WO2018028589A1 - Poly-conjugate and preparation method therefor, and pharmaceutical composition comprising same and use thereof - Google Patents

Abstract

Provided are a poly-conjugate and a preparation method therefor, and a pharmaceutical composition comprising the poly-conjugate and the use thereof. The poly-conjugate comprises: an amino acid polymer as a main chain and an active pharmaceutical molecule as a branched chain, wherein the active pharmaceutical molecule is 7-ethyl-10-hydroxycamptothecine and linked to the amino acid polymer through one amino acid at 20-hydroxyl. The method for preparing the poly-conjugate comprises the following steps of: 1) adding the amino acid polymer, a compound represented by formula A, a catalyst and a condensing agent into a solvent; 2) bringing the solution obtained in step 1) into contact with a poor solvent, and then filtering same; 3) adding the filter cake obtained in step 2) into the solvent, and bringing same into contact with a deprotection agent to obtain a poly-conjugate. The pharmaceutical composition comprises the poly-conjugate, and a pharmaceutically acceptable adjuvant. The poly-conjugate and pharmaceutical composition are useful in the treatment of a tumour or cancer.

Description

Polyconjugate and preparation method thereof, and pharmaceutical composition comprising the same, and use thereof Technical field

The present invention relates to a multimeric binder and a process for the preparation thereof, and a pharmaceutical composition comprising the same, and more particularly to the use of irinotecan active metabolite SN38 and amino acid polymer A formed multimeric conjugate that can be used in medical antitumor drugs.

Background technique

Irinotecan, a clinically used hydrochloride (trihydrate), can be used as a first-line treatment for adult metastatic colorectal cancer. For patients who have failed 5-Fu chemotherapy, irinotecan can be used as a second line. Therapeutic medication. At the same time, irinotecan is being used in a variety of clinical trials for gastric cancer, esophageal cancer, and extensive small cell lung cancer.

The English chemical name of irinotecan hydrochloride trihydrate is: (S)-4,11-diethyl-3,4,12,14-tetrahydro-4-hydroxy-3,14-dioxo1H-pyrano[3',4':6,7]-indolizino[1,2-b]quinolin-9-yl-[1,4'bipiperidine]-1'-carboxylate,monohydrochloride,trihydrate,the chemical formula is: C ₃₃ H ₃₈ N ₄ O ₆ · HCl·3H ₂ O, molecular weight: 677.19, its chemical structural formula is:

Irinotecan is metabolized in the body to form its active metabolite 7-ethyl-10-hydroxycamptothecin. The CAS accession number is 86639-52-3. The active metabolite is also commonly known as SN38. The chemical structure of SN38 is :

Attempts have been made to administer SN38 directly as an active therapeutic agent to treat related tumor diseases, but in view of the poor water solubility of SN38, it is very difficult to administer injectables. Therefore, it seems to be an effective solution to make SN38 into its water-soluble derivative. For example, CN1429121A and CN101507820A disclose the formation of a combination of camptothecin or an analog thereof such as SN38 and polyglutamic acid.

However, as in the study by Puja Sapra et al. (Puja Sapra, et al., Novel Delivery of SN38 Markedly Inhibits Tumor Growth in Xenografts, Including a Camptothecin-11-Refractory MSN38el, Clin Cancer Res 2008; 14(6) March 15, 2008: 1888) As mentioned, the biologically active lactone-type SN38 has the possibility of ring opening in its E ring, which forms a carboxylic acid type of inactive substance, namely:

The hydroxyl group at position 20 on the E ring is shown in the above SN38 structure, which may be referred to herein as a 20-hydroxy group.

The above-mentioned ring opening to form a carboxylic acid type needs to be avoided as much as possible. Unfortunately, the prior art solutions are not ideal in the face of the above problems.

Accordingly, those skilled in the art are still expecting a new method of treating tumors, such as a new method for the clinical application of the irinotecan active metabolite SN38, and expecting such a method, including the therapeutic agent provided by the method having one or The advantages of many aspects.

Summary of the invention

The object of the present invention is to overcome the above-mentioned deficiencies of the prior art and to provide a novel method for treating tumors, for example, to provide an activity of irinotecan active metabolite SN38 (i.e., 7-ethyl-10-hydroxycamptothecin). A method of treating a disease as described above; and the therapeutic agent provided by the method has one or more advantages. In the present invention, it has been surprisingly found that SN38 multimeric conjugates formed by the combination of polymeric macromolecules (e.g., amino acid polymers) with SN38 exhibit at least one completely unexpected property. The present invention has been completed based on this finding.

In order to achieve the above object, in one aspect, the present invention provides a multimeric binder, said plurality The polyconjugate includes: an amino acid polymer as a main chain and an active drug molecule as a branch, wherein the active drug molecule is 7-ethyl-10-hydroxycamptothecin, which passes an amino acid at the 20-hydroxyl group Attached to the amino acid polymer.

In another aspect, the present invention also provides a method of preparing the above polyconjugate, comprising the steps of:

1) adding an amino acid polymer, a compound represented by Formula A, a catalyst, and a condensing agent to a solvent;

2) contacting the solution obtained in the step 1) with a poor solvent, and then filtering;

3) adding the filter cake obtained in step 2) to a solvent and contacting the deprotecting agent to obtain a multimeric binder;

Wherein R' is a side chain of a natural amino acid.

In another aspect, the invention also provides a pharmaceutical composition comprising the above polymeric composition, and a pharmaceutically acceptable adjuvant.

In another aspect, the invention also provides the use of the above polymeric compositions and pharmaceutical compositions for treating a tumor or cancer.

The polyconjugate according to the present invention has better solubility than its corresponding active drug, and has excellent chemical stability and excellent biological effects, and specific effects will be described in detail in the following description and examples.

DRAWINGS

The drawings are intended to provide a further understanding of the invention, and are intended to be a In the drawing:

Figure 1: Efficacy trial of nude mice using human colon cancer HCT116 cells: each group was administered as a tail vein Time-dependent changes in tumor volume after surgery.

Figure 2: Nude mouse pharmacokinetic experiments using human colon cancer HCT116 cells: time-dependent changes in plasma drug concentration and released SN-38 concentration.

Figure 3: Nude mouse pharmacokinetic experiments using human colon cancer HCT116 cells: time-dependent changes in drug concentration and released SN-38 concentration in tumors.

detailed description

Specific embodiments of the present invention will be described in detail below. It is to be understood that the specific embodiments described herein are merely illustrative and not restrictive.

The endpoints and any values of the ranges disclosed herein are not limited to the precise range or value, and such ranges or values should be understood to include values that are close to the ranges or values. For numerical ranges, the endpoint values of the various ranges, the endpoint values of the various ranges and the individual point values, and the individual point values can be combined with one another to yield one or more new ranges of values. The scope should be considered as specifically disclosed herein.

In the present invention, the term "polyconjugate" may also be referred to as "conjugate", "polymer" or the like, and refers to a chemical substance including an active drug molecule and an amino acid polymer, the activity The drug molecule moiety is bonded to the amino acid polymer moiety by chemical bonding (including direct binding or indirect bonding through an intermediate small molecule), for example, by an ester bond or an amide bond, and thus, the polyconjugate of the present invention is essentially a The compound, ie, the multimeric conjugate of the invention may also be referred to as a compound. The term "poly" has the meanings well known to those skilled in the art, for example, it can be understood as polymers, polymers, polymeric substances, polymeric compounds, polymeric macromolecules and the like.

In one aspect, the invention provides a multimeric conjugate comprising: an amino acid polymer as a backbone and an active drug molecule as a branch, wherein the active drug molecule is 7-B A benzyl-10-hydroxycamptothecin which is linked to the amino acid polymer by an amino acid at the 20-hydroxyl group.

其在20-羟基处通过一个氨基酸与所述氨基酸聚合物连接。本发明的发明人发现，若将活性药物分子和氨基酸聚合物直接键合，如此形成的多聚结合物的结构十分不稳定，药物活性分子很容易从氨基酸聚合物上脱落，并且容易发生E环开环，使得该药物活性分子的药效显著降低；但是若将活性药物分子和氨基酸聚合物通过一个氨基酸间接地键合，如此形成的多聚结合物的结构将非常稳定，因而获得令人满意的药效。其中，该氨基酸亦可称为桥接氨基酸，其与活性药物分子和氨基酸聚合物的连接方式可以为：所述氨基酸的一个羧基与活性药物分子的20-羟基连接，且所述氨基酸的一个氨基与氨基酸聚合物连接。通过上述方法，活性药物分子和氨基酸聚合物可以通过一个氨基酸间接地结合在一起。Specifically, the active drug molecule 7-ethyl-10-hydroxycamptothecin is SN38, and its chemical structure is

It is linked to the amino acid polymer by an amino acid at the 20-hydroxyl group. The inventors of the present invention have found that if the active drug molecule and the amino acid polymer are directly bonded, the structure of the thus formed multimeric bond is very unstable, the drug active molecule is easily detached from the amino acid polymer, and the E ring is liable to occur. The ring opening makes the pharmacological effect of the pharmaceutically active molecule significantly lower; however, if the active drug molecule and the amino acid polymer are indirectly bonded through one amino acid, the structure of the thus formed polymer conjugate will be very stable and thus satisfactory. The efficacy of the drug. Wherein, the amino acid may also be referred to as a bridging amino acid, and the method of linking with the active drug molecule and the amino acid polymer may be: one carboxyl group of the amino acid is linked to the 20-hydroxyl group of the active drug molecule, and an amino group of the amino acid is Amino acid polymer linkage. By the above method, the active drug molecule and the amino acid polymer can be indirectly bonded together by one amino acid.

