KR20080088335A - Nk cell activating molecules, nk cells and pharmaceutical compositons comprising the same - Google Patents

Abstract

NK(natural killer) cell-activating molecules are provided to increase cell-killing ability of NK cells by increasing the expression of natural cytotoxicity receptors such as NCR1, NCR2 and NCR3 and death ligand on the surface of NK cell, so that the NK cells are useful as anticancer and antiviral agents. Compounds represented by the formula(1) are useful as NK cell-activating molecules, wherein R1 is hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl or alkoxy; R2 and R3 are each independently hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl, alkoxy, amino, cyano, carboxyl, aryl or -R23R24; R23 is linear or branched alkylene, linear or branched alkenylene, or linear or branched alkynylene; R24 is hydroxy, alkylamino, dialkylamino, amino, cyano, carboxyl, optionally substituted aryl, heteroaryl or hetero ring compound, thiourea, isothiourea, alkylthio, thiocyanate, sulfamoyl or alkylsulfonyl; and R4 to R11 are each independently hydrogen, alkyl, cycloalkyl or alkoxy. The NK cells are treated by the NK cell-activating molecules through culture in a medium containing the NK cell-activating molecules at 30-37 deg. C for 1-48 hours.

Description

NK cell activating molecules, NK cells and pharmaceutical compositons comprising the same

The present invention increases the expression of Natural Cytotoxicity Receptor (NCR) and Suicide Ligand in Natural Killer Cells (hereinafter referred to as NK cells). Compounds capable of activating; Its production method; NK cells treated with the compound; Treating NK cells with the compound; And it relates to a pharmaceutical composition comprising the compound or NK cells.

NK cells function to remove host cells infected with tumor cells, bacteria, intracellular parasites or viruses without prior sensitization by antigens, reject inappropriate bone marrow transplantation, and regulate T cell immune responses. It is a behavioral cell that acts in the first line of immune system defense mechanisms. NK cells are particularly sensitive to target cells (e.g., tumor cells or viruses) that do not express the major histocompatibility complex (MHC), or which have significantly reduced expression of class I MHC. Infecting host cells), and is distinct from T cells or B cells, which are the same immune cells in terms of expression of CD markers and cytokine profile (Ljunggren et al., In search of the 'missing self': MHC molecules and NK cell recognition.Imunmun.Today. 1990; 11: 237-44). NK cells are known to play an important role in the body's anticancer immunity, showing a particularly strong killing ability against blood cancer, various solid cancers.

The mechanisms by which NK cells or cytotoxic T cells are cytotoxic to target cells are relatively well known. The representative mechanism is that these cells recognize target cells through activating receptors and secrete cytotoxic substances such as perforin or granzyme A, B to induce cell death or apoptosis of target cells. will be. NK cells, like cytotoxic T cells, have a mechanism for inducing apoptosis by recognizing the target receptor's death receptors through the death ligands (Fas-L, TNF, and TRAIL) on the surface of NK cells. Using such killings, kills tumor cells or infected cells directly. In addition, NK cells also serve to enhance adaptive immunity by delivering debris from target cells to T cells through macrophages or other antigen presenting cells.

The immunological function of NK cells is governed by the balance of the stimulatory signal that causes its killing function and the inhibitory signal that inhibits the killing function. Specifically, NK cells that receive strong activation signals attack and remove target cells, and NK cells that receive strong inactivation signals keep target cells alive.

The killing functions of NK cells include ADCC (antibody-dependent cell cytoxicity) and natural killing (Natural Killing). ADCC induced by the antigen-antibody complex is mediated by Fcγ receptor III (FcγRIII, CD16) and spontaneous killing is mediated by NK cell activating receptors. While ADCC proceeds via Pl3-kinase (phosphatidylinositol-3-kinase), the specific mechanism is somewhat different in that natural killing occurs through protein kinase C (PKC). However, both ADCC and spontaneous killing require the activation of PTK (Protein Tyrosine Kinase) and have in common that they are blocked by inactivation signals delivered by inactivating receptors of NK cells. Since the killing function of the NK cells depends on the balance between the activation signal and the inactivation signal, the NK cells can be distinguished and removed from normal host cells and infected or tumorized cells.

Target specificity of NK cells is determined by inactivating receptors, and inactivating receptors of NK cells, like T cell receptors, also recognize MHC of target cells. NK cells do not attack cells in which the MHC first antigen is normally expressed, but cells in which the MHC first antigen is not expressed or whose expression is significantly reduced are regarded as abnormal cells and attack. NK cell receptors with these functions have been identified in human and rat NK cells, and cDNA is cloned to reveal the primary structure of the protein. In general, since expression of the MHC first antigen is significantly reduced in tumor cells and host cells infected with viruses, the function of the inactivated receptors of NK cells is essential for selectively removing tumor cells or virus infected host cells.

In humans, there are several NK cell inactivation receptors identified so far. The above examples are specific to HLA-A, HLA-B, and HLA-C, and are inactive KIR (inhibitory Killer Ig-like receptors) belonging to the immunoglobulin superfamily, which interact with the MHC first antigen. KIR2DL (p58 KIR) and KIR3DL (p70 KIR). It has been reported that KIR2DL binds to HLA-C protein and KIR3DL binds to HLA-A and KLA-B. KIR is mainly expressed in NK cells, but is also expressed in some T cells, and it is known that down-regulation of not only the killing function of NK cells but also the immune response of T cells. Another example of an NK cell inactivating receptor is NKG2A and NKG2B of the C-type lectin line, which are complexed with CD94 molecules to be specific for human leukocyte antigen type E alleles. Intracellular parts of these inhibitory receptors contain immunoreceptor tyrosine inhibitory motifs (ITIMs) that inhibit NK cell activity.

Examples of NK cell activating receptors are IgG Fc receptors, which are specific for the CD16 and HLA-E alleles involved in ADCC and are co-receptors for p50 KIR of NKG2C, NKG2E, and activated KIR lines that bind CD94 molecules. 2B4, NKp80, etc. are mentioned. These NK cell activation receptors have a short intracellular portion, which must be combined with adapter molecules such as FcR γ-chains, CD3 ζ-chains, DAP10 or DAP12, which have immunoreceptor tyrosine-based activation motifs (ITAMs) to activate activation signals. I can deliver it.

In addition to the above, NK cells include NCR1 (NKp46), NCR2 (NKp44), and NCR3 (NCR1), which are natural cytotoxicity receptors (NCR), which play the greatest role in NK cells attacking tumor cells or cells infected with viruses. NKp30) and activating receptors such as NKG2D (Bottino C, et al. The human natural cytotoxicity receptors (NCR) that induce HLA class I-independent NK cell triggering. Hum. Immunol. 2000; 61: 1-6; Pende D , et al. Role of NKG2D in tumor cell lysis mediated by human NK cells: cooperation with natural cytotoxicity receptors and capability of recognizing tumors of nonepithelial origin.Eur. J. Immunol. 2001; 31: 1076-86; Moretta A, et al (2001) Activating receptors and coreceptors involved in human natural killer cell-mediated cytolysis.Annu. Rev. Immunol. 19, 197-223.). NCR is a recent discovery among NK cell activating receptors, where (1) the expression of receptors is mainly restricted to NK cells, and (2) redirected killing when cross-linking NCR with monoclonal antibodies. assay) can confirm the target cell killing function of NK cells, and (3) inducing the natural killing of NK cells in that NK cell mediated cytotoxicity is suppressed when the receptor is masked by monoclonal antibody It is believed to be a key activating receptor (Moretta, A. et al. Natural cytotoxicity receptors that trigger human NK-cell-mediated cytolysis. Immunol. Today. 2000; 21 (5): 228-34; Biron et al., (1999 Natural killer cells in antiviral defence: function and regulation by innate cytokines.Annu. Rev. immunol. 17, 3546-3556 .; De Maria A, et al. (2001) Identification, molecular cloning and functional characterization of NKp46 and NKp30 natural cytotoxicity recep tors in Macaca Fascicularis NK cells.Eur. J. Immunol. 31, 3546-3556.).

NCR1 is expressed in all activated and inactivated NK cells, but not in T cells or other cells. In addition, NCR1 is specifically expressed not only in humans but also in NK cells such as monkeys, mice or cows. The expression of analogs in NK cells such as mice suggests that NK cell activation mechanisms through NCR1 are preserved between species. NCR1 is also the only receptor expressed on human NK cells that can recognize ligands on the surface of rat cells. NCR1 is a receptor belonging to the Ig superfamily with two Ig domains, and its intracellular portion consists of 30 amino acids. In addition, since it does not contain ITAMs, it has a positively charged arginine (Arg) at the cell membrane transit site and binds to the CD3ζ-chain to transmit a signal. NCR1 induces calcium flow (Ca ^{2 +} mobilization), cytotoxicity and cytokine secretion in NK cells (Sivori, S. et al. ( 1997) P46, a novel natural killer cell-specific surface molecule which mediates cell activation. J. Exp. Med. 186, 1129-1136.). The shielding of the receptors is also known to inhibit lysis against several tumors, including lung cancer, liver cancer, breast cancer, melanoma and EBV-transformed cell lines. Gazit et al. Recently discovered that NCR1 is an immunoreceptor that plays a key role in the removal of Influenza virus in vivo (Gazit R, et al. (2006) Lethal influenza infection in the absence of the natural killer cell receptor gene Ncr1. Nature Immunol. 7, 517-523.). It has also been shown that NCR1, together with NCR2, recognizes haemagglutinin of influenza virus as a ligand to activate NK cells. (Mandelboim O, et al. (2001) Recognition of haemagglutinins on virus-infected cells by NKp46 activates lysis by NK cells.Nature 409, 1055-1060 .; Arnon TI, et al. (2001) Recognition of viral haemagglutinins by NKp44 but not by NKp 30. Eur. J. Immunol. 31, 2680-2689.). Vankayalapati et al. Also found that NCR1 on the surface of NK cells plays a critical role in killing cells infected with Mycobacterium tuberculosis (Vankayalapati R, et al. (2002) The NKp46 receptor contributes to NK cell lysis of mononuclear phagocytes infected with an intracellular bacterium J. Immunol. 168, 3451-3457.).

The second NCR, NCR2, is expressed only in activated NK cells incubated with IL-2, but not in lymphocytes or αβ T cells isolated from peripheral blood, and thus may be a special marker of activated NK cells. Unlike NCR1 and NCR3, NCR2 has not yet been found in non-human species and has the characteristic of being expressed in some γδ T cells in addition to activated NK cells. NCR2 belongs to the Ig superfamily, which has one Ig domain in the extracellular part, and because there is no ITAMs in the intracellular part, NCR2 transmits a signal by binding to DAP12 through Lysine, which is positively charged at the cell membrane transit site. (Cantoni, C. et al (1999) NKP44, a trigering receptor involved in tumor cell lysis by activated human natural killer cells, is a novel member of the immunoglobulin superfamily. J. Exp. Med. 189, 787-796.) .

The third NCR, NCR3, works with NCR1 and NCR2 to induce cytotoxicity against several target cells and is expressed in all NK cells freshly isolated or incubated with IL-2. The extracellular part of NCR3 is an Ig superfamily composed of one Ig domain, and because it does not have ITAMs in the intracellular part, it binds to the CD3ζ-chain and transmits a signal. Analogues also exist in mouse NK cells, indicating that the mechanism of NK cell activation through NCR3 is also conserved between species (Pende, D. et al. Identification and molecular characterization of NKp30, a novel triggering receptor involved in natural). cytotoxicity mediated by human natural killer cells.J.Exp. Med. 1999; 190: 1505-16).

There is a close correlation between the expression level of NCRs and apoptosis of NK cells. That is, NK cells that express strongly NCR show strong cell killing ability against target cells, whereas weakly expressing NK cells show weak cell killing ability (Sivori, S. et al. NKp46 is the major triggering receptor involved in the natural cytotoxicity of fresh of cultured human NK cells: correlation between surface density of NKp46 and natural cytotoxicity against autologous, allogeneic or xenogeneic target cells.Eur. J. Immunol. 1999; 29: 1647-55; Moretta L, Moretta A (2004 Unraveling natural killer cell function: triggering and inhibitory human NK receptors.EMBO J. 23, 255-259.). In addition, NK extracellular expression of each NCR is characterized by being closely related to each other and expressed in the same aspect. That is, NK cells that express strongly NCR1 express strongly NCR3, and when activated, express strongly NCR2. On the other hand, NK cells weakly expressing NCR1 also weakly express NCR3 and weakly express NCR2 upon activation.

The research that has been conducted to date on the expression and regulation of NCR is as follows. First, the CD3ζ-chain, an adapter molecule that binds NCR1 and NCR3, does not affect the expression of these receptors on the surface of NK cells, but is essential for the expression of NCR2 (Pessino, A. et al. Molecular cloning of NKp46: a novel member of the immunoglobulin superfamily involved in triggering of natural cytotoxicity. J. Exp. Med. 1998; 188: 953-60). Cortisol, well known as human corticosteroid, inhibits the expression of NCR1 and NCR3 in NK cells, and prolactin or IL-2, an immune enhancing cytokine, is known to increase the expression of NCR1 and NCR3. (Mavoungou E, Bouyou-Akotet MK, Kremsner PG (2004) Effects of prolactin and cortisol on natural killer (NK) cell surface expression and function of natural cytotoxicity receptors (NKp46, NKp44 and NKp30). Clinical and Exp. Immunol. 139 , 287-296.). In particular, IL-2 has been known as a cytokine for activating NK cells for a long time, and many animal and clinical trials have been performed. Prolactin has also been shown to enhance anticancer function of NK cells (Sun et al., (2003) Recombinant human prolactin improves anti-tumor effects of murine natural killer cells in vitro and in vivo. Neuroimmunomodulation 10, 169-176.). It has also been reported that TGFβ1 decreases NCR3 and NKG2D expression on the surface of NK cells and consequently reduces the cytotoxicity of NK cells. Recently, Mavoungou et al. Reported that the cytotoxicity of NK cells was increased by IL-2 or prolactin, but decreased by cortisol (Mavoungou et al., (2004) Effects of prolactin and cortisol on natural killer (NK)). cell surface expression and function of natural cytotoxicity receptors (NKp46, NKp44 and NKp30) .Clinical and Exp. Immunol. 139, 287-296.). Taken together, these findings suggest that hormones such as IL-2, prolactin, TGFβ1, or cortisol regulate NK cell activity through NCR expression control.Increasing NCR expression increases NK cell apoptosis and, conversely, NCR It can be seen that the decrease in the expression of apoptosis of NK cells also decreases. In addition, this fact suggests that if the expression of NCR in NK cells is increased, the cytotoxicity of NK cells is also increased, and thus can be usefully used as a therapeutic agent for cancer or viral diseases.

In addition to the above facts, most of NK cells express a lot of NCR1, but there are also differences in the expression of NCR1 among NK cells, such as the presence of weakly expressing NK cells. However, aside from the above facts, studies on the mechanism by which NCR is expressed and regulated on the surface of NK cells are insignificant.

NK cells have the function of initiating and modulating adaptive immune responses as well as the killing function of target cells described above (Raulet DH (2004) Interplay of natural killer cells their receptors with the adaptive immune response. Immunol. 5, 996-1002; Delgi-Esposti MA & Smyth MJ (2005) Close encounters of different kinds: dendritic cells and NK cells take center stage.Nature Rev. Immunol. 5, 112-124.). In other words, NK cells are known to induce T-cell immune responses through IFN-γ secretion early in infection (Martin-Fontecha A, et al. (2004) Induced recruitment of NK cells to lymph nodes provides IFN-g for TH1 priming.Nature Immunol. 5, 1260-1265.), NK cells have recently been shown to interact with NK cells and myeloid cells to play an important role in the development of the next immune response. Research is ongoing on how to regulate adaptive immune responses. NK cells also interact with monocytes, such as dendritic cells or macrophage, and as a result, these immune cells interact with each other to activate next-generation immune responses, such as adaptive immune responses. It is known to let. The interaction and activation between NK cells and monocytes is known to be mediated by receptors on the cell surface as well as water soluble factors, and NCR3 is known to play an important role. These results suggest that activation of NK cells can activate not only natural immune responses but also adaptive immune responses involving T cells. Therefore, if NK cells are activated by increasing NCR expression on the surface of NK cells, antigen-presenting cells such as dendritic cells or macrophages may be activated, and ultimately, T cells may be activated.

The present invention has been made in consideration of the above-described contents, and synthesizes a compound capable of increasing the cytotoxicity of NK cells by increasing the expression of NCR and death ligand on the surface of NK cells, and analyzing the mechanism of action thereof. It aims at the application to the pharmaceutical use, such as anticancer agent and antiviral agent.

The present invention as a means for solving the above problems, a compound represented by the following formula (1), a pharmaceutically acceptable salt or solvate thereof; And it provides a method for producing the compound.

[Formula 1]

The present invention provides NK cells treated with the compound of the present invention as another means for solving the above problems.

