CN115969975A - Application of PHGDH inhibitor in prevention and/or treatment of colorectal cancer metastasis - Google Patents

Abstract

The invention belongs to the field of biological medicine, and discloses application of PHGDH serving as a target in inhibiting colorectal cancer metastasis. The invention also discloses an application of the PHGDH inhibitor, the PHGDH inhibitor is used as an active ingredient for inhibiting colorectal cancer metastasis, can prevent and/or treat colorectal cancer metastasis, effectively prolongs the life of a patient while improving the life quality of the patient, and has good clinical application prospect.

Description

Application of PHGDH inhibitor in prevention and/or treatment of colorectal cancer metastasis

Technical Field

The invention belongs to the field of biological medicines, and relates to application of a PHGDH inhibitor in inhibiting colorectal cancer metastasis, which is used for preventing and/or treating colorectal cancer metastasis.

Background

Colorectal cancer is one of the most common digestive tract tumors, and seriously threatens human health. The liver is the most major target organ for colorectal cancer metastasis, and about 40% to 70% of colorectal cancer patients eventually develop liver metastasis and/or colorectal cancer metastasis throughout the course of the disease. Part of colorectal cancer patients are detected as liver metastases accompanied by the liver metastases at the definite diagnosis time, and are concurrent liver metastases and/or colorectal cancer metastases; in addition, liver metastasis was not detected in patients with confirmed diagnosis, but was sporadic after primary focal radical treatment of colorectal cancer. Liver metastasis is the leading cause of death in colorectal cancer patients, and therefore, colorectal cancer liver metastasis is also a major and difficult point in colorectal cancer treatment.

Colorectal cancer liver metastasis is a complex process, and is characterized by the invasion of tumor cells from primary colorectal sites into blood vessels, the escape of tumor cells (CTC) from the body in blood circulation to kill and invade the liver by immunity, the formation of new metastases in the liver and other processes. In order to improve the prospect of treating and preventing colorectal cancer metastasis, the development of the medicine which can effectively inhibit colorectal cancer metastasis and reduce the high mortality rate caused by colorectal cancer metastasis is urgently needed, and the medicine has important significance.

Disclosure of Invention

The technical problem to be solved by the invention is to overcome the defects of the drugs for treating colorectal cancer metastasis in the prior art, and a large number of experimental screening and research prove that a phosphoglycerate dehydrogenase (PHGDH) inhibitor can be used as an effective component for inhibiting colorectal cancer metastasis. PHGDH, also known as phosphoglycerate dehydrogenase, catalyzes the oxidation of 3-phosphoglycerate to 3-phosphooxopyruvate, a key step in serine biosynthesis. The PHGDH inhibitor can inhibit the enzyme activity of PHGDH, thereby inhibiting the synthesis of serine and downstream products thereof. The PHGDH inhibitors reported so far can be used for inhibiting the growth of colorectal cancer, but the application of the PHGDH inhibitors in inhibiting colorectal cancer metastasis has not been reported. The mechanism by which PHGDH inhibitors inhibit cancer growth is: inhibition of PHGDH prevents glucose-derived serine synthesis, inhibits nucleotide synthesis in the cell, and thus leads to cell cycle arrest. The mechanism by which PHGDH inhibitors inhibit cancer metastasis is: inhibiting PHGDH inhibits serine synthesis pathway, and reduces downstream S-adenosylmethionine (SAM) content, inhibiting expression of cell migration related gene, thereby inhibiting cancer metastasis.

To achieve the above objects, the present invention aims to provide the use of PHGDH as a target for the preparation of a medicament for the prevention and/or treatment of colorectal cancer metastasis.

One aspect of the present invention is to provide the use of PHGDH inhibitors in the preparation of a medicament for inhibiting colorectal cancer metastasis, and to provide novel methods and pharmaceutical compositions for preventing and/or treating colorectal cancer metastasis.

One aspect of the present invention is to provide a PHGDH inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof and its use in the preparation of a medicament for inhibiting colorectal cancer metastasis.

Further preferably, the PHGDH inhibitor is of formula C ₂₀ H ₂₃ F ₃ N ₄ The structure of the compound of S (CAS number 1916571-90-8) is shown in formula I:

in another aspect of the present invention is provided a pharmaceutical composition for inhibiting colorectal cancer metastasis, comprising a therapeutically effective amount of the PHGDH inhibitor or its pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof as described above in combination with at least one pharmaceutically acceptable excipient.

