Composition of polyethylene glycol maleimide derivative and polymerization inhibitor

Abstract

The present invention provides a composition of a polyethylene glycol maleimide derivative and a polymerization inhibitor. In particular, the present invention provides a composition of an 8-arm polyethylene glycol maleimide derivative and a phenolic polymerization inhibitor. The ingredient and content of the polymerization inhibitor in the composition are reasonably chosen, thereby significantly increasing stability of the polyethylene glycol maleimide derivative, effectively avoiding the undesirable effect of gel solidifying due to polymerization during storage and transportation, and extending a pot life and shelf life of a product thereof.

Claims

1. A pharmaceutical conjugate, comprising a composition of a polyethylene glycol maleimide derivative and a polymerization inhibitor and a drug, wherein the mass ratio of the polymerization inhibitor to the polyethylene glycol maleimide derivative is ≥0.1 μg:1 g, the polyethylene glycol maleimide derivative contains at least one terminal maleimide group.

2. The pharmaceutical conjugate according to claim 1, wherein said polymerization inhibitor is selected from one or a combination of two or more of a radical polymerization inhibitor, a phenolic polymerization inhibitor, an inorganic compound polymerization inhibitor, and an organometallic compound polymerization inhibitor.

3. The pharmaceutical conjugate according to claim 1, wherein the mass ratio of the polymerization inhibitor to the polyethylene glycol maleimide derivative is from 0.1 μg to 10 mg:1 g.

4. The pharmaceutical conjugate according to claim 1, wherein the polyethylene glycol maleimide derivative has the following structure: PEG-X-MAL (I), wherein PEG is polyethylene glycol residue, X is a linking group of PEG and MAL, and is selected from: one or a combination of two or more of —(CH.sub.2).sub.r—, ##STR00016## —(CH.sub.2).sub.rO—, —(CH.sub.2).sub.rCO—, —(CH.sub.2).sub.rNH—, —(CH.sub.2).sub.rCONH—, —(CH.sub.2).sub.rNHCO—, —(CH.sub.2).sub.rS—, —(CH.sub.2).sub.rCOO— and —(CH.sub.2).sub.rOCO—, r is an integer from 0 to 10, MAL is a maleimide group ##STR00017## R.sub.1 and R.sub.2 are independently selected from: —H, a C1-6 alkyl group, a C1-6 alkoxy group, a C3-6 cycloalkyl group, and a C4-10 alkylene cycloalkyl group.

5. The pharmaceutical conjugate according to claim 4, wherein said X is selected from the group consisting of: a single bond, —CH.sub.2—, —CH.sub.2CH.sub.2—, —CH.sub.2CH.sub.2CH.sub.2—, —CH(CH.sub.3)—, —CH.sub.2CH(CH.sub.3)—, —CH.sub.2CH.sub.2CH(CH.sub.3)—, —(CH.sub.2).sub.rO—, —(CH.sub.2).sub.rCO—, —(CH.sub.2).sub.rNH—, —(CH.sub.2).sub.rCONH— and —(CH.sub.2).sub.rNHCO—; and/or, the r is an integer from 0 to 5.

6. The pharmaceutical conjugate according to claim 4, wherein said PEG is a linear polyethylene glycol residue having a structure shown in formula II or III: ##STR00018## wherein p and q are independently selected from the group consisting of an integer of 1-2400; or, the PEG is a Y-shape or U-shape polyethylene glycol residue having a structure of formula IV or V: ##STR00019## wherein i and k are independently selected from an integer of 1-1200; or PEG is a multi-branched polyethylene glycol residue having the structure shown in formula VI: ##STR00020## wherein n is an integer from 1 to 800, 1 is an integer from 0 to 7, and j is an integer from 1 to 8, R is a core molecule of a multi-branched polyethylene glycol, and R is selected from the group consisting of: pentaerythritol, oligo-pentaerythritol, methyl glucoside, sucrose, diethylene glycol, propylene glycol, glycerol, and polyglycerol residues; wherein Y is a terminal group selected from the group consisting of: C1-C6 alkoxy group, hydroxyl, carboxyl group, succinimide carbonate group, succinimide acetate group, succinimide propionate group, and succinimide succinate group, succinimide group, dithiopyridyl group, propionic acid group, aldehyde group, thiol ester group, acryloxy group, acrylic acid group, azido group, glutaric acid group, alkynyl, 4-nitrophenyl carbonate, silane, and carboxymethyl.

7. The pharmaceutical conjugate according to claim 6, wherein said polyethylene glycol maleimide derivative has the following structure: ##STR00021## and, j=8, PEG is an eight-arm polyethylene glycol residue.