As another means for solving the above problems of the present invention,

It provides a method for treating NK cells, comprising culturing the NK cells in the culture medium to which the compound of the present invention is added.

The present invention is another means for solving the above problems,

It provides a pharmaceutical composition comprising the compound of the present invention or NK cells treated with the compound, and a pharmaceutically acceptable carrier.

The compound of formula 1 of the present invention activates NCR and suicide ligands of NK cells. As such, NK cells in which NCR and suicide are activated have increased killing ability against target cells, and thus may be usefully used for medical purposes such as anticancer agents or antiviral agents. The present invention firstly, unlike the prior art that targets hormones or cytokines as a substance that modulates the activity of NK cells, first identified the fact that the organic compound of a specific structural formula increases the cell killing ability of NK cells, second It is distinguished from the prior art in that the first compound was found to dramatically increase the expression of NCR on the surface of NK cells, and as a result, the cell killing ability of NK cells is significantly increased. Third, the compounds of the present invention can significantly increase the expression of NCR1 as well as NCR2 and NCR3 on the cell surface by up to 30 times compared to conventional IL-2 or prolactin, which increased the expression of NCR by about 1.5 to 2 times. . Fourth, the compound of the present invention significantly increases the target cell killing ability of NK cells by significantly increasing the expression of not only NCR but also suicide, and therefore, the result of its use for medical use is also remarkably excellent.

The present invention relates to a compound represented by the following formula (1), a pharmaceutically acceptable salt or solvate thereof. The compound of the present invention is excellent in expressing NCR and suicide ligands FasL and TRAIL on NK cells. Since NK cells with activated NCR and suicide ligands on the cell surface have increased killing ability against tumor cells and infected cells, the compounds of the present invention or NK cells treated with the compounds are useful for medical applications such as the treatment of cancer and infectious diseases. Can be applied effectively. Hereinafter, in the present specification, 'compound represented by Chemical Formula 1' is used in the same meaning as 'NK cell Activating Molecule' or 'NKAM'.

[Formula 1]

In which R ₁ represents hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl or alkoxy;

R ₂ and R ₃ each independently represent hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl, alkoxy, amino, cyano, carboxyl, aryl or -R ₂₃ R ₂₄ ;

R ₂₃ in the above is linear or branched alkylene; Straight or branched alkenylene; And linear or branched alkynylene

R ₂₄ is hydroxy; Alkylamino; Dialkylamino; Amino; Cyano; Carboxyl; Substituted or unsubstituted aryl, heteroaryl or heterocyclic compounds; Thiourea; Isothiourea; Alkylthio; Thiocyanato; Sulfamoyl or alkylsulfonyl;

R ₄ to R ₁₁ each independently represent hydrogen, alkyl, cycloalkyl or alkoxy.

As used herein, the term “alkyl” means substituted or unsubstituted straight, branched, cyclic and / or complex alkyl consisting of any carbon-carbon single bond; "Alkenyl" means substituted or unsubstituted straight, branched, cyclic and / or complex alkenyl comprising any carbon-carbon double bond; "Alkynyl" means a substituted or unsubstituted straight, branched, cyclic and / or complex alkynyl containing any carbon-carbon triple bond. The term 'cycloalkyl', as used herein, also forms a ring and may be substituted or unsubstituted, which may include any unsaturated bond and any heteroatom (eg O, N or S) in or on the ring. Cyclic cycloalkyl; "Aryl" is a substituent derived from an aromatic compound, and means a substituted or unsubstituted phenyl, biphenyl, naphthyl or anthracenyl ring system, and the like; 'Heteroaryl' means a substituted or unsubstituted 5-12 membered heterocyclic or aryl ring containing one or more heteroatoms selected from the group consisting of O, N and S, and includes furyl, pyrrolyl, pyrrolidinyl, Thienyl, pyridinyl, piperidyl, indolyl, quinolyl, thiazole, benzthiazole and triazoles, and the like.

In addition, in the definition of the substituents of the compounds of the present invention, alkyl is preferably alkyl having 1 to 12 carbon atoms, more preferably alkyl having 1 to 8 carbon atoms, most preferably alkyl having 1 to 4 carbon atoms; Cycloalkyl is preferably cycloalkyl having 3 to 12 carbon atoms, more preferably cycloalkyl having 3 to 8 carbon atoms; Alkoxy is preferably alkoxy having 1 to 12 carbon atoms, more preferably alkoxy having 1 to 8 carbon atoms, and most preferably alkoxy having 1 to 4 carbon atoms; Alkenyl is preferably alkenyl having 2 to 12 carbon atoms, more preferably alkenyl having 2 to 8 carbon atoms, and most preferably alkenyl having 2 to 4 carbon atoms; Alkynyl is preferably alkynyl having 2 to 12 carbon atoms, more preferably alkynyl having 2 to 8 carbon atoms, and most preferably alkynyl having 2 to 4 carbon atoms.

Preferred examples of R ₁ in the compound of Formula 1 include hydrogen, halogen, hydroxy, alkyl having 1 to 12 carbon atoms, cycloalkyl having 3 to 12 carbon atoms, alkenyl having 2 to 12 carbon atoms, alkynyl having 2 to 12 carbon atoms, or And alkoxy having 1 to 12 carbon atoms, hydrogen or alkyl having 1 to 8 carbon atoms is more preferred, and hydrogen, methyl or ethyl is most preferred.

In addition, preferred examples of R ₂ or R ₃ in the compound of Formula 1 include hydrogen, hydroxy, alkyl having 1 to 12 carbon atoms, cycloalkyl having 3 to 12 carbon atoms, alkenyl having 2 to 12 carbon atoms, and alky having 2 to 12 carbon atoms. Aryl, alkoxy having 1 to 12 carbon atoms, amino, cyano, carboxyl, phenyl, naphthyl or -R ₂₃ R ₂₄ , wherein a preferred example of R ₂₃ is straight or branched chain alkyl having 1 to 12 carbon atoms Lene, and preferred examples of R ₂₄ include hydroxy, alkylamino having 1 to 12 carbon atoms; Dialkylamino having 2 to 24 carbon atoms; Aryl, heteroaryl, any one selected from the group consisting of phenyl, indolyl, pyrrolyl, furanyl, pyridinyl, piperidinyl, pyranyl, pyridazinyl, pyrimidinyl, pyrazinyl, piperazinyl and triazolyl Or heterocyclic compounds; Thiourea; Or isothiourea, wherein the aryl, heteroaryl, or heterocyclic compound is composed of hydroxy, amino, cyano, arylalkyl, aryl unsubstituted or substituted with alkyl, and alkyl substituted or unsubstituted with carboxyl. It may be substituted with one or more selected from the group.

More preferred examples of R ₂ or R ₃ are hydrogen, alkyl of 1 to 8 carbon atoms, cycloalkyl of 3 to 8 carbon atoms, alkenyl of 2 to 8 carbon atoms, alkynyl of 2 to 8 carbon atoms, alkoxy of 1 to 8 carbon atoms. Or —R ₂₃ R ₂₄ , wherein R ₂₃ may be a straight or branched chain alkylene having 1 to 8 carbon atoms, and an example of R ₂₄ may be hydroxy or alkylamino having 1 to 8 carbon atoms. ; Dialkylamino having 2 to 16 carbon atoms; Any one aryl, heteroaryl or heterocyclic compound selected from the group consisting of phenyl, pyridinyl and triazolyl; Thiourea; Or isothiourea, wherein the aryl, heteroaryl or heterocyclic compound is hydroxy, amino, cyano, arylalkyl having 7 to 12 carbon atoms, aryl and carboxyl unsubstituted or substituted with alkyl having 1 to 4 carbon atoms. It may be substituted with one or more selected from the group consisting of alkyl having 1 to 12 carbon atoms unsubstituted or substituted with.

R ₂ or R ₃ is most preferably hydrogen, alkyl of 1 to 4 carbon atoms, cycloalkyl of 3 to 6 carbon atoms, alkenyl of 2 to 4 carbon atoms, alkynyl of 2 to 4 carbon atoms, of 1 to 4 carbon atoms. Alkoxy or —R ₂₃ R ₂₄ , wherein R ₂₃ is straight or branched alkylene having 1 to 4 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 4 carbon atoms; Dialkylamino having 2 to 8 carbon atoms; Substituted or unsubstituted triazolyl; Thiourea or isothiourea, wherein the triazolyl substituent is at least one selected from the group consisting of alkyl having 1 to 4 carbon atoms unsubstituted or substituted with benzyl and carboxyl.

Further preferred examples of R ₄ to R ₁₁ in the compound of Formula 1 include hydrogen, alkyl having 1 to 12 carbon atoms, cycloalkyl having 3 to 12 carbon atoms or alkoxy having 1 to 12 carbon atoms, and hydrogen, having 1 to 8 carbon atoms. More preferably alkyl or alkoxy having 1 to 8 carbon atoms, most preferably hydrogen, methyl, ethyl, methoxy or ethoxy.

The present invention also in the compound of Formula 1,

R ₁ is hydrogen or alkyl of 1 to 8 carbon atoms,

R ₂ and R ₃ are each independently hydrogen, alkyl of 1 to 8 carbon atoms, cycloalkyl of 3 to 8 carbon atoms, alkenyl of 2 to 8 carbon atoms, alkynyl of 2 to 8 carbon atoms, alkoxy of 1 to 8 carbon atoms, or- R ₂₃ R ₂₄ is represented;

In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 8 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 8 carbon atoms; Dialkylamino having 2 to 16 carbon atoms; Any one aryl, heteroaryl or heterocyclic compound selected from the group consisting of phenyl, pyridinyl and triazolyl; Thiourea; Or isothiourea;

The aryl, heteroaryl or heterocyclic compound is hydroxy, amino, cyano, arylalkyl having 7 to 12 carbon atoms, aryl unsubstituted or substituted with alkyl having 1 to 4 carbon atoms and 1 to 1 carbon atoms unsubstituted or substituted with carboxyl. May be substituted with one or more selected from the group consisting of 12 alkyl;

R ₄ to R ₁₁ are each independently hydrogen, alkyl having 1 to 8 carbons or alkoxy having 1 to 8 carbons, and a pharmaceutically acceptable salt or solvate thereof.

The present invention also in the compound of Formula 1,

R ₁ is hydrogen, methyl or ethyl,

R ₂ and R ₃ are each independently hydrogen, alkyl having 1 to 4 carbon atoms, cycloalkyl having 3 to 6 carbon atoms, alkenyl having 2 to 4 carbon atoms, alkynyl having 2 to 4 carbon atoms, alkoxy having 1 to 4 carbon atoms, or- R ₂₃ R ₂₄ is represented;

In the above, R ₂₃ represents a linear or branched alkylene having 1 to 4 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 4 carbon atoms; Dialkylamino having 2 to 8 carbon atoms; Substituted or unsubstituted triazolyl, thiourea or isothiourea;

The substituent of triazolyl is at least one selected from the group consisting of alkyl having 1 to 4 carbon atoms unsubstituted or substituted with benzyl and carboxyl;

R ₄ to R ₁₁ are each independently hydrogen, methyl, ethyl, methoxy or ethoxy, and a pharmaceutically acceptable salt or solvate thereof.

More preferably, the compound of formula 1 of the present invention,

3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione;

3- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione;

3- [1- (2-hydroxyethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -pyrrole-2,5-dione;

3- [1- (2-dimethylaminoethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione;

3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione;

3-[(1-benzyl-1H- [1,2,3] triazol-4-ylmethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl ) -Pyrrole-2,5-dione;

(4- {3- [4- (1H-Indol-3-yl) -2,5-dioxy-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxy-1H-indole -1-ylmethyl}-[1,2,3] triazol-1-yl) acetic acid;

3- [1- (2-dimethylaminoethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione;

3- [1- (2-hydroxyethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione;

2- (2- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} -ethyl) isothiourea;

2- (3- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} propyl) isothiourea;

3- [1- (3-hydroxypropyl) -1H-indol-3-yl] -4- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione;

3- [1- (3-dimethylaminopropyl) -1H-indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione;

2- [3- (3- {4- [1- (3-carbamimidoylsulfanylpropyl) -1H-indol-3-yl] -2,5-dioxo-2,5-dihydro-1H- Pyrrole-3-yl} -5-methoxyindol-1-yl) -propyl] -isothiourea;

3- [1- (4-hydroxybutyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -pyrrole-2,5-dione;

2- (4- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} butyl) isothiourea;

3- [1- (4-dimethylaminobutyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3- yl) pyrrole-2,5-dione;

3,4-bis [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] pyrrole-2,5-dione;

3- [1- (4-hydroxybutyl) -1H-indol-3-yl] -4- [1- (4-hydroxybutyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione;

2- [4- (3- {4- [1- (4-carbamimidolsulfanylbutyl) -1H-indol-3-yl} -2,5-dioxo-2,5-dihydro-1H- Pyrrole-3-yl] -5-methoxyindol-1-yl) -butyl] -isothiourea;

3- [1- (4-Dimethylaminobutyl) -1 H-indol-3-yl] -4- [1-4-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione;

3- [1- (4-Dimethylaminobutyl) -1 H-indol-3-yl] -4- [1-4-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione;

3- [1- (4-Dimethylaminobutyl) -1H-indol-3-yl] -4- [1-3-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione;

3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1-methyl-1H-indol-3-yl) -pyrrole-2,5-dione ;

3- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -4- [1-methyl-1H-indol-3-yl) -pyrrole-2,5-dione ;

2- (3- {5-methoxy-3- [4- (1-methyl-1H-indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrole-3- Il] -indol-1-yl} -propyl) -isothiourea; or

2- (4- {5-methoxy-3- [4- (1-methyl-1H-indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrole-3- Il] -indol-1-yl} -butyl) -isothiourea.

Most preferably, the compound of formula 1 of the present invention

3- [1- (3-hydroxypropyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione;

3- [1- (2-dimethylaminoethyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione;

2- (2-3- [4- ( 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl] -5-methoxyindole-1 -Yl-ethyl) isothiourea;

3- [1- (3-dimethylaminopropyl) - 1H-indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione;

2- [3- (3-4- [1- (3-carbamimidoylsulfanylpropyl) -1H-indol-3-yl] -2,5-dioxo-2,5-dihydro-1H-pyrrole -3-yl-5-methoxyindol-1-yl) -propyl] -isothiourea;

2- (4-3- [4- ( 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl] -5-methoxyindole-1 -Ylbutyl) isothiourea;

3- [1- (4-dimethylaminobutyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione;

3,4-bis [1- (3-dimethylaminopropyl) -5-methoxy- 1H -indol-3-yl] pyrrole-2,5-dione;

3- [1- (3-dimethylaminopropyl) -5-methoxy- 1H -indol-3-yl] -4- [1-methyl- 1H -indol-3-yl) -pyrrole-2,5-dione ;

2- (3-5-methoxy-3- [4- (1-methyl- 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl ] -Indol-1-yl-propyl) -isothiourea; or

2- (4-5-methoxy-3- [4- (1-methyl- 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl ] -Indol-1-yl-butyl) -isothiourea.

The compound of Formula 1 of the present invention may be prepared and used in the form of a pharmaceutically acceptable salt or solvate, wherein the preparation of salts or solvates may be used with inorganic or organic acids or solvents commonly used in the art. Can be.

Examples of inorganic or organic acids that can be used in the preparation of pharmaceutically acceptable salts of the compounds of formula 1 of the present invention include inorganic acids such as hydrochloric acid, bromic acid, sulfuric acid, phosphoric acid or nitric acid; Or organic acids such as citric acid, acetic acid, lactic acid, tartaric acid, maleic acid, gluconic acid, succinic acid, formic acid, trifluoroacetic acid, oxalic acid, fumaric acid, methanesulfonic acid, benzenesulfonic acid, paratoluenesulfonic acid or camphorsulfonic acid. It doesn't happen.

In the preparation of the solvates, pharmaceutically acceptable solvents commonly used in the art may be used without limitation. Examples of the solvent include water or ether.

The present invention also relates to a method for preparing the compound represented by Chemical Formula 1.

That is, the present invention relates to a method for preparing a compound of Formula 1 ', comprising reacting a compound of Formula 2 with a compound of Formula 3.

[화학식 2]

[Formula 2]

[화학식 3]

[Formula 3]

[화학식 1‘]

[Formula 1 ']

Wherein R ₁ and R ₄ to R ₁₁ are the same as defined in Formula 1,

R ₂ ^′ and R ₃ ^′ each independently represent hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl, alkoxy, amino, cyano, carboxyl, aryl or —R ₂₃ ^' R ₂₄ ^' ;

R ₂₃ ′ in the above is a linear or branched alkylene; Straight or branched alkenylene; And linear or branched alkynylene, and R _{24 ′} is hydroxy; Amino; Cyano; Carboxyl; Substituted or unsubstituted aryl, heteroaryl or heterocyclic compounds; Alkylthio; Thiocyanato; Sulfamoyl or protecting group. At this time, examples of the protecting group include alkylsulfonyloxy; Tetraalkylsiloxy; Ethoxy carbonyl; Or acetoxy.