In another preferred embodiment, the dosage form of the pharmaceutical composition is selected from: injection, injectable sterile powder, tablet, capsule, spirit, powder, granule, syrup, solution, tincture, aerosol, powder spray, or suppository.

In the present invention, the pharmaceutical composition may also be administered in combination with at least one immune system modulator and/or at least one agent that inhibits colorectal cancer metastasis.

In another aspect, the invention provides the use of the pharmaceutical composition in the preparation of a medicament for preventing and/or treating colorectal cancer metastasis.

In another aspect of the invention there is provided a method of inhibiting colorectal cancer metastasis in vitro (preferably non-therapeutically), the method comprising: administering to a subject in need of inhibition of colorectal cancer metastasis an effective amount of a PHGDH inhibitor as described above or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof.

Compared with the prior art, the invention has the beneficial effects that:

the invention creatively uses the PHGDH inhibitor, in particular the PHGDH inhibitor shown in the formula I, for inhibiting colorectal cancer metastasis. The experiment of inhibiting colorectal cancer metastasis in vitro shows that the PHGDH inhibitor of the invention has the unexpected effect of inhibiting colorectal cancer metastasis, achieves the purposes of effectively treating colorectal cancer metastasis and improving the prognosis of patients, makes up the defects of treatment means such as operation, radiotherapy, chemotherapy and the like, and has no toxic or side effect. The PHGDH inhibitor has great clinical application value in preparing the medicine for preventing and/or treating colorectal cancer metastasis, and can be used as an effective method for intervening colorectal cancer metastasis.

Drawings

FIG. 1 shows the effect of PHGDH inhibitors on colorectal cancer cell metastasis in vitro.

FIG. 2 shows the effect of PHGDH inhibitors on the growth and metastasis of colorectal cancer in mice in vivo. Wherein A is the weight of subcutaneous tumor; b is the number of metastatic nodules on the liver; c is paraffin embedding and HE staining to prove that the metastatic nodules on the liver are tumor tissues.

Detailed Description

The invention provides a novel active ingredient for colorectal cancer metastasis treatment. Combined with the years of research on the catalytic action of PHGDH by the inventor, a series of in vivo and in vitro functional experiments surprisingly find that the inhibition of PHGDH can effectively inhibit the metastasis of colorectal cancer cells, and has excellent effect of inhibiting the metastasis of colorectal cancer. The inventors determined the metastatic effect of PHGDH inhibitors on colorectal cancer cells in vitro by the method of inoculating colorectal cancer cells into Transwell chamber and staining, and confirmed that PHGDH inhibitors could inhibit liver metastasis of colorectal cancer by constructing mouse colorectal cancer tumor model, measuring mouse subcutaneous tumor weight and counting the number of metastatic nodules on liver.

The invention provides an application of PHGDH as a target in preparing a medicament for preventing and/or treating colorectal cancer metastasis.

The PHGDH as a target specifically comprises the following components: the PHGDH is used as an action object, and the medicines or the preparations are screened to find the medicine capable of inhibiting the PHGDH as a candidate medicine for preventing and/or treating colorectal cancer metastasis. The PHGDH inhibitor represented by formula I is obtained by screening PHGDH as an action object, and can be used as a medicine for inhibiting colorectal cancer metastasis. In addition, PHGDH can be used as an active ingredient such as an antibody drug, a nucleic acid drug, etc.

The invention provides application of a PHGDH inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof in medicines for preventing and/or treating colorectal cancer metastasis.

Further, the inhibitor is a small molecule compound, an antibody, a nucleic acid or a lentivirus-encapsulated sequence.

Such nucleic acids include, but are not limited to: antisense oligonucleotides, double-stranded RNA (dsRNA), ribozymes, small interfering RNA prepared by endoribonuclease III or short hairpin RNA (shRNA).

The amount of the drug for colorectal cancer metastasis administered is a dose sufficient to reduce transcription or translation of the human PHGDH gene, or to reduce expression, activity or function of the human PHGDH protein, such that expression of the human PHGDH gene is reduced by at least 50%, specifically 55%, 60%, 65%, 70%, 75%, 80%, 90%, 95%, 99% or 100%.

The medicament necessarily comprises the PHGDH inhibitor as an effective component of the aforementioned effects.

In the medicament, the effective component for playing the functions can be only a PHGDH inhibitor, and other molecules capable of playing the functions can also be contained.

