Uses of Hyaluronan Conjugates

Abstract

The present specification discloses the use of hyaluronan (HA) conjugates for treating cancer.The present specification also discloses the use of hyaluronan conjugates for treating cancer.The hyaluronan conjugates are nimesulide-HA conjugates having monosaccharide or 1-4 disaccharide units of hyaluronic acid.

Description

[Technical field]

[0001] The present disclosure relates to hyaluronic acid conjugates, and more particularly, to hyaluronic acid conjugates for treating cancer. [Background technology]

[0002] Cancer is one of the leading causes of death worldwide, with approximately 10 million cancer deaths in 2020. According to statistics from the World Health Organization (WHO), the five most common new cancer cases in 2020 were breast cancer (2.26 million), lung cancer (2.21 million), colorectal cancer (1.93 million), prostate cancer (1.41 million), and skin cancer (non-melanoma) (1.2 million). The most common causes of cancer death in 2020 were lung cancer (1.8 million deaths), colorectal cancer (935,000 deaths), liver cancer (830,000 deaths), stomach cancer (769,000 deaths), and breast cancer (685,000 deaths).

[0003] Currently, there are four standard methods for treating cancer: radiation therapy, chemotherapy, surgery, and / or immunotherapy. Among them, surgery and chemotherapy remain the most commonly employed means of cancer treatment. Determining the goal of treatment is an important first step. The main goal is generally to cure the cancer or to significantly extend lifespan. Surgery is often performed with the aim of completely removing the cancer without causing significant damage to other parts of the body, but its effectiveness is often limited by the tendency of cancer to invade adjacent tissues or spread to distant sites through micrometastases. Chemotherapy is the treatment of cancer with cytotoxic drugs that can destroy cancer cells. However, these drugs may not be useful in cancer treatment because cancer cells acquire resistance or become insensitive to one or more cytotoxic drugs. In addition, some cancers (e.g., lung cancer) have poor prognosis or are insensitive to current chemotherapeutic agents. Therefore, there have been many attempts to find other effective drugs or means to successfully treat cancer, especially drug-resistant or chemotherapeutic-insensitive cancers.

[0004] Thus, there is a need in the art for agents or compounds that may be useful in the treatment of cancer.

[0005] (overview) The following presents a simplified summary of the disclosure in order to provide the reader with a basic understanding. This summary is not an extensive overview of the disclosure, and it does not identify key / critical elements of the invention or delineate the scope of the invention. Its sole purpose is to present some concepts disclosed herein in a simplified form as a prelude to the more detailed description that is presented below.

[0006] In one aspect, the disclosure relates to a method of treating cancer in a subject in need of treatment.

[0007] In some embodiments, the method includes administering to a subject an effective amount of a hyaluronan conjugate, wherein the hyaluronan conjugate includes a hyaluronan moiety of (-4GlcUAβ1-3GlcNAcβ1-)n, where n=1 to 4, and a substituent (e.g., a cytotoxic agent) can be attached to one or more carboxyl groups of D-glucuronic acid (4GlcUA).

[0008] In some other embodiments, the hyaluronan conjugate comprises a hyaluronan moiety of 3GlcNAcβ1-(4GlcUAβ1-3GlcNAcβ1-)n or (4GlcUAβ1-3GlcNAcβ1-)n-4GlcUAβ1 (wherein n=1 to 4), and a substituent (e.g., a cytotoxic drug) may be attached to one or more carboxyl groups of D-glucuronic acid (4GlcUA).

[0009] According to some embodiments of the present disclosure, the method includes administering to a subject an effective amount of a hyaluronan conjugate, wherein the hyaluronan conjugate includes only two disaccharide units and has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0010] In some embodiments, the hyaluronan conjugate has the structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0011] In some embodiments, the hyaluronan conjugate has the structure: [ka] or a pharma- ceutically acceptable salt thereof, wherein X is 1 and Y is 0, 1, 2 or 3.

[0012] In various embodiments, the subject is a mammal, including a human.

[0013] According to some embodiments of the present disclosure, the cancer is breast cancer, lung cancer, or colorectal cancer. For example, the lung cancer can be non-small cell lung cancer (e.g., squamous cell carcinoma, adenocarcinoma, and large cell carcinoma) or small cell lung cancer.

[0014] In some embodiments, the hyaluronan conjugate is administered by intravenous injection.

[0015] In yet another aspect, the present disclosure relates to a pharmaceutical composition for treating cancer.

