JP6194322B2 - Lactate dehydrogenase inhibitors and pharmaceuticals containing the same - Google Patents

Description

  The present invention relates to a lactate dehydrogenase inhibitor containing a compound typified by stylpentol as an active ingredient and its pharmaceutical use.

  Malignant tumor (cancer) is still a disease with a high mortality rate, and development of a new therapeutic agent is desired. In human cancer, lactate dehydrogenase (LDH), a glycolytic metabolic enzyme, is highly expressed, especially its isoform 5 (LDH-5; also known as LDH-A, M-LDH) ) Expression is remarkable (Non-patent Document 1). Furthermore, it has also been shown that if the expression level of LDH-5 in this cancer cell is reduced by the gene silencing method, proliferation of the cancer cell can be suppressed (Non-patent Document 2). That is, it is now a well-known fact that the in vivo enzyme lactate dehydrogenase LDH-5 is a cancer growth control protein.

  Based on these findings, inhibitors of lactate dehydrogenase have recently been developed (Non-Patent Documents 3 to 6). These inhibitors have also been shown to inhibit cancer cell growth at the experimental level. However, there are no reports of clinical application of these compounds to humans.

  Since lactate dehydrogenase is a glycolytic metabolic enzyme, it is also an enzyme involved in energy production and lipid metabolism. In fact, it is known that inhibition of lactate dehydrogenase in the brain is involved in metabolic control of blood sugar levels and neutral fats (triglycerides) (Non-Patent Documents 7 to 8). That is, the inhibitor of lactate dehydrogenase is also involved in the control of various metabolic diseases (obesity, diabetes, hyperlipidemia, etc.).

  On the other hand, epilepsy is a neurological disease with a high morbidity rate of about 1% of the total population. In addition, 30% of epilepsy patients do not succeed with existing antiepileptic drugs. Against this background, various antiepileptic drugs have been developed, and more than 20 active ingredients are currently used as antiepileptic drugs.

  As a result of the development of many antiepileptic drugs, several examples of “second pharmaceutical use” in which the antiepileptic drug (compound) is also effective in different diseases have been reported. For example, zonisamide (CAS: 68291-97-4) was originally developed as a therapeutic agent for epilepsy (Patent Document 1), but subsequently treated for ischemic brain injury (Patent Document 2) and a therapeutic drug for neurodegenerative diseases (Patent Document 1). The utility as document 3) is shown. In particular, zonisamide as a therapeutic agent for neurodegenerative diseases is actually used as a therapeutic agent for Parkinson's disease, and therefore, the second pharmaceutical use of an epilepsy therapeutic agent has high industrial utility value.

  Styripentol (CAS: 49763-96-4, 4-dimethyl-1-[(3,4-methylenedioxy) -phenyl] -1-penten-3-ol, see formula (II) below) A new drug that was developed as an active drug (Patent Document 4), and subsequently has an inhibitory effect on intractable epilepsy (Dravet syndrome) (Non-Patent Documents 9 and 10), and is currently used in Europe and Japan as an epilepsy treatment drug. is there. Recently, it has been shown to be useful as a neuroprotective agent (Patent Document 5). However, there are no reports on pharmaceutical uses other than neurological diseases.

JP-A-53-077057 Japanese Patent Laid-Open No. 03-007226 International Publication WO99 / 33465 US Patent 3,910,959 US Patent Application Publication 2010/0240588

Goldman et al, Cancer Res 24: 389-399, 1964 Fantin et al, Cancer Cell 9: 425-434, 2006 Le et al, Proc Natl Acad Sci USA 107: 2037-2042, 2010 Granchi et al, J Med Chem 54: 1599-1612, 2011 Granchi et al, Eur J Med Chem 46: 5398-5407, 2011 Farabegoli et al, Eur J Pharm Sci 47: 729-738, 2012 Lam et al, Science 309: 943-947, 2005 Lam et al, Nat Med 13: 171-180, 2007 Perez et al, Epilepsia 40: 1618-1626, 1999 Chiron et al, Lancet 356: 1638-1642, 2000

