CZ307096A3 - Methods of video analysis by making use of metal complexes of nitrogen-containing macrocyclic ligands - Google Patents

Abstract

The present invention is directed to complexes represented by formula (I) wherein R, R', R1, R'1, R2, R'2, R3, R'3, R4, R'4, R5, R'5, R6, R'6, R7, R'7, R8, R'8, R9, R'9, R10, R11, R12, R13, R14, M, X, Y, Z and N are defined herein for use as contrast agents in diagnostic imaging.

Description

Methods of imaging analysis using metal complexes of nitrogen-containing macrocyclic ligands

Technical field

The invention relates to compounds effective as contrast agents in imaging. The invention relates to magnetic resonance imaging (MRI) of human or non-human animal subjects using metal complexes of substituted, 15 membered nitrogen-containing macrocyclic ligands as contrast agents. The invention relates to divalent manganese complexes in substituted 15-membered nitrogen-containing macrocyclic ligands as co-agents for MRI.

X-rays have long been used to visualize human and non-human animal tissues, i.e., the patient's internal organs, with the patient positioned between the X-ray source and the film sensitive to the beams. Where the pigs interfere with the passage of the rays, the film is less exposed and the resulting exposed film shows the state of the organ.

More recently, nuclear magnetic resonance (NMR) has also been developed as an imaging technique, ie MRI. MRI avoids the harmful effects sometimes accompanying exposure to X-rays. To improve X-ray imaging, contrast agents are administered to patients prior to scanning, either orally or parenterally. After a certain period of time required to distribute the contrast agent in the patient, sensing is performed. In order to obtain a good image, it is desirable that the time from receipt of the contrast agent be minimal. On the other hand, the efficiency of kle2 is with time, so that it is desirable that the excretion is relatively slow to ensure a sufficient time period during which imaging can be performed.

In the NMR imaging process, body fluid protons relax by two mechanisms, described as T and T. The rate at which the relaxation process occurs can be altered for some water molecules, producing values different from the norm.

Compounds that enhance NMR imaging, referred to as contrast agents, are generally paramagnetic in nature. These may be organic free radicals or transition or lanthanide metals having one to seven unpaired electrons.

A prerequisite for any ligand that binds metal to form a contrast agent is that the resulting contrast agent must be stable to prevent metal loss and subsequent accumulation in the body. Other conditions include the ability to reversibly bind water, which increases its contrast properties of the substance and reduces the amount of substance required. This ability is clearly important as the interaction between any two nuclear spins in space decreases inversely proportionally to the sixth power of the distance.

U.S. Patent No. 4,647,447 discloses the use of a NMR imaging contrast agent that contains salts of a complexing acid anion and a paramagnetic metal anion. A preferred embodiment is the incorporation of diethylenetriaminepentaacetic acid (Gd DTPA) gadolinium chelate, which is now commercially available from Nycomed Salutar, Inc. under the trade name Magnevist for use as a contrast agent for NMR. From the data reported there, these substances appear to be functional. However, this substance is rapidly excreted by the kidneys, so the timing of the injection is extremely critical. In addition, the substance is hardly absorbed by any solid organ, such as the heart, pancreas or liver.

Many gadoline contrast agents are already known, but there remains the possibility that small amounts of free lanthanides will be released into the body upon decomposition. Since it is not a metal normally found in the body, little is known about its long-term effects.

Other nitrogen-containing macrocyclic ligands have been proposed for use as NMR contrast agents. S. C. Jackeis et al., Aqueous Prcton NMR Relaxation Snhancements by Manganese and Macrocyclic Complexes? 51 ruc ture-R eiaxi Víty Relationships, Inorg. Chem., 31, 234-239 (1992) discloses 15-membered nitrogen-containing ring complexes. The disadvantage of these compounds, however, is that they are insufficiently stable, possibly colored, and thus are unsuitable for application as MRI contrast agents.

Therefore, it would be highly desirable to develop alternative contrast agents that overcome one or more of the above disadvantages.

It has now been discovered that metal complexes of substituted, nitrogen-containing macrocyclic ligands having increased kinetic, thermodynamic and oxidative stability, and which can be substituted to control lipophilicity, i.

bio-distribution, eliminates the problems of the above-mentioned contrast agents while ensuring good contrast efficacy. SUMMARY OF THE INVENTION

It is an object of the present invention to provide a magnetic resonance imaging (MRI) contrast agent with enhanced kinetic stability, i.e., the rate at which the paramagnetic metal dissociates from the metal complexes of the invention. It is a further object of the invention to provide contrast agents for MRIs in which the biodistribution of the contrast agent can be controlled. Yet another object of the invention is the preparation of MRI contrast agents with enhanced oxidation stability and enhanced hydrogen bonding. Yet another object of the invention is to prepare metal complexes which are useful as x-ray or ultrasound contrast agents and which can be used in scintigraphy and radiotherapy.

The present invention provides a magnetic resonance imaging method comprising administering to a human or non-human animal subject a contrast agent comprising a physiologically compatible paramagnetic metal complex of the invention and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle, and generating a magnetic residual image of at least a portion of the subject.

The invention further provides a method of diagnostic imaging by administering to a human or non-human animal subject a diagnostic agent representing a physiologically compatible heavy metal complex of the invention and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle, and producing an X-ray, sonographic or scintigraphic image. at least a portion of the subject.

The invention further provides a method of radiotherapy carried out on a human or non-human animal subject by administering to the subject a radioactive agent representing a physiologically compatible radioactive metal complex of the invention and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle.

Exemplary embodiments of the invention

The invention is further described by way of example embodiments, the metal complexes of the invention being used as MRI contrast agents, as X-ray diagnostic agents, sonographic or scintigraphic imaging analysis, or as radiotherapeutic agents, are represented by the formula;

where R, R ' ¹ , R, R ', R, R ', R, R ¹ , R, R ', G ' ~ 7 ' groups: hydrogen, alkyl, alkenyl, cycloalkylalkyl

and R 'are independently selected from alkenyl, alkynyl, cycloalkyl, cyclo, cycloalkylcycloalkyl, cycloalkenylalkyl, alkylcycloalkyl, alkenylcycloalkyl, alkylcycloalkenyl, alkenylcycloalkenyl, heterocycle, aryl and aralkyl;

groups and groups attached to α-carbon α-amino acids; either R or R 'a R or R ¹ , R or R ¹ a R or R ¹ , R ^No_ .1 1 Li 3 3 J 4 ' or R ¹ a R: or R ¹ , R or R 'and R' or R ¹ and R or 5 tS G ~ 7 7B and 9 R ¹ and R o or: R ’ together with carbon atoms they are on navá- inflammation, forms nez independently saturated, partially saturated or nen- carbonated cycles having 3 to 20 carbon atoms; R and R ¹ and R ', R' and R 1 , R and R ', R and R', R and R ¹ _f R and R ¹ , R 2. 3 3 4 4 5 <5 ň 'v and R ', R' -7 and R O l 8 and R _g and R _g together with the carbon atoms, on which are linked F independently form saturated, partly ; saturated or unsaturated ring structures having 3 to 20 uh- atoms Líku; or bleached from R, R ', R, R', R, R ', R, R ’, R, ^r 1 1 2 2 3 3 R ', R, R ', R , R ', R, R', R, R ¹ , R, R ¹ , R, R, -1 75 & X o 1 L R, R and R , together with another R, R ¹ , R, R ¹ , R R ’.., L 2 _I 3 1 4 1 i - R- "R", R, R ', R, R', R, R ', R, R-, R, R ¹ , R, and R ', R 9 1 o , R 1 X ' R, R and R which are 1, 2 .L 3 14 established on different

the carbon or nitrogen atom in the macrocyclic ligand may be bonded to form a chain represented by the formula ch ch ₂ m -e CH ₂ κ ₂ * _{2 y y} where w, x, y and z are independent integers from 0 to 10, and M, L and J are independently selected from the group consisting of alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroarl, alkaryl, alkheteroaryl, aza, amide, ammonium, thia, sulfonyl, sulfinyl, sulfonamide, phosphonyl, phosphinyl, phosphino, phosphonium keto, ester, carbamate, urea, thiocarbonyl, borate, borane, borase, silyl, siloxy, silaza, and combinations thereof;

R and R, Ρ. , R, R, and R are independently selected from the group consisting of hydrogen, alkyl and alkyl substituted with -OR, -COOR, -CONR. R or POΗ, wherein R and R are independently hydrogen or alkyl; and wherein at least two of R, R ', R, R', R, R ', R, R', R, R ', R, R ¹ , R, R', R, R ¹ , R, R ¹ , R ¹ and R ¹ 'are other than hydrogen.

Θ N 9 9 ^JJ

X, Y and Z represent suitable ligands or charge neutralizing anions derived from a monodentate or polydentate coordination ligand or ligand system or their corresponding anion (e.g., benzoic acid or benzoate anion, phenol or phenoxide anion, alcohol or alkoxide anion). and Z are independently selected from the group consisting of halide, oxo, aguo, hydroxo, alcohol, phenol, dioxygen, peroxo, hydroperoxo, alkylperoxo, aryperoxo, ammonia, alkylamino, arylamino, heterocycloalkylamino, heterocycloarvlamino, amine oxides, hydrazine, alkylhydrazine, arylhydrazine, nitrogen dioxide, cyanide, cyanate, thiocyanate, isocyanate, isothiocyanate, alkyl nitrile, aryl nitrile, alkylisonitrile, arylisonitrile, nitrate, nitrite, azido, alkylsulfonic acid, arylsulfonic acid, alkylsulfoxide, arylsulfoxide, alkyllarylsulfenulfide, alkylsulfenesulfoxide, alkylsulfenesulfoxide, alkylsulfulfide, alkylsulfulfide arylsulfinic acid, alkylthiol carboxylic acid, arylthiol carboxylic acid, alkylthiolthiocarboxylic acid, arylthiolthiocarboxylic acid, alkylcarboxylic acid (e.g. acetic acid, trifluoroacetic acid, oxalic acid), arylcarboxylic acid (e.g. benzoic acid, phthalic acid), urea, alkylurea, arylurea, alkylthiourea, alkyllaryl ary1thíomočovina, alkylarylthíomočovina, sulfate, sulfite, hyčrogensíran, bisulfite, thiosulfate, thiosulfite, hydrosulfite (hyčrogensířičitan), alkyl phosphine, aryl phosphine, alkyl phosphine oxides, arylfosfinoxid, alkylarylfosfinoxid, alkylfosfinsulfid, arylfosfinsulfid, alkylarylfosfinsulfid, alkvlfosfonová acid, alkylphosphonic acid, Alkyl phosphonium acid-phosphinic acid alkylfosfinitá acid, aryl phosphinic acid, phosphate, thiophosphate, phosphite, pyrophosphite, triphosphate, hydrogen phosphate, dihydrogen phosphate, alkylguanidines, arylguanidino, alkylarylguanidino, alkylcarbamate, arylcarbamates, alkylaryl carbamates, alkylthiocarbamates, arylthiokarbamát, alkylarylthiokarbamát, alkyldithiocarbamates, aryldithiokarbamát, alkylaryldithiokarbamát, bicarbonate, carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate, bromite, hypobromite, tetrahalomanganan, tetrafluoroborate, hexafluorophosphate, hexafluoroantimonate, hypophosphite, iodate, periodate, metaborate, tetraarylborate, tetraalkylborate, tartrate, salicylate, succinate, citrate, ascorbate, saccharinate, amino acid, hydroxamic acid, thiotosylate and ion exchange resin anions, or systems wherein one or more of X; are independently attached to one or more R groups wherein n is an integer from 0 to 3. Preferred ligands from which X, Y and Z are selected include halide, organic acid, anion of nitrate or bicarbonate.

The metal atoms or anions, M which are suitable for use in the complexes of the invention as MRI contrast agents are paramagnetic metals with atomic numbers 21-29, 42-44 and 57-71. Of the complexes for use as MRI contrast agents, the preferred those containing Eu, Gd, Dy, Ho, Cr, Mn or Fe as the metal, preferably Gd (III) or Mn (II), and most preferably Μη (II).

The metal atoms or anions, M, which are suitable for use in the complexes of the invention, as X-ray or ultrasound contrast agents, are heavy metals with atomic numbers 20-32, 42-44, 49 and 57-83. as X-ray or ultrasound contrast agents, those containing a non-radioactive metal having atomic numbers 42-44, 49 and 57-83, and preferably Gd, Dy or Yb, are preferred.

The metal atoms or anions, M, in the complexes of the invention which are suitable for use in scintigraphy and radiotherapy are the radioactive metals of any of the radioactive isotopes entering the complexes, preferably with atomic numbers 20-32, 42-44, 49 and 57-83 . In scintigraphy the most preferred metals are ^99m Tc or in radiotherapy the most preferred metals are ¹⁵³ Sm, ^{s 7} Cu or ⁹ ° Y.

As used herein, the term alkyl, alone or in combination, refers to a straight or branched chain alkyl group containing 1 to about 22 carbon atoms, preferably 1 to 18 carbon atoms, and most preferably 1 to 12 carbon atoms. Examples of such groups include, but are not limited to: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, isoamyl, hexyl, octyl, nonyl, decyl, dodecyl, tetradecyl, hexadecyl, octadecyl and eicosyl. By alkenyl, alone or in combination, is meant an alkyl group having one or more double bonds. Examples of such alkenyl groups include, but are not limited to: ethenyl, propenyl, 1-butenyl, cis-2-butenyl, trans-2-butenyl, iso-butylenyl, cis-2-pentenyl, trans-2-pentenyl,

2,3-dimethyl-2-butenyl, 1-pentenyl, 1-hexenyl, 1-octenyl, decenyl, dodecenyl, tetradecenyl, hexadecenyl, cis- and trans-9-octadecenyl, 1,3-pentadienyl, 2,4-pentadienyl, 2,3 pentadienyl, 1,3-hexadienyl, 2,4-hexadienyl, 5,8,11,14-eicosatetraenyl and 9,12,15-octadecatrienyl. As used herein, the term alkynyl, alone or in combination, refers to an alkyl group having one or more triple bonds. Examples of such alkynyl groups include, but are not limited to: ethynyl, propynylpropargyl), 1-butynyl, 1-octinyl, 9-octadecinyl, 1,3-pentadiinyl, 2,4-pentadinyl,

1,3-hexadiinyl and 2,4-hexadiinyl. As used herein, the term cycloalkyl, alone or in combination, refers to a cycloalkyl group containing from 3 to about 10, preferably from 3 to about 8, and most preferably from 3 to about 6 carbon atoms. Examples of such cycloalkyl groups include, but are not limited to: cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl and perhydronaphthyl. As used herein, the term cycloalkylalkyl means an alkyl group as defined above which is substituted with a cycloalkyl group as defined above. Examples of such cycloalkylalkyl groups include, but are not limited to: cyclohexylmethyl, cyclopentylmethyl, (4-isopropylcyclohexyl) methyl, (4-1-butyl cyclohexyl) methyl, 3-cyclohexylpropyl, 2-cyclohexylmethylpentyl, 3-cyclopentylmethylhexyl, 1- (4-neopentylcyclohexyl) methylhexyl and 1- (4-isopropylcyclohexyl) methylheptyl. As used herein, the term cycloalkylcycloalkyl means a cycloalkyl group as defined above that is substituted with another cycloalkyl group as defined above. Examples of such cycloalkylcycloalkyl groups include, but are not limited to: cyclohexylcyclopentyl and cyclohexylcyclohexyl. As used herein, the term cycloalkenyl, alone or in combination, is intended. a cycloalkyl group having one or more double bonds. Examples of such cycloalkenyl groups include, but are not limited to: cyclopentenyl. cyclohexenyl, cyclococenyl, cyclopentadienyl, cyclohexadienyl and cyclooctadienyl. As used herein, the term cycloalkenylalkyl means an alkyl group as defined above which is substituted with a cycloalkenyl group as defined above. Examples of such cycloalkenylalkyl groups include, but are not limited to: 2-cyclohexen-1-ylmethyl, 1-cyclopenten-1-ylmethyl, 2- (1-cyclohexen-1-yl) ethyl, 3- (1-cyclopenten-1-yl) propyl, 1- (1-cyclohexen-1-ylmethyl) pentyl, 1- (1-cyclopenten-1-yl) hexyl, 6- (1-cyclohexen-1-yl) hexyl, 1- (1-cyclopenten-1-yl) nonyl and 1- (1-cyclohexen-1-yl) nonyl. If it is here. As used herein, the terms alkylcycloalkyl and alkenylcycloalkyl, is meant a cycloalkyl group as defined above which is substituted with an alkyl or alkenyl group as defined above. Examples of alkylcycloalkyl and alkenylcykloalkylových groups include, but are not limited to, 2-ethylcyclobutyl _r l-methylcyclopentyl, l-hexylcykopentyl, L-methylcykohexyl 1- (9oktadecenyl) cyclopentyl and l- (9oktadecenyl) cyclohexyl. As used herein, the terms alkylcycloalkenyl and alkenylcycloalkenyl means a cycloalkenyl group as defined above which is substituted with an alkyl or alkenyl group as defined above. Examples of such alkylcycloalkenyl and alkenylcycloalkenyl groups include, but are not limited to:

1- methyl-2-cyclopentyl-l-phenyl-cyclopent hexy1-2 1-e 2 thy1- cyclohexenyl, l-butyl-2 ₇ cyclohexenyl, 1- (9-octadecenyl) -2cyklohexenyl and 1- (2-pentenylj- As used herein, the term aryl, alone or in combination, means a phenyl or naphthyl group to which optionally one or more substituents selected from: alkyl, cycloalkyl, cycloalkenyl, phenyl, naphthyl may be attached. , heterocycle, alkoxyaryl, alkaryl, alkoxy, halogen, hydroxy, amine, cyano, nitro, alkylthio, phenoxy, ether, trifluoromethyl and the like, for example phenyl, p-tolyl, 4-methoxyphenyl, 4- (tert-butoxy) phenyl, 4-fluorophenyl, 4-chlorophenyl, 4-hydroxyphenyl, 1-naphthyl,

2-naphthyl and the like. If it is here. the term aralkyl, alone or in combination, means an alkyl or cycloalkyl group as defined above in which one hydrogen atom is replaced by an aryl group as mentioned above, such as benzyl, 2-phenylethyl and the like. As used herein, the term heterocycle refers to a ring structure containing at least one other kind of atom in addition to the carbon atom in the ring. The most common of these non-carbon atoms are nitrogen, oxygen and sulfur. Examples of heterocycles include, but are not limited to: pyrrolidinyl, piperidyl, imidazolidinyl, tetrahydrofuryl, tetrahydrothienyl, furyl, thienyl, pyridyl, quinolyl, i.soquinolyl, pyridazinyl, pyrazinyl, indolyl. imidazolyl,. Oxazolyl, thiazolyl, pyrazolyl, pyridinyl, benzoxadiazolyl, benzothiadiazolyl, triazolyl and terazolyloy groups. The term saturated, partially saturated or unsaturated ring refers to fused ring structures in which the 2 ring carbons are also part of a 15 membered macrocyclic ligand. The ring structure may contain 3 to. 20 atoms ^? carbon, preferably 5 to 8 carbon atoms, and may also contain one or more kinds of atoms other than a carbon atom. The most common of these other types of atoms are nitrogen, oxygen and sulfur atoms. The ring structure may also comprise more than one ring. The term saturated, partially saturated or unsaturated ring structure refers to a ring structure in which one ring carbon is also part of a 15 membered macrocyclic ligand. The ring structure may comprise

· Fourth 'to 20 carbon atoms, preferably 5-8 carbon atoms, ₄ a.může also contain nitrogen, oxygen possibly sulfur. The term anion of an organic acid refers to anions of carboxylic acids having probably 1 to 18 carbon atoms.