The present invention also relates to a method for producing a compound characterized in that it further comprises the step of reacting the compound of Formula 1 'with a nucleophile. At this time, examples of the nucleophile include, but are not limited to, one or more selected from the group consisting of alkyl amines, dialkyl amines, thioureas and isothioureas.

The present invention also relates to a process for producing a compound of formula 1 'further comprising the step of reacting with an electrophile. At this time, examples of the electrophile include dialkyl sulfate, preferably dimethyl sulfate.

Hereinafter, a method for preparing the compound of Formula 1 will be described in detail.

Specific reaction conditions (eg, solvent, reactant, catalyst, addition amount, reaction temperature, reaction time, etc.) at each step of the method for preparing a compound of the present invention are not particularly limited, and the field of synthesis of organic compounds The person skilled in the art can easily adopt and use suitable reaction conditions in accordance with the desired compound.

For example, in the reaction of the compounds of the formulas (2) and (3), the compound of the formula (3) is used in an amount of 1.2 to 1.5 equivalents relative to the compound of the formula (2), and 12 to 15 hours at a temperature of 0 to 25 ℃ It is preferred, but not limited to, to carry out the reaction.

In addition, in the step of reacting the compound of Formula 1 'with the nucleophile, the nucleophile is used in an amount of 1.2 equivalents to 1.5 equivalents relative to the compound of Formula 1', and the reaction is carried out for 10 to 12 hours at a temperature of 80 to 85 ℃ It is preferred to perform, but not limited to. In addition, in the step of reacting the compound of Formula 1 'with an electrophile, an electrophile is used in an amount of 1.2 equivalents to 1.5 equivalents relative to the compound of Formula 1', for 8 hours to 12 hours at a reaction temperature of 60 to 65 ℃ It is preferable to carry out the reaction, but not limited thereto.

In addition, the solvent used in each step of the production method of the present invention is also not particularly limited, it is possible to use a solvent commonly used in the synthesis of organic compounds. Examples of solvents that can be used are t-butyl alcohol, dimethylacetamide, dichloromethane, chloroform, diethyl ether, dioxane, acetonitrile, benzene, toluene, acetone, dimethylformamide (DMF) or tetrahydrofuran (THF) And acetone, dimethylformamide (DMF) or tetrahydrofuran (THF) are preferred among the above.

The invention also relates to a process for the preparation of the compound of formula 2 used in the above process. That is, the compound of Formula 2 may be prepared by reacting a compound of Formula 4 with oxalyl chloride and sodium methoxide.

[화학식 4]

[Formula 4]

Wherein R _{2 ′} and R ₄ to R ₇ are the same as defined in Chemical Formula 2.

Reaction conditions for the preparation of the compound of Formula 2 may also be appropriately selected and applied depending on the reaction compound and the desired compound. For example, it is preferable to use oxalyl chloride in an amount of 1.2 equivalents to 1.5 equivalents and sodium methoxide in an amount of 3 equivalents to 4 equivalents based on the compound of Formula 4, and for 1 hour to 3 at a temperature of -65 to 25 ° C. It is preferred to carry out the reaction for a time, but not limited thereto.

The invention also relates to a process for the preparation of the compound of formula 3 used in the above process. That is, the compound of Formula 3 may be prepared through the step of reacting the compound of Formula 5 with a base (for example, potassium hydroxide (KOH), etc.).

[화학식 5]

[Formula 5]

Wherein R ₃ ′ and R ₈ to R ₁₁ are the same as defined in Chemical Formula 3.

Reaction conditions for the preparation of the compound of Formula 3 may also be appropriately selected and applied depending on the reaction compound and the desired compound. For example, the base is preferably used in an amount of 7 to 8 equivalents based on the compound of Formula 5, and the reaction is performed at a temperature of 85 to 90 ° C. for 2 to 3 hours, but is not limited thereto. The base used is also not particularly limited, and conventional ones in this field can be used without limitation, but potassium hydroxide (KOH) and the like are preferable.

The present invention also relates to NK cells treated with the compound of formula 1 of the present invention. The compounds of the present invention can significantly increase the expression of NCR that induces natural cytotoxicity on NK cells and suicide ligands that react with suicide receptors on target cells, thereby causing host cells infected with tumor cells or viruses. Can increase the killing ability of NK cells against.

The compounds of the present invention will be described in detail with reference to an example of a mechanism for increasing the killing ability of NK cells as follows. Treatment of NK cells with the compound of formula 1 of the present invention, as shown in Figure 1, significantly increases the expression of NCR1, NCR2 and NCR3 on the surface, the increased NCR ultimately increases the cell killing capacity of NK cells . This function of the compounds according to the invention is a unique phenomenon that is not observed in cytokines or PMA, which are known as NK cell activators. In fact, the test results of the present inventors confirmed that various cytokines or PMAs hardly increase the expression of NCR, and that LPS, an immune cell activator without specificity, does not significantly affect the expression of NCR. NK cells have the function of effectively killing cancer cells and cells infected with various viruses, which function is proportional to the expression of NCR on the surface. Therefore, the compound according to the present invention or NK cells treated by the present invention can kill virus-infected cells much better, and greatly increase NCR expression on the surface of NK cells, thereby enhancing the ability of NK cells to kill cells to target cells. The compound of the present invention can be usefully used as an anticancer agent or an antiviral agent.

The present invention also relates to a method for treating NK cells with a compound according to the present invention, and more particularly, to a method for treating NK cells identified from a host in a culture medium to which the compound of the present invention is added. . Culture conditions, such as a culture solution used at this time is not particularly limited, and those commonly used in this field may be used.

The host includes humans, monkeys, cows, sheep, goats, horses, pigs, rabbits, dogs, cats, rats, mice, or marmots. Examples of the culture medium used include RPMI, MEM. Or DMEM, and the like, but is not particularly limited as long as it is commonly used in the culture of animal cells such as NK cells. In addition, in the culture process of the NK cells, the culture temperature is preferably 30 to 37 ℃, the culture time is preferably 1 hour to 48 hours, but is not limited thereto. The amount of the compound added in the treatment may be appropriately selected depending on the intended use and is not particularly limited, but is preferably in the range of 1.0 to 10.0 μM for NK cells 10 ⁵ to 10 ⁶ cells / ml .

The invention also relates to a pharmaceutical composition comprising a compound of formula 1, a pharmaceutically acceptable salt or solvate thereof, as described above, or NK cells treated with the compound.

As described above, the compound of formula 1 of the present invention increases the expression of NCR and suicide ligand of NK cells, and NK cells with increased expression of NCR significantly increase the killing ability against cancer cells or target cells infected with virus. do. Therefore, the compound of the present invention or NK cells treated by the present invention can be used for medical purposes through various methods. As an example of the method, a method of obtaining a therapeutic effect by activating NK cells in vivo by first administering a pharmaceutical composition comprising the compound of Formula 1 of the present invention directly to a patient by oral administration or injection, etc. Can be. Secondly, the method of using the compound of the present invention for NK cell activation when developing a cell therapy (for example, by treating with the compound of the present invention in the process of culturing NK cells in vitro when developing a cell therapy using NK cell). Activating NK cells and then grafting the activated NK cells to the patient).

The pharmaceutical composition of the present invention can be particularly useful as an anticancer agent or an antiviral agent, and can also be useful for the treatment of infectious diseases caused by intracellular parasitic bacteria such as tuberculosis and leprosy.

Specifically, the pharmaceutical composition of the present invention can be used for various solid cancers such as various diseases associated with tumors, for example, lung cancer, liver cancer, stomach cancer, colon cancer, bladder cancer, prostate gland, breast cancer, ovarian cancer, cervical cancer, thyroid cancer, and melanoma. In addition, it can be usefully used for the treatment of various blood cancers, including leukemia. In addition, NCR-activated NK cells have increased killing ability against virus-infected host cells, and thus infectious diseases caused by viruses or bacteria, such as AIDS, bird flu, flu, and CMV infectious diseases and tuberculosis or leprosy. It can also be usefully used to treat bacterial diseases.

Carriers used in the composition according to the present invention include carriers and vehicles commonly used in the pharmaceutical field, and specifically, ion exchange resins, alumina, aluminum stearate, lecithin, serum proteins (eg, human serum albumin), buffer substances (E.g. various phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids), water, salts or electrolytes (e.g. protamine sulfate, disodium hydrogen phosphate, carbohydrogen phosphate, sodium chloride and zinc salts) , Colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose based substrates, polyethylene glycol, sodium carboxymethylcellulose, polyarylates, waxes, polyethylene glycols or wool, and the like. The compositions of the present invention may also further comprise lubricants, wetting agents, emulsifiers, suspending agents, preservatives and the like in addition to the above components.

The pharmaceutical compositions of the present invention may be prepared in parenteral dosage forms such as oral dosage forms or injections.

Examples of formulations for oral administration include tablets, troches, lozenges, water soluble or oily suspensions, prepared powders or granules, emulsions, hard or soft capsules, syrups or elixirs. For formulation into tablets and capsules, lactose, saccharose, sorbitol, mannitol, starch, amylopectin, binders such as cellulose or gelatin, excipients such as dicalcium phosphate, disintegrants such as corn starch or sweet potato starch, magnesium stearate Lubricating oils, such as calcium stearate, sodium stea fumarate or polyethylene glycol wax. In addition, the capsule formulation may contain a liquid carrier such as fatty oil in addition to the above-mentioned substances.

Formulations for oral administration include sterile aqueous solutions, non-aqueous solvents, suspensions, emulsions, and lyophilized preparations. Non-aqueous solvents and suspending solvents may be used vegetable oils such as propylene glycol, polyethylene glycol or olive oil, injectable esters such as ethyl oleate.

In one embodiment, the composition according to the invention may be prepared in an aqueous solution for parenteral administration. Preferably, a buffer solution such as Hanks' solution, Ringer's solution, or physically buffered saline may be used. Aqueous injection suspensions can be added with a substrate that can increase the viscosity of the suspension, such as sodium carboxymethylcellulose, sorbitol or dextran.

Another preferred embodiment of the compositions of the present invention may be in the form of sterile injectable preparations of aqueous or oily suspensions. Such suspensions may be formulated according to techniques known in the art using suitable dispersing or wetting agents (eg Tween 80) and suspending agents. Sterile injectable preparations may also be sterile injectable solutions or suspensions (eg solutions in 1,3-butanediol) in nontoxic parenterally acceptable diluents or solvents. Vehicles and solvents that may be used include mannitol, water, Ringer's solution, and isotonic sodium chloride solution. In addition, sterile, nonvolatile oils are conventionally employed as a solvent or suspending medium. For this purpose any non-irritating non-volatile oil can be used including synthetic mono or diglycerides.

The effective dose of the pharmaceutical composition according to the present invention is preferably 0.2 to 200 mg / kg, and may be administered once to several times a day. The dosage needs to be changed, and those skilled in the art will be able to appropriately change the dosage in view of the patient's constitution specificity and body weight, the type and depth of the disease, the nature of the formulation, the nature of the drug administration and the duration or interval of administration. Can be.

Hereinafter, the present invention will be described in more detail through examples according to the present invention, but the scope of the present invention is not limited to the examples given below.

< Example  1> 3- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) pyrrole-2,5- Dion (5)  Produce

[1- (3-hydroxypropyl) -5- Methoxy - 1H Indole-3-yl]- Oxoacetic acid methyl  Preparation of Ester (4)

After adding NaH (407 mg, 12.79 mmol) to DMF, 5-methoxy indole (1) (1.00 g, 6.79 mmol) was slowly added at 0 ° C. and maintained for 5 minutes. 3-bromopropanol (0.71 ml, 8.15 mmol) was then added slowly, held at the same temperature for 10 minutes, then warmed to 70 ° C. and heated to reflux for 4 hours. The reaction was neutralized with ammonium chloride (5 ml), then diluted with water (40 ml) and extracted three times with ethyl acetate (100 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 10: 1) to give compound (2) in the form of a colorless oil in 79% yield (1.10 g, 5.36 mmol).

Compound (2) (1.10 g, 5.63 mmol) was added to THF (15 ml) followed by the slow addition of NaH (214 mg, 5.36 mmol) at 0 ° C. TBSCl (tert-butyldimethylsilyl chloride) (969 mg, 6.43 mmol) was slowly added to the mixture, and the reaction was maintained at the same temperature for 5 minutes, followed by 2 hours at room temperature. The reaction was neutralized with ammonium chloride (6 ml), then diluted with water (30 ml) and extracted three times with ethyl acetate (80 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 15: 1) to give the title compound (3) in the form of a colorless oil in 93% yield (1.60 g, 5.00 mmol).

Compound (3) (520 mg, 1.63 mmol) was added to dichloromethane (20 ml), and slowly added oxalyl chloride (0.24 ml, 1.96 mmol) at 0 ° C. for 1 hour at the same temperature. Reacted. Then, the temperature of the reaction vessel was lowered to -65 ° C, sodium methoxide (to which 100 mg of sodium was added to 2.5 ml of methanol) was slowly added and reacted for 1 hour while keeping the temperature the same. Subsequently, it was reacted again at room temperature for 1 hour. The reaction was extracted three times with dichloromethane (80 ml) and then dichloromethane was removed. The residue was purified by column chromatography (hexane: ethyl acetate = 1: 1) to give the title compound (4) in the form of a green oil in 84% yield (400 mg, 1.37 mmol).

3- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) pyrrole-2,5- Dion (5)  Produce

A mixture of compound (4) (180 mg, 0.62 mmol) and indol-3-ylacetamide (200 mg, 1.15 mmol) was dissolved in THF (12 ml) and then t-butoxy potassium at 0 ° C. (potassium tert- butoxide) (3 ml) was added slowly. The mixture was kept at the same temperature for 10 minutes and reacted at room temperature for 12 hours. 2M hydrochloric acid (5 ml) was added, followed by further reaction for 1 hour. The reaction was then diluted with water (25 ml) and extracted three times with ethyl acetate (50 ml). The residue was purified by column chromatography (hexane: ethyl acetate = 1: 2) to give the title compound (5) in the form of a red solid in 62% yield (160 mg, 0.85 mmol).

MS (ESI +); 438 [M + Na] ⁺

MP; 243 ~ 245 ℃

¹ H NMR (400 MHz); DMSO- d ₆ δ = 11.60 (1H, s), 10.86 (1H, s), 7.83 (1H, s), 7.65 (1H, s), 7.36 (1H, d, J = 8.0 Hz), 7.30 (1H, d, J = 8.8 Hz), 6.97 (1H, t, J = 7.2 Hz), 6.88 (1H, d, J = 4.0 Hz), 6.64 (1H, t, J = 7.6 Hz), 6.57-6.55 (1H, m), 6.09 (1H, s), 4.25 (2H, t, J = 6.8 Hz), 2.98 (3H, s), 1.90-1.83 (2H, m)

IR: 3490, 2911, 2850, 1651, 1499, 1440, 1411, 1258, 1090 cm ^-1

< Example  2> 3- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5- Dion (7)  Produce

3- ( 1H -Indol-3-yl) -4-5- Methoxy -1- [3- ( Methanesulfonyloxy )profile]- 1H Indole-3- Ilpyrrole -2,5- Dion (6)  Produce

Compound (5) prepared in Example 1 (70 mg, 0.17 mmol) was dissolved in DMF (4 ml), then triethylamine (0.07 ml, 0.51 mmol) was added slowly at 0 ° C. After 3 minutes, methanesulfonyl chloride (0.04 ml, 0.34 mmol) was added and reacted at the same temperature for 10 minutes, followed by reaction at room temperature for 6 hours. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml). The residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the title compound (6) in the form of a red solid in 94% yield (80 mg, 0.16 mmol).

3- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione ( 7)  Produce

Compound (6) (80 mg, 0.168 mmol) and 2M of dimethylamine (2.0 ml) were dissolved in DMF (4 ml) and then heated to reflux at 80 ° C. for 13 hours. The reaction mixture was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. Then recrystallization method using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) gave the desired compound (7) in red solid form in 88% yield (65 mg, 0.147 mmol).

MS (ESI +); 433 [M + H] ⁺

MP; 208 ~ 210 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 11.61 (1H, s), 10.84 (1H, s), 7.80 (1H, s), 7.67 (1H, s), 7.35 (1H, d, J = 8.0 Hz), 7.30 (1H , d, J = 8.8 Hz), 6.97 (1H, t, J = 7.2 Hz), 6.84 (1H, d, J = 4.0 Hz), 6.61 (1H, t, J = 7.6 Hz), 6.57-6.54 (1H , m), 6.11 (1H, s), 4.21 (2H, t, J = 6.4 Hz), 2.30 (3H, s), 2.09 (6H, s), 1.86-1.81 (2H, m)

IR: 3301, 1753, 1708, 1663, 1622, 1532, 1487, 1454, 1434, 1393, 1336, 1091 cm ^-1

< Example  3> 3- [1- (2- Hydroxyethyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) -pyrrole-2,5- Dion (11)  synthesis

 A mixture of compound (10) (450 mg, 1.62 mmol) and indol-3-ylacetamide (380 mg, 2.18 mmol) was dissolved in THF (15 ml) and t-butoxy potassium (4.8 ml) at 0 ° C. Was added slowly. The reaction was carried out at the same temperature for 10 minutes, and the temperature was further raised to room temperature to react for 12 hours. Then 2M hydrochloric acid (5 ml) was added and reacted for 1 hour, after which the reaction was terminated. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 2) to give the desired compound (11) in the form of a red solid in 58% yield (380 mg, 0.95 mmol).

