Use of a kidney-type glutaminase inhibitor and its combination

Abstract

This invention relates to renal glutaminase inhibitors and their combined use. The invention utilizes molecular docking technology to analyze, screen, and identify specific KGA inhibitor compounds from a known compound library. These compounds exhibit good KGA inhibitory properties and, when combined with specific genistein, can significantly enhance the therapeutic effect on fibrosarcoma tumors. The formulation of these compounds into a kit shows excellent application prospects.

Description

Technical Field

[0001] This application relates to the field of biopharmaceuticals, specifically to the use of renal glutaminase inhibitors and their combination therapy. Background Technology

[0002] Renal glutaminase (GLS) is encoded by the GLS gene located on human chromosome 2 and has different splicing variants. The longest one is called renal glutaminase (KGA). The human KGA sequence was deduced from the sequencing of a large number of isolated cDNAs, and more targeted studies have been conducted on the identified mouse and pig KGA analogs. The GLS gene has been found to consist of 19 exons, exceeding 82 kb in length. Exons 1-15 are spliced ​​together to form GAC, while exons 1-14 and 16-19 are spliced ​​together to form KGA. GAM mRNA contains 2-3 introns, distinguishing it from GAC / KGA starting from residue 162. The C-terminal sequence of GAM is “VSFYIFLS”, encoded by introns 2-3. Introns are very rare as alternative splicing forms in mammalian cells; therefore, it cannot be determined whether GAM is a result of gene defects in isolated cDNA cells.

[0003] The role of renal glutaminase (GLS) in cancer has been a key area of ​​research. The Cancer Genome Atlas (TCGA) pan-oncogene expression data showed that GLS is highly expressed in acute myeloid leukemia, adrenocortical carcinoma, triple-negative breast cancer, colorectal cancer, clear or papillary renal cell carcinoma, lung adenocarcinoma, melanoma, mesothelioma, pancreatic cancer, sarcoma, and thyroid cancer. In particular, small molecule inhibitor and gene silencing experiments revealed that GLS is crucial for the growth of cancer cells in acute myeloid leukemia, breast cancer, colorectal cancer, renal cancer, lung cancer, melanoma, and pancreatic cancer. Although TCGA data showed relatively low GLS expression in glioblastoma cell lines, they remained highly sensitive to glutaminase inhibitors in vitro. These results indicate that GLS is essential for the growth of various cancer cell lines, and inhibition of this enzyme represents a potentially valuable therapeutic strategy.

[0004] For 80 years after the discovery of glutaminase, the only chemical inhibitor that garnered significant attention was DON (6-diazo-5-oxo-1-isoleucine). DON is a glutamine mimic that irreversibly binds to the catalytic serine residue of two glutaminase isoenzymes. While other small molecules have been used to inhibit GLS (particularly membrane-impermeable molecules used to determine subcellular localization), DON has been the most widely used inhibitor over the past decade. However, DON's broad target range and severe toxicity limit its application. In 2007, another GLS inhibitor, BPTES, was discovered. BPTES is a long, highly flexible, and C2-symmetric molecule that binds to GLS in a 1:2 ratio at the dimer binding interface. BPTES has an affinity for GLS approximately 1000 times greater than LGA. However, to date, the only GLS enzyme used for research is a recombinant mutant GLS enzyme, and BPTES has a binding site similar to LGA. Since the crystal structure complex of BPTES and GLS was reported, BPTES has been a hot research topic, with the focus mainly on improving inhibitor affinity and enhancing its drug properties.

[0005] Furthermore, in a new study, researchers from institutions including the University of Tokyo, Keio University, and Kyushu University in Japan discovered that inhibiting kidney-type glutaminase (KGA)-dependent glutamine breakdown in mice can eliminate senescent cells. Specifically, they used RNA interference (RNAi) to identify enzymes necessary for the survival of senescent cells and subsequently induced their death. This research involved using RNA interference to identify enzymes required for the survival of senescent cells. This prompted them to carefully study glutamine metabolism, particularly glutaminase 1 (GLS1). Tests showed that it is crucial for the survival of senescent cells. The researchers then inhibited the glutaminase 1 pathway in test mice. After allowing these changes time to take effect, inhibiting this pathway led to the death of senescent cells. In the long term, this also reduced age-related organ problems and obesity-related health issues.