The term halide refers to chloride or bromide.

The total charge of the complex can vary from negative to positive with respect to nitrogen or carbon substitution of the respective charged groups on the macrocyclic structure.

While the divalent manganese complexes of the invention exist as monocations in the methanolic solution, the axial anions are labile and in. in vivo, they can rapidly exchange with endogenous charged or uncharged ligands. Taking into account the nature of the arrangement of the metal center formed of divalent, manganese, total. The charge -complex- can be adjusted to suit the r-3 as needed. enhance the desired pharmaceutical properties, such as shmolality, tissue distribution and removal. without decomposition. For example, if the complex carries only neutral functional groups, such as N- or C-alkyl substitution, then the overall charge of the complex will be determined by the manganese center /, and will be positive. Multipositive complexes are obtainable by incorporating terminal cations, for example, protonated aminoalkyl groups. These types of complexes may bind to endogenous anions, anionic proteins, cell membranes, and the like. When two terminal anionic groups are attached, for example, carboxylates, phenolates, phosphorus, melted, sulfonates, and the like, the total charge of the complex can be . Another possibility occurs when three or more terminal anionic groups are attached; then the result is an anionic complex. End groups can be designed to axially chelate and formally. displaced axial anions, or they may be. designed specifically not to chelate but to retain charge.

Substituents on the complex of the invention, i.e. R groups other than hydrogen, are those that result in complexes with improved stability, controlled lipophilicity, increased hydrogen bridge formation, and greater macrocyclic ligand rigidity.

When it comes to macrocycle rigidity, groups that increase rigidity ultimately improve stability and increase relaxation inside and outside the sphere. Examples of groups that. macrocycle enhancers include, but are not limited to: cycloalkyl groups such as trans-cyclohexaho, and multiple alkyl groups such as pentamethyl.

With regard to hydrogen bonds, the groups that strengthen them result in an increase in the water retention time of the metal complex by:. provide alternative binding sites. Examples of hydrogen bonding groups are, but are not limited to, hydroxyalkyl, for example hydroxymethyl.

The lipophilicity of the complexes, i.e., the biodistribution of the complexes of the invention, can be controlled by positioning different numbers and types of ... substituents, e.g. In this way, the complexes of the invention can be targeted to certain tissues or organs in the body by determining the type and number of substituents.

The kinetic stability of the metal complex is important because complexes that are not sufficiently kinetically stable dissociate and release the free metal into the body. The kinetic stability, k _{a ± s} (M 2 sec ¹ ), can be controlled by positioning different numbers and types of substituents other than hydrogen. The complexes of the present invention have a pc of 1400 M ^ sec ^-1 , ie. the complexes of the invention are at least twice as stable as compared to complexes in which all R ¹ are hydrogen. Na16 more type plated. the number of substituents may be chosen such that the complexes are at least 1000 times kinetically more stable than those in which all R 'are hydrogen. Examples of groups that improve kinetic stability include, but are not limited to. cycloalkyl groups, e.g. trans-cyclohexano, and multiple alkyl groups, e.g. methyl. The oxidation stability of metal complexes is a particular problem for manganese complexes and is important because the complexes are not. sufficiently oxidatively stable will switch from divalent to trivalent manganese. Since manganese (III) complexes are colored, it is necessary to keep the complexes in a divalent form. manganese, - to create an effective contrast agent. By choosing type a. the amount of substituents can control the oxidation stability, ^E ₂ (v) ·. In general, it is desirable to select a type and number of substituents such that E _{1 / z} is greater than probably 0.7v. .....

- The number of R groups attached to the carbon atoms., I.e. R - R _g and R '^ - R ^ ·, which are other than hydrogen is preferably at least three, and preferably alaspoň fifth,

One group of currently preferred compounds are those wherein: at least one of either R ₁ or R ¹ and R or R 1, R 2. or R '. and R or R ', R or R ¹

2 2 3 3 4 4 SS and R or R ', R or R' and R or R 'and R or R' β € 7 7 8 β 9 9 and R or R 'together with the carbon atoms to which they are attached, form a saturated ring having 5 to 8 carbon atoms; and all of the remaining R groups are independently selected from each other: hydrogen, alkyl, or alkyl substituted with -OR 7 or -NR R, wherein R and R are independently hydrogen or alkyl. Po1516 15 16 ^JJ chat saturated cycles can vary from one to 5, but preferably at least two, and preferably saturated cycles have a ring of 6 carbon atoms, i.e., A cyclohexane group. An example of such a merger is shown in the following formula:

Another group in at least two groups

R *

R 'of the presently preferred compounds are those wherein of R, R', R, R ', R, R', R, R ', R', R, R ', R, R', R and R 'are alkyl. '/' Θ a 7 9 9 ^{2 J} or alkyl substituted by -OR or -NR R wherein R and R ^JJ J. S 15 16 '1' S 1 FC are independently hydrogen or alkyl. A suitable number of R groups which are alkyl or substituted alkyl is preferably at least 3, and more preferably at least 5. An example of such a compound is shown in the following formula:

HIM

A first embodiment of the invention relates to the aforementioned metal complexes in which at least one group of R ₁₀ ,

R, R, and R are other than hydrogen.

A second embodiment of the invention relates to a magnetic resonance imaging method, which is to be understood

a) administering to the human or non-human animal subject a contrast agent comprising a physiologically compatible complex of the invention and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle; and

b) producing a magnetic resonance image of at least a portion of the human or non-human animal subject.

A third embodiment of the invention relates to a method of diagnostic imaging comprising

(a) administering to the human or non-human animal subject a contrast agent comprising a physiologically compatible complex of the invention and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle, and

(b) an X-ray, sonographic or scintigraphic image of at least a portion of the human or non-human animal subject is produced.

A fourth embodiment relates to a method of radiotherapy carried out on a human or non-human animal subject, comprising administering to a human or non-human animal subject a radioactive substance comprising a physiologically compatible complex of the invention wherein M is a radioactive metal and a nontoxic, pharmaceutically acceptable carrier, adjuvant or vehicle.

The macrocyclic ligand used herein for comparison with the complexes of the invention wherein all R 'are H can be prepared according to general synthetic scheme A below, using methods known in the art for preparing certain intermediates and certain ligands. See, for example, Richman et al., J.,. Am. Chem. Soc., 96, 2268 (1974); Atkins et al., Org.

Synth., 58, 86 (1978); Thus, the triazaalkane is tosylated in a suitable solvent system to give the corresponding tris (N-tosyl) derivative. The corresponding base is then added to form the corresponding disulfonamide anion The diasulfonamide anion is then reacted with di-O-tosylated di-N-tosyl-diazaalkanediol to form the corresponding petatosylpentaazacycloalkane. The tosyl groups are then removed and the resulting compound is reacted with a divalent manganese compound substantially free of water and oxygen to form the corresponding divalent manganese complex with pentaazacyclloalkane. .

Macrocyclic ligands useful in the complexes of the present invention wherein R, R ', R', R ', R', R, R 'R and R''. they may be H or any of the functions as described above. to be; l prepared according to the general peptide ^method: shown in

Scheme B, which will be presented below. Process for preparing cyclic peptide precursors from the corresponding linear peptides. they are the same or are significant modifications of methods known in the art. See, for example, Veber, D.F. et al., J. Org. chem.,

44, 3101 (1979). The general method outlined in Scheme B below is an example using the sequential preparation of a functionalized linear polypeptide from the N-terminus to the C-terminus in the solution phase. Alternatively, the reaction sequence to prepare the linear pentapeptide may be performed as a solid state preparation using methods known in the art. The sequence of reactions may be conducted from the C-terminus to the N-terminus. and using convergent approaches, for example, pairing of di-α and tripeptides as desired. So 9

The Boc-protected amino acid reacts. with an amino acid ester using standard peptide coupling reagents.

The new Boc-dipeptide ester is then saponified to the free acid, which condenses again with another amino acid ester. The resulting Boc-tri-peptide ester is saponified again, and this method continues until the free acid is prepared with the Boc-protected pentapeptide. The Boc protecting group is removed under standard conditions and the resulting pentapeptide or salt thereof is converted to the cyclic pentapeptide. _; the pentapeptide is then reduced to pentaazacyclopentadecane using lithium aluminum hydride or borane. The final ligand is then reacted with a divalent manganese compound under substantially anaerobic conditions. thereby forming a corresponding divalent manganese complex with pentaazacyclopentadecane.

Scheme B was used to synthesize the complexes in Examples 3 and 5.

The R groups of macrocycles produced by the cyclic peptide pathway, i. R _x, R, R _and R _'3, R _s, R _s, R _7, R \, _R and _R' may be derived from D or L forms of the following amino acids: alanine, aspartic acid, arginine , asparagine, cysteine, glycine, glutamic acid, glutamine, histidine, isoleucine, leucine, lysine, methionine, proline; phenylalanine, serine, trypropane, threonine, tyrosine, valine and optionally R groups of unnatural α-amino acids, for example alkyl, ethyl, butyl, tert. butyl, cycloalkyl, phenyl, alkenyl, allyl, alkynyl, aryl, heteroaryl, polycycloalkyl, polycycloaryl, polycycloheteroaryl, imines, aminoalkyl, hydroxyalkyl, hydroxyl, phenol, amine oxides, thioalkyl, carboalkoxyalkyl, carboxylic acids and derivatives thereof, keto, ether, aldehyde, amine, nitrile, halogen, thiol, sulfoxide, sulfone, sulfonic acid, sulfide, disulfide, phosphonic acid, phosphinic acid, phosphine oxides, sulfonamides, amides, amino acids, peptides, proteins, carbohydrates, nucleic acids, fatty acids, lipids, nitro, hydroxylamines , hydroxamic acids, thiocarbonyl, borates, boranes, borase, silvl, siloxy, silaza and combinations thereof,

The macrocyclic ligands useful in the complexes of the invention may also be prepared via the formation of the dicarboxylic acid dichloride shown in Scheme C below. In this case, the triazaalkane is tosylated in a suitable solvent system to form the corresponding tris (N-tosyl derivative. Such a derivative is compounded with a suitable base to form the corresponding disulfonamide anion. The disulfonamide anion is dialkylated with a suitable electrophile to form a dicarboxylic acid derivative. A dicarboxylic acid derivative is prepared from this dicarboxylic acid derivative which is combined with a suitable substance to form a dicarboxylic acid dichloride The desired vicinal diamine is obtained by any one of several routes One useful route is the preparation of the aldehyde by reaction with cyanide in the presence of ammonium chloride, followed by treatment with an acid to form alpha ammonium nitrile, which is reduced in the presence of an acid and then treated with a suitable base to form the vicinal diamine Condensation of the dicarboxylic acid dichloride with the vicinal diamine present. Suitable bases form a tris (tosyl) diamide macrocycle The tosyl groups are removed and the amides are reduced and the resulting compound is reacted with a divalent manganese compound under strictly anhydrous and anaerobic conditions to form the corresponding substituted divalent manganese pentaazacycloalkane complex.

Scheme C was used to synthesize the complexes in the example.

Vicinal diamines were prepared by the aforementioned route (known as Strecker synthesis) and vicinal diamines were purchased when commercially available. Any method for preparing vicinal diamines can be used.

The macrocyclic ligands useful in the complexes of the invention can also be prepared by the bis (haloacetamide route shown in Scheme D below). Thus, the triazaalkane is tosylated in a suitable solvent system to give the corresponding tris (N-tosyl) derivative. A bis (haloacetamide), for example bis (chloroacetamide) vicinal diamine, is prepared by reacting the diamine with an excess of a haloacetyl halide such as chloroacetyl in the presence of a base, and the disulfonamide anion tris (N-tosyl) triazaalkane reacts with bis (chloroacetamide) diamine. Tosyl groups are removed and amides are reduced, and the resulting compound is reacted with a divalent manganese compound under strictly anhydrous and anaerobic conditions to form the corresponding substituted pentaazacyclic complex. of a divalent manganese.

Scheme D is an alternative route for the synthesis of the complex of Example 2.

The macrocyclic ligands useful in the complexes of the invention wherein R, R ', R, R' are part of a cis- or transcykloalkylového ring system, and R, R ', R, R' ₇ and R @ _g, R @ ¹ may be hydrogen or any functional group so far described, can be prepared according to the pseudopepouid method shown in Scheme E below. Cis-1,2-diaminocycloalkane or trans- (R, R) -1,2-diaminocycloalkane or trans (S, S) -1,2-diaminocycloalkane can be used in combination with any amino acid in this method. the stereochemistry of the linked cycloalkane ring and the substituents R, R ', R, R' and R, R ', and the other functional functions and stereochemistry were defined in any way. For example, trans- (R, R) -1,2-diamino-cyclohexane was monotosylated; subjected to reaction with Boc anhydride to provide differentiated N-Boc, N-tosyl derivatives Sulfonamide was alkylated with methyl bromoacetate using sodium hydride as the base and saponified to the free acid N-tosylglycine-containing cyclohexančiamine serves as a dipeptide substitution in standard peptide synthesis in solution phase Coupling with a functionalized amino acid ester thus provides the corresponding pseudo-tripeptide, and two consecutive TFA cleavage will yield a pseudopentapeptide that may have N- and C end groups deprotected in one step, using HCl / AcOH. DPPA-mediated cyclization followed by LiAlH 4 reduction or borane provides the appropriate macrocyclic ligand. This ligand system reacts with a manganese (TT) containing compound, such as, for example, manganese chloride under strict anaerobic conditions, to form a corresponding functional-functionalized manganese pentaazacyclloalkane complex. The macrocyclic ligands useful in the complexes of this invention where _R, R, R. ,, R, and _R, R, R, R _g are part of the ring system * cis or trans cycloalkyl and R9, R 9 can be H or any The previously described function may be prepared according to the iterative pseudo-peptide method shown in Scheme F below. Cis-1,2-diaminocycloalkane or trans- (R, R) -1,2-diaminocycloalkane or trans- (S, S) -1,2-diaminocycloalkane can be used in any combination using this method and in combination with any amino acid. This makes it possible to define any method of stereochemistry to each other both of the cycloalkane-linked rings and of the substituents R, R, R, RR, R ' as well as the occupancy and stereochemistry. Thus, the dipeptide substitute for (S, S) -1,2-diaminocyclohexyl-N-tosylglycine prepared from (S, S) -1,2-diaminocyclohexyne exactly as in Scheme E for (R, R) -1,2-diaminocyclohexane , may be coupled to a functionalized amino acid ester to form the corresponding pseudotripeptide. Cleavage of TFA provides a pseudotripeptide salt of TFA that is paired with (R, R) -diaminocyclohexyl-N-tosylglycine. The saponification and cleavage of TFA provides a pseudo-pentapeptide containing a bis-cyclohexane group. DPPA mediated cyclization of the corresponding bis-cyclohexano-linked macrocyclic ligand. This ligand system reacts with a divalent manganese compound, for example manganese chloride, under stringent anaerobic conditions to form the corresponding functionalized pentaazacyclloalkane complex of divalent manganese.

Scheme A

KjX N HHi H

X 0 «

TjCI, pyrWln «V /» Λ

TiHH M HKTl

It

TiCVEljH

CH 3 Cl 2

t HiS0 _fc ltt * C t HiOM

Scheme Β

BocNH

EDC-HCl, HC6T. DMF. TEA, AT or ethyl chloroformate, DMF. TEA, ^0Q C +

NaOH, H 2 O CH 3 OH

BocNH

EDC-HCl. ΗΟΘΤ. DMF. TEA. RT or ethyl chloroformate DMF, TEA, 0 ° C

Scheme B (continued)

NaO «. H ₃ O CH ₃ OH

BccNH

/ UAIH.

THF.A or

BH ₃ . THF

Scheme C

R,

Scheme D

and

Scheme E

'NHj

NHTs (Boc) ₂ 0

O ^ NHTs' NHSoc

CCH-,

NaH, DMF

Ts O

NHSoc

OH ^{Λ; ΗΝ} > Λ ^

R _s

EDO

->

EDO

Scheme E (continued)

*

MnCÉ

MaOH

Scheme F i!

Ts O

Z \ y

NHBoc

F

F

OH

Ts O Rj m _n l Ru OEi

KOH

Ts O Rj R'9

Cl λ ° ^h

TFA

-N NHBoc

O

TS O Rg Rg ^R 1 OH OH

N, N-N ⁵ n ₂ -tfa

Diagram F (continued)

Macrocyclic ligands useful in the preparation of the bridged complexes of the invention may be prepared according to the schemes in the following examples.

—I / 1 Λ 1 rt -3 * ι f f r r r r r r r r r,,,,,- r.r * ·, Λ k · »*. Λ * Λ 1, · "· I I-ry-ι -Ί riCXAJ. Uxy Aj. .x. yj ur X vivuuxOAO ^ ii y. v y ut;

wherein R or R ', and R _g or R' _q are connected to each other via a bridged or second ring system in a bicyclic sense, wherein S- _L ", R ₁₃ and may be H or any of the functional groups described so far and R" defined as R, R and R, can be prepared according to the method described in Scheme G below. The TFA salt of the trans - (R, R) -1,2-diaminocyclohexyl-N-tosylglycine dipeptide replacement is paired with Z-Glu (OtBu). -OH using EDC in DMF. This tripeptide is then saponified and coupled to Orn (Boc) -OMe HCl using EDC in DMF. The resulting tetrapeptide is then deprotected by hydrogenolysis of the Z group and coupled to Z-Gly again using EDC in DMF. By saponification and hydrogenolysis, a deprotected pseudopentapeptide is formed which is cyclized with DPPA. The side chains are then deprotected in one step using TFA and the other bridging cyclization and again performed using DPPA as the condensing agent. Reduction with lithium hydride provides a bicyclic ligand system that reacts with manganese chloride under strict anaerobic conditions to form the corresponding functionalized divalent manganese hexaazabicycloalkane complex.

Scheme G

Diagram G (continued)

NR, 13cc

O

OtSu

O

Diagram G (continued)

The macrocyclic ligands useful in the complexes of the invention wherein the nitrogen substituents R and R or R 'are linked to each other via a bridging or second ring system and R _a3 , R 6 and R ₁₇ may be H or any functional group so far described may be prepared. according to the method described below in Scheme H. The TFA salt of the trans- (R, R) -1,2-diaminocyclohexyl-N-tosylglycine dipeptide replacement is paired with Z-Glu (OtBu) -OH using EDC in DMF. saponified and coupled with N- [3- (Boc-amino) propyl] -Gly-OMe HCl using EDC in DMF The resulting tetrapeptide is then deprived of N-protection by hydrogenolysis of the Z group and paired with Z-Gly again using EDC in DMF By saponification and hydrogenolysis, a pseudopentapeptide-deprotected protease is formed which is cyclized with DPPA, and the side chains are deprotected in one step using TFA and the other bypass cyclization, and again performed using DPPA as the condensation agent. Reduction of lithium aluminum hydride provides a bicyclic ligand system that reacts with manganese (II) chloride under strict anaerobic conditions to form the corresponding functionalized divalent manganese hexaazabicycloalkane complex.

