Straight chain orBranched alkyl or alkenyl, or Ar₁Ar here₁Selected from 2-furyl, 2-thienyl or phenyl;

x is selected from oxygen and sulfur;

y is oxygen; and

z is a straight or branched alkyl or alkenyl group, where the alkyl chain is substituted at one or more positions with Ar as defined above₁Substitution, C₃-C₆Cycloalkyl radical, Ar₂Ar here₂Selected from 2-, 3-or 4-pyridyl or phenyl having 1 to 3 substituents independently selected from hydrogen and C₁-C₄A substituent of an alkoxy group.

The compounds of the present invention exist in two forms of stereoisomers, i.e., enantiomers and diastereomers. There are two stereochemical structures at position 1 of formula I, i.e., R-form or S-form, with S being preferred. Included within the scope of the present invention are enantiomers, racemates and diastereomeric mixtures. Enantiomers and diastereomers may be separated by methods well known to those skilled in the art.

It is now known that immunophilins like FKBP have been preferentially considered as peptides containing Xaa-Pro-Yaa, where Xaa and Yaa are both lipophilic amino acid residues, as described in Schreiber et al, J.Org.chem., 1990, 55, 4984-4986, and Harrison & Stein Biochemistry 1990, 29, 3813-3816. Thus, a modified prolyl peptidomimetic compound with a lipophilic substituent should bind with high affinity to the hydrophobic core of the FKBP active site, inhibiting its rotamase activity.

Preferred compounds of the invention contain such R₁The known shape and size of the groups, which are compared to the hydrophobic core of the FKBP active center, are not stereochemically large. Therefore, if R is₁Are large and/or carry many substituents, they have much less affinity for binding to the FKBP active site.

Preferred compounds of the invention include:

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3, 4, 5-trimethoxyphenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3, 4, 5-trimethoxyphenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-methylenedioxyphenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-methylenedioxyphenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-cyclohexyl-1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-cyclohexyl-1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R)1, 3-diphenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-2- [ 2-furyl ] -ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-2- [ 2-thienyl ] -ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-2- [ 2-thiazolyl ] -ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-2-phenyl) ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 2- (3, 4, 5-trimethoxyphenyl) -1-ethyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4-pyridyl) -1-propyl ester,

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -N- ([ 2-thienyl ] glyoxyl) pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxobutyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1-Cyclohexylglyoxyloyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1- (2-thienyl) glyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester.

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-dichlorophenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-dichlorophenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R) -1, 3-diphenyl-1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R) -1-cyclohexyl-3-phenyl-1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R) -1-cyclohexyl-3-phenyl-1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R) -1- (4, 5-dichlorophenyl) -3-phenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-2-cyclohexyl) ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (1, 2-dioxo-4-cyclohexyl) butyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 1, 7-diphenyl-4-heptanoate,

(2S) -1- (3, 3-dimethyl-1, 2-dioxo-4-hydroxybutyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-phenylalanine ethyl ester,

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-leucine ethyl ester,

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-phenylglycine ethyl ester,

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-phenylalanine phenyl ester,

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-phenylalanine benzyl ester, and

1-1- (3, 3-dimethyl-1, 2-dioxopentyl) -L-proline ] -L-isoleucine ethyl ester.

Particularly preferred N-glyoxyl prolyl ester compounds are selected from the following:

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-prop-2- (E) -enyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 2- (3, 4, 5-trimethoxyphenyl) -1-ethyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4-pyridyl) -1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester,

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -N- ([ 2-thienyl ] glyoxyl) pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester,

(2S) -1- (3, 3-dimethyl-1, 2-dioxobutyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester,

(2S) -1-Cyclohexylglyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, and

(2S) -1- (2-thienyl) glyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester.

The compound of the present invention may use inorganic or organic acids thereof. Salts obtained with inorganic or organic bases. Such acid salts include: acetate, adipate, alginate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, citrate, camphorate, camphorsulfonate, cyclopentanepropionate, digluconate, lauryl sulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, oxalate, pamoate, pectate, propionate, succinate, tartrate, thiocyanate, tosylate, and undecanoate. Base salts include ammonium salts, alkali metal salts such as sodium and potassium salts, alkaline earth metal salts such as calcium and magnesium salts, salts with organic bases such as dicyclohexylamine salts, N-methyl-D-glucamine salts, and salts with amino acids such as arginine, lysine, and the like. Nitrogenous bases can also form quaternary ammonium salts with agents such as lower alkyl halides, such as methyl, ethyl, propyl and butyl chlorides, bromides and iodides, dialkyl sulfates, such as dimethyl, diethyl, dibutyl and diamyl sulfates, long chain halides, such as decyl, lauryl, myristyl and stearyl chlorides, bromides and iodides; aralkyl halides such as benzyl and phenethyl bromide and the like. Water-soluble or oil-soluble or dispersible products are thereby obtained.

The neurotrophic compounds of the present invention may be administered periodically to patients in need of treatment for neurological disorders or other reasons in which it is desirable to stimulate neuronal regeneration and growth, such as various peripheral neuropathies and neurological disorders involving neurodegeneration. Can also be administered to mammals other than human to treat various mammalian neurological diseases.