[0006] However, there is currently not enough research on inhibitors of renal glutaminase GLS, and the available options are also limited. Summary of the Invention

[0007] On the one hand, the present invention provides a specific KGA inhibitor.

[0008] The KGA inhibitor is a compound.

[0009] The structural formula of the KGA inhibitor compound of the present invention is shown below:

[0010]

[0011] The compound can also be synthesized according to the prior art CN102030700B.

[0012] Furthermore, the present invention provides a pharmaceutical composition for treating fibrosarcoma tumors, the pharmaceutical composition comprising the KGA inhibitor compound of the present invention and pharmaceutical excipients.

[0013] Furthermore, the pharmaceutical excipients described herein can be those widely used in the pharmaceutical manufacturing field. Excipients primarily serve to provide a safe, stable, and functional pharmaceutical composition, and can also provide methods to ensure that the active ingredient dissolves at a desired rate after administration to a subject, or to promote the effective absorption of the active ingredient after administration to a subject. The pharmaceutical excipients may be inert fillers, or provide a function such as stabilizing the overall pH of the composition or preventing the degradation of the active ingredient. The pharmaceutical excipients may include one or more of the following: binders, suspending agents, emulsifiers, diluents, fillers, granulating agents, adhesives, disintegrants, lubricants, anti-adhesion agents, flow aids, wetting agents, gelling agents, absorption delay agents, dissolution inhibitors, enhancers, adsorbents, buffers, chelating agents, preservatives, colorants, flavoring agents, and sweeteners.

[0014] Furthermore, substances that can be used as pharmaceutically acceptable excipients include, but are not limited to, ion exchangers, aluminum, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffering substances such as phosphates, glycine, sorbic acid, potassium sorbate, mixtures of partial glycerides of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silicates, magnesium trisilicate, polyvinylpyrrolidone, polyacrylates, waxes, polyethylene-polyoxypropylene-blocking polymers, lanolin, sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as carboxymethyl cellulose. Sodium cellulose, ethyl cellulose, and cellulose acetate; gum powder; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil, and soybean oil; glycols such as propylene glycol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic salts; Ringer's solution; ethanol, phosphate buffer solutions, and other non-toxic and suitable lubricants such as sodium lauryl sulfate and magnesium stearate; colorants, release agents, coatings, sweeteners, flavorings and spices, preservatives, and antioxidants.

[0015] Substances that can be used as pharmaceutically acceptable excipients include, but are not limited to, ion exchangers, aluminum, aluminum stearate, lecithin, serum proteins such as human serum albumin, buffering substances such as phosphates, glycine, sorbic acid, potassium sorbate, mixtures of partial glycerides of saturated vegetable fatty acids, water, salts or electrolytes such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silicates, magnesium trisilicate, polyvinylpyrrolidone, polyacrylates, waxes, polyethylene-polyoxypropylene-blocking polymers, lanolin, sugars such as lactose, glucose, and sucrose; starches such as corn starch and potato starch; cellulose and its derivatives such as carboxymethyl cellulose. Sodium thiosulfate, ethyl cellulose and cellulose acetate; gum powder; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols such as propylene glycol and polyethylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffers such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic salts; Ringer's solution; ethanol, phosphate buffer solution, and other non-toxic and suitable lubricants such as sodium lauryl sulfate and magnesium stearate, colorants, release agents, coatings, sweeteners, flavorings and spices, preservatives and antioxidants.

[0016] The pharmaceutical compositions of the present invention can be prepared using any method known to those skilled in the art, based on the disclosure. For example, conventional mixing, dissolving, granulation, emulsification, grinding, encapsulation, embedding, or lyophilization processes.

[0017] The therapeutically effective dose of the compositions of this invention can vary depending on various factors, such as the method of administration, target site, and patient condition. Therefore, when using the compositions of this invention in humans, the dosage should be determined to be an appropriate amount that balances safety and efficacy. The dosage for human use can also be estimated from the effective dose determined through animal studies. Considerations for determining the effective dose are described in references such as Hardman and Limbird, Goodman and Gilman's The Pharmacological Basis of Therapeutics, 10th edition (2001), Pergamon Press; and E.W. Martin, Remington's Pharmaceutical Sciences, 18th edition (1990), Mack Publishing.