8. The pharmaceutical conjugate of claim 7 wherein R has the structure of formula IX or X: ##STR00022##

9. The pharmaceutical conjugate according to claim 6, wherein PEG has a molecular weight of from 1 to 80 kDa.

10. The pharmaceutical conjugate according to claim 2, wherein the radical polymerization inhibitor is selected from one or more of 1,1-diphenyl-2-trinitrophenylhydrazine, 4,4′-dimethoxydiphenyl oxynitride, 4,4′-dinitrodiphenyl oxynitride, di-tert-butyl oxynitride, 2,2,6,6-tetramethyl-4-hydroxyl piperidine oxynitride and benzoyl peroxide; and/or said phenolic polymerization inhibitor is selected from one or more of: hydroquinone, methyl hydroquinone, p-tert-butylcatechol, 2-tert-butyl hydroquinone, 4-methoxyphenol, 2,5-di-tert-butyl hydroquinone, 2,6-di-tert-butyl-4-methylphenol, 4,4′-dihydroxybiphenyl, pyrogallol and bisphenol A; and/or said inorganic compound polymerization inhibitor is selected from one or more of: ferric chloride, cuprous chloride, copper chloride, copper sulfate, titanium trichloride, titanium chloride, sodium sulfate, sodium sulfide and ammonium thiocyanate; and/or said organometallic compound polymerization inhibitor is selected from one or more of: naphthenic acid copper, chromium acetate, nickel acetate, copper dimethyldithioformate, and copper dit-butyldithiocarbamate.

11. The pharmaceutical conjugate according to claim 1, wherein the radical polymerization inhibitor is 1,1-diphenyl-2-trinitrophenylhydrazine; and/or, the phenolic polymerization inhibitor is hydroquinone or p-tert-butyl phenol; and/or the inorganic compound polymerization inhibitor is copper chloride; and/or the organometallic compound polymerization inhibitor is nickel acetate.

12. The pharmaceutical conjugate according to claim 1, wherein the polymerization inhibitor is a phenolic polymerization inhibitor.

13. The pharmaceutical conjugate according to claim 1, wherein the polymerization inhibitor is selected from one or more of: hydroquinone, methyl hydroquinone, p-tert-butylcatechol, 2-tert-butyl hydroquinone, 4-methoxyphenol, 2,5-di-tert-butyl hydroquinone, 2,6-di-tert-butyl-4-methylphenol, 4,4′-dihydroxybiphenyl, pyrogallol and bisphenol A.

14. The pharmaceutical conjugate according to claim 1, wherein the polymerization inhibitor is hydroquinone or p-tert-butyl-phenol.

15. The pharmaceutical conjugate according to claim 1, wherein the drug is selected from the group consisting of amino acids, polypeptides, proteins, sugars, organic acids, alkaloids, flavonoids, terpenoids, terpenoids, phenylpropanoid phenols, steroids and steroids.

16. The pharmaceutical conjugate according to claim 15, wherein the drug is a polypeptide or protein drug.

17. The pharmaceutical conjugate according to claim 16, wherein the polypeptide or protein drug contains free sulfhydryl groups; and/or, a sulfhydryl group is introduced at a specific site in the polypeptide or protein drug.

Description

DRAWINGS

(1) FIG. 1 shows the GPC pattern of 8ARM(TP)-PEG-MAL-40K with a polymerization inhibitor added after 2 days irradiation.

(2) FIG. 2 shows the effect of the type and concentration or content of the polymerization inhibitor on the time of 8ARM(TP)-PEG-MAL-40K before the complete deterioration.

(3) FIG. 3 shows the effect of the type and concentration or content of the polymerization inhibitor on the time of 8ARM(TP)-PEG-MAL-20K before the complete deterioration.

(4) FIG. 4 shows the effect of the type and concentration or content of the polymerization inhibitor on the time of 8ARM(TP)-PEG-MAL-10K before the complete deterioration.

(5) FIGS. 5A and 5B show the GPC detection spectrum in Example 5, wherein FIG. 5A is the GPC detection spectrum with hydroquinone, and FIG. 5B is the GPC detection spectrum without hydroquinone.

DETAILED DESCRIPTION OF THE INVENTION

(6) Unless defined otherwise, all technical and scientific terms used in the present invention have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains.

(7) “Alkyl” refers to a hydrocarbon chain radical that is linear or branched and free of unsaturated bonds. The C.sub.1-C.sub.6 alkyl means an alkyl having 1 to 6 carbon atoms, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, t-butyl, n-pentyl, isopentyl, neopentyl, tert-amyl, n-hexyl, isohexyl and the like.

(8) “Alkoxy” means a substituent formed by substituting the hydrogen in hydroxy group with an alkyl group, and C1-C6 alkoxy group means an alkoxy group having 1 to 6 carbon atoms, such as methoxy or ethoxy, propoxy, butoxy, and the like.

