NZ227985A - Intermediate activated ionophores carrying a thiol group - Google Patents
Intermediate activated ionophores carrying a thiol groupInfo
- Publication number
- NZ227985A NZ227985A NZ22798585A NZ22798585A NZ227985A NZ 227985 A NZ227985 A NZ 227985A NZ 22798585 A NZ22798585 A NZ 22798585A NZ 22798585 A NZ22798585 A NZ 22798585A NZ 227985 A NZ227985 A NZ 227985A
- Authority
- NZ
- New Zealand
- Prior art keywords
- ionophore
- thiol group
- ionophores
- residue
- formula
- Prior art date
Links
- 230000000236 ionophoric effect Effects 0.000 title claims description 25
- 239000002555 ionophore Substances 0.000 title claims description 23
- 125000003396 thiol group Chemical group [H]S* 0.000 title claims description 11
- 229930191564 Monensin Natural products 0.000 claims description 10
- GAOZTHIDHYLHMS-UHFFFAOYSA-N Monensin A Natural products O1C(CC)(C2C(CC(O2)C2C(CC(C)C(O)(CO)O2)C)C)CCC1C(O1)(C)CCC21CC(O)C(C)C(C(C)C(OC)C(C)C(O)=O)O2 GAOZTHIDHYLHMS-UHFFFAOYSA-N 0.000 claims description 10
- 229960005358 monensin Drugs 0.000 claims description 10
- GAOZTHIDHYLHMS-KEOBGNEYSA-N monensin A Chemical compound C([C@@](O1)(C)[C@H]2CC[C@@](O2)(CC)[C@H]2[C@H](C[C@@H](O2)[C@@H]2[C@H](C[C@@H](C)[C@](O)(CO)O2)C)C)C[C@@]21C[C@H](O)[C@@H](C)[C@@H]([C@@H](C)[C@@H](OC)[C@H](C)C(O)=O)O2 GAOZTHIDHYLHMS-KEOBGNEYSA-N 0.000 claims description 10
- 150000001875 compounds Chemical class 0.000 claims description 8
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims description 8
- 238000006243 chemical reaction Methods 0.000 claims description 7
- DANUORFCFTYTSZ-UHFFFAOYSA-N epinigericin Natural products O1C2(C(CC(C)(O2)C2OC(C)(CC2)C2C(CC(O2)C2C(CC(C)C(O)(CO)O2)C)C)C)C(C)C(OC)CC1CC1CCC(C)C(C(C)C(O)=O)O1 DANUORFCFTYTSZ-UHFFFAOYSA-N 0.000 claims description 6
- 238000000034 method Methods 0.000 claims description 6
- DANUORFCFTYTSZ-BIBFWWMMSA-N nigericin Chemical compound C([C@@H]1C[C@H]([C@H]([C@]2([C@@H](C[C@](C)(O2)C2O[C@@](C)(CC2)C2[C@H](CC(O2)[C@@H]2[C@H](C[C@@H](C)[C@](O)(CO)O2)C)C)C)O1)C)OC)[C@H]1CC[C@H](C)C([C@@H](C)C(O)=O)O1 DANUORFCFTYTSZ-BIBFWWMMSA-N 0.000 claims description 6
- 125000002777 acetyl group Chemical group [H]C([H])([H])C(*)=O 0.000 claims description 5
- AHTFMWCHTGEJHA-UHFFFAOYSA-N s-(2,5-dioxooxolan-3-yl) ethanethioate Chemical compound CC(=O)SC1CC(=O)OC1=O AHTFMWCHTGEJHA-UHFFFAOYSA-N 0.000 claims description 5
- AVXURJPOCDRRFD-UHFFFAOYSA-N Hydroxylamine Chemical compound ON AVXURJPOCDRRFD-UHFFFAOYSA-N 0.000 claims description 4
- 239000001257 hydrogen Substances 0.000 claims description 3
- 229910052739 hydrogen Inorganic materials 0.000 claims description 3
- 241001125671 Eretmochelys imbricata Species 0.000 claims 1
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims 1
- 229920002521 macromolecule Polymers 0.000 description 5
- 239000000047 product Substances 0.000 description 5
- 230000008878 coupling Effects 0.000 description 4
- 238000010168 coupling process Methods 0.000 description 4
- 238000005859 coupling reaction Methods 0.000 description 4
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 3
- 239000000543 intermediate Substances 0.000 description 3
- 239000012071 phase Substances 0.000 description 3
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 125000003158 alcohol group Chemical group 0.000 description 2
- 238000012412 chemical coupling Methods 0.000 description 2
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 2
- 238000011065 in-situ storage Methods 0.000 description 2
- 239000005445 natural material Substances 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- SVOAXTTVWPHOQW-UHFFFAOYSA-N 2-acetylsulfanylbutanedioic acid Chemical compound CC(=O)SC(C(O)=O)CC(O)=O SVOAXTTVWPHOQW-UHFFFAOYSA-N 0.000 description 1
- 229930182504 Lasalocid Natural products 0.000 description 1
- 241001655322 Streptomycetales Species 0.000 description 1
- TUCNEACPLKLKNU-UHFFFAOYSA-N acetyl Chemical compound C[C]=O TUCNEACPLKLKNU-UHFFFAOYSA-N 0.000 description 1
- 239000012736 aqueous medium Substances 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 125000002843 carboxylic acid group Chemical group 0.000 description 1
- 239000013065 commercial product Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 125000001183 hydrocarbyl group Chemical group 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- BBMULGJBVDDDNI-OWKLGTHSSA-N lasalocid Chemical compound C([C@@H]1[C@@]2(CC)O[C@@H]([C@H](C2)C)[C@@H](CC)C(=O)[C@@H](C)[C@@H](O)[C@H](C)CCC=2C(=C(O)C(C)=CC=2)C(O)=O)C[C@](O)(CC)[C@H](C)O1 BBMULGJBVDDDNI-OWKLGTHSSA-N 0.000 description 1
