Derivatives bicyclopentadiene acids or their pharmaceutically acceptable salts and method of production thereof

 

(57) Abstract:

Usage: as contrast agents in magnetic resonance imaging. Products: derivatives bicyclopentadiene acids of the formula I, where R is the group - (C)n(x) (y) - T, x, and y is H or COOH, n is an integer 1, 2 or 3, T - COOH, - C6H4R4or P (O) (R1) OH, R1- OH, or-O-(C1- C5)alkyl, R4- NO2or NH2provided that one T must be a group P(O) (R1)OH, or their pharmaceutically acceptable salts. Reagent 1: compound of formula 1, in which at least one group R is a hydrogen. Reagent 2: phosphorous acid or its ester. Reaction conditions: in the midst of a solvent in the presence of formaldehyde. 2 C. and 18 h.p. f-crystals, 4 PL.

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the compound of formula I.

The invention relates to ligands, which are bicyclopentadiene acids, and their complexes and conjugates for use as contrast agents in magnetic resonance imaging (MRI). Some of the ligands and complexes are also useful as a means of caring for the oral cavity and as agents for the inhibition of scale formation in water systems processing. For best p which is a non-invasive technique for diagnosis, which gives a well-resolved image of the cross section of the soft tissues of the animal body, preferably, human. This technique is based on the property of certain atomic nuclei (e.g. protons of water), which possess a magnetic moment (as defined using mathematical equations: see G. M. Barrow, physical Chemistry, 3rd edition, Mc Graw-Hill, HY (1973)), to be installed in one line (centered) in a magnetic field. After centering the equilibrium state can deviate from the norm when using an external radio frequency (RF) pulse, which causes the protons to avoid alignment of the magnetic field. When the RF pulse is terminated (end), the nuclei return to their equilibrium state, and the time required for this to have occurred, known as the relaxation time. The relaxation time consists of two parameters, known as spirestone (TI) and pinpinela (T2), relaxation, and these relaxation measurements give information on the degree of molecular organization and interaction of protons with the environment.

Since the content of water in living tissue is essential, and there are varying content and environment among tissue types, get the promotion. The larger the differences in the relaxation time (T1 and T2) of the protons present in the scanned tissue, the greater the contrast of the output image (J. Magnetic Resonance, 33, 83-106 (1979)).

It is known that paramagnetic chelates with symmetric e-the main condition can have a dramatic effect on the relaxation rate T1 and T2 are superimposed on each water protons, and that the effectiveness of the chelate in this respect lies partly with the number of unpaired electrons, which gives the magnetic moment (Magnetic Resonance Annual, 231-266, Raven Press, NY (1985). It was also shown that, when a paramagnetic chelate of this type is introduced into the animal organism, its effect on T1 and T2 of different tissues can be viewed directly on magnetic resonance (MR) images with increased contrast is observed in the region of localization of the chelate. Therefore, to increase the diagnostic information obtained with the aid of MRI, it was proposed to introduce an animal stable nontoxic paramagnetic chelates (Frontiers of Biol. Energetics, 1, 752-759 (1978); J. Nucl. Med., 25, 506-513 (1984); Proc. of NMR Imaqinq Symp. (Oct. 26-27, 1980); F. A. Cotton et al. , Adv. Inorg. Chem., 634-639, 1966). Paramagnetic chelates metals used this way are called agents of contrast enhancement or contrast agents.

+3), iron (Fe+3), manganese (Mn+2and (Mn+3) and chromium (Cr+3because these ions have the greatest impact on water protons due to their large magnetic moments. In the uncomplexed form (for example, GdCl3) ions of these metals are toxic to animals, causing prevented their use in the form of a simple salt. Therefore, the main role of the organic chelating agent (also called a ligand) is to make the paramagnetic metal is non-toxic to the animal while maintaining its desirable effect on the relaxation rate T1 and T2 of the surrounding water protons.

Knowledge and experience in the field of MRI are very extensive, so the following brief description, without intending to be exhaustive, is given only as an overview of this area and other compounds, which are probably similar in structure. In U.S. patent 4899755 disclosed rotate (shift) time proton NMR relaxation in the liver or bile ducts animals using Fe+3-ethylene-bis-(2-hydroxyphenylglycine) complexes and their deposition is inkarbaeva acid. In U.S. patent 4880008 (application with partial continuation of U.S. patent 4899755) revealed additional data visualization of the liver tissue of rats, but not shown any additional complexes. U.S. patent 4980148 reveals gadolinium complexes for MRI, which are acyclic compounds (C. J. Broan et al., J. Chem. Soc., Chem. Commun., 1739-1741, 1990) describe some bifunctional macrocyclic Postanovlenie compounds (C. J. Broan etc., J. Chem. Soc., Chem. Commun., 1738-1739, 1990) describe compounds that are trisalicylate (I. K. Adzamli etc., J. Med. Chem., 32, 139-144, 1989) describe acyclic phosphonate derivatives of gadolinium complexes for NMR imaging.

Currently the only industrial contrast agent available in the United States, is a complex of gadolinium with diethylenetriaminepentaacetic acid (TRA-Gd3+- MAGNEVISTTMcompany Schering AG). MAGNEVISTTMit is considered non-specific perfusion agent, because it is freely distributed in extracellular fluid with subsequent effective output through the renal system. MAGNEVISTTMproved valuable in the diagnosis of brain damage, because accompanying the gap gematogenny to MAGNEVISTTM'the Guerbet produces industrial for sale macrocyclic perfusion agent (DOTAREMTM), which is currently available only in Europe. In various stages of development there are a number of other potential contrast agents.

Now unexpectedly been found that contrast agents can be different bicyclopentadiene ligands. In addition, these ligands can modify their charge, due to the structure of the ligand and selected metal, which may affect their ability to be more specific to a particular site (site). In particular, the invention is directed to new ligands, which are bicyclopentadiene the compounds of formula:

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where

R is

< / BR>
where

X and Y represent independently H, OH, (1 - 3)alkyl or COOH;

n is an integer 1, 2 or 3;

provided that: when n is 2, then the sum of X and Y must be equal to or more than H; and when n is 3, then the sum of X and Y must be equal to three or more H;

T represents H, (1 - 18)C alkyl, COOH, OH, SO3H,

< / BR>
where

R1is OH, (1 - 5) C-alkyl or-O- (1 - 5) C Alki is about, bromoacetamide or carboxyl;

R2represents H or OH; provided that when R2is OH, then R containing R2must be all of X and Y is H; provided that at least one T must present P/O/R1OH, and provided that when one T represents a group

< / BR>
then one radical X or Y this R radical can represent COOH, and all other X and Y given R radical should represent H;

A represents CH, N, C-Br, C-Cl, C-OR3C-OR8N+-R5X,

< / BR>
R3is H, (1 - 5)C alkyl, benzyl, or benzyl substituted by at least one of R4;

R4has the values defined above;

R5is (1 - 16)C alkyl, benzyl, or benzyl substituted by at least one of R4;

R8is (1 - 16)C alkylamino;

X-represents Cl-, Br-l-or H3CO.-2;

Q and Z independently represent CH, N, N+-R5X, C-CH2-OR3or C-C(O)-R6;

R5has the values defined above;

R6represents-O-(1 - 3)alkyl, OH or other7;

R7is (1 - 5)C alkyl or biologically active material;

X-the ima is A or Z is N or N+-R5X, then the other two groups should represent CH;

b) when A is C-Br, C-Cl, C-OR3or C-OR8then Q and Z must be CH;

c) the amount of the radicals R4, R7and R8when they are present, may not exceed one; and

d) only one of Q or Z may be C-C (O)-R6and when one of Q or Z is C-C (O)-R6then A must be CH.

When the above ligands of formula 1 have at least two of the R radicals T is equal to PO3H2[P(O)R1OH, where R1is OH, and the third T is equal to H, COOH or (1 - 18)C alkyl; A, Q and Z are CH; n is 1; and X and Y independently represent H or (1 - 3)C alkyl; then the ligands useful for the care of the oral cavity. Especially preferred are those compounds in which the three R radical T is P(O)R1OH, where R1is OH, n is 1; and X and Y are H. the Use of these ligands is discussed and in other States that are together on the consideration of applications.