  As shown in the prior art, lactate dehydrogenase is an in vivo protein that controls cancer and metabolic diseases. Several inhibitors of lactate dehydrogenase have also been developed. However, there are no lactate dehydrogenase inhibitors that have been shown to be safe for humans. Because lactate dehydrogenase is a glycolytic enzyme that is very important in the energy metabolic pathway, serious side effects (such as nutritional deficiencies and various organ abnormalities associated with it) This is because the worst is death). In fact, mice deficient in LDH-5 (LDH-A) are known to be fetal lethal (http://www.sanger.ac.uk/mouseportal/phenotyping/MBCB/ viability-at-weaning /), an extremely important protein for life. Against this background, inhibitors of lactate dehydrogenase reported so far have been observed to inhibit cancer cell growth at the laboratory level, but serious side effects are expected at the current technical level. Therefore, it is not realistic as a therapeutic agent for humans.

  That is, if there is a lactate dehydrogenase inhibitor with minimal side effects on humans, its industrial use such as cancer drugs and metabolic disease drugs is immeasurable, but there are no reports of such inhibitors. An object of the present invention is to provide a lactic acid dehydrogenase inhibitor that can be an active ingredient of a therapeutic drug for cancer or a therapeutic drug for metabolic diseases, and has minimal side effects on humans.

  The present inventors have made a bold hypothesis that a compound having a lactate dehydrogenase inhibitory action exists among pharmaceuticals (specifically, antiepileptic drugs) with limited side effects on humans. If present, it becomes an invention of a lactate dehydrogenase inhibitor with minimal side effects. Since epilepsy is a brain disease characterized by excessive excitement of brain electrical activity, most antiepileptic drugs act on regulatory proteins of electrical activity such as channels, synapses, and transporters. On the other hand, lactate dehydrogenase is a glycolytic metabolic enzyme. That is, an antiepileptic drug and a lactate dehydrogenase inhibitor are a combination that is difficult to imagine in the current state of the art. However, the present inventors have confirmed that the anti-epileptic drug Stilpentol (approved in Europe in 2007 and approved in Japan on September 28, 2012) is an inhibitor of lactate dehydrogenase (such as LDH-5), And it discovered that it actually had an anticancer action, and completed this invention.

  In one aspect, the present invention provides a lactate dehydrogenase inhibitor containing a specific compound as an active ingredient. In a further aspect, the present invention provides a pharmaceutical containing the lactate dehydrogenase inhibitor as an active ingredient. The present invention includes the following inventions.

  [1] An inhibitor of lactate dehydrogenase (LDH), comprising a compound represented by the formula (I).

In the formula (I), R ¹ and R ² represent one of hydrogen and the other of a hydroxyl group, or oxygen bonded together to a carbon atom through a double bond.
[2] The inhibitor according to [1], wherein the compound represented by the formula (I) is a compound (styripentol) represented by the formula (II).

[3] The inhibitor according to [1] or [2], wherein the lactate dehydrogenase is isoform 1 (LDH-1) or isoform 5 (LDH-5).
[4] The inhibitor according to any one of [1] to [3], wherein the lactate dehydrogenase is derived from a human.

[5] An anticancer agent comprising the lactate dehydrogenase according to any one of [1] to [4] as an active ingredient.
[6] A therapeutic drug for metabolic diseases, comprising the lactate dehydrogenase according to any one of [1] to [4] as an active ingredient.

  In addition, the invention as described above is a method for inhibiting lactate dehydrogenase that reacts with the specific compound, a treatment method for administering the lactate dehydrogenase inhibitor, and a lactate dehydrogenase inhibitor for the specific compound. It is obvious to those skilled in the art that it can be converted into an invention expressed as use, use of the lactate dehydrogenase inhibitor as an active ingredient of a pharmaceutical (in the manufacture of a pharmaceutical), and the like.