- 229960000320 lasalocid Drugs 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000000655 nuclear magnetic resonance spectrum Methods 0.000 description 1
- 239000012074 organic phase Substances 0.000 description 1
- 239000003960 organic solvent Substances 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 229910052760 oxygen Inorganic materials 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 239000012429 reaction media Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Landscapes
- Heterocyclic Carbon Compounds Containing A Hetero Ring Having Oxygen Or Sulfur (AREA)
Description
New Zealand Paient Spedficaiion for Paient Number £27985 22 7985 NO DRAWINGS Priority D3te(s): <•"* Complete Specification Filed, hr.
Class; ^Q7?.HT?./ , .Q k>/.
Col m ^ / oVjl t % » W * •*» I ♦'« • • • W«TTi / ••«•■»•#»•»••••• PubHCio. 0K« MY.M...
P.O. Journal. No: Under the provisions of Regulation 23 (1) the Specification has been antedated to 19 §5.,.
Initials Patents Form No. 5 O NEW ZEALAND PATENTS ACT 1953 COMPLETE SPECIFICATION O INTERMEDIATE ACTIVATED lONOPHORES We, SANOFI of 40, Avenue George V, 75008 Paris, France a French company, hereby declare the invention, for which we pray that a patent may be granted to us, and the method by which it is to be performed, to be particularly described in and by the following statement *" i / 22 7 9 8 5 Intermediate Activated Ionophores The present invention relates to activated carboxylic ionophores, useful as intermediates in the synthesis of conjugates in which a monovalent carboxylic ionophore is associated by means of a covalent bond with a macromolecule. Such conjugates are described and claimed in the applicants New Zealand patent specification No. 212118.
In the abovementioned specification, the replacement of carboxylic ionophores with conjugates in which a monovalent carboxylic ionophore is associated by means of a covalent bond with a macromolecule such as an antibody, a fragment of an antibody, a protein, a peptide or alternatively a mixed macromolecule obtained by the covalent coupling of an antibody with a protein is disclosed.
The ionophores used are known molecules, especially natural substances isolated from strains of Streptomycetes bacteria which contain a hydrocarbon skeleton incorporating oxygen heterocycles. There is always a carboxylic acid group at one end of the chain and there are one or more alcohol groups at the other end (B.C. Pressman, Annual review of Biochemistry, 45, 501-530, 1976).
These natural substances, as well as certain semisynthetic compounds derived therefrom, share an 227985 ionophoric activity, i.e. the capacity to enable metal ions (especially monovalent ions) to be transferred from a hydrophilic phase to a lipophilic phase, the said phases being immiscible. with the ionophore can be carried out by a variety of methods with the proviso that the chosen method: - preserves the respective biological activities of the components of the conjugate; - ensures a satisfactory reproducibility and a good coupling yield; - makes it possible to control the value of the ratio ionophore/macromolecule in the conjugate obtained; and corresponding to these characteristics, it is possible to choose those which involve one or more thiol groups for forming the bond. In this case, it is possible to use a thiol group which has been introduced artificially into one of the compounds to be coupled, and to introduce into the other compound one or more groups which are capable of reacting with the thiol groups, in an aqueous medium with a pH of between 5 and 9, at a temperature not exceeding 30°C, to produce a stable, covalent and clearly defined bond. r The chemical coupling of the macromolecules - leads to a stable and water-soluble product. Among all the chemical coupling methods 22798 Thus, by the present invention there is provided a method whereby a thiol group is introduced artificially into the ionophores by reaction with S-acetyl-mercaptosuccinic anhydride, which is capable of acylating one of the alcohol groups of the ionophore. It will then be possible to free this thiol group by reaction with hydroxylamine to remove the acetyl protecting radical, in a manner already described (Archives of Biochemistry and Biophysics, 119, 41-49, 1967).