When the above ligands of formula 1 are:

the R radical of at least two T is P(O)R1OH, where R1is OH and the other R radical, T is COOH or P/O/ R1OH, and n, R1X, Y, A, Q and Z have ukazannye, and in the other two R radicals T represents COOH or P(O)R1OH, and n, R1X, Y, A, Q and Z have the above meanings; or

in the R radical is three T are P(O)R1OH, where R1is (1 - 5)C alkyl or-O-(1 - 5)C alkyl, and n, R1X, Y, A, Q and Z have the meanings given above; then the ligands are useful as contrast agents.

Especially preferred are those compounds of formula 1,

where X and Y are H; n is 1; or

A, Q and Z are CH.

Bifunctional ligands of formula 1 is desirable to obtain the conjugates of the present invention. Such ligands should have:

one R radical, where T is the fragment is

< / BR>
where

R2and R4have the meanings given above, especially when the two R radicals, not containing R4radical, both radical T is P/O/R1OH, where R1has the values defined above, or where two R radicals, not containing R4radical, one T represents COOH, and the other T radical is P(O)R1OH, where R1has the values defined above, preferably the fragment above T radical, where one of X and Y this radical represents COOH; and

preference is dstanley C-OR3or C-OR8where and R3and R8have the meanings given above, or

< / BR>
where

R4has the values defined above; or

A is CH, and one of Q or Z is CH and the other is C-C//-R6where R6has the values defined above;

especially those compounds in which R6is other7where R7represents the biologically active material.

The ligands of formula I can be complex with various metal ions, such as gadolinium /Gd+3/ iron /Fe+3/ manganese /Mn+2/, moreover, it is preferable Gd+3. Complexes formed in this way can be used by themselves or can join via covalent bonds to larger molecules, such as dextran, polypeptide or biologically active molecules, including antibodies or fragments thereof, and used for diagnostic purposes. Such conjugates and complexes are useful as contrast agents.

Complexes and conjugates of the invention may be constructed to provide specific General charge, which has been successful in influencing the in vivo biologicaly and contrast of the image is always

in the three radicals R T is P(O)R1OH, where R1is OH, and n is 1; or

two R radicals T is P(O)R1OH, where R1is OH, in the third R-radical T is COOH and n is 1; or

two R radicals T is P(O)R1OH, where R1is OH, in the third R-radical T is P(O)R1OH, where R1is (1-5)C alkyl, and n is 1; or

two R radical T is P(O) R1OH, where R1is OH, in the third R-radical T is P(O)R1OH, where R1represents-O-(1-5)C alkyl, and n is 1; or

(B) the Total charge of -1 when

in one R radical T is P(O)R1OH, where R1is OH and the other two R radicals T is P(O)R1OH, where R1represents-O-(1-5)C alkyl, and n is 1; or

in one R radical T is P(O)R1OH, where R1is OH, and in the other two R radicals T is P(O)R1OH, where R1is (1-5)C alkyl, and n is 1; or

in one R radical T is P(O) R1OH, where R1is OH, and the other two R radicals T represents COOH, and n is 1; or

(C) overall neutral charge - when

in the three R radicals T depict what is P(O)R1OH, where R1is (1-5)C alkyl, and n is 1; or

(D) the total charge of +1 when

one of the A, Q or Z is N+-R5X-where R5and X-have the meanings defined above; and one R radical T fragment is P(O)R1OH, where R1is (1-5)C alkyl or-O-/(1-5)C alkyl/; in the other two R fragment T represents COOH or P(O) R1OH, where R1is (1-5)C alkyl, -O-((1-5)C alkyl); and X and Y radicals are h

As the complexes and conjugates can be formulated in pharmaceutically acceptable form for the appointment of an animal.

Perhaps the use of ligands of the formula I with ions of other metals for the diagnosis of pathological States such as cancer. The use of these complexes and conjugates discussed in another application, are jointly considered.

The compounds of formula I are numbered for nomenclatural purposes as follows:

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One aspect of the invention concerns the development of contrast agents with synthetic modification paramagnetic chelate, providing the opportunity for site-specific delivery of a contrast agent into the desired tissue. Moreover, the advantage is a false contrast, resulting from non-specific perfusion, which may or may not be visible with extracellular agents. The specificity of the ligand of formula I can be regulated by adjusting the total charge and lipophilic nature of the complex. The total interval of the charge of the complex is from -3 to +1. For example, for a complex having 2 or more groups of PO3H2total charge is highly negative and expected uptake by bone; whereas, when the overall charge of the complex is O (is thus neutral) complex may have the ability to cross the blood-brain barrier and may be possible absorption by the brain.

Tissue specificity may also be realized by using ionic or covalent joining of the chelate to a naturally occurring or synthetic molecule having specificity against the desired target tissue. One of the possible applications of this approach is to use paired with a chelate of a monoclonal antibody which is transported paramagnetic chelate to the diseased tissue, allowing visualization using MRI. In addition, the accession paramagnetic Hel to improved contrast relative to unrelated chelate. Recent work of the author Lauffer (U.S. patent 4880008 and 4899755) demonstrated that variations of the liquid can result in specific tissues agents, and that increased lipophilic character of favorable non-covalent interaction with blood proteins, resulting in increased relaxivity.

In addition to this, these contrast agents of formula I, which are neutral in charge, especially preferred for the formation of the conjugates of the present invention, as reduced to a minimum the undesirable ionic interactions between chelate and protein, which retains immunoreactivities antibodies. In addition, these neutral complexes reduce the osmotic pressure relative to DTRA-Gd+3that may mitigate or eliminate the discomfort from the injection.

Not wishing to be bound by any theory, believe that when I charged complex of the invention (for example, maybe -2 or -3 for bones, -1 for the liver, or +1 for the heart), the variation of the ionic charge of the chelate may affect biologicaly. Thus, if the antibody or other guide fragment is also specific to the same site or sitzposition in the definition of formula I, additionally are defined as follows, (1-3 C alkyl, (1-5)alkyl, (1-18)C alkyl includes alkyl groups, as straight and branched chain. Animal includes a warm-blooded mammal, preferably human.

Biologically active material refers to the dextrans, peptides or molecules that have a specific affinity for a receptor, or preferably, antibodies or fragments of antibodies.

Antibody refers to any polyclonal, monoclonal, chimeric antibody or heterodontidae, preferably by monoclonal antibody; "antibody fragments include Fab fragments and F/ab'/2fragments, and any part of the antibody having specificity against the desired epitope or epitopes. When you use the term conjugate chelate radioactive metal/antibody or conjugate, we mean that the antibody includes whole antibodies and/or fragments of antibodies, including semi-synthetic variants or variants, obtained by the methods of genetic engineering. Possible antibodies are 1116-NS-19-9 (anti-colorectal carcinoma), 1116-NS-3d (anti-CEA), 703D4 (anti-human lung cancer), 704AI (anti-human lung cancer), CC49 (anti-TAG-72), CC83 (anti-TAG-72) and B72.3. under the Deposit numbers ATCC HB 8059, ATCC CRL-8019, ATCC HB 8301, ATCC HB 8302, ATCC HB 9459, ATCC HB 9453 and ATCC HB 8108, respectively.

In the sense as it is used here, the term complex refers to the complex of the compounds of formula I, associated in complex or complex with a metal ion, where the at least one metal atom is chelated or bound in a chelate complex; conjugate refers to the chelate ion bromide, which is covalently attached to the antibody or antibody fragment. Terms bifunctional coordinator, bifunctional chelating or chelate forming agent and functionalized chelating agent are used interchangeably and refer to compounds that have chelating fragment, capable of forming a chelate complex of a metal ion and fragment covalently linked to a chelating fragment that is able to serve as a means of covalent joining the antibody or antibody fragment.

Bifunctional chelating agents described herein (represented by formula 1) can be used to form a chelate or bind to chelate metal ions, forming chelates with metal ions (called here also complexes). The complexes, due to the presence of options is to join a biologically active materials, such as dextran, molecules that have a specific affinity for receptors or preferably covalently join the antibodies or fragments of antibodies. Thus, the complexes described herein, can covalently join the antibodies or fragments of antibodies or to have specific affinity to receptors and are referred to here as the conjugates.

In the same sense as it is used in this description, the term "pharmaceutically acceptable salt" means any salt or mixture of salts of the compounds of formula 1, which are sufficiently non-toxic to be useful in the treatment or diagnosis of animals, preferably mammals. Thus, according to this invention are useful salts. Typical representatives of these salts obtained or produced using standard reactions with both organic and inorganic sources include, for example, salts of sulfuric, hydrochloric, phosphoric, acetic, succinic, citric, lactic, Malinova, fumaric, palmitic, holeva, pambou, mucus, glutamine, gluconic acid, o-camphoric, glutaric, glycolic, phthalic, tartaric, formic, lauric, stearic, salicylic, metasolv is evidence of salt formed using standard reactions from organic or inorganic sources, such as ammonium or 1-deoxy-1-/methylamine/D glucitol, alkali metal ions, alkaline earth metal ions and other ions. Especially preferred are salts of compounds of formula 1, when salt is a salt of potassium, sodium, ammonium. Also includes mixtures of the above salts.