MS (ESI +); 424 [M + Na] ⁺

MP; 241 ~ 243 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 11.58 (1H, s), 10.86 (1H, s), 7.90 (1H, s), 7.60 (1H, s), 7.37 (1H, d, J = 8.0 Hz), 7.30 (1H, d, J = 8.8 Hz), 7.02-6.97 (2H, m), 6.70 (1H, t, J = 7.6 Hz), 6.57-6.54 (1H, m), 6.06 (1H, s), 4.23 (2H, t, J = 6.8 Hz), 3.71-3.70 (2H, m), 2.94 (3H, s)

IR; 3497, 2924, 2856, 1655, 1499, 1440, 1405, 1258, 1095 cm ^-1

< Example  4> 3- [1- (2- Dimethylaminoethyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) pyrrole-2,5- Dion (13)  synthesis

 Compound 12 (50 mg, 0.104 mmol) and 2M dimethyl amine (2.0 ml) were dissolved in DMF (3 ml) and then heated to reflux at 80 ° C. for 10 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml). Subsequently, ethyl acetate was removed and washed three times with hexane, followed by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to give 94% of the title compound (13) as a red solid. Obtained (42 mg, 0.098 mmol).

MS (ESI +); 429 [M + H] ⁺

MP; 230 ~ 232 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 11.60 (1H, s), 10.85 (1H, s), 7.93 (1H, s), 7.65 (1H, s), 7.35 (1H, d, J = 8.0 Hz), 7.30 (1H , d, J = 8.8 Hz), 6.97 (1H, t, J = 7.2 Hz), 6.87 (1H, d, J = 4.0 Hz), 6.20 (1H, t, J = 7.6 Hz), 6.57-6.54 (1H , m), 6.11 (1H, s), 4.27 (2H, t, J = 6.4 Hz), 3.02 (3H, s), 2.57 (2H, t, J = 6.4 Hz), 2.21 (6H, s)

IR; 3477, 1757, 1716, 1663, 1438, 1409, 1389, 1336, 1258, 1091, 1054 cm ^-1

< Example  5> 3- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) -1- Methylpyrrole Synthesis of -2,5-dione (9)

Compound (5) (50 mg, 0.120 mmol), dimethyl sulfate (0.013 ml) and potassium carbonate (32 mg, 0.232 mmol) prepared in Example 1 were dissolved in acetone (3 ml) After heating, the mixture was heated to reflux at 60 ° C. for 8 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (20 ml), then ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the title as a red solid. Compound (9) was obtained in 95% yield (49 mg, 0.114 mmol).

MS (ESI +); 452 [M + Na] ⁺

MP; 115 ~ 117 ℃

¹ H NMR (400 MHz); CD ₃ CN: δ = 9.75 (1H, s), 7.84 (1H, s), 7.71 (1H, s), 7.40 (1H, d, J = 8.0 Hz), 7.25 (1H, d, J = 8.8 Hz) , 7.01 (1H, t, J = 7.2 Hz), 6.91 (1H, d, J = 4.0 Hz), 6.66 (1H, t, J = 7.6 Hz), 6.62-6.59 (1H, m), 6.20 (1H, s), 4.21 (2H, t, J = 6.4 Hz), 3.50-3.46 (2H, m), 3.08 (6H, s), 2.01-1.98 (2H, m)

IR; 3452, 2929, 2864, 2803, 1654, 1495, 1434, 1413, 1389, 1254, 1127, 1090, 1054 cm ^-1

< Example  6> 3-[(1-benzyl- 1H -[1,2,3] Triazole -4- days) methyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) -pyrrole-2,5-di On (2 Synthesis of 0)

1- (1-benzyl- 1H -[1,2,3] Triazole -4- methyl ) -5- Methoxy - 1H - Of indole (18)  synthesis

NaH (160 mg, 4.07 mmol) was added to DMF (16 ml), and 5-methoxy-indole (1) (500 mg, 3.40 mmol) was slowly added at 0 ° C, and reacted at the same temperature for 5 minutes. Then propargyl bromide (0.35 ml, 4.08 mmol) was added slowly, tetrabutylammonium iodide (10 mg) was added immediately and reacted for 10 minutes, followed by 8 hours at 70 ° C. Heated to reflux. The reaction was neutralized with ammonium chloride (5 ml), then diluted with water (40 ml) and extracted three times with ethyl acetate (100 ml). The residue was purified by column chromatography (hexane: ethyl acetate = 10: 1) to give the title compound (17) as a colorless oil in a yield of 79% (500 mg, 2.70 mmol).

The compound (17) (230 mg, 1.24 mmol) and benzyl azide (250 mg, 1.87 mmol) were dissolved in THF (10 ml) and copper sulfoxide (80 mg, 0.32 mmol). ) And sodium ascorbate (370 mg, 1.87 mmol) were added followed by a slow addition of water (3 ml). The reaction was maintained at room temperature for 8 hours, the reaction was neutralized with ammonium chloride (5 ml), then diluted with water (30 ml) and extracted three times with ethyl acetate (100 ml). The residue was purified by column chromatography (hexane: methylene chromide = 1: 1) to give the title compound (18) in the form of a colorless oil in 96% yield (380 mg, 1.19 mmol).

[1- (1-benzyl- 1H -[1,2,3] Triazole -4- Methyl ) -5- Methoxy - 1H Indole-3-yl]- Oxoacetic acid  Synthesis of Methyl Ester (19)

Compound (18) (700 mg, 2.20 mmol) prepared above was added to dichloromethane (20 ml), and oxalyl chloride (0.23 ml, 2.64 mmol) was slowly added at 0 ° C, and reacted for 1 hour. Then, the temperature of the reaction vessel was lowered to -65 ° C, and sodium methoxide (to which 100 mg of sodium was added to 2.5 ml of methanol) was slowly added while maintaining the temperature, and reacted for 1 hour. Then it was reacted again at room temperature for 1 hour and the reaction was terminated. The reaction was extracted three times with methylene chloride (80 ml). Dichloromethane was removed and the residue was purified by column chromatography (hexane: methylene chromide = 1: 1) to give the title compound (19) in the form of a yellow solid in 95% yield (850 mg, 2.10 mmol).

3-[(1-benzyl- 1H -[1,2,3] Triazole -4- days) methyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) -pyrrole-2,5- Dion (20)  synthesis

A mixture of compound (19) (500 mg, 1.24 mmol) and indol-3-ylacetamide (250 mg, 1.43 mmol) prepared above was dissolved in THF (15 ml) and t-butoxy potassium (6.0) at 0 ° C. ml) was added slowly. The mixture was kept at the same temperature for 10 minutes and reacted again at room temperature for 13 hours. Then 2M hydrochloric acid (8 ml) was added and then further reacted for 1 hour. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 2) to give the target compound (20) in the form of a red solid. Obtained in 46% yield (300 mg, 0.56 mmol).

MS (ESI +); 551 [M + Na] ⁺

MP; 280 ~ 282 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 11.60 (1H, s), 10.87 (1H, s), 8.05 (1H, s), 7.95 (1H, s), 7.62 (1H, s), 7.40-7.25 (7H, m) , 6.95 (1H, t, J = 7.6 Hz), 6.88 (1H, d, J = 8.4 Hz), 6.58-6.54 (2H, m), 6.05 (1H, s), 5.56 (2H, s), 5.50 ( 2H, s), 2.94 (3H, s)

IR; 3532, 1720, 1650, 1455, 1123, 1030, 1015 cm ^-1

< Example  7> (4-3- [4- ( 1H -Indol-3-yl) -2,5- Deoxy -2,5- Dehydro - 1H -Pyrrole-3-yl] -5- Methoxy - 1H -Indole-1- Methyl Synthesis of-[1,2,3] triazol-1-yl) acetic acid (23)

[4- (5- Methoxyindole -One- Methyl )-[1,2,3] Triazole Synthesis of -1-yl] -acetic acid ethyl ester (21)

A mixture of the above compound (17) (230 mg, 1.24 mmol) and azide acetic acid ethyl ester (200 mg, 1.55 mmol) was dissolved in THF (10 ml), and sulfoxide copper (80 mg, 0.32 mmol) and ascor Sodium vinate (370 mg, 1.87 mmol) was added and the reaction was carried out with slowly adding water (3 ml). The reaction was held for 8 hours at room temperature and the reaction was neutralized with ammonium chloride (5 ml). The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (100 ml), and then purified by column chromatography (hexane: methylene fluoride = 1: 1) to give the title compound (21) as a colorless oil. Obtained in% yield (380 mg, 1.21 mmol).

[1- (1- Ethoxycarbonylmethyl - 1H -[1,2,3] Triazole -4- Methyl ) -5- Methoxy - 1H Indole-3-yl]- Oxoacetic acid  Methyl ester ( 22)  synthesis

Compound (21) prepared above (1.0 g, 3.18 mmol) was added to dichloromethane (20 ml), and oxalyl chloride (0.27 ml, 3.18 mmol) was slowly added at 0 ° C, followed by reaction at the same temperature for 1 hour. I was. The temperature of the reaction vessel was then lowered to −65 ° C., followed by the slow addition of sodium methoxide (adding 100 mg of sodium to 2.5 ml of methanol) and reacting for 1 hour. After reacting again for 1 hour at room temperature, the reaction was terminated and the reaction was extracted three times with methylene chloride (80 ml). The residue was purified by column chromatography (hexane: methylene chromide = 1: 1) to give the title compound (22) in the form of a yellow solid in 86% yield (1.10 g, 2.75 mmol).

(4-3- [4- ( 1H -Indol-3-yl) -2,5- Deoxy -2,5- Dehydro - 1H -Pyrrole-3-yl] -5- Methoxy - 1H -Indole-1- Methyl Synthesis of-[1,2,3] triazol-1-yl) acetic acid (23)

A mixture of compound (22) (500 mg, 1.24 mmol) and indol-3-ylacetamide (250 mg, 1.43 mmol) prepared above was dissolved in THF (15 ml) and t-butoxy potassium (0 6.0 ml) was added slowly. After holding for 10 minutes at the same temperature, it was reacted for 13 hours at room temperature. After the passage of time, 2M hydrochloric acid (8 ml) was added, followed by further reaction for 1 hour, and the reaction was terminated. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml). The residue was purified by column chromatography (methylene chloride: ethyl acetate = 1: 2) to give the title compound (23) in the form of a red solid in 41% yield (230 mg, 0.503 mmol).

MS (ESI +); 519 [M + Na] ⁺

MP; 180 ~ 182 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 11.60 (1H, s), 10.88 (1H, s), 8.00 (1H, s), 7.61 (1H, s), 7.42 (1H, d, J = 9.2 Hz), 7.35 (1H , d, J = 8.0 Hz), 6.98 (1H, t, J = 7.6 Hz), 6.93 (1H, d, J = 8.0 Hz), 6.65 (1H, t, J = 7.6 Hz), 6.57-6.55 (1H , m), 6.05 (1H, s), 5.54 (2H, s), 5.24 (2H, s), 2.94 (3H, s)

IR; 3530, 1716, 1646, 1450, 1189, 1119, 1091, 1049, 1017 cm ^-1

< Example  8> 3- [1- (2- Dimethylaminoethyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H Indole-3-yl) -1-meth Tilpi Roll-2,5- Dion (16)  synthesis

Compound (13) (30 mg, 0.070 mmol), dimethyl sulfate (0.008 ml) and potassium carbonate (20 mg, 0.145 mmol) were dissolved in acetone (3 ml) and then heated to reflux at 60 ° C. for 8 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (20 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the target compound (16) in the form of a red solid. Obtained 97% yield (30 mg, 0.068 mmol).

MS (ESI +); 443 [M + H] ⁺

MP; 141 ~ 143 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.72 (1H, s), 7.52 (1H, s), 7.34 (1H, d, J = 8.0 Hz), 7.25 (1H, d, J = 8.8 Hz), 6.97 (1H, t, J = 7.2 Hz), 6.90 (1H, d, J = 4.0 Hz), 6.66 (2H, m), 6.21 (1H, s), 4.58 (2H, t, J = 6.4 Hz), 3.59 (2H, t, J = 6.4 Hz), 3.05 (12H, s)

IR; 1634, 1462, 1140, 1123, 1091 cm ^-1

< Example  9> 3- [1- (2- Hydroxyethyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) -1- Methylpyrrole -2,5- Dion (15)  synthesis

Compound (11) (20 mg, 0.050 mmol), dimethyl sulfate (0.005 ml) and potassium carbonate (14 mg, 0.101 mmol) were dissolved in acetone (3 ml) and then heated to reflux at 60 ° C. for 8 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (20 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the title compound as a red solid (15). Was obtained in 96% yield (20 mg, 0.048 mmol).

MS (ESI +); 438 [M + Na] ⁺

MP; 117 ~ 119 ℃

¹ H NMR (400 MHz); CD ₃ CN: δ = 9.75 (1H, s), 7.90 (1H, s), 7.67 (1H, s), 7.41 (1H, d, J = 8.0 Hz), 7.25 (1H, d, J = 8.8 Hz) , 7.04 (1H, t, J = 7.2 Hz), 6.91 (1H, d, J = 4.0 Hz), 6.66 (1H, t, J = 7.6 Hz), 6.62-6.59 (1H, m), 6.20 (1H, s), 4.26 (2H, t, J = 6.4 Hz), 3.85-3.81 (2H, m), 3.07 (6H, s)

IR; 1687, 1622, 1540, 1479, 1442, 1152, 1123, 1095 cm ^-1

< Example  10> 2- (2-3- [4- ( 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl] -5- Methoxyindole -1-yl-ethyl) Isothiourea Synthesis of 14

Methanesulfonic acid  2-3- [4- ( 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3day] -5- Methoxy -Indole-1- Ethyl  ester( 12)  synthesis

Compound (11) (170 mg, 0.42 mmol) was dissolved in DMF (4 ml), and triethyl amine (0.07 ml, 0.51 mmol) was added slowly at 0 ° C., and after 3 minutes, methanesulfonyl chloride (0.04 ml, 0.34) mmol) was added. The reaction was carried out at the same temperature for 10 minutes and again at room temperature for 6 hours. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the target compound (12) in the form of a red solid. Obtained 94% yield (180 mg, 0.37 mmol).

2- (2-3- [4- ( 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl] -5- Methoxyindole -1-yl-ethyl) Isothioure Synthesis of Ah 14

Compound (12) (120 mg, 0.25 mmol) and thiourea (38 mg) prepared above were dissolved in DMF (3 ml) and then heated to reflux at 80 ° C. for 10 hours. The reaction was diluted with water (15 ml) and extracted twice with ethyl acetate (20 ml) to remove material present in the organic layer. Then water was removed and washed three times with ethyl acetate, followed by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to give 96% of the desired compound (14) as a red solid. Yield of (110 mg, 0.24 mmol) was obtained.

MS (ESI +); 460 [M + H] ⁺

MP; 180 ~ 182 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.80 (1H, s), 7.67 (1H, s), 7.34 (1H, d, J = 8.0 Hz), 7.24 (1H, d, J = 8.8 Hz), 7.01 (1H, t, J = 7.2 Hz), 6.93 (1H, d, J = 4.0 Hz), 6.67-6.60 (2H, m), 6.17 (1H, s), 4.52 (2H, t, J = 6.4 Hz), 3.60 (2H, t, J = 6.8 Hz), 3.01 (3H, s)

IR; 1703, 1659, 1528, 1483, 1193, 1140, 1128, 1091, 1061 cm ^-1

Example 11 2- (3-3- [4- ( 1H -Indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -Pyrrole-3-yl] -5-methoxyindol-1-ylpropyl) Isothiourea Synthesis of 8

Compound (6) (68 mg, 0.137 mmol) and thiourea (10 mg) were dissolved in DMF (3 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was diluted with water (15 ml) and extracted twice with ethyl acetate (20 ml), then the material present in the organic layer was removed. Then water was removed and washed three times with ethyl acetate, followed by 92% of the desired compound (8) in the form of a red solid by recrystallization using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml). Obtained in (60 mg, 0.126 mmol).