  In the prior art, lactate dehydrogenase inhibitors were compounds that could have serious side effects that could be life-threatening. However, in the present invention, it was found that steripentol administered to humans as an actual pharmaceutical product is a lactate dehydrogenase inhibitor. Side effects such as somnolence, loss of appetite, and ataxia have been reported to stiripentol (from the package insert of the anti-epileptic drug Diacommit [brand name]). That is, a “lactate dehydrogenase inhibitor with limited side effects” that could not be expected from the prior art is provided.

  Lactic acid dehydrogenase LDH-5 is an in vivo protein that is involved in the growth of cancer cells. By inhibiting its activity, the growth of cancer cells is suppressed. The use of styripentol as a cancer drug (second pharmaceutical use) ”is easily assumed, and it has been confirmed that it actually has an anticancer effect. Considering the social background in which the birth of a new anticancer drug is strongly desired due to its high mortality rate, styripentol has already been confirmed to be safe for humans as an epilepsy treatment. Is considered to have extremely high industrial utility value. In addition, since lactate dehydrogenase is a molecule on the metabolic pathway, it can be expected as a therapeutic drug for metabolic diseases.

FIG. 1 is a graph showing the enzyme activity of lactate dehydrogenase activity (LDH) when 20 types of antiepileptic drugs are allowed to act (see Examples). The enzyme activity (control value) when no antiepileptic drug was used was 100%. FIG. 2 is a graph showing the production rate of pyruvic acid from lactic acid by LDH-1 and LDH-5 when styripentol is allowed to act (see Examples). A, B: Michaelis Menten plot. C, D: Line weaver bark plot. FIG. 3 is a graph showing the production rate of lactic acid from pyruvic acid by LDH-1 and LDH-5 when styripentol is allowed to act (see Examples). A, B: Michaelis Menten plot. C, D: Line weaver bark plot. FIG. 4 is a graph showing the growth-inhibiting action when various types of cultured cells (cancer cells and normal cells) are administered with stilipentol (see Examples).

-Lactate dehydrogenase inhibitor-
The lactate dehydrogenase inhibitor of the present invention contains a compound represented by the formula (I) as an active ingredient.

In the formula (I), R ¹ and R ² represent one of hydrogen and the other of a hydroxyl group, or oxygen bonded together to a carbon atom through a double bond. Specifically, the compound represented by the formula (I) includes the compound represented by the formula (II) and the compound represented by the formula (III).

  A particularly preferred compound in the present invention is a compound represented by the formula (II), that is, styripentol. Styripentol has stereoisomers represented by formula (IIa) and formula (IIb). In the present invention, stilipentol may be a mixture of both stereoisomers (racemate) or a purified product of one of the stereoisomers.

  In addition, there is a possibility that a compound represented by the formula (I), particularly an analog (derivative or similar compound) of styripentol can also be used as a lactate dehydrogenase inhibitor according to the present invention. The inhibitory activity against lactic acid dehydrogenase (the amino acid sequence and the three-dimensional structure of the protein may differ somewhat depending on the species or isozyme from which it is derived) and the toxicity to the administration target can be evaluated by known methods. A suitable analog may be selected in consideration of the result.

  The compound represented by the formula (I) can be synthesized according to a known method (for example, see Patent Document 1). Stilipentol, which is used as an active ingredient of antiepileptic agents, can be easily obtained.

  When preparing the lactate dehydrogenase inhibitor of the present invention, other compounds having inhibitory activity of lactate dehydrogenase can be used in combination, if necessary, but are represented by the formula (I) It is preferable to use only the compound, particularly only highly safe stypentol.

  Although the use of the lactate dehydrogenase inhibitor of the present invention is not particularly limited, as described later, it is preferable to use it as an active ingredient of a pharmaceutical for treating a specific disease. For other purposes, it may be used to inhibit the activity of lactate dehydrogenase in any cell in vivo, ex vivo, in vitro. For example, the lactate dehydrogenase of the present invention may be used by adding it to cells cultured in vitro for the development of pharmaceuticals or the study of biological reactions involving lactate dehydrogenase. Included in the use of dehydrogenase inhibitors.