That is, the PHGDH inhibitor is the only active ingredient or one of the active ingredients of the drug.

The medicament can be a single-component substance or a multi-component substance.

The form of the drug is not particularly limited, and can be various forms of substances such as solid, liquid, gel, semifluid, aerosol and the like.

The drug is primarily directed to a subject that is a mammal. The mammal is preferably a rodent, artiodactyla, perissodactyla, lagomorpha, primate, or the like. The primate is preferably a monkey, ape or human.

Further preferably, the PHGDH inhibitor has the molecular formula of C ₂₀ H ₂₃ F ₃ N ₄ The compound of S has a CAS number of 1916571-90-8 and a structure shown in formula I:

in the compound and the application thereof, the active compound, namely PHGDH inhibitor or pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate has the activity of inhibiting colorectal cancer metastasis and has the effect of preventing and/or treating colorectal cancer metastasis.

In the compound and the application thereof, the PHGDH inhibitor can be used in the form of pharmaceutically or physiologically acceptable salt or ester. By "pharmaceutically acceptable salt" is meant generally any salt which is physiologically tolerable (meaning generally non-toxic, in particular as a result of counterions) when used in an appropriate manner in therapy, in particular when applied or used in humans and/or mammals. These physiologically acceptable salts may be formed with cations or bases and in the context of the present invention, especially when administered in humans and/or mammals, they are to be understood as being salts formed by at least one compound provided according to the invention, usually an acid (deprotonated), such as an anion, and at least one physiologically tolerated cation, preferably an inorganic cation. In the context of the present invention, salts with alkali metals and alkaline earth metals, and salts with ammonium cations (NH 4 +) may be specifically included, and specifically may include, but are not limited to, salts with (mono) or (di) sodium, (mono) or (di) potassium, magnesium or calcium. These physiologically acceptable salts may also be formed with anions or acids, and in the context of the present invention, in particular when administered in humans and/or mammals, they are to be understood as being salts formed by at least one compound, usually protonated, such as a cation and at least one physiologically tolerable anion, provided according to the present invention. The salts and esters of the PHGDH inhibitor include but are not limited to salts and esters formed with: hydrochloric acid, hydrobromic acid, sulfuric acid, citric acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, oxaloacetic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, or isethionic acid. Salts of halides are also suitable. Other salts include: salts with alkali metals or alkaline earth metals, such as sodium, potassium, calcium or magnesium.

In the compounds and the use thereof of the present invention, the term "prodrug" refers to a prodrug that undergoes a metabolic or chemical reaction in a human body to be converted into the active PHGDH inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof after being taken by an appropriate method.

In the compound and the application thereof, the active compound can be combined with other treatment means, such as operations, radiotherapy, chemotherapy and targeted treatment.

In the compounds and uses thereof of the present invention, when the active compound is used in combination with other therapeutic agents, the active compound is co-administered with the other therapeutic agents. "coadministration" means simultaneous administration via the same or different routes, or sequential administration via the same or different routes, in the same formulation or in two different formulations. By "sequential" administration is meant having a time difference in seconds, minutes, hours, or days between the administration of two or more different compounds.

The invention also provides a pharmaceutical composition comprising the PHGDH inhibitor or the pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof as described above, and optionally one or more pharmaceutically acceptable carriers and vehicles. Such acceptable carriers, vehicles such as sterile water or physiological saline, stabilizers, excipients, antioxidants (ascorbic acid, etc.), buffers (phosphoric acid, citric acid, other organic acids, etc.), preservatives, surfactants (PEG, tween, etc.), chelating agents (EDTA, etc.), binders, and the like. Moreover, other low molecular weight polypeptides may also be present; proteins such as serum albumin, gelatin, and immunoglobulin; amino acids such as glycine, glutamine, asparagine, arginine, and lysine; saccharides or carbohydrates such as polysaccharides and monosaccharides; sugar alcohols such as mannitol and sorbitol. When an aqueous solution for injection is prepared, for example, physiological saline, an isotonic solution containing glucose or other auxiliary drugs, such as D-sorbitol, D-mannose, D-mannitol, sodium chloride, may be used in combination with an appropriate solubilizer, such as alcohol (ethanol, etc.), polyhydric alcohol (propylene glycol, PEG, etc.), nonionic surfactant (Tween 80, HCO-50), etc.