Scheme Η

CMe

EDC. You

NR15B0C

NaOH

Ts O \

.N

NR _l5 3cc

I η I with O .N 'i

OH - NH

NR ₁₄ Z eoc

OWe * NH nr. ₄ :

O ^^ OtSu

Hj, NR € NR, s3oc, NR ₁₅ 3cc

About me

EDC

Ts O

K ^ AJ

X ^ N

NH

O

OtBu ^Ri * ⁵ ί Υ ΥΥ

Y

O OlBu

Diagram H (continued)

1. NaOH

2. Hj. POO

NHR, _S- TFA

TFA

NR ₁₅ 3oc

Diagram H (continued)

LiAlHi or BH '!

The macrocyclic ligands useful in the complexes of the invention where the nitrogen substituents R, and R are connected together through bridging 'or second ring system, and _R, R ^1, R _7, R \, R, _, R' ₉ and R 'may be H or any functional group described so far may be prepared according to the method described below in Scheme I. The cyclohexyl-containing pentaazamacrocyclic ligand prepared according to Scheme E is reacted with ditosylsulfonamide or diethanolamine prepared from diethanoamine and tosyl chloride. This over-phosphating cyclization can be controlled using Cs 2 CO as a base in a DMF solvent. This bicyclic ligand system then reacts with manganese chloride under strict anaerobic conditions to form the corresponding functionalized divalent manganese hexaazabicycloalkane complex.

Diagram

MnC! ₂ . CH ₃ OH

The pentaazamacrocycles of the invention may have one or more asymmetric carbon atoms and are therefore capable of existing in the form of optical isomers as well as in the form of their racemic and non-racemic mixtures. Optical isomers may be obtained by resolution of racemic mixtures by conventional means, for example by formation of diastereoisomeric salts by treatment with an optically active acid. Examples of the appropriate acids are tartaric, diacetyltartaric, dibenzoyltartaric, detroluyltartaric and camphorsulfonic acid and then separation of the mixture of diastereoisomers by crystallization followed by liberation of the optically active bases from these salts. Another method for separating optical isomers involves using a chiral chromatography column optimally selected to maximize separation of enantiomers. Yet another available method involves the synthesis of covalent diastereoisomeric molecules by reacting one or more secondary amino groups (s) of the invention with an optically pure acid in activated form or an optically pure isocyanate. The diastereoisomers formed can be separated by conventional methods, for example chromatography, distillation, crystallization or sublimation, and then hydrolyzed to obtain an enantiomerically pure ligand. The optically active compounds of the invention may also be obtained using optically active starting materials, for example natural amino acids.

The methods of diagnostic analysis of the invention include administering to the human or non-human animal subject or host, the contrast enhancing compounds of the invention, in an amount sufficient to produce the desired contrast (or displacement) and then subjecting the host to diagnostic analysis. by NMR; including especially preferred NMR imaging analysis (MRI). Further, the complexes of the invention are useful in diagnostic X-ray imaging analysis, ultrasound analysis, or scintigraphic analysis. Although the complexes of the present invention are primarily described as contrast enhancers, they may also act as NMR shift agents and such use in accordance with the methods disclosed herein.

The complexes of the present invention used as contrast agents are administered in an amount sufficient to produce the desired effect. 1 H-NMR is that amount which produces an NMR signal, i.e., the amount of a given complex that changes the relaxation times of the NMR. the spin-lattice, spin-spin or spin-echo signals or for the shift agent selectively shifts the spectral positions of the resonant core relative to other similar cores. This change is carried out in a manner following the increase in signal received from the analyzed subjects either by reducing the above relaxation times or by increasing them to the host area. or the host itself to which the complex has been administered. In another embodiment, the amount of complex affecting the NMR signal is an amount that, in conjunction with changing the relaxation time of the patient's NMR signals, also alters these relaxation times sufficiently to achieve a higher resolution or contrast between the parts of the patient into or without .

The relaxation time Τ _χ (called spin-lattice) measures the speed at which the magnetic energy is transferred from the resonant nuclei. 1 to all other energy degrees of freedom, except for other resonant cores. The relaxation time ± spin (spin) measures the rate of magnetization transfer to other resonant nuclei.

Another measurable parameter is the proton density in the medium.

In the first consideration it represents the amount of free water contained in the sample.

The NMR image represents the distribution of these parameters δ

T, T 'and combinations thereof. The contrast between a given tissue and the surrounding area

ΐ these tissues increase as a function of tissues containing more or ί

less water or mobile protons and different relaxation j

times. It is also possible to modify the contrast by changing one or both

Parametrů multiple parameters (experimental spin responses help ’1 \ function, or magnetization back-up allowing local measurement

TJ.

: Experience has shown that it is advisable to modify the relaxation t

; time to improve the contrast of the acquired image, for example with the contrast agents mentioned herein. Proton density!

• (in the practice of water or lipid protons) it differs slightly between i

individual organs, and often less between normal and pathological tissues. However, relaxation characteristics depend on; a number of factors (microscopic dynamics of molecules, chemical exchange, paramagnetic disturbances, etc.) that are

Even more variable.

ii Detailed discussion of NMR and theoretical assertions at ¹ taking appropriate parameters for diagnostic analysis, e.g. MRI, for example, is disclosed in U.S. Patent No. 4,749,560 which is incorporated herein by reference. listed here in the literature. X-ray imaging diagnosis, ultrasound diagnostics, scintigraphic imaging

The lysis and radiotherapy using the complexes of the invention are all performed in accordance with well established techniques known to those skilled in the art.

In addition, the method of diagnostic analysis of the invention makes it possible to achieve tissue or organ specific diagnostics. For example, contrast agents may exhibit organ or tissue specificity, for example, biodifferential distribution / / for example. in myocardial tissue when the complexes of the invention are lipophilic in nature.

*. ^b ~ ~ - '. ,. The complexes of the invention may be administered to the host as a pharmaceutical enhancing amount in a pharmaceutical composition. The pharmaceutical compositions comprise a contrast enhancing dose of the contrast agents of the invention together with a non-toxic pharmaceutically acceptable carrier, adjuvant or vehicle. The compositions may be administered by well known routes including oral, intravenous (if the mixture is soluble), intramuscular, intra-oral. nasal, intradermal, subcutaneous, parenteral, enteral, and the like. Depending on the route of administration, the pharmaceutical composition may require protective coatings.

Pharmaceutical forms suitable for injectable use include sterile aqueous solutions or dispersions and sterile powders for the extemporaneous preparation of sterile injectable solutions or dispersions. In all cases, the final form of the solution must be sterile and fluid. Typical carriers include a solvent or dispersion medium containing, for example, water, buffered aqueous solutions (i.e., biocompatible buffers), ethanol, polyol (glycerol, propylene glycol, polyethylene glycol, and the like), suitable mixtures thereof, surfactants and vegetable oils. Sterilization may be accomplished by any technique known in the art, including, but not limited to: addition of antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol, sorbic acid, thiomersal, and the like. Further, isotonic agents such as sugars or sodium chloride may be included in the composition.

The production of sterile injectable solutions comprising the subject contrast agent is accomplished by incorporating these in the desired amount into the appropriate solvent, as appropriate, with the various ingredients listed above, followed by sterilization, preferably filter sterilization. To obtain a sterile powder, the above solutions are vacuum dried or lyophilized as necessary.

Solid dosage forms for oral administration may include capsules, tablets, pills, powders, granules and gels. In such solid dosage forms, the active compound may be admixed with at least one inert diluent, for example, sucrose, lactose or starch. Such dosage forms may also include, as is conventional practice, other ingredients other than an inert diluent, for example, lubricating agents, for example magnesium stearate. In the case of capsules, tablets and pills, the dosage forms may also comprise buffering agents. Tablets and pills may additionally be prepared with an enteric coating.

Liquid dosage forms for oral administration may include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs containing an inert diluent commonly used in the art, for example water. Such compositions may also include adjuvants such as wetting agents, emulsifying and suspending agents, and sweetening, flavoring, and perfuming agents.

The contrast agents of the invention are thus formulated for proper and effective administration in pharmaceutically effective amounts with a pharmaceutically acceptable carrier, adjuvant or vehicle at a dose which results in enhanced contrast. These amounts are preferably about 1 micromole per 1 mole of contrast agent per liter, eventually administering doses of about 0.001 to 5 mmol / kg body weight. Preferred compositions provide an effective dosage of contrast agents in the range of about 0.001-5 mmol / kg for NMR diagnostics, preferably between about 0.005-0.5 mmol / kg; in the range of probably 0.1-5 mmol / kg for X-ray diagnostics; and in the range of probably 0.1-5 mmol / kg for ultrasound diagnostics. For scintigraphic diagnostics, the dosage of contrast medium should generally be lower than for NMR diagnostics, ie. MRI. Conventional doses known to those skilled in the art may be used for radiotherapy.

As used herein, the term pharmaceutically acceptable carrier, adjuvant or vehicle means any of all solvents, dispersants, coatings, antibacterial and antifungal agents, isotonic agents and the like. The use of such media and substances is well known in the art.

Exemplary equivalents of the formulas above for compounds and derivatives as well as intermediates are compounds that are otherwise equivalent and having the same general properties, for example, tautomers of compounds and those wherein one or more different R groups is a simple variation of substituents as well as

i i

i í

are defined herein, for example, wherein R is a higher alkyl group than indicated, or wherein tosyl groups are other nitrogen or oxygen protecting groups, or wherein O-tosyl is a halide. Anions having a charge different from 1, for example carbonate, phosphate and dibasic phosphate, may be used in place of charged anions even if they do not have an adverse effect on the overall activity of the complex. However, the use of anions having a charge different from 1 results in a minor modification of the general formula above. In addition, where a substituent is determined or may be hydrogen, the exact chemical nature of a substituent other than hydrogen at that position, for example, a hydrocarbyl group or halogen, hydroxy, amino and the like, is not critical as long as it does not adversely affect overall activity or process. synthesis.

The chemical reactions described above are generally described in terms of their broadest application to prepare the compounds of the invention. In some cases, the reaction may not be applicable as described for each candidate substance. The compounds for which this is applicable can be readily recognized by those skilled in the art. In all such cases, either the reactions may be carried out by conventional modifications known to those skilled in the art, for example by appropriately protecting interfering groups, altering alternative conventional reagents, routinely modifying the reaction conditions, and the like, or other compounds mentioned herein may be used to prepare the corresponding compounds. otherwise customary reactions otherwise; usually. In all methods of preparation, all starting materials are known or readily obtainable from known starting materials Without further elaboration, it is believed that one skilled in the art can use the same. . previous description to use this. invention in its entirety. Another preferred specific embodiment is. therefore, it is to be construed as merely illustrative and in no way restrict the rest of the possible cases.

DETAILED DESCRIPTION OF THE INVENTION

All reagents were used as obtained without purification unless otherwise indicated. All NMR spectra were obtained on nuclear magnetic resonance. the Varian VXR-300 or VXR ₇ 400 spectrometer. Qualitative and quantitative mass spectroscopy was performed on a Finnigan MAT90, Finnigan 45.00, and VG40-250T per instrument. using m-nitrobenzyl alcohol (NBA) or m-nitrobenzyl alcohol / LiCl (NBA + Li). Melting points (bt) are unadjusted. . '

The following abbreviations, which apply, to amino acids and their protective. are in accordance with the recommendations of the IUPAC - IUB Commission on Biochemical Nomenclature (Biochemistry, ii, 1726 (1972)) and with normal use. .

Ala . L-alanine. DAla D-alanine Gly glycine Ser L-serine DSer D-serine ppg propargylglycine Tyr L-tyrosine

Bzl benzyl Boc tert-butoxycarbonyl Et ethyl TFA trifluoroacetate DMF dimethylformamide HOBT H 0 1-hydroxy- (1 H) -benzotriazole monohydrate EDC HCl 1- (3-Dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride TEA triethylamine DMSO dimethylsulfoxide THF tetrahydrofuran DPPA diphenylphosphoryl azide DMPU dimethylpropyleneurea C concentration, g / cm ³ DME 1,2-dimethoxyethane

The abbreviation Cvc stands for 1,2-cyclohexanodiamine (stereochemistry, ie R, R or S, S is given as such). This allows the use of three letter letter peptide nomenclature for pseudopeptides containing a 1,2-cyclohexanodiamine residue.

Example 1 (comparative)

And, Synthesis 1.4. Of 7-tris (o-toluenesulfonyl) -1,4,7-triazaheotane This the compound was synthesized according to the procedure of At- kins, T.J. .; Richman, J.E .; and Oetle W. F .; Org. Synth., 58,

86-98 (1978). To a stirred solution of p-toluenesulfonyl chloride (618 g, 3.24 mol) in pyridine (1500 mL) at 0 ° C was added 1,4,7-triazaheptane (95.5 g, 0.926 mol) in pyridine (150 mL). in a dry argon atmosphere, maintaining a temperature of 50 ° C. This addition required 30 minutes. After the mixture was allowed to cool slowly to room temperature with stirring for an hour, H 2 O (2 L) was slowly added to the cooled (ice bath) mixture. The heavy white precipitate that formed was filtered off and washed thoroughly with H 2 O. The light yellow solid was dissolved in DMP (3 L) and 0.1 N HCl (4 L) was slowly added at ° C. The slurry precipitate was filtered and the pale žlutná solid washed thoroughly with H? O and dried in vacuo to yield 486 g (93% yield) of product: mp 180-1 ° C ^{Χ Η} NMR _(DMSO-d) δ 2 39 (s, 3H), 2.40 (s, 6H), 2.84 (m, 4H), 3.04 (t, J = 6.9Hz,

H), 7.40 (d, J = 8.1 Hz, 4 H) 7.59 (d, J = 8.3 Hz, 2H),

7.67 (m, 6H).

B. Synthesis of 1,4,7-tris (p-toluenesulfonyl) -1,4,7-triazahgptan-i

-7-disodium salts

This compound was synthesized according to the procedure of Atkins, T. J .; Richman, J.E .; and Oetle W. F .; Org. Synth., 58, 86-98 (1978), to a mechanically stirred slurry

The 1,4,7-tris (p-toluenesulfonyl) -1,4,7-triazaheptane prepared as in Example 1A (486 g, 0.859 mol) in ethanol (1150 mL), heated to reflux in a dry argon atmosphere, was as near as possible a solution of sodium ethoxide (prepared by dissolving sodium metal (39.5 g, 1.72 mol) in absolute ethanol (1.0 L)) was added faster. The clear brown solution that rapidly formed was cooled to room temperature and ethyl ether (1.0 L) was added. The crystals were filtered under a dry argon overlay, washed with a mixture of ethanol and ethyl acetate 3: 1 and ethyl ether. Then the crystals were dried in vacuo to yield 509 g (97% yield) of product as a white powder: ^X H NMR (DMSO-d 6 2.30 (s, 6 Η) 2, '36 (s, 3 .H) 2.63 (t, J = 8.7 Hz, 4 H), 2.89 (t, J = 7.2 Hz, 4 H), 7.11 (d, J = 8.1 Hz, 4 H) 7.28 (d, J = 8.0 Hz ', 2 Η), 7.46 (m, 6H) C ^ Synthesis of 3,6-bis (p-toluenesulfonyl) -3,6-diazaoctane-1,8- dip-toluenesulfonate

To a stirred solution of p-toluenesulfonyl chloride (566 g, 2.97 mol) and triethylamine (300 g, 2.97 mol) in CH 2 Cl 2 (2.0 L) at 0 ° C under a dry argon atmosphere was added 3.times. 6-diazaoctane-1,8-diol (100 g 0.675 mol) in portions, maintaining a temperature of &lt; 10 ° C. The addition took 30 minutes. The mixture was allowed to warm to room temperature with stirring for an additional 18 h and then poured onto ice (1000 g). The CH 2 Cl 2 layer was from. separated, washed with 10% HCl, H 2 O and saturated NaCl solution and dried (MgSO 4). Roztok'byl -.zkoncentrován in vacuo to a volume of - 1.5 1st Crystallization by addition of hexane (4 L) gave 477 ^_r! · G- (92% yield) of the product as colorless needles: bt

151-3 ° C ^X H-NMR (CDCl 5 : 2.43 (S, 12 Η), 3.29 (s, 4 Η), 3.36 (t, J = 5.2 Hz, 4 ohms), 4.14 (t, J = 5.2 Hz, 4 Η), 7.33 4 (d, J = 7.8 Hz, 8H) 7.71 (d, J = 8.2) Hz, 4 H), 7.79 (d,

J = 8.3 Hz, 4 H).

D, Synthesis of 1,4.7.10.13-penta (p-toluenesulfones 11-1.4.7,10,13pentaazacyclopentadecane

This compound was synthesized according to the procedure. Richman, J.E .; and Atkins, TJ; J. Am. Chem. Soc., 96, 2268-70 (1974). To a stirred solution of 1,4,7-tris (p-toluenesulfonyl) -1,4,7-triazaheptane-1,7-disodium salt prepared as in Example 1B (146 g, 0.240 mol) in anhydrous DMF (2250 mL) was added dropwise. to a solution of 3,6-bis (p-toluenesulfonyl) -3,6-diazaoctane-1,8-di-p-toluenesulfonate prepared as in Example 1C (184 g, 0.240 mol) in anhydrous DMF (1020 mL) over 3 hours ) in an atmosphere of dry argon, maintaining a temperature of 100 ° C. After stirring for an additional hour at 100 ° C, the solution was concentrated in vacuo to a volume of 1.5 L. At 80 ° C, H. was slowly added _; 0 (500 mL) to crystallize the product. The resulting slurry was slowly cooled to 0 ° C and additional water (1250 mL) was added. The solid was filtered, washed well with EtOAc and then with 90% ethanol and dried in vacuo. The off-white solid was dissolved in CH 2 Cl 2, the insoluble impurities were removed by filtration, and the filtrate was washed with H 2 O and then dried (MgSO 4). The solvent was removed in vacuo to give a yellow solid which was purified by recrystallization from CH 2 Cl 2 -hexane to give 164 g (63% yield) of the product as a white crystalline solid: mp 290-3 ° C, ^X 1 H NMR (CDCl 3) δ 2.44 (s, 15H), 3.27 (s, 20H), 7.32 (d, J = 8.3Hz, 10H), 7.66 (d, J = 8.3 Hz, 10 H).

E, Synthesis of 1,4,7,10,13-pentaazacyclopentadecane

A mixture of 1,4,7,10,13-penta (p-toluenesulfonyl) -1,4,7,10,13pentaazacyclopentadecane prepared as in Example ID (168 g, 0.170 mol) and concentrated H 2 SO 4 (500 mL) was heated at 100 ° C with stirring and under a dry argon atmosphere for 70 hours. To the resulting dark brown solution was added dropwise ethanol (500 mL) with stirring at 0 ° C, followed by ethyl ether (3 L). The white solid was filtered and washed with ethyl ether. The solid was dissolved in H 2 O (500 mL) and the resulting solution washed with ethyl ether. After reducing the volume of the solution in vacuo to 200 mL, the pH was adjusted to 10-11 with 10 N NaOH and the solvent removed in vacuo. Ethanol (500 ml) was then added and removed to dryness in vacuo. The resulting tan oil was extracted with hot THF (2 x 500 mL) and filtered at room temperature. The filtrates were combined and the solvent removed in vacuo to give the crude product as a yellow crystalline solid, which was then redissolved in CH 2 Cl 2 and filtered to remove insoluble impurities. Recrystallization from cold (-20 ° C) CH 2 CN gave 11.3 g (31% yield) of the product as colorless needles; 108-9 ° C, NMR (CDCl 3 δ 1.74 (brS, 5H), 2.73 (s, 20H); Exact Mass (M + Li): Calcd, 222.2270;

222.2269 (CHN Li).