Suitable ionophores are monensin, nigericin, grisonixin and lasalocid; particular preference is given to monensin and nigericin, which have the formulae: nigericin The artificial introduction of the thiol is carried out by reacting the ionophore with S-acetyl-mercaptosuccinic anhydride of the formula: 0 |j ^ s-co-ch3 ' o \— ■. li :s^s 227985 in an inert organic solvent at approximately ambient temperature.
This reaction forms an ester bond between one « of the hydroxyls of the ionophore, which is the primary hydroxy1 in the case of monensin and nigericin, and the acetylmercaptosuccinic acid to give a compound of the formula: l°-0-C0-CH-S-C0-CH3 CH2COOH in which I° represents the residue of the ionophore from which one of the hydroxyls, which is the primary hydroxyl in the case of monensin and nigericin, has been removed.
The thiol group of the above compound can be freed by reaction with hydroxylamine, and the resulting product of the formulas i'-o-co-ch-sh ch2cooh in which 1° is as defined above, can be used in situ for the coupling operation.
The said activated ionophores, which are represented by the formula: 1-0-c0-ch-s-w ch2cooh iii .-"■vVlv - \ s# r ■ in which W is hydrogen or an acetyl group, are new and represent another feature of the present invention.
The group: I"-0-C0-CH- ch2cooh is indicated hereafter as I and denotes "the ionophore to be coupled." The following example illustrates the invention without limiting its scope.
Example: Monensin into which a thiol has been introduced.
The monensin used is a commercial product. It was modified as follows: 693mg of monensin are dissolved in chloroform and then reacted with 350mg of S-acetylmercaptosuccinic anhydride (SAMSA). The reaction is left to proceed for 1/2 hour at ambient temperature. The reaction medium is subsequently evaporated to dryness in vacuo, the residue is then taken up in ethyl acetate and the mixture is washed extensively with water. The organic phase is then dried and pumped to dryness in vacuo. The product is obtained in crystalline form after drying in vacuo. It is identified by its NMR spectrum and its mass spectrum.
The product thus obtained (S-acetyl activated monensin) has the formula III above in which 1° is the 227985 residue of monensin from which the primary hydroxyl has been removed, and W is an acetyl group.
The acetyl radical can be released by reaction with hydroxylamine at a final concentration of 50mM, and the product thus obtained, corresponding to the formula III above in which 1° is the residue of monensin from which the primary hydroxyl has been removed, and W is hydrogen, can be used directly for the coupling operation.
However, for the preparation of the ionophoric conjugate it is preferable to free the thiol group in situ in the presence of the S-acetyl activated monensin and the activated antibody. 227085
Claims (5)
1. Activated carboxylic ionophores containing a thiol group, optionally acetylated, artificially introduced by the action of S-acetylmercaptosuccinic anhydride on the ionophore and optional release of the acetyl group by reaction with hydroxylamine > said activated carboxylic ionophore corresponding to the formula: i'-o-co-ch-s-w I ch2c00h in which 1° represents the residue of an ionophore from which one of the hydroxyls has been removed, and W represents hydrogen or an acetyl group.
2. A compound as claimed in claim 1 having the formula: i°-0-c0-ch-s-w ch2c00h o m which I represents the residue of an ionophore from which the primary hydroxyl has been removed, and W is as defined in claim !•
3. A compound as claimed in claim 2 having the formula: 1°-o-co-ch-s -v ch2cooh in which 1° is the residue of monensin or nigericin from which the primary hydroxyl has been removed, and -9- 227.9 85 wherein W is as defined in claim 1.
4. An activated carboxylic ionophore as claimed in claim 1- as specifically set forth herein.
5. A process for producing a compound as claimed in claim 1 substantially as herein described. SANOFI By their Attorneys BALDWIN. SON & CAREY u
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| FR8408073A FR2564839B1 (en) | 1984-05-23 | 1984-05-23 | CONJUGATES COMBINING BY COVALENT BINDING A MONOVALENT CARBOXYLIC IONOPHORE AND A MACROMOLECULE AND THEIR USE AS POTENTIALIZERS OF IMMUNOTOXINS |
| FR8411208A FR2567521B1 (en) | 1984-07-13 | 1984-07-13 | NEW ACTIVE IONOPHORES |
| NZ212118A NZ212118A (en) | 1984-05-23 | 1985-05-20 | Conjugate of macromolecule and ionophore; use as immunotoxin potentiator |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| NZ227985A true NZ227985A (en) | 1990-11-27 |
Family
ID=27251220
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| NZ22798585A NZ227985A (en) | 1984-05-23 | 1985-05-20 | Intermediate activated ionophores carrying a thiol group |
Country Status (1)
| Country | Link |
|---|---|
| NZ (1) | NZ227985A (en) |
-
1985
- 1985-05-20 NZ NZ22798585A patent/NZ227985A/en unknown
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