MS (ESI +); 474 [M + H] ⁺

MP; 230 ~ 232 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.74 (1H, s), 7.70 (1H, s), 7.35 (1H, d, J = 8.0 Hz), 7.24 (1H, d, J = 8.8 Hz), 7.01 (1H, t, J = 7.2 Hz), 6.88 (1H, d, J = 4.0 Hz), 6.64-6.60 (2H, m), 6.28 (1H, s), 4.32 (2H, t, J = 6.4 Hz), 3.08 (3H, s), 3.02 (2H, t, J = 6.8 Hz), 2.22-2.19 (2H, m)

IR: 1654, 1622, 1479, 1148, 1095, 1119 cm ^-1

Example 12 3- [1- (3-hydroxypropyl)- 1H -Indol-3-yl] -4- [1- (3-hydroxypropyl) -5-methoxy- 1H -Indol-3-yl] -pyrrole-2,5- Dion (31)  synthesis

Examples The compound synthesized in 1 (4) (500 mg, 1.44 mmol) and 2- [1- (3-hydroxypropyl) - 1H-indol-3-yl] -acetamide (800 mg, 3.45 mmol) of The mixture was taken up in THF (20 ml) and t-butoxy potassium (10.0 ml) was added slowly at 0 ° C. Subsequently, the mixture was maintained at the same temperature for 10 minutes, and then reacted at room temperature for 12 hours. 2M hydrochloric acid (5 ml) was added, followed by reaction for 1 hour, and the reaction was terminated. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (31) in the form of a red solid. Obtained 44% yield (300 mg, 0.64 mmol).

MS (ESI +); 496 [M + Na] ⁺

MP; 190 ~ 192 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.76 (1H, s), 7.56 (1H, s), 7.35 (1H, d, J = 8.0 Hz), 7.21 (1H, d, J = 8.8 Hz), 7.03 (1H, t, J = 7.2 Hz), 6.97 (1H, d, J = 8.0 Hz), 6.68 (1H, t, J = 7.2 Hz), 6.11 (1H, s), 4.25-4.19 (4H, m), 3.54-3.47 ( 4H, m), 2.00-1.89 (4H, m)

IR; 3351, 1708, 1659, 1618, 1532, 1471, 1140, 1123, 1095 cm ^-1

Example 13 3- [1- (3-dimethylaminopropyl)- 1H -Indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy- 1H Synthesis of -indol-3-yl] -pyrrole-2,5-dione (33)

Methanesulfonic acid  3- (3-4- [1- ( Methanesulfonyloxy -profile)- 1H -Idol-3-yl] -2,5- Dioxo -2,5- Dehydro - 1H Synthesis of -pyrrole-3-yl-5-methoxy-indol-1-yl) propyl ester (32)

Compound (31) (250 mg, 0.53 mmol) prepared in Example 12 was dissolved in DMF (6.0 ml), and triethyl amine (0.30 ml, 2.12 mmol) was slowly added at 0 ° C., and methane chloride was added after 3 minutes. Ponyyl (0.16 ml, 2.12 mmol) was added. The reaction was kept at the same temperature for 10 minutes and further reacted at room temperature for 6 hours. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (32) in the form of a red solid in 81% yield (270 mg, 0.43 mmol).

3- [1- (3-dimethylaminopropyl)- 1H -Indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy- 1H -Indol-3-yl] -pyrrole-2,5-di On (3 3) Synthesis

Compound (32) prepared above (150 mg, 0.238 mmol) and 2M dimethyl amine (1.5 ml) were dissolved in DMF (4.0 ml) and then heated to reflux at 80 ° C. for 13 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. And recrystallization using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) gave the desired compound (33) in the form of a red solid in 88% yield (110 mg, 0.208 mmol). .

MS (ESI +); 550 [M + Na] ⁺

MP; 182 ~ 184 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 10.87 (1H, s), 7.82 (1H, s), 7.63 (1H, s), 7.44 (1H, d, J = 8.0 Hz), 7.31 (1H, d, J = 8.8 Hz ), 7.04 (1H, t, J = 7.6 Hz), 6.93 (1H, d, J = 8.0 Hz), 6.68 (1H, t, J = 7.6 Hz), 6.59-6.56 (1H, m), 6.06 (1H) , s), 4.21-4.17 (4H, m), 2.94 (3H, s), 2.09-205 (16H, m), 1.85-1.74 (4H, m)

IR; 3399, 1659, 1054, 1033, 1013 cm ^-1

< Example  14> 2- [3- (3- {4- [1- (3- Carbamimidoylsulfanylpropyl ) -1H-indol-3-yl] -2,5- Dioxo -2,5- Dehydro -1H-pyrrole-3-yl} -5- Methoxyindole -1-yl) -propyl]- Of isothiourea (34)  synthesis

Compound (32) (100 mg, 0.159 mmol) and thiourea (48 mg) prepared in Example 13 were dissolved in DMF (3.5 ml) and then heated to reflux at 80 ° C. for 13 hours. The reaction was diluted with water (15 ml) and extracted twice with ethyl acetate (20 ml), then the material in the organic layer was removed. Then water was removed and washed three times with ethyl acetate, followed by 91% of the desired compound (34) in the form of a red solid by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml). Obtained in (85 mg, 0.144 mmol).

MS (ESI +); 590 [M + H] ⁺

MP; 120 to 122 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.82 (1H, s), 7.57 (1H, s), 7.45 (1H, d, J = 8.4 Hz), 7.31 (1H, d, J = 8.8 Hz), 7.12-7.08 (2H, m), 6.75 (1H, t, J = 7.2 Hz), 6.68-6.65 (1H, m), 6.12 (1H, s), 4.38-4.31 (4H, m), 3.10-2.98 (7H, s), 2.26 -2.23 (2H, m), 2.16-2.13 (2H, m)

IR; 3211, 1761, 1712, 1650, 1532, 1487, 1442, 1397, 1344, 1189, 1054 cm ^-1

Example 15 3- [1- (4-hydroxybutyl) -5-methoxy- 1H -Indol-3-yl] -4- ( 1H Synthesis of -indol-3-yl) -pyrrole-2,5-dione (27)

Synthesis of acetic acid 4- (5-methoxyindol-1-yl) -butyl ester (24)

NaH (600 mg, 15.29 mmol) was added to DMF (30 ml), and 5-methoxy indole (1.5 g, 10.19 mmol) was slowly added at 0 ° C., and reacted for 5 minutes while maintaining the temperature. 4-Bromo butyl acetate (1.90 ml, 13.25 mmol) was slowly added to the reaction, followed by reaction for 10 minutes with tetrabutylammonium iodide (10 mg), followed by further reaction at room temperature for 2 hours. I was. The reaction was neutralized with ammonium chloride (6.0 ml), then diluted with water (40 ml) and extracted three times with ethyl acetate (100 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 10: 1) to give the title compound (24) in the form of a green oil in 93% yield (2.5 g, 9.59 mmol).

[1- (4- Acetoxybutyl ) -5- Methoxy - 1H Indole-3-yl]- Oxoacetic acid methyl  Synthesis of Ester (25)

Compound (24) prepared above (2.50 g, 9.57 mmol) was added to dichloromethane (20 ml), and oxalyl chloride (1.0 ml, 11.48 mmol) was slowly added at 0 ° C, and then the temperature was maintained at 1 The reaction was carried out for a time. Subsequently, after the temperature of the reaction vessel was lowered to -65 ° C, sodium methoxide (which added 150 mg of sodium to 4.0 ml of methanol) was slowly added to react for 1 hour, followed by an additional 1 hour at room temperature. The reaction was extracted three times with methylene chloride (80 ml) and dichloromethane was removed. The residue was purified by column chromatography (hexane: ethyl acetate = 1: 1) to give the title compound (25) in the form of a yellow oil in a yield of 54% (1.80 g, 5.18 mmol). In the reaction, Compound 26 in the form of a yellow oil was further formed in 41% yield (1.20 g, 3.93 mmol).

3- [1- (4-hydroxybutyl) -5-methoxy- 1H -Indol-3-yl] -4- ( 1H Synthesis of -indol-3-yl) -pyrrole-2,5-dione (27)

A mixture of compound (25) (500 mg, 1.44 mmol) and indol-3-ylacetamide (300 mg, 1.72 mmol) prepared above was dissolved in THF (15 ml) and t-butoxy potassium (4.3) at 0 ° C. ml) was added slowly. After maintaining at the same temperature for 10 minutes, the reaction was carried out at room temperature for 12 hours. Then 2M hydrochloric acid (5 ml) was added, followed by further reaction for 1 hour. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the target compound (27) in the form of a red solid. Obtained 74% yield (460 mg, 1.07 mmol).

MS (ESI +); 452 [M + Na] ⁺

MP; 220 ~ 222 ℃

¹ H NMR (400 MHz); DMSO- d ₆ δ = 11.61 (1H, s), 10.87 (1H, s), 7.85 (1H, s), 7.68 (1H, s), 7.37 (1H, d, J = 4.0 Hz), 7.30 (1H, d, J = 8.8 Hz), 6.98 (1H, t, J = 7.6 Hz), 6.90 (1H, d, J = 8.0 Hz), 6.65 (1H, t, 7.6 Hz), 6.12-6.16 (1H, m), 4.20 ( 2H, t, J = 6.4 Hz), 3.40 (2H, s), 1.77 (2H, t, J = 6.4 Hz), 1.38-1.39 (2H, m)

IR; 3412, 1712, 1659, 1532, 1487, 1340, 1205, 1054, 1021, 1013 cm ^-1

Example 16 2- (4-3- [4- ( 1H -Indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H Synthesis of -pyrrole-3-yl] -5-methoxyindol-1-ylbutyl) isothiourea (30)

Methanesulfonic acid 4-3- [4- ( 1H -Indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H Synthesis of -pyrrole-3-yl] -5-methoxy-indol-1-yl-butyl ester (28)

Compound (27) (800 mg, 1.86 mmol) prepared in Example 15 was dissolved in DMF (10 ml), and triethyl amine (0.50 ml, 3.72 mmol) was added slowly at 0 ° C., and after 3 minutes, methane chloride Sulfonyl (0.29 ml, 3.72 mmol) was added. The reaction was carried out at the same temperature for 10 minutes, and further reacted at room temperature for 6 hours. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml). The ethyl acetate was then removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (28) in the form of a red solid in 95% yield (900 mg, 1.77 mmol).

2- (4-3- [4- ( 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl] -5- Methoxyindole -1 day part Teal) Isothiourea Synthesis of 30

`Compound (28) (500 mg, 1.03 mmol) and thiourea (140 mg) prepared above were dissolved in DMF (6.0 ml) and then heated to reflux at 80 ° C for 10 hours. The reaction was diluted with water (15 ml), extracted twice with ethyl acetate (20 ml) and the material in the organic layer was removed. Then water was removed and washed three times with ethyl acetate, followed by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to 92% of the desired compound (30) in the form of a red solid. Yield (460 mg, 0.94 mmol) was obtained.

MS (ESI +); 488 [M + H] ⁺

MP; 182 ~ 184 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.78 (1H, s), 7.68 (1H, s), 7.35 (1H, d, J = 8.0 Hz), 7.21 (1H, d, J = 8.8 Hz), 6.99 (1H, t, J = 7.6 Hz), 6.89 (1H, d, J = 4.0 Hz), 6.64-6.58 (2H, m), 6.20 (1H, s), 4.20 (2H, t, J = 6.8 Hz), 3.10 (2H, t, J = 7.2 Hz), 3.04 (3H, s), 1.95-1.93 (2H, m), 1.67-1.63 (2H, m)

IR; 3387, 3285, 3179, 1699, 1650, 1605, 1528, 1475, 1417, 1336, 1193, 1091, 1058 cm ^-1

< Example  17> 3- [1- (4- Dimethylaminobutyl ) -5- Methoxy - 1H -Indol-3-yl] -4- ( 1H -Indol-3-yl) pyrrole-2,5- Dion (29)  synthesis

Compound (28) (400 mg, 0.79 mmol) and 2M dimethyl amine (0.8 ml) prepared in Example 16 were dissolved in DMF (6 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. Then recrystallization method using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) gave the desired compound (29) in the form of a red solid in 94% yield (340 mg, 0.74 mmol).

MS (ESI +); 457 [M + H] ⁺

MP; 100 to 102 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.78 (1H, s), 7.63 (1H, s), 7.33 (1H, d, J = 4.4 Hz), 7.18 (1H, d, J = 8.8 Hz), 6.99 (1H, t, J = 7.6 Hz), 6.94 (1H, d, J = 4.0 Hz), 6.63 (1H, t, J = 7.6 Hz), 6.59-6.60 (1H, m), 6.18 (1H, s), 4.15-4.12 ( 2H, m), 3.01 (3H, s), 2.28 (2H, t, J = 7.2 Hz), 1.80-1.76 (2H, m), 1.47-1.44 (2H, m)

IR; 3305, 2946, 1761, 1699, 1659, 1614, 1528, 1479, 1446, 1332, 1134, 1123, 1086 cm ^-1

Example 18 3,4-bis [1- (3-dimethylaminopropyl) -5-methoxy- 1H Synthesis of -indol-3-yl] pyrrole-2,5-dione (40)

 2- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indole-3-day] Acetamide  Synthesis of 37

After dissolving NaH (130 mg, 3.22 mmol) in DMF (20 ml), 5-methoxyindol-3-ylacetonitrile (500 mg, 2.685 mmol) (35) was added slowly at 0 ° C. for 5 minutes. Reacted. Then 3-bromopropanol (0.35 ml, 4.03 mmol) was added slowly, reacted for 10 minutes at the same temperature, and then heated to reflux at 70 ° C. for 4 hours. The reaction was neutralized with ammonium chloride (5 ml), diluted with water (40 ml) and then extracted three times with ethyl acetate (100 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 1) to afford the desired compound (36) in the form of a colorless oil in 76% yield (500 mg, 2.046 mmol).

A mixture of the prepared compound (36) (500 mg, 2.046 mmol) and potassium hydroxide (800 mg, 14.26 mmol) was dissolved in tert-butyl alcohol (7.0 ml) and heated to reflux at 85 ° C. for 2 hours. 2M hydrochloric acid (6.0 ml) was then added, diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (37) in oil form in 84% yield (450 mg, 1.716 mmol).

3,4-bis [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] pyrrole-2,5- Dion (38)  synthesis

A mixture of compound (37) (500 mg, 1.91 mmol) prepared and compound (4) (400 mg, 1.37 mmol) prepared in Example 1 was dissolved in THF (20 ml) and t-butoxy potassium at 0 ° C. (4.1 ml) was added slowly. It was kept at the same temperature for 10 minutes and reacted at room temperature for 12 hours. Then 2M hydrochloric acid (5 ml) was added and reacted for 1 hour. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (38) in the form of a red solid. Obtained 42% yield (290 mg, 0.576 mmol).

Methanesulfonic acid  3- (3-4- [1- (3- Methanesulfonyloxy -Propyl) -5- Methoxy - 1H -Indol-3-yl] -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl-5- Methoxy Synthesis of -indol-1-yl) -propyl Ester (39)

Dissolved Compound (38) (160 mg, 0.318 mmol) in DMF (6.0 ml) was added triethyl amine (0.17 ml, 1.272 mmol) slowly at 0 ° C., and after 3 minutes methanesulfonyl chloride (0.10 ml, 1.272 mmol) was added. Subsequently, the reaction was maintained at the same temperature for 10 minutes, and further reacted at room temperature for 6 hours. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (50 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (39) in the form of a red solid in 85% yield (200 mg, 0.270 mmol).

3,4-bis [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] pyrrole-2,5- Dion (40)  synthesis

The prepared compound (39) (40 mg, 0.054 mmol) and 2M of dimethyl amine (0.2 ml) were dissolved in DMF (3.0 ml), then heated to reflux at 80 ° C. for 12 hours, and the reaction was diluted with water (10 ml). Diluted and extracted three times with ethyl acetate (30 ml). After ethyl acetate was removed and washed three times with hexane, 99% of the desired compound (40) in the form of a red solid was recrystallized using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml). Yield (110 mg, 0.0538 mmol) was obtained.

MS (ESI +); 558 [M + H] ⁺

MP; 100 to 102 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.68 (1H, s), 7.22 (1H, d, J = 8.4 Hz), 6.65-6.62 (1H, m), 6.31 (1H, s), 4.12 (2H, t, J = 7.2 Hz), 3.30 (3H, s), 2.26-2.22 (2H, m), 2.17 (6H, s), 1.92-1.88 (2H, m)

IR; 1708, 1622, 1487, 1454, 1193, 1144, 1127, 1091, 1054, 1025 cm ^-1

< Example  19> 3- [1- (4- Hydroxybutyl )- 1H -Indol-3-yl] -4- [1- (4- Hydroxybutyl ) -5- Methoxy - 1H -Indol-3-yl] -pyrrole-2,5- Dion  Synthesis of 41

A mixture of compound (24) (800 mg, 2.30 mmol) and indol-3-ylacetamide-N-butylacetate (620 mg, 2.15 mmol) prepared in Example 15 was dissolved in THF (20 ml) and at 0 ° C. t-butoxy potassium (9.2 ml) was added slowly. It was then kept at the same temperature for 10 minutes and then reacted at room temperature for 13 hours. Then 2M hydrochloric acid (5 ml) was added, followed by further reaction for 1 hour. The reaction was diluted with water (20 ml) and extracted three times with ethyl acetate (70 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 4) to give the desired compound (41) in the form of a red solid. Obtained 61% yield (702 mg, 1.40 mmol).