In the pharmaceutical composition of the present invention, the PHGDH inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof may be a single active ingredient, or may be combined with other active ingredients to form a combined preparation. Preferably, the pharmaceutical composition may also be used in combination with compounds and agents useful for inhibiting colorectal cancer metastasis.

The content of the active ingredient (PHGDH inhibitor or its pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate) in the pharmaceutical composition of the present invention is generally a safe and effective amount which should be adjusted by those skilled in the art, for example, the amount of the active ingredient administered will generally depend on the body weight of the patient, the type of application, the condition and severity of the disease, for example, the amount of the active ingredient administered will generally be 1 to 1000mg/kg/day, 20 to 200mg/kg/day, 1 to 3mg/kg/day, 3 to 5mg/kg/day, 5 to 10mg/kg/day, 10 to 25mg/kg/day, 25 to 30mg/kg/day, 30 to 40mg/kg/day, 40 to 60mg/kg/day, 60 to 80mg/kg/day, 80 to 100mg/kg/day, 100 to 150mg/kg/day, 150 to 200mg/kg/day, 300mg/kg/day, 500mg/kg/day, or 500 mg/kg/day.

One skilled in the art can consider the effective amount to be administered depending on the severity of the condition and the health and age of the subject. An effective amount will typically vary from 0.01ng/kg body weight to about 100mg/kg body weight.

The active ingredients provided by the present invention or pharmaceutical compositions containing the active ingredients may be adapted for any form of administration, and may be administered orally or parenterally, for example, by pulmonary, nasal, rectal and/or intravenous injection, more specifically intradermal, subcutaneous, intramuscular, intraarticular, intraperitoneal, pulmonary, buccal, sublingual, buccal, nasal, transdermal, vaginal, oral or parenteral administration; the injection administration includes intravenous injection, intramuscular injection, subcutaneous injection and the like, transdermal administration and the like.

As used herein, the dosage form of the pharmaceutical composition is selected from: injection, injectable sterile powder, tablet, pill, capsule, lozenge, spirit, powder, granule, syrup, solution, tincture, aerosol, powder spray, or suppository. Those skilled in the art can select a suitable formulation according to the administration mode, for example, a formulation suitable for oral administration may be, but is not limited to, a pill, a tablet, a chewable agent, a capsule, a granule, a solution, a drop, a syrup, an aerosol, a powder spray, etc., and a formulation suitable for parenteral administration may be, for example, a solution, a suspension, a reconstitutable dry preparation, a spray, etc., and for rectal administration, a suppository may be, for example, a sterile powder for injection, etc.

Wherein the tablet, lozenge, pill, capsule, etc. may also contain the following components: binders, such as gums, acacia, corn starch or gelatin; excipients, such as dicalcium phosphate; disintegrating agents such as corn starch, potato starch, alginic acid, and the like; lubricants, such as magnesium stearate; sweetening agents such as sucrose, lactose or saccharin may be added, or flavoring agents such as peppermint, oil of wintergreen, or cherry flavoring. When the unit dosage form is a capsule, it may contain, in addition to the materials described above, a liquid carrier. Various other materials may be present in the form of coatings or to modify the physical form of the unit dosage form. For example, tablets, pills, or capsules may be coated with shellac, sugar or both. A syrup or elixir may contain the active compound, sucrose as a sweetening agent, methyl and propylparabens as preservatives, a colorant and flavoring such as cherry or orange flavor. Any material used in preparing any unit dosage form should be pharmaceutically pure and substantially non-toxic in amounts used. In addition, the active compounds may be incorporated into sustained release preparations or formulations.

The invention also provides a method for preventing and/or treating colorectal cancer metastasis by administering a PHGDH inhibitor to a subject.

The subject may be a mammal or a mammalian colorectal cancer cell. The mammal is preferably a rodent, artiodactyla, perissodactyla, lagomorpha, primate, or the like. The primate is preferably a monkey, ape or human. The colorectal cancer cell can be an ex vivo colorectal cancer cell.

The subject may be a patient suffering from colorectal cancer metastasis or an individual for whom prevention and/or treatment of colorectal cancer metastasis is desired. Or the subject is a patient with colorectal cancer metastasis or an isolated cancer cell of an individual for whom prevention and/or treatment of colorectal cancer metastasis is desired.

The PHGDH inhibitor may be administered to a subject before, during, or after receiving colorectal cancer metastasis therapy.