The compounds of formula 1 are obtained using different processes. Typical General synthetic path or approach to such processes is illustrated by means of diagrams of the reactions below.

According to scheme 1 (see end of text) are compounds of formula 1, where X and y = H, n = 1 (but would also apply if n = 2 or 3, with appropriate replacement of the reagent), T = PO3H2and Q, A and Z = CH.

According to scheme 2 (see end of text) are compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate substitution of reagents), T =

< / BR>
where

R1= -O-(1-5)C alkyl; and Q, A and Z = CH.

Scheme 3 (see end of text) are compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable when n = 2 or 3, with a corresponding change in the reagent), T =

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dedimania formula 1, in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -O-(1-5)C alkyl or (1-5)C alkyl; A = C-Br, and Q and Z = CH.

Scheme 5 (see end of text) are compounds of formula 1 in which X and Y = H, n = 1 (but this applies also, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -O-(1-5)C alkyl or (1-5)C alkyl; A =

< / BR>
R4= H, NO2, NH2or Q and Z = CH.

According to the scheme 6 (see end of text) are obtained the compounds of formula 1, where X and Y = H, n = 1 (but this applies also, if n = 2 or 3, with appropriate replacement reagent) T =

< / BR>
where

R1= -O-(1-5)C alkyl/ or (1-5)C alkyl;

A = C OR8where R8= (1-5)C alkylamino; and Q and Z = CH.

According to scheme 7 (see end of text) are obtained the compounds of formula 1 in which X and Y = H, n = 1, but it is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -OH, -O-/(1-5)C alkyl/ or (1-5)C alkyl;

Z = C-C(O)-R6where R6= OH; and Q and A = CH.

According to scheme 8 (see end of text) are obtained the compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

A = CH.

Scheme 9 (see end of text) are obtained the compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -OH, -O-(1-5)alkyl, (1-5)C alkyl;

A = N or N - R5; R5= (1-16)C alkylhalogenide; and Q and Z = CH.

According to scheme 10 (see end of text) are obtained the compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -OH, -O-(1-5)C alkyl or (1-5)C alkyl;

Q = N - R5; R5= (1-16)C alkylhalogenide; and A and Z = CH.

According to scheme 11 (see end of text) are obtained the compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), T =

< / BR>
where

R1= -OH, -O-(1-5)C alkyl or (1-5)C alkyl;

Q = N - R5; R5= (1-16)C alkylhalogenide; and A and Z = CH.

Scheme 12 (see end of text) are compounds of formula 1 in which X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), R at position 3 is T =

< / BR>
where

R1= -OH or-O-(1-5)C alkyl; and the other two R radicals have T = COOH; and A, Q and Z = CH.

Scheme 13 (see end of text) p is Yong reagent), R in the 3-position and 6-position is T =

< / BR>
where

R1= OH or-O-(1-5)C alkyl and the other R radicals in the 9 position is T = COOH; and A, Q and Z = CH.

Scheme 14 (see end of text) are obtained the compounds of formula 1, where X and Y = H, n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), R radicals in the 3 and 9 position have T =

< / BR>
where

R1= -OH or-O-(1-5)C alkyl; and R radical in the 6 position is T = COOH; and A, Q and Z = CH.

Scheme 15 (see end of text) are obtained the compounds of formula 1, where n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), R radicals in the 3 and 9 position have T =

< / BR>
where

R1= -OH or-O-(1-5)C alkyl; and X and Y = H; R radical in the 6 position is T =

< / BR>
where

R4= NO2or NH2and one of X or Y = H and the other = COOH; and A, Q and Z = CH.

Scheme 16 (see end of text) are obtained the compounds of formula 1, where n = 1 (but the method is also applicable, if n = 2 or 3, with appropriate replacement of the reagent), R radicals in the 3 and 6 position have T =

< / BR>
where

R1= -OH or-O-(1-5)C alkyl; and X and Y = H;

R radical in the 9 position is T =

< / BR>
where

R4= NO2or NH2; and one of X or Y = H and the other = COOH;

A, Q and Z = CH.

R radical in the 6 position is T=

< / BR>
where

R1=-OH; And X and Y = H;

R radical in the 3 and 9 positions is T = COOH; and A, Q and Z = CH.

In the above schemes description of the overall process illustrates the particular stage that can be used to perform the desired reaction stage.

The scheme of the synthesis of 1 starts with haloiding industrial available bis-piridinovogo alcohol 1 using thionyl chloride. Similar procedures for the conversion of alcohol into electrophilic substrate, such as processing toluensulfonate, HBr or Hcl, should also result in similarly reactive product that works well in subsequent reactions ring closure. The literature describes numerous procedures macrocyclization, and the desired tetraazamacrocycle were prepared in accordance with the method of Stetter and others, Tetrahedron, 37, 767 - 772 1981. Since then, have been published in more General techniques that give good yield similar macrocycles using milder conditions (A. D. Sherry and others, J. Ong. Chem. , 54, 2990 - 2992, 1989). Detailimage intermediate macrocycle [(3) to obtain (4)] was carried out in acidic conditions with a good yield. The techniques of restorative detailrow the STI. Phosphonomethylglycine to obtain a derivative of Tris-aminophosphonic acid (5, RTMR) was carried out under typical conditions of use of Mannich bases using phosphorous acid and formaldehyde.

In addition to the derivatives of phosphonic acid, phosphonate esters (for example, formula 6 can be obtained in organic conditions in alcohols or aprotic solvents (for example, acetonitrile, benzene, toluene, tetrahydrofuran) and using the desired dialkylphosphate as a nucleophilic agent (see Scheme 2). Depending on the reactivity of amine, the reaction can be conducted at a temperature between -10 and about 100oC. In addition to the above, trialkylphosphine can be used in similar conditions, manniche, giving phosphonate ester by oxidation of phosphorus (III) in phosphorus (V) with simultaneous displacement of one mole of alcohol (Arbuzov reaction). These reactions can be carried out in the presence or in the absence of solvent. When the solvent used for reactions of alcohols or dialkyl - or trialkyl-postitem favorably to the use of alcohol, which is the corresponding complex phosphonate ester, in order from the evidence also using N-alkylation galoidkyetonov in such solvents, as acetonitrile, chloroform, dimethylformamide, tetrahydrofuran or 1,4-dioxane, with or without added dinucleophiles base such as potassium carbonate, at room temperature or above. The resulting ester intermediate compound is then easily hydrolyzed in basic conditions (aqueous hydroxide, pH 8 to 14, 30 - 110oC), giving the corresponding semi-acid derivative.

According to the scheme 3 macrocyclic methylphosphonous acid (10 and 11) are obtained in conditions similar to the conditions described in scheme 2. Condensation using diethoxymethylsilane as a nucleophilic agent and paraformaldehyde may be conducted in solvents such as tetrahydrofuran, dimethylformamide, dioxane, acetonitrile, or in alcoholic media. The resulting Phosphinates ester is then hydrolyzed to the acid (6 norms. HCl, 80 - 100oC) or major (stoichiometric amount of base, 40 - 100oC) conditions, giving appropriate methylphosphonous acid. Alternatively, to obtain phosphinate derivatives with increased lipophilic character, can be used a method developed by the authors A. D. Sherry and others (Inorg. Chem., 1991) using the ture in pyridine link 12-membered tetraazamacrocycle. So, Haidamaka acid (Sigma chemical company, 12) can turn into a bis-kaleidotile derivative (13) with the appropriate substitution in the 4-position of pyridyl. Conversion, leading to this intermediate compound is General in its nature, and its reception is described by the authors Takalo and others (Acta Chemica Scandinavica B 42, 373 - 377, 1988). Subsequent macrocyclization with the use of this intermediate product (15) can be performed using standard reaction in DMF at 100oC with sodium trichotillomania a triamine, or at room temperature with trichotillomania free base and potassium carbonate, sodium carbonate or cesium carbonate as the base, giving the products similar to the products described previously. The subsequent reactions leading to phosphonate semi-acids and phosphinate functionality are identical transformations and the conditions described in the previous schemes.