MS (ESI +); 524 [M + Na] ⁺

MP; 102 ~ 104 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.24 (s, 1H), 7.51 (s, 1H), 7.31 (d, J = 8.0 Hz, 1H), 7.16 (d, J = 8.8 Hz, 1H), 7.02-6.96 (m, 2H), 6.65 (t, J = 8.0 Hz, 1H), 6.56-6.54 (m, 1H), 4.13-4.07 (m, 4H), 3.51-3.44 (m, 4H), 1.85-1.74 (m, 4H) , 1.48-1.30 (m, 4H)

IR; 3305, 1699, 1650, 1622, 1471, 1197, 1144, 1119, 1086 cm ^-1

< Example  20> 2- [4- (3-4- [1- (4- Carbamimidol sulfanyl butyl )- 1H Indole-3-yl-2,5- Dioxo -2,5-di He Draw 1H -Pyrrole-3-yl] -5- Methoxyindole -1-yl) -butyl]- Isothiourea  Synthesis of 43

Compound (41) (240 mg, 0.38 mmol) and thiourea (117 mg) prepared in Example 19 were dissolved in DMF (4.0 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was then diluted with water (15 ml), extracted twice with ethyl acetate (20 ml) and the material in the organic layer was removed. The water was then removed and washed three times with ethyl acetate, then recrystallized with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to give the desired compound (43) in the form of a red solid 80 Obtained in% yield (210 mg, 0.34 mmol).

MS (ESI +); 618 [M + H] ⁺

MP; 101 ~ 103 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.76 (s, 1H), 7.51 (s, 1H), 7.36 (d, J = 8.4 Hz, 1H), 7.22 (d, J = 8.8 Hz, 1H), 7.05 (t, J = 7.2 Hz, 1H), 6.99 (d, J = 8.0 Hz, 1H), 6.69 (t, J = 7.2 Hz, 1H), 6.62-6.59 (m, 1H), 6.08 (s, 1H), 4.16-4.09 ( m, 4H), 3.09-3.01 (m, 4H), 2.95 (s, 3H), 1.90-1.79 (m, 4H), 1.64-1.56 (m, 4H)

IR; 3305, 3187, 1699, 1642, 1524, 1393, 1340, 1193, 1050 cm ^-1

< Example  21> 3- [1- (4- Dimethylaminobutyl )- 1H -Indol-3-yl] -4- [1- (4- Dimethylaminobutyl ) -5- Methoxy - 1H -Indol-3-yl] -pyrrole-2,5- Dion  Synthesis of 44

Compound (41) (200 mg, 0.32 mmol) and 2M of dimethyl amine (1.28 ml) prepared in Example 19 were dissolved in DMF (6.0 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. Then recrystallization method using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) gave the desired compound (44) in the form of a red solid in 80% yield (150 mg, 0.27 mmol).

MS (ESI +); 556 [M + H] ⁺

MP; 78 ~ 80 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.77 (s, 1H), 7.54 (s, 1H), 7.33 (d, J = 8.0 Hz, 1H), 7.18 (d, J = 8.8 Hz, 1H), 7.03-7.00 (m, 2H), 6.67 (t, J = 7.2 Hz, 1H), 6.60-6.57 (m, 1H), 6.12 (s, 1H) 4.11-4.05 (m, 4H), 2.95 (s, 3H), 2.33-2.15 ( m, 16H), 1.78-1.66 (m, 4H), 1.46-1.40 (m, 4H)

IR; 2933, 2766, 1708, 1618, 1528, 1462, 1332, 1135, 1119, 1091 cm ^-1

< Example  22> 3- [1- (4- Hydroxybutyl )- 1H -Indol-3-yl] -4- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] -pyrrole-2,5- Dion  Synthesis of 45

A mixture of compound (4) (300 mg, 1.03 mmol) and indol-3-ylacetamide-N-butylacetate (450 mg, 1.29 mmol) prepared in Example 1 was dissolved in THF (20 ml) and 0 ° C. T-butoxy potassium (6.0 ml) was added slowly. It was kept at the same temperature for 10 minutes and then reacted at room temperature for 12 hours. Then 2M hydrochloric acid (8.0 ml) was added, followed by reaction for 1 hour, and the reaction was terminated. The mixture was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 4) to obtain the target compound (45) in the form of a red solid. Obtained 44% yield (210 mg, 0.43 mmol).

MS (ESI +); 510 [M + Na] ⁺

MP; 75 ~ 77 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.77 (s, 1H), 7.55 (s, 1H), 7.35 (d, J = 8.4 Hz, 1H), 7.23 (d, J = 8.8 Hz, 1H), 7.06-6.99 (m, 2H), 6.68 (t, J = 7.2 Hz, 1H), 6.59 (dd, J = 2.4, 8.8 Hz, 1H), 6.12 (d, J = 2.4 Hz, 1H), 4.25 (t, J = 6.8 Hz, 2H), 4.12 (t, J = 7.2 Hz, 2H), 3.55-3.48 (m, 4H), 2.97 (s, 3H), 2.02-1.98 (m, 2H), 1.82-1.78 (m, 2H), 1.47 -1.44 (m, 2H)

IR; 3318, 1740, 1732, 1720, 1695, 1630, 1536, 1495, 1462, 1385, 1193, 1119, 1095, 1054 cm ^-1

< Example  23> 3- [1- (4- Dimethylaminobutyl )- 1H -Indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -pyrrole-2,5- Dion  Synthesis of 47

Methanesulfonic acid  3- (3-4- [1- (4- Methanesulfonyloxybutyl )- 1H -Indol-3-yl] -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl-5- Methoxy Synthesis of -indol-1-yl) -propyl Ester (46)

Compound (45) (210 mg, 0.43 mmol) prepared in Example 22 was dissolved in DMF (10 ml). Triethyl amine (0.24 ml, 1.72 mmol) was then added slowly at 0 ° C. and methanesulfonyl chloride (0.13 ml, 1.72 mmol) was added after 3 minutes. The reaction was carried out at the same temperature for 10 minutes and further reaction at room temperature for 10 hours. The reaction was then diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml). Ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (46) in the form of a red solid in 70% yield (200 mg, 0.31 mmol).

3- [1- (4- Dimethylaminobutyl )- 1H -Indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -pyrrole-2,5- Dion  Synthesis of 47

Compound (46) (60 mg, 0.09 mmol) and 2M dimethyl amine (1.0 ml) prepared above were dissolved in DMF (5.0 ml) and then heated to reflux at 80 ° C. for 8 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. Then recrystallization method using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) gave the desired compound (47) in red solid form in 99% yield (50 mg, 0.09 mmol).

MS (ESI +); 542 [M + H] ⁺

MP; 73 ~ 75 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.78 (s, 1H), 7.60 (s, 1H), 7.37 (d, J = 8.4 Hz, 1H), 7.24 (d, J = 8.8 Hz, 1H), 7.05 (t, J = 7.2 Hz, 2H), 7.00 (d, J = 7.2 Hz, 1H), 6.70 (t, J = 7.2 Hz, 1H), 6.60 (dd, J = 2.4, 8.8 Hz, 1H), 6.15 (d, J = 2.4 Hz, 1H), 4.20 (t, J = 6.8 Hz, 2H), 4.14 (t, J = 7.2 Hz, 2H), 2.84 (s, 3H), 2.29-2.25 (m, 4H), 2.21 (s, 6H), 2.18 (s, 6H), 1.99-1.95 (m, 2H), 1.76-1.71 (m, 2H), 1.48-1.46 (m, 2H)

IR; 3318, 2946, 1704, 1626, 1540, 1471, 1393, 1221, 1140, 1123, 1095 cm ^-1

< Example  24> 3- [1- (3-hydroxypropyl) -5- Methoxy - 1H -Indol-3-yl] -4- (1- methyl - 1H -Indol-3-yl) -pyrrole-2,5- Dion  Synthesis of 51

A mixture of compound (4) (500 mg, 1.50 mmol) and 2- (1-methyl-1H-indol-3-yl) -acetamide (50) (300 mg, 1.59 mmol) prepared in Example 1 was THF. (20 ml) and t-butoxy potassium (7.0 ml) was added slowly at 0 ° C. It was then kept at the same temperature for 10 minutes and reacted at room temperature for 12 hours. 2 M hydrochloric acid (8.0 ml) was added, and the reaction was terminated after 1 hour of reaction. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 4) to give the desired compound (51) in the form of a red solid. Obtained 53% yield (340 mg, 0.79 mmol).

MS (ESI +); 452 [M + Na] ⁺

MP; 222 ~ 224 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 10.87 (s, 1H), 7.82 (s, 1H), 7.74 (s, 1H), 7.40 (d, J = 8.4 Hz, 1H), 7.29 (d, J = 8.8 Hz, 1H ), 7.03 (t, J = 7.2 Hz, 1H), 6.77 (d, J = 8.0 Hz, 1H), 6.63 (t, J = 7.2 Hz, 1H), 6.56 (dd, J = 2.4, 8.8 Hz, 1H ), 6.12 (d, J = 2.4 Hz, 1H), 4.61 (t, J = 2.4 Hz, 1H), 4.24 (t, J = 7.2 Hz, 2H), 3.82 (s, 3H), 3.36-3.35 (m , 2H), 3.00 (s, 3H), 1.88-1.84 (m, 2H)

IR; 3383, 2251, 2124, 1752, 1703, 1642, 1532, 1462, 1336, 1217, 1131, 1095, 1050, 1029, 1005 cm ^-1

< Example  25> 3- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -4- [1- methyl - 1H -Indol-3-yl) -pyrrole-2,5- Dion  Synthesis of 53

Methanesulfonic acid  3-5- Methoxy -3- [4- (1- methyl - 1H -Indol-3-yl] -2,5- Dehydro - 1H Synthesis of -pyrrole-3-yl-indole-propyl ester (52)

Compound (51) (340 mg, 0.79 mmol) prepared in Example 24 was dissolved in DMF (10 ml) and triethyl amine (0.22 ml, 1.58 mmol) was added slowly at 0 ° C. After 3 minutes methanesulfonyl chloride (0.12 ml, 1.58 mmol) was added and reacted at the same temperature for 10 minutes, followed by further reaction at room temperature for 10 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml). The ethyl acetate was then removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (52) in the form of a red solid in 80% yield (300 mg, 0.59 mmol).

3- [1- (3-dimethylaminopropyl) -5- Methoxy - 1H -Indol-3-yl] -4- [1- methyl - 1H -Indol-3-yl) -pyrrole-2,5- Dion  Synthesis of 53

Compound (52) prepared above (120 mg, 0.24 mmol) and 2M dimethyl amine (0.24 ml) were dissolved in DMF (5.0 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml), then ethyl acetate was removed and washed three times with hexane. Recrystallization method then gave the desired compound (53) in the form of a red solid in 93% yield (100 mg, 0.22 mmol) using ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml).

MS (ESI +); 457 [M + H] ⁺

MP; 180 ~ 182 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 10.83 (bs, 1H), 7.78 (d, J = 13.6 Hz, 1H), 7.40 (d, J = 8.0 Hz, 1H), 7.29 (d, J = 8.8 Hz, 1H), 7.02 (t, J = 7.2 Hz, 1H), 6.75 (d, J = 8.0 Hz, 1H), 6.61 (t, J = 8.8 Hz, 1H), 6.56 (dd, J = 2.4, 8.8 Hz, 1H), 6.14 (d, J = 2.4 Hz, 1H), 4.21 (t, J = 6.4 Hz, 2H), 3.82 (s, 3H), 3.02 (s, 3H), 2.09-2.05 (m, 8H), 1.84-1.80 (m, 2H)

IR; 2917, 1708, 1622, 1532, 1462, 1328, 1221, 1152, 1123, 1091, 1054 cm ^-1

< Example  26> 2- (3- {5- Methoxy -3- [4- (1- methyl - 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl] -indol-1-yl} -propyl)- Isothiourea  Synthesis of 54

Compound (52) (40 mg, 0.08 mmol) and thiourea (12 mg) prepared in Example 25 were dissolved in DMF (3.0 ml) and then heated to reflux at 80 ° C. for 12 hours. The reaction was diluted with water (15 ml), extracted twice with ethyl acetate (20 ml) and the material in the organic layer was removed. Then water was removed and washed three times with ethyl acetate, followed by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to give 91% of the desired compound (54) in the form of a red solid. Yield (35 mg, 0.07 mmol) was obtained.

MS (ESI +); 488 [M + H] ⁺

MP; 103 ~ 105 ℃

¹ H NMR (400 MHz); CD ₃ OD: δ = 7.77 (s, 1H), 7.68 (s, 1H), 7.36 (d, J = 8.4 Hz, 1H), 7.26 (d, J = 8.8 Hz, 1H), 7.07 (t, J = 8.0 Hz, 1H), 6.86 (d, J = 8.0 Hz, 1H), 6.67-6.62 (m, 2H), 6.25 (d, J = 2.4 Hz, 1H), 4.35 (t, J = 6.4 Hz, 2H) , 3.84 (s, 3H), 3.08 (s, 3H), 3.04 (t, J = 8.0 Hz, 2H), 2.27-2.21 (m, 2H)

IR; 3318, 1753, 1704, 1650, 1618, 1532, 1454, 1348, 1197, 1119, 1058, 1095 cm ^-1

< Example  27> 2- (4- {5- Methoxy -3- [4- (1- methyl - 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrol-3-yl] -indol-1-yl} -butyl)- Isothiourea  Synthesis of 56

3- [1- (4- Hydroxybutyl ) -5- Methoxy - 1H -Indol-3-yl] -4- (1- methyl - 1H Synthesis of -indol-3-yl) -pyrrole-2,5-dione (55)

A mixture of compound (24) (225 mg, 0.65 mmol) and 2- (1-methyl- 1H -indol-3-yl) -acetamide (50) (250 mg, 1.32 mmol) prepared in Example 15 was diluted with THF. (20 ml) and slowly added t-butoxy potassium (4.0 ml) at 0 ° C. After holding at the same temperature for 10 minutes, the reaction was carried out at room temperature for 12 hours. Subsequently, 2M hydrochloric acid (8.0 ml) was added, followed by reaction for 1 hour, and the reaction was terminated. The reaction was diluted with water (30 ml) and extracted three times with ethyl acetate (50 ml), and then the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (55) in the form of a red solid. Obtained in 52% yield (150 mg, 0.34 mmol).

Methanesulfonic acid  4-5- Methoxy -3- [4- (1- methyl - 1H -Indol-3-yl] -2,5- Dioxo -2,5- Dehydro - 1H Synthesis of -pyrrole-3-yl] -indol-1-yl-butyl ester (56)

Compound (55) (150 mg, 0.34 mmol) prepared above was dissolved in DMF (10 ml), and triethyl amine (0.14 ml, 0.70 mmol) was slowly added at 0 ° C., and after 3 minutes, methanesulfonyl chloride (0.04 ml, 0.50 mmol) was added. The reaction was carried out at the same temperature for 10 minutes, and further reacted at room temperature for 8 hours. The reaction was then diluted with water (10 ml) and extracted three times with ethyl acetate (30 ml). Then ethyl acetate was removed and the residue was purified by column chromatography (hexane: ethyl acetate = 1: 3) to give the desired compound (56) in the form of a red solid in 96% yield (170 mg, 0.33 mmol).

2- (4-5- Methoxy -3- [4- (1- methyl - 1H -Indol-3-yl) -2,5- Dioxo -2,5- Dehydro - 1H -Pyrrole-3-yl] -indol-1-yl-butyl)- Isothiourea  Synthesis of 56

Compound (56) (34 mg, 0.07 mmol) and thiourea (10 mg) prepared above were dissolved in DMF (3.0 ml) and then heated to reflux at 80 ° C. for 7 hours. The reaction was diluted with water (15 ml), extracted twice with ethyl acetate (20 ml) and the material in the organic layer was removed. Then water was removed and washed three times with ethyl acetate, followed by recrystallization with ethyl acetate (3 ml), hexane (5 ml) and methanol (1 ml) to 92% of the desired compound (57) in the form of a red solid. Yield (30 mg, 0.06 mmol) was obtained.