[0018] The compositions of the present invention may comprise carriers, diluents, excipients, and mixtures of two or more of the above substances conventionally used in biological agents. Pharmaceutically acceptable carriers may be any carrier suitable for in vivo delivery of the composition, without particular limitation. For example, compounds described in the Merck Index (13th edition, Merck & Co. Inc.), saline solutions, sterile water, Ringer's solution, buffered saline, dextran solution, maltodextrin solution, glycerol, ethanol, and mixtures of one or more of the above components may be used, and other conventional additives (such as antioxidants, buffer solutions, bactericides, etc.) may be added if necessary. Furthermore, diluents, dispersants, surfactants, binders, and lubricants may be added to prepare injections (such as aqueous solutions, suspensions, emulsions), pills, capsules, granules, or tablets. Additionally, compositions may be formulated according to appropriate methods in the art or methods disclosed in Remington's Pharmaceutical Science (Mack Publishing, Easton PA, 18th edition, 1990), depending on the disease or ingredient.

[0019] The compositions of the present invention may additionally contain one or more active ingredients having the same or similar functions. Relative to the total weight of the composition, the compositions of the present invention may contain from about 0.0001 wt% to about 10 wt%, preferably from about 0.001 wt% to about 1 wt%.

[0020] Depending on the intended method, the compositions of the present invention may be administered parenterally (e.g., intravenously, subcutaneously, intraperitoneally, or orally) or orally. Dosage may vary depending on factors such as body weight, age, sex, health status, diet of a particular patient, duration of administration, method of administration, clearance, and severity of disease. The daily dose of the compositions of the present invention is from about 0.0001 mg / mL to about 10 mg / mL, preferably from about 0.0001 mg / mL to about 5 mg / mL, and more preferably once daily or in divided doses several times a day.

[0021] Furthermore, the present invention also provides a kit for treating fibrosarcoma tumors, the kit comprising the KGA inhibitor compound of the present invention and genistein.

[0022] Furthermore, the ratio of the dosage of the two drugs is 1:1 to 1:5, more preferably 1:2.

[0023] Furthermore, the present invention also provides the use of KGA inhibitor compounds and genistein in the preparation of a medicament for treating fibrosarcoma tumors.

[0024] Furthermore, the dosage of the medicine box is a pharmacologically effective amount.

[0025] The pharmaceutically effective dose may be the total amount of both, but not limited to, from about 0.00001 mg / kg to about 10 mg / kg, preferably from about 0.0001 mg / kg to about 1 mg / kg. The dosage may vary depending on factors such as weight, age, sex, health status, diet of the specific patient, duration of administration, method of administration, clearance rate, and disease severity.

[0026] Individuals may be vertebrates, preferably mammals, more preferably laboratory animals (such as rats, rabbits, guinea pigs, hamsters, dogs, and cats), and most preferably great apes (such as chimpanzees and gorillas). Administration may be oral or parenteral. For parenteral administration, intraperitoneal injection, rectal injection, subcutaneous injection, intravenous injection, intramuscular injection, epidural injection, intracerebral injection, or intrapleural injection may be selected.

[0027] Furthermore, based on the mechanism of this invention, the medicine box can also be used for the treatment of other cancers.

[0028] As used in this invention, the term "cancer" refers to solid tumors and hematologic malignancies selected from the group consisting of: breast cancer, prostate cancer, cervical cancer, ovarian cancer, gastric cancer, colorectal cancer (i.e., including colon and rectal cancer), pancreatic cancer, liver cancer, brain cancer, neuroendocrine cancer, lung cancer, kidney cancer, hematologic malignancies (e.g., leukemia), melanoma, and sarcoma. More particularly preferred cancers are selected from the group consisting of: hematologic malignancies, prostate cancer, breast cancer, cervical cancer, ovarian cancer, colorectal cancer, melanoma, and lung cancer. In a particularly preferred embodiment, the cancer is acute myeloid leukemia (AML) or prostate cancer.