The novel compounds of the present invention are potent inhibitors of rotamase activity and have an excellent degree of neurotrophic activity. This activity can be used to stimulate damaged neurons, promote neuronal regeneration, prevent neurodegeneration, and treat several neurological disorders known to be associated with neuronal degeneration and peripheral neuropathy. Neurological disorders that may be treated include, but are not limited to: trigeminal neuralgia, glossopharyngeal pain, facial paralysis, myasthenia gravis, muscular dystrophy, amyotrophic lateral sclerosis, progressive muscular atrophy, progressive hereditary global muscular atrophy, herniated, ruptured or prolapsed invertebral disc syndrome, cervical spondylosis, plexus abnormalities, thoracic outlet damage syndrome, peripheral nerve disorders such as diseases caused by lead, dapsone, ticks, prophyria or acute infectious polyneuritis, alzheimer's disease and parkinson's disease.

For this purpose, the compounds of the invention can be administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted depot formulation, in the form of generally nontoxic pharmaceutically acceptable carriers, adjuvants and vehicles. The term parenteral as used herein includes subcutaneous, intravenous, intramuscular, intraperitoneal, intrathecal, intraventricular, intrasternal and intracranial injection or infusion techniques.

In order to target the central nervous system to make the treatment effective, the immunophilin-drug complex should readily cross the blood-brain barrier when administered peripherally. Compounds of the invention that do not penetrate the blood-brain barrier can be effectively administered by intraventricular routes.

The pharmaceutical compositions may be in the form of sterile injectable preparations, such as sterile injectable aqueous or oleaginous suspensions. The suspension may be formulated according to the known art using suitable dispersing or wetting agents and suspending agents. The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic pharmaceutically acceptable diluent or solvent, for example, a solution of 1, 3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, ringer's solution and isotonic sodium chloride solution. In addition, sterilized fixed oils are conventionally employed as a solvent or suspending medium. For this purpose, any number of nonvolatile oils may be used, including synthetic mono-or diglycerides. Fatty acids such as oleic acid and its glyceride derivatives find use in the preparation of injectable olive oil or castor oil, particularly the polyethoxylates thereof. These oil solutions or suspensions may also contain a long chain alcohol diluent or dispersant.

These compounds can be administered orally in the form of capsules or tablets, or, for example, in the form of aqueous suspensions or solutions. In the case of oral tablets, carriers which are commonly used include lactose and corn starch. Lubricating agents such as magnesium stearate are also typically added. In the case of oral capsules, useful diluents include lactose and dried corn starch. When aqueous suspensions for oral use are desired, the active ingredient will generally be employed in conjunction with emulsifying and suspending agents, and, if desired, certain sweetening and/or flavoring and/or coloring agents may be added.

The compounds of the present invention may also be administered in the form of suppositories for rectal administration of the drug. Such compositions may be prepared by mixing the drug with a suitable non-irritating excipient which is solid at room temperature and liquid at rectal temperature and will melt in the rectum to release the drug. Such materials include cocoa butter, beeswax and polyethylene glycols.

The compounds of the present invention may also be administered topically, particularly in the treatment of conditions involving areas or organs readily accessible by topical application, including neurological conditions of the eye, skin or lower intestinal tract. Suitable topical formulations are readily prepared for each of these areas.

For ophthalmic use, the compounds may be formulated as micronized suspensions in isotonic sterile pH adjusted saline, or, and preferably, as solutions in isotonic and sterile pH adjusted saline, with or without the addition of preservatives such as benzyl chloride alkanolates. In addition, the compound can be formulated into ointment for eye use, such as vaseline ointment.

For topical application to the skin, the compounds of the present invention may be formulated as a suitable cream containing the compound of the present invention dissolved or suspended in a mixture of one or more of the following: mineral oil, liquid petrolatum, white petrolatum, propylene glycol, polyethylene oxide propylene oxide compounds, emulsifying wax and water. In addition, the compounds may be formulated as suitable lotions or creams containing the compounds of the invention dissolved or suspended in one or more of the following mixtures of components: mineral oil, sorbitan, monostearate, tween 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.

Local administration to the lower intestinal tract is carried out as rectal suppositories (see above) or as appropriate enemas.

The dosage of the active ingredient compound useful in the treatment of the above conditions is from about 0.1mg to about 10,000mg, preferably from 0.1mg to about 1,000 mg. The amount of active ingredient that may be employed in conjunction with the carrier materials to produce a single formulation is dependent upon the host treated and the particular mode of administration.

It will be understood, however, that the specific dose for any particular patient will vary with a number of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, rate of excretion, drug combination and the severity of the particular disease undergoing therapy and the form of administration.

The compounds of the present invention may be administered with other neurotrophic agents, such as Neurotrophic Growth Factor (NGF), glial-derived growth factor, brain-derived growth factor, ciliary neurotrophic factor, and neurotropic-3. The dosage of the other neurotrophic agents will be related to the aforementioned factors as well as the neurotrophic effects of the combination.

Ki testing method

The inhibitory effect of the compounds of the present invention on peptidyl-proline isomerase (rotamase) activity can be determined by the methods described in literature [ hating (Harding, M.W.) et al, Nature 341: 758-; holt et al, journal of the American society for chemistry (J.am.chem.Soc.) 115: 9923-9938 ] according to the known method described in (1). The values obtained are apparent Ki values, as listed in table 1. The cis-trans isomerism of the alanine-proline bond on the model enzyme substrate N-succinyl-Ala-Ala-Pro-Phe-p-nitroanilide was monitored spectrophotometrically in a chymotrypsin binding assay which releases p-nitroanilide from the trans substrate. The inhibition of this reaction by the addition of different concentrations of inhibitor was measured and the data analyzed to obtain apparent Ki values with the change in the first order rate constant as a function of inhibitor concentration.