Scheme 4 describes 4-galodriel substituted macrocycles (16), which can be subjected to substitution in the 4 position peredelnogo fragment, as described in scheme 5. So, organometallics Pd (II) complexes can be used to accelerate or facilitate the reaction of the combination between the f is for this conversion uses anhydrous conditions with triethylamine as solvent and a reaction temperature between about 10 and 30oC to achieve optimal outputs. The same product can also be obtained when using phenylacetylide Cu /I/ in anhydrous pyridine at a temperature between about 80 and 110oC. in Addition, for substitution in the pyridine nucleus may apply standard techniques anion alkylation, using, for example, sodium alcoholate in DMF or dioxane at a temperature of from about 80 to 100oC, using bases, such as potassium carbonate or sodium hydroxide. Macrocyclic tetraazamacrocycles connection(24, 25, 26, 27, 28), derivateservlet this way, compatible with the transformations described in the previous schemes, resulting in similar phosphonate chelating agents.

Variation 4-peredelnogo substitution is described in scheme 6, in which 4-hydroxypyridine fragment (29) alkiliruya romancelatina, giving associated with the intermediate ester of sodium (31), which later turns into a macrocyclic structure. This type of reception alkylation best carried out in anhydrous conditions in an aprotic solvent such as tetrahydrofuran (THF) and using dinucleophiles base, such to the described second Chaubet, etc. for acyclic analogues (Tetrahedron Letters, 31 (40), 5729 - 5732, 1990). Macrocyclic nitrile obtained in this way can be restored to the primary amine (36) using standard procedures followed by protection of the primary amine 2-/tert-butoxycarbonyloxyimino/-2-phenylacetonitrile (BOC-ON; 37). Subsequent funktsionalizatsiya macrocyclic secondary amines(38, 39, 40, 41, 42, 43) can then be carried out using the discussed techniques with the additional requirement of removal of the BOC protective group using triperoxonane acid as described in scheme 6.

As illustrated in figure 7, the functionalization can be carried out also in the 3-position of the pyridine ring in the macrocyclic structure. Newkome and others (Tetrahedron, 39(12), 2001 - 2008, 1983) the previously described synthesis of ethyl 2,6-haloalkalitolerant (45), which serves as primary source material in this synthetic method. So, trichotillomania macrocyclic intermediate compound (46) can metasilicates in acidic conditions (HBr) AcOH, 25 - 115oC) with simultaneous hydrolysis, giving a derivative of nicotinic acid (48), or the recovery of ester in boiling ethanol before detailirohke results in 3-hydroxym the th scheme of the functionalization of the secondary amine, giving various types of phosphonate chelating agents of the formula 1 (49, 50, 51, 52, 53).

In contrast, 3-hydroxymethylene similar favorable protected before functionalization of macrocyclic amines. Figure 8 shows benzyl (Bz), a protective group, because it must be resistant to harsh acidic conditions existing at the stage of detailirohke. After will be held functionalization of secondary amines, as described in the previous schemes, the benzyl group is removed under mild conditions of catalytic hydrogenation (58).

Microsilica derivatives can also be obtained, as shown in the diagrams 12-14, when the same molecule is present as a carboxylate, and phosphonate chelating functionality. Thus, different degrees carboxylate functionality can be entered using the typical procedures for water alkylation using bromoxynil acid. After this stage, other amines can phosphonomethylglycine using the procedures discussed in the previous schemes, using formaldehyde and phosphorous acid, dialkylphosphate or trialkylphosphites.

In schemes 15 and 16 shows the synthetic is hydrated Deputy. In a typical case, macrocyclic amine mono-N-functionalities in an organic solvent such as acetonitrile or DMF, at room temperature using dinucleophiles base such as potassium carbonate. Additional functionalization of the remaining provisions of nitrogen is then carried out using the methods and conditions described in the previous schemes. After entering the desired chelating fragments, the nitro group is reconstructed using platinum oxide and hydrogen in water. In this form of the chelating agent is compatible with the technology of conjugation, which makes it possible to join a larger synthetic or natural molecules.

Scheme 17 illustrates the synthesis of macrocyclic compounds 4, in which the amines in positions 3 and 9 are introduced into a reaction with at least two moles of sodium salt hydroxymethanesulfinic acid in water at a pH of about 9, giving the corresponding macrocyclic compound, in which position 3 and 9 represent the sodium salt methanesulfonic acid (119). The sulfonic acid group is then replaced using sodium cyanide to form the corresponding cyanomethane derivative (120). Cyano group is hydrolyzed in bateley reaction with a derivative of phosphorous acid and formaldehyde with the formation of phosphonic acid 6-position (121), with subsequent acid hydrolysis at elevated temperature, cyanotic groups and any present-derived fragment of phosphorous acid. The resulting connection is a macrocycle with two carboxylic acid groups in positions 3 and 9 and phosphonoacetate group in position 6. Phosphonomethylglycine may also be conducted using the methods discussed above.

Metal ions used for the formation of the complexes of this invention are Gd+3, Mn+2, Fe+3and are industrial or commercially available, for example, are delivered by the company Aldrich chemical company. Present the anion is a halide ion, preferably chloride, or free from salts (metal oxide).

Paramagnetic nuclide invention means a metal ion, which detects the spin angular moment and/or orbital angular moment. Two types of moments are combined, giving the observed paramagnetic moment in a way that depends to a great extent from atoms bearing an unpaired electron, and to a lesser extent,from the environment of these atoms. Paramagnetic nuclides, which are found useful in the practice of this invention, are

The complexes are obtained by methods well known in the art. For example, see the work of Chelating Agents and Metal Chelates, Dwyer and Mellor, Academic Press (1964), Chapter 7. See how to get amino acids in the work of the Synthetic Production and Utilization of Amino Acids (publications Cameco and others), John Willy Andes sons (1974). One example of a receiving system includes the interaction bicyclopentadiene acid with a metal ion in aqueous conditions at a pH from 5 to 7. The complex is formed by a chemical bond and results in a stable paramagnetic nuclide composition (composition), such as stable disassociating paramagnetic nuclide from the ligand.

The complexes of the present invention are assigned when the molar ratio of ligand to metal is at least about 1:1, preferably from 1:1 to 3:1, more preferably from 1:1 to 1.5:1. A large excess of ligand is undesirable because uncomplexed ligand can be toxic to the animal or may result in cardiac arrest or gipocalziemiceski convulsions.

Antibodies or fragments of antibodies that can be used in the conjugates described in this description can be obtained using the Dom hybridization, well known in the art (Kohler and Milsten Nature, 256, 495-497 (1975); Eur. J. Immunol., 6, 511-519, 1976). Such antibodies typically have highly specific reactivity. In aiming at this the antibody conjugates can be used antibodies directed against any desired antigen or hapten. Preferably the antibodies used in the conjugates, are monoclonal antibodies, or fragments thereof, having high specificity to the desired epitope (or epitopes). Antibodies used in the present invention may be directed against, for example, tumors, bacteria, fungi, viruses, parasites, Mycoplasma, differentiation antigens and other antigens on the cell membranes, pathogen surface antigens, toxins, enzymes, allergens, drugs and any biologically active molecules. Some examples of antibodies or fragments of antibodies are 1116-NS-19-9, 1116-NS-3d, 703D4, 704A1, CC49, SS and B72.3. All these antibodies are deposited in ATSS. A more complete list can be found in U.S. patent 4193983. The conjugates of the present invention is particularly preferred for the diagnosis of various cancers.

The invention is used with a physiologically acceptable carrier, excipient or zapolnitelejj in suspension, injectable solution or other suitable preparative forms. Can be used physiologically acceptable suspendresume environment with or without adjuvants.

For diagnostic use "effective amount" preparative form. Also examines methods of diagnosis in vivo, using preparative forms of the invention.

Other areas of use of certain chelating agents of the present invention include removing unwanted metals (e.g. iron) from the body, attaching to polymeric carriers for a variety of purposes, such as diagnostic agents, and the removal of metal ions by selective extraction. The ligands of formula 1, having at least two R radicals T equal to T/O/R1OH, may be used in less than stoichiometric amount. Similar use area known for compounds (U.S. patent NN 2609390, 3331773, 3336221 and 3434969).

The invention will be explained with regard to the following examples, whose purpose is purely an illustration of the invention.

Some of the terms used in the following examples are defined as follows:

LC = liquid is Oh with full manual Q-SeparateTManion-exchange column (h cm).

