Pyrrolopyrimidine derivative or pharmaceutically acceptable salt and application thereof

Abstract

The invention discloses a pyrrolopyrimidine derivative or a pharmaceutically acceptable salt and application thereof. The general formula of the pyrrolopyrimidine derivative is shown in the specification. Wherein R1 is substituted or unsubstituted phenyl or a 5-membered or 6-membered heterocyclic group of heteroatoms of oxygen, nitrogen and sulfur, and substituent groups are selected from halogen atoms, C1-C6 alkyl groups, C1-C3 alkoxy groups, cyano groups and nitro groups; x is selected from the group consisting of,,,,,, and the like; y1 is a C2-C5 alkenyl group, a C2-C5 alkynyl group or a halogen substituted C1-C4 alkyl group; y2 is a substituted or unsubstituted C1-C6 alkyl group, and a substituent group is selected from a halogen atom and a C1-C3 alkoxy group. The compound provided by the invention can inhibit one or more of JAK3, TAK1 and ITK, and is further applied to preparation of drugs for treating diseases caused by abnormal activation of one or more kinases.

Description

Technical Field

[0001] This invention relates to the field of chemical and pharmaceutical technology. More specifically, this invention relates to a pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof and its applications. Background Technology

[0002] Autoimmune diseases (such as rheumatoid arthritis, psoriatic arthritis, ulcerative colitis, Crohn's disease, alopecia areata, atopic dermatitis, multiple sclerosis, and systemic sclerosis) and some hematologic malignancies (such as peripheral T-cell lymphoma and adult T-cell leukemia) are a group of diseases that seriously threaten human health. Their pathogenesis is complex, and the core is closely related to immune system dysfunction and abnormal activation of inflammatory signaling pathways. These diseases not only lead to organ dysfunction and decreased quality of life, but some severe cases can also be life-threatening. Although currently used clinical treatments can alleviate symptoms to some extent, they generally have limited efficacy, are prone to drug resistance, and have significant side effects. For example, the broad-spectrum inhibitory effect of traditional immunosuppressants may lead to an increased risk of infection, and some targeted drugs may cause adverse reactions such as anemia and dyslipidemia due to insufficient selectivity. Therefore, there is an urgent need to develop new therapeutic drugs with clear mechanisms of action, strong targeting, and high safety.

[0003] In the network of immune regulation and inflammatory response, several key signaling molecules have become core targets for disease treatment. Among them, Janus Kinase 3 (JAK3), Transforming Growth Factor-β Activated Kinase 1 (TAK1), and Interleukin-2 Inducible Tyrosine Kinase (ITK) occupy crucial node positions.

[0004] JAK3 belongs to the non-receptor tyrosine protein kinase JAK family. Unlike other members of the family (JAK1, JAK2, TYK2) which are widely distributed and have multi-cytokine activation characteristics, JAK3 has high specificity: it is only activated by γ-chain cytokines (IL-2, IL-4, IL-7, IL-9, IL-15, IL-21) and is specifically expressed in T lymphocytes, B lymphocytes, natural killer (NK) cells, bone marrow cells, and thymocytes. Its activation process only induces JAK1 to form a dimer with it, which activates downstream signals through cross-tyrosine phosphorylation, thereby catalyzing the phosphorylation of receptor tyrosine residues, recruiting and activating signal transduction and transcription activators (STATs), and ultimately regulating gene transcription, participating in key immune processes such as leukocyte maturation and activation, cytokine and immunoglobulin production, and lymphocyte growth and development.

[0005] As a central switch in the autoimmune inflammatory network, TAK1 plays a key regulatory role in inflammatory signal transduction. When stimulated by danger signals such as TNF-α, IL-1β, or TLR ligands, TAK1 is rapidly activated via K63 ubiquitination, subsequently phosphorylating downstream key nodes such as NF-κB, JNK, p38, and ERK, driving the transcription of pro-inflammatory cytokines such as IL-6, IL-17, and TNF-α, as well as matrix metalloproteinases, forming a positive feedback loop that amplifies inflammation. In rheumatoid arthritis, TAK1 is highly activated in synovial fibroblasts and macrophages, promoting angiogenesis, synovial hyperplasia, and cartilage and bone erosion. In multiple sclerosis, its activation is a crucial "gatekeeper" for peripheral immune cells entering the central nervous system. Furthermore, TAK1 participates in the survival and proliferation of T and B lymphocytes, which is essential for maintaining immune tolerance.