950mL of ice-cold assay buffer (25mM HEPES, pH 7.8, 100mM NaCl), 10mL of FKBP (2.5mM in 10mM Tris-Cl (pH7.5), 100mM NaCl, 1mM dithiothreitol), 25mL of chymotrypsin (50mg/mL in 1mM HCl), and 10mL of different concentrations of test compound in dimethyl sulfoxide were added to a plastic cuvette. 5mL of substrate (succinyl-Ala-Phe-Pro-Phe-p-nitroanilide, 5mg/mL in 2.35mL of LiCl trifluoroethanol) was added to initiate the reaction.

The relationship between the light absorption at 390nm and time was monitored by a spectrophotometer for 90 seconds to determine the velocity constant from the data of light absorption and time.

The data for these tests are listed in table I.

TABLE 1

No.	R1	R	Ki
				1	1, 1-dimethylpropyl radical	3-phenyl propyl group	42
2	1, 1-dimethylpropyl radical	3-phenyl-prop-2- (E) -enyl	125
				3	1, 1-dimethylpropyl radical	3- (3, 4, 5-Trimethoxyphenyl) -propyl	200
4	1, 1-dimethylpropyl radical	3- (3, 4, 5-Trimethoxyphenyl) -prop-2- (E) -enyl	65
				5	1, 1-dimethylpropyl radical	3- (4, 5-methylenedioxy) -phenylpropyl	170
6	1, 1-dimethylpropyl radical	3-(4，5-Methylenedioxy) -phenyl-prop-2- (E) -enyl	160
				7	1, 1-dimethylpropyl radical	3-Cyclohexylpropyl group	200
8	1, 1-dimethylpropyl radical	3-cyclohexyl-prop-2- (E) -enyl	600
				9	1, 1-dimethylpropyl radical	(1R) -1, 3-phenyl-1-propyl	52
10	2-furyl radical	3-phenyl propyl group	4000
				11	2-thienyl radical	3-phenyl propyl group	92
12	2-thiazolyl group	3-phenyl propyl group	100
				13	Phenyl radical	3-phenyl propyl group	1970
14	1, 1-dimethylpropyl radical	3- (2, 5-dimethoxy) phenyl-propyl	250
				15	1, 1-dimethylpropyl radical	3- (2, 5-dimethoxy) phenyl-prop-2- (E) -enyl	450
16	1, 1-dimethylpropyl radical	2- (3, 4, 5-trimethoxyphenyl) ethyl	120
				17	1, 1-dimethylpropyl radical	3- (3-pyridyl) propyl group	5
18	1, 1-dimethylpropyl radical	3- (2-pyridyl) propyl group	195
				19	1, 1-dimethylpropyl radical	3- (4-pyridinyl) propyl	23
20	Cyclohexyl radical	3-phenyl propyl group	82

21	Tert-butyl radical	3-phenyl propyl group	95
				22	Cyclohexylethyl group	3-phenyl propyl group	1025
23	Cyclohexylethyl group	3- (3-pyridyl) propyl group	1400
				24	Tert-butyl radical	3- (3-pyridyl) propyl group	3
25	1, 1-dimethylpropyl radical	3, 5-Diphenylpropyl radical	5
				26	Cyclohexyl radical	3- (3-pyridyl) propyl group	9
27	2-thienyl radical	3- (3-pyridyl) propyl group	1000
				28	Tert-butyl radical	3, 3-Diphenylpropyl radical	5
29	Cyclohexyl radical	3, 3-Diphenylpropyl radical	20
				30	2-thienyl radical	3, 3-Diphenylpropyl radical	150

FKBP-12 and cyclohexadecanetriphosphate receptor (IP) in mammalian cells₃R) and ryanodine receptor (RyR). It is believed that the neurotrophic compounds of the present invention dissociate FKBP-12 from these complexes, causing "leakage" of the calcium channel (see, Camylon Cameron et al, 1995). Calcium flow is involved in axonal extension, so IP may be involved in the neurotrophic effects of drugs₃R receptors and ryanodine receptors. Due to the binding site of the drug to FKBP-12 and to IP₃The binding sites of R receptors are the same, we canIt is hypothesized that the drug replaces this channel from FKBP-12.

Chick embryo dorsal root ganglion culture and axonal outgrowth

Dorsal root ganglia were isolated from chick embryos incubated for 10 days. On 12-well plates coated with thin-layer matrix gel, high glucose medium was added with Leberweitz (Liebovitz)15 medium supplemented with 2mM glutamine and 10% fetal bovine serum, in addition to 10. mu.M cytosine beta-D arabinofuranoside (AraC) at 37 ℃ and 5% CO₂Culturing whole ganglion explants under conditions of (1). After 24 hours, the dorsal root ganglia were treated with various concentrations of nerve growth factor, immunophilin ligand, or shared nerve growth factor medications. After 48 hours of drug treatment, the ganglia were observed under phase contrast or Hoffman contrast with a Zeiss (Zeiss) Axiovert inverted microscope, micrographs of explants were taken and the number of axons overgrowth was calculated. Axons longer than the diameter of the dorsal root ganglion were scored as positive values, and the total number of axons counted under each experimental condition was counted. 3-4 dorsal root ganglia were cultured per well, each treatment was done in duplicate.