X O 2 S

F. Chloride [manganese (II) dichloro (1,4,7, IQ, 13-pentaazaclopentadecane) 1

A solution of 1,4,7,10,13-pentaazacyclopentadecane prepared as in Example 1E (2.0 g, 9.3 mmol) and anhydrous manganese chloride (1.2 g, 9.3 mmol) in anhydrous methanol (50 mL) ) was refluxed in a dry nitrogen atmosphere for 3 hours. The solution was filtered and the solvent removed in vacuo. The resulting solid was recrystallized from ethanol-ethyl ether to give 2.79 g (88% yield) of the product as an off-white crystalline solid: FAB mass spectrum (NBA) m / z (relative intensity) 340 (M +, 2), 305 / 307 ((M-Cl) * 100/45)); Anal. calculated for C C HH ^Cl ^MnN ^:: C,

35.17; Η, 7.38; Cl, 20.76; Ν 20.60. Found: C, 34.95;

,3 7.31; Cl, 20.49; Ν 20.22.

Example 2

A. Synthesis of dimethyl 3,6,9-trig (p-toluenesulfonyl) -3,6,9-triazaundecanedioate

1,4,7-Tris (p-toluenesulfonyl) -1,4,7-triazaheptane-1,7-disodium salt prepared as in Example 1B (30 g, 49.2 mmol) was dissolved in dry N, N-dimethylformamide (180 mL) under argon. After cooling to 0 ° C in an ice bath, methyl chloroacetate (15.40 g, 141.9 mmol) was added dropwise over a 10 minute period. The reaction mixture became cloudy at the end of the addition, and was allowed to stir overnight while the ice bath was warmed to room temperature. The solvent was evaporated under reduced pressure to give a brown oil, which was dissolved in ethyl acetate (450 mL) to give a cloudy solution. This solution was washed twice with water (500 mL, then 300 mL). The combined aqueous layers were back extracted with ethyl acetate (300 mL). The combined ethyl acetate layers were washed twice with saturated brine (200 mL), filtered and evaporated to dryness. This residue was dissolved in dichloromethane (200 mL), evaporated to dryness and placed on a vacuum line. Recrystallization from chloroform-methanol and washing with methanol and ether gave an off-white solid weighing 27.46 g. An additional amount of slightly darker solid (4.7 g) was obtained from the filtrate to remove the solvent and recrystallize as above. Total yield was 32.2 g (93% yield): mp 141-2 ° C ^Χ Η NMR (CDC1J δ 2.42 and 2.44 (2 s, 9 Η), 3.41 (br s, 8 Η ), 3.60 (s, 6 Η), 4.07 (s, 4 Η), 7.26 - 7.35 (m, 6 H), 7.63 - 7.74 (m, 6 H).

B, Synthesis of 3,6,9-tris (p-toluenesulfonyl) -3,6,9-triazaundecanedioic acid

The dimethyl 3,6,9-tris (p-toluenesulfonyl) -3,6,9-triaza-unecandioate prepared as in Example 2A (16 g, 22.5 mol) was slurried in tetrahydrofuran slurry (100 mL). Sodium hydroxide (2N, 160 mL) was added dropwise over 1 h. After 72 hours, the solvent was evaporated under reduced pressure and hydrochloric acid (1 N) was added to lower the pH to 4. This aqueous phase was extracted several times with ethyl acetate, combined ethyl acetate layers They were washed twice with brine, dried (MgSO, filtered and evaporated to give a white solid, 14.22 g (93% yield): mp 177-80 ° C ^X H NMR (DMSO-d) δ 2.38 and 2 , 40 (2 s, 9 Η), 3.10 (m, 4 Η), 3.73 (s, 4 Η), 7.37 and 7.41 (2d, J = 7.9, 8.2 Hz) ,

Δ), 7.61 and 7.66 (2d, J = 8.2, 8.0 Hz, 6 H).

C, Synthesis of 3,6,9-tris (p-toluenesulfonyl) -3,6,9-triazaundecanedioyl dichloride

3,6,9-Tris (p-toluenesulfonyl) -3,6,9-triazaundecanedioic acid prepared as in Example 2B (40.5 g, 59.4 mmol) was placed in a round-bottom flask under argon and added oxalyl chloride (400 g, 3.15 mol). This mixture, initially cloudy, cleared after a few hours and was stirred overnight at room temperature. At the end of this time, it was heated to 40 ° C for 30 min. Oxalyl chloride was removed on a rotary evaporator. To the resulting solid was added di 60 chloromethane (50-60 mL) to dissolve and then removed on a rotary evaporator. This procedure was repeated twice to give 40.5 g (99% yield) of a white solid: m.p.

136-7 ° C

3.38 (m, ^X H NMR (CDC1J 6 2.43 and 2.46 (2 s, 9 Η), 3.30 4 Η), 3.40 - 3.43 (m, 4 Η), 4, 56 (S, 4), 7.30

7.40 (m, 6H), 7.71 (d, J = 8.2 Hz, 6 H).

D. Synthesis of trans-5,6-cyclohexano-l, 10,13-tristep-toluenesulfonyl) -1,4,7,10,13-pentaazaoyclopentadecane-3,8-dione

Dry dichloromethane (150 mL) was placed in a 1 L, 4-neck round bottom flask under argon equipped with two separating funnels. The 3,6,9-Tris (p-toluenesulfonyl) -3,6,9-triazaundecanedioyl dichloride prepared as in Example 2C (5.07 g, 7.05 mmol) was dissolved in dry dichloromethane (150 mL) and added to one. from separating funnels. 3,6,9-Trans-1,2-diaminocyclohexane (0.805 g, 7.05 mmol) and triethylamine (1.96 mL, 14.1 mmol) were dissolved in dry dichloromethane (150 mL) and added to a second separatory funnel. . After cooling the flask containing dichloromethane in an ice bath to an internal temperature of 0 to 5 ° C, the contents of the separatory funnel were added simultaneously to the stirred mixture over 2.25 hours. A white precipitate appeared before the addition was complete. At the end of this time, the ice bath was removed and the reaction mixture was stirred at room temperature overnight. The reaction mixture was filtered and a white precipitate was identified as pure product. The filtrate was washed twice with water (100 mL), once with brine (100 mL), dried (MgSO 4), filtered, and the solvent was removed under reduced pressure. Recrystallization from dichloromethane-hexane gave another product, together

with the original clot ligament 2.85 254 -4 ° C, ^{Γ Η} NMR (DMSO-d) that δ i, (m, 4 H), 2.42 and 2.43 (2s, 9 H), J = 16.5 Hz, 4H), 4.01 (d, J = (m, 8 H), 7.71 (d, J = 8.3) Hz, Hz, 2 H).

g ( 53 % yield) b. t. 15 (br with, 4 H), 1.52 1.75 3.04 (m, 8 H), 3.51 (d + m, 16.5 Hz, 2 H), 7.35 - 7.53 4 H), 7.80 (br d, J = 10.5

E. Synthesis of trans-1,2-bis (chloroacetamido) cyclohexane A three-necked three-necked flask equipped with a magnetic stirrer and two 1-liter separating funnels was charged with 1,2-diaminocyclohexane (35.0 g, 0.310 mol) dissolved in chloroform (375 mL) and water (185 mL). Both separatory funnels were each filled separately with chloroacetyl chloride (75 mL, 0.94 mol) in chloroform (440 mL) and potassium carbonate (120.5 g, 0.87 mL) in water (4 L) was added in four portions during the addition. . The flask was cooled in an ice salt bath and reagents were added for two hours. The ice salt bath was removed, water (600 mL) was added and the reaction mixture was stirred for 2.5 hours. The mixture was separated and the aqueous layer extracted several times with chloroform. The combined chloroform layers were washed with water and then brine. The organic layer was dried (sodium sulfate) and concentrated in vacuo to give an off-white solid. The solid was washed with ether to give 55.32 g (67% yield) of a white solid after drying under vacuum: mp 202-3 ° C, ¹ H NMR (CDCl 3 δ 1.27-1.50 (m, 4) H), 1.75-1.95 (m, 2H), 2.03-2.20 (m, 2H), 3.72-3.87 (m, 2H), 4.05 (s, 4H), 6.81 (br s, 2H)

F. Synthesis of frans-5,6-cyclohexano-1,10,13-tris (p-toluenesulfonyl) -1,4,7,10,13-pgntaazacyclopentadecane-3,8-dione (alternative method)

A five-liter, three-necked flask equipped with a magnetic stirrer and a one-liter separator was dried and placed in a dry argon atmosphere. To a solution of trans-1,2-bis (chloroacetamido) cyclohexane prepared as in Example 2E (6.68 g, 250 mmol) in anhydrous DMF (1.25 L) was added 1,4,7 solution at room temperature over 12 hours. -tris (p-toluenesulfonyl) -1,

4,7-triazaheptane-1,7-disodium salt prepared as in Example 1B (15.2 g, 25 mmol) in anhydrous DMF (1.25 L). After stirring for an additional 2 h, the solvent was removed in vacuo. The solid residue was triturated with chloroform (1 L) and filtered to give a white solid. The solid was recrystallized from acetonitrile to give 7.22 g (38% yield) of fluffy white crystals: mp 254

- 5 ° C; NMR (DMSO-d6) δ 1.15 (br s, 4H), 1.52-1.75 (m,

H), 2.42 and 2.43 (2s, 9H), 3.04 (m, 8H), 3.51 (d + m, J = 16.5 Hz, 4H), 4.01 ( d, J = 16.5 Hz, 2H). 7.35 - 7.53 (m, 8 H), 7.71 (d J = 8.3 Hz, 4 H), 7.80 (br d, J = 10.5 Hz,

H).

G, Synthesis of trans-2,3-cyclohexane-1,4,7,10,13-pentaazacycletrans-5,6-cyclohexano-1,10,13-tris (p-toluenesulfonyl) 1,4,7,10,13 -pentaazacyclopentadecane-3,8-dione, prepared as in Example 2D (1.755 g, 2.32 mmol), was suspended in 1,2-dimethoxyethane (dme, 40 mL) under an argon atmosphere, and the flask was placed in a water bath . Aluminum lithium hydride (0.5 M in dm, 55 mL, 27.5 mmol) was added over five minutes. Five minutes later, jacket heating was started, and fifteen minutes later reflux occurred. The reaction became almost colorless after heating under reflux for several minutes and later turned yellow with a white precipitate. Heating at reflux was continued for 43.5 hours and then the reaction mixture was allowed to cool to room temperature. The reaction was quenched by careful addition of water (0.86 mL) using a water bath to cool. Five minutes later, a 15% aqueous sodium hydroxide solution (0.86 mL) was added followed by water (2.6 mL). The slightly yellowish color was largely lost during this process. An hour later, tetrahydrofuran (55 mL) was added and stirring continued for 2 h. The quenched reaction mixture was filtered. The filtrate was evaporated under reduced pressure and placed on a vacuum line to give a yellow-white solid. This solid was dissolved in dichloromethane and then filtered, then concentrated to a solid and placed in a vacuum line. Was recrystallized from hot acetonitrile under argon to produce 0.316 g (50% yield) white needles: mp 112-3 ° C (under nitrogen) ^X H NMR (CDC1J δ 0.97 (m, 2 Η), 1.22 (m, 2 Η), 1.39 - 1.96 (3 m, 7 Η), 2.11 (m, 4 Η), 2.49 (m, 2 Η), 2.54 - 2.88 ( several m, 12H), 2.94 (m, 2H) Exact Mass (Μ + H) *: Calculated 270.2658, Found 270.2658 (C Η N).

32 5

H, Synthesis of Manganese (II) dichloro (trans-2,3-cyclohexano-1,4,7,

10,13-pentaazacyclopentadecane) 1 trans-2,3-Cyclohexano-1,4,7,10,13-pentaazacyclopentadecane added as in Example 2G (301 mg, 1.12 mol) was added to hot anhydrous MeOH (50 mL). ml), containing anhydrous magnesium chloride (140 mg, 1.12 mmol) under a dry nitrogen atmosphere. After refluxing for 2 hours, the solution was stirred at room temperature overnight and dried. The white solid was dissolved in warm acetone (15 mL) and the solution was filtered. The solution was evaporated to dryness and the white solid was washed with ethyl ether. The solid was dried under vacuum to give 0.36 g (82% yield) of the product: FAB mass spectrum (NBA) m / z (relative intensity) 394 (Μ *,

1), 359/361 [(M-Cl) &lt; + &gt;,100/29]; Anal. calculated for

C, N, MnCl: C, 42.54; H, 7.91; N, 17.72. Found: C,

42.56; H, 8.17; N, 17.42.

Example 3

A. Synthesis of Boc-DAla-Ala-QEt

To a solution of Boc-DAla (25.0 g, 132.1 mmol) in DMF (1450 mL) was added HOBT HO (19.8 g, 129.3 mmol) and EDC-HCl (28.0 g, 146.3) mmol) and the resulting solution was allowed to stir at room temperature for 30 minutes. To this solution was added alanine ethyl ester hydrochloride (20.3 g, 132.1 mmol) and TEA (20.4 mL, 146.3) and the reaction mixture was allowed to stir for three days (for convenience). The DMF was evaporated and the residue was partitioned between water (500 mL) and ethyl acetate (500 mL). The ethyl acetate solution was washed with 1 N NaHSO 4 (250 mL), water (250 mL), saturated NaHCO ₃ (250 mL), brine (250 mL) and dried over Na 2 SO ₄ . Filtration and concentration yielded 31.7 g. (83% yield) of the desired dipeptide as a white foam: NMR _(DMSO-dg) 6 1.14 (d, J =

7.4 Hz, 3 1,1), 1.16 _: (t, J = 7.4 Hz, 3 ohms), 1.24 (d, J = 7.0) Ht, 3 Η) 1.36 (s, 9 Η), 3.96 - 4.09 (m, 3 Η), 4.17 - 4.22 (apparent · quintet, J = 7.4 Hz, 1 Η), 6.77 (d, J = 7.7 Hz, 1 Η), 8.09 . (dy J = 7.0 Hz, .1 Η), MS (LCI, CH ₄ ( ) m / z (relation- intensity - intensity) = 317 (5) + C ₂ H _s ], 289 (60) [M + H] +. .

B, Synthesis of Boc-Ala-Ala-OH

To a suspension of the dipeptide (15.0 g, 93.6 mmol) in THF (192 mL) was added 0.5 N NaOH (192 mL). To the resulting solution was added di-t-butyl dicarbonate (26.6 g, 121.7 mmol) in one portion. The pH of the reaction was maintained for 5 hours at about 10 and the mixture was allowed to stir overnight. The pH of the reaction was again adjusted to probably 10 'and the solution was extracted with ethyl acetate (2 x 100 mL). The pH of the aqueous layer was adjusted, probably. 3.5 aqueous potassium bisulfate and this mixture was extracted with ethyl acetate (3 x 100 mL). The combined extracts were dried (MgSO _4), filtered and concentrated to give 20.7 g (85% yield) of desired product as a white powder: ¹ H NMR _(DMSO-dg) δ 1.16 (d, J = 6, 8 Hz, 3 H), 1.28 (d, J = 7.3 Hz, 3 H), 1.38 (s, 9 Η), 3.95 -4.49 (m, l Η), 4 20 _: (quintet, J = 7.3 Hz, 1H), 6.87 (d, J = 8.0 Hz, 1H), 8.00 (d, 7.3 Hz, 1H); MS (HRFAB, NBA-Li). m / z = 267.1557 [M + Li] ^&lt; ⁺ &^gt;; 267.1532 calculation for

C, Synthesis of DAla-Ala-OET TFA

The protected dipeptide (31.4 g, 109. mmol) was dissolved in methylene chloride (200 mL) and TFA (66 mLK) was added. The resulting solution was allowed to stir for 30 minutes at room temperature and then concentrated. 2 x 200 ml), dissolved in ether and eluted by addition of excess hexanes: The solvents were decanted and the residue was pumped under high vacuum for 12 hours to give 39.6 g (100% yield, containing residual TFA) of the desired salt TFA as an orange oil: NMR _(DMSO-d6) <5 1.16 (t, J = 7.0 Hz, 3 H), 1.28. (d, J = 7.0 Hz, 3H) 1.34 (d, J = 7.0 Hz, 3 H), 3.86 (bs, 1 H), 4.07 (q, J = 7.0 Hz, 2 H), 4.26 (quintet) J - 7.0 Hz, 1H), 8.21 (bs, 3.H), 8.86 (d, J = 7.4 Hz, 1H); MS (LRCI, CH3) m / z (relative intensity) 217 (5) [M + C_H _s] * 189 (40) [M + H] +.

D, Synthesis of Boc-Ala-Ala-DAla-Ala-OEt

To a solution of Boc-Ala-Ala-OH (20.1 g, 77.2 mmol) in DMF (850 mL) was added ΗΟΒΤ · Η ₃ Ο (13.1 g, 85.4 mmol) and EDC-HCl (16 mL). , 4 g, 85.4 mmol). To this solution was added DAla-Ala-OEt · TFA (23.3 g, 77.2 mmol) followed by TEA (11.9 mL, 85.4 mmol). and the resulting mixture was then stirred for 12 h. The DMF was evaporated and the residue was dissolved in ethyl acetate (300 mL) and washed with 1 N potassium bisulfate (150 mL), water (150 mL), saturated sodium bicarbonate (150 mL) and brine (150 mL). The ethyl acetate layer was dried (MgSO 4), filtered and concentrated to reduce the volume in half and crystallization could begin. Isolation by filtration gave 20.5 g (62% yield) of the desired tetrapeptide as a white solid: NMR (DMSO-d ₆ ) δ 1.13 (d, J = 7.0 Hz,

3. H), 1.17 (two coincidence d, J = 7.0 Hz, 6 H), 1.25 (d, J = 7.4 Hz, 3 H), 3.91-4, 30 (m, 6H), 6.87 (d, J = 7.0 Hz, 1H), 7.92 (d, J = 6.3 Hz, 1H), 8.07 (d, J = 7., 3. Hz, 1H), 8.09 (d, J = 6.6 Hz, 1H); MS (HRFAB, NBA-Li) m / z = 437.2600 [M + Li] +; 437.2588 calcd.