[0006] ITK, a member of the Tec kinase family, is specifically anchored on the inner membrane of T cells, NK cells, and mast cells, acting as a "signal amplifier." At the physiological level, after TCR-CD3 recognizes an MHC peptide, Lck phosphorylates ITK and localizes it to the LAT-SLP-76 signaling backbone, subsequently phosphorylating PLC-γ1, triggering calcium flux, PKC activation, and nuclear translocation of NFAT and NF-κB. This drives the secretion of cytokines such as IL-2, IFN-γ, and IL-17, and regulates immune synapse formation. Simultaneously, ITK participates in Th2 / Th17 cell differentiation through allosteric reactions, maintaining humoral immune homeostasis. Abnormal ITK function is closely related to various diseases: its deficiency can lead to Th2 / Th17 immunodeficiency, while upregulation of expression / activity is associated with the development and progression of autoimmune diseases such as allergic asthma, atopic dermatitis, and systemic lupus erythematosus, as well as tumors such as peripheral T-cell lymphoma.

[0007] Further research has confirmed that the three core signaling pathways, including JAK3, TAK1, and ITK, exhibit clear parallel and highly active synergistic pathogenic characteristics in autoimmune diseases and related tumors such as rheumatoid arthritis, psoriasis, multiple sclerosis, and peripheral T-cell lymphoma: JAK3 maintains the survival and proliferation of T / B / NK cells through γc chain cytokine signaling, providing the cellular basis for abnormal immune responses; TAK1, as a central switch of the inflammatory network, amplifies the innate immune inflammatory response by activating the NF-κB and MAPK pathways, forming a cytokine storm; and ITK, as a T-cell receptor signaling-specific kinase, regulates Th2 / Th17 differentiation and cytokine secretion, determining the initiation intensity of the immune response. Therefore, developing triple inhibitors of JAK3, TAK1, and ITK can achieve progressive complementarity at the "cytokine-receptor-transcriptional" level: JAK3 interrupts cytokine signaling upstream, ITK inhibits pathogenic T cell differentiation and migration in the middle, and TAK1 suppresses cytokine storms downstream. This allows for the attainment of more durable immunomodulators at extremely low doses and reduces drug resistance and toxicity caused by compensatory upregulation of single targets. Thus, developing novel, metabolically stable triple inhibitors that specifically inhibit JAK3, TAK1, and ITK can provide a highly efficient and complementary intervention strategy for the treatment of autoimmune diseases. Summary of the Invention

[0008] One object of the present invention is to provide a pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof that can simultaneously inhibit JAK3, TAK1 and ITK, and thus be used in the preparation of drugs for diseases caused by abnormal activation of the above three kinases.

[0009] To achieve these objects and other advantages of the present invention, according to one aspect of the present invention, the present invention provides pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof, the general formula of which is shown below; Wherein, R1 is a 5- or 6-membered heterocyclic group of substituted or unsubstituted phenyl or heteroatoms of oxygen, nitrogen and sulfur, and the substituent is selected from halogen atom, C1-C6 alkyl, C1-C3 alkoxy, cyano, nitro; X is selected from , , , , , , , , , , ; Y1 is a C2-C5 alkenyl, C2-C5 alkynyl, or halogen-substituted C1-C4 alkyl; Y2 is a substituted or unsubstituted C1-C6 alkyl group, and the substituent is selected from halogen atoms and C1-C3 alkoxy groups.

[0010] According to another aspect of the invention, the use of pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof is also provided for the preparation of medicaments that inhibit at least one of JAK3, TAK1, and ITK.

[0011] According to another aspect of the invention, the use of pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof is also provided for the preparation of medicaments for treating diseases caused by JAK3, TAK1, and ITK.