[0029] Furthermore, the drug in the said medicine box is a solid dispersion. Solid dispersions, particularly MBP and / or spray-dried products obtainable according to the provided methods, can be used in various forms for administering poorly water-soluble drugs (e.g., compound A), especially for oral dosage forms. Exemplary dosage forms include powders or granules that can be dried or reconstituted by adding water to form pastes, slurries, suspensions, or solutions for oral ingestion; tablets, capsules, or pills. Various additives can be mixed, ground, or granulated with the solid dispersions as described in this invention to form materials suitable for the above dosage forms. Potentially beneficial additives generally fall into the following classes: other matrix materials or diluents, surfactants, drug complexing agents or solubilizers, fillers, disintegrants, binders, lubricants, and pH modifiers (e.g., acids, bases, or buffers). Examples of other matrix materials, fillers, or diluents include lactose, mannitol, xylitol, microcrystalline cellulose, calcium pyrophosphate, and starch. Examples of surfactants include sodium dodecyl sulfate and polysorbate 80. Examples of drug complexing agents or solubilizers include polyethylene glycol, caffeine, xanthracene, gentianic acid, and cyclodextrin. Examples of disintegrants include sodium carboxymethyl starch, sodium alginate, sodium carboxymethyl cellulose, methylcellulose, and croscarmellose sodium. Examples of binders include methylcellulose, microcrystalline cellulose, starch, and gums such as guar gum and astragalus gum. Examples of lubricants include magnesium stearate and calcium stearate. Examples of pH modifiers include acids such as citric acid, acetic acid, ascorbic acid, lactic acid, aspartic acid, succinic acid, phosphoric acid, etc.; bases such as sodium acetate, potassium acetate, calcium oxide, magnesium oxide, trisodium phosphate, sodium hydroxide, calcium hydroxide, aluminum hydroxide, etc., and buffers that typically contain a mixture of an acid and a salt of said acid. At least one function of including this pH modifier is to control the dissolution rate of the drug, matrix polymer, or both, thereby controlling the local drug concentration during dissolution.

[0030] Beneficial effects

[0031] This invention utilizes molecular docking technology to analyze, screen, and identify specific KGA inhibitor compounds from a known compound library. These compounds exhibit good KGA inhibitory properties.

[0032] Combination therapy is a common and effective form of cancer treatment, but choosing which drugs to combine it with is not easy.

[0033] The inventors, through the identification of dozens of drug combinations, accidentally discovered that the compound, when used in combination with a specific genistein, could significantly improve the treatment efficacy of fibrosarcoma tumors. However, the synergistic effect was not significant when used with other common drugs such as paclitaxel.

[0034] The compounds of the present invention have good safety, and the preparation method is also mature and known in the art. Attached Figure Description

[0035] Figure 1 Tumor suppression effect diagram of each treatment group Detailed Implementation

[0036] Specific embodiments of the invention will now be described in more detail with reference to the accompanying drawings. While specific embodiments of the invention are shown in the drawings, it should be understood that the invention can be implemented in various forms and should not be limited to the embodiments set forth herein. Rather, these embodiments are provided to enable a more thorough understanding of the invention and to fully convey the scope of the invention to those skilled in the art.

[0037] Example 1: Screening of inhibitors of recombinant human KGA protein

[0038] The amino acid sequence of the human KGA functional active region is as follows.

[0039] IPDFMSFTSH IDELYESAKK QSGGKVADYIPQLAKFSPDL WGVSVCTVDG QRHSTGDTKV

[0040] PFCLQSCVKP LKYAIAVNDL GTEYVHRYVG KEPSGLRFNK LFLNEDDKPH NPMVNAGAIV

[0041] VTSLIKQGVN NAEKFDYVMQ FLNKMAGNEY VGFSNATFQS ERESGDRNFA IGYYLKEKKC

[0042] FPEGTDMVGI LDFYFQLCSI EVTCESASVM AATLANGGFCPITGERVLSP EAVRNTLSLM

[0043] HSCGMYDFSG QFAFHVGLPA KSGVAGGILL VVPNVMGMMC WSPPLDKMGN SVKGIHFCHD

[0044] LVSLCNFHNY DNL; Detai Biotechnology was commissioned to perform recombinant expression of Pichia pastoris to obtain the functionally active protein KGA. After identification as an active recombinant protein by SDS-PAGE and human kidney-type glutaminase ELISA kit (catalog number 2H-KMLJh315182, brand Camilo), the protein concentration was adjusted to 1 mg / mL for later use.