[0016] In some embodiments, the pharmaceutical composition comprises an effective amount of the conjugate of hyaluronic acid described above and a pharma- ceutically acceptable excipient.

[0017] Subject matter encompassed within other aspects of the present disclosure includes the use of conjugates of hyaluronic acid in the manufacture of a medicament for use in the treatment of cancer, and conjugates of hyaluronic acid for use in the treatment of cancer.

[0018] Many of the attendant features and advantages of the present disclosure will be better understood by reference to the following detailed description considered in conjunction with the accompanying drawings.

[0019] The specification will be better understood from the following detailed description taken in conjunction with the accompanying drawings. [Brief description of the drawings]

[0020] [Figure 1] FIG. 1 is a line graph showing the tumor cytotoxicity of HA conjugates against A549 cell line (*: p<0.05 vs. Nim; #: p<0.05 vs. HA4). [Diagram 2] FIG. 2 is a line graph showing the tumor cytotoxicity of HA conjugates against the MDA-MB-231 cell line (*: p<0.05 vs. Nim; #: p<0.05 vs. HA4). [Diagram 3] FIG. 3 is a line graph showing the tumor cytotoxicity of HA conjugates against HT-29 cell lines (*: p<0.05 vs. Nim; #: p<0.05 vs. HA4). DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0021] (explanation) The detailed description provided below in conjunction with the accompanying drawings is intended to be illustrative of the present embodiment and is not intended to represent the only manner in which the present embodiment may be constructed or utilized. The description sets forth the functions of the present embodiment and the sequence of steps for constructing and executing the present embodiment. However, the same or equivalent functions and sequences may be accomplished by different embodiments.

[0022] For convenience, certain terms employed in the specification, examples, and appended claims are collected here. Unless otherwise defined herein, scientific and technical terms employed in this disclosure shall have the meanings commonly understood and used by those of ordinary skill in the art.

[0023] Unless otherwise required by context, it is understood that the singular includes the plural of the same and the plural includes the singular. Also, as used herein and in the claims, the terms "at least one" and "one or more" have the same meaning and include one, two, three, or more. Furthermore, as used throughout this specification and the appended claims, the phrases "at least one of A, B, and C," "at least one of A, B, or C," and "at least one of A, B, and / or C" are intended to cover A only, B only, C only, A and B together, B and C together, A and C together, and A, B, and C together.

[0024] Notwithstanding that the numerical ranges and parameters setting forth the broad scope of the invention are approximations, the numerical values ​​set forth in the specific examples are reported as precisely as possible. However, any numerical value contains a certain degree of error resulting from the standard deviation found in their respective testing measurements. Additionally, as used herein, the term "about" generally means within 10%, 5%, 1%, or 0.5% of a given value or range. Additionally, the term "about" means within an acceptable standard error of the mean as would be expected by one of ordinary skill in the art. Unless otherwise specified in the Examples / Examples or otherwise expressly indicated, all numerical ranges, amounts, values, and percentages for amounts of materials, lengths of time, temperatures, operating conditions, proportions of amounts, and the like disclosed herein are to be understood as being modified in all instances by the term "about." Thus, unless specifically indicated to the contrary, the numerical parameters set forth in the present disclosure and the appended claims are approximations that may be varied as desired. At a minimum, each numerical parameter should be interpreted at least in light of the number of reported significant digits and by applying ordinary rounding techniques. Ranges may be expressed herein as from one endpoint to another endpoint or between two endpoints. All ranges disclosed herein include the endpoints unless otherwise specified.

[0025] The terms "treatment" and "treating" as used herein may refer to preventative (e.g., prophylactic), curative or palliative measures. In particular, the term "treating" as used herein refers to the application or administration of the hyaluronan conjugate or a pharmaceutical composition comprising the same to a subject having a pathology, a symptom associated with a pathology, a disease or disorder secondary to a pathology, or a predisposition to a pathology, with the purpose of partially or fully alleviating, ameliorating, relieving, delaying the onset, inhibiting the progression, reducing the severity, and / or reducing the incidence of one or more symptoms or characteristics of the particular disease, disorder, and / or condition. Treatment may be administered to subjects who do not show signs of a disease, disorder, and / or condition and / or to subjects who show only early signs of a disease, disorder, and / or condition, with the purpose of reducing the risk of developing pathologies associated with the disease, disorder, and / or condition.