In the present invention, the amino acid polymer may also be referred to as a polyamino acid, wherein the amino acid in the polyamino acid may be selected from a carboxylic acid type amino acid, an amino type amino acid, or a combination thereof, thereby forming a carboxylic acid type polyamino acid, Or a polyamino acid such as an amino polyamino acid. The carboxylic acid type amino acid is selected from the group consisting of aspartic acid (Asp), glutamic acid (Glu), or a combination thereof in any ratio; the amino type amino acid may be selected from the group consisting of ornithine (Orn) and lysine (Lys). , arginine (Arg), hydroxylysine (Hyl), or a combination thereof in any ratio. These amino acids may include their D-type, L-form, and DL hybrid types. Methods for the preparation of polyamino acids are well known to those skilled in the art. For example, US Pat. No. 5,610,264 discloses the synthesis of polyaspartic acid, and S. Roweton et al. (Journal of Environmental Polymer Degradation, Vol. 5, No. 3, 1997: 175). A method for synthesizing polyaspartic acid is disclosed. Similarly, other polyamino acids are also known in many literatures for their synthesis, and by controlling the conditions of these synthetic methods, it is advantageous to obtain a polyamino acid having a desired degree of polymerization, i.e., it is advantageous to obtain a desired molecular weight or molecular weight range. Polyamino acid. Further, pharmaceutically acceptable salts of these polyamino acids, such as a sodium salt (for example, a sodium salt of polyaspartic acid), a hydrochloride (for example, a hydrochloride of polyornithine), or the like can also be used.

According to a preferred embodiment of the present invention, in order to make the active drug molecule and the amino acid polymer easier and more stable, preferably, the monomer of the amino acid polymer is aspartic acid, or aspartate A combination of both acid and glutamic acid. Further, when the monomer of the amino acid polymer contains both aspartic acid and glutamic acid, in order to increase the stability of the polyconjugate, aspartic acid in aspartic acid and glutamic acid The content may be greater than 40 mole percent, such as greater than 50 mole percent, such as greater than 60 mole percent, such as greater than 70 mole percent. It has been surprisingly found that in the case where the amino acid forming the polyamino acid polymer is aspartic acid, or both aspartic acid and glutamic acid are used and the content of glutamic acid is less than 60 mol%, The polyconjugate has excellent stability, especially in which the E ring of SN38 is stable, and it is not easy to open the ring and lose biological activity. In addition, since glutamate is an important excitatory neurotransmitter in the central nervous system, it is the highest content of amino acids in brain tissue. Glutamate acts as a neurotransmitter, and there are two types of ionic and metabotropic forms. Receptors, different types of receptors produce different physiological effects in their specific ways and interact with other regulatory substances. Excessive accumulation of glutamate in the brain can have a neurotoxic effect on the nervous system. Polyglutamate degrades glutamate in the body. When applied in practice, it will bring certain biological toxicity to the body with increasing dose, while polyaspartic acid does not have these defects.

According to the present invention, there is no particular limitation or requirement on the optical rotation of the amino acid of the amino acid polymer as long as it does not affect the basic chemical properties of the amino acid polymer. Preferably, the amino acid of the amino acid polymer is a D-form amino acid, an L-form amino acid, a DL mixed amino acid, or a combination thereof.

According to the present invention, the number average molecular weight of the amino acid polymer is not particularly limited and may be a number average molecular weight of a conventional amino acid polymer in the art. Preferably, the amino acid polymer has a number average molecular weight of from 1000 to 100000 Daltons, such as from 10,000 to 80,000 Daltons, such as from 10,000 to 50,000 Daltons, such as from 10,000 to 40,000 Daltons.

According to the multimeric conjugate of the present invention, the active drug molecule is linked to the amino acid polymer by an amino acid at the 20-hydroxyl group. The manner of linking the amino acid to the active drug molecule and the amino acid polymer is particularly limited as long as the active drug molecule and the amino acid polymer can be stably bridged together. Preferably, one carboxyl group of the amino acid forms an ester bond with the 20-hydroxyl group of the active drug molecule, or an amino group of the amino acid forms an amide bond with a carboxyl group on the amino acid polymer. More preferably, one carboxyl group of the amino acid forms an ester bond with the 20-hydroxyl group of the active drug molecule, and an amino group of the amino acid forms an amide bond with a carboxyl group on the amino acid polymer. More specifically, an amino group and a carboxyl group of the amino acid as described above may be provided by an amino group and a carboxyl group inherent to the amino acid, or may also be provided by an amino group and a carboxyl group in the R group of the amino acid.

According to the present invention, the amount of the active drug molecule bound to each of the amino acid polymers is not particularly limited as long as the amount of the active drug molecule can reach a pharmaceutically reasonable effective amount. In a preferred embodiment, one or more of said active drug molecules are bound to each of said amino acid polymers, said active drug molecule being present in an amount of from 3 to 60% by weight, based on 100% by weight of the polymeric binder. That is, one molecule of the active drug molecule can be bound to each monomer in each amino acid polymer such that one or more of the active drug molecules can be bound to each amino acid polymer. Further, the content of the active drug molecule may be, for example, 10 to 50% by weight, for example 10 to 40%, for example, 10 to 35%, based on 100% by weight of the polyconjugate.

In accordance with the above description of the multimeric conjugates of the invention, in a preferred embodiment, the multimeric conjugates of the invention have the general formula:

among them,

R' is a side chain of a natural amino acid;

n is an integer from 2 to 150;

PA is the amino acid polymer.

According to the present invention, the R' may be a side chain selected from the following amino acids (ie, an R group of an amino acid): glycine, alanine, valine, leucine, isoleucine, valine, styrene Amino acid, tryptophan, methionine, serine, threonine, cysteine, tyrosine, asparagine, glutamine, aspartic acid, glutamic acid, lysine, arginine, group Amino acid. Particularly, when the amino acid is glycine, R' is hydrogen, that is, in the description of the present invention, hydrogen is regarded as a side chain of glycine.

According to the invention, n means the amount of active drug molecule bound to the amino acid polymer. In a preferred embodiment, n is an integer from 2 to 150, such as an integer from 5 to 75, such as an integer from 5 to 60, such as an integer from 5 to 50, such that the amount of active drug molecule is within the ranges described above ( That is, within 3 to 60% by weight).

In another aspect, the present invention also provides a method of preparing the above polyconjugate, comprising the steps of:

1) adding an amino acid polymer, a compound represented by Formula A, a catalyst, and a condensing agent to a solvent;

2) contacting the solution obtained in the step 1) with a poor solvent, and then filtering;

3) adding the filter cake obtained in step 2) to a solvent and contacting the deprotecting agent to obtain a multimeric binder;

Wherein R' is a side chain of a natural amino acid.

Specifically, the amino acid polymer in the step (1), the compound represented by the formula A, the catalyst, and the condensing agent may be added to the solvent in an arbitrary order, and the compound represented by the formula A may be regarded as an intermediate of the active drug molecule SN38. Both the amino acid polymer and the active drug molecule form a multimeric bond by chemical bonding in the presence of a catalyst and a condensing agent. During the reaction, it can be carried out simultaneously with stirring (for example, magnetic stirring), so that the reaction is more sufficient.

According to the present invention, the kind of the solvent is not particularly limited as long as the amino acid polymer and the active drug can be sufficiently dissolved; similarly, the kind of the poor solvent is not particularly limited as long as the reaction product can be precipitated. Preferably, the solvent is an organic solvent; more preferably, the solvents in the step (1) and the step (4) are each independently N,N-dimethylformamide and/or dimethyl sulfoxide; The poor solvent in the step (3) is at least one of water, acetone, methanol, ethanol, isopropanol, diethyl ether, methyl tert-butyl ether, tetrahydrofuran, dichloromethane, and ethyl acetate.

According to the present invention, there is no particular limitation on the kind and amount of the catalyst, and it may be a catalyst which is commonly used in the art. Preferably, the catalyst is pyridine (Py), 4-dimethylaminopyridine (DMAP), triethylamine (TEA), N,N-diisopropylethylamine (DIPEA) and nitrogen methylmorpholine (NMM) At least one of them. More preferably, the molar ratio of the catalyst to the active drug is from 0.01 to 4:1, such as from 0.5 to 2:1, such as from 0.8 to 1.5:1, such as from 1.0 to 1.2:1.

According to the present invention, the kind and amount of the condensing agent are not particularly limited and may be a condensing agent which is commonly used in the art. Preferably, the condensing agent is dicyclohexylcarbodiimide (DCC), diisopropyl At least one of carbodiimide (DIC) and 1-ethyl-(3-dimethylaminopropyl) carbonyldiimide hydrochloride (EDCI). More preferably, the molar ratio of the condensing agent to the active drug is from 0.5 to 4:1, such as from 0.8 to 1.5:1, such as from 1.0 to 1.2:1.

According to the present invention, there is no particular limitation on the kind and amount of the deprotecting agent, and a deprotecting agent which is commonly used in the art. Preferably, the deprotecting agent is at least one of tetrabutylammonium fluoride, tetraethylammonium fluoride, tetramethylammonium fluoride, and hydrogen fluoride pyridine solution.