E, Synthesis of Boc-Ala-Ala-DAla-Ala-OH

A solution of Boc-Ala-Ala-DAla-Ala-OEt (10.9 g, 25.3 mmol) in n-methanol (100 mL) was treated with 2.5 M sodium hydroxide (20.0 mL, 50 mL), 0 ml) and the resulting solution was allowed to stir for two hours at room temperature. At the same time, the pH of the solution was reduced to probably 3. aqueous potassium bicarbonate was added and the resulting mixture was extracted with ethyl acetate (3 x 100 mL). The combined extracts were dried (MgSO 4), filtered and concentrated. to give 6.8 g (67% yield) of the desired acid. as a solid: NMR ^{Χ Η} (DMSO-d _s) 6 1.17 (d, J = 7.2 Hz, 3H), 1.20 (two coincidental d, J = 7.1 Hz,

H), 1.28 (d, J = 1.3 Hz, 3 H), 1.38 (s, 9 H), 3.90-4.00 (m, 1H), 4.17-4, 30 (m, 3 H), 6.93 (d, J = 6.7 Hz, 1H),

7.96 (d, J = 6.7 Hz, 1H), 8.04 (d, J = 7.4 Hz, 1H), 8.07 (d = 7.8 Hz, 1 Hz); MS (HRFAB, NBA-Li) m / z = 409.2331 [M + Li] +; 409.2353 calcd for C 24 H 25 N 4 O 4 Li.

F. Synthesis of Boc-Ala-Ala-DAla-Ala-DAla-OBzl

To a solution of Boc-Ala-Ala-DAla-Ala-OH (6.5 g, 16.3 mmol) in DMF (180 mL) was added ΗΟΒΤΊΓΟ (2.86 g, 18.7 mmol) and EDC HCl (3 mL). 58 g, 18.7 mmol). The resulting solution was allowed to stir for 15 minutes at room temperature and treated with DAla-OBzl p-toluene sulfonate salt (5.57 g, 18.7 mmol) and TEA (2.6 mL, 18.7 mmol). The mixture was then allowed to stir for 12 hours. The DMF was evaporated and the residue was partitioned between ethyl acetate (300 mL) and water (300 mL). The ethyl acetate layer was washed with 1 N NaHSO 4 (150 mL), water (150 mL), saturated NaHCO ₃ (150 mL), and brine (150 mL). The ethyl acetate layer then byla'vysušena MgSO _4, filtered and zkoiicentrována to give 9.0 g (100% yield) of the desired compound as a white powder: ^X H 'NMR _(DMSO-dg) 1.17 (d, J = 7, 3 Hz, 3 H) i, 21 (two coincidence d, J = 7.0 Hz, 6 Η), 1.22 (d, J =

7.0 Hz, 3 Η), 1.32 (d, J = 7.3 Hz, 3 Η), 1.37 (s, 9 Η),

3.90 - 4.09 (m, 1H), 4.18 -. 4.34 (m, 4Η), 5.13 (ABq, J =

12.7 Hz, = 10.5 Hz, 2 Η), 6.94 (d, J = 7.3 Hz, 1 Η), 7.30

- 7,41. (m, 5 Η), 7.97 (d, J = 7.0 Hz, 1 Η), 8.10 - 8.18 (m,

2H), 8.25 (d, J = 6.9 Hz; 1H) MS (HRFAB, NBA-Li) m / z = 570.3140 [M + Li] +; 517.3115 calcd for C C HH ^N ^L Li.

G. Synthesis of Ala-Ala-DAla-Ala-DAlarHCl

Boc-Ala-Ala-DAla-Ala-OEt (10.4 g, 18.7 mmol) was dissolved in acetic acid (225 mL) and treated with concentrated hydrochloric acid (75 mL). The resulting solution was then allowed to stir at room temperature for 14 hours. The reaction mixture was then concentrated, coevaporated with water (50 mL) and azeotroped with toluene coevaporation (2 x 100 mL) to give 7.8 g (96% yield) of the freed pentapeptide hydrochloride as a white powder; ¹ H NMR (D ₂ O) d 1.29-1.39 (m, 1H), 1.47 (d, J = 7.0 Hz, 3 H), 4.06 (q, J = 7, 0 Hz, 1H), 4.18-4.38 (m, 4H); MS (LRFAB, NBA-HCl) 374 [M + H] +.

H, Cyclo- (Ala-Ala-DAla-Ala-DA (a -) synthesis)

To a solution of Ala-Ala-DAla-Ala-DAla · HCl (7.8 g, 19.0 mmol) in DMF (2400 mL) at -40 ° C was added DPPA (6.29 g, 22.8 mmol). mmol) and enough TEA to adjust the pH to about 8 (measured by dropping the reaction mixture on moistened hydrion paper). This solution was allowed to stand at -23 ° C for 48 hours and at 8 ° C for 48 hours. During this time, the pH was again maintained at probably 8 by regular addition of TEA. At the end of this time, the reaction mixture was poured into water (2400 mL) and mixed with the mixed ion exchange resin (1200 g) for 6 hours. The resin was removed by filtration and the filtrate was concentrated to a volume of probably 100 ml. Ether (500 mL) was added and the precipitated white solid was collected by filtration and washed with excess ether. The solid was then triturated by stirring with THF (100 mL) for 12 h (to remove traces of DMF), filtered and thoroughly dried to give 3.15 g (47% yield) of the desired cyclic peptide as a fine white powder; 1 H NMR (DMSO-d 6) 1.08-1.25 (m, 12H), 1.24 (d, J = 7.3Hz, 3H), 4.00-4.10 (m, 1H) H), 4.26-4.30 (m, 2H), 4.34 (q, J = 7.2 Hz, 1H), 4.41 (q, J = 7.6 Hz, 1H) 7.58 (d, J = 7.0 Hz, 1H), 7.83 (d, J = 8.4 Hz, 1H), 8.22 (d, J = 6.2 Hz, 1H) ), 8.33 (d, J = 7.81, 1H), 8.49 (d, J = 6.8 Hz, 1H); MS (HRFAB, NBA-HCl) m / z = 356.1989 . (Μ + Η) *; 356.1934 calculation for C _ig H _2s N _s O _s (Μ + H)

Synthesis of (2S, .5R, 8S, 11R, 14, S) Rpentamethyl-1,4,7,10,13pentaazacyclopentadecane

To a stirred suspension of cyclo- (Ala-Ala-DAla-Ala-DAla-) (3.10 g, 8.70 mmol) in THF (70 mL) at room temperature was added aluminum hydride (108 mL of a 1.0 M solution in THF, 108 mL). Resulting. the mixture was stirred at room temperature for 2 hours and then at room temperature. 16 hours under reflux. The mixture was then cooled to probably -20 ° C and quenched by the dropwise addition of saturated sodium sulfate (probably 30 mL). The resulting mixture was concentrated to a dry white powder and this powder was triturated with ether (2 x 150 mL). The combined triturates were concentrated and recrystallized from acetonitrile to afford 1.10 g (44% yield) of the desired ligand as a white solid. 1 H NMR (CDCl ₃ ) d 0.96 (d, J, = 5.2 Hz, 3 Η), 1.00 (two coincidence d, J = 5.0 Hz, 6 Η), 1 , 02 (two coincidence d, J = 5.0 Hz, 6 Η),

I, 30-1.55 (bm, 2H), 1.85-2.15 (bs, 3 ohms), 2.05 -2.19 (m, 5 ohms), 2.42-3.00 { complex m, 12H); MS (HRFAB, NBA-HCl) m / z = 286.3013 (Μ + Η) +, 286.2971 calcd for C Η N (Μ + H) +.

3GS

J. Synthesis of Manganese (JI) dichloro- (2S, 5R, _as ^ 11Κ, 14-3Ϊ-penta-methyl-1,4.7,10,13 -pentaazacyclopentadecane)

To a stirred solution of anhydrous MnCl 2 (79.0 mg, 0.62 mmol) in hot ethanol (5 mL) was added (2S, 5R, 8S, 11R,

14S) -pentaazacyclopentadecane (177 mg, 0.62 mmol). The solution was refluxed for one hour and stirred at room temperature for a further 16 h. The solution was filtered through celite, concentrated to half volume and combined with ether (30 mL). The white crystals were isolated by filtration and dried under vacuum to give 187 mg (73% yield) of the complex as a white solid: MS (LRFAB, NBA) m / z (relative intensity) = 410 (5) [M] +,

375/377 (100/30) [M cr

Anal. calculated for

CΗΗNMnCl: C, 43.75; H, 8.57; N, 17.01. Found: C,

43.74; H, 8.60; N, 16.97.

Example 4

A. Synthesis of N- (p-toluenesulfonyl) - (R, R) -1,2-diaminocyclohexane

To a stirred solution of (R, R) -1,2-diaminocyclohexane (300 g, 2.63 mol) in CH 2 Cl 2 (5.00 L) at -10 ° C was added dropwise a solution of p-toluenesulfonyl chloride dropwise over 7 hours. (209 g, 1.10 mol) in CH? Cl _from (5.00 1), maintaining the temperature between -5? -10 ° C. The mixture was allowed to warm to room temperature overnight while stirring. The mixture was concentrated in vacuo to a volume of 3 L and the white solid was filtered off. The solution was then washed with H 2 O (10 x 1 L) and dried

MgSO4. Removal of solvent in vacuo gave 286 g (97.5% yield) of product as a yellow crystalline solid: ^X H NMR (CDC1J d 0.98 to 1.27 (m, 4 H), 1.54 to 1.66 (m (2H), 1.81-1.93 (m, 2H), 2.34 (dt, J = 4.0, 10.7 Hz, 1H), 2.42 (s, 3H), 2.62 (dt, J = 4.2, 9.9 Hz, 1 H), 7.29 (d, J = 8.1 Hz, 2H), 7.77 (d, J = 8.3 Hz) MS (LRFAB-DTT

- DTE) m / z = 269 [D + H] *.

B. Synthesis of N- (p-toluenesulfonyl) -Ν '- (Boc) - (R, R) -1,2-diaminocyclohexane

To a stirred, solution of N- (p-toluenesulfonyl) - (R, R) -1,2-diamino-cyclohexane prepared as in Example 1A (256 g _r 0.955 mol) in THF (1.15 L) was added 1 N aqueous NaOH (1.15 L, 1.15 mol), then di-t-butyl dicarbonate (229 g, 1.05 mol) was added and the resulting mixture was stirred overnight The layers were separated and the aqueous layer was adjusted to pH 2 with 1 N HCl and saturated NaCl, the aqueous solution was then extracted with CH 2 Cl 2 (2 x 500 mL), and the extracts and the THF layer were combined and dried over MgSO 4 The solvent was evaporated in vacuo to give a yellow solid. - ether-hexane mixture to give 310 g (88.1% yield) of the product as a white crystalline solid: mp: 137-139 ° C; Ή NMR (CDCl 3) 1.04-1.28 (m, 4 Η) ), 1.44 (s, 9 Η), 1.61 - 1.69 (m, 2 Η), 1.94 - 2.01 (m, 2 Η), 2.43 (s, 3 Η), 2.86 (brs,

Η), 3.30 (br d, J = 9.6 Hz, 1 Η), 4.37 (br d, J = 6.7 Hz,

Δ), 5.48 (br d, J = 4.6 Hz, 1 Η), 7.27 (d, J = 9.7 Hz,

Δ), 7.73 (d, J = 8.1 Hz, 2H); MS (LRFAB, NBA-Li) m / z = 375 [M + Li] +.

Synthesis of Boo- (R, R) -Cyc (Ts) -gly; QMe

To a stirred solution of N- (p-toluenesulfonyl) -1 '- (Boc) (R, R) -1,2-diaminocyclohexane prepared as in Example IB (310 g, 0.841 mol) in anhydrous DMF (3.11 L) at NaH (37.4 g - 60% in oil, 0.934 mol) was added portionwise at 0 ° C and the resulting mixture was stirred for 30 minutes. Methyl bromoacetate (142 g, 0.925 mol) was then added dropwise over 45 minutes and the mixture was allowed to warm to room temperature with stirring overnight. After stirring for 17 h, the solvent was distilled off in vacuo and the residue was dissolved in ethyl acetate (3 L) and H 2 O (1.1). The ethyl acetate solution was washed with saturated NaHCO 3 (11), saturated NaCl (500 mL), and dried over MgSO ₄ . The solvent was removed in vacuo and the resulting oil was dissolved in ether. Crystallizers with the addition of hexanes gave 364 g (98% yield) of the product (TLC (98: 2 CHCl ₃ -MeOH / silica gel / UV determination) showed the product contained about 5% starting material) as colorless needles: bt pure sample 15 , 1. - 2 ° C; ¹ H NMR (CDCl 3) δ 1.11-1.22 (m, 4H); 1.45 (s, 9µ), 1.64-1.70 (m, 3µ), 2 , 16 - 2.19 (m, 1H), 2.43 (s, 3 Η), 3.34 - 3.40 (m, 2 Η), 3.68 (s, 3 Η), 4.06 (s) ABq, J = 18.5 Hz, = 155 Hz, 2 Η), 4.77 (br s, 1H), 7.30 (d, J = 8.3 Hz, 2 Η), 7.82. (d, J = 8.3 Hz, 2H); MS (LRFAB-DTT-DTE) m / z = 441 [M + H] +.

D-Synthesis of Boc- (R.R 1 -Cyc (Ts) -Gly-OH

To a stirred solution of impure Boc- (R, R) -CyC (Ts) -Gly-OMe prepared as in Example 1C (217 g, 0.492 mol) in MeOH (1.05 L) was slowly added 2.5 N aqueous NaOH (295 mL, 0.737 mol) and the resulting solution was stirred for two hours. The solvent was removed in vacuo and the residue was dissolved in H _and O (1.5 L). The solution was filtered to remove a small amount of solid and was washed with ether (7 x 11) to remove impurity (Compound 1B) which, after drying the combined washings over MgSO 4 and evaporating the solvent in vacuo, yielded 8.37 g. PH The aqueous solution was then adjusted to 2 with 1 N HCl and the product was extracted with ethyl acetate (3 x 1 L). The extracts were combined, washed with saturated NaCl (500 mL; and dried over MgSO. The solvent was removed in vacuo and the residue of ethyl acetate removed by coevaporation with ether (500 mL) and then CH 2 Cl 2 (500 mL) to yield 205 g ( 97.6 yield) of the product as a white foam;

1 H NMR (CDCl ₃ ) δ 1.15 - 1,22 (m, 4 Η), 1.48 (with, 9 H) 1.68 (m, 3 ohms), 2.12 - 2,15 (m, 1 Η), 2.43 (with, 3 H) 3.49 (m, 2), 3.97 (ABq, J = 17.9 Hz, = 69.6 Hz,

4.79 (br s, 1 Η), 7.31 (d, J = 8.1 Hz, 2 Η), 7.77 (d, J =

8.3 Hz, 2 H), 8.81 (br s, 1H); MS (LRFAB, NBA-Li) m / z =

433 [M + Li]

Synthesis of N- (p-toluenesulfonyl) - (S.S) -1,2-diaminocyclohexane

To a stirred solution of (S, S) -1,2-diaminocyclohexane (300 g, 2.63 mol) in CH 2 Cl 2 (5.00 L) at -10 ° C was added dropwise a solution of p-toluenesulfonyl chloride over 8 hours. (209 g, 1.10 mol) in CH 2 Cl 2 while maintaining the temperature at -5 to -10 ° C. The mixture was allowed to warm to room temperature overnight with stirring. The mixture was concentrated in vacuo to a volume of 31a and the white solid was filtered off. The solution was then washed with H 2 O (10 x 1 L) and dried over MgSO 4. Removal of solvent in vacuo gave 289 g (98.3% yield) of product as a yellow crystalline solid: NMR ^Τ Η (CDC1 ₃₎ 0.98 - 1.27 (m, 4 Η), 1.55 to 1.66 ( m, 2 Η), 1.81 - 1.94 (m, 2 Η), 2.32 (dt, J = 4.0, 10.9 Hz, 1 Η), 2.42 (s, 3 Η) 2.61 (dt, J = 4.0, 9.9 Hz, 1 Η), 7.30 (d, J = 7.9 Hz, 2 Η), 7.77 (d, J =

8.3 Hz, 2 Η); MS (LRFAB GT-HCl) m / z = 269 (M + H) +.

F. Synthesis of N- (p-toluenesulfonyl) -1 '- (Boc) - (S, S) -1,2-diaminocyclohexane

To a stirred solution of N- (p-toluenesulfonyl) - (S, S) -1,2-diaminocyclohexane prepared as in Example 1E (289 g, 1.08 mol) in THF (1.29 L) was added 1 N aqueous solution NaOH (1.29 L, 1.29 mol) was then added di-t-butyl dicarbonate (258 g, 1.18 mol) and the resulting mixture was stirred overnight The solid was filtered and washed with THF. The aqueous layer was adjusted to pH 2 with 1 N HCl and saturated with NaCl The aqueous solution was then extracted with CH 2 Cl 2 (2 x 500 mL) and the extracts and THF layer were combined, washed with saturated NaCl (500 mL) and dried over MgSO 4. Crystallization with ether added 364 g (91.9% yield) of the product as colorless needles: mp 137-139 ° C; 1 H NMR (CDCl 3) δ 1.06- 1.31 (m, 4H), 1.44 (s, 9H), 1.60-1.69 (m, 2H),

1.95-1.99 (m, 2H), 2.42 (s, 3H), 2.86 (brs, 1H), 3.30 (br d, J = 2.6 Hz, 1 H) H), 4.41 (br d, J = 7.3 Hz, 1 H), 5.54 (br d, J = 5.4 Hz, 1 H), 7.28 (d, J = 8.1) Hz, 2 H), 7.73 (d,

J = 8.3 Hz, 2H); MS (LRFAB NBA-HCl) m / z = 369 [M + H].

G, Synthesis of Boc- (S, S) -Cyc (TsI-qlysQMe

To a stirred solution of N- (p-toluenesulfonyl) -N '- (Boc) - (S, Ξ) -1,2-diaminocyclohexane prepared as in Example 1F (364 g, 0.989 mol) in anhydrous DMF (3.66 L) NaH (47.4 g - 60% in oil, 1.19 mol) was added portionwise at 0 ° C and the resulting mixture was stirred for 1.5 h. The mixture was warmed to room temperature, stirred for an additional 30 min and then cooled back to 0 ° C. Methyl bromoacetate (189 g, 1.24 mol) was added dropwise and the mixture was allowed to warm to room temperature overnight with stirring. After stirring for 17 h, the solvent was removed in vacuo and the residue was dissolved in a mixture of ethyl acetate (3 L) and H 2 O (1 L). The layers were separated and the ethyl acetate solution was washed with saturated NaHCO 3 (1 L), Η ₌ 0 (1 L), saturated NaCl (2 x 500 mL) and dried over MgSO ₄ . The solvent was removed in vacuo and the resulting oil was dissolved in ether. Crystallization with the addition of hexanes gave 290 g of crude product as yellow needles. An additional 180 g was obtained from the filtrate as an oil. TLC (98: 2 CHCl ₃ -MeOH / silica gel / UV assay) showed that both the solid and the oil contained the starting material. ^Χ Η NMR (CDCl δ 1.06 to 1.29 (m, 4 Η), 1.44 (s, 9 Η), 1.58 - 1.66 (m, 3 Η), 2.17 - 2 19 (m, 1 Η), 2.43 (s, 3 Η), 3.28 - 3.43 (m, 2 Η), 3.68 (s, 3 Η), 4.25 (ABq, J = 18.5 Ηζ, _Δν -115 Hz, 2 Η), 4.76 (br s, 1 Η), 7.31 (d, J = 8.3 Hz, 2 Η), 7.83 (d, J = 8.3 Hz, 2 H); MS (LRFAB NBA-Li) m / z = 447 [Μ + H] +.