As used herein, the colorectal cancer (also referred to as colorectal cancer, colon cancer, rectal cancer, or intestinal cancer) refers to a cancer when a cancer occurs in the colon or rectum (a portion of the large intestine).

As used herein, the colorectal cancer metastasis refers to the invasion of the lymphatic vessels from the primary site by colorectal cancer tumor cells, and the blood vessels or other pathways are brought to grow to form tumors of the same type as the primary site tumor. The colorectal cancer liver metastasis means that colorectal cancer tumor cells invade the liver from a primary part through various possible ways, and form tumors of the same type as the primary part in the liver.

The present invention further provides a method of reducing, delaying or inhibiting colorectal cancer metastasis comprising administering said PHGDH inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof to an individual in need of treatment.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the methods of the present invention. The preferred embodiments and materials described herein are exemplary only.

According to the research results, a new method for diagnosing and treating the gene is further explored and developed, so that more choices can be provided for diagnosing and treating colorectal cancer metastasis patients.

PHGDH inhibitors

Refers to a molecule having an inhibitory effect on PHGDH. Having inhibitory effects on PHGDH include, but are not limited to: inhibiting the expression or activity of PHGDH.

Specifically, the inhibition of the expression of PHGDH may be the inhibition of transcription or translation of the PHGDH gene, and specifically, may be: making the gene of PHGDH non-transcribed or reducing the transcription activity of the gene of PHGDH, or making the gene of PHGDH non-translated or reducing the translation level of the gene of PHGDH.

Inhibiting the activity of PHGDH means to decrease the viability of PHGDH. Preferably, the viability of PHGDH is reduced by at least 10%, preferably by at least 30%, more preferably by at least 50%, even more preferably by at least 70%, and most preferably by at least 90% compared to that before inhibition.

The regulation of gene expression of PHGDH can be performed by one skilled in the art using conventional methods, such as gene knock-out, homologous recombination, interfering RNA, etc.

The inhibition of PHGDH gene expression was confirmed by PCR and Western Blot analysis of the expression level.

Preferably, the expression of the PHGDH gene is reduced by at least 10%, preferably by at least 30%, more preferably by at least 50%, more preferably by at least 70%, still more preferably by at least 90%, most preferably completely absent from the PHGDH gene as compared to the wild type.

Small molecule compounds

The invention refers to a compound which is composed of several or dozens of atoms and has the molecular mass of less than 1000.

Preparation of medicine for preventing or treating colorectal cancer metastasis

Nucleic acid molecules that reduce PHGDH gene expression in colorectal cancer cells can be utilized; and/or, a PHGDH gene interfering nucleic acid construct; and/or the PHGDH gene interferes with lentivirus to be used as an effective component for preparing a medicament for preventing or treating colorectal cancer metastasis. Generally, the medicament may comprise one or more pharmaceutically acceptable carriers or excipients in addition to the active ingredients, according to the requirements of different dosage forms.

By "pharmaceutically acceptable" is meant that the molecular entities and compositions do not produce adverse, allergic, or other untoward reactions when properly administered to an animal or human.

The "pharmaceutically acceptable carrier or adjuvant" should be compatible with the active ingredient, i.e., capable of being blended therewith without substantially diminishing the effectiveness of the drug as is normally the case. Specific examples of some substances that can serve as pharmaceutically acceptable carriers or adjuvants are sugars, such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose and its derivatives, such as sodium methylcellulose, ethylcellulose and methylcellulose; powdered gum tragacanth; malt; gelatin; talc; solid lubricants, such as stearic acid and magnesium stearate; calcium sulfate; vegetable oils such as peanut oil, cottonseed oil, sesame oil, olive oil, corn oil and cocoa butter; polyhydric alcohols such as propylene glycol, glycerin, sorbitol, mannitol, and polyethylene glycol; alginic acid; emulsifiers, such as Tween; wetting agents, such as sodium lauryl sulfate; a colorant; a flavoring agent; tableting agents, stabilizers; an antioxidant; a preservative; pyrogen-free water; isotonic saline solution; and phosphate buffer, and the like. These materials are used as needed to aid in the stability of the formulation or to aid in the enhancement of the activity or its bioavailability or to produce an acceptable mouthfeel or odor upon oral administration.

References herein to "comprising," "including," and the like are to be construed in an inclusive sense as opposed to an exclusive or exhaustive sense; i.e., meaning "including but not limited to".