According to the present invention, the method of preparing the above-described multimeric binder may further include various common post-treatment processes in the art. Preferably, the method further comprises: 4) separating and extracting the resulting multimeric conjugate. More preferably, the separation extraction is carried out by dialysis or ultrafiltration. This method can easily remove unbound free active drug. Such dialysis or ultrafiltration separation methods are well known to those skilled in the art.

In the method of the present invention, the condensing agent DCC and the catalyst DMAP reaction conditions can be used to condense the free carboxyl group of the carboxylic acid type polyamino acid such as polyaspartic acid with the amino group in the molecular structure of the active pharmaceutical intermediate, thereby forming the active drug SN38 and A polyconjugate of a polyamino acid that is chemically bonded. Other methods known to those skilled in the art that can chemically bond the active drug of the present invention to the amino acid polymer can also be used in the present invention.

It has been found that the multimeric conjugates produced by the process of the invention have a significantly higher solubility in solubility compared to their corresponding active drugs. For example, by the method of the present invention, the solubility of the SN38 multimeric conjugate obtained by the present invention (in terms of SN38) is significantly higher than that of the SN38 original drug.

In another aspect, the present invention also provides a pharmaceutical composition comprising the above-described multimeric composition, and a pharmaceutically acceptable excipient.

Wherein, the pharmaceutically acceptable excipient may be an optional pharmaceutically acceptable excipient, which may be based on the dosage form of the pharmaceutical composition and other various needs, and is polymerized based on the total weight of the pharmaceutical composition. The content of the composition may also be determined according to actual needs, for example, from 1 to 99% by weight.

A pharmaceutical composition according to the present invention, which may be in the form of any pharmaceutical preparation in the pharmaceutical industry, such as, but not limited to, an oral preparation such as a tablet, a capsule, a granule, an oral solution, an oral suspension, etc., an injection preparation For example, small-volume injections, large-volume injections, sterile powder preparations (such as freeze-dried powder injections, etc.), inhaled preparations such as nasal inhalation preparations (for example, inhalation powders or inhalation solutions) Liquid agent).

Furthermore, the choice of the excipients may depend on the particular form of formulation presented by the pharmaceutical composition. For example, when the pharmaceutical composition is a tablet, a capsule or a granule or the like, it may optionally include a diluent (for example, starch, lactose, microcrystalline cellulose, etc.), a disintegrating agent (for example, carboxymethyl cellulose). Sodium, sodium carboxymethyl starch, low-substituted hydroxypropyl cellulose, etc.), binders (such as hydroxypropyl methylcellulose, polyvinylpyrrolidone, polyethylene glycol, etc.), lubricants or glidants (eg Magnesium stearate, talc, stearic acid, colloidal silica, etc.). Flavoring agents, coloring agents, coating agents and the like may also optionally be included.

Further, for example, when the pharmaceutical composition is a small-volume injection solution or a large-volume injection solution or the like, a solvent such as water, ethanol, glycerin, propylene glycol, polyethylene glycol having a molecular weight of 200 to 600, or the like may be contained therein, and optionally The ground contains an osmotic pressure adjusting agent such as sodium chloride or glucose.

For another example, when the pharmaceutical composition is a sterile powder preparation or the like, an excipient such as mannitol, glucose, dextran, sodium chloride, sucrose or lactose may be contained therein.

The formulation and preparation of these pharmaceutical compositions of the invention in the form of pharmaceutical preparations can be accomplished by methods and experience well known to those skilled in the art. For example, the tablet can be prepared by a method such as wet granulation tableting, dry granulation tableting, powder direct compression, or the like. Further, for example, for a freeze-dried powder injection, an appropriate amount of mannitol may be added according to the requirements of the preparation and dissolved in water for injection, followed by lyophilization.

In a preferred embodiment, the pharmaceutical composition is a pharmaceutical composition for injection. More preferably, the pharmaceutically acceptable excipient comprises water for injection, 0.8-1.2% by weight of sodium chloride injection, and 3 to 7% by weight of glucose injection.

In another aspect, the invention also provides the use of the above polymeric compositions and pharmaceutical compositions for the treatment of tumors and cancer.

Preferably, the tumor or cancer is at least one of colorectal cancer, pancreatic cancer, breast cancer, ovarian cancer, lung cancer, kidney cancer, stomach cancer, and liver cancer. In one embodiment, the tumor or cancer is colorectal cancer. In one embodiment, the multimeric conjugate is for use in combination with 5-fluorouracil and folinic acid in the treatment of patients with advanced colorectal cancer who have not received chemotherapy; or the multimeric conjugate is used as a single drug, 5-fluorouracil chemotherapy regimen for patients who failed treatment.

Any of the embodiments of any of the aspects of the invention may be combined with other embodiments as long as they do not contradict each other. Furthermore, in any of the embodiments of any of the aspects of the invention, any of the technical features may be applied to the technical features in other embodiments as long as they do not contradict each other.

Any of the technical features of any of the aspects of the invention or any one of the aspects of the invention are equally applicable to any one of any other embodiment or any other aspect, as long as they do not contradict each other, of course When applicable, the corresponding features may be appropriately modified as necessary. Various aspects and features of the present invention are further described below.

All documents cited in the present invention are hereby incorporated by reference in their entirety, and if the meanings expressed by these documents are inconsistent with the present invention, the expression of the present invention shall prevail. Moreover, the various terms and phrases used in the present invention have the ordinary meanings well known to those skilled in the art, and even though the present invention is intended to provide a more detailed description and explanation of the terms and phrases herein, such terms and phrases are Inconsistent with the well-known meaning, the meaning expressed in the present invention shall prevail.

In the present invention, these polyamino acids are also commercially available, and for example, polyamino acids having a desired degree of polymerization or molecular weight or molecular weight range can be easily obtained from Sigma-Aldrich, for example, products available from Sigma-Aldrich. Poly-L-aspartic acid having a molecular weight of 16,000 to 50,000 Daltons of P6762. In the present invention, the polyamino acid used is commercially available, unless otherwise stated.

The SN38 used in the present invention can be easily obtained commercially, for example, from Sigma-Aldrich Co., Ltd., or from Hubei Yuancheng Pharmaceutical Co., Ltd. In the present invention, SN38 used was purchased from the latter unless otherwise specified.

In one embodiment of the invention, the present invention overcomes SN38 and investigates the design of a novel polymeric SN38 multimeric conjugate of a polymeric macromolecule in combination with a camptothecin such as SN38.

In one embodiment of the multimeric combination of the invention, a plurality of SN38 can be combined on one polymeric macromolecule.

SN38 has similar biological activity as irinotecan. Irinotecan is a derivative of semi-synthetic camptothecin and is an antitumor drug that specifically inhibits DNA topoisomerase I. It is metabolized by a carboxylesterase to SN38 in most tissues, while the latter acts on purified topoisomerase I more strongly than irinotecan and is also highly cytotoxic to several murine and human tumor cell lines. In irinotecan. SN38 or irinotecan can induce single-strand DNA damage, thereby blocking the DNA replication fork, thereby producing cytotoxicity. This cytotoxicity is time-dependent and specifically acts on the S phase.

In vitro, irinotecan and SN38 were not found to be efficiently recognized by P-glycoprotein [sup]MDR[/sup], and cytotoxicity was observed in cell lines resistant to doxorubicin and vinblastine. effect.

In addition, in vivo experiments, irinotecan showed a broad spectrum of anti-tumor activity in the murine tumor model (P03 pancreatic ductal adenocarcinoma, MA-16/C breast cancer, C38 and C51 colon adenocarcinoma) and anti-human xenograft Tumor activity (Co-4 colon adenocarcinoma, MX-1 breast cancer, St-15 and SC-6 gastric adenocarcinoma), irinotecan on tumors expressing P-glycoprotein [sup]MDR[/sup] Alkali and doxorubicin-resistant P388 leukemia) also have anti-tumor activity.

In addition to its anti-tumor activity, the most relevant pharmacological action is to inhibit acetylcholinesterase.

After using this drug, the intensity of major side effects (such as leukopenia and diarrhea) is related to the area under the curve of the parent drug and its metabolite SN38. In monotherapy, hematologic toxicity (leukocyte and neutrophil decreased to the lowest point) or the extent of diarrhea was significantly associated with the area under the curve of irinotecan and its metabolite SN38. The pharmacokinetic properties of irinotecan and SN38 (active metabolites) were studied in Phase I clinical trials. Sixty patients received a recommended dose regimen, ie 30-minute intravenous infusion of this product 100-750 mg /m ² . The kinetics of irinotecan is non-dose dependent. Patients enrolled in clinical studies received different irinotecan dosing regimens with similar pharmacokinetics. Its plasma decay mode is both two-compartment and three-chamber. The mean plasma half-life in the three-compartment model was 12 minutes in the first phase, 2.5 hours in the second phase, and 14.2 hours in the final phase. At the end of the infusion of the recommended dose of 350 mg/m ² , irinotecan and SN38 reached plasma peak concentrations of 7.7 μg/mL and 56 μg/mL, respectively, and the area under the curve was 34 μg/h/mL, 451 μg/h/ mL, its steady-state distribution volume is large and remains relatively stable as a function of dose, averaging 157 L/m ² . The average body clearance rate averaged 15 L/h/m ² and remained stable during the different treatment cycles of the same patient. SN38 has a large variation in its drug metabolism parameters in different individuals. The 24-hour average urinary excretion rates of irinotecan and SN38 were 19.9% and 0.25%, respectively, of the dose used. Phase II clinical trials on the pharmacokinetics of irinotecan were performed in 72 patients with cancer. The pharmacokinetic parameters calculated by the restricted sampling model are very close to those of the Phase I study. In vitro, the plasma protein binding rates of irinotecan and SN38 were 65% and 95%, respectively. When the patient's bilirubin is 1.5-3 times the upper limit of normal, the irinotecan clearance is reduced by about 40%. When irinotecan was administered at 200 mg/m ² in these patients, the plasma drug concentration was the same as that of irinotecan 350 mg/m ² in cancer patients with normal liver function. In combination with 5-fluorouracil/leucovorin, the pharmacokinetic properties of irinotecan are not altered.