H, Synthesis of Boc- (S, S) -Cyc (Ts) -Gly-OH

To a stirred solution of impure Boc- (S, S) -Cyc (Ts) -Gly-OMe prepared as in Example 1G (197 g, 0.447 mol) in MeOH (925 mL) was slowly added 2.5 N aqueous NaOH (268 mL). ml, 0.670 mol) and the resulting mixture was stirred for two hours. The solvent was removed in vacuo and the residue was dissolved in H 2 O (1 L). The solution was washed with ether (4 x 1 L) to remove impurity (compound 1F) which, after drying the combined washings over MgSO 4 and removing the solvent in vacuo, yielded 14.8 g. The pH of the aqueous solution was then adjusted to 2. with 1 N HCl and the product was extracted with ethyl acetate (3 x 1 L). The extracts were combined, washed with saturated NaCl and dried over MgSO 4. The solvent was removed in vacuo to yield 171 g (89.7% yield) of product as an oil which crystallized on standing: ^X H NMR (CDCl) δ 1.10 to 1.22 (m, 4 Η), 1, 45 (S, 9 Η), 1.55 - 1.68 (m, 3 Η), 2.13 - 2.16 (m, 1 Η), 2.43 (s, 3 Η), 3.39 - 3.41 (m, 2Η), 4.00 (ABq, J = 18.1Ηζ, Αν ·

80.4 Hz, 2 Η), 4.82 (br s, 1 Η), 7.31 (d, J = 8.3 Hz, 2 Η),

7.75 (d, J = 8.3 Hz, 2 H), 9.28 (br s, 1H); MS (LRFAB NBA

- Li) m / z = 433 [M + Li] +

I, Synthesis of Boc- (S, 5) -Cyc (Ts) -Gly-Gly-QEt

To a stirred solution of Boc- (S, S) -Cyc (Ts) -Gly-OH prepared as in Example IH (26.7 g, 62.5 mmol) in anhydrous degassed DMF (690 mL) was added HOBT (10.1 g, 75.0 mmol) and EDC-HCl (14.4 g, 75.0 mmol). After the resulting solution was stirred for 30 min, glycine ethyl ester hydrochloride (9.60 g, 68.8 mol) was added and the pH was adjusted to 8 by TEA. After stirring for 2.75 days, the solvent was removed in vacuo. The residue was dissolved in a mixture of ethyl acetate (1 L) and H 2 O (11) and the layers were separated. The aqueous layer was extracted with ethyl acetate (1 L) and the extracts were combined. The ethyl acetate solution was washed with 0.1 N HCl (1 L), saturated NaHCO ₃ (1 L), saturated NaCl (500 mL) and dried over MgSO 4.

The solvent was removed in vacuo to give 30.2 g (94.4% yield) of the product as a white foam: ¹ H NMR (CDCl 3) d 1.19-1.23 (m, 3H), 1.28 (t, J) = 7.05 Hz, 3H),

1.42 (s, 11K), 1.63-1.71 (m, 2H), 2.16-2.18 (m, 1H),

2.43 (s, 3H), 3.50-3.57 (m, 2H), 3.83 (ABq, J = 17.7 Hz, = 35.7 Hz, 2H), 4.01 (dABq) , J = 6.05, 17.92 Hz, av ₌ 28.9 Hz,

H), 4.20 (q, J = 7.3 Hz, 2H), 4.88 (br, 1H), 7.31 (d,

J = 8.3 Hz, 2H), 7.36 (br s, 1H), 7.73 (d, J = 8.3 Hz,

H); MS (LRFAB NBA-HCl) m / z = 512 [M + H] +. .

J. Synthesis of (S.S) -Cyc (Ts) -Gly-Glv-OEt TFA salt

To a stirred solution of Boc- (S, S) -Cyc (Ts) -Gly-Gly-OEt prepared as in Example II (30.1 g, 58.8 mmol) in CH 2 Cl 2 (265 mL) was added TFA ( 63 ml) and the resulting solution was stirred for 30 minutes. The solvent was evaporated in vacuo and the residual TFA was co-evaporated with CH 2 Cl 2 (2 x 1 L) and ether (1 L).

The oil was then triturated with ether (2 x 1 L) and decanted. The resulting foam was dried under vacuum to give 33.7 g (expected quantitative yield) of the product as a dark powder: ¹ H NMR (CDCl 3 δ 0.96 - 1.23 (m, 4 H), 1.25 (t, J) = 7.3 Hz, 3 H), 1.51 - 1.66 (m, 3 H), 2.12 - 2.26 (m, 1

H), 2.41 (S, 3H), 2.98-3.10 (brs, 1H), 3.67-3.71 (m,

H), 4.04 (ABq, J = 17.7 Ηζ, ^{ν -} 154 Hz, 2 H), 4.04 (d, J =

4.4 Hz, 2 H), 4.17 (q, J = 7.3 Hz, 2 H), 7.29 (d, J = 8.3 Hz, 2 H), 7.70 (d, J = 8.3 Hz, 2H), 8.04 (br s, 1H), 8.14 (br s, 3H); MS (LRFAB NBA-HCl) m / z = 412 [M + H] +.

K. Synthesis of Boc- (R.R) -Cyc (Ts) -Gly- (S.S) -Cvc (Ts) -Gly-Gly-OEt

To a stirred solution of Boc- (R, R) -Cyc (Ts) -Gly-OH prepared as in Example ID (25.1 g, 58.8 mmol) in anhydrous, degassed DMF (650 mL) was added HOBT (9, 54 g, 70.6 mmol) and EDC-HCl (13.5 g, 70.6 mmol). After the resulting solution was stirred for 30 min, (S, S) -Cyc (Ts) -Gly-Gly-OEt TFA salt prepared as in Example 1J (33.6 g, 58.8 mmol) was added and the pH was adjusted with TEA After stirring for 2.75 days, the solvent was removed in vacuo. The residue was dissolved in a mixture of ethyl acetate (11) and H 2 O (1 L) and the layers were separated.The ethyl acetate solution was washed with 0.1 N HCl (2 x 1 L), saturated NaHCO ₃ (2 x 11), saturated NaCl ( 500 ml) and dried over MgSO 4 The solvent was evaporated in vacuo to give 47.5 g (98.4% yield) of the product as a dark foam ^Χ Η NMR (CDCl 3 δ 1.12 - 1.83 (τη, 26 Η)) , 2.21 - 2.24 (m, 2 Η), 2.42 (s, 3 K), 2.43 (s, 3 Η), 3.36 - 3.51 (br S,

Η), 3.68 - 3.96 (m, 6 Η), 4.00 (d, J = 5.4 Hz, 2 Η), 4.19 (q, J = 7.1 Hz, 2 Η) 4.72 (br s, 1 Η), 6.78 (br s, 1 Η), 7.31 (d, J = 8.1 Hz, 4 Η), 7.46 (br s, 1 Η) 7.79 (m, 4H); MS (LRFAB NBA-HCl) m / z = 820 [Μ + H] +,

L, Synthesis of Boc- (R, R) -Cyc (Ts) -Gly- (3, S) -Cyc (Ts) -Gly-Gly-QH

To a stirred solution of Boc- (R, R) -Cyc (Ts) -Gly- (S, S) Cyc (Ts) -Gly-Gly-OEt prepared as in Example 1K (47.4 g,

57.8 mol) in MeOH (240 mL) was added 2.5 N aqueous NaOH (34.7 mL, 86.7 mol) and the resulting mixture was stirred for two hours. The solvent was removed in vacuo and the residue was dissolved in H ₃ O (1 L). The solution was washed with ether (2 x 1 L) and the pH of the aqueous solution was adjusted to 2 with 1

N HCl. The solution was then saturated with NaCl and extracted with ethyl acetate. (3 x 11). The combined extracts were dried over MgSO ₄ and the solvent was evaporated. removed in vacuo. Residual ethyl acetate removed by coevaporation with CH ₂ C1 ₂ and the resulting foam was dried in vacuo to give 45.7 g (99.7% yield) of product as a dark, powder ^X H NMR (CDC1J δ 1.16. - 1.75 (m, 23H),

2.13-2.17 (m, 2H), 2.41 (s, 3H), 2.42 (s, 3H), 3.49

4.16 (m, 10H), 4.53 (brs, 1H), 7.01 (brs, 1H), 7.30 (d, J = 8.1 Hz, 4H) 7.40 (br s, 1H), 7.79 (d, J = 8.1 Hz, 2H), 7.86 (d, J = 7.7 Hz, 2H), 10.40 ( br s, 1H); MS '(LRFAB NBA-HCl) m / z = 792 [M + H] +.

· Γ.

M. Synthesis of (R, R) -Cyc (Ts) -Gly- (S.S) -Cyc (Ts) -Gly-Gly-OH-TFA salts.

To a stirred solution of Boc- (R, R) -Cyc (Ts) -Gly- (S, S) Cyc (Ts) -Gly-Gly-OH. prepared as in Example IL, (45.5 g,

57.5 mol) in CH ₂ C1 ₂ (260 mL) .byl. TFA (60 mL) was added; The resulting solution was stirred for 30 minutes and the solvent was removed in vacuo. The residual TFA was removed by coevaporation with CH _and Cl ₂ (3 x 1 L) and trituration of the resulting foam with ether (1 L, 2 x 750 mL), decanting the ether each time. After drying under vacuum, 47.4 g (100% yield) of the product was obtained as an off-white powder: NMR (CDCl 3 d 1.05-1.31 (m, 9H)),

1.48-2.63 (m, 5H), 2.11-2.21 (m, 2H) H), 2.40 (s, 3H), 2.42 (s, 3H), 3.25 (br. S, 1 H), 3.60 - 3.80 (m, 3H), 3.83 4.19 (m, 6H). 6.94 (br s, 1 H), 7.31 (m, 4 H), 7.69 (m,

H), 7.83 (br s, 3H), 13.17 (br s, 2H); MS (LRFAB DTT-DTE) m / z = 692 [M + H] +.

N. Synthesis of cyclo-f (R, R) -Cyc (Ts) -Gly- (S.S) -CvclTs) -Gly-Glyl

To a stirred solution of (R, R) -Cyc (Ts) -Gly- (S, S) -Cyc (Ts) Gly-Gly-OH-TFA salt prepared as in Example 1M (32.2 g, 40.0 mmol) in anhydrous degassed DMF (10.0 L) at -78 ° C was added DPPA (13.4 g, 48.8 mmol). of the solution was adjusted to 8 with TEA and the solution was allowed to stand at -78 ° C for 6 h. The pH was readjusted to 8 with TEA and the solution was warmed to -45 ° C for 24 h. After re-adjusting the pH as before, the solution was allowed to warm to -40 ° C over 24 h. The pH was readjusted as above and the solution was allowed to stand at -20 ° C for 24 h. The pH was readjusted as above and the solution was allowed to warm to 2 ° C for 24 h. The pH dropped only slightly. The pH was readjusted as before and the solution was allowed to stand at 2 ° C for an additional 24 h, during which the pH was no longer changed. The solution was equally divided into 6 4 L beakers and H ₂ O (1.1 L) was added to each. A total of 5.00 kg of mixed ion exchange resin (evenly divided into 6 beakers) was then added to the solution and mixed for 6 h. The resin was then filtered and washed with DMF. The solvent was then removed in vacuo and the solid residue was dissolved in MeOH (100 mL) and filtered to remove the fine suspension. The solution was then concentrated in vacuo to a volume of 25 mL and ether was added periodically during crystallization to give 22.2 g (82.5% yield) of the product as colorless needles;

b, mp 190-200 ° C; ¹ H NMR (CDCl 3) δ 0.87 - 2.13 (m, 16 Η), 2.41 (s, 3 Η), 2.45 (s, 3 Η), 3.56 - 3.97 (m, 10 Η), 6.66 (br s, 1 H), 7.18 (br s, 1 Η), 7.34 (d, J = 8.1 Hz, 4 Η),

7.65 (br s, 1 Η), 7.71 (d, J = 7.3 Hz, 2 Η), 7.89 (d, J =

7.3 Hz, 2H); MS (LRFAB NBA-Li) m / z = 680 [M + 1] +.

O, Synthesis of 2,3- (R, R) -8,9- (S.S) -bis-cyclohexano-1,4,7,10,13pentaazacyclopentadecane

To a stirred solution of the cyclo - [(R, R) -Cyc (Ts) -Gly- (S, S) Cyc (Ts) -Gly-Gly] prepared as in Example 1 N (19.4 g,

A solution of 1.0 M LiAlH 4 in THF (345 mL, 345 mmol) was added dropwise over 30 min. The yellow homogeneous solution was refluxed for 20 hours (during which it became heterogeneous) and then cooled to 0 ° C. The mixture was then quenched by dropwise addition of 10% sodium sulfate (50 mL) while cooling in an ice bath. The solids were filtered off under argon and THF was removed in vacuo to give a rapidly crystallizing oil. The solids were then refluxed with anhydrous THF (1 L) for 1 hour and the mixture was filtered and the solvent removed in vacuo as before. The solids were then refluxed with a mixture of THF (1 L) and MeOH (500 mL) for 1 h and treated as before. The extractions were then dissolved in anhydrous THF, combined and the solids filtered. The solvent was evaporated in vacuo and the yellow foam was dried by azeotropic distillation of H 2 O with toluene (1.75 L) under vacuum at 90 ° C. The solids were then refluxed with hexanes (1 L) for 30 min and the hot solution transferred to a dark flask and the solvent evaporated in vacuo to give 6.1 g of an oil which crystallized on standing. The remaining solids were refluxed with hexanes as before to give 1.4 g of an oil which crystallized on standing. The solids were then dissolved in MeOH and toluene (1 L) was added. The solvent was evaporated in vacuo and any remaining H 2 O was removed by azeotropic distillation with toluene (1 L) and then hexanes (3 x 1 L). The resulting white powder was heated to reflux with hexanes (1 L) for 2 h under argon and filtered into a dark flask. The solvent was evaporated in vacuo to give 1.7 g of an oil which crystallized on standing. Crystalline residues from all three extracts were dissolved in hexanes and mixed. A small amount of turbidity was filtered off and the solution concentrated to give 5.3 g (57% yield) of the product as a pale yellow crystalline solid. Recrystallization from acetonitrile gave 4.47 g (48% yield) of a colorless crystalline solid: mp 107-8 ° C; ^Χ Η NMR (CDCl @) 5 0.95 to 1.01 (m, 4 Η), 1.19 to 1.24 (m, 4 Η), 1.70 to 1.73 (m, 4 Η)

1.97 (br s, 5 ohms), 2.03-2.14 (m, 8 ohms), 2.49-2.58 (m, 6 ohms)

Δ), 2.74-2.80 (m, 2H), 2.85-2.90 (m, 2H), 2.94-2.99 (m, 2H); MS (LRFAB, NBA) m / z = 324 [M + H] analysis calculated for C 17 H 17 N 3 O: C, 66.83; H, 11.53; N, 21.65. Found: C, 66.80; H, 11.44; N, 21.71.

P, Synthesis of [manganese (II) dichloro (2,3- (R, R) -8,9- (S, S) -bis-cyclohexano-1,4,7,10,13-pentaazacyclopentadecanyl chloride)

To a stirred solution of anhydrous MnCl 2 (1.67 g, 13.3 mol) in hot methanol (120 mL) was added 2,3- (R, R) -8,9- (S, S) bis-cyclohexano-l, The 4,7,10,13-pentaazacyclopentadecane prepared as in Example 10 (4.30 g, 13.3 mmol) was refluxed and then stirred at room temperature overnight. Crystallization from ether gave 5.11 g (85.6% yield) of the product as an off-white crystalline solid: MS (LRFAB, NBA) m / z (relative intensity) = 448. (2) [M] +, 413/415. ( 100.33} [M-Cl] +;

Anal. for ^C 37 ^H 37 ^N with ^Mn Cl ₂ : C, 48.11; H, 8.30; N, 15.59, Cl, 15.78. Found: C, 48.18; H, 8.32; N,

15.56, Cl, 15.75,

Example 5

A, Synthesis of Boc-DSer (OBzl) -OMe '

To Boc-DS.er (OBzl) -OH (15.0 g, 50.8 mmol) with ACN (250 mL) was added CS ₂ CO ₃ (33.0 g, 102 mmol) and methyl iodide (6.32 mL). (102 mmol) and the resulting mixture was then stirred at room temperature. half three hours. The reaction mixture was then filtered and concentrated. The rest was divided between. water ^(250: -'ml) and ethyl acetate (250 mL). The ethyl acetate layer was dried (MgSO ₄ ), filtered and concentrated to give 15.0 g (95% yield) of the desired methyl ester as a white solid: 1 H NMR (DMSO-d 6) δ 1.40 (s, 9H) ), 3, .64 (s, 3Η),

3.58-3.73 (m, 2 Η), 4.32 (bg, J = 6.2 Hz, 1 Η), 4.49. (s, 2H), 7.16 (bd, J = 6.2 Hz, 1H), 7.25-7.38 (m, 5H); MSIHRFAB, NBA-Li) m / z = 316.1768. [M + Li] +; 316.1736 calcd for C _i6 H ₂₃ O ₅ NLI.

B. Synthesis of DSer (OBzl) -OMe »TFA

Boc-DSer (OBzl) -OMe (15.0 g, 48.6 mmol) was dissolved in methylene chloride (125 mL) and treated with TFA (32 mL) .The resulting mixture was stirred for 30 min at room temperature and concentrated. The oil was triturated with ether (400 mL) to remove residual TFA to afford 15.9 g of {> 100% yield, contains a small amount of residual TFA) of the desired TFA salt as a colorless oil: ^R H NMR _(DMSO-dg) δ 3.72 (s, 3 Η), 3.73 - 3.83 (m, 2 Η), 4.36 (bt, J = 3.3 Hz, 1 Η),

5.50 (AB q, J = 12 Hz, \ _v = 24 Hz 2 Η), 7.27 - 7.37 (m, 5 H), 8.72 (bs, 3H); MSfHRFAB, NBA-HCl) 210.1159 [Μ + H] +; 210.1130 calculated for

C. Synthesis of Boc-Ser (OBzl) -DSer (QBzl) -OMe

Boc-Ser (OBzl) -OH (14.5 g, 49.2 mmol) was dissolved in DMF (550 mL) and treated with HOBT H 2 O (9.03 g, 59.0 mmol) followed by EDC HCl (11.3 g, 59.0 mmol). The resulting solution was stirred for 20 minutes at room temperature and treated with DSer (OBzl} -OMe 'TFA (15.9 g, 49.2 mmol) and TEA (8.22 mL, 59 mmol) and then for 12 hours The DMF was evaporated and the residue was taken up in ethyl acetate (300 mL) and washed with 1 N sodium hydrogen sulfate (100 mL), water (100 mL), saturated sodium bicarbonate (100 mL), brine (100 mL) and dried. (MgSO₄). Filtration and concentration afforded 20.8 g (87% yield) of the desired dipeptide as a white solid: ^X H NMR _(DMSO-dg) δ 1.37 (s, 9 Η), 3.57 - 3 , 75 (m, 4 Η), 3.62 (s, 3 Η), 4.35 - 4.50 (m, 1 Η), 4.46 (s, 3 H) 4.57 - 4.63 ( m, 1 Η), 6.88 (d, J = 8.1 Hz, 1 H), 7.20-7.34 (m, 10 Η), 8.34 (d, J = 8.1 Hz, MS (HRFAB, NBA-Li) m / z = 493.2559 [M + Li] +; 493.2526 calcd for C ₂₆ H ₃₄ N ₂ O _1.7 Li.