  Lactate dehydrogenase (LDH) is a protein having a molecular weight of about 140 kD, and is a tetramer composed of two types of subunits, M (muscle or A) and H (heart or B). Due to their binding mode, lactate dehydrogenase has five isozymes of LDH-1 (H4), LDH-2 (H3M1), LDH-3 (H2M2), LDH-4 (H1M3) and LDH-5 (M4). Exists. The lactate dehydrogenase inhibitor of the present invention can be used for any of the above lactate dehydrogenases, LDH-1 (LDH-B), which is a tetramer of only H, and a tetramer of only M It shows inhibitory activity against both of LDH-5 (LDH-A), but it shows higher inhibitory activity against LDH-5.

  In addition, the lactate dehydrogenase inhibitor of the present invention is not limited to human-derived lactate dehydrogenase but also other animals having lactate dehydrogenase, such as mice, rats, guinea pigs, rabbits, goats, cats, dogs, pigs. And lactate dehydrogenase derived from mammals such as monkeys.

Lactate dehydrogenase catalyzes the interconversion of lactic acid and pyruvate, and at the same time, the interconversion of NADH and NAD ⁺ occurs simultaneously (see the following formula). In addition, what exists normally in the living body is L-lactic acid. In the brain, the conversion of pyruvate to L-lactic acid is performed by LDH-5 (LDH-A) in astrocytes (astrocytic cells), and L-lactic acid taken up in neurons (nerve cells) is converted to pyruvate. The conversion is performed by LDH-1 (LDH-B). In cancer cells, energy (ATP) is actively produced by conversion of pyruvic acid to lactic acid.

Whether stilipentol is an inhibitor of lactate dehydrogenase depends on whether the enzyme activity of lactate dehydrogenase decreases (usually with statistical significance) when reacted with the lactate dehydrogenase. Can be confirmed. Here, since stilipentol causes non-competitive (non-antagonistic) inhibition against lactate dehydrogenase, Vmax (the substrate concentration was made infinite depending on the presence or absence of an inhibitor based on the Michaelis-Menten equation kinetics) Reaction rate) decreases, while Km (concentration of substrate when Vmax / 2 is reached) does not change. Vmax and Km can be obtained by a known method. For example, the concentration of NAD ⁺ produced in the reaction solution from the conversion of pyruvate to lactic acid, or conversely, the concentration of NADH produced in the reaction solution from the conversion of lactic acid to pyruvate is determined by spectrophotometry. Each reaction rate can be calculated by measuring using a meter and applying it to the Michaelis-Menten equation. It is also possible to calculate Vmax (1 / Vmax: y intercept) and Km (-1 / Km: x intercept) by converting the Michaelis-Menten equation to the Lineweaver-Burk equation and creating the plot. .

−Pharmaceuticals−
The pharmaceutical product of the present invention is caused by a disease in which a lactate dehydrogenase inhibitor has an effect of treatment or (recurrence) prevention, that is, expression of lactate dehydrogenase or enhanced activity, and the activity of lactate dehydrogenase is reduced. By inhibiting the disease, it is possible to target a diseased tissue or symptom of the disease that exhibits effects such as treatment, improvement, mitigation, or prevention.