The data for this experiment are presented in table II. A representative photograph of example 17 is shown in FIG. 1, and the dose response curve for this example is shown in FIG. 2.

TABLE II

Outgrowth of axons in dorsal root ganglia of chickens

Example No. 2	ED50Neurite outgrowth, nM
		1	53
2	105
		3	149
4	190
		5	10
6	75
		10	0.46
11	0.015
		14	2
15	0.8
		16	0.015
17	0.05
		18	30
19	6
		20	0.13
21	0.025
		22	0.66
23	1100
		24	0.014
25	0.50
		26	2
27	500
		28	0.50
29	10
		30	100

Sciatic nerve axonotomy (Axotomy)

Spraque-Dawley males, 6 weeks old, were anesthetized, the sciatic nerve was exposed and then crushed near the hip with forceps. Test compounds or vehicles were injected subcutaneously daily immediately before and 18 days after injury. Sciatic nerve sections were stained with holms silver stain to calculate axon counts and ruxol fast blue stain was used to count myelination levels. After 18 days of injury, there was a dramatic decrease in axon counts in vehicle-treated animals, 50% compared to the uninjured control, and a dramatic decrease in myelination, 90% compared to the uninjured control.

The drug of example 1 (subcutaneous 30mg/kg) was administered daily before and 18 days after injury, with a significant increase in both axonal count (5% reduction compared to the uninjured control group) and myelination (50% reduction compared to the control group) compared to vehicle-treated animals. The significant effect of the compound of example 1 is consistent with its potent activity in inhibiting rotamase activity and stimulating neurite outgrowth in the chick embryo dorsal root ganglion. These results are presented in table 3, "pretend" indicates that the control animals received vehicle alone but not injured; by "excipient" is meant an animal that is injured and that receives only excipient (i.e., no drug). Example 1 demonstrates a clear similarity to sham-treated animals, showing a potent neuroregenerative effect of this class of compounds in vivo. Inactive are those compounds that are not active as FKBP12 inhibitors. Animals treated with this compound were similar to vehicle-treated wounded animals, consistent with the results observed in example 1 for nerve regeneration directly caused by its inhibition of FKBP 12. These data are shown in table III.

TABLE III

Treatment of	Number of axons (% of control)	Myelin level
			Camouflage	100	100
Injury of the skin
			+ Excipient (subcutaneous)	50	10
+ example 1(30mg/kg subcutaneous)	100	50
			+ Inactive (30mg/kg subcutaneous)	25	25

Model of paralysis agitans MPTP mouse

Mice with MPTP injury to dopaminergic neurons were used as an animal model for paralysis agitans. 4-week-old male CD1 mice were given a dose of 30mg/kg MPTP intraperitoneally for 5 days. Example 17 (10-40 mg/kg) or vehicle was administered together with MPTP subcutaneously for 5 days, and further administered 5 days after the MPTP treatment was stopped. Animals were sacrificed 18 days after MPTP treatment, striatum isolated and homogenized. Binding of [3H ] -CFT, a radioligand for dopamine transporter, to the striatal membrane was performed to quantify the levels of dopamine transporter (DAT) following injury and drug treatment. Immunostaining was performed with anti-tyrosine hydroxylase Ig on sagittal and coronal sections of the brain, and viable and restored dopaminergic neurons were counted. In the group of animals treated with MPTP and vehicle, a significant loss of functional dopaminergic termini was observed compared to the group of non-injured animals. The TH-stained dopaminergic neurons of the injured group of animals that received the compound of example 17 showed near-quantitative recovery.

FIGS. 4 and 5 show the quantification of DAT levels, while FIGS. 6-8 show the regeneration effect of the compound of example 17 in this model. FIG. 4 shows that functional dopaminergic termini are significantly restored compared to animals receiving MPTP but not Gilford (Guilford) compounds, as determined by the [3H ] -CFT binding assay. Figure 5 presents this data in bar chart form. It was shown that the group of animals receiving 40mg/kg of the compound of example 17 in addition to MPTP showed a restoration of [3H ] -CFT binding of more than 90%. As shown in FIGS. 6-8, immunostaining for tyrosine hydroxylase (visible as a marker for dopaminergic neurons) in the striatum, the substantia, and the mid-proximal forebrain tract indicated a clear and significant recovery of functional neurons in the group of animals receiving the compound of example 17 compared to the group of animals receiving the nociceptive but not the drug (i.e., MPTP/vehicle).

The following examples are preferred illustrative embodiments of the present invention and are not intended to limit the invention thereto. The molecular weights of all polymers are average molecular weights. Unless otherwise indicated, all percentages are by weight based on the final delivery system or formulation prepared, and the total is 100% by weight.

Examples

The compounds of the present invention can be prepared by utilizing different synthetic sequences of established chemical transformations. The general route for preparing the present compounds is depicted in scheme I. N-glyoxylproline derivatives were prepared by reacting L-proline methyl ester with oxalato-methyl ester monoacyl chloride as shown in scheme I. The obtained oxalate can react with various nucleophilic carbons to obtain an intermediate compound. These intermediates are then reacted with various alcohols, amides or protected amino acid residues to give the propyl esters and amides of the invention.