[0026] The terms "subject" and "patient" are used interchangeably herein and are intended to mean animals, including human species, treatable by the hyaluronan conjugates described herein, pharmaceutical compositions containing same, and / or methods of the invention. The term "subject" or "patient" includes any mammalian species that may benefit from the present disclosure. As used herein, "mammal" refers to all members of the class Mammalia, including humans, primates, domestic and farm animals such as rabbits, pigs, sheep, and cows, as well as zoo, sport, or pet animals, and rodents such as mice and rats. The term "non-human mammal" refers to all members of the class Mammalia, except humans. In one exemplary embodiment, the patient is a human. The term "subject" or "patient" is intended to refer to both males and females, unless one gender is specifically indicated.

[0027] The terms "application" and "administration" are used interchangeably herein and refer to the application of a hyaluronan conjugate or pharmaceutical composition of the invention to a subject in need of treatment.

[0028] The term "effective amount" as used herein refers to an amount of the hyaluronan conjugate of the present invention sufficient to produce a desired therapeutic effect. An effective amount of a drug is not required to cure a disease or condition, but provides treatment for a disease or condition, such as delaying, hindering, or preventing the onset of the disease or condition, or ameliorating the symptoms of the disease or condition. An effective amount may be divided into one, two, or more doses in a form suitable for administration once, twice, or more times over a specified period of time. The specific effective or sufficient amount will vary depending on factors such as the particular condition being treated, the physical condition of the patient (e.g., the patient's weight, age, or sex), the type of mammal or animal being treated, the duration of the treatment, the characteristics of the concomitant therapy (if any), and the particular dosage form and structure of the compound or its derivatives used. An effective amount may be expressed, for example, as the total mass of the hyaluronan conjugate or the equivalent mass of 4-aminonimesulide in the hyaluronan conjugate (e.g., in grams, milligrams, or micrograms), or as a ratio of the mass of the hyaluronan conjugate or the equivalent mass of 4-aminonimesulide in the hyaluronan conjugate to body weight, for example, milligrams per kilogram (mg / kg).

[0029] Additionally, according to the examples provided below, the hyaluronan conjugates are administered by intravenous injection, however, this is merely an illustration of how the invention may be practiced and the disclosure is not limited thereto.

[0030] For example, the hyaluronan conjugates can be formulated with pharma- ceutically acceptable excipients into pharmaceutical compositions suitable for the desired mode of administration. Certain pharmaceutical compositions prepared in accordance with the inventive concepts disclosed and claimed herein are in single unit dosage forms suitable for oral, mucosal (e.g., intranasal, sublingual, intravaginal, buccal, or rectal), parenteral (e.g., subcutaneous, intravenous, bolus injection, intramuscular, or intraarterial), intravitreal, or transdermal administration to a patient. Examples of dosage forms include, but are not limited to, tablets, caplets, capsules such as soft gelatin capsules, cachets, troches, lozenges, dispersions, suppositories, ointments, poultices (compresses), pastes, powders, dressings, creams, salves, solutions, patches, aerosols (e.g., nasal sprays or inhalers), gels, liquid dosage forms suitable for oral or mucosal administration to a patient, such as suspensions (e.g., aqueous or non-aqueous liquid suspensions, oil-in-water emulsions, or water-in-oil liquid emulsions), solutions, elixirs, and sterile solids (e.g., crystalline or amorphous solids) that can be reconstituted to provide liquid dosage forms suitable for parenteral administration to a patient. As can be appreciated, these pharmaceutical compositions are also within the scope of the present disclosure.

[0031] The term "pharmaceutical acceptable excipient" as used herein means a pharma- ceutically acceptable material, composition, or vehicle, such as a liquid or solid filler, diluent, carrier, solvent, encapsulating material, etc., involved in carrying or transporting a subject agent, such as from one organ or part of the body to another. Each excipient must be "acceptable" in the sense of being compatible with the other components of the formulation. The pharmaceutical formulation comprises a compound of the invention in combination with one or more pharma- ceutically acceptable ingredients. The excipient may be in the form of a solid, semi-solid or liquid diluent, cream or capsule. These pharmaceutical formulations are further objects of the present invention. Usually, the amount of active compound is 0.1-95% by weight of the formulation, preferably 0.2-20% by weight for parenteral preparations and 1-50% by weight for oral preparations. For clinical use of the method of the present invention, the pharmaceutical composition of the present invention is formulated into a formulation suitable for the intended route of administration.