  A malignant tumor (cancer) is mentioned as a typical example of the disease used as the application object of the pharmaceutical of this invention. That is, the pharmaceutical agent of the present invention is an anticancer agent in one embodiment. The energy production of cancer cells greatly depends on the metabolism of pyruvate to lactic acid even in the absence of hypoxia, and as a result, the expression of lactate dehydrogenase, particularly LDH-5, is significantly enhanced. Therefore, by inhibiting the enzyme activity of LDH-5 by administering a lactate dehydrogenase inhibitor, it is possible to kill or suppress the growth of cancer cells. Examples of malignant tumor (cancer) include malignant lymphoma (Hodgkin lymphoma, non-Hodgkin lymphoma, etc.), gastric cancer, penile cancer, pharyngeal cancer (nasopharyngeal cancer, oropharyngeal cancer, hypopharyngeal cancer, etc.) ), Vulvar cancer, pituitary fibroma, hepatocellular carcinoma, thymoma, mycosis fungoides, unknown primary cancer, myelodysplastic syndrome, uterine cancer (cervical cancer, uterine body cancer, uterus) Sarcoma), chorionic disease, esophageal cancer, renal pelvic cancer, ureteral cancer, glioma, renal cell cancer, pancreatic cancer, pancreatic endocrine tumor, testicular tumor, prostate cancer, colon cancer ( Cecal cancer, colon cancer, rectal cancer, etc.), multiple myeloma, bile duct cancer, gallbladder cancer, vagina cancer, mesothelioma, acoustic schwannoma, soft tissue sarcoma, breast cancer, brain tumor, lung cancer, leukemia ( Acute myeloid leukemia, acute lymphoblastic leukemia, adult T-cell leukemia lymphoma, chronic myeloid leukemia, chronic lymphocytic leukemia ), Skin cancer (malignant melanoma, etc.), bladder cancer, chronic myeloproliferative disease, ovarian cancer, ovarian germ cell tumor, etc. As long as it is a thing, it will not specifically limit. As an example, the anticancer agent according to the present invention exhibits excellent anticancer activity against blood cancer such as multiple myeloma and leukemia.

  Another representative example of the disease to which the pharmaceutical of the present invention is applied is a metabolic disease involving energy production and lipid metabolism via glycolysis (pyruvic acid, lactic acid), such as obesity, diabetes, and hyperlipidemia. It is done.

  A pharmaceutical product is usually prepared as a pharmaceutical composition. The pharmaceutical product (pharmaceutical composition) of the present invention contains at least a lactate dehydrogenase inhibitor as an active ingredient, but may further contain other drugs or functional ingredients as necessary. For example, if it is an anticancer agent, you may use together with the lactate dehydrogenase inhibitor other compounds which have anticancer activity. Moreover, you may contain the suitable carrier (excipient, diluent) and other various additives in preparing a pharmaceutical as needed.

  The pharmaceutical dosage form and administration method are not particularly limited, and may be appropriate depending on the use of the pharmaceutical. For example, dosage forms for oral administration such as tablets, capsules, soft capsules, granules, powders, fine granules, (dry) syrups, solutions, suspensions, and for subcutaneous, intramuscular or intravenous administration It is possible to select from dosage forms for parenteral administration such as injections, infusions, and suppositories.

  It is also preferable to prepare a targeting agent so that a lactate dehydrogenase inhibitor acts in a specific tissue. For example, if it is an anticancer agent, it may be prepared as a liposome preparation designed to be selectively delivered to cancer cells.

  A pharmaceutical composition of such a dosage form can be produced by a general production method. For example, if it is a dosage form for oral administration, additives such as excipients, disintegrants, binders, lubricants, suspending agents, tonicity agents, emulsifiers, sweeteners, fragrances, coloring agents, etc. It can manufacture by mixing said active ingredient with a conventional method and shape | molding. Among these, as an excipient, cellulose derivatives (for example, crystalline cellulose, methylcellulose, hydroxypropylcellulose, hydroxypropylmethylcellulose, etc.), polyvinylpyrrolidone, dextrin, starch, lactose, mannitol, sorbitol, vegetable oils (for example, corn oil, cottonseed oil, coconut oil) , Almond oil, olive oil, peanut oil, etc.), oil esters such as medium chain fatty acid glyceride oil, mineral oil, glycerin esters such as tricaprylin and triacetin, alcohols such as ethanol, saline, propylene glycol, polyethylene glycol, Animal fats and oils, petrolatum and the like can be mentioned.

  On the other hand, in the case of injections, ampules are prepared by dissolving the above active ingredients in an appropriate diluent (physiological saline, glucose injection, lactose injection, mannitol injection, etc.) and sterilizing by filtration sterilization. In addition, it can also be manufactured as a powder injection mixed with sodium chloride or an injection in a lyophilized form based on the Japanese Pharmacopoeia. In addition, as additives, adjuvants such as polyethylene glycol and surfactants, and carriers such as ethanol, liposomes, and cyclodextrins can be used.