Scheme I

Example 1

Synthesis of (2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid-3-phenyl-1-propyl ester (example 1)

Synthesis of (2S) -1- (1, 2-dioxo-2-methoxyethyl) -2-pyrrolidinecarboxylic acid methyl ester

A solution of L-proline methyl ester hydrochloride (3.08 g; 18.60mmol) in anhydrous dichloromethane was cooled to 0 ℃ and treated with triethylamine (3.92 g; 38.74 mmol; 2.1 eq). After the resulting slurry was stirred for 15 minutes under a nitrogen atmosphere, a solution of oxalic acid-methyl ester monoacid chloride (3.20 g; 26.12mmol) in dichloromethane (45mL) was added dropwise. The resulting mixture was stirred at 0 ℃ for 1.5 hr. After filtering to remove solidsThe organic phase was washed with water and MgSO₄Dried and concentrated. The crude residue was chromatographed on a silica gel column eluting with 50% ethyl acetate in hexane to give 3.52g (88%) of a reddish oily product, a mixture of cis-trans amide rotamers, giving the data for trans-rotamers¹H NMR(CDCl₃): d 1.93(dm, 2H); 2.17(m, 2H); 3.62(m, 2H); 3.71(s, 3H); 3.79, 3.84(s, 3H, total); 4.86(dd, 1H, J ═ 8.4, 3.3).

Synthesis of (2S) -1- (1, 2-dioxo-3, 3-dimethylpentyl) -2-pyrrolidinecarboxylic acid methyl ester

A solution of methyl (2S) -1- (1, 2-dioxo-2-methoxyethyl) -2-pyrrolidinecarboxylate (2.35 g; 10.90mmol) in 30mL of Tetrahydrofuran (THF) was cooled to-78 deg.C and treated with 14.2mL of a 1.0M solution of 1, 1-dimethylpropylmagnesium chloride in THF. Stirring the resulting homogeneous mixture at-78 deg.C for 3hr, pouring saturated NH₄Cl solution (100mL) and extracted with ethyl acetate. The organic phase was washed with water, dried and concentrated. The solvent was removed from the obtained crude material, and the crude material was eluted on a silica gel column with 25% ethyl acetate in hexane to obtain 2.10g (75%) of oxalate as a colorless oil.¹H NMR(CDCl₃): d 0.88(t, 3H); 1.22, 1.26(s, 3H each); 1.75(dm, 2H); 1.87-2.10(m, 3H); 2.23(m, 1H); 3.54(m, 2H); 3.76(s, 3H); 4.52(dm, 1H, J ═ 8.4, 3.4).

Synthesis of (2S) -1- (1, 2-dioxo-3, 3-dimethylpentyl) -2-pyrrolidinecarboxylic acid

A mixture of methyl (2S) -1- (1, 2-dioxo-3, 3-dimethylpentyl) -2-pyrrolidinecarboxylate (2.10 g; 8.23mmol), 1N LiOH (15mL) and methanol (50mL) was stirred at 0 ℃ for 30 minutes and then at room temperature overnight. The mixture was acidified to pH1 with 1N HCl, diluted with water, and extracted with 100mL dichloromethane. The organic extracts were washed with brine and concentrated to give 1.73g (87%) of a snow white solid which was not further purified.¹H NMR(CDCl₃): d 0.87(t, 3H); 1.22, 1.25(s, 3H each); 1.77(dm，2H)；2.02(m，2H)；2.17(m，1H)；2.25(m，1H)；3.53(dd，2H，J＝10.4，7.3)；4.55(dd，1H，J＝8.6，4.1)。

Synthesis of (2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester (example 1)

A mixture of (2S) -1- (1, 2-dioxo-3, 3-dimethylpentyl) -2-pyrrolidinecarboxylic acid (600 mg; 2.49mmol), 3-phenyl-1-propanol (508 mg; 3.73mmol), dicyclohexylcarbodiimide (822 mg; 3.98mmol), camphorsulfonic acid (190 mg; 0.8mmol) and 4-dimethylaminopyridine (100 mg; 0.8mmol) in dichloromethane (20mL) was stirred under nitrogen overnight. The reaction mixture was filtered through celite to remove solids and concentrated in vacuo, and the crude product was purified by flash chromatography (25% ethyl acetate/hexanes) to give 720mg (80%) of the compound of example 1 as a colorless oil.¹H NMR(CDCl₃)：d 0.84(t，3H)；1.19(s，3H)；1.23(s，3H)；1.70(dm，2H)；1.98(m，5H)；2.22(m，1H)；2.64(m，2H)；3.47(m，2H)；4.14(m，2H)；4.51(d，1H)；7.16(m，3H)；7.26(m，2H)。