(9) “Cycloalkyl” means an alicyclic hydrocarbon, such as those containing 1 to 4 monocyclic and/or fused rings. It may contain 3 to 18 carbon atoms, preferably 3 to 10 carbon atoms, such as cyclopropyl, cyclohexyl or adamantyl and the like. C3-C6 cycloalkyl in the present invention means a cycloalkyl having 3 to 6 carbon atoms, such as cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

(10) In addition, some specific groups and their chemical structures involved in the present invention correspond to the following: hydroxyl group, —OH;

(11) ##STR00010##
(wherein Q.sub.1 may be an alkyl group or a heterocyclic group such as methyl, ethyl, n-propyl, tert-butyl, pyridyl (such as

(12) ##STR00011##
etc.);

(13) ##STR00012##
glutarate, such as

(14) ##STR00013##
(wherein Q.sub.2 may be the same or different alkyl or alkoxy group, such as methyl, ethyl, propyl, butyl, pentyl, methoxy, ethoxy, propyl Oxyl, butoxy, etc., preferably, Q.sub.2 is methyl, ethyl, n-propyl, methoxy, ethoxy, n-propoxy, etc.);

(15) ##STR00014##

(16) In the definition of a linking group in the present invention, the “combination” means a group formed by linking two or more of the listed linking groups by a chemical bond. For example, the combination of —(CH.sub.2).sub.r— and —(CH.sub.2).sub.rNHCO— may be —(CH.sub.2).sub.rNHCO(CH.sub.2).sub.r— and specifically, the combination of —CH.sub.2— and —CH.sub.2CH.sub.2NHCO— may be —CH.sub.2CH.sub.2NHCOCH.sub.2—, —CH.sub.2CH.sub.2CH.sub.2NHCO—. The “combination” is used to define the chemical structure of the linking group, and does not involve the preparation steps, the order of linking groups in the combination, etc.

(17) The polyethylene glycol maleimide derivative described in the present invention is a polyethylene glycol to which a maleimide group is attached, as shown in formula I of the present invention, specifically as shown in formula XI; the linkage can be achieved by a covalent bond or by a linking group, and the reaction for achieving the linkage is well known to those skilled in the art, which is not specifically defined in the present invention.

(18) The “composition of a polyethylene glycol maleimide derivative and a polymerization inhibitor” described in the present invention may further contain other components, such as ultraviolet absorbers (UV-P) to improve product quality or function. Those skilled in the art can add other components according to actual needs. Moisture, impurities and the like are inevitably involved in the process of production, transportation or storage of the polyethylene glycol maleimide derivatives and the compositions thereof and these substances are not specifically defined in the present invention.

(19) The technical solutions of the present invention will be described clearly and completely with reference to the embodiments of the present invention. It is obvious that these embodiments are only a part of the possible embodiments of the present invention, and not all of the embodiments.

(20) All other embodiments obtained by a person of ordinary skill in the art based on the embodiments of the present invention without involvement of inventive step are within the scope of the present invention.

(21) The compounds used in the present invention are either commercially available or can be prepared according to the disclosed preparation methods, which are not meant to limit the scope of the invention.

(22) The polyethylene glycol derivative used in the examples was supplied by JENKEM Technology Co., Ltd. (Beijing). All others are commercially available reagents.

Example 1

(23) Hydroquinone, DPPH, nickel acetate, copper chloride and was respectively obtained and mixed with 8ARM(TP)-PEG-MAL-40K (250 mg) to form composition powders, wherein hydroquinone, DPPH, nickel acetate and copper chloride were all at 100 ppm.

(24) The above composition powders were separately placed in a watch glass and placed in a light box at a constant temperature (2750 Lux, 18.5° C.) for two days, and the GPC results are shown in FIG. 1.

(25) According to FIG. 1, the GPC concentration or content comparison results after 2 days of irradiation of 8ARM(TP)-PEG-MAL-40K (to which a polymerization inhibitor was added) are shown in Table 1.

(26) TABLE-US-00001 TABLE 1 GPC concentration or content after 2 days irradiation of 8ARM(TP)-PEG-MAL-40K with polymerization inhibitor added Addition of a polymerization inhibitor nickel copper hydroquinone DPPH acetate chloride 8ARM(TP)-P 83.9 77.3 67.9 54.3 EG-MAL-40K purity(%)

(27) As shown in Table 1, among the above four polymerization inhibitors, with the addition of hydroquinone, 8ARM(TP)-PEG-MAL-40K had the lowest degree of purity reduction, with the best inhibiting effect.

Example 2

(28) Different types and concentrations or content of additives were obtained and mixed with 8ARM(TP)-PEG-MAL-40K (250 mg) to form composition powders: hydroquinone, p-tert-butyl-phenol, a mixture of hydroquinone and UV absorber (hydroquinone & UV-P, the mass ratio is 1:1), DPPH, nickel acetate, copper chloride, wherein the concentrations or content of each additive were at 1, 20, 100, 400 and 1000 ppm.