[0032] As used herein, the term "hyaluronic acid" (HA) (also referred to as hyaluronate or hyaluronan) is an anionic, non-sulfated glycosaminoglycan (-4GlcUAβ1-3GlcNAcβ1-) consisting of at least one disaccharide unit, specifically D-glucuronic acid and N-acetyl-D-glucosamine. In some embodiments of the present disclosure, the HA comprises two disaccharide units (i.e., HA tetrasaccharide). The term "HA derivative" refers to HA having any modification of the hydroxyl, carboxyl, amide, or acetylamino groups of one or more disaccharide units of HA. According to certain embodiments of the present disclosure, the HA conjugate is in the form of a metal salt, preferably an alkali metal salt, more preferably a sodium or potassium salt.

[0033] According to certain embodiments of the present disclosure, the hyaluronan conjugate comprises a hyaluronan moiety of (-4GlcUAβ1-3GlcNAcβ1-)n, where n=1-4, and a substituent (i.e., a cytotoxic agent) can be attached to one or more carboxyl groups of D-glucuronic acid (4GlcUA). For example, the cytotoxic agent is hydrogenated nimesulide.

[0034] In some other embodiments, the hyaluronan conjugate comprises a hyaluronan moiety of 3GlcNAcβ1-(4GlcUAβ1-3GlcNAcβ1-)n or (4GlcUAβ1-3GlcNAcβ1-)n-4GlcUAβ1, where n=1-4, and a substituent (e.g., a cytotoxic agent) can be attached to one or more carboxyl groups of D-glucuronic acid (4GlcUA). For example, the cytotoxic agent is hydrogenated nimesulide.

[0035] Specifically, according to certain embodiments, the hyaluronan conjugate comprises only two disaccharide units and has the following structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0036] In some embodiments, the hyaluronan conjugate has the structure: [ka] or a pharma- ceutically acceptable salt thereof.

[0037] In some embodiments, the hyaluronan conjugate has the structure: [ka] or a pharma- ceutically acceptable salt thereof, wherein X is 1 and Y is 0, 1, 2 or 3.

[0038] As can be appreciated, when the hyaluronan moiety has more than one carboxyl group, the "degree of substitution (DS)" of the HA conjugate (i.e., the ratio of substituents (i.e., hydrogenated nimesulide) attached per carboxyl group of the HA moiety) can be from 20 to 100%. Specifically, for an HA conjugate having a hyaluronan moiety of (-4GlcUAβ1-3GlcNAcβ1-)4, the DS is 25, 50, 75, or 100% when the HA conjugate has 1, 2, 3, or 4 substituents, respectively. In another example, for an HA conjugate having a hyaluronan moiety of (4GlcUAβ1-3GlcNAcβ1-)4-4GlcUAβ1, the DS is 20, 40, 60, 80, or 100% when the HA conjugate has 1, 2, 3, 4, or 5 substituents, respectively.

[0039] The present disclosure is based, at least in part, on the discovery that Nimesulide-HA tetrasaccharide conjugate (NIM-Tetra) exhibits unexpectedly superior tumor cytotoxicity against certain cancer cell lines compared to Nimesulide alone or other Nimesulide-HA conjugates.

[0040] In view of the above, the present disclosure proposes a method for treating cancer, particularly breast cancer, lung cancer and colorectal cancer.Also described herein is the use of said hyaluronan conjugate in the treatment of cancer, and its use in the manufacture of medicines for said therapeutic purposes.Said medicines (i.e. pharmaceutical compositions comprising said hyaluronan conjugates) are of course subjects within the scope of this application.

[0041] The following examples are provided to illustrate certain aspects of the present invention and to aid those skilled in the art in practicing the present invention. These examples are not to be construed as limiting the scope of the present invention in any manner. Without further elaboration, it is believed that those skilled in the art can utilize the present invention to its fullest extent based on the description herein. EXAMPLES

[0042] Example 1

[0043] Synthesis and characterization of NIM-HA conjugates

[0044] (1) Preparation of hydrogenated nimesulide (H-NIM)

[0045] Hydrogenated nimesulide (N-(4-amino-2-phenoxyphenyl)methanesulfonamide, or 4-aminonimesulide) was synthesized from commercially available nimesulide (N-(4-nitro-2-phenoxyphenyl)methanesulfonamide). Briefly, 500 mg of nimesulide was completely dissolved in 20 ml of ethyl acetate, and then 200 mg of 5% Pd / C (palladium on carbon) was added to the solution as a catalyst. The air was removed from the bottle while stirring, and the air was replaced with hydrogen gas up to 1 atm, and then the mixture was stirred for 24 hours to obtain hydrogenated nimesulide (H-NIM).