The following illustrative examples were prepared using the procedure of example 1:

example 2

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-prop-2- (E) -enyl ester, 80%,¹H NMR(360MHz，CDCl₃)：d 0.86(t，3H)；1.21(s，3H)；1.25(s，3H)；1.54-2.10(m，5H)；2.10-2.37(m，1H)；3.52-3.55(m，2H)；4.56(dd，1H，J＝3.8，8.9)；4.78-4.83(m，2H)；6.27(m，1H)；6.67(dd，1H，J＝15.9)；7.13-7.50(m，5H)。

example 3

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3, 4, 5-trimethoxyphenyl) -1-propyl ester, 61%,¹H NMR(CDCl₃)：d 0.84(t，3H)；1.15(s，3H)；1.24(s，3H)；1.71(dm，2H)；1.98(m，5H)；2.24(m，1H)；2.63(m，2H)；3.51(t，2H)；3.79(s，3H)；3.83(s，3H)；4.14(m，2H)；4.52(m，1H)；6.36(s，2H)。

example 4

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3, 4, 5-trimethoxyphenyl) -1-prop-2- (E) -enyl ester, 66%,¹H NMR(CDCl₃)：d 0.85(t，3H)；1.22(s，3H)；1.25(s，3H)；1.50-2.11(m，5H)；2.11-2.40(m，1H)；3.55(m，2H)；3.85(s，3H)；3.88(s，6H)；4.56(dd，1H)；4.81(m，2H)；6.22(m，1H)；6.58(d，1H，J＝16)；6.63(s，2H)。

example 5

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-methylenedioxyphenyl) -1-propyl ester, 82%,¹H NMR(360MHz，CDCl₃)：d 0.86(t，3H)；1.22(s，3H)；1.25(s，3H)；1.60-2.10(m，5H)；3.36-3.79(m，2H)；4.53(dd，1H，J＝3.8，8.6)；4.61-4.89(m，2H)；5.96(s，2H)；6.10(m，1H)；6.57(dd，1H，J＝6.2，15.8 )；6.75(d，1H，J＝8.0)；6.83(dd，1H，J＝1.3，8.0)；6.93(s，1H)。

example 6

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4, 5-methylenedioxyphenyl) -1-prop-2- (E) -enyl ester, 82%,¹H NMR(360MHz，CDCl₃)：d 0.86(t，3H)；1.22(s，3H)；1.25(s，3H)；1.60-2.10(m，5H)；2.10-2.39(m，1H)；3.36-3.79(m，2H)；4.53(dd，1H，.J＝3.8，8.6)；4.61-4.89(m，2H)；5.96(s，2H)；6.10(m，1H)；6.57(dd，1H，J＝6.2，15.8)；6.75(d，1H，J＝8.0)；6.83(dd，1H，J＝1.3，8.0)；6.93(s，1H)。

example 8

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3-cyclohexyl-1-prop-2- (E) -enyl ester, 92%,¹H NMR(360MHz，CDCl₃): d 0.86(t, 3H); 1.13-1.40(m +2 singlets, 9H total); 1.50-1.87(m, 8H); 1.87-2.44(m, 6H); 3.34-3.82(m, 2H); 4.40-4.76(m, 3H); 5.35-5.60(m, 1H); 5.60-5.82(dd, 1H, J ═ 6.5, 16).

Example 9

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid (1R) -1, 3-diphenyl-1-propyl ester, 90%,¹H NMR(360MHz，CDCl₃)：d 0.85(t，3H)；1.20(s，3H)；1.23(s，3H)；1.49-2.39(m，7H)；2.46-2.86(m，2H)；3.25-3.80(m，2H)；4.42-4.82(m，1H)；5.82(td，1H，J＝1.8，6.7)；7.05-7.21(m，3H)；7.21-7.46(m，7H)。

example 10

(2S) -1- (1, 2-dioxo-2- [ 2-furyl ] -ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 99%,¹H NMR(300MHz，CDCl₃)：d 1.66-2.41(m，6H)；2.72(t，2H，J＝7.5)；3.75(m，2H)；4.21(m，2H)；4.61(m，1H)；6.58(m，1H)；7.16-7.29(m，5H)；7.73(m，2H)。

example 11

(2S) -1- (1, 2-dioxo-2- [ 2-thienyl ] -ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 81%,¹H NMR(300MHz，CDCl₃)：d 1.88-2.41(m，6H)；2.72(dm，2H)；3.72(m，2H)；4.05(m，1H)；4.22(m，1H)；4.64(m，1H)；7.13-7.29(m，6H)；7.75(dm，1H)；8.05(m，1H)。

example 13

(2S) -1- (1, 2-dioxo-2-phenyl) ethyl-2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 99%,¹H NMR(300MHz，CDCl₃)：d 1.97-2.32(m，6H)；2.74(t，2H，J＝7.5)；3.57(m，2H)；4.24(m，2H)；4.67(m，1H)；6.95-7.28(m，5H)；7.51-7.64(m，3H)；8.03-8.09(m，2H)。

example 14

(2S)-1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-propyl ester, 99%,¹H NMR(300MHz，CDCl₃)：d 0.87(t，3H)；1.22(s，3H)；1.26(s，3H)；1.69(m，2H)；1.96(m，5H)；2.24(m，1H)；2.68(m，2H)；3.55(m，2H)；3.75(s，3H)；3.77(s，3H)；4.17(m，2H)；4.53(d，1H)；6.72(m，3H)。

example 15

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2, 5-dimethoxyphenyl) -1-prop-2- (E) -enyl ester, 99%,¹H NMR(300MHz，CDCl₃)：d 0.87(t，3H)；1.22(s，3H)；1.26(s，3H)；1.67(m，2H)；1.78(m，1H)；2.07(m，2H)；2.26(m，1H)；3.52(m，2H)；3.78(s，3H)；3.80(s，3H)；4.54(m，1H)；4.81(m，2H)；6.29(dt，1H，J＝15.9)；6.98(s，1H)。