  The content of the active ingredient in the pharmaceutical composition may be adjusted within an appropriate range, but is usually 0.05 to 99% by weight, preferably 0.1 to 70% by weight, based on the total weight of the pharmaceutical composition. More preferably, the amount is 0.1 to 50% by weight. Further, a pharmaceutically acceptable carrier may be contained in an amount of usually 1 to 99.95% by weight, preferably 30 to 99.9% by weight, more preferably 50 to 99.9% by weight.

  Such a pharmaceutical product of the present invention may be administered in an appropriate amount and number of active ingredients in consideration of the purpose, age of the subject to be administered (patient), body weight, severity of disease, administration route, pharmacokinetics, and the like. .

Method The activity of lactate dehydrogenase was measured by a general method based on the light absorption of NADH. Lactic acid dehydrogenase is an enzyme that interconverts pyruvate and lactic acid. At that time, NADH and NAD are also mutually converted. For lactic acid dehydrogenase activity from lactic acid to pyruvate, lactic acid (1-20 mM), NAD (200 μM) and lactate dehydrogenase were added, and the reaction rate of the produced NADH was measured. On the other hand, for the lactate dehydrogenase activity from pyruvate to lactic acid, pyruvate (0.05-1.0 mM), NADH (200 μM) and lactate dehydrogenase were added, and the decreasing reaction rate of NADH was measured. Both reactions were performed in 100 mM phosphate buffered saline (pH 7.4) at 37 ° C, and the absorbance at 340 nm of NADH was measured in a quartz cell (total volume: 3 ml) using a spectrophotometer. The reaction rate was determined by measuring for 3.5 minutes at second intervals. As a lactate dehydrogenase, screening was first performed with mammalian lactate dehydrogenase (from porcine heart; 0.03 units), and the hit compounds were analyzed in detail with two types of human lactate dehydrogenases (human LDH-1, LDH-5). investigated. The amount of lactate dehydrogenase is set so that the maximum rates of the two types of lactate dehydrogenase are approximately the same. Regarding the activity of lactate → pyruvic acid, LDH-1 is 0.01 units, LDH-5 is 0.03 units, and pyruvin. Regarding the activity of acid → lactic acid, LDH-1 was 0.005 units and LDH-5 was 0.01 units. The inhibitory effect of lactate dehydrogenase was examined by adding various antiepileptic compounds (500 μM) to these enzyme reaction systems. The analysis was performed using Igor Pro 6, the reaction rate was the Michaelis-Menten equation, and the Lineweaver-Burk plot was a regression analysis with a straight line.

Furthermore, anticancer activity against cultured cells was examined by the following method. Cell lines derived from multiple myeloma cells (KMS11, KMS12PE), cell lines derived from human acute T cell leukemia cells (Jurkat), cell lines derived from chronic myelogenous leukemia cells (K562), cells derived from B lymphocytes of Burkitt lymphoma patients The effect of stiripentol was examined on the strain (Raji) and non-cancer cell-derived cell fibroblast cell line (Hs27). Per well of 96-well multiplate, suspended cells were seeded with 1x10 ⁵ cells, and adherent cells were seeded with 5x10 ⁴ cells. The next day, styripentol was added to final concentrations of 250 μM, 125 μM, 62.5 μM, and 31.5 μM (DMSO as a solvent was 0.2%). In addition, a control was added to add only a solvent that does not contain styripentol. These were cultured for 2 days, and the number of each cell was counted. The cell proliferation inhibitory effect was evaluated by standardizing the number of cells in each concentration group containing styripentol, assuming that the number of cells in control after 2 days of culture was 100%.

Results First of all, there are 20 antiepileptic drugs (acetazolamide, carbamazepine, diazepam, ethosuximide, ferbamate, gabapentin, bromide, lamotrigine, levetiracetam, oxcarbazepine, phenobarbital, phenytoin, primidone, rufinamide, stipentolmate, tiagaprotemate, pyramate , Vigabatrin, zonisamide), the inhibitory effect of lactate dehydrogenase activity from lactic acid to pyruvate was investigated. Lactic acid concentration was screened at 20 mM using lactate dehydrogenase from porcine heart. As a result, although most antiepileptic compounds did not show any action, only stilipentol showed inhibitory activity (Fig. 1; n = 4 to 5 for each antiepileptic drug. Enzyme activity without control [control value]).