[0046] The purity of H-NIM was determined by thin layer chromatography (TLC) on a TLC plate precoated with silica gel 60F254 under UV irradiation at 254 nm. The mobile phase was hexane:ethyl acetate=2:1.

[0047] The Pd / C catalyst was removed by filtration, and the filtrate was concentrated on a rotary evaporator to remove residual solvent. The hydrogenated product was then dissolved in a hexane-ethyl acetate (1:1) solution and further purified on a silica gel column. The column was eluted with hexane-ethyl acetate = 1:1. The colored fraction was collected and lyophilized. The concentration and structure of the obtained product were confirmed using UV and NMR, respectively.

[0048] (2) Synthesis of NIM-HA tetrasaccharide conjugate

[0049] HA tetrasaccharide (HA4) with two disaccharide units of D-glucuronic acid and N-acetylglucosamine was reacted with 4-aminonimesulide (hydrogenated nimesulide, H-NIM) to obtain NIM-HA tetrasaccharide conjugate (NIM-Tetra). Briefly, 50 mg of HA4 (0.06437 mmol) was dissolved in 3 mL of DDW. Then, 5 mL of DMSO was added to the HA4 solution. 9.1 mg of Oxyma (0.06437 mmol) was dissolved in 0.5 mL of DMSO and added to the HA4 solution under stirring for 6 min. Then, 17.9 mg of H-NIM (0.07081 mmol) was dissolved in 0.5 mL of DMSO and added to the HA4 solution. 14 μL of DIC (0.09656 mmol) was dissolved in 0.5 mL of DMSO under stirring for 1 min and added to the HA4 solution. The final reaction mixture was stirred at room temperature for 48 hours, and the resulting solution was freeze-dried for 3 days and separated and purified by preparative HPLC (Waters 600) using an Insertsil ODS-3 column. The NIM-Tetra thus synthesized has the following structure: [ka] It has.

[0050] (3) Synthesis of NIM-HA disaccharide conjugate

[0051] A HA disaccharide having one disaccharide unit of D-glucuronic acid and N-acetylglucosamine was reacted with 4-aminonimesulide (hydrogenated nimesulide, H-NIM) to obtain the following structure: [ka] The NIM-HA disaccharide (NIM-Di) conjugate having the formula:

[0052] (4) Synthesis of NIM-HA monosaccharide conjugate

[0053] Briefly, glucuronic acid (194.14 Da) was reacted with 4-aminonimesulide (hydrogenated nimesulide, H-NIM) to give the following structure: [ka] The NIM-HA monosaccharide (454.4 Da) (NIM-Mono) conjugate having the formula:

[0054] Example 2

[0055] Tumor cytotoxicity of NIM-HA conjugates against MDA-MB-231 cells

[0056] In this example, MDA-MB-231 cells (a human breast cancer cell line) were used to evaluate the tumor cytotoxicity of various NIM-HA conjugates.

[0057] Plate 5 × 10 MDA-MB-231 cells in 100 µL of L-15 medium per well in a 96-well plate. 3 The plates were seeded at a density of 100 μL and incubated overnight. The next day, the medium was discarded and replaced with fresh medium (final DMSO concentration: 0.8%) containing 0–150 μM nimesulide (NIM), NIM-Tetra, HA tetrasaccharide, or NIM-HA disaccharide (NIM-Di) conjugates, and the plates were incubated for 24 h. The medium was then discarded, 100 μL medium containing 10% MTT solution (MTT stock: 5 mg / mL) was added, and the plates were incubated at 37 °C in the dark for 4 h. After that, the medium was carefully discarded, 200 μL DMSO was added, and the plates were shaken for 10 min before reading the plates at 570 nm.

[0058] As summarized in Figure 1, the results show that NIM-Tetra exhibits significantly superior tumor cytotoxicity against MDA-MB-231 cells compared to Nimesulide alone, HA tetrasaccharide alone, or NIM-HA disaccharide (NIM-Di) conjugate. NIM-HA disaccharide (NIM-Di) also exhibited tumor cytotoxic activity comparable to Nim.

[0059] Example 3

[0060] Tumor cytotoxicity of NIM-HA conjugates against A549 cells

[0061] In this example, A549 cells (a human lung cancer cell line) were used to evaluate the tumor cytotoxicity of various NIM-HA conjugates.