D. Synthesis of Ser (QBzl) -DSer (OBzl) -OMe TFA

Boc-Ser (OBzl) -D-Ser (OBzl) -OMe (20.4 g, 42.0 mmol) was dissolved in methylene chloride (170 mL) and TFA (43 mL) was added. The resulting mixture was stirred for 30 min at room temperature and concentrated. The residue was triturated with ether (400 mL) to remove excess TFA to give 22.7 g (> 100% yield, containing excess TFA) of the desired TFA salt: ¹ H NMR (DMSO-d 3 O 3, 59-3, 64 (m, 1H), 3.67 (s, 3H), 3.7.0

3.85 (m, 3H), 4.23 (bs, 1H), 4.40-4.58 (m, 4H).

- 4.70 (m, ΊΗ), 7.20-7.39 (m, 10H), 8.41 (bs, 3H),

9.09 (d, J = 7.81 Hz, 1H); MS (HRFAB, NBA-HCl) m / z =

387.1927 [Μ + H] *; 387.1920 calcd for C 11 H NO.

.E. Synthesis of Boc-Dser (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe

To a solution of Boc-DSer (OBzl) -OH in DMF (480 mL) was added

HOBT H ₂ O (7.96 g, 52.0 mmol) followed by EDOHCl (9.97 g, 52.0 mmol) and the resulting solution was stirred for 20 minutes at room temperature. To this solution was added Ser (OBzl) -DSer (OBzl) OMeTFA (21.7 g, 43.3 mmol) and TEA (7.25 mL, 52.0 mmol), and the resulting mixture was then stirred for 16 hours. stirred. The DMF was evaporated and the residue was partitioned between water (100 mL) and ethyl acetate (300 mL). The ethyl acetate solution was washed with 1 N hydrogen chloride with sodium Iran (150 mL), water (150 mL), saturated sodium bicarbonate (150 mL) and brine (150 mL), dried (Na ₂ SO ₄ ), filtered and concentrated to afford 26 6 g (93% yield) of the desired tripeptide as a white foam: · NMR (DMSO-d6) 1.39 (s, 9H), 3.56

3.74 (complex m, 6H), 3.64. (s, 3H). 4.35-4.43 (m, 1H)

H), 4.43-4.50 (m, 6 H), 4.63 (m, 1H), 4.73 tm, 1H), 6.94 (d, J = 7.8 Hz, 1 H) H), 7.24-7.36 (m, 15H), 8.13 (d, J = 8.2 Hz, 1 H), 8.51 (d, J = 7.8 Hz, 1H) ; MS (HRFAB, NBA-Li) m / z = 670.3326 (M + Li); 670.3316 calculated calc.

C Η N 0 Li.

45 3 9

F, Synthesis of DSer (OBzl) -Ser (QBzl) -DSer (OBzl) -QMe TFA salts

To a solution of Boc-DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe (26.4 g, 39.7 mmol) in methylene chloride (220 mL) was added TFA (55 mL) and the resulting mixture was stirred for 30 min at room temperature and concentrated. The 2-residue was triturated with ether (300 mL) and the ether triturate was decanted to give 26.2 g (97% yield) of the desired TFA salt as an orange oil after vacuum drying: ¹ H NMR (DMSO-d) δ 3.55 - 3.83 (complex m, 6 Η), 3.65 (s, 3 Η), 4.21 (bs,

Η), 4.40 - 4.58 (m, 6 Η), 4.60 - 4.67 (m, 1 Η), 4.85

- 4.94 (m, 1 Η), 7.20 - 7.40 (m, 15 Η), 8.37 (bs, 3 Η), 8.78 (d, J = 7.8 Hz, 1 Η) ), 8.84 (d, J = 8.3 Hz, 1H); MS (HRFAB, NBA-Li) m / z - 570.2790 [M + H] +; 570.2792 calcd.

G, Synthesis of Boc-Ser (OBzl) -DSer (OBzl) -Ser (QBzl) -DSer (OBzl) -OMe

To a solution of Boc-Ser (OBzl) -OH (11.2 g, 38.0 mmol) was added HOBT · H _z O (6.99 g, 45.6 mmol) followed by EDC HCl (8.74 g,

45.6 mmol) and the resulting solution was stirred for 30 minutes at room temperature. To this solution was then added DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe (25.8 g, 38.0 mmol) and TEA (6.36 mi, 45.6 mmol). The resulting solution was then allowed to stir for 16 hours. The DMF was evaporated and the residue was partitioned between water (200 mL) and ethyl acetate (400 mL). The ethyl acetate solution was washed with 1 N sodium bisulfate (200 mL), water (200 mL), saturated bicarbonate. bicarbonate (200 mL) a.solankou (200 mL), - dried (MgSO₄), filtered and concentrated to give 30.2 g (95% yield) of the desired tetrapeptide as a white foam: ^X H NMR (DMSO-d _g ) δ 1.38 (s, 9 Η),

3.53 3.76 (complex m, 8 ohms), 3.64 (s, 3 ohms), 4.34-4.42 (m,

Η), 4.40 - 4.51 (m, 8 Η), 4.58 - 4.66 (m, 1 Η), 4.67

- 4.77 (m, 1 Η), 4.74 - 4.82 (apparent q, J = 7.8 Hz, 1 Η),

6.89 (d, J = 8.2 Hz, 1 H), 7.22-7.38 (m, 20 H), 8.14 (d, J = 7.6 Hz, 1 H), 8 34 (d, J = 7.8 Hz, 1H), 8.55 (d,

J = 7.8 Hz, 1H); MS (HRFAB, NBA-Li) m / z = 847.4095 [M + Li] +; 847.4106 calcd for C H _{4 S ss} N ₄ O _ii Li.

H, Synthesis of Ser (OBzl) -bSer (OBZI) -Ser (QBzl) -DSer (QBzl) -OMe «TFA

Boc-Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe (30.2 'g, 35.9 mmol) was dissolved in methylene chloride (250 mL) and treated with TFA (63 mL). The resulting mixture ^was: stirred po'30 min at room temperature and zkoiicentrována. Trituration with ether afforded 30.0 g (98% yield) of the desired TFA salt as an oily residue 'Ή NMR (DMSO-d _e) δ

3.50 - 3.85 (complex m, 8 Η), 3.65 (s, 3 Η), 4.21 (bs,

H), 4.40-4.55 (m, 8H), 4.58-4.66 (m, 1H), 4.80

- 4.95 (m, 2H), 7.20 - 7.40 (m, 20 ohms), 8.35 (bs, 3 ohms),

8.54 (d, J = 8.2 Hz, 1 H, 8.69 (d, J = 7.4 Hz, 1 H), 8.86 (d, J = 8.2 Hz, 1 H); MS (HRFAB, NBA-Li) m / z = 747.3590 [M + Li] +; 747.3581 calcd for C ₄₁ H 44 _{S 4} O ₉ Li.

I. Synthesis of Boc-Ser (QBzl) -Ser (QBzl) -DSer (OBzl) -Ser (ORzl) 89

DSer (OBzl) -QMe

To a solution of Boc-Ser (OBzl) -OH (10.1 g, 34.2 mmol) in DMF (380 mL) was added ΗΟΒΤΉ ^ Ο (6.28 g, 41.0 mmol) followed by EDC HCl (7, 86 g, 41.0 mmol) and the resulting mixture was stirred for 20 minutes at room temperature. To this solution was then added Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe TFA (29.2 g, 34.2 mmol) and TEA (5.71 mL, 41.0). mmol) and the reaction mixture was allowed to stir for 12 hours. The DMF was evaporated and the residue was partitioned between water (200 mL) and ethyl acetate (400 mL). The ethyl acetate layer was washed with 1 N sodium bisulfate (200 mL), water (200 mL), saturated sodium bicarbonate (200 mL), and brine (200 mL), dried (MgSO 4), filtered and concentrated to give 33.7 g (97% yield) of the desired pentapeptide as a white powder: ¹ H NMR (DMSO-d) δ 1.39 (s, 9 H), 3.55-3.77 (complex m, 10 H),

3.64 (s, 3H), 4.32-4.42 (m, 1H), 4.40-4.54 (m, 10H),

4.60-4.68 (m, 1H), 4.49-4.84 (m, 3H), 7.00 (d, J =

7.7 Hz, 1H), 7.20-7.40 (m, 25H), 8.11 (d, J = 7.4 Hz,

H), 8.30 (d, J = 7.8 Hz, 1H), 8.35 (d, J = 7.8 Hz, 1H),

8.58 (d, J = 7.8 Hz, 1H); MS (LRFAB, NBA-Li) m / z = 1024;

IM + Li]

J, Synthesis Ser (OBzl) -Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (QBzl) »HCl

Boc-Ser (OBzl) -Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -OMe (31.7 g, 31.1 mmol) was dissolved in acetic acid (760 mL) and was treated with concentrated HCl (190 mL). The resulting solution was then stirred for 12 hours at 90 ° C. Concentration gave 29.3 g (100%) of the desired, deprotected peptide salt as a white foam;

¹ H NMR (DMSO-d 6)? -d ₆ ) δ 3.55 - 3.90 (complex m, 10 Η), 4.20 (m, 1 Η), 4.38 -4.61 (m, 11H), 4.73 - 4.85 (m, 3 Η), 7.20- 7.40 (m, 25H) Η), 8.34 - 8.42 (m, 2H) , 8.49 (m, 4), 8.95 (d, J = 7.7) Hz, 1H); MS (LRFAB, NBA) - HC1) m / z = 905 [M &lt; + &gt; H] ’.

K, Synthesis of cyclo-rSer (OBzl) -Ser (OBzl) -DSer (OBzl) -Ser (OBzl) DSer (OBzl) -1

A solution of Ser (OBzl) -Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (OBzl) - HCl (29.4 g, 31.3 mmol) in anhydrous, degassed DMF (4000 mL) was cooled to -40 ° C and treated with DPPA (8.22 mL, 38.1 mmol) followed by TEA (4.53 mL,

32.5 mmol). The reaction mixture was stored at -20 ° C for 48 h and then for an additional 43 h at 0 ° C. During this time, the pH was maintained at probably 8 by periodic addition of TEA (measured by dropping the reaction mixture on moistened Hydrion paper). After this time, the reaction mixture was diluted with water (1000 mL) and stirred with the mixed ion exchange resin (1600 g) for 6 hours. The resin was filtered off and the filtrate was concentrated to a volume of probably 200 ml. The product was then treated by adding water (500 ml). The solid was filtered and washed with ether (250 ml) to afford after vacuum drying 20.5 g (74% yield) of the desired cyclic peptide as a white powder: NMR _(DMSO-d6, major conformer) ύ

3.50 - 3.83 (m complex, 11 H), 4.32 - 4.82 (m complex, 14 H), 7.18 - 7.38 (m, 25 Η), 7.56 (d, J = 6.8 Hz, 1 H), 7.94 (d, J = 8.7 Hz, 1 H), 8.50 (d, J = 6.0 Hz, 1 H), 8.78 (m 2 H); MS (LRFAB, NBA-Li) m / z = 893 [M + Li] +.

L, synthesis. 12S, 5R, 8S, 11R, 14S) foam (benzyloxymethyl) -1,

4, 7, 10, 13-Pentaazacyclopenadadane.

To a solution of cyclo- [Ser (OBzl) -Ser (OBzl) -DSer (OBzl) -Ser (OBzl) -DSer (OBzl) -] (7.00 g, 7.90 mmol) in THF (100 mL) was add aluminum hydride (10 mL of 1.0 M solution in THF, 100 mmol) for 5 minutes at room temperature. The resulting solution was then heated at reflux for 16 h. The mixture was cooled to probably -20 ° C and carefully quenched with.

% sodium sulfate solution (30 mL). The resulting mixture was concentrated to a white powder and this powder was completely dried by coevaporation with toluene (3 x 50 mL). The residue was triturated with ether (2 x 100 mL) and the combined trisurates were concentrated to give 6.1 g ( 95% yield) of the desired ligand as a yellow oil NMR (CDCl ₃ ) δ 1.95 (bs,

.5H), 2.30-3.60 (complex, m, 25H), 4.20-4.40 (m, 10H), 7.00-7.38 (m, 25H) ; MS (HRFAB, NBA-Li) m / z = 822.5174 [M + Li] +; 822.5146 calcd for C _Sa H _{6 S} n ₅ O _with Li.

M. Synthesis of TMangan (II) dichloro (2S, 5R, 8S, 11R, 14S) - oenta (hydroxymethyl) -1, 4, 7, 10, 13-pentazacylcyclopentane

To a solution of manganese chloride (221 mg, 1.76 mmol) in methanol (75 mL) was added 2S, 5R, 8S, 11R, 14S) -penta (benzyloxymethyl) -1,4,7,10,13 -pentaazacyclopentadecane (1S). (43 g, 1.75 mmol) and the reaction mixture was refluxed for 2 h and then stirred at room temperature for 12 h. The methanol solution was allowed to evaporate and the residual oil was redissolved in ethanol (10 mL) and water (1 mL) in a Parr bottle. This mixture was treated with 10% Pd on activated carbon (1 g) and then hydrogenolyzed at 60 ° C and 65 psi for 16 hours. Filtration, concentration and recrystallization from ethanol-ether gave 212 mg (29% yield) of the desired complex as a white solid; MS (HRFAB, NBA) m / z = 455.1718 [M-Cl] ^&lt; ⁺ &^gt;; 455.1707 calcd for C ₃₅ N _if _H _g MnCl.

Example 6

Relaxation measurements were performed in the complexes of the invention (Examples 2-5) and in the comparative complex (Example 1) and the results are shown in Table I.

Proton relaxation times (T) of the sample in 100 mM Hepes buffer, pH = 7.4 were determined at 40 ° C from a monoexponential curve obtained from inverse-recovery pulse sequences (180 ° - ^c - 90 °) on a Bruker PC 120 / 125/10 VTs NMR. The spectrometer was calibrated for each sample to ensure correct duration of 90 ⁰ and 180 ⁰ radio frequency pulses and adequate magnetic field strength, adapted to the operating frequency of the 20 MHz system. Relaxation (R) was obtained from the slope of the 1 / T curve plotted against the concentration of the paramagnetic compound.

The relaxation time (T) of each sample in 100 mM Hepes buffer, pH = 7.4, was measured at 40 ° C using

Purcel-Meiboom-Gill pulse sequence on the same Bruker instrument. Relaxivity (R) was obtained from 1 / T dependence on the paramagnetic concentration.

Table 1

Sample Relaxation, mM ^-1 sec “ ¹ R R 2 Example '1 (Comparative) 1.82 2.37 Example 2 2.09 2.49 Example 3 2.76 3.86 Example 4 2.10 2.61 Example 5 .3,48 5.93

The results shown in Table I demonstrate that the complexes of the present invention (Examples 2-5) improved relaxivity compared to the comparison complex (Example 1).

Example 7

The kinetic stability of the complexes of the invention (Examples 24) and the comparative complex (Example 1) was measured and the results are shown in Table II.

The kinetic stability constant (k _aie ) for each complex can be determined by observing the rate at which copper appears to replace manganese as the metal center of the manganese macrocyclic complexes at different pHs.

The Beckman DU-70 UV / VIS spectrometer was set to scan wavelengths from 700 to 200 nanometers. Complex concentrations were kept low enough to remain within the linearity limits of observable absorbances for DU-70, which are below 2.00.

Since copper has an observable absorption band within the experimental wavelength range and the binding energies are much higher than the manganese binding energies, the replacement of copper with manganese as the metal center is almost instantaneous.

Table II

Example #

(comparative)

Kinetic stability, k (NT sec ' ^T ) dis

2814

1375

17.7

31.5

ND ^{1 1} Not determined

The data in Table II show that the complexes of the invention (Examples 2-4) are substantially kinetically more stable than the comparative complex (Example 1),

Example 8

The oxidation potentials of the complexes of the invention (Examples 2-4) and the comparison complex (Example 1) were measured and the results are shown in Table III.

Cyclic voltammograms were performed in methanol containing 0.18 M tetrabutylammonium chloride under a nitrogen atmosphere using a glassy carbon electrode with a platinum reference electrode and a ferrocene internal standard.

Table III

Example #

(comparative) ¹ Not determined

Oxidation potential, E (volts) 0.70

0.75

0.85

0, 74

ND ¹

The data in Table III show that the divalent manganese complexes of the invention (Examples 2-4) are more oxidatively stable than the comparative magnesium complex (Example 1).

Example 9

The partition coefficient (log P) of the complexes of the invention (Examples 2-5) and the comparison complex (Example 1) was measured and the results are shown in Table

IV.

The partition coefficient was determined by measuring the manganese content in each of the buffered and octanol phases. About 5 mg of the sample was dissolved in 5 ml of a buffered solution saturated with octanol (10 mM Hepes / 150 mM NaCl, pH = 7.4). The sample solution was mixed with 5 ml buffer saturated octanol and shaken overnight. The combined solution was centrifuged to separate the two phases. 2 ml of the solution (in duplicate) were taken from each phase and digested with a mixture of acids. The prepared solutions then

were measured for manganese 2a Use ICP-AES, ICP-MS or both. Log P values were calculated as a ratio Mn (octane) / Mn (buffer).

Table IV Example # Partition coefficient, log P 1 (comparative) -2.9 2 -1.9 3 -1.1 4 -0.76 5 -3.2

Table IV Example # Partition coefficient, log P 1 (comparative) -2.9 2 -1.9 3 -1.1 4 -0.76 5 -3.2 ¹ Not specified

The data in Table IV show that the log P, i.e. lipophilia or biodistribution, of the complexes of the invention (Examples 2-5) can be controlled by controlling the substituents on the macrocycle.