[0045] The KGA inhibitor compound (Formula 1) obtained through molecular docking technology was adjusted to a final concentration of 0-100 μM in the enzyme reaction system. Specifically, it was prepared as: K₂PO₄ 0.15 M.

[0046] Tris-Acetate pH 8.65 0.00mM, Bovine Serum Albumin (BSA) 0.10mg / mL, EDTA 0.25mM, DTT 1.00mM, NAD 4.00mM, Glutamine 0-20mM, Glutamate Dehydrogenase (GDH) 1 unit, KGA

[0047] 1.00 μM, compounds 0-100 μM.

[0048] Prepare reaction systems without KGA enzyme and the compound according to the above concentrations. The reaction group without the compound serves as the blank control. In the final step, KGA protein is added, and the OD value change at 340 nm is measured within 10 min at 37°C. The final volume of the enzyme reaction system is 100.00 μL, with three replicates per reaction. Prepare reaction systems without KGA enzyme and the compound following the same steps. Incubate the compound and KGA enzyme at 37°C for 10 min, then add the enzyme reaction system to start the reaction, with a final volume of 100.00 μL. Measure the OD value change within 10 min, with three replicates per reaction condition. Measure the inhibitory OD value of the compound on the enzyme reaction system. BPTES is used as a positive control compound, and its inhibitory activity against KGA is measured using the same steps. Calculate the IC50 value of the compound against KGA. The results are shown in Table 1.

[0049] Table 1 IC50 values ​​of compounds against KGA

[0050] Compound type IC50 (μM) KGA inhibitor compound (Formula 1) 2.03±0.12 BPTES positive control 8.67±0.56

[0051] As can be seen from Table 1, the KGA inhibitor compound (Formula 1) of the present invention has good inhibitory activity against KGA.

[0052] Example 2: Cytotoxicity detection of KGA inhibitor compound (Formula 1)

[0053] Human fibrosarcoma cells HT1080 were purchased from Yunclone (Beijing) Biotechnology Co., Ltd., model YKLHT-1080. Cells frozen in liquid nitrogen were rapidly thawed in a 37°C water bath and transferred to 10cm cell culture dishes. 10mL of medium containing 10% FBS was added, and the cells were cultured at 37°C with 5% CO2 for 4 hours. After 4 hours, the supernatant and non-adherent cells were aspirated and discarded. The cells were rinsed once with isotonic FBS phosphate buffer and discarded. 10mL of cell culture medium containing 10% FBS was added. When the cell density reached approximately 80%, the cells were digested with trypsin and counted. The cells were diluted to a density of 50,000 / mL with medium containing 10% FBS and added to 96-well plates at 100.00μL / well (5000 cells / well) using a pipette (100.00μL of FBS was added to the outermost ring of the 96-well plate). The cells were cultured in a cell culture incubator for 12 hours. 5) The compound was then cultured in cell culture medium (containing 10% FBS). FBS was diluted to a concentration gradient of 200.00 μM, 100.00 μM, 50.00 μM, 25.00 μM, 12.50 μM, 6.25 μM, 3.12 μM, 1.56 μM, and 0.78 μM. Cell culture medium was aspirated from the 96-well plates (as thoroughly as possible) and cultured in three replicates at each concentration. Cells were incubated at 37°C for 48 h. CCK-8 solution was added to the cell culture medium without FBS to bring the final concentration of CCK-8 to 10%. 100.00 μL of 10% CCK-8 cell culture medium was added to the cell culture medium in the 96-well plates and incubated at 37°C for 30 min. The OD value of each well was measured at 450 nm, and the IC50 value of cell proliferation inhibition was calculated using GraphPad Prism 5.0. BPTES was used as a control. The results are shown in Table 2.

[0054] Table 2. IC50 values ​​of compounds against HT1080 tumor cells.