[0062] Plate A549 cells in 100 µL of F-12K medium at 3 × 10 cells per well in a 96-well plate. 3 The plates were seeded at a density of 100 μL and incubated overnight. The next day, the medium was discarded and replaced with fresh medium (final DMSO concentration: 0.8%) containing 0–150 μM nimesulide, NIM-Tetra, HA tetrasaccharide, or NIM-HA disaccharide (NIM-Di) conjugate, and the plates were incubated for 24 h. The medium was then discarded, 100 μL medium containing 10% MTT solution (MTT stock: 5 mg / mL) was added, and the plates were incubated at 37 °C in the dark for 4 h. The medium was then carefully discarded, 200 μL DMSO was added, and the plates were shaken for 10 min before reading the plates at 570 nm.

[0063] As summarized in Figure 2, the results show that both NIM-Tetra and NIM-HA disaccharide (NIM-Di) conjugates exhibit significantly superior tumor cytotoxicity against A549 cells compared to nimesulide alone or HA tetrasaccharide alone. NIM-HA monosaccharide (NIM-Mono) also exhibited tumor cytotoxic activity comparable to NIM. NIM-HA disaccharide (NIM-Di) also exhibited superior tumor cytotoxic activity to nimesulide.

[0064] Example 4

[0065] Tumor cytotoxicity of NIM-HA conjugates against HT-29 cells

[0066] In this example, HT-29 cells (a human colorectal cancer cell line) were used to evaluate the tumor cytotoxicity of various NIM-HA conjugates.

[0067] HT-29 cells in 100 μL of MyCoy's-5 medium were plated in a 96-well plate at 3 × 10 cells per well. 3 The plates were seeded at a density of 100 μL and incubated overnight. The next day, the medium was discarded and replaced with fresh medium (final DMSO concentration: 0.8%) containing 0–150 μM Nimesulide (NIM), NIM-Tetra, or HA tetrasaccharide, and the plates were incubated for 24 h. After that, the medium was discarded, 100 μL medium containing 10% MTT solution (MTT stock: 5 mg / mL) was added, and the plates were incubated at 37 °C in the dark for 4 h. The medium was then carefully discarded, 200 μL DMSO was added, and the plates were shaken for 10 min before reading the plates at 570 nm.

[0068] As summarized in FIG. 3, the results show that NIM-Tetra exhibits significantly superior tumor cytotoxicity against HT-29 cells compared to Nimesulide alone or HA tetrasaccharide alone.

[0069] In summary, the results provided in Examples 2-4 of the present disclosure demonstrate that the NIM-Tetra conjugate of the present invention exhibits desirable tumor cytotoxicity against several cancers, such as breast, lung, and colorectal cancer. NIM-HA disaccharide (NIM-Di) exhibits desirable cytotoxicity in several cancers, such as breast, lung, and colorectal cancer.

[0070] It will be understood that the above description of the embodiments is given by way of example only, and that various modifications may be made by those skilled in the art. The above specification, examples and data provide a complete description of the structure and use of the exemplary embodiments of the invention. Although various embodiments of the invention have been described with a degree of particularity or with reference to one or more individual embodiments, those skilled in the art may make numerous modifications to the disclosed embodiments without departing from the spirit or scope of the invention.

Claims

1. A pharmaceutical composition for treating cancer in a subject in need thereof, comprising an effective amount of a hyaluronan conjugate and a pharma- ceutical acceptable excipient, wherein the hyaluronan conjugate is 1 to 4 disaccharide units of D-glucuronic acid and N-acetylglucosamine, one or more hydrogenated nimesulides, each covalently attached to said D-glucuronic acid in one of said disaccharide units.

2. The hyaluronan conjugate has the structure 【Chemistry 1】 or a pharma- ceutically acceptable salt thereof.

3. The hyaluronan conjugate has the structure 【Chemistry 2】 or a pharma- ceutically acceptable salt thereof.

4. The hyaluronan conjugate has the structure 【Chemistry 3】 or a pharma- ceutically acceptable salt thereof, wherein X is 1 and Y is 0, 1, 2 or 3, and both ends of the substituent are hydroxyl groups.

5. The pharmaceutical composition of claim 1 , wherein the subject is a human.

6. The pharmaceutical composition of claim 1 , wherein the cancer is breast cancer, lung cancer, or colorectal cancer.

7. 2. The pharmaceutical composition of claim 1, wherein the degree of substitution of the hyaluronan conjugate is 25-100%.

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