(29) The above composition powders were separately placed in a watch glass and placed in a light box at a constant temperature (2750 Lux, 18.5° C.) to record the time required before complete deterioration (i.e., when a gel insoluble in methanol occurred), and the result was as shown in FIG. 2 (Note: If no additives were added, 8ARM(TP)-PEG-MAL-40K were completely deteriorated after two days of irradiation).

(30) It can be seen from FIG. 2 that the effect of phenolic polymerization inhibitors such as hydroquinone and p-tert-butyl-phenol are better than the other polymerization inhibitors, and the mixture of hydroquinone and UV-P has the similar effect as that of hydroquinone alone.

Example 3

(31) Composition powders containing different concentrations or content of hydroquinone, DPPH, nickel acetate, copper chloride and 8ARM(TP)-PEG-MAL-20K (250 mg), were separately taken, wherein the concentration or content of each polymerization inhibitor were at 1, 20, 100, 400, 1000 ppm.

(32) The above composition powders were separately placed in a watch glass and placed in a light box at a constant temperature (2750 Lux, 18.5° C.) to record the time required before complete deterioration (i.e., when a gel insoluble in methanol occurred), and the result was as shown in FIG. 3 (Note: If no inhibitor is added, 8ARM(TP)-PEG-MAL-20K would completely deteriorate after two days of irradiation).

Example 4

(33) A composition powder containing different concentrations or content of hydroquinone, DPPH, nickel acetate, copper chloride and 8ARM(TP)-PEG-MAL-10K (250 mg) was separately taken, wherein the concentration or content of each polymerization inhibitor was at 1, 20, 100, 400, 1000 ppm.

(34) The above composition powders were separately placed in a watch glass and placed in a light box at a constant temperature (2750 Lux, 18.5° C.) to record the time required before complete deterioration (i.e., a gel insoluble in methanol occurred), and the result was as Shown in FIG. 4 (Note: If no inhibitor is added, 8ARM(TP)-PEG-MAL-10K would completely deteriorate after one day of irradiation).

(35) It can be seen from Examples 2-4 that the above polymerization inhibitors, particularly hydroquinone, have a good stabilizing effect on polyethylene glycol maleimide derivatives of different molecular weights.

(36) The 8ARM(TP)-PEG-MAL used in the embodiment of the present invention has the following structure:

(37) ##STR00015##

(38) Enhanced stability, prolonged pot life and storage period could be achieved all the same for the maleimide derivative of polyethylene glycol of other configurations and molecular weights, when mixed with the polymerization inhibitor in the examples of the present invention. Those experimental data are not shown herein.

Example 5

(39) 1 g of 8ARM(TP)-MAL-40K and 10 mg of hydroquinone were weighed and put into 50 ml single mouth round-bottomed flask and dissolved by 10 ml of dichloromethane. In addition, 1 g of 8ARM(TP)-MAL-40K were weighed and put into a 50 ml single mouth round-bottomed flask and dissolved by 10 ml dichloromethane. After the two systems were placed under an incandescent lamp for 48 hours, 40 mg thiosalicylic acid was added respectively. The reaction lasted for two hours. Samples were taken for GPC test, the GPC test spectrum is shown in FIGS. 5A-5B, and the corresponding spectrum data are shown in Table 2 and Table 3. The GPC test results in Table 4 show that the purity of pharmaceutical conjugate in the reaction system with hydroquinone is 89.8%, and that in the reaction system without hydroquinone is 87%, and the percentages of the high molecular weight impurity and the low molecular weight impurity are less than those in the system without hydroquinone.

(40) TABLE-US-00002 TABLE 2 Peak Retention number time Area Height Concentration 1 13.986 40155 1083 5.789 2 14.877 622103 16701 89.819 3 16.281 30363 962 4.384 Total 692621 18747

(41) TABLE-US-00003 TABLE 3 Peak Retention number time Area Height Concentration 1 13.975 77989 1972 7.306 2 14.882 928480 25133 86.981 3 16.296 60980 1535 5.713 Total 1067449 28640

(42) TABLE-US-00004 TABLE 4 Main High molecular Low molecular product weight impurity weight impurity GPC test (%) (%) (%) With Hydroquinone 89.8 5.8 4.4 without Hydroquinone 87 7.3 5.7

(43) The above results show that the addition of polymerization inhibitor (such as hydroquinone) in the reaction system can improve the purity of pharmaceutical conjugate products.

(44) The above is only the preferred embodiment of the present invention, and is not intended to limit the present invention. Any modifications, equivalent substitutions, etc., which are within the spirit and principles of the present invention, should be included in the scope of the present invention.