[0012] The present invention has at least the following beneficial effects: The pyrrolopyrimidine derivatives or pharmaceutically acceptable salts of the present invention, through precise structural design, have their respective groups synergistically adapted to the binding pockets of JAK3, TAK1, and ITK targets, exhibiting extremely strong targeting. In vitro experiments have confirmed that the compounds have an inhibitory activity against JAK3 reaching IC50. 50 <1 nM, it also has good inhibitory effects on ITK and TAK1, and exhibits excellent hepatic microsomal metabolic stability (half-life >40 minutes). In vivo studies on atopic dermatitis show that the representative compound 2a has significantly better therapeutic effects than the marketed drugs litexitinib and torvatinib, effectively reducing epidermal thickness and mast cell infiltration. This class of compounds addresses the limitations of existing drugs in terms of limited efficacy, drug resistance, and significant side effects, providing a novel treatment option with a clear mechanism of action and high safety for autoimmune diseases and related hematological malignancies, with broad prospects for clinical application.

[0013] Other advantages, objectives and features of the present invention will become apparent in part from the following description, and in part from those skilled in the art through study and practice of the invention. Attached Figure Description

[0014] Figure 1 The therapeutic effect of compound 2a on a mouse model of DNCB-induced atopic dermatitis (AD) is shown in Figure A: Representative graphs of H&E and TB staining in skin and ear tissues. Figure B: Statistical graphs of H&E and TB staining in skin and ear tissues. Comparison with the model group (model)*** p <0.001, **** p <0.0001; compared to the normal group ### p <0.001. Detailed Implementation

[0015] The present invention will now be described in further detail so that those skilled in the art can implement it based on the description.

[0016] It should be understood that terms such as "having," "comprising," and "including" used in the embodiments of this application do not exclude the presence or addition of one or more other elements or combinations thereof. All directional indications (such as up, down, left, right, front, back, etc.) in the embodiments of this application are only used to explain the relative positional relationship and movement of components in a specific posture. If the specific posture changes, the directional indication will also change accordingly. When an element is referred to as "fixed to" or "set on" another element, it can be directly on the other element or may have an intervening element present. When an element is referred to as "connected to" another element, it can be directly connected to the other element or indirectly connected to the other element through an intervening element. Descriptions involving "first," "second," etc., in the embodiments of this application are for descriptive purposes only and should not be construed as indicating or implying their relative importance or implicitly specifying the number of indicated technical features. Therefore, a feature defined with "first" or "second" may explicitly or implicitly include at least one of those features.

[0017] It should be noted that the technical solutions of the various embodiments of this application can be combined with each other, but only if they are based on the ability of those skilled in the art to implement them. When the combination of technical solutions is contradictory or cannot be implemented, it should be considered that such combination of technical solutions does not exist and is not within the scope of protection claimed by this application.

[0018] Embodiments of this application provide pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof, the general formula of which is shown below; Wherein, R1 is a 5- or 6-membered heterocyclic group of substituted or unsubstituted phenyl or heteroatoms of oxygen, nitrogen and sulfur, and the substituent is selected from halogen atom, C1-C6 alkyl, C1-C3 alkoxy, cyano, nitro; X is selected from , , , , , , , , , , ; Y1 is a C2-C5 alkenyl, C2-C5 alkynyl, or halogen-substituted C1-C4 alkyl; Y2 is a substituted or unsubstituted C1-C6 alkyl group, and the substituent is selected from halogen atoms and C1-C3 alkoxy groups.