Reference herein to a "therapeutically effective amount" generally refers to an amount which, after an appropriate period of administration, is effective to treat the disease conditions as set forth above.

References herein to "therapeutic" and "prophylactic" are to be understood in their broadest sense. The term "therapeutic" does not necessarily imply that the mammal is receiving treatment until complete recovery. Similarly, "prophylactic" does not necessarily mean that the subject will not eventually metastasize. Thus, treatment and prevention includes alleviation of the symptoms of, or prevention or reduction of the risk of, a particular disorder. The term "prevention" is to be understood as reducing the severity of the onset of a particular condition. Treatment may also reduce the severity of existing conditions or the frequency of acute episodes.

In the present invention, the subject or subject to be treated therapeutically or prophylactically is preferably a mammal, such as, but not limited to, a human, a primate, a livestock animal (e.g., sheep, cattle, horse, donkey, pig), a pet animal (e.g., dog, cat), a laboratory test animal (e.g., mouse, rabbit, rat, guinea pig, hamster) or a captured wild animal (e.g., fox, deer). The subject is preferably a primate. The subject is most preferably a human.

The active ingredient, pharmaceutical composition and method thereof of the present invention can be used for preventing colorectal cancer metastasis, not only for preventing the establishment of cancer lesions in the liver at the early stage of colorectal cancer metastasis, but also for administration prior to colorectal cancer metastasis as a preventive treatment method. Alternatively, the occurrence of colorectal cancer metastasis can be avoided by pre-administering the active ingredient or pharmaceutical composition.

Before the present embodiments are further described, it is to be understood that the scope of the invention is not limited to the particular embodiments described below; it is also to be understood that the terminology used in the examples is for the purpose of describing particular embodiments, and is not intended to limit the scope of the present invention; in the description and claims of the present application, the singular forms "a", "an" and "the" include plural referents unless the context clearly dictates otherwise.

Test object

All mice were housed in a pathogen-free facility at the central innovation of molecular cell science of the academy of sciences of china. Animals were randomly assigned to control and experimental groups. All animal experiments were approved by the Institute of Animal Care and Use (IACUC) at the central agency of molecular cell science of the Chinese academy of sciences, and met all relevant ethical norms.

Example 1 Effect of PHGDH inhibitors of formula I on colorectal cancer cell metastasis in vitro

HCT116 cells were treated with PHGDH inhibitor of formula I (NCT-503, 10. Mu.M) and changes in cell transfer were detected.

Mixing HCT116 (1X 10) ⁵ ) Cells were seeded in the upper chamber of a Transwell chamber with 8- μ M pore size and cultured using serum-free medium, in which 10 μ M of PHGDH of formula I was added to the experimental group (+) and the corresponding DMSO was added to the blank control group (-). Complete medium containing 10% fetal bovine serum was used in the plates. After 12 hours of culture, cells from the Transwell chamber were fixed with 4% paraformaldehyde. The cells that did not migrate to the top of the screen were removed with a cotton swab and the cells at the bottom of the screen were stained with 0.4% crystal violet. At the same time, cells were plated individually onto plates without Transwell chambers to determine the total number of attached cells. The relative transfer of cells was calculated by dividing the number of transferred cells by the total number of cells and further normalized to the control group. For each experiment, the number of cells under the microscope (magnification 100 ×) in 5 random fields was counted and 3 independent Transwell chambers were analyzed.

HCT116 cells were treated with PHGDH inhibitors, which were found to inhibit metastasis of HCT116 cells (fig. 1).

1.2 Effect of PHGDH inhibitors of formula I on colorectal cancer liver metastasis in vivo

Fresh colorectal cancer patients tumor tissue was from Shanghai cancer center, university of double denier. The surgically excised tumor tissue was immediately placed in DMEM media supplemented with 2%v/v penicillin/streptomycin and transferred to a biosafety cabinet on ice for manipulation. Cutting the tumor tissue into 60mm ³ And (3) selecting appropriate tissues for small blocks, and inoculating the tissues to the lateral abdomen subcutaneous of the BALB/c nude mouse. When the xenograft reached 6 weeks, mice were operated and tumors dissected, which were cut into small pieces (60 mm) ³ ) And transplanted again into a new generation of mice for constitutionA colorectal cancer patient-derived xenograft (PDX) model was constructed, and 1 week after tumor tissue implantation into mice, experimental group (+): the PHGDH inhibitor (NCT-503, 25mg/kg body weight) was injected intraperitoneally into mice, and the blank control (-): the reagents containing 5% ethanol, 35% polyethylene glycol 300 and 60% aqueous solution of hydroxypropyl- β -cyclodextrin as a 30% solution were injected intraperitoneally into mice. Once daily for 3 weeks.