  Therefore, next, we examined whether or not stilipentol has an inhibitory effect on human lactate dehydrogenase, including detailed kinetic analysis. First, we investigated the effect of styripentol on lactic acid dehydrogenase activity from lactic acid (1-20 mM) to pyruvate. It was found that styripentol has an inhibitory effect on both human LDH-1 and human LDH-5. (FIG. 2A, B; n = 3-7 at each point). In particular, the inhibitory effect on human LDH-5 was remarkable. Furthermore, by creating a Lineweaver-Burk plot from these data, it was also clarified that the lactic acid → pyruvate conversion inhibition mode by stilipentol is non-competitive inhibition (FIGS. 2C and 2D). Next, the action of stilpentol was examined for the lactate dehydrogenase activity from pyruvate (0.05-1.0 mM) to lactic acid. With regard to the enzyme activity in the reverse direction (pyruvic acid → lactic acid), it has been clarified that styripentol shows an inhibitory action in both human LDH-1 and human LDH-5 (FIGS. 3A and 3B; n = 3-8). With regard to the enzyme activity in the opposite direction, the inhibitory effect of stilipentol on human LDH-5 was more prominent. Furthermore, by creating a Lineweaver-Burk plot from these data, it became clear that the pyruvate-lactate conversion inhibition mode by stilipentol is also non-competitive inhibition (FIGS. 3C and D).

  From the above, it was revealed that stilipentol is an inhibitor of lactate dehydrogenase (LDH-1, LDH-5).

  It is a well-known fact that inhibitors of lactate dehydrogenase (especially LDH-5 inhibitors) have anticancer effects (Le et al, Proc Natl Acad Sci USA 107: 2037-2042, 2010; Granchi et al, J Med Chem 54: 1599-1612, 2011; Granchi et al, Eur J Med Chem 46: 5398-5407, 2011; Farabegoli et al, Eur J Pharm Sci 47: 729-738, 2012). Therefore, the anticancer effect of stilipentol was examined using cancer-derived cultured cells (FIG. 4). Even when stilipentol was administered to fibroblast Hs27 as normal cells, no significant growth inhibitory effect was observed at concentrations up to 250 μM (FIG. 4A; n = 9). On the other hand, when styripentol was administered to multiple types of cancer cells, a clear growth inhibitory action was observed at a lower concentration of styripentol compared to the normal cells shown in FIG. 4A (FIGS. 4B-F; each). N = 6-9 for cell types). In particular, Jurkat (FIG. 4B; n = 9), which is a leukemia cell, and KMS11 (FIG. 4E; n = 6), KMS12PE (FIG. 4F; n = 9), which are multiple myeloma cells. The effect was observed.

  In summary, it has become clear that stilipentol is not only an inhibitor of lactate dehydrogenase, but also has an anticancer effect.

Claims (6)

An inhibitor of lactate dehydrogenase (LDH), comprising a compound represented by formula (I).
In the formula (I), R ¹ and R ² represent one of hydrogen and the other of a hydroxyl group, or oxygen bonded together to a carbon atom through a double bond. The inhibitor according to claim 1, wherein the compound represented by the formula (I) is a compound represented by the formula (II) (stilpentol).
  The inhibitor according to claim 1 or 2, wherein the lactate dehydrogenase is isoform 1 (LDH-1) or isoform 5 (LDH-5).   The inhibitor according to any one of claims 1 to 3, wherein the lactate dehydrogenase is derived from a human.   An anticancer agent comprising the lactate dehydrogenase according to any one of claims 1 to 4 as an active ingredient.   A therapeutic agent for metabolic diseases comprising the lactate dehydrogenase according to any one of claims 1 to 4 as an active ingredient.