[0019] For example, R1, as the hydrophobic end functional group of the pyrrolopyrimidine core skeleton, has a structure that synergistically regulates the binding efficiency of the molecule to the target through electronic and steric effects, adapting to the active center pocket characteristics of JAK3, TAK1, and ITK: the unsubstituted phenyl is the basic hydrophobic skeleton, and the conjugated aromatic structure can form a stable π-π stacking interaction with the hydrophobic region of the target protein. Among substituted phenyl groups, halogen substitution (4-fluorophenyl, 3-chlorophenyl, 2-bromophenyl) modulates the electron cloud density of the benzene ring by leveraging the electronegativity of the halogen atom. Its small size does not increase steric hindrance. The strong electronegativity of 4-fluorophenyl can enhance the interaction between the molecule and the target polar residues. C1-C6 alkyl substitution (4-methylphenyl, 3-ethylphenyl) improves the binding stability of the molecule with the target hydrophobic pocket by increasing hydrophobicity. Long-chain alkyl groups (such as 3-propylphenyl) can be adapted to the target hydrophobic channels. The electron-donating effect of C1-C3 alkoxy substitution (4-methoxyphenyl, 3-ethoxyphenyl) can optimize the electron distribution of the benzene ring and form favorable interactions with the target electropositive amino acid residues. The strong electron-withdrawing properties of cyano (4-cyanophenyl) and nitro (3-nitrophenyl) can form specific interactions with the target hydrogen bond donor, significantly improving the binding affinity. In 5 / 6-membered heterocyclic groups (furan-2-yl, furan-3-yl, thiophene-3-yl, pyridin-4-yl) containing oxygen, nitrogen, and sulfur heteroatoms, the lone pair electrons of the heteroatom can form hydrogen bonds with the target site. The rigid structure of the heterocycle fixes the molecular conformation, while the introduction of polar sites balances lipophilicity and water solubility. The electronegativity of the oxygen atom works synergistically with the hydrophobic structure of the heterocycle to adapt to the polar-hydrophobic mixed binding region of the target site.

[0020] X is a key bridging bond connecting the pyrrolopyrimidine core skeleton and the central nitrogen atom (tertiary amine), directly affecting the spatial distance between the core skeleton and the central nitrogen atom, and thus regulating the compatibility of Y1 and Y2 with the target site through their respective connecting chains. The optional structures of X are four-membered / five-membered / six-membered / seven-membered ring systems containing N / O heteroatoms, all of which are directly bonded to the central nitrogen atom through single bonds, as follows: N-containing four-membered ring (azacyclobutane): The four-membered ring has extremely high rigidity, which can fix the relative spatial conformation of the core skeleton and the central nitrogen atom to the maximum extent, avoiding target site binding disorder caused by excessive molecular flexibility, and adapting to the spatially compact target active center; the lone pair electrons of the N atom on the ring (which is not directly bonded to the central nitrogen atom) can enhance the electron cloud density of the ring system, indirectly increasing the electrostatic interaction strength between the central nitrogen atom and the acidic residues of the target. Oxygen-containing four-membered ring (oxobutane): The strong electronegativity of the oxygen atom can efficiently transfer the electronic effect between the core skeleton and the central nitrogen atom, enhancing the polar interaction between the molecule and the target. The rigid structure of the four-membered ring provides stable hydrophobic support, precisely restricting the spatial orientation of the Y1 and Y2 side chains, ensuring their fit with the target pocket. N-containing six-membered ring (piperidine ring): The rigid structure of the six-membered ring can fix the relative spatial position of the core skeleton and the central nitrogen atom, avoiding conformational disorder of the target binding caused by excessive molecular flexibility. The N atom on the ring (not directly bonded to the central nitrogen atom) can help regulate the electron cloud density of the ring, indirectly enhancing the electrostatic interaction between the central nitrogen atom and the acidic residues of the target. The spatial orientation (cis / trans) of the substitution sites on the ring can precisely control the fitting angle between the Y1 and Y2 side chains and the target binding pocket. Oxygen-connected cyclohexyl group: This structure uses the oxygen atom as a connecting bridge, with one end bonded to the pyrrolopyrimidine core skeleton, and the other end of the cyclohexyl group directly connected to the central nitrogen atom by a single bond. The electronegativity of the oxygen atom optimizes electron transfer between the core framework and the central nitrogen atom, enhancing the polar interaction between the molecule and the target. The saturated six-membered ring structure of the cyclohexyl group provides stable hydrophobic support, and cis-trans isomerism allows for precise control of the spatial orientation of the side chains, enabling Y1 and Y2 to fit the target pocket. X can also be selected from cyclopentyl groups linked by oxygen atoms, oxygen-containing six-membered rings, nitrogen-containing five-membered rings, and nitrogen-containing seven-membered rings.