In one embodiment of the invention, the multimeric conjugate of the invention is obtained by combining a carboxylic acid type polyamino acid with SN38. For example, it is composed of a carboxylic acid on a polyamino acid molecule and a 20-hydroxyl group on the SN38 molecule via a bridged amino acid to form an ester bond. For example, polyaspartic acid and SN38 through glycine Taking the formed multimeric binder as an example, an exemplary chemical bonding mode is:

Wherein PA represents polyaspartic acid and R' is hydrogen (i.e., the bridging amino acid is glycine), and the size or range of n can be easily adjusted depending on the ratio of the feed and the control reaction conditions.

The amino group of the SN38 intermediate can be reacted with the carboxyl group of polydeptonate in a chemical condensation reaction using DCC as a condensing agent, and the H ₂ O molecule is removed from the _two molecules to form an amide bond (-CON-). A plurality of free carboxyl groups on the amino acid peptide chain may be bonded to a desired amount of the SN38 intermediate according to specific conditions of the condensation reaction (for example, the amount of the feed, the reaction temperature, the reaction time, etc.), for example, the obtained polyamino acid and the SN38 intermediate. In a two-part polymeric combination, the weight of the SN38 intermediate may range from 3 to 60% by weight of the multimeric binder (eg, 10-50%, such as 10-40%, such as 10-35%), followed by The resulting multimeric conjugate is reacted with a basic material to form a pharmaceutically acceptable salt, for example to form a sodium salt. Polyconjugates formed by chemical bonding rather than physical bonding (e.g., hydrogen bonding) have been confirmed in the following by the ultraviolet and HPLC methods. Typical examples of the above carboxylic acid type polyamino acid include, but are not limited to, D-aspartic acid (Asp), L-aspartic acid, DL-aspartic acid, and combinations thereof, such as aspartame in any configuration. A carboxylic acid type polyamino acid in which an acid and any configuration of glutamic acid are polymerized in an arbitrary ratio, for example, in a mixing ratio of 1-99%.

In one embodiment of the present invention, the multimeric conjugate of the present invention is obtained by combining a carboxypolyamino acid with SN38. For example, it is formed by a free carboxyl group on a polyamino acid molecule by bridging an amino acid with a 20-hydroxy group on the SN38 molecule under the action of a reagent.

In the method of preparing the substance of the present invention, various raw materials used for the reaction can be prepared by those skilled in the art based on the prior art, or can be obtained by a method known in the literature, or can be commercially obtained. The intermediates, starting materials, reagents, reaction conditions and the like used in the reaction scheme of the present invention can be appropriately changed according to the knowledge of those skilled in the art.

The polyconjugate of the present invention can be used in combination with other active ingredients as long as it does not produce other For example, it does not produce an allergic reaction.

The multimeric conjugates of the invention may be used as the sole drug or may be used in combination with one or more other agents which have a synergistic and/or synergistic effect on the substance of the invention. Combination therapy can be achieved by administering the individual therapeutic components simultaneously, sequentially or separately.

The term "pharmaceutical composition" as used herein is meant to include a product comprising specified amounts of each of the specified ingredients, as well as any product produced directly or indirectly from a specified amount of each specified combination of ingredients. In the present invention, the term "pharmaceutical composition" can be used interchangeably with "composition."

The actual dosage level of each active ingredient in the pharmaceutical compositions of the present invention can be varied so that the resulting active mass is effective to achieve the desired therapeutic response for a particular patient, compound, and mode of administration. The dosage level will be selected based on the activity of the particular active substance, the route of administration, the severity of the condition being treated, and the condition and past medical history of the patient to be treated. However, it is the practice in the art that the dosage of the active substance be started from a level lower than that required to achieve the desired therapeutic effect, and the dosage is gradually increased until the desired effect is obtained.

A therapeutically and/or prophylactically effective amount of a multimeric conjugate of the invention may be applied in pure form when used in the treatment and/or prevention or other treatment and/or prophylaxis of the invention described above. Alternatively, the multimeric conjugate can be administered as a pharmaceutical composition comprising the multimeric conjugate of interest and one or more pharmaceutically acceptable excipients. The term "therapeutically and/or prophylactically effective amount" of a multimeric conjugate of the invention refers to a sufficient amount of a multimeric conjugate to treat a disorder with a reasonable effect/risk ratio suitable for any medical treatment and/or prevention. It will be appreciated, however, that the total daily usage of the multimeric conjugates and pharmaceutical compositions of the present invention will be determined by the attending physician within the scope of sound medical judgment. The specific therapeutically effective dose level for any particular patient will depend on a number of factors, including the disorder being treated and the severity of the disorder; the specific polyconjugate used; the specific Pharmaceutical composition; age, weight, general health, sex and diet of the patient; time of administration, route of administration and excretion rate of the particular pharmaceutical composition employed; duration of treatment; use in combination with the pharmaceutical composition employed Or other drugs used at the same time; and similar factors well known in the medical field. For example, it is art practice that the dosage of the pharmaceutical composition begins at a level below that required to achieve the desired therapeutic effect, gradually increasing the dosage until the desired effect is achieved. In general, the dosage of the multimeric conjugate of the present invention for use in mammals, particularly humans, may range from 0.0001 to 1000 mg/kg body weight per day, for example from 0.001 to 500 mg per day, based on the amount of the camptothecin compound SN38. Kg body weight/day, for example between 0.001 and 100 mg/kg body weight/day.

Pharmaceutical compositions containing an effective amount of a multimeric conjugate of the invention can be prepared using pharmaceutical carriers well known to those skilled in the art. The invention therefore also provides a pharmaceutical composition comprising a multimeric conjugate of the invention formulated together with one or more non-toxic pharmaceutically acceptable carriers. The pharmaceutical compositions may be specially formulated for oral administration in solid or liquid form for parenteral injection or for rectal administration.

The pharmaceutical composition may be formulated into a plurality of dosage forms for ease of administration, for example, oral preparations (such as tablets, capsules, solutions or suspensions); injectable preparations (such as injectable solutions or suspensions, Or an injectable dry powder, which can be used immediately before injection. The carrier in the pharmaceutical composition includes: a binder for oral preparation (such as starch, usually corn, wheat or rice starch, gelatin, methylcellulose, sodium carboxymethylcellulose, and/or polyvinylpyrrolidone) , diluent, lubricant (such as silica, talc, stearic acid or a salt thereof, usually magnesium stearate or calcium stearate, and / or polyethylene glycol), and if necessary, also contains disintegration Such as starch, agar, alginic acid or a salt thereof, usually sodium alginate, and / or effervescent mixture, cosolvents, stabilizers, suspending agents, coloring agents, flavoring agents, etc., antiseptic for injectable preparations Agents, solubilizers, stabilizers, etc.; bases, diluents, lubricants, etc. for topical preparations. The pharmaceutical preparations can be administered orally or parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or topically), and if certain drugs are unstable under gastric conditions, they can be formulated into enteric coated tablets.

More specifically, the pharmaceutical compositions of the present invention can be administered orally, rectally, parenterally, intracereally, intravaginally, intraperitoneally, topically (e.g., by powder, ointment or drops), buccally to humans and other mammals. Or as an oral spray or nasal spray. The term "parenteral" as used herein, refers to a mode of administration including intravenous, intramuscular, intraperitoneal, intrasternal, subcutaneous, and intraarticular injections and infusions.

Pharmaceutical compositions suitable for parenteral injection may include physiologically acceptable sterile aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, and steril Powder. Examples of suitable aqueous or nonaqueous vehicles, diluents, solvents or vehicles include water, ethanol, polyols (such as propylene glycol, polyethylene glycol, glycerol, etc.), vegetable oils (such as olive oil), injectable organic esters such as oils. Ethyl acetate and suitable mixtures thereof.

These pharmaceutical compositions may also contain adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. The action of microorganisms can be ensured by various antibacterial and antifungal agents, such as parabens, chlorobutanol, phenol, sorbic acid and the like. It is also desirable to include isotonic agents, for example, sugars, sodium chloride, and the like. Prolonged absorption of the injectable pharmaceutical form can be brought about through the use of materials which delay absorption, such as aluminum monostearate and gelatin.

Suspensions may contain suspending agents in addition to the active compound, such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol and polyoxyethylene sorbitan ester, microcrystalline cellulose, aluminum metahydroxide, bentonite , agar and tragacanth or a mixture of these substances.

In some cases, in order to prolong the action of the drug, it is desirable to slow the absorption of the drug by subcutaneous or intramuscular injection. This can be achieved by using a liquid suspension of poorly water-soluble crystals or amorphous materials. Thus, the rate of absorption of the drug depends on its rate of dissolution, which in turn may depend on crystal size and crystalline form. Alternatively, delayed absorption of a pharmaceutical form for parenteral administration is accomplished by dissolving or suspending the drug in an oil vehicle.