example 16

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 2- (3, 4, 5-trimethoxyphenyl) -1-ethyl ester, 97%,¹H NMR(300MHz，CDCl₃)：d 0.84(t，3H)；1.15(s，3H)；1.24(s，3H)；1.71(dm，2H)；1.98(m，5H)；2.24(m，1H)；2.63(m，2H)；3.51(t，2H)；3.79(s，3H)；3.83(s，3H)；4.14(m，2H)；4.52(m，1H)；6.36(s，2H)。

example 17

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester, 80%,¹H NMR(CDCl₃300 MHz): d 0.85(t, 3H); 1.23-1.26(s, 3H each); 1.63-1.89(m, 2H); 1.90-2.30(m, 4H); 2.30-2.50(m, 1H); 2.72(t, 2H); 3.53(m, 2H); 4.19(m, 2H); 4.53(m, 1H); 7.22(m, 1H); 7.53(dd, 1H); 8.45.

example 18

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (2-pyridinyl) -1-propaneThe amount of ester, 88%,¹H NMR(CDCl₃300 MHz): d 0.84(t, 3H); 1.22-1.27(s, 3H each); 1.68-2.32(m, 8H); 2.88(t, 2H, J ═ 7.5); 3.52(m, 2H); 4.20(m, 2H); 4.51(m, 1H); 7.09-7.19(m, 2H); 7.59(m, 1H); 8.53(d, 1H, J ═ 4.9).

Example 19

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3- (4-pyridyl) -1-propyl ester, 91%,¹H NMR(CDCl₃，300MHz)：d 6.92-6.80(m，4H)；6.28(m，1H)；5.25(d，1H，J＝5.7)；4.12(m，1H)；4.08(s，3H)；3.79(s，3H)；3.30(m，2H)；2.33(m，1H)；1.85-1.22(m，7H)；1.25(s，3H)；1.23(s，3H)；0.89(t，3H，J＝7.5)。

example 20

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 91%,¹H NMR(CDCl₃300 MHz): d 1.09-1.33(m, 5H); 1.62-2.33(m, 12H); 2.69(t, 2H, J ═ 7.5); 3.15(dm, 1H); 3.68(m, 2H); 4.16(m, 2H); 4.53, 4.84(d, 1H total); 7.19(m, 3H); 7.29(m, 2H).

Example 21

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 92%,¹H NMR(CDCl₃，300MHz)：d 1.29(s，9H)；1.94-2.03(m，5H)；2.21(m，1H)；2.69(m，2H)；3.50-3.52(m，2H)；4.16(m，2H)；4.53(m，1H)；7.19(m，3H)；7.30(m，2H)。

example 22

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3-phenyl-1-propyl ester, 97%,¹H NMR (CDCl₃，300MHz)：d 0.88(m，2H)；1.16(m，4H)；1.43-1.51(m，2H)；1.67(m，5H)；1.94-2.01(m，6H)；2.66-2.87(m，4H)；3.62-3.77(m，2H)；4.15(m，2H)；4.86(m，1H)；7.17-7.32(m，5H)。

example 23

(2S) -1- (2-cyclohexylethyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester, 70%,¹H NMR(CDCl₃，300MHz)：d 0.87(m，2H)；1.16(m，4H)；1.49(m，2H)；1.68(m，4H)；1.95-2.32(m，7H)；2.71(m，2H)；2.85(m，2H)；3.63-3.78(m，2H)；4.19(m，2H)；5.30(m，1H)；7.23(m，1H)；7.53(m，1H)；8.46(m，2H)。

example 24

(2S) -1- (2-tert-butyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester, 83%,¹H NMR(CDCl₃，300MHz)：d 1.29(s，9H)；1.95-2.04(m，5H)；2.31(m，1H)；2.72(t，2H，J＝7.5)；3.52(m，2H)；4.18(m，2H)；4.52(m，1H)；7.19-7.25(m，1H)；7.53(m，1H)；8.46(m，2H)。

example 25

(2S) -1- (3, 3-dimethyl-1, 2-dioxopentyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, 99%,¹H NMR(CDCl₃300 MHz): d 0.85(t, 3H); 1.21, 1.26(s, 3H each); 1.68-2.04(m, 5H); 2.31(m, 1H); 2.40(m, 2H); 3.51(m, 2H); 4.08(m, 3H); 4.52(m, 1H); 7.18-7.31(m, 10H).

Example 26

(2S) -1- (2-cyclohexyl-1, 2-dioxoethyl) -2-pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester, 88%,¹H NMR(CDCl₃，300MHz)：d 1.24-1.28(m，5H)；1.88-2.3 5(m，11H)；2.72(t，2H，J＝7.5)；3.00-3.3 3(dm，1H)；3.69(m，2H)；4.19(m，2H)；4.55(m，1H)；7.20-7.24(m，1H)；7.53(m，1H)；8.47(m，2H)。

example 27

(2S) -N- ([ 2-thienyl ] glyoxyl) pyrrolidinecarboxylic acid 3- (3-pyridyl) -1-propyl ester, 49%,¹H NMR (CDCl₃，300MHz)：d 1.81-2.39(m，6H)；2.72(dm，2H)；3.73(m，2H)；4.21(m，2H)；4.95(m，1H)；7.19(m，2H)；7.61(m，1H)；7.80(d，1H)；8.04(d，1H)；8.46(m，2H)。