DMF = dimethylformamide,

AcOH = acetic acid,

1CP = inductively coupled plasma,

g = g/s/,

mg = milligrams,

kg = kilograms,

ml = milliliter/s/,

μl = microliter/s/.

General procedure pH stability

Was made of the original solution 159Gd Cl3(or 153 SmCl3) by adding 2 ál 310-1M 159 Gd Cl30.1 norms. HCl to 2 ml 310-4M GdCl3solution media. Then he prepared the corresponding solutions of the ligands in deionized water. Then were prepared complexes 1:1 ligand/metal by combining the ligand dissolved in 100 to 500 μl of deionized water, 2 ml of the original 159 GdCl3solution followed by thorough mixing to obtain an acidic solution (pH 2). The pH of the solution is then raised to 7.0 using 0.1 norms. NaOH. Then we determined the percentage of metal in the form of a complex by passing the sample solution of the complex through a SephadexTMG - 50 column, elution with 4:1 malevil solution (85% NaCl/NH4OH and collection 2x3 ml fractions. A quantitative measure of radioactivity in the United buervenich solutions were then compared with the number remaining is elicina pH of aliquots of a solution of the complex using IM NaOH or IM HCl and determine the percentage of metal, existing in the form of the complex, using the method of ion exchange as described above. According to the experimental comparison, it is known that the Sm results are identical in the case of complexation and bearsdley of dilandau of the present invention, the source materials.

Example A. Obtaining 2,6-bis(chloromethyl) pyridine.

To 100 ml of thionyl chloride, which was cooled (ice bath), was added 24 g (to 0.17 mol) of 2,6-bis (hydroxymethyl) pyridine. After 30 min the reaction mixture was warmed up to room temperature, then was heated under reflux for 1.5 hours, After cooling the reaction mixture to room temperature, the solid that formed was filtered off, washed with benzene and dried in vacuum. The solid then was kind of balanced out with saturated sodium bicarbonate, filtered and dried, yielding 23.1 g (71,5%) of the desired product in the form of not quite white crystalline solid, so pl. 74,5 - 75,5oC, and further characterized by the following data:

1H NMR (CDCl3) 4,88 (C., 4H), 7,25 - to 7.95 (m, 3H).

Example B. Obtaining 3,6,9-Tris/p-tamilselvan/-3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1 (15), 11, 13-the triens.

The solution in the dim and was heated to 100oC in nitrogen atmosphere. To the solution was added dropwise over 45 min to 2 g to (11.4 mmol) of 2,6-bis(chloromethyl/pyridine (obtained by the procedure of example A) in 37 ml of DMF. When the addition was completed, the reaction mixture is stirred at 40oC for 12 hours To the reaction mixture was then added 50 to 75 ml of water, which has resulted in immediate dissolution of NaCl, followed by precipitation of the desired product. The resulting suspension was then filtered, and the solid was rinsed with water and dried in vacuum. The target product was obtained as a pale reddish brown powder, 6.5 g (86%), so pl. 168 - 170oC Razlog. and it was further characterized as follows:

1H NMR (CDCl3) 2,40 (C., 3H), 2,44 (C., 6H), to 2.75 (m, 4H), 3,30 (m, 4H), 4,28 (C., 4H), 7,27 (D., 2H), 7,34 (D., 4H), 7,43 (D., 2H), 7,65 (D., 4H), of 7.75 (t, 1H); and

13C NMR 21,48, 47,29 50,37, 54,86, 124,19, 127,00, 127,11 129,73, 135,04, 135,74, 138,95, 143,42, 143,73, 155,15.

Example C. 3, 6, 9, 15-tetraazabicyclo [9.3.1.] pentadec-1(15), 11, 15-the triens.

A solution of HBr and AcOH were prepared by using a mixture of 48% HBr and glacial AcOH in the ratio of 64: 35. To 112 ml of HBr/AcOH mixture was added 5.5 g (8.2 mmol) 5,6,9-Tris/p-tamilselvan/-3,6,9,15-tetraazabicyclo [9.3.1.] pentadec-1(15), 11,13-triens (obtained with the aid of which shivani within 72 hours The reaction mixture was then cooled to room temperature and concentrated to about 1/10 of the initial volume. The remaining solution was mixed vigorously, and added 15 - 20 ml of diethyl ether. Formed not quite white solid, which was filtered with diethyl ether and dried in vacuum. Dry tetrahydropyrimidine salt was then dissolved in 10 ml of water, brought to a pH of 9.5 with NaOH (50% weight/weight) and continuously extracted with chloroform for 4 hours. After drying over anhydrous sodium sulfate, the chloroform was evaporated, giving a light reddish brown oil, which slowly crystallized upon standing at room temperature, giving 1.2 g (71%) of the target product, so pl. 86 - 88oC, which was characterized by the following data:

1H NMR (CDCl3) of 2.21 (m, 4H), at 2.59 (m, 4H), 3,06 (C., 3H), 3,85 (C., 4H), 6.89 in (D., 2H), 7,44 (t, 1H), and

13C NMR 48,73, 49,01, 53,63, 119,67, 136,29, 159,54.

Example D. Obtaining 3,6,9,15-tetraazabicyclo [9,3,1]-pentadec-1 (15), 11,13-triene-3,9-dimethylaniline acid.

A suspension of 500 mg (2.4 mmol) of 3,6,9,15-tetraazabicyclo [9.3.1]-pentadec-1(15), 11,13-triens (obtained by the procedure of example C) were mixed in 6 ml of water, and the pH was brought on the, the pH was brought to 9 with 50% aqueous sodium hydroxide. After stirring for 3 h at room temperature data13C NMR indicated complete conversion to the target bis-methansulfonate product.

Example E. Obtaining 3,6,9,15-tetraazabicyclo [9.3.1]-pentadec-1 (15), 11,13-triene-3,9-dimethylaniline.

To the reaction mixture containing 3,6,9,15-tetraazabicyclo [9.3.1]-pentadec-1(15), 11,13-triene-3,9-dimethylsulfonium acid from example D, was added 47 mg (9.6 mmol) of sodium cyanide. The reaction mixture was stirred at room temperature for 24 h

13C NMR data showed that the conversion to bis-nitrile was completed. The reaction mixture was then filtered, extremerules three times with 25 ml portions of chloroform, dried over anhydrous magnesium sulfate and concentrated, giving a viscous oil. The oil was then dissolved in chloroform, were pulverized with cyclohexane and concentrated, giving in the form of a white powder, 530 mg (78%) target dimethylaniline product.

Example f-3 ,9-bis/sodium methansulfonate/-3,6,9,15-tetraazabicyclo-[9.3.1] pentadec-1/15/, 11,13-triens (PC25).

An aqueous solution (10.0 ml) of 3,6,9,15-tetraazabicyclo [9.3.1] pentad is Oh HCl, and the mixture was mixed for 10 min to ensure complete dissolution. The resulting solution had a pH of 8.6. To the solution were then added to 1.37 g (10.2 mmol) HOCH2SO3Na with 5 ml of deionized water. The solution was heated at 60oC for 10 min, and the pH value fell to 5.6. After cooling, the pH was brought to 9.0 with 1M aqueous sodium hydroxide, then lyophilization was carried out, giving the desired product as a white solid in quantitative yield, which was characterized by the following data:

1H NMR (D2O) 2,87 (t, 4H), 3,18 (t, 4H), cent to 8.85 (C., 4H), 4,11 (C., 4H), 7,03 (D., 2H), 7,55 (t, 1H); and

13C NMR (D2O) 48,52, 54,04, 58,92, 79,09, 123,90, 141,37, 161,89.

Example G. Obtaining 3,9-bis(metileritrol)-3,6,9,15-tetraazabicyclo [9.3.1]-pentadec-1(15), 11,13-triens.

To aqueous solution, 10.0 ml, 3,9-bis/sodium methansulfonate/-3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1 (15), 11,13-triens (obtained by the procedure of example F), and 2.26 g (5 mmol) was added 0.6 g (12,24 mmol) of sodium cyanide. The mixture was mixed for 3 h at room temperature. The pH of the reaction mixture was about 10. The pH value was brought to above 13 using concentrated aqueous hydroxide h and filtered. After removal of solvent and concentration in vacuo the desired product was separated in the form of a waxy white powder, 1.0 g (71%) and was characterized by the following data:

1H NMR (CDCl3) 2,03 (width, C., 4H), of 2.64 (m, 4H), 3,82 (C., 4H), 3,90 (C., 4H), 7,14 (D., 2H), a 7.62 (t, 1H); and

13C NMR (CDCl3) 46,08, 46,64, 52,89, 60,78, 115,31, 122,02, 137,57, 157,33.