[0021] Y1 is a C2-C5 alkenyl, C2-C5 alkynyl, or halogen-substituted C1-C4 alkyl group, indirectly connected to the central nitrogen atom through corresponding linking structures. Its structural characteristics directly affect the binding of the side chain to the target hydrophobic sub-pocket: C2-C5 alkenyl (vinyl, propenyl, pentenyl): The carbon-carbon double bond has a certain rigidity, which can restrict the free rotation of the acyl chain (carbonyl + alkenyl), allowing the acrylamide structure in which Y1 is located to be precisely embedded into the target hydrophobic sub-pocket; after the carbon chain length extends from C2 to C5, it can adapt to the target hydrophobic channel at different depths. Among them, the long chain structure of pentenyl can form a stronger hydrophobic interaction with the deep hydrophobic region of the target. At the same time, the π electron cloud of the double bond can bind to the target aromatic amino acid residues through π-π stacking, balancing steric hindrance and binding efficiency, and improving binding stability. C2-C5 alkynyl groups (ethynyl, propynyl, pentylyyl): have a stronger linear rigid structure, and the electron cloud density of the carbon-carbon triple bond is higher than that of the double bond. The "-CO-C≡C-" structure formed with the carbonyl group can form a stronger π-π stacking interaction with the aromatic amino acid residues of the target. The linear long chain of C5 alkynyl can further reduce steric hindrance, making it easier for the acyl chain to penetrate into the deep hydrophobic region of the target. At the same time, the long chain structure increases the contact area between the molecule and the target, significantly enhancing the binding specificity. Halogen-substituted C1-C4 alkyl groups (fluoromethyl, chloroethyl, trifluoropropyl, etc.): The strong electronegativity of halogen atoms (fluorine, chlorine, etc.) can regulate the electron cloud distribution of the side chain and form specific interactions with the polar residues of the target. The hydrophobic skeleton of C1-C4 alkyl groups can enhance the binding stability of the molecule with the hydrophobic subpocket of the target. Among them, polyhalogen substituents such as trifluoromethyl can simultaneously optimize the lipophilic and water solubility of the molecule and improve the ability to penetrate biological membranes, while monohalogen substituents such as chloroethyl can maintain hydrophobicity and precisely regulate the fitting angle between the side chain and the target pocket through the spatial effect of halogen atoms, further improving the binding selectivity.

[0022] Y2 represents substituted or unsubstituted C1-C6 alkyl groups, indirectly connected to the central nitrogen atom via a carbonyl-methylene-alkyl structure. Its structure primarily regulates the lipophilic and hydrophilic properties of the molecule, influencing in vivo absorption and target binding specificity: Unsubstituted C1-C6 alkyl groups (methyl, ethyl, butyl): Methyl groups exhibit minimal steric hindrance, moderately increasing hydrophobicity without affecting molecular conformation; the combination of amino and methyl groups further enhances water solubility. Long-chain alkyl groups (butyl, hexyl) enhance hydrophobic interactions, suitable for scenarios where long-chain hydrophobic channels exist within the target binding pocket. Halogen-substituted C1-C6 alkyl groups: The strong hydrophobicity and electronegativity of trifluoromethyl groups simultaneously optimize lipophilic and electronic properties, enhancing the molecule's ability to cross biological membranes; chloromethyl groups combine the electronegativity of chlorine atoms with the hydrophobicity of methyl groups, balancing molecular polarity and lipophilicity, further optimizing drug distribution and target binding efficiency in vivo. C1-C3 alkoxy-substituted C1-C6 alkyl: The electron-donating effect of alkoxy groups increases molecular polarity and improves water solubility; the oxygen atom of the methoxy group can form hydrogen bonds with the target site; the steric effect brought about by carbon chain elongation can further optimize the compatibility between the side chain where Y2 is located and the polar site of the target, and improve the binding specificity.