Injectable depot formulations are prepared by forming microencapsule matrices of the drug in biodegradable polymers such as polylactide-polyglycolide. The rate of drug release can be controlled based on the ratio of drug to polymer and the nature of the particular polymer employed. Examples of other biodegradable polymers include polyorthoesters and polyanhydrides. Injectable depot formulations are also prepared by embedding the drug in liposomes or microemulsions which are compatible with body tissues.

The injectable preparation can be sterilized, for example, by filtration with a bacteriophage or by incorporating a sterilizing agent in the form of a sterile solid composition which can be dissolved or dispersed in sterile water or other sterilized form before use. Injectable medium.

The compound of the present invention or a pharmaceutical composition thereof can be administered orally or parenterally. Oral administration may be a tablet, a capsule, or a coating, and an enteral preparation may be an injection or a suppository. These formulations are prepared according to methods familiar to those skilled in the art. The excipients used in the manufacture of tablets, capsules, and coatings are conventional excipients such as starch, gelatin, gum arabic, silica, polyethylene glycol, solvents for liquid dosage forms such as water, ethanol, propylene glycol, vegetable oils (such as corn). Oil, peanut oil, olive oil, etc.). There are other adjuvants in the formulation containing the compound of the present invention, such as surfactants, lubricants, disintegrants, preservatives, flavoring agents, and pigments. The dose containing the multimeric conjugate of the present invention in tablets, capsules, coatings, injections and suppositories is calculated as the amount of camptothecin-like compound SN38 present in the unit dosage form. The active compound of the invention is generally present in the unit dosage form in an amount of from 0.01 to 5000 mg, preferably in a unit dosage form containing from 0.1 to 500 mg, more preferably in a unit dosage form containing from 1 to 500 mg. In particular, solid dosage forms for oral administration that can be provided by the present invention include capsules, tablets, pills, powders, and granules. In such solid dosage forms, the active compound may be mixed with at least one inert pharmaceutically acceptable excipient or carrier such as sodium citrate or dicalcium phosphate and/or the following: a) filler or extender; b) Bonding Agents such as carboxymethylcellulose, alginate, gelatin, polyvinylpyrrolidone, sucrose and gum arabic; c) humectants such as glycerin; d) disintegrants such as agar, calcium carbonate, potato or tapioca, alginic acid, certain Some silicates and sodium carbonate; e) solution retarders such as paraffin; f) absorption accelerators such as quaternary ammonium compounds; g) wetting agents such as cetyl alcohol and glyceryl monostearate; h) adsorbents such as kaolin and bentonite And i) lubricants such as talc, calcium stearate, magnesium stearate, solid polyethylene glycol, sodium lauryl sulfate, and mixtures thereof. In the case of capsules, tablets and pills, buffers may also be included in the dosage form.

Solid compositions of a similar type may be used as fillers in soft and hard capsules using such excipients as lactose and high molecular weight polyethylene glycols and the like.

The solid dosage forms of tablets, dragees, capsules, pills, and granules can be prepared with coatings and shells such as enteric coatings and other materials known in the art of pharmaceutical preparations. These solid dosage forms may optionally contain opacifying agents and may be formulated such that they are only or preferentially released in a certain portion of the intestinal tract in a delayed manner. Examples of the embedding composition that can be used include high molecular substances and waxes. If appropriate, the active substance may also be formulated in microencapsulated form with one or more of the above-mentioned excipients.

Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs. The liquid dosage form may contain, in addition to the active substance, an inert diluent commonly used in the art, such as water or other solvents, solubilizers and emulsifiers such as ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate. Ester, propylene glycol, 1,3-butanediol, dimethylformamide, oils (especially cottonseed oil, peanut oil, corn oil, germ oil, olive oil, castor oil and sesame oil), glycerin, tetrahydrofurfuryl alcohol, poly Fatty acid esters of ethylene glycol and sorbitan and mixtures thereof. The oral compositions may contain, in addition to inert diluents, excipients such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and flavoring agents.

Compositions for rectal or vaginal administration are preferably suppositories. Suppositories can be prepared by mixing the multi-conjugates of the invention with a suitable non-irritating excipient or carrier such as cocoa butter, polyethylene glycol or suppository wax, which are solid at room temperature but liquid at body temperature Thus, it can be melted in the rectal or vaginal cavity to release the active compound.

The multimeric conjugates of the invention and pharmaceutical compositions thereof are also contemplated for topical administration. Dosage forms for topical administration of the multimeric conjugates of the invention include powders, sprays, ointments and inhalants. The active substance is mixed under sterile conditions with a pharmaceutically acceptable carrier and any required preservative, buffer or propellant. Ophthalmic formulations, eye ointments, powders and solutions are also contemplated as being within the scope of the invention.

The multimeric conjugates of the invention may also be administered in the form of liposomes. Liposomes are known as known in the art It is usually made from phospholipids or other lipids. Liposomes are formed from single or multiple layers of hydrated liquid crystal dispersed in an aqueous medium. Any non-toxic, physiologically acceptable and metabolizable lipid capable of forming liposomes can be used. The compound of the present invention in the form of a liposome may contain, in addition to the compound of the present invention, a stabilizer, a preservative, an excipient or the like. Preferred lipids are natural and synthetic phospholipids and phosphatidylcholines (lecithins), which may be used alone or together. Methods of forming liposomes are well known in the art. See, for example, Prescott, Ed., Meth SN 38s in Cell Biology, Volume XIV, Academic Press, New York, N. Y. (1976), p.

The inventors have surprisingly found that the multimeric conjugates of the invention exhibit satisfactory beneficial effects both biologically and/or physically and/or chemically.

The invention is further illustrated by the following specific examples and test examples, but it should be understood that these examples are only intended to be more specific and not to be construed as limiting the invention in any way.

The present invention provides a general and/or specific description of the materials and test methods used in the tests. While many of the materials and methods of operation used to accomplish the objectives of the present invention are well known in the art, the present invention is still described in detail herein. It will be apparent to those skilled in the art that, hereinafter, the materials and methods of operation of the present invention are well known in the art unless otherwise specified.

A. Method for testing polyconjugates

Test Method 1: UV-Vis Spectrophotometry

UV-visible spectrophotometry was performed on the SN38, polyaspartic acid, and polyaspartic acid-SN38 polyconjugates. The results showed that polyaspartic acid would not be more than 250-400 nm. The absorbance of the polyconjugate or SN38 contributes, and in this region the polyaspartate-SN38 polyconjugate binds to the typical absorption peak wavelength of both the prodrug SN38. These results indicate that the three materials have their unique spectral behavior and spectral contributions under spectrophotometry, making it possible to legally and quantitatively analyze three substances by HPLC.

In addition, the inventors have found in further experiments that the polyamino acids obtained by the combination of aspartic acid-glutamic acid have similar properties to the above-mentioned polyaspartic acid, SN38 and their polyconjugates for SN38. The UV-visible spectrophotometric spectral scanning features, the three types of substances have their unique spectral behavior and spectral contribution under spectrophotometry, making it possible to legally and quantitatively analyze three types of substances by HPLC.

Test Method 2: High Performance Liquid Chromatography (HPLC)

Column: Aijieer Venusil XBP C44.6*250mm, 5μm, 300A

Protective column core: Philomon C4

Mobile phase A: 5 mM Na ₂ HPO ₄ aqueous solution, adjusted to pH 7.0

Mobile phase B: acetonitrile

Column temperature: 40 ° C

Flow rate: 1ml/min

Detector: DAD (measured at 266 nm wavelength when measuring SN38 and its polyconjugate)

Sample concentration: 1mg/ml

Injection volume: 10μl

Gradient elution conditions:

T(min) A (%) B (%) 0 95 5 5 95 5 6 80 20 15 80 20 30 5 95 35 5 95 35.01 95 5 40 95 5

According to the above HPLC method, the following four substances and their chromatographic behavior were tested: (A) SN38, (B) polyaspartic acid, (C) polyaspartic acid-SN38 polyconjugate, (D) poly-binding Mixture of free material with free SN38.

The results show:

(A) High-performance liquid chromatogram of SN38 (25 μg/ml) showing a peak at about 18.3 min of SN38;

(B) High-performance liquid chromatogram of polyaspartic acid (PA, 75 μg/ml) showed a small chromatographic peak at about 5.2 min, and the peak of the chromatogram was small because the PA absorption was weak at the detection wavelength of 254 nm;

(C) Polyaspartic acid-SN38 polyconjugate (PA-SN38, product of Preparation 1, 100 μg/ml, A high-performance liquid chromatogram containing the concentration of SN38 corresponding to the concentration of SN38 in the above (A) SN38 sample solution, a small peak was detected at about 18.3 min (free SN38 < 0.15%), but at about 5.2. A peak with a peak area corresponding to that in (A) is shown at min. This indicates that the PA-SN38 polyconjugate has substantially no free SN38 (the area normalization method has a peak area percentage of less than 0.15%, ie, free SN38 < 0.15%), and the method of the present invention can obtain a high purity PA. - SN38 poly-conjugate (purity greater than 99% in terms of SN38); while PA-SN38 is chemically bonded to form a stable compound, cannot be separated under chromatographic conditions, and due to the strong polarity of the polyamino acid, the PA-SN38 The chromatographic retention behavior of the multimeric binder is close to that of PA and peaks at PA. In addition, the percentage of SN38 in the multimer conjugate, that is, the active drug content, can be calculated by the HPLC method, and the active drug content of the preparation example 1 is 23.1% by weight;

(D) High performance liquid chromatogram of polyaspartic acid-SN38 polyconjugate and free SN38 physical mixture (PA-SN38, 100 μg/ml; SN38, 25 μg/ml) detected at approximately 18.4 min The SN38 peak corresponding to the amount of SN38 added, and the PA-SN38 peak corresponding to the amount of PA-SN38 added was detected at about 5.2 min. This indicates that the PA-SN38 polyconjugate has independent chromatographic behavior with both free SN38, and the two do not interfere with each other.