Example H. the Receipt of 3,9-bis/metileritrol/-6- /methylenediphosphonate/-3,6,9,15-tetraazabicyclo [9.3.1] -pentadec-1(15), 11,13-triene-3,9-dimethylaniline.

3,9-bis/metileritrol/-3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11, 13-triene (obtained by the procedure of example G), 285 mg (1.0 mmol) was combined with 60 ml (2.0 mmol, excess) of paraformaldehyde and 0,354 ml (372 mg, 3.0 mmol, excess) trimethylphosphite. The mixture is gently stirred for 10 min to obtain a suspension, and then was heated to 90oC for 1 h After the excess reagents and by-products were removed under vacuum (1 h at 125oC/0.01 mm RT. Art.) the resulting dark brown residue was dissolved in 20 ml of chloroform and was washed with deionized water (I ml). The organic layer was dried over anhydrous magnesium sulfate, filtered and excess solvent was evaporated in vacuo, giving trebunie:

1H NMR (CDCl3) 2,61 (Shir.S., 8H), 2,73 (D., 2H), 3,62, and 3,68 (C., 6H), of 3.73 (SD, 4H), 3,84 (C., 4H), 7,06 (D., 2H), EUR 7.57 (t, 1H); and

13C NMR (CDCl3) 44,44, 50,74, 51,03, 51,85, 52,51, 60,28, 115,61, 122,27, 137,24, 156,61.

Example 1. Getting 3,6,9,15-tetraazabicyclo[9.3.1] pentadec-1(15), 11,13-triene-3,6,9-methylenediphosphonate.

A mixture of 1 g (4.8 mmol) of 3,6,9,15-tetraazabicyclo[9.3.1] pentadec-1(15), 11,13-triens (obtained by the procedure of example (C), 4.8 g (28.8 mmol) of triethylphosphite and 864 mg (28.8 mmol) of paraformaldehyde was heated at 90oC with constant stirring for 45 minutes the Reaction mixture was concentrated in vacuo, and the viscous oil was chromatographically on a column of basic alumina with elution by chloroform. After concentration of the organic solvent the desired product was separated in the form of a colorless oil, 2.0 g (64%) and was characterized by the following data:

1H NMR (CDCl3) of 1.23 (m, 18H), 2,77 (m, 12H), 3.04 from (doctor, 6H), 4,13 (m , 12H), 7,17 (D., 2H), 7,60 (t, 1H); and

13C NMR (CDCl3) 16,43, 50,03, 50,31, 50,43, 50,77, 51,23, 51,38, 52,63, 53,30, 60,86, 60,92, 61,63, 61,74, 61,83, 61,93, 62,32, 76,46, 76,97, 77,18, 77,48, 122,50, 137,10, 157,18; and

31P NMR 24,92 (C., 2P), 24,97 (C., 1P).

Example J. Getting 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11,13-triene-3,6,9-methylenedi/n-propyl/phosphonate.

oC with stirring for 1 h the Resulting homogeneous solution was concentrated in vacuo, giving a viscous oil, which was chromatographically on a column of neutral alumina with elution by chloroform. After concentration of the organic solvent the desired product was separated in the form of a colorless oil, 320 mg (90%), and was characterized by the following data:

1H NMR (CDCl3) to 0.88 (m, 18H), to 1.61 (m, 12H), of 2.72 (m, 12H), 3,03 (D., 6H), of 3.97 (m, 12H), 7,13 (d. 2H), 7,55 (t, 1H); and

13C NMR (CDCl3) 9,96, 23,73, 49,84, 50,14, 50,26, 50,57, 51,11, 51,23, 52,43, 53,01, 60,78, 60,84, 67,27, 67,40, 122,48, 137,04, 157,16; and

31P NMR 24,98 (3P).

Example K. Getting 3,6,9,15-tetraazabicyclo[9.3.1] pentadec-1(15), 11,13-triene-3,6,9-methylenedi/n-butyl/phosphonate.

A mixture of 500 mg (2.4 mmol) of 3,6,9,15-tetraazabicyclo[9.3.1] pentadec-1(15), 11,13-triens (obtained by the procedure of example (C), 2.0 g (8 mmol) of tributylphosphine and 240 mg (8 mol) of paraformaldehyde was heated at 100oC with stirring for 1 h the Resulting viscous solution was concentrated in vacuo, yielding an oil which centering of the organic solvent the desired product was separated in the form of a colorless oil, 1,25 g (65%), which was characterized by the following data:

1H NMR (CDCl3) is 0.84 (m, 18H), of 1.27 (m, 12H), was 1.58 (m, 12H), to 2.57 (m, 12H), 3,01 (d. 6H), 3,99 (m, 12H), 7,12 (D., 2H), 7,54 (t, 1H); and

13H NMR (CDCl3) 13,42, 13,46, 18,50, 18,59, 32,16, 32,43, 49,88, 50,03, 50,16, 50,63, 51,11, 51,27, 52,48, 53,16, 60,71, 60,78, 65,38, 65,48, 65,58, 122,46, 136,96, 157,14; and

31P NMR ARE 24.88 (2P), 24,93 (1P).

Example L. Getting 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11,13-triene-3[(4-nitrophenyl)acetate].

To a solution of 2.5 ml of chloroform, which was rapidly mixed, and 200 mg (0.97 mmol) of 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11,13-triens (obtained by the procedure of example C) was added in one portion 266 mg (0.97 mmol) of bromine-/4-nitrophenyl/acetate and 2.5 ml of chloroform. The reaction mixture was stirred for 24 h at room temperature. The solution was concentrated in vacuo, giving semi-solid substance, which was chromatographically on silicagel column with elution with a mixture of chloroform/methanol/ammonium hydroxide (16: 4: 1). After concentration of the organic eluent was allocated the desired product as a pale yellow solid, 250 mg (64%), which was characterized by the following data:

13C NMR (CDCl3) 45,67, 45,90, 45,97, 51,65, 52,08, 52,28, 53,78, 69,54, 119,03, 119,icicle [9.3.1] pentadec-1 (15), 11, 13-triene-3,6,9-trimethylindolenine acid (RTMR).

A mixture of 2.06 g (10 mmol) of 3, 6, 9, 15-tetraazabicyclo [9.3.1]-pentadec-1(15), 11, 13-the triens (obtained by the procedure of example (C), 11.3 g (138 mmol) of phosphoric acid and 15 g (152 mmol) of concentrated HCl was heated to careful boiling (103oC) with constant stirring, followed by adding dropwise (2 ml/min) 12.2 g (150 mmol, 15 ml) of aqueous formaldehyde (37%). After complete addition the reaction mixture is stirred at the temperature of reflux distilled for 16 h, cooled to room temperature and concentrated to a thick viscous oil. The product was then purified using LC anion-exchange chromatography (0-30% formic acid, 3 ml/min, retention time = 32 min). Combined fractions were dried by freezing, giving 4.8 g (99%) of the desired product as a white solid, so pl. 275-280oC, which was characterized by the following data:

1H NMR (D2O) and 2.83 (m, 6H), 3.46 in (m, 10H), 7,28 (D., 2H) 7,78 (t, 1H); and

13C NMR 53,61, 53,81, 55,27, 57,93, 62,20, 125,48, 143,08, 152,31; and

31P 8,12 (2P), 19,81 (1P).

Example 2. Obtaining a complex of 153 Sm-3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11, 13-triene-3,6,9-trimethylindolenine acid (153Sm-RTMR).

3H2O (310-4M 0.01 norms. HCl) containing isotopic indicator 153 SmCl3. After thorough mixing was determined by the percentage of metal in the form of a complex by passing the sample solution of the complex through the column with SephadexTMwhen elution with a mixture of 4:1 salt solution of 0.85% NaCl (NH4OH and collection 2x3 ml fractions. The amount of radioactivity in the combined solution after elution was compared with the magnitude of the substances remaining on the resin. Under these conditions, the complex was removed eluent, and uncomplexed metal remains on the resin. This method was determined that the complexation was 98%. The sample solution was passed through the resin, were used to study pH. Then pH was determined stability using the General procedure described above.

Example 3. Getting 3,9-dioxyna acid-6-/methylenephosphonate acid/-3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1 (15), 11, 13-the triens (RSAR).