MS (ESI +); 502 [M + H] ⁺

MP; 82 ~ 84 ℃

¹ H NMR (400 MHz); DMSO- d ₆ : δ = 7.78 (s, 1H), 7.63 (s, 1H), 7.34 (d, J = 8.4 Hz, 1H), 7.23 (d, J = 8.8 Hz, 1H), 7.06 (t, J = 8.0 Hz, 1H), 6.86 (d, J = 8.0 Hz, 1H), 6.66-6.59 (m, 2H), 6.18 (d, J = 2.4 Hz, 1H), 4.23 (t, J = 6.4 Hz, 2H ), 3.82 (s, 3H), 3.30 (t, J = 8.0 Hz, 2H), 3.29 (s, 3H), 2.00-1.95 (m, 2H), 1.71-1.66 (m, 2H)

IR; 3326, 3199, 1753, 1699, 1654, 1609, 1528, 1446, 1336, 1193, 1152, 1127, 1095, 1062 cm ^-1

< Test Example  1> The compound of the present invention NCR3 Of the effect on the expression of

Test Example  1-1

To confirm the effect of the compounds of the present invention on the expression of NCR3 (NKp30) on the surface of NK cells, tests were performed using immunofluorescence staining and flow cytometry. Specifically, 5 μM of compounds (Examples 2, 4, 10, 13, 14, 16, 17, and 18) randomly selected from the prepared examples were added, and cells were cultured for 24 hours. . The cultured NK cells were washed twice with PBS, and 10 µl of PE-conjugated anti-NCR3 antibody (Beckman Coulter) and 30 µl at 4 ° C in 90 µl of PBS containing 0.1% BSA. The reaction was carried out for a minute. The reaction was then washed twice with PBS to measure fluorescence intensity with FACStar (Becton Dickinson) and analyzed using WinMDI 2.8 program, the results are shown in FIGS. 2 and 3.

2 is a result of measuring the expression of NCR3 on the surface of NK cells treated with the compound of the present invention using a flow cytometry, in which the thick black line is an isotype control and the green line is basically NCR3 to be expressed as, gray filled portion represents NCR3 expressed after treatment with the compound of the present invention. As can be seen from FIG. 2, NCR3 was expressed in freshly isolated or 24-hour cultured NK cells (green line), but was not treated on the surface of NK cells treated with the compound prepared in Example. Compared to the case, a significant amount of NCR3 was expressed (gray filled portion). The result of analyzing the results of FIG. 2 by relative MFI (relative mean fluorescence intensity ratio) is shown in Table 1 below, which is graphically represented in FIG. 3.

Relative MFI ratio (fold) of each Example when G mean value of untreated NK cells is set to 1 Unprocessed NK  cell One Example  2 5.5 Example  4 About 3.5 Example  10 About 3 Example  13 About 6 Example  14 About 4 Example  16 About 6 Example  17 About 3 Example  18 5.5

As can be seen from Table 1 and FIG. 3, NK cells treated with the compounds of Examples 2, 13, 16, and 18 were observed to express about 6 times more NCR3 than untreated cells. In addition, in the case of the compounds of Examples 4, 10, 14 and 17, expression of NCR3 of about 3 times or more was observed.

Test Example  1-2

5 μM of each of the selected compounds (Examples 2, 20, 21, 23, and 25 to 27) of the prepared examples were added, and NK cells having different donors from those used in Test Example 1-1 were incubated for 4 hours. . The cultured NK cells were washed twice with PBS, and 10 µl of PE-conjugated anti-NCR3 antibody (Beckman Coulter) and 30 µl at 4 ° C in 90 µl of PBS containing 0.1% BSA. The reaction was carried out for a minute. Subsequently, the reaction was washed twice with PBS to measure fluorescence intensity with FACScalibur (Becton Dickinson), and analyzed using WinMDI 2.8 program. The results are shown in FIG. 4.

Figure 4 is a result of measuring the expression of NCR3 on the surface of NK cells treated with a compound of the present invention using a flow cytometer, the thick black line in the figure is the isotype control, the green line is basically expressed NCR3, filled with gray Portions represent NCR3 expressed after treatment with a compound of the present invention. As a result of analyzing the results of FIG. 4 by relative MFI (relative mean fluorescence intensity ratio), the results are shown in Table 2, and these are graphically represented in FIG. 5.

Unprocessed NK  When the G mean value of the cells is set to 1, Example  relative MFI  Rate (times) Unprocessed NK  cell One Example  2 7.52 Example  20 3.02 Example  21 3.57 Example  23 3.69 Example  25 9.33 Example  26 10.43 Example  27 16.48

As can be seen from Table 2 and FIG. 5, the compounds of the present invention significantly increased the expression of NCR3 on the surface of NK cells. That is, in NK cells treated with the compounds of Examples 2 and 25 to 27, expression of NCR3 of about 8 times or more was observed than that of untreated, especially in Examples 26 and 27, 10 times or more expression of NCR3 was observed. It became.

< Test Example  2> the compound of the present invention NCR1 Of the effect on the expression of

Compounds of randomly selected examples (Examples 2, 4, 5, 6, 10, 11, 13, 14, 16, 17, and 18 to analyze the effect of the compounds of the invention on the expression of NCR1 on the surface of NK cells ), The test was carried out in the same manner as in Test Example 1-1, and the results are shown in Figures 6 and 7.

NCR1 is a receptor expressed in both activated and inactivated NK cells, as shown in FIG. 6. Significant expression was observed in all freshly isolated or NK cells cultured for 24 hours (green line). As in the case of NCR3, it was confirmed that the surface of NK cells treated with the compound of Example 2 expressed much higher amounts of NCR1 as compared to the untreated (grey-filled portion). Quantitatively analyzing the results of FIG. 6 by comparing relative MFIs are shown in Table 3 and FIG. 7.

Relative MFI ratio (fold) of each Example when G mean value of untreated NK cells is set to 1 Unprocessed NK  cell One Example  2 About 2.3 Example  4 Approximately 1.4 Example  5 1.3 Example  6 1.3 Example  10 About 1.6 Example  11 1.3 Example  13 About 2.4 Example  14 Approximately 1.4 Example  16 About 2.5 Example  17 Approximately 1.4 Example  18 Approximately 1.8

As can be seen from the results of Table 2 and FIG. 7, NCR1 expression was significantly increased even when treated with the compounds of Examples 2, 13 and 16 (about 2 times or more), and other cases were compared with the untreated cases. When a large amount of NCR1 was confirmed that it was expressed.

< Test Example  3> The compound of the present invention NCR2 Of the effect on the expression of

In order to confirm the effect of the compound of the present invention on the expression of NCR2 in NK cells, the test was performed in a similar manner to Test Example 1 using the compound of Example 2. In addition, NK cells were previously treated with PMA, which is known as an NK cell activator, and compared with the untreated case, the results are shown in FIGS. As can be seen from FIG. 8, NCR2 was hardly expressed in untreated NK cells or NK cells treated with PMA, whereas expression of NCR2 was induced very strongly in NK cells treated with the compound of Example 2, respectively. Quantitative analysis of the relative MFI ratio is shown in Table 4 and FIG. 9.

Unprocessed NK  When the G mean value of the cells is set to 1, Example  relative MFI  Rate (times) Unprocessed NK  cell One PMA One Example  2 About 30

As shown in Table 4 and FIG. 9, it was confirmed that NK cells treated with the compound of Example 2 of the present invention express NCR2 at least about 30-fold higher than untreated or treated with PMA.

As can be seen from the results of Test Examples 1 to 3, the compounds of the present invention significantly increased the extracellular expression of NCR1, 2 and 3. The relative MFI ratio, which quantitatively analyzes the increase of each NCR expression, shows that the expression amount after treatment with the compound of the present invention is somewhat different depending on the type of NCR. This difference appears because NCR2 is rarely expressed in normal NK cells, while NCR3 is very weakly expressed. As such, direct comparison is difficult due to the difference in the expression state according to the type of NCR. However, when the amounts of NCR1, 2, and 3 expressed in NK cells treated with the compound of the present invention are compared, all NCRs are strongly expressed. You can check it.

< Test Example  4> Cytokines NCR Of the effect on the expression of

To analyze the effect of cytokines on the expression of NCR in NK cells, tests were performed using immunofluorescence staining and flow cytometry, similar to Test Example 1. The cytokines used in the tests were IL-2 (1 ng / ml), IL-15 (1 ng / ml), IL-8 (10 ng / ml), IL-12 (10 ng / ml), IL-18 ( 10 ng / ml), IFN-α1 (10 ng / ml) and IFN-α2b (10 ng / ml). When the G average value of NCR expression of untreated NK cells in simple culture was set to 1, after 24 hours of treatment with each cytokine, the amount of NCR expression on the surface of NK cells was expressed as a relative MFI ratio.

As can be seen from FIG. 10, only IL-12 of the cytokines used in the test slightly increased the expression of NCR3 (FIG. 10 (A)), while IL-15 and IFN-α2 slightly increased the expression of NCR1 ( 10 (B)), both cases were weak. In addition, in the case of NK cells treated with other cytokines, the expression levels of NCR1 and NCR3 were little different from those of untreated NK cells (FIG. 10).

As described above, cytokines such as IL-2, IL-12 and IL-15 are known to play an important role in the differentiation and development of NK cells. Accordingly, in the above test examples, the effects of cytokines such as IL-2, IL-8, IL-12, IL-15, IL-18, IFNa1 and IFN-a2b on NCR expression on the surface of NK cells were examined. As a result, it was confirmed that all of these cytokines had no significant effect on the expression of NCR (FIG. 10). In other words, some of the cytokines (such as IL-12, IL-15 and IFN-α2) slightly increased the expression of NCR1 or NCR3, but all were difficult to see as meaningful results. These results indicate that the compounds of the present invention activate NK cells in a different way than cytokines affect the differentiation, development, growth and proliferation of NK cells.

< Test Example  5> The compound of the present invention NK  Of cancer cells On cell killing ability  Analysis of the impact

Test Example  5-1 ( PMA  And Example  2, compound)

The effect of NK cells treated with PMA 100 ng / ml and 5 μM of the compound of Example 2 on the apoptosis of hepatocellular carcinoma was confirmed through a JAM test. Specifically, target cells were seeded at 1 × 10 ⁴ cells / well in 96-well plates and cultured so that HepG2 and Hep3B adhered to the bottom. Removing the cell culture medium, and ³ [H] - thymidine ^{(3 [H] -thymidine) (} NEN) was allowed to uptake (uptake) with 20 μCi / well. The remaining ³ [H] -thymidine was removed and cocultured with a ratio of NK cells and target cells 3: 1. After 2 hours the cells attached to the bottom of the plate were removed from the bottom using 0.125% trypsin / 0.5 mM EDTA (Gibco BRL). The target cells are adsorbed onto a glass fiber filter (Wallac Oy), washed well, and then melted and melted the Meltilex (Wallac) to cover the filter, and the amount of radiation is emitted by β-Wallac. Measured.

Next, using the following formula (1), the amount of NK cells causing cell death was calculated.

Equation (1):% DNA fragmentation = ((1-experimental value) / negative control value) x 100

Then treated with the compound of Example 2 The cytotoxicity of NK cells against cancer cells was analyzed. HepG2 and Hep3B, which are liver cancer cell lines, were used as target cells. After NK cells were treated with the compound of Example 2, the cells were mixed and cultured with liver cancer cell lines, respectively, and the amount of dead liver cancer cell line was quantified. In addition, the cell death of hepatocellular carcinoma cell line by NK cells was measured using a jam test, and is shown in FIG. As shown in FIG. 11, untreated NK cells and NK cells treated with PMA were found to kill HepG2 or Hep3B liver cancer cell lines within about 20% for 2 hours, but NK cells treated with the compound of Example 2 were nearly 40%. It was confirmed that the mortality was about 2 times or more.

Test Example  5-2 ( Example  2 and Example  27 compounds)

As a target cell, HepG2 and Hep3B, which are liver cancer cell lines, and HeLa, which are cervical cancer cell lines, were used, and NK cells provided by donors different from Test Example 5-1 were treated with the compounds of Examples 2 and 27 for 4 hours. Except for the above, the effect of the compounds of Examples 2 and 27 of the present invention on the killing ability of NK cells against cancer cells was measured by a method similar to Test Example 5-1, and the results are shown in FIG. 12.

As can be seen from Figure 12, NK cells treated with the compounds of Examples 2 and 27 of the present invention as compared to untreated NK cells was found to have excellent killing ability against cancer cell lines. In addition, the killing ability of the NK cells treated with Example 27 was better than the NK cells treated with Example 2. This result is because, as shown in Test Example 1, etc., the compound of Example 27 increased NCR expression on the surface of NK cells more than the compound of Example 2, which also increased the natural killing ability of NK cells.

< Test Example  6> The compound of the present invention NK  Suicide on Cells Of ligands Extracellular  Analysis of the impact on expression

The effect of the compounds of the present invention on the expression of FasL and TRAIL, which are suicide ligands on the surface of NK cells, was measured using immunofluorescence staining and flow cytometry. Specifically, NK cells were incubated with 5 μM of the compound of Example 2 for 24 hours. Subsequently, the cultured NK cells were washed twice with PBS, and PE-conjugated anti-FasL antibody or PE-bound anti-TRAIL antibody bound to 90 μl of PBS containing 0.1% BSA. 10 µl of (PE-conjugated anti-TRAIL antibody) was added and reacted at 4 ° C for 30 minutes. The reaction was then washed twice with PBS, fluorescence intensity was measured with FACScalibur (Becton Dickinson), and the results were analyzed using the WinMDI 2.8 program. NK cells were treated with the compound of Example 2 as described above, and after 24 hours, the amounts of FasL and TRAIL expressed on the surface of NK cells were analyzed by flow cytometry. The analysis results were compared through the relative MFI, and the results are shown in FIG. 13. As can be seen from Figure 13, NK cells treated with the compound of the present invention was found to express FasL about 30 times or more, TRAIL more than about 40 times more than untreated NK cells.

Such suicide ligands of NK cells have a function of inducing apoptosis by recognizing suicide receptors of target cells such as cancer cells. Accordingly, the NK cells treated with the compounds of the present invention increase the expression of suicide ligands in addition to increasing the expression of NCR, indicating that killing ability against cancer cells and the like can be further increased.

< Test Example  7> Analysis of anticancer effects on mice of the compounds of the present invention

In order to confirm the anticancer effect of the compound of the present invention on the mouse, the difference in survival time between the control group and the group injected with the compound of Example 2 (treatment group) in the tumor mouse model was investigated. As a tumor mouse model, 8-week-old C57BL / 6 mice were injected subcutaneously with 1 × 10 ⁶ of malignant melanoma cancer cell line B16BL6 cells. 4.4 μg (2 nmole) of the compound of Example 2 of the present invention was dissolved in PBS, and this was performed 5 times at 2 to 3 days each, once 2 days before cancer cell injection, 1 time on cancer cell injection, and 3 times after cancer cell injection. Killed (treated group). As a control, one injected with only PBS five times at the same time was used. Survival rates for 2 months after cancer cell injection were examined and the results are shown in FIG. 14. Tumors began to grow after about 2 weeks in both the control and treatment groups, and no significant difference in tumor size was observed between the two groups. However, in the control group, all died 35 days before cancer cell injection, whereas in the treatment group, about one third died at about the same time as the control group, but about one third lived about one week longer than the control group, / 3 survived without dying for the experimental period (63 days). These results suggest that the compounds of the present invention play a role in prolonging the survival time of tumor mouse models, and have excellent in vivo anticancer effects.

< Test Example  8> Mouse in vivo of the compound of the present invention Antitransition effect

Example  2, compound

The mouse cancer metastasis model was obtained by intravenously injecting 1 x 10 ⁵ malignant melanoma cancer cell line B16BL6 cells into the tail of 8-week-old C57BL / 6 mice. In the treatment group, 4.4 μg (2 nmole) or 22 μg (5 nmole) of the compound of Example 2 dissolved in PBS was added once every two days before cancer cell injection, once every day of cancer cell injection, and three times after cancer cell injection, two to three days apart. A total of 5 treatments were performed. The control group was injected with PBS five times at the same time as the treatment group. On day 18 after cancer cell injection, control and treatment mice were simultaneously caught and lung melanoma was compared. In each group, statistics were counted with one melanoma spot and one case with the least number of melanoma spots. As a result, as shown in FIG. 15, the average number of melanoma spots metastasized to the lungs of the control group was 60, whereas in the group treated with 4.4 μg (2 nmole) or 22 μg (5 nmole) of the compound of Example 2 The spots were 34.9 and 26.3, respectively, indicating that metastasis was effectively prevented.

Example  27 compounds

The mouse cancer metastasis model was obtained by intravenously injecting 1 x 10 ⁵ malignant melanoma cancer cell line B16BL6 cells into the tail of 8-week-old C57BL / 6 mice. In the treatment group, 5 μg (2 nmol) or 25 μg (5 nmol) of the compound of Example 27 dissolved in PBS was added once every 2 days before cancer cell injection, once every day of cancer cell injection, and 3 times each after cancer cell injection. Five treatments were performed. The control group was injected with PBS five times at the same time as the treatment group. On the 21st day after the cancer cell injection, the control and treatment mice were simultaneously caught to compare the melanoma in the lungs, except for the one with the most and the least with melanoma spots. As a result, as shown in FIG. 16 (A), the average number of melanoma spots metastasized to the lungs of the control group was 78, whereas 5 μg (2 nmole) or 25 μg (5 nmole) of the compound of Example 27 was treated. In the case of the group, an average of 33.6 and 32.6 spots was shown, indicating that metastasis was effectively prevented. Figure 16 (B) summarizes the above results to show the anti-transduction effect of the compounds of Examples 2 and 27 as a percentage.