example 28

(2S) -1- (3, 3-dimethyl-1, 2-dioxobutyl) -2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, 99%,¹H NMR(CDCl₃，300MHz)：d 1.27(s，9H)；1.96( m，2H)；2.44(m，4H)；3.49(m，1H)；3.64(m，1H)；4.08(m，4H)；4.53(dd，1H)；7.24(m，10H)。

example 29

(2S) -1-Cyclohexylglyoxyloyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, 91%,¹H NMR(CDCl₃，300MHz)：d 1.32(m，6H)；1.54-2.41(m，10H)；3.20(dm，1H)；3.69(m，2H)；4.12(m，4H)；4.52(d，1H)；7.28(m，10H)。

example 30

(2S) -1- (2-thienyl) glyoxyl-2-pyrrolidinecarboxylic acid 3, 3-diphenyl-1-propyl ester, 75%,¹H NMR (CDCl₃300 MHz): d 2.04(m, 3H); 2.26(m, 2H); 2.48(m, 1H); 3.70(m, 2H); 3.82-4.18(m, 3H total); 4.64(m, 1H); 7.25(m, 11H); 7.76(dd, 1H); 8.03(m, 1H).

The requisite substituted alcohols can be prepared by a number of methods well known to those skilled in the art of organic synthesis. As shown in scheme II, the alkyl or aryl aldehydes can be recarburized to phenylpropanol by reaction with methyl (triphenylphosphidenyl) acetate to provide various trans-cinnamates which can be reduced to saturated alcohols by reaction with excess lithium aluminum hydride or subsequent reduction of the double bond by catalytic hydrogenation and reduction of the saturated ester with a suitable reducing agent. On the other hand, trans-cinnamate can be reduced to allyl alcohol using diisobutylaluminum hydride.

Scheme II

Longer chain alcohols can be prepared by the recarburization of benzaldehyde and higher aldehydes. Alternatively, these aldehydes may be prepared by conversion of the corresponding phenylacetic acid or higher acids with phenylethanol and higher alcohols.

A general method for the synthesis of acrylic esters such as methyl (3, 3, 5-trimethoxy) -trans-cinnamate is:

a solution of 3, 4, 5-trimethoxybenzaldehyde (5.0 g; 25.48mmol) and methyl (triphenylphosphanylidene) acetate (10.0 g; 29.91mmol) in tetrahydrofuran (250mL) was refluxed overnight. After cooling, the reaction mixture was diluted with 200mL of ethyl acetate, washed with 2X 200mL of water, dried and concentrated in vacuo. The crude residue was chromatographed on a silica gel column eluting with 25% ethyl acetate/hexane to give 5.63g (88%) of a white crystalline solid cinnamate,¹H NMR(300MHz，CDCl₃)：d 3.78(s，3H)；3.85(s，6H)；6.32(d，1H，J＝16)；6.72(s，2H)；7.59(d，1H，J＝16)。

a general method for the synthesis of saturated alcohols from acrylic esters is exemplified by (3, 4, 5-trimethoxy) phenyl propanol.

A solution of methyl (3, 3, 5-trimethoxy) -trans-cinnamate (1.81 g; 7.17mmol) in THF (30mL) was added dropwise to a solution of lithium aluminum hydride (14mmol) in THF (35mL) while stirring under argon. After completion of the dropwise addition, the mixture was heated to 75 ℃ to react for 4 hr. After cooling, the reaction was quenched by careful addition of 15mL of 2N NaOH and an additional 50mL of water. The resulting mixture was filtered through celite to remove solids, and the filter cake was washed with ethyl acetate. The combined organic fractions were washed with water, dried and concentrated in vacuo and purified on a silica gel column eluting with ethyl acetate to give 0.86g (53%) of alcohol as a clear oil.¹H NMR(300MHz，CDCl₃)：d 1.23(br，1H)；1.87(m，2H)；2.61(t，2H，J＝7.1)；3.66(t，2H)；3.80(s，3H)；3.83(s，6H)；6.40(s，2H)。

A general procedure for the synthesis of trans-allylic alcohols from acrylic esters is exemplified by (3, 4, 5-trimethoxy) phenylprop-2- (E) -enol.

A solution of methyl (3, 3, 5-trimethoxy) -trans-cinnamate (1.35 g; 5.35mmol) in toluene (25mL) was cooled to-10 deg.C and treated with a solution of diisobutylaluminum hydride in toluene (11.25mL, 1.0M solution; 11.25mmol), the reaction mixture was stirred at 0 deg.C for 3hr, then quenched with 3mL of methanol, followed by 1N HCl until the pH was brought to 1. The reaction mixture was extracted with ethyl acetate, and the organic phase was washed with water, dried and concentrated. Purification was carried out on a silica gel column eluting with 25% ethyl acetate/hexane to give 0.96g (80%) of the product as a viscous oil,¹H NMR(360MHz，CDCl₃)：d 3.85(s，3H)；3.87(s，6H)；4.32(d，2H，J＝5.6)；6.29(dt，1H，J＝15.8，5.7)；6.54(d，1H，J＝15.8)6.61(s，2H)。

having thus described the invention, it will be obvious that the same may be varied in many ways. Such variations are not to be regarded as a departure from the spirit and scope of the invention, and all such modifications are intended to be included within the scope of the following claims.

Claims (1)

1. The compound (2S) -1- (1, 2-dioxo-3, 3-dimethylpentyl) -2-pyrrolidinecarboxylic acid.