Concentrated means of hydrochloric acid solution (37%, 5 ml) 3,9-bis/metileritrol/-6-/methylenediphosphonate/-3,6,9,15-tetraazabicyclo [9.3.1]pentadec-1(15), 11, 13-TA the solution was evaporated to dryness, with subsequent joint by evaporation and deionized water (2x10 ml) to remove excess hydrochloric acid. The final product stood out as a dark brown solid after freeze-drying the concentrated aqueous solution, which was characterized by the following data:

1H NMR (D2O) 2,68 (Shir.S., 4H), and 3.31 (Shir.S., 4H), 4,08 (C., 4H), 4,55 (C., 4H), 7,16 (D., 2H), 7,68 (t, 1H); and

13C NMR (D2O) 52,35, 54,04, 57,02, 59,24, 62,26, 125,52, 143,64, 152,36, 171,54; and

31P NMR (D2O) 20,03.

Example 4. Getting 3,6,9,15-tetraazabicyclo [9.3.1.] pentadec-1(15), 11,13-triene-3,6,9-methylenediphosphonate Tris (potassium salt (RMINE).

To water of 0.1 norms. to a solution of potassium hydroxide (2 ml) was added 250 mg (0.38 mmol) of 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11,13-triene-3,6,9-methylenediphosphonate (obtained by the procedure of example 1). The solution was heated at 90oC for 5 h, the Reaction mixture was cooled to room temperature, filtered, dried by freezing, giving the desired product in the form of not quite white solids, 252 mg (97%), which was characterized by the following data:

13C NMR (D2O) 18,98, 19,82, 51,78, 52,06, 53,08, 54,46, 54,68, 57,01, 58,22, 60,24, 63,19, 63,25, 63,36, 63,49, 63,59, 63,95, 64,18, 64,25, 66,80, 126,62, 141,63, 159,40; I,13-triene-3,6,9-methylene/n-propyl/phosphonate Tris/potassium salt (RMRP).

To aqueous solution of potassium hydroxide (0.5 ml of 1 norms./dioxane /0.5 ml) was added 81 mg (to 0.108 mmol) of 3,6,9,15-tetraazabicyclo [9.3.1]-pentadec-1(15), 11,13-triene-3,6,9-methylene/n-propyl/phosphonate /received using the procedure of example J). The solution was heated under conditions of reflux distilled within 24 hours, the Reaction mixture was cooled to room temperature and was extracted with diethyl ether. The ether extract was then concentrated in vacuo, giving the desired product in the form of not quite white solids, 48.6 mg (60%), which was characterized by the following data:

31P NMR 20,49 (C., 3P).

Example 6. Getting 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1(15), 11.13-triene-3,6,9-methylene/n-butyl/phosphonate Tris/sodium salt/ (PMBHE).

To aqueous solution of 35 ml of 1 norms. potassium hydroxide was added 3,21 g (3.88 mmol) of 3,6,9,15-tetraazabicyclo /9.3.1/ pentadec-1 (15),11.13-triene-3,6,9-methylenedi/n-butyl/phosphonate (obtained by the procedure of example J). The solution was heated under conditions of reflux distilled for 5 days. The reaction mixture was cooled to room temperature, filtered, and the filtrate was subjected to drying by freezing, giving painted in a cream color solid. This substance is then suspendibility in 160 ml was metanationals, giving semi-solid substance. This substance was taken in 150 ml of chloroform m were dried over anhydrous sodium sulfate and filtered. After concentration in vacuo the product stood out as not quite white solids, 1.86 g (62%), and was characterized by the following data:

IH NMR (D2O) to 0.68 (m, 9H), 1.14 in (m, 6H), to 1.37 (m, 6H), was 2.76 (D., 6H), to 3.41 (m, 12H), to 3.73 (m, 6H), 7,24 (D., 2H), 7,76 (t, 1H); and

13C YARM (D2O) 15,76, 15,80, 21,12, 21,20, 34,96, 35,06, 35,14, 52,08, 52,53, 53,38, 53,48, 54,49, 54,75, 57,70, 57,76, 61,86, 67,65, 67,75, 67,98, 68,08, 125,15, 142,93, 152,25; and

31P Yarm 9,73 (C., 2P) 21,00 (C., 1P).

Example 7. Getting 3,6,9,15-tetraazabicyclo[9.3.1] pentadec-1(15), 11,13-triene - 3[(4-nitrophenyl)acetate]-6,9-methylenediphosphonate.

A solution of 250 mg (of 0.62 mmol, 3,6,9,15-tetraazabicyclo [9.3.1] pentadec-1 (15), 11,13-triene-3-[(4-nitrophenyl) acetate] (obtained using the procedure of example L), 624 mg (3.7 mmol) of triethylphosphite and 111 mg (3.7 mmol) of paraformaldehyde were mixed with 100oC for 1 h the Resulting homogeneous solution was concentrated in vacuo, giving a viscous oil. The oil was dissolved in 10 ml of chloroform and washed with water (3x5 ml). The organic layer was dried over anhydrous magnesium sulfate, filtered, and the filtrate was concentrated in Vacu the Yarm (CDCl3) 24,67, (C., 2P), are 24.88 (C., 1P).

The biodistribution. The General procedure.

Rat Sprague Dawley were left to acclimatize for five days, then inheritability 100 μl of a solution of the complex through the tail vein. During injection the rats weighed between 150 and 200 g After 30 min, rats were killed by cervical displacement and cut. The amount of radioactivity in each tissue was determined by counting in a NaI scintillation counter, connected to a multichannel analyzer. The number of calculations were compared with the calculations in 100 ál of standard samples to determine the percentage of the dose in each tissue or organ.

Percentage of dose in the blood was estimated by taking blood for 7% of body weight. Percentage dose in bone was estimated by multiplying the percentage of the dose in the hips 25. Percentage dose muscle was estimated by making muscles for 43% of body weight.

In addition to the biodistribution in organs chelates of the compounds of formula I were evaluated on the effectiveness of localization in the bones, as phosphonates are known for their ability to bind to hydroxyapatite.

Example 1. The percentage injected dose of complex of example 2 (153 Sm-PCTMP) in several tissues is given below). Numeric values Bones 34,87

Liver - 0,99

Kidney - 1,42

Spleen - 0,07

Muscle - 4,77

Blood - 6,27

Example 2. The percentage injected dose of complex of example 5 (153 Sm-PMPHE) in several tissues is given below. Numeric values represent the average from at least 3 rats 2 h after injection:

Fabric - Average

Bones - 10,86

Liver - 4,14

Kidney - 1.55V

Spleen - 0,05

Muscle - 1,19

Blood - 0,25

Heart - 0,12

Light - 0,12

Brain - 0,00

Stomach - 0,44

Small intestine - 10,71

Colon - 2,17

Example 3. The percentage injected dose of complex of example 6 (153 Sm-PMBHE) in several tissues is given below. Numeric values represent the average from at least 3 rats 2 h after injection:

Fabric - Average

Bones of 3.73

Liver - 2,70

Kidney - 0,43

Spleen - 0,05

Muscle - 1,09

Blood - 0,14

Heart - 0.02

Light - 0.04

Brain - 0,00

Stomach - 0,08

Small intestine - 57,89

Colon - 0,77

Example 4. The percentage injected dose of complex of example 3 (153 Sm-PC2A1) in some tissues (given below). Numerical values represent the average from at least 3 rats 2 h after injection:

Fabric - Average

Bones - 47,98

Liver - 1,46

Kidney - 0,93<

Stomach - 0,25

Small intestine - 13,10

Colon - 0,12

Experiments on visualization. The General procedure.

First were prepared injectable solutions (0.5 M) by dissolving the appropriate amount of each complex in 2 ml of deionized water. The pH of the solution was then brought to 7.4 using 1M HCl or NaOH depending on the need. Then determined the total content of Gd each solution using ICP analysis.

Shot rats Sprague Dawley were intramuscularly injected with one of the solutions of the metal described above, at a dose of 0.05 - 0.1 mmol Gd/kg of body weight. Then through various time intervals was filmed image and compared with penjelasannya control in 0 time.

Example 2. The complex Gd-RTMR (obtained in example 2) showed renal activation and bone localization in the shoulders, spine and sternum.