(E) The (C) polyaspartic acid-SN38 multimeric conjugate sample was allowed to stand at 60 ° C for 10 hours while being measured, and the results showed that the PA-SN38 peak detected at about 5.2 min became small, and was about A peak appeared at 4.5 min, and the peak of the E ring opening degradation of SN38 in the polyconjugate was detected by liquid-mass spectrometry.

The above HPLC method can also be extended to poly-conjugates of SN38 and other amino acid polymers such as poly(aspartic acid-glutamic acid) polymer, even without substantially changing the HPLC conditions. Qualitative and quantitative analysis of the polyconjugate (the main peak at about 5.2 min and the E ring open-loop degradation product at about 4.5 min will deviate accordingly). This deviation is mainly due to the difference in the amino acid polymer used. , does not affect the measurement results).

B. Preparation of the poly-conjugate

Preparation Example 1: Chemical Synthesis of Polyaspartic Acid-Glycine-SN38 Polyconjugate

Poly-DL-aspartic acid (20000 Da) was obtained by a known method, and this was prepared as an amino acid polymer. Glycine is used as a bridging amino acid. The schematic synthetic route is as follows:

(polyaspartic acid-SN38 polyconjugate)

Synthesis step: 800.0 mg of poly-L-aspartic acid (molecular weight 28000 Da) was taken and dissolved in 5 ml of N,N-dimethylformamide. Further, 206.7 mg of the intermediate 03 was added to the above N,N-dimethylformamide solution, and the intermediate 03 was dissolved by stirring. Another 62.5 mg of condensing agent DCC and 37.0 mg of catalyst DMAP were added to the above N,N-dimethylformamide solution, and magnetically stirred at room temperature (23-25 ° C) for 12 hours. The solution was added to poor solvent water, stirred for 30 minutes, filtered, and the filter cake was dissolved in DMF, and 156.7 mg of deprotecting agent tetrabutylammonium fluoride was added thereto, and stirring was continued for 6 hours. 100 ml of 0.5 M NaHCO ₃ solution was added to the above reaction solution, magnetically stirred for 1 hour, and the above alkaline solution was poured into a dialysis bag, and after dialysis against deionized water for 45 hours, the solution was passed through a 0.2 μm filter membrane, frozen and Drying gave 820.4 mg of polyaspartic acid-SN38 polyconjugate.

The hydrogen nuclear magnetic resonance (1H-NMR) spectrum (not shown) of the polyaspartic acid-SN38 polyconjugate obtained in the present preparation was determined to have a line corresponding to the polyconjugate.

The carbon nuclear magnetic resonance (13C-NMR) spectrum (not shown) of the polyaspartic acid-SN38 polyconjugate obtained in this Preparation Example was determined to have a line corresponding to the polyconjugate.

A high performance liquid chromatography (HPLC) UV detection (not shown) of the polyaspartic acid-SN38 polyconjugate obtained in this Preparation was determined to be consistent with the expected results.

The high-performance liquid chromatography (HPLC) mass spectrometry (MS) map of the polyaspartic acid-SN38 polyconjugate obtained in this Preparation Example was determined to show some fragments formed by SN38 (not shown). This indicates that SN38 is present in the multimeric binder and elutes in the HPLC system along with the amino acid polymer portion.

The final product obtained in this preparation example was determined to have a bound active drug content of 23.1% by weight and a free active drug content of <0.15% (the peak area of the SN38 peak in the HPLC chart divided by the peak area of the PA-SN38 peak was multiplied by 100% of the percentage obtained is calculated).

Preparation 2: Chemical synthesis of polyaspartic acid-glycine-SN38 polyconjugate

Refer to the method of Preparation Example 1, except that the amino acid polymer was changed to poly-D-aspartic acid (molecular weight: 5000 Da), glycine was used as the bridging amino acid, acetone was used as the poor solvent, TEA was used as the catalyst, and DIC was used as the condensing agent. Tetraethylammonium fluoride was used as a deprotecting agent. The resulting multimeric conjugate was 85.8% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 17.9% by weight and a free active drug content of <0.18%.

Preparation Example 3: Chemical Synthesis of Polyaspartic Acid-Glycine-SN38 Polyconjugate

Refer to the method of Preparation Example 1, except that the amino acid polymer was changed to poly-L-aspartic acid (molecular weight 100,000 Da), glycine was used as the bridging amino acid, DIPEA was used as the catalyst, EDCI was used as the condensing agent, and tetramethyl fluorination was used. Ammonium acts as a deprotecting agent. The resulting multimeric conjugate was 88.3% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 31.1% by weight and a free active drug content of <0.25%.

Preparation Example 4: Chemical Synthesis of Poly(Glutamic Acid/Aspartate)-Glycine-SN38 Polyconjugate

Refer to the method of Preparation Example 1, except that the amino acid polymer is changed to poly(glutamic acid/aspartic acid) (glutamic acid is 35% of the total amino acid molar amount, molecular weight is 20,000 Da), and glycine is used as the bridging amino acid, and methanol is used. As a poor solvent, a hydrogen fluoride pyridine solution was used as a deprotecting agent. The resulting multimeric conjugate was 82.4% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 24.5% by weight and a free active drug content of <0.22%.

Preparation Example 5: Chemical Synthesis of Poly(Glutamic Acid/Aspartic Acid)-Glycine-SN38 Polyconjugate

Refer to the method of Preparation Example 1, except that the amino acid polymer was changed to poly(glutamic acid/aspartic acid) (glutamic acid accounted for 23.5% of the total amino acid molar amount, molecular weight 20000 Da), glycine was used as the bridging amino acid, and ethanol was used. As a poor solvent. The resulting multimeric conjugate was 82.4% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 24.5% by weight and a free active drug content of <0.22%.

Preparation Example 6, Chemical Synthesis of Poly(Glutamic Acid/Aspartic Acid)-Glycine-SN38 Polyconjugate

Refer to the method of Preparation Example 1, but the amino acid polymer is changed to poly(glutamic acid/aspartic acid) (Valley The amino acid accounts for 35.8% of the total amino acid molar amount, and the molecular weight is 20,000 Da). Glycine is used as the bridging amino acid, and isopropyl alcohol is used as the poor solvent. The resulting multimeric conjugate was 84.8% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 18.7% by weight and a free active drug content of <0.20%.

Preparation Example 7, Chemical Synthesis of Amino Acid Polymer-Bridged Amino Acid-SN38 Multimeric Conjugate

Reference was made to the method of Preparation Example 1, except that the bridged amino acid glycine was changed to lysine, and diethyl ether was used as a poor solvent. The resulting multimeric conjugate was 86.3% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 15.9% by weight and a free active drug content of <0.15%.

Preparation Example 8, Chemical Synthesis of Amino Acid Polymer-Bridged Amino Acid-SN38 Polyconjugate

Referring to the method of Preparation 1, but the amino acid polymer was changed to poly(glutamic acid/aspartic acid) (glutamic acid accounted for 15.5% of the total amino acid molar amount, molecular weight 10000 Da), and the bridging amino acid glycine was changed to glutamic acid. Methyl tert-butyl ether was used as a poor solvent. The resulting multimeric conjugate was 84.3% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 28.4% by weight and a free active drug content of <0.15%.

Preparation Example 9, Chemical Synthesis of Amino Acid Polymer-Bridged Amino Acid-SN38 Multimeric Conjugate

Referring to the method of Preparation 1, but the amino acid polymer was changed to poly(glutamic acid/aspartic acid) (glutamic acid accounted for 7.5% of the total amino acid molar amount, molecular weight 25000 Da), and the bridged amino acid glycine was changed to alanine. Tetrahydrofuran was used as a poor solvent. The resulting multimeric conjugate was 87.3% based on the small molecule active drug. The obtained product was determined to have a bound active drug content of 32.7% by weight and a free active drug content of <0.15%.

Preparation Example 10: Chemical Synthesis of Polyaspartic Acid-Glycine-SN38 Polyconjugate

Reference is made to the method described in the description of Example 18 of CN1429121A, except that the amino acid polymer used in Preparation Example 1 of the present invention is used, respectively, to obtain a sample of the multimeric binder. The resulting multimeric conjugates are all in the range of 80-90% based on the small molecule active drug. The obtained product was determined to have a combined active drug content in the range of 10-30%, and the free active drug content was less than 0.5%. The product obtained in this example has excellent E-ring stability, indicating that the technical effects of the present invention can also be obtained by using the precursor of the present invention in combination with the multi-component obtained by the different processes in the prior art.