The experimental results are as follows:

mice were sacrificed 5 weeks after inoculation, subcutaneous tumors were removed and weighed, and the results are shown in fig. 2A, and as can be seen from fig. 2A, the tumor weight of the mice in the experimental group was significantly lower than that of the blank control group, indicating that the PHGDH inhibitor of the present invention can inhibit the growth of colorectal cancer tissues (p <0.001, which is statistically significant). The mouse liver was removed, and the number of metastatic nodules on the liver was counted and counted, with the results shown in FIG. 2B; liver tissue was fixed with 4% paraformaldehyde, paraffin-embedded and HE-stained, demonstrating that liver metastases were indeed tumor tissue, as shown in fig. 2C; as can be seen from fig. 2B and fig. 2C, the PHGDH inhibitor of the present invention can significantly inhibit liver metastasis of colorectal cancer, with statistical differences.

While the invention has been described with respect to a preferred embodiment, it will be understood by those skilled in the art that the foregoing and other changes, omissions and deviations in the form and detail thereof may be made without departing from the scope of this invention. Those skilled in the art can make various changes, modifications and equivalent arrangements, which are equivalent to the embodiments of the present invention, without departing from the spirit and scope of the present invention, and which may be made by utilizing the techniques disclosed above; meanwhile, any changes, modifications and variations of the above-described embodiments, which are equivalent to those of the technical spirit of the present invention, are within the scope of the technical solution of the present invention.

Claims (10)

1. Use of PHGDH as a target for the preparation of a medicament for the prevention and/or treatment of colorectal cancer metastasis.
2. Use of a phgdh inhibitor or a pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof for the manufacture of a medicament for the prevention and/or treatment of colorectal cancer metastasis.
3. 3. The use of claim 2 wherein the pharmaceutically acceptable salts, esters comprise salts or esters of said PHGDH inhibitor with: hydrochloric acid, hydrobromic acid, sulfuric acid, citric acid, tartaric acid, phosphoric acid, lactic acid, pyruvic acid, acetic acid, succinic acid, oxalic acid, fumaric acid, maleic acid, oxaloacetic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, or isethionic acid.
4. 4. The use of claim 2 wherein the PHGDH inhibitor is capable of specifically inhibiting the expression of PHGDH or inhibiting the activity of PHGDH;

   wherein, inhibiting the expression of PHGDH means inhibiting the transcription or translation of the PHGDH gene, including: making the gene of PHGDH non-transcribed or reducing the transcription activity of the gene of PHGDH, or making the gene of PHGDH non-translated or reducing the translation level of the gene of PHGDH; the expression of PHGDH is reduced by at least 10% compared to before inhibition;

   wherein, inhibiting the activity of PHGDH means reducing the activity of PHGDH; the PHGDH activity is reduced by at least 10% compared to that before inhibition.
5. 5. The use of claim 4 wherein the expression or activity of PHGDH is reduced by 50% or more.
6. 6. The use of claim 2 wherein the PHGDH inhibitor has the formula C ₂₀ H ₂₃ F ₃ N ₄ S, the structure is shown as the following formula I:

   
7. 7. the use of claim 2, wherein the metastasis is hepatic metastasis.
8. 8. The use of claim 2 wherein the PHGDH inhibitor prevents, inhibits and/or blocks metastasis of colorectal cancer by inhibiting the activity of PHGDH, inhibiting the serine synthesis pathway.
9. 9. A pharmaceutical composition for preventing and/or treating colorectal cancer metastasis, comprising the PHGDH inhibitor or its pharmaceutically acceptable salt, ester, isomer, prodrug, polymorph or solvate thereof according to any of claims 1 to 8, and a pharmaceutically acceptable carrier, vehicle.
10. 10. The pharmaceutical composition of claim 9, wherein the metastasis is hepatic metastasis; and/or the pharmaceutical composition is an injection, an injectable sterile powder, a tablet, a pill, a capsule, a lozenge, spirit, a powder, a granule, a syrup, a solution, a tincture, an aerosol, a powder cloud agent, or a suppository.

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