Other embodiments of the invention will become apparent (clear) specialists in this field in the consideration of this specification or practice of the implementation described in the invention. It should be borne in mind that the descriptions and examples serve only to illustrate, the true essence and scope of the invention is ormula I

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where the R group

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where X and Y are independently H or COOH;

n is an integer 1, 2 or 3, provided that when n = 2, then the sum of X and Y must be equal to two or more H, when n = 3, then the sum of X and Y must be equal to three or more H;

T - COOH group

< / BR>
where R1Is-OH or-O-C1- C5-alkyl;

R4- NO2or NH2provided that at least one T must be a group P(O)R1OH, and provided that when T represents one

< / BR>
then one of X or Y is specified, the R radical can represent COOH, and all other X and Y radicals R represent H, and provided that the sum of R4radicals, when they are present, may not exceed 1,

or their pharmaceutically acceptable salts.

2. Connection on p. 1, in which at least two R radicals are T representing P(O)R1OH, where R1is OH, and the third T is COOH, n = 1, and X and Y each represents h

3. Connection on p. 1, in which the three R radicals have T representing P(O)R1OH, where R1is OH, and X and Y each represents H, namely: 3,6,9,15-tetradecyl [9.3.1] pentadec-1(15), 11,13-triene-3,6,9-trimethylantimony acid or its pharmaceutically acceptable salt.

what item R the radical T is COOH, and n = 1.

5. Connection on p. 1, in which the two R radicals T is P(O)R1-OH, where R1is OH, and in the third R-radical T is P(O)R1OH, where R1represents-O-C1- C5-alkyl, and n = 1.

6. Connection on p. 1, in which the R radical is at least one T is P(O)R1OH, where R1is OH and the other two R radicals T represents COOH or P(O)R1OH, and n, R1X and Y have the meanings given in paragraph 1.

7. Connection on p. 6, in which one R radical is at least one T is P(O)R1OH, where R1is OH and the other two R radicals T are P(O)R1OH, where R1represents-O-C1- C5-alkyl, and n = 1.

8. Connection on p. 6, in which one R radical T is P(O)R1OH, where R1is OH and the other two R radicals T represents COOH, n = 1.

9. Connection on p. 1, in which the R radical is three T are P(O)R1OH, where R1represents-O-C1- C5-alkyl, and n, R1X and Y have the meanings given in paragraph 1.

10. Connection on p. 1, in which the three R radicals T is P(O)R1OH, where R1represents-O-C1- C5

13. Connection on p. 1, in which the three R radicals X, Y and n have the meanings given in paragraph 1, and T represents one group

< / BR>
where R4matter specified in paragraph 1,

and the other two T have the meanings given in paragraph 1.

14. Connection on p. 13, in which n = 1.

15. Connection on p. 14, in which the R radical, which contains T fragment which contains the group R4there is also one of their X and Y specified R radical representing COOH.

16. Connection on p. 15, in which the two R radicals, which do not contain the radical R4all other X and Y radicals are h

17. Connection on p. 16, in which the two R radicals, not containing R4both radicals T are P(O)R1OH, where R1matter specified in paragraph 1, and are the same.

18. Connection on p. 1, in which X and Y represent H, T represents COOH or

< / BR>
where R1is OH or-O-C1-C5- alkyl.

19. The method of obtaining compounds of General formula I on p. 1, characterized in that the compound of the formula I, in which at least one R group is hydrogen, is subjected to reaction with fosforiliruyusciye agent, namely phosphorous cyclotouriste acceptable salt.

20. The method according to p. 19, characterized in that as fosforiliruyusciye agent is used as a compound of General formula P(OR1)3in which R1is OH or-O-C1- C5-alkyl, and the process is executed in the environment of the solvent.

 

Same patents:

The invention relates to organic chemistry, particularly to a technology for higher esters alkylphosphonic acids of General formula

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where

R is alkyl (C1-C2halogenated;

R', R" are alkyl (C4-C8

The invention relates to new derivatives methylenephosphonic acid, especially to new, halogensubstituted Amida and ether-Amida (ester-Amida) methylenephosphonic acid, methods of producing these new compounds, as well as to pharmaceutical compositions containing these new compounds

-chloroethyl)vinylphosphonate" target="_blank">

The invention relates to the chemistry of organophosphorus compounds, specifically to methods for di-(-chloroethyl)vinylphosphonate (Vinnitsa) by dehydrochlorination di(-chloroethyl)--chloroethylphosphonic (isomerate) in the presence of a nucleophilic reagent by heating

The invention relates to 2-sharonlee - 4,5,6,7 - tetrahydro-2-sharinaletisha-phosphonates and-Phosphinates, inhibiting enzymatic activity; to compositions containing these compounds, their use and the treatment of disruptive disorders, and to methods for their preparation

The invention relates to new derivatives methylenephosphonic acid of General formula I

< / BR>
in which R1, R2, R3and R4independently are C1-C10the alkyl straight or branched chain, optionally unsaturated, C3-C10-cycloalkyl, optionally unsaturated, aryl, aralkyl, silicom SiR3or hydrogen, in formula I, at least one of the groups R1, R2, R3and R4is hydrogen and at least one of the groups R1, R2, R3and R4different from hydrogen

The invention relates to a technology for obtaining esters vinylphosphonic acid, which finds wide use as a reactive flame retardants and plasticizers in the production of various polymer materials
The invention relates to the field of chemistry of organophosphorus compounds, in particular, to methods for producing uranyl trinational the salt postemergency acid, used in medicine and cosmetics as an anti-virus drug

The invention relates to new derivatives methylenephosphonic acid, especially to new, halogensubstituted Amida and ether-Amida (ester-Amida) methylenephosphonic acid, methods of producing these new compounds, as well as to pharmaceutical compositions containing these new compounds

The invention relates to compounds of General formula (I):

,

where A is -(CH2)ngroup, and n includes the interval between 1 and 10, R is an acyl residue of a known anti-inflammatory compounds belonging to the class of salicylic, akriluksusnoy, arylpropionate, Anthranilic, 4,5-dihydroxy - or 4,5,8-trihydroxy-9,10-dihydro-9,10-dioxo-2-intracisternally and nicotinic acid

The invention relates to new derivatives methylenephosphonic acid of General formula I

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in which R1, R2, R3and R4independently are C1-C10the alkyl straight or branched chain, optionally unsaturated, C3-C10-cycloalkyl, optionally unsaturated, aryl, aralkyl, silicom SiR3or hydrogen, in formula I, at least one of the groups R1, R2, R3and R4is hydrogen and at least one of the groups R1, R2, R3and R4different from hydrogen

The invention relates to a method for oksietilidendifosfonovaya acid formula

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a highly effective combined and used in power, oil, fragrance, textile, household, medicine, production of mineral fertilizers

FIELD: organophosphorus compounds, medicine.

SUBSTANCE: invention relates to new biologically active phosphonate compounds. Invention describes phosphonate compound of the formula:

wherein R1 and R'1 represent independently hydrogen atom (-H) substituted possibly with -O-(C1-C24)-alkyl, -O-(C1-C24)-alkenyl, -O-(C1-C24)-acyl, -S-(C1-C24)-alkyl, -S-(C1-C24)-alkenyl or -S-(C1-C24)-acyl wherein at least one among R and R'1 doesn't represent -H and wherein indicated alkenyl or acyl comprise from 1 to 6 double bonds; R2 and R'2 represent independently -H substituted possibly with -O-(C1-C7)-alkyl, -O-(C1-C7)-alkenyl, -S-(C1-C7)-alkyl, -S-(C1-C7)-alkenyl, -O-(C1-C7)-acyl, -S-(C1-C7)-acyl, -N-(C1-C7)-acyl, -NH-(C1-C7)-alkyl, -N-((C1-C7)alkyl)2, oxo-group, halogen atom, -NH2, -OH or -SH; R3 represents phosphonate derivative of nucleoside or biphosphonate; X represents compound of the formula:

L represents a valence bond or a bifunctional binding molecule of the formula: -J-(CR2)t-G- wherein t is a whole number from 1 to 24; J and G represent independently -O-, -S-, -C(O)O- or -NH-; R represents -H, unsubstituted or substituted alkyl or alkenyl; m means a whole number from 0 to 6; n = 0 or 1. Also, invention describes pharmaceutical compositions comprising phosphonate compounds, method for treatment of osteoporosis in mammal, method for increasing mineral osseous density, method for prophylaxis of apoptosis of osteoblasts and osteocytes in mammal, method for treatment of viral infection in mammal, method for treatment of growing neoplasm in mammal and method for proliferation of cells. Invention provides preparing new compounds eliciting useful biological properties.

EFFECT: valuable medicinal properties of phosphonate compounds.

17 cl, 2 dwg, 7 tbl, 21 ex

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