All of the multimeric conjugates obtained in Preparation Example 1 - Preparation Example 10 above can be represented by the following formula, according to the bound active pharmaceutical content, the theoretical amount of each substance, the unbound SN38, and the unbound ammonia. The calculation of the parameters of the base acid polymer, etc., can be calculated that their n values are in the range of 5-75:

Preparation Example 11, Preparation of Amino Acid Polymer-SN38 Polyconjugate

Reference is made to the methods of Preparation Examples 1-10 and the prior art methods (for example, the method described in Chinese Patent No. CN104721830A), except that the amino acid polymer is directly linked to the hydroxyl group at position 20 of SN38 without using an intermediate bridge. Amino acid, 10 samples of multimeric conjugates were obtained. The resulting multimeric conjugates are all in the range of 80-90% based on the small molecule active drug. The obtained product was determined to have a combined active drug content in the range of 10-30%, and the free active drug content was less than 0.5%.

Preparation Example 12, Preparation of Polyconjugate

Reference was made to the methods of Examples 1-3, respectively, except that the amino acid polymer used was polyglutamic acid, and three kinds of multimeric conjugate samples were obtained. The resulting multimeric conjugates are all in the range of 80-90% based on the small molecule active drug. The obtained product was determined to have a combined active drug content in the range of 10-30%, and the free active drug content was less than 0.5%.

Preparation Example 13, Preparation of Polyconjugate

Refer to the methods of Examples 4-6, respectively, except that the amino acid polymer used is poly(glutamic acid/aspartic acid) and glutamic acid accounts for 50.3%, 68.2%, and 85.4%, respectively, of the total amino acid molar amount. Three multimeric conjugate samples were obtained. The resulting multimeric conjugates are all in the range of 80-90% based on the small molecule active drug. The obtained product was determined to have a combined active drug content in the range of 10-30%, and the free active drug content was less than 0.5%.

Preparation Example 14, Preparation of Polyconjugate

The polyconjugate polyglutamic acid-glycine-SN38 was prepared according to the method described in Example 18 of the CN1429121A specification. The resulting multimeric conjugate was 82.7% in terms of small molecule active drug. The obtained product was determined to have a bound active drug content of 15.4% and a free active drug content of <0.25%.

As can be seen, the polyconjugates obtained in Preparation Examples 1-14 all have higher yields and bound active drug contents, and the free active drug content is lower. Further, the preparation scale of each of the above preparation examples can be easily enlarged and the respective performance parameters are not significantly changed, and can be effectively repeated.

C, test case

Test Example 1: Chemical stability of the polyconjugate of the present invention

The various multimeric conjugates obtained in the above preparation examples of the present invention were allowed to stand at 35 ° C for 8 months, and the content of the poly conjugate was determined by the HPLC method described above according to the present invention, and the poly conjugate molecules were simultaneously determined. The relative percentage of the E-ring open-loop degradation product of SN38 relative to the main component.

For the SN38 polycondensate, the percentage of the 8-month relative to the month of 0 was used as the evaluation index; for the E-ring open-loop degradation product, the percentage increase with respect to the month of August was used as the evaluation index. These indicators were used to evaluate the stability of the polyconjugate.

According to the results of Test Example 1, all the amino acid polymer-bridging amino acid-SN38 polyconjugates obtained in Preparation Example 1 - Preparation Example 10 were in the range of 96.8-98.5% in August, and E in August. The percent increase of ring-opening degradation products was in the range of 23.2-38.6%, showing excellent chemical stability; for all the polyconjugates obtained in Preparation Example 11, the percentage of August was in the range of 89.2-92.6%. In August, the percentage increase of E-ring open-loop degradation products was in the range of 142.2-176.5%, indicating that these poly-conjugates (ie, poly-conjugates containing no bridging amino acids) have a lower content of main components and an increase in degradation products. For all the polyadhesives obtained in Preparation Example 12-Preparation Example 14, the percentage of August was in the range of 93.1-94.3%, and the percentage increase of the ring opening degradation products of E ring in August was in the range of 237.1-282.7%. It is shown that the content of the main component of these multimeric binders (i.e., the polyadduct in which the amino acid polymer is glutamic acid or mostly glutamic acid) is reduced, and the degradation products are greatly increased. Thus, it is seen that the preferred samples of the present invention (Preparation Examples 1-10) are superior in stability to the samples of Reference Examples (Preparations 11-14).

Test Example 2: Biological experiment

Test Example 2-1: Biological effect test

Although irinotecan is clinically used for colorectal cancer, it is also effective for lung cancer, and its metabolite SN38 has the same anticancer spectrum. The TGD of all the multimeric conjugates obtained in Preparation Examples 1-14 (which reflects tumor growth retardation) was tested in accordance with the method described in Example 8 of CN101507820A. In addition, different doses of free SN38 were used to determine TGD in the same manner, and dose-response curves were plotted with free SN38 and TGD. Based on the dose-response curve, the amount of polyconjugate required to achieve the same effect (converted to the amount of SN38 contained in the polyconjugate) was calculated, and the amount of polyconjugate was calculated to achieve the same effect. Relative to the percentage of free SN38 dose, if the value is less than 100%, it indicates The dose of polyconjugate required to achieve the same biological effect is smaller than the free SN38 dose, and the lower the value, the better the biological effect of the polyconjugate.

According to the results of Test Example 2, the percentage of all the multimeric conjugates obtained in Preparation Examples 1-10 relative to the free SN38 dose was in the range of 68-77%, and all the poly conjugates obtained in Preparation Examples 12-14 were relative to the free SN38. The percentage of the dose is in the range of 84-96%, for example, the percentage of all polyconjugates obtained in Preparation 11 relative to the free SN38 dose is in the range of 93-96%. Thus, the biological effects of the multimeric conjugates obtained in Preparation Examples 1-10 were very good.

Test Example 2-2: Toxicity test

The samples obtained by using the above various preparation examples (Preparation Examples 1-10) of the present invention, SN-38, and the cited examples (Preparation Examples 11-14) were used as test articles, and the SPF grade was 6-8 weeks. KM mice were experimental animals and the maximum tolerated dose (MTD) of the study drug was tested by single administration of tail vein injection. The results showed that the maximum tolerated dose of SN-38 mice was 14 mg/kg, and the maximum tolerated dose of the samples prepared in the present invention (Preparation Examples 1-10) was above 80 mg/kg, and the cited examples (Preparations 11-14) The MTD in the sample is also only 20-40 mg/kg. This result indicates that the toxicity of the polyconjugate prepared by the present invention is much less than that of the SN38 and the cited sample. The toxicity of the cited sample is twice or more higher than that of the sample of the present invention, and the toxicity of the SN38 is 5.7 times or more of the sample of the present invention. high.

Test Example 2-3: Efficacy test of human colon cancer HCT116 tumor-bearing mice:

The following experiment was carried out using the polyaspartic acid-SN38 multimeric conjugate (Sample 1) obtained in Preparation Example 1.

HCT116 cells were cultured in a 37 ° C, 5% CO ₂ incubator with McCoy's 5a medium containing inactivated 10% fetal calf serum, 100 U/ml penicillin and 100 μg/ml streptomycin and 2 mM glutamine. The initial concentration of the cell culture was 1 × 10 ⁶ /ml, and the cells were passaged every 3 to 4 days after the cells were over. Tumor cells in the logarithmic growth phase are used for inoculation of tumors in vivo. HCT116 tumor cells resuspended in serum-free McCoy's 5a culture medium were inoculated subcutaneously into the right flank of the experimental animals at 5 × 10 ⁶ /100 μl. When the tumor grew to about 120 mm ³ , animals with a relatively uniform tumor volume were selected and administered in groups of 4, with 8 rats in each group.

Tumor measurement and experimental indicators: The tumor was measured twice a week using a vernier caliper to measure the long diameter and short diameter of the tumor. The volume was calculated as: volume = 0.5 × long diameter × short diameter ² . Relative tumor volume (RTV) and relative tumor volume increase ratio (%, T/C) were calculated based on tumor volume. RTV = Vt / V0, where Vt is the mean tumor volume on day t after group administration, and V0 is the mean tumor volume on the day of grouping. T/C=TRTV/CRTV×100, where TRTV is the treatment group RTV and CRTV is the solvent control group RTV. The tumor growth inhibition rate (%, TGI) was calculated as follows: (1-T/C) × 100%.

The tail volume was administered, and the tumor volume and the change in body weight of the following four groups were measured:

Group 1: solvent control group (administered once in the first week and second week); group 2: positive control group irinotecan 80 mg/kg (administered once in the first week and the second week); Group 3: polyaspartic acid-SN38 polyconjugate (Sample 1) 50 mg/kg (based on SN38 content) (administered only once in the first week); Group 4: polyaspartic acid-SN38 Polyconjugate (Sample 1) 40 mg/kg (based on SN38 content) (administered once each in the first week and the second week). The test results are shown in Figure 1 respectively.

As can be seen from Figure 1, the use of a single dose of 50 mg/kg of sample 1 was more pronounced in inhibiting tumor growth than the use of two doses of 80 mg/kg of irinotecan, while using 40 mg/kg of both. The second dose of sample 1 showed a very excellent tumor suppressing effect. Sample 1 40 mg/kg (qwk × 2) group and single dose group 50 mg / kg showed very significant anti-tumor effect, the tumor growth inhibition rate on day 26 was 93.1% and 90.7%, respectively (compared with the solvent control group) P < 0.001); continued observation showed that the tumor weight of the irinotecan group and the sample 1 (40 mg / kg, qwk × 2) group were significantly different. At the end of the experiment, the tumor of the sample 1 (40 mg / kg, qwk × 2) group The delay time (TGD) was 36 days, and the tumor delay time (TGD) for the sample 1 (50 mg/kg single administration) group was 20 days, while the irinotecan group was only 14 days. In summary, the use of the product of the present invention has a very satisfactory effect in treating tumors.