Claims (14)

PATENT CLAIMS A magnetic resonance imaging method comprising: (a) administering to the human or non-human animal subject a contrast agent comprising a physiologically compatible complex represented by the formula: wherein R, R ', R _1, R' _FS r _2, r _2, r _3, R _'3, R _4, R' _4, R _5, R _5, R _6, R _'6, R y, R ' _? , R _g , R ' ₈ , R _g and R' ₉ are independently selected from hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkylalkyl, cycloalkylcycloalkyl, cycloalkenylalkyl, alkylcycloalkyl, alkenylcycloalkyl, alkylcycloalkenyl, alkenylcycloalkenyl, heterocycle, aryl and aralkyl groups and groups attached to α-carbon of α-amino acids; either R _x or R ' _x and R ₂ or R' ₂ , R ₃ or R ' ₃ and R ₄ or R' ₄ , R _g or R ' ₅ and R ₆ or R' _g , R _? or R ' ₇ and R _g or R' _g and R _g or R ' _g and R or R' together with the carbon atoms to which they are attached form independently saturated, partially saturated or unsaturated cycles having 3 to 20 carbon atoms; R and R ', and R' _1, R ₂ and R ' ₂ , R ₃ and R' ₃ , R ₄ and R ' ₄ , R ₅ and R' ₅ , R ₆ and R ' _g , R _? -98a R _'7, R g and R _a and R _g, and R' ₉ together with the carbon atoms to which they are attached independently form a saturated, partially saturated or unsaturated ring structure having 3 to 20 carbon atoms; or one of R, R ', R ₁ , R'-p R ₂ , R' ₂ , R g, R ' ₃ , R ₄ , R 4' ^E 5 'R 5' R g <R g 'R 7t Η' / ^R 12 ' ^R 13' ^{and R} 14 'together with another R, F, ¹ , R _x , R 4, R ₂ , R' ₂ , R3> R 3 'R4' ^R 4 ' ^R 5' ^R 5 'Rg1 ^Rg ' R7i ^R * 7 'Rg>R'g>Rg> ^R ' g from Rjo ' ^R11 ' ^r 12 ' ^R 13 ^{and R} 14' ^{where the bond} is attached to a different carbon or nitrogen atom in the macrocyclic ligand may be bonded to form a chain represented by the formula CH CH, x x H CH CH, h CH, h CH CH, ~ _T where w, x, y and z are independent of each other integers from 0 to 10, and M, L and J are independently selected from the group consisting of alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, alkaryl, alkheteroaryl, aza, amide, ammonium, thia, sulfonyl, sulfinyl, sulfonamide phosphonyl, phosphinyl, phosphino, phosphonium, keto, ester, carbamate, urea, thiocarbonyl, borate, borane, borase, silyl, siloxy, silaza, and combinations thereof; and R q, R ^r yr 12 ^'R 13 ^{and R} 14 3 ^are independently from each other selected from the group consisting of hydrogen, alkyl and alkyl substituted ^-οκ 15 "' ůOORgg, -C0NR ₁₅ R ₁₆ or POgH _2, wherein R ₅ and R ₁₆ are independently hydrogen or alkyl; and wherein at least two of R, R ', Rq / ^E 2 'R * 2' R3 'R'3' R41 R'4 'R5 f R'5' Rg 1 Rg 'R71 R * ₇ , Rg, R'g, Rg ^and R'g are other than hydrogen ; wherein X, Y and Z are independently selected ligands selected from the group consisting of halide, oxo, aquo, hydroxo, alcohol, phenol, dioxygen, peroxo, hydroperoxo, alkylperoxo, aryperoxo, ammonia, -99alkylamino, arylamino, heterocycloalkylamino, heterocycloarylamino, amine oxides, hydrazine, alkylhydrazine, arylhydrazine, nitrogen dioxide, cyanide, cyanate, thiocyanate, isocyanate, isothiocyanate, alkyl nitrile, arylnitrile, alkylizonitrile, arylisonitrile, nitrile, nitrite, nitrite, arylsulfonic acid, alkylsulfoxide, arylsulfoxide, alkylarylsulfoxide, alkylsulfenic acid, arylsulfenic acid, alkylsulfinic acid, arylsulfinic acid, alkyl thiolcarboxylic acid, arylthiolcarboxylic acid, alkylthiolthiocarboxylic acid, arylthiolthiocarboxylic acid, alkylthiolthiocarboxylic acid, alkylthiolthiocarboxylic acid, arylthiolthiocarboxylic acid, , arylthiourea, alkylarylthiourea, sulphate, sulphite, hydrosulphate, bisulphite, thiosulphate, thiosulphite, hydrosulphite (hydrosulphite), alkylphosphine, arylphosphine, alkylphosphine oxide, arylphosphine oxide, alkylarylphos finoxide, alkylphosphine sulfide, arylphosphine sulfide, alkylarylphosphine sulfide, alkylphosphonic acid, arylphosphonic acid, alkylphosphinic acid, arylphosphinic acid, alkylphosphinic acid, phosphate, thiophosphate, alkyl phosphate, pyrophosphate, triphosphate, di-phosphate, di-phosphate alkylaryl carbamates, alkylthiocarbamates, arylthiokarbamát, alkylarylthiokarbamát, alkyldithiocarbamates, aryldithiokarbamát, alkylaryldithiokarbamát, hydrohenuhličitan, carbonate, perchlorate, chlorate, chlorite, hypochlorite, perbromate, bromate, bromite, hypobromite, tetrahalomanganan, tetrafluoroborate 100ritan, hexafluorophosphate, hexafluoroantimonate, phosphorane, iodate, periodate, metaborate, tetraarylborate, tetraalkylborate, tartrate, salicylate, succinate, citrate, ascorbate, saccharinate, amino acid, hydroxamic acid, thiotosylate and ion exchange anions CH resins or their corresponding anions, or one or more groups of X, Y and Z are independently attached to one or more R groups, wherein n is an integer from 0 to 3; M is a paramagnetic metal selected from the group consisting of metals having atomic numbers 21-29, 42-44 and 57-71; and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle; and (b) producing a magnetic resonance image of at least a portion of said subject. The method of claim 1, wherein M is divalent Mn or trivalent Gd. The method of claim 2, wherein M is divalent Mn. The method of claim 1, wherein at least three of R, R ', ^R 11 ^R _V ^R 2 ^{of R} 2 ^R 3 ^R 3 ^R 3 ^R 4 ^R 4 ^R 5 ^R 5 ^R 6 'ό'' R _7, Rg, R θ ^1, R g and R _g, are other than hydrogen The process according to claim 1, wherein at least one of either R 1 or R ₁ and R ₂ or R ₂ , ^r 3 or R ₃ and R ₄ or R ₁₄ , R ₅ or R ₁₅ and R 8 or Rg, R ₇ or R _'7 and R _Q or R _g and R _g or R' ₉ and R or R 'together with the carbon atoms to which they are attached independently form a saturated cycle having 5 to 8 carbon atoms; and all of the remaining R groups are each other Independently selected from the following: hydrogen, alkyl, or alkyl substituted with -OR ₁₅ or -NR ₁₅ R ₁₆ , wherein R ₁₅ and R ₁₆ are independently hydrogen or alkyl. The method of claim 5, wherein said complex is represented by the formula: The method of claim 1, wherein at least two groups of R, R ', ^R ' 1 ' ^R 2' ^ * 2 '^ 3' ^R 3 ' ^R 4' ^ 4 '^ 5' ^ * 5 '^ 6' ^ 6 '^ 7' R _{7, and} R, R _', Rg and Rg are alkyl or alkyl substituted with -OR ₁₅ or -NR ₁₅ R _16, wherein R ₁₅ and R ₁₆ are independently hydrogen or alkyl. The method of claim 7, wherein said complex is represented by the formula: HIM' -1029. Delivery method representing: (a) administering to a human or non-human animal subject a substance representing a physiologically compatible complex represented by the formula: r ₅ r _s ^Ri2 K » R ' ₆ , FJ, R' ₇ , Rg, R ' _g , Rg and R' _g are independently selected from: hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkylalkyl, cycloalkylcycloalkyl, cycloalkenylalkyl, alkylcycloalkyl, alkenylcycloalkyl, alkylcycloalkenyl , alkenylcycloalkenyl, heterocycle, aryl and aralkyl groups and groups attached to α-carbon α-amino acids; either R 6 or R 7 and R ₂ or R ' ₂ , R ₃ or R' ₃ and R ₄ or R ' ₄ , R ₅ or R' ₅ and R ₆ or R ' ₆ , R ₇ or R' ₇ and R 8, or R ' _Q and R _g or R' _g and R or R 'together with the carbon atoms to which they are attached form independently saturated, partially saturated or unsaturated cycles having 3 to 20 carbon atoms; R and R ', R ^ and R ^, R ₂ and R' 2 / R 3 ^and R, R₁ and R _'4, R' 5 and Rg / Rg and Rg, ^ 7 and R _'7, R g and R _g and R g and R ' ₉ together with the carbon atoms to which they are attached independently form saturated, partially saturated or unsaturated crown structures having 3 to 20 carbon atoms; or one of R, R ', R 1, R 1, R ₂ , R' ₂ , R ₃ , R ' ₃ , R ₄ , -103- ^R '4' ^R 5 ' ^R * 5 ' ^R 6' ^R '6' ^R 7 ' ¹ R 7, R g, R _g, Eg i R'g, ^R 10 ' r _i2, r ₁₃ and Rj ₄ , together with others R, R ', ^R 'R', / ^ 2 ' ^R 3 ' ^R 3' R ₄ Z ^R, ₄ from R5 / K ' ₅ , ^R 6 ' ^R ' 6 ' ^R 7' Rg, Rg • ^R 's ^R 11 ' ^R 12' ^R 13 ^a R ₁₄ which is established on the carbon or nitrogen atom in the macrocyclic ligand may be bonded to form a chain represented by the formula -e ₂ HHT ch ch ch kl -e, -e j s t _r where w, x, y and z are each independently integers from 0 to 10 and M, L and J are independently selected from the group consisting of alkyl , alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, alkaryl, alkheteroaryl, aza, amide, ammonium, thia, sulfonyl, sulfinyl, sulfonamide, phosphonyl, phosphinyl, phosphino, phosphonium, keto, ester, carbamate, urea, thiocarbonyl, borate, borane , borase, silyl, siloxy, silaza, and combinations thereof; and ^R L0 'll- ^{R f R} l2 ^R 13 ^{and R} 14 3 ^are independently from each other selected from the group consisting of hydrogen, alkyl and substituted ^alkyl-0R 15' ^C00R i5 / ^-COÍÍR 15 ^R 16 ^no ^ ° P ° 3H ₂ wherein R ₁₆ and R ₁₆ are independently hydrogen or alkyl; and wherein at least two of R, R ', ^R 1' ^R 1 ^'R 2' ^R 2 ^'R 3' ^R '3' ^R 4 ^'R 4' ^R 5 ^{'R <5' R} 6 ^'R 6 ^R ^ 7 'R _{7, Rg,} Rg, _Rg and R ₉ are other than hydrogen; wherein X, Y and Z are independently selected ligands from the group consisting of halide, oxo, aquo, hydroxo, alcohol, phenol, dioxygen, peroxo, hydroperoxo, alkylperoxo, arylperoxo, ammonia, alkylamino, arylamino, heterocycloalkylamino, heterocycloarylamino, amine oxides, hydrazine , alkylhydrazine, arylhydrazine, nitrogen dioxide, cyanide, cyanate, thiocyanate, isocyanate, isothiocyanate, alkyl nitrile, arylnitrile, alkylisonltril, 104 arylisonitrile, nitrate, nitrite, azido, alkylsulfonic acid, arylsulfonic acid, alkylsulfoxide, arylsulfoxide, alkylarylsulfoxide, alkylsulfenic acid, arylsulfenic acid, alkylsulfinic acid, arylsulfinic acid, alkylthiol carboxylic acid, arylthiol carboxylic acid, arylthiol carboxylic acid, arylthiol carboxylic acid, arylthiol carboxylic acid, arylthiol carboxylic acid , urea, alkyl urea, aryl urea, alkylaryl urea, thiourea, alkylthiourea, arylthiourea, alkylarylthiourea, sulphate, sulfite, hydrosulfite, hydrosulfite, thiosulfate, thiosulfite, alkylsulfosulfide, alkylphosphine, alkylphosphine, arylphosphine, alkylphosphine, arylphosphine, arylphosphine alkylphosphonic acid, arylphosphonic acid, alkylphosphinic acid, arylphosphinic acid, alkylphosphinic acid, arylphosphinic acid, phosphate, thiophosphate, phosphite, pyrophosphite, triphosphate, hydrogen phosphate, dihydrogen phosphate, alkylguanidino, arylguanidino, alkylarylguanidino, alkylcarbamate, arylcarbamate, alkyllarylcarbamate, alkylthiocarbamate, alkylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate, alkyllarylthiocarbamate; halate, chlorate, hypochlorite, hypochlorite, bromate, bromate, bromate, bromate, tetrahalomanganate, tetrafluoroborate, hexafluorophosphate, hexafluoroantimonate, phosphate, iodate, periodate, metaborate, tetraarylborate, tetraalkylborate, tartrate, succinate, siccate, siccate, siccate, 105 the hydroxamic acid, thiotosylate and ion exchange resin anions or their corresponding anions, or one or more of X, Y and Z is independently bonded to one or more R groups wherein n is an integer from 0 to 3; M is a heavy metal selected from the group consisting of metals having atomic numbers 20-32, 42-44, and 57-83; and a non-toxic, pharmaceutically acceptable carrier, adjuvant or vehicle; and (b) forming an X-ray, ultrasound or scintigraphic image of at least a portion of said subject. The method of claim 9, wherein M is a radioactive metal isotope selected from the group consisting of ^99m Tc and ^{11: L} In and said image is a scintigraphic image. The method of claim 9, wherein at least three of R, R ', R, R ¹ , R, R', R, R ', R, R', R, R ', R, R', R, XX 2 2 3 3 ¹ 4 4 S 5 6 β 7 ¹ R ', R, R', R and R 'are other than hydrogen. YS Θ 3 9 ^JJ (2) n where R, R ', R, R', R, R ', R, R', R, R ', R, R', R ', _χ , x' 2 3 ' 3 4 4 3 R, R _'and R ^ to R _g are independently selected from 106 groups: hydrogen, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkylalkyl, cycloalkylcycloalkyl, cycloalkenylalkyl, alkylcycloalkyl, alkenylcycloalkyl, alkylcycloalkenyl, alkenylcycloalkenyl, heterocycle, aryl and aralkyl groups and α-amino-linked groups; either R or R ¹ and R or R ', R or R' and R or R ', R or R' and R 'or R', R or R 'and R or R' and R or R 'and R or R together with the carbon atoms to which they are attached form independently saturated, partially saturated or unsaturated cycles having 3 to 20 carbon atoms; R and R ', R and R', R and R ', R and R', R and R ', R and R', R and R ^1, R and R _'7, R _g and R' and R _g and R 1 together with the carbon atoms to which they are attached independently form saturated, partially saturated or unsaturated ring structures having 3 to 20 carbon atoms; or one of R, R ', R ^, R ^, _R2, R' _2, R _g, R _'3, R₄, R ', R, R', R, R ', R, R', R, R ', R, R', R, R, ^{R 12} ' ^R 13 ^{and R} m' together with another R, R ', R 1 (R 1 = R ₂ , R ' ₂ , R. R ', R, R', R, R ', R, R', R, R ', R, R ^1, R R ', R, R, R', R ', and R', which is attached to a different carbon or nitrogen atom in the macrocyclic ligand, may be bonded to form a bridging represented by the formula ^: CH, d ₇ wherein w, x, y and z are each independently integers from 0 to 10 and M, L and J are independently selected from the group zai hrnující alkyl, alkenyl, alkynyl, aryl, cycloalkyl, heteroaryl, alkaryl, alkheteroaryl, aza, amide, ammonium, thia, sulfonyl, sulfinyl, sulfonamide, phosphonyl, phosphinyl, phosphino, 107 phosphonium, keto, ester, carbamate, urea, thiocarbonyl, borate, borane, borase, silyl, siloxy, silaza, and combinations thereof; and R _io, R _XI, R _i2, R _x3 and R _x4 are each independently selected from hydrogen, alkyl and alkyl substituted with -OR, -COOR, -CONR R or PO Η wherein R and R are wearing independently hydrogen or alkyl; and wherein at least two of R, R ', R, R *, R, R', R, R ¹ R, R ', R, R', R, R ', R, R' ₇ , R _e , R ' _e, R _s and R ₉ are other than hydrogen; wherein X, Y and Z are independently selected ligands from the group consisting of halide, oxo, aquo, hydroxo, alcohol, phenol, dioxygen, peroxo, hydroperoxo, alkylperoxo, aryperoxo, ammonia, alkylamino, arylamino, heterocycloalkyl amino, heterocycloarylamino, amine oxides, hydrazine, alkyl hydrazine, arylhydrazine, nitrogen dioxide, cyanide, cyanate, thiocyanate, isocyanate, isothiocyanate, alkyl nitrile, aryl nitrile, alkyl isonitrile, aryl isonitrile, nitrate, nitrite, azido, alkylsulfonic acid, arylsulfonic acid, alkylsulfoxide, arylsulfoxide, arylsulfoxide aryl sulfoxide, alkylsulfenic acid, arylsulfenic acid, alkylsulfinic acid, arylsulfinic acid, alkylthiol carboxylic acid, arylthiolcarboxylic acid, alkylthiolthiocarboxylic acid, arylthiolthiocarboxylic acid, alkylcarboxylic acid, arylcarboxylic acid, alkyl urea, alkylurea, arylurea, arylurea, arylurea, arylurea, arylurea na, sulfate, sulphite, hydrosulfate, hydrosulfite, thiosulfate, thiosulfite, hydrosulfite (hydrosulfite), alkylphosphine, arylphosphine, alkylphosphine oxide, arylphosphine oxide, alkylarylphosphine oxide, alkylphosphine108 sulfide, arylphosphine sulfide, alkylphosphine sulfonic acid, alkylphosphine phosphine sulfide, alkylphosphine sulfonic acid, acid, aryl phosphinite acids, phosphates, thiophosphates, phosphites, pyrofosfit, triphosphate, hydrogen phosphate, dihydrogen phosphate, alkylquanidine, arylguanidino, alkylarylguanidino, alkylcarbamate, arylcarbamates, alkylaryl carbamates, alkylthiocarbamates, arylthiokarbamát, alkylarylthiokarbamát, alkyldithiocarbamates, aryldithiokarbamát, alkylaryldithiokarbamát, hydro ěňuhličitan, carbonate, perchlorate , chlorate, chlorite, hypochlorite, hypochlorite, bromate, bromite, bromate, tetrahalomanganate, tetrafluoroborate, hexafluorophosphate, hexafluoroantimonate, hypophosphite, iodate, periodate , metaborate, tetraarylborate, tetraalkylborate, tartrate, salicylate, succinate, citrate, ascorbate, saccharinate, amino acid, hydroxamic acid, thiotosylate and ion exchange resin anions or their corresponding anions, or one or more of X, Y and Z are independently bonded to one or more R groups wherein n is an integer from 0 to 3; M is a paramagnetic metal selected from the group consisting of metals having atomic numbers 21-29, 42-44 and 57-71; NEJM and one group of R _io, R _{xi, i =} R, R _i3 R and _X4 is other than hydrogen. The compound of claim 12, wherein M is divalent Mn or trivalent Gd. A compound according to claim 12, wherein at least three of R 1, R ', _R', R · R _=, R _{a, R,} R 'RR' RR 'RR · 109 R ₇ , R ' ₇ , R _a , R' _e , R ₉ and R ' _s are other than hydrogen. A compound according to claim 12, wherein at least one of either R or R 'and R or R ¹ , R or R' and R or R ^f , R or R 'and R or R', R or R 'and R or R 'and R or R' _g and R or R 'together with the carbon atoms to which they are attached independently form a saturated cycle having 5 to 8 carbon atoms; and all of the remaining R groups are independently selected from: hydrogen, alkyl, or alkyl substituted _with -OR or -NR _L5 R _Ie wherein R _is _ and R are independently hydrogen or alkyl. The compound of claim 12 wherein at least two of R, R ', R', R ', R', R ', R' R, R ', R, R', R and R bound by -OR or -NR R ^{J '} X 5 16' R ¹ R ¹ R ' R ₉ are alkyl or alkyl substituokde R _i5 and R _IE are independently hydrogen or alkyl.