[0055] Compound type IC50 (μM) KGA inhibitor compound (Formula 1) 5.13±0.34 BPTES positive control 48.17±0.76

[0056] As shown in Table 2, the KGA inhibitor compound (Formula 1) of this invention exhibits significantly higher inhibitory activity against tumor cell proliferation than the previously reported positive control compound BPTES. It demonstrates superior activity in inhibiting tumor cell growth.

[0057] Example 3: Validation of the therapeutic effects of the compound and its combination therapy

[0058] Balb / C nude mice were provided by Beijing Vital River Laboratory Animal Co., Ltd.

[0059] Human fibrosarcoma HT1080 cells in logarithmic growth phase were digested with 0.25% trypsin at room temperature, centrifuged to remove the supernatant, washed twice with serum-free culture medium, counted, and the cell density was adjusted to approximately 5 × 10⁶ cells / year.10 A cell suspension of / L cells was prepared. 0.2ml of the prepared cell suspension was drawn and subcutaneously inoculated into the right axilla of Balb / C nude mice. Inoculation continued until the tumor volume reached 100mm². 3 At approximately 10:00 AM, the animals were divided into groups and given the drug.

[0060] Mice with successfully inoculated human fibrosarcoma were randomly divided into a model control group, high-, medium-, and low-dose compound groups, a positive control group, a combination therapy group, and a genistein monotherapy group. The compound was administered at doses of 2 mg / kg / 2 days, 1 mg / kg / 2 days, and 0.5 mg / kg / 2 days, twice weekly for 3 weeks via intraperitoneal injection. The control group received the same compound, with cyclophosphamide administered at 2 mg / kg / 2 days twice weekly via intraperitoneal injection for 3 weeks. The combination therapy group received the compound in combination with genistein (purchased from Sigma), with the compound at 1 mg / kg / 2 days and genistein at 2 mg / kg / 2 days, twice weekly via intraperitoneal injection for 3 weeks. The genistein monotherapy group received genistein at 2 mg / kg / 2 days twice weekly via intraperitoneal injection for 3 weeks. The model group received saline injections. Animals were sacrificed 24 hours after the last administration, tumors were excised and their weight measured. The average tumor weight and tumor inhibition rate were calculated for each group. Tumor inhibition rate (%) = (average tumor weight in the control group - average tumor weight in the treatment group) / average tumor weight in the control group * 100%. Results are as follows: Figure 1 As shown.

[0061] from Figure 1 It can be seen that, compared with the model group, each treatment group has a significant tumor-suppressing effect (P<0.01). Furthermore, regarding the dosage of the compound, the tumor inhibition rate gradually increases with increasing dosage. The therapeutic effect of the compound is also better than that of the positive control group. Combining the compound of this invention with the screened genistein drug exhibits a significant synergistic therapeutic effect, with a tumor inhibition rate reaching (98.14±2.92)%, which is far superior to existing drugs and demonstrates better therapeutic efficacy.

[0062] Before the mice in each treatment group were sacrificed, no obvious toxic side effects were observed. After the mice were sacrificed, liver samples were taken to test the corresponding enzyme activity levels, and no obvious symptoms of liver toxicity were found. This indicates that the compound has good safety.

[0063] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them. Although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features therein. Such modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

[0064] In the description of this specification, references to terms such as "one embodiment," "some embodiments," "example," "specific example," or "some examples," etc., indicate that a specific feature, structure, material, or characteristic described in connection with that embodiment or example is included in at least one embodiment or example of the invention. In this specification, illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in one or more embodiments or examples. Although embodiments of the invention have been shown and described, those skilled in the art will understand that various changes, modifications, substitutions, and variations can be made to these embodiments without departing from the principles and spirit of the invention, the scope of which is defined by the claims and their equivalents.

Claims

1. Use of a KGA inhibitor compound and genistein in the preparation of a kit for treating fibrosarcoma tumors, wherein the KGA inhibitor compound has the following structural formula:

2. The use as described in claim 1, characterized in that... The ratio of KGA inhibitor compound to genistein in the kit is 1:

2.

3. The use as described in claim 1, characterized in that... The fibrosarcoma tumor was caused by HT1080.

4. Use of a KGA inhibitor compound in the preparation of a medicament for treating fibrosarcoma tumors, wherein the structural formula of the KGA inhibitor compound is shown below:

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