Test Example 2-4: Pharmacokinetic test of human colon cancer HCT116 tumor-bearing mice

In the test case 2-3, the sample 1 was treated, and the two groups of animals on the 40th day after the drug withdrawal (group A and group B, respectively) were administered with a dose of 40 mg/kg through the tail vein, respectively, at various time points. Animal plasma and tumor tissues were taken, and the drug concentration (as SN-38 concentration) and the released SN38 concentration in plasma and tumor tissues were analyzed by HPLC-MS, and the results are shown in Fig. 2 and Fig. 3. It can be seen from Fig. 2 and Fig. 3 that the results of group A and group B are almost identical, and the animal plasma remains in the tumor-bearing animal after treatment with sample 1 and on the 40th day after stopping the drug (ie, corresponding to the 0 mark of the abscissa). The presence of sample 1 and SN38 can be detected (ie, corresponding to the position indicated by the arrow at 0 o'clock), and the concentration is substantially the same as the concentration 48 hours after the current administration, indicating that sample 1 has a long length in the animal. Long-lasting slow release effect. Same sample 1 in the tumor group The intramuscular drug is cleared slowly, so sample 1 can also inhibit tumor growth for a long time after a single administration.

In addition, comparing Fig. 2 and Fig. 3, it can be concluded that the drug concentration of SN38 in the tumor tissue reached 47 times of the plasma concentration at 6 hours after administration, and the drug of the sample 1 in the tumor tissue at 48 hours. The concentration was 376 times of the plasma concentration, indicating that sample 1 was specifically enriched in tumor tissues and showed excellent targeting effects.

Test Example 2-5: Pharmacokinetic test of healthy animals

Healthy CD-1 mice were used as experimental animals, two groups, 3 in each group, a total of 6 animals, with SN-38 as the positive control, the dose was 10 mg/kg, and the sample 1 group was administered 40 mg/kg. They were all administered by tail vein injection, and continuous blood sampling was used to detect the concentration of drug in plasma. The heart, liver, spleen, lung, kidney, brain, colon and other tissues were collected at the last time point and analyzed for drug concentration.

The results showed that the plasma half-life of SN-38 released from sample 1 was 13.3 hours, the half-life of sample 1 in plasma was 16.4 hours, and the half-life of positive control SN-38 in plasma was 15.9 hours. From the perspective of tissue distribution, 48 hours after administration, SN-38 was detected in the SN-38 group of the drug substance in heart, liver, spleen, lung, kidney, brain and colon, and sample 1 was specifically distributed in the above organs, such as kidney cancer. For liver cancer, lung cancer, etc., according to the distribution of sample 1 in the main organs, corresponding indications can be developed.

D, part of the composition example

Composition Example 1: Preparation of freeze-dried powder injection

Formulation: 50 parts by weight of polyconjugate, 200 parts by weight of mannitol

Method: Add the polyconjugate and mannitol to the appropriate amount of water for injection, dissolve, adjust the pH of the solution to 5.0-5.5 with 1M hydrochloric acid solution or 1M sodium hydroxide solution, add water for injection to the liquid. The content was 7.5%. The bacteria were sterilized by 0.45 μm and 0.22 μm microporous membrane filtration, and the liquid was dispensed into a vial under aseptic conditions (the amount of SN38 was 50 mg per bottle), and the stopper was placed in a freeze dryer. Inside, freeze-dry to a moisture content of less than 3%, tampon, that is.

Composition Example 2: Preparation of freeze-dried powder injection

Formulation: 50 parts by weight of polyconjugate, 250 parts by weight of lactose

Method: Add the polyconjugate and lactose to an appropriate amount of water for injection, dissolve, adjust the pH of the solution to 4.5-5.0 with 1M hydrochloric acid solution or 1M sodium hydroxide solution, and add water for injection to the solid in the liquid. The content is 10%. The bacteria were sterilized by 0.45 μm and 0.22 μm microporous membrane filtration, and the liquid was dispensed into a vial under aseptic conditions (the amount of SN38 was 50 mg per bottle), and it was stoppered and placed in a frozen state. In the dryer, freeze-dry to a moisture content of less than 3%, squeezing, that is.

Composition Example 3: Preparation of Injection

Formulation: 50 parts by weight of polyconjugate, 10 parts by weight of citric acid, and 1000 parts by weight of water for injection

Method: The polyconjugate and citric acid are added to an appropriate amount of water for injection to dissolve, and the pH of the solution is adjusted to 3.5-4.0 with 1 M hydrochloric acid solution or 1 M sodium hydroxide solution, and water for injection is added to the total amount. The bacteria were sequentially filtered through a 0.45 μm and 0.22 μm microporous membrane filter, and the drug solution was dispensed into an ampoule under aseptic conditions (the amount of each bottle was converted into SN38 was 50 mg), and sealed.

From this, it can be seen that the multi-conjugate of the present invention can be made into different dosage forms for use according to different conditions according to actual conditions or needs.

The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the specific details of the above embodiments, and various simple modifications can be made to the technical solutions of the present invention within the scope of the technical idea of the present invention. These simple variants All fall within the scope of protection of the present invention.

It should be further noted that the specific technical features described in the above specific embodiments may be combined in any suitable manner without contradiction. To avoid unnecessary repetition, the present invention has various possibilities. The combination method will not be described separately.

In addition, any combination of various embodiments of the invention may be made as long as it does not deviate from the idea of the invention, and it should be regarded as the disclosure of the invention.

Claims (20)

A multimeric binder, wherein the multimeric binder comprises: an amino acid polymer as a main chain and an active drug molecule as a branch, wherein the active drug molecule is 7-ethyl-10-hydroxyl A cardinine which is linked to the amino acid polymer by an amino acid at the 20-hydroxyl group. The multimeric conjugate according to claim 1, wherein the monomer of the amino acid polymer is aspartic acid, or a combination of both aspartic acid and glutamic acid. The multimeric binder according to claim 1, wherein the amino acid of the amino acid polymer is a D-form amino acid, an L-form amino acid, a DL mixed amino acid, or a combination thereof. The multimeric binder according to claim 1, wherein the amino acid polymer has a number average molecular weight of from 1,000 to 100,000 Daltons. The multimeric conjugate according to claim 1, wherein one carboxyl group of said amino acid forms an ester bond with a 20-hydroxyl group of said active drug molecule, and/or an amino group of said amino acid and said amino acid polymer The carboxyl group forms an amide bond. The multimeric conjugate according to claim 1, wherein one or more of said active drug molecules are bound to each of said amino acid polymers, said active drug molecule based on 100% by weight of a polymeric binder It is 3-60% by weight. The multimeric conjugate according to any one of claims 1 to 6, which has the following formula:

among them, R' is a side chain of a natural amino acid; n is an integer from 2 to 150; PA is the amino acid polymer. A method of preparing the multimeric binder of any of claims 1-7, comprising the steps of: 1) adding an amino acid polymer, a compound represented by Formula A, a catalyst, and a condensing agent to a solvent; 2) contacting the solution obtained in the step 1) with a poor solvent, and then filtering; 3) adding the filter cake obtained in step 2) to a solvent and contacting the deprotecting agent to obtain a multimeric binder;

Wherein R' is a side chain of a natural amino acid. The method according to claim 8, wherein the solvents in the step (1) and the step (3) are each independently N,N-dimethylformamide and/or dimethyl sulfoxide; and the step (2) The poor solvent in the) is at least one of water, acetone, methanol, ethanol, isopropanol, diethyl ether, methyl tert-butyl ether, tetrahydrofuran, dichloromethane, and ethyl acetate. The method according to claim 8, wherein the catalyst is at least one of pyridine, 4-dimethylaminopyridine, triethylamine, N,N-diisopropylethylamine and nitrogen methylmorpholine. The method according to claim 8 or 10, wherein the molar ratio of the catalyst to the active drug is from 0.01 to 4:1. The method according to claim 8, wherein said condensing agent is dicyclohexylcarbodiimide, diisopropylcarbodiimide and 1-ethyl-(3-dimethylaminopropyl)carbonyl At least one of diimine hydrochloride. The method according to claim 8 or 12, wherein the molar ratio of the condensing agent to the active drug is from 0.5 to 4:1. The method according to claim 8, wherein the deprotecting agent is at least one of tetrabutylammonium fluoride, tetraethylammonium fluoride, tetramethylammonium fluoride, and hydrogen fluoride pyridine solution. The method according to claim 8, further comprising: 4) separating and extracting the resulting multimeric conjugate, wherein the separation extraction is carried out by dialysis or ultrafiltration. A pharmaceutical composition comprising the multimeric composition of any of claims 1-7, and a pharmaceutically acceptable excipient. The pharmaceutical composition according to claim 16, which is a pharmaceutical composition for injection. The pharmaceutical composition according to claim 17, wherein the pharmaceutically acceptable excipient comprises water for injection, 0.8 to 1.2% by weight of sodium chloride injection, and 3 to 7% by weight of glucose injection. Use of the multimeric composition of any of claims 1-7 or the pharmaceutical composition of any of claims 16-18 for the treatment of a tumor or cancer. The use according to claim 19, wherein the tumor or cancer is at least one of colorectal cancer, pancreatic cancer, breast cancer, ovarian cancer, lung cancer, kidney cancer, stomach cancer, and liver cancer.

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