1 is a view showing the mechanism of action of the NK cell activating material of the present invention.

2 is a diagram showing the results of measuring the expression level of NCR3 by flow cytometry when NK cells were treated with the compound of the present invention in Test Example 1-1.

3 is a diagram showing the expression level of NCR3 in relative MFI ratio when NK cells were treated with the compound of the present invention in Test Example 1-1.

4 is a diagram showing the results of measuring the expression level of NCR3 by flow cytometry when NK cells were treated with the compound of the present invention in Test Example 1-2.

FIG. 5 is a diagram showing the expression level of NCR3 in relative MFI ratio when NK cells were treated with the compound of the present invention in Test Example 1-2.

Figure 6 is a diagram showing the result of measuring the expression level of NCR1 by flow cytometry when treated with NK cells with a compound of the present invention.

7 is a diagram showing the expression of NCR1 in relative MFI ratios when treated with NK cells with the compounds of the present invention.

8 is a diagram showing the results of measuring the expression level of NCR2 in untreated NK cells, compounds of the present invention and PMA treated NK cells by flow cytometry.

FIG. 9 is a diagram showing the results of FIG. 8 in relative MFI ratios.

10 is a diagram showing the expression of NCR of untreated NK cells and cytokine-treated NK cells in relative MFI ratios.

11 and 12 are diagrams showing the effect of the compounds of the present invention on the killing ability of NK cells against cancer cells.

Figure 13 shows the effect of the compounds of the present invention on the expression of suicide ligands on NK cells.

14 is a diagram showing the anti-cancer effect of the mouse of the present invention in vivo.

15 and 16 are diagrams showing anti-metastatic effects in mice in vivo of the compounds of the present invention.

Claims (29)

A compound represented by the following formula (1), a pharmaceutically acceptable salt or solvate thereof: [Formula 1]

In which R ₁ represents hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl or alkoxy; R ₂ and R ₃ each independently represent hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl, alkoxy, amino, cyano, carboxyl, aryl or -R ₂₃ R ₂₄ ; R ₂₃ is linear or branched alkylene; Straight or branched alkenylene; And straight or branched alkynylene; R ₂₄ is hydroxy; Alkylamino; Dialkylamino; Amino; Cyano; Carboxyl; Substituted or unsubstituted aryl, heteroaryl or heterocyclic compounds; Thiourea; Isothiourea; Alkylthio; Thiocyanato; Sulfamoyl or alkylsulfonyl; R ₄ to R ₁₁ each independently represent hydrogen, alkyl, cycloalkyl or alkoxy. The method of claim 1, R ₁ is hydrogen, halogen, hydroxy, alkyl having 1 to 12 carbon atoms, cycloalkyl having 3 to 12 carbon atoms, alkenyl having 2 to 12 carbon atoms, alkynyl having 2 to 12 carbon atoms or alkoxy having 1 to 12 carbon atoms. Compounds, pharmaceutically acceptable salts or solvates thereof. The method of claim 1, R ₂ and R ₃ are each independently hydrogen, hydroxy, alkyl having 1 to 12 carbon atoms, cycloalkyl having 3 to 12 carbon atoms, alkenyl having 2 to 12 carbon atoms, alkynyl having 2 to 12 carbon atoms, and having 1 to 12 carbon atoms. Alkoxy, amino, cyano, carboxyl, phenyl, naphthyl or -R ₂₃ R ₂₄ ; In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 12 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 12 carbon atoms; Dialkylamino having 2 to 24 carbon atoms; Aryl, heteroaryl, any one selected from the group consisting of phenyl, indolyl, pyrrolyl, furanyl, pyridinyl, piperidinyl, pyranyl, pyridazinyl, pyrimidinyl, pyrazinyl, piperazinyl and triazolyl Or heterocyclic compounds; Thiourea; Or isothiourea, The aryl, heteroaryl or heterocyclic compound may be substituted with one or more selected from the group consisting of hydroxy, amino, cyano, arylalkyl, aryl unsubstituted or substituted with alkyl, and alkyl substituted or unsubstituted with carboxyl. Compound, a pharmaceutically acceptable salt or solvate thereof. The method of claim 3, wherein R ₂ and R ₃ are each independently hydrogen, alkyl of 1 to 8 carbon atoms, cycloalkyl of 3 to 8 carbon atoms, alkenyl of 2 to 8 carbon atoms, alkynyl of 2 to 8 carbon atoms, alkoxy of 1 to 8 carbon atoms, or- R ₂₃ R ₂₄ is represented, In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 8 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 8 carbon atoms; Dialkylamino having 2 to 16 carbon atoms; Any one aryl, heteroaryl or heterocyclic compound selected from the group consisting of phenyl, pyridinyl and triazolyl; Thiourea; Or isothiourea, The aryl, heteroaryl, or heterocyclic compound is hydroxy, amino, cyano, arylalkyl having 7 to 12 carbon atoms, aryl unsubstituted or substituted with alkyl having 1 to 4 carbon atoms and 1 or unsubstituted carbon atoms with carboxyl A compound, a pharmaceutically acceptable salt or solvate thereof, which may be substituted with one or more selected from the group consisting of alkyl of 12 to 12. The method of claim 4, wherein R ₂ and R ₃ are each independently hydrogen, alkyl having 1 to 4 carbon atoms, cycloalkyl having 3 to 6 carbon atoms, alkenyl having 2 to 4 carbon atoms, alkynyl having 2 to 4 carbon atoms, alkoxy having 1 to 4 carbon atoms, or- R ₂₃ R ₂₄ is represented, In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 4 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 4 carbon atoms; Dialkylamino having 2 to 8 carbon atoms; Substituted or unsubstituted triazolyl, thiourea or isothiourea, Wherein the substituent of triazolyl is at least one selected from the group consisting of alkyl having 1 to 4 carbon atoms unsubstituted or substituted with benzyl and carboxyl, a pharmaceutically acceptable salt or solvate thereof. The method of claim 1, R ₄ to R ₁₁ are each independently hydrogen, alkyl having 1 to 8 carbon atoms or alkoxy having 1 to 8 carbon atoms, a pharmaceutically acceptable salt or solvate thereof. The method of claim 1, R ₁ is hydrogen or alkyl of 1 to 8 carbon atoms, R ₂ and R ₃ are each independently hydrogen, alkyl of 1 to 8 carbon atoms, cycloalkyl of 3 to 8 carbon atoms, alkenyl of 2 to 8 carbon atoms, alkynyl of 2 to 8 carbon atoms, alkoxy of 1 to 8 carbon atoms, or- R ₂₃ R ₂₄ ; In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 8 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 8 carbon atoms; Dialkylamino having 2 to 16 carbon atoms; Any one aryl, heteroaryl or heterocyclic compound selected from the group consisting of phenyl, pyridinyl and triazolyl; Thiourea; Or isothiourea; The aryl, heteroaryl or heterocyclic compound is hydroxy, amino, cyano, arylalkyl having 7 to 12 carbon atoms, aryl unsubstituted or substituted with alkyl having 1 to 4 carbon atoms and 1 or unsubstituted carbon atoms with carboxyl. To one or more selected from the group consisting of alkyl of 12 to 12; R ₄ to R ₁₁ are each independently hydrogen, alkyl having 1 to 8 carbons or alkoxy having 1 to 8 carbons, a pharmaceutically acceptable salt or solvate thereof. The method of claim 7, wherein R ₁ is hydrogen, methyl or ethyl, R ₂ and R ₃ are each independently hydrogen, alkyl having 1 to 4 carbon atoms, cycloalkyl having 3 to 6 carbon atoms, alkenyl having 2 to 4 carbon atoms, alkynyl having 2 to 4 carbon atoms, alkoxy having 1 to 4 carbon atoms, or- R ₂₃ R ₂₄ ; In the above, R ₂₃ represents a straight or branched chain alkylene having 1 to 4 carbon atoms, R ₂₄ is hydroxy, alkylamino having 1 to 4 carbon atoms; Dialkylamino having 2 to 8 carbon atoms; Substituted or unsubstituted triazolyl, thiourea or isothiourea; The substituent of triazolyl is at least one selected from the group consisting of alkyl having 1 to 4 carbon atoms unsubstituted or substituted with benzyl and carboxyl; A compound, a pharmaceutically acceptable salt or solvate thereof, wherein R ₄ to R ₁₁ are each independently hydrogen, methyl, ethyl, methoxy or ethoxy. The method of claim 1, Compound of Formula 1, 3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione; 3- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione; 3- [1- (2-hydroxyethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -pyrrole-2,5-dione; 3- [1- (2-dimethylaminoethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione; 3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione; 3-[(1-benzyl-1H- [1,2,3] triazol-4-ylmethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl ) -Pyrrole-2,5-dione; (4- {3- [4- (1H-Indol-3-yl) -2,5-dioxy-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxy-1H-indole -1-ylmethyl}-[1,2,3] triazol-1-yl) acetic acid; 3- [1- (2-dimethylaminoethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione; 3- [1- (2-hydroxyethyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -1-methylpyrrole-2,5-dione; 2- (2- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} -ethyl) isothiourea; 2- (3- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} propyl) isothiourea; 3- [1- (3-hydroxypropyl) -1H-indol-3-yl] -4- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione; 3- [1- (3-dimethylaminopropyl) -1H-indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione; 2- [3- (3- {4- [1- (3-carbamimidoylsulfanylpropyl) -1H-indol-3-yl] -2,5-dioxo-2,5-dihydro-1H- Pyrrole-3-yl} -5-methoxyindol-1-yl) -propyl] -isothiourea; 3- [1- (4-hydroxybutyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) -pyrrole-2,5-dione; 2- (4- {3- [4- (1H-Indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrol-3-yl] -5-methoxyindole- 1-yl} butyl) isothiourea; 3- [1- (4-dimethylaminobutyl) -5-methoxy-1H-indol-3-yl] -4- (1H-indol-3-yl) pyrrole-2,5-dione; 3,4-bis [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] pyrrole-2,5-dione; 3- [1- (4-hydroxybutyl) -1H-indol-3-yl] -4- [1- (4-hydroxybutyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione; 2- [4- (3- {4- [1- (4-carbamimidolsulfanylbutyl) -1H-indol-3-yl} -2,5-dioxo-2,5-dihydro-1H- Pyrrole-3-yl] -5-methoxyindol-1-yl) -butyl] -isothiourea; 3- [1- (4-Dimethylaminobutyl) -1 H-indol-3-yl] -4- [1-4-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione; 3- [1- (4-Dimethylaminobutyl) -1 H-indol-3-yl] -4- [1-4-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione; 3- [1- (4-Dimethylaminobutyl) -1H-indol-3-yl] -4- [1-3-dimethylaminopropyl) -5-methoxy-1 H-indol-3-yl] -pyrrole- 2,5-dione; 3- [1- (3-hydroxypropyl) -5-methoxy-1H-indol-3-yl] -4- (1-methyl-1H-indol-3-yl) -pyrrole-2,5-dione ; 3- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -4- [1-methyl-1H-indol-3-yl) -pyrrole-2,5-dione ; 2- (3- {5-methoxy-3- [4- (1-methyl-1H-indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrole-3- Il] -indol-1-yl} -propyl) -isothiourea; or 2- (4- {5-methoxy-3- [4- (1-methyl-1H-indol-3-yl) -2,5-dioxo-2,5-dihydro-1H-pyrrole-3- Il] -indol-1-yl} -butyl) -isothiourea, a pharmaceutically acceptable salt or solvate thereof. The method of claim 9, Compound of Formula 1, 3- [1- (3-hydroxypropyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione; 3- [1- (2-dimethylaminoethyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione; 2- (2- {3- [4- ( 1H -Indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl] -5-methoxyindole- 1-yl} -ethyl) isothiourea; 3- [1- (3-dimethylaminopropyl) - 1H-indol-3-yl] -4- [1- (3-dimethylaminopropyl) -5-methoxy-1H-indol-3-yl] -pyrrole -2,5-dione; 2- [3- (3- {4- [1- (3-carbamimidoyl sulfanyl propyl) - 1H - indol-3-yl] -2,5-dioxo-2,5-dihydro - 1H - Pyrrole-3-yl} -5-methoxyindol-1-yl) -propyl] -isothiourea; 2- (4- {3- [4- ( 1H -Indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrol-3-yl] -5-methoxyindole- 1-yl} butyl) isothiourea; 3- [1- (4-dimethylaminobutyl) -5-methoxy- 1H -indol-3-yl] -4- ( 1H -indol-3-yl) pyrrole-2,5-dione; 3,4-bis [1- (3-dimethylaminopropyl) -5-methoxy- 1H -indol-3-yl] pyrrole-2,5-dione; 3- [1- (3-dimethylaminopropyl) -5-methoxy- 1H -indol-3-yl] -4- [1-methyl- 1H -indol-3-yl) -pyrrole-2,5-dione ; 2- (3- {5-methoxy-3- [4- (1-methyl- 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrole-3- Il] -indol-1-yl} -propyl) -isothiourea; or 2- (4- {5-methoxy-3- [4- (1-methyl- 1H -indol-3-yl) -2,5-dioxo-2,5-dihydro- 1H -pyrrole-3- Il] -indol-1-yl} -butyl) -isothiourea, a pharmaceutically acceptable salt or solvate thereof. A method for preparing a compound of Formula 1 ′, comprising the step of reacting a compound of Formula 2 with a compound of Formula 3: [Formula 2]

[Formula 3]

[Formula 1 ’]

Wherein R ₁ and R ₄ to R ₁₁ are as defined in claim 1, R ₂ ′ and R ₃ ′ each independently represent hydrogen, halogen, hydroxy, alkyl, cycloalkyl, alkenyl, alkynyl, alkoxy, amino, cyano, carboxyl, aryl or —R ₂₃ 'R ₂₄ '; R ₂₃ ′ in the above is a linear or branched alkylene; Straight or branched alkenylene; And linear or branched alkynylene, and R ₂₄ ′ is hydroxy; Amino; Cyano; Carboxyl; Substituted or unsubstituted aryl, heteroaryl or heterocyclic compounds; Alkylthio; Thiocyanato; Sulfamoyl; Or a protecting group. The method of claim 11, The method of claim 1, further comprising the step of reacting the compound of formula (1) with the nucleophile. The method of claim 11, The method of claim 1, further comprising the step of reacting the compound of formula (1) with an electrophile. The method of claim 11, A compound of formula (2) is obtained by reacting a compound of formula (4) with oxalyl chloride and sodium methoxide: [Formula 4]

Wherein R ₂ ′ and R ₄ to R ₇ are as defined in claim 11. The method of claim 11, A compound of formula 3 is prepared by reacting a compound of formula 5 with a base: [Formula 5]

Wherein R ₃ ′ and R ₈ to R ₁₁ are as defined in claim 11. NK cells treated with the compound of any one of claims 1 to 10. Comprising the step of culturing the NK cells in the culture medium to which the compound of any one of claims 1 to 10 added A method of treating NK cells identified from a host. The method of claim 17, The host is a human, monkey, cow, sheep, goat, horse, pig, rabbit, dog, cat, rat, mouse or morphot. The method of claim 17, The culture medium is RPMI, MEM or DMEM. The method of claim 17, The culture temperature is 30 to 37 ℃, the culture time is characterized in that 1 to 48 hours. The method of claim 17, 1 to 10 μM of the compound is added per 105 to 106 cells / ml of NK cells. A compound of any one of claims 1 to 10, a pharmaceutically acceptable salt or solvate thereof; And Pharmaceutical composition for the treatment of a disease, characterized in that it comprises a pharmaceutically acceptable carrier. The method of claim 22, Disease cancer; Or an infectious disease caused by a virus or bacterium. The method of claim 23, The cancer is lung cancer, liver cancer, stomach cancer, colon cancer, bladder cancer, prostate cancer, breast cancer, ovarian cancer, cervical cancer, thyroid cancer, melanoma or hematologic cancer. The method of claim 23, Infectious diseases caused by viruses or bacteria are AIDS, bird flu, flu, CMV infectious diseases, tuberculosis or leprosy. A pharmaceutical composition for the treatment of a disease comprising the NK cell of claim 16 and a pharmaceutically acceptable carrier. The method of claim 26, Disease cancer; Or an infectious disease caused by a virus or bacterium. The method of claim 27, The cancer is lung cancer, liver cancer, stomach cancer, colon cancer, bladder cancer, prostate cancer, breast cancer, ovarian cancer, cervical cancer, thyroid cancer, melanoma or hematologic cancer. The method of claim 27, Infectious diseases caused by viruses or bacteria are AIDS, bird flu, flu, CMV infectious diseases, tuberculosis or leprosy.

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