[0023] The central nitrogen atom (tertiary amine nitrogen atom) is the linking hub of the molecule, directly bonding to X: the lone pair electrons of the nitrogen atom can form strong electrostatic interactions or hydrogen bonds with the acidic amino acid residues in the active centers of JAK3, TAK1, and ITK, significantly enhancing the binding affinity. The basicity of the tertiary amine can precisely regulate the pKa value of the molecule, causing the compound to be partially protonated at physiological pH (7.4), which improves water solubility (facilitating intestinal absorption or distribution in body fluids) while retaining a certain degree of lipid solubility (facilitating entry into target cells across biological membranes). The nitrogen atom is sp3 hybridized, and after directly connecting to X, it drives Y1 and Y2 to form a tetrahedral spatial conformation, fixing the relative positions of R1 (core backbone end) and Y1 and Y2 (side chain ends), avoiding the loss of activity due to conformational isomerism when the molecule binds to the target site.

[0024] Embodiments of this application also provide the use of pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof for the preparation of medicaments that inhibit at least one of JAK3, TAK1, and ITK.

[0025] For example, given the synergistic pathogenicity of the JAK3, TAK1, and ITK signaling pathways, compound capsules can be prepared by selecting compounds that have inhibitory activity against all three kinases and combining them with appropriate excipients to achieve synergistic inhibition of multiple targets.

[0026] Excipient composition: The filler is microcrystalline cellulose (40%-60%), which has both filling and binding functions; the disintegrant is sodium carboxymethyl starch (5%-8%), which promotes the disintegration of the capsule contents in vivo; the lubricant is talc (1%-2%), which improves the flowability of the powder.

[0027] Preparation steps: ① Place the active ingredient (15%-25%), filler, and disintegrant in a three-dimensional mixer and mix for 15 minutes until homogeneous; ② Add lubricant and continue mixing for 5 minutes; ③ Use a fully automatic capsule filling machine to fill the mixed powder into No. 0 empty capsules, each containing 20-40mg of active ingredient; ④ After polishing the capsules, package them with aluminum-plastic blister packs to obtain the final product.

[0028] Embodiments of this application also provide the use of pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof for the preparation of medicaments for treating diseases caused by JAK3, TAK1, and ITK.

[0029] For example, using sucrose pellet core as a carrier, two or three active ingredients and binders are alternately coated on the surface of the pellet core using a lamination method, and then a sustained-release coating layer (ethyl cellulose, accounting for 10%-15%) is coated on it to control the drug release rate and achieve synchronous peak blood drug concentrations and long-term maintenance of the three components.

[0030] The auxiliary materials consist of: a polyvinyl ketone aqueous solution as the binder; a slow-release coating solution containing ethyl cellulose, polyethylene glycol 400 as the plasticizer, and ethanol as the solvent; and microcrystalline cellulose as the filler.

[0031] Preparation steps: ① Place the sucrose pellet core in a fluidized bed coating machine, spray in a povidone aqueous solution as a binder, and simultaneously sprinkle in a mixture of active ingredients and microcrystalline cellulose powder for layer coating; ② Spray in a sustained-release coating solution; ③ After drying, sieve micro-pellets of 150-300μm, fill them into capsules, each containing 10-20mg of each active ingredient, and the product is obtained.

[0032] The diseases described in this embodiment may be one or more of the following: rheumatoid arthritis, psoriatic arthritis, ulcerative colitis, Crohn's disease, alopecia areata, atopic dermatitis, multiple sclerosis, systemic sclerosis, peripheral T-cell lymphoma, adult T-cell leukemia, and ankylosing spondylitis.

[0033] The following is a description of a specific embodiment.

[0034] Tables 1-5 list typical pyrrolopyrimidine derivatives.

[0035] Table 1 Table 2 Table 3 Table 4 Table 5 Example 1: Method for preparing pyrrolopyrimidine derivatives or pharmaceutically acceptable salts thereof To facilitate the description of the synthetic routes and methods in the examples later, the abbreviations of the raw materials or reagents used are listed in the table below.

[0036] Table 6 Reagents and Abbreviations The detailed instructions are as follows: At room temperature, intermediate 1 (1.0 eq), DIEA (1.5 eq), (1r,4r)-4-hydroxycyclohexyl)carbamate tert-butyl ester (1.2 eq), and an appropriate amount of n-BuOH as solvent were added to the reaction flask. The mixture was heated to 80 °C and stirred for 3 h, then the reaction was stopped. The reaction solution was extracted with EA / H2O, the oil layer was collected, the oil layer was washed several times with saturated brine, dried over anhydrous sodium sulfate, the liquid was collected by filtration, the oil layer was evaporated, and intermediate 3 was purified by normal-phase silica gel column chromatography, which yielded a yellow oil.

[0037] At room temperature, intermediate 3 (1.0 eq), boric acid derivative (1.1 eq), Pd(PPh3)4 (0.05 eq), and K2CO3 (3.0 eq) were added to a three-necked flask. After purging the air in the flask multiple times with N2, a mixed solvent of 1,4-dioxane / H2O (7 / 3, v / v) was injected into the flask using a syringe. The mixture was heated under reflux for approximately 5 h. Insoluble matter was removed by diatomaceous earth filtration, and the filtrate was collected and removed under reduced pressure. The filtrate was extracted with EtOAc and water, and the oil layer was collected. The oil layer was then washed three times with saturated brine, dried over anhydrous Na2SO4, filtered, and the liquid was collected. The oil layer was then evaporated to dryness, and product 4 was purified using a normal-phase silica gel column chromatography.

[0038] At room temperature, intermediate 4 (1.0 eq), TBAF3·H2O (6.0 eq), and an appropriate amount of DMF were added to the reaction flask. The temperature was raised to 75°C and stirred. The reaction was stopped after TLC monitoring showed completion. The reaction solution was extracted with EA / H2O, the oil layer was collected, and the oil layer was washed several times with saturated brine. After drying with anhydrous sodium sulfate, the liquid was collected by filtration, and the oil layer was concentrated under reduced pressure to obtain the intermediate after the SEM protecting group was removed. The intermediate was then dissolved with EA, and a saturated HCl solution of EA was added. The mixture was stirred overnight, and the deprotection of the starting material was monitored by TLC. The solvent was removed by concentration under reduced pressure, and the solid was collected to obtain intermediate 5.

[0039] At room temperature, add 1.0 eq of medium 5 and an appropriate amount of dry DMF to the reaction flask, then add 5.0 eq of DIEA and 1.0 eq of halogenated reagent dropwise, and react overnight at room temperature. Extract the reaction solution with DCM and water, collect the oil layer, wash the oil layer three times with saturated brine, dry the oil layer with anhydrous Na2SO4, filter and collect the liquid, concentrate the oil layer under reduced pressure, and purify and collect product 7 using a normal-phase silica gel column.

[0040] At room temperature, add 1.0 eq of medium-based 6 and an appropriate amount of dried DMF to the reaction flask, followed by dropwise addition of DIEA (2.0 eq) and an acyl chloride derivative (1.0 eq). React overnight at room temperature. Extract the reaction solution with DCM and water, collect the oil layer, wash the oil layer three times with saturated brine, dry the oil layer with anhydrous Na2SO4, filter and collect the liquid, concentrate the oil layer under reduced pressure, and purify and collect product 7 using a normal-phase silica gel column.

[0041] Wherein, R1 is a 5- or 6-membered heterocyclic group of substituted or unsubstituted phenyl or heteroatoms of oxygen, nitrogen and sulfur, and the substituent is selected from halogen atom, C1-C6 alkyl, C1-C3 alkoxy, cyano, nitro; R2 is Y2 is a substituted or unsubstituted C1-C6 alkyl group, and the substituent is selected from halogen atoms and C1-C3 alkoxy groups.

[0042] Structural identification data for some compounds are shown in Table 7.

[0043] Table 7 Table 8 Table 9 Example 2: In vitro biochemical inhibition of JAK kinase activity experiment Materials: JAK3, ITK, and TAK1 kinases (Carna); peptides FAM-P22 and FAM-P30 (GL Biochem); ATP, DMSO, and EDTA (Sigma); 96-well plates (Corning).

[0044] method: 1. Prepare an alkaline buffer and a termination buffer containing 1x kinase.

[0045] 1) 1x kinase base buffer contains 50 Mm HEPES, pH 7.5; 0.0015% Brij-35; 10 Mm MgCl2; 2 Mm DTT.

[0046] 2) The termination buffer contains 100 Mm HEPES, pH 7.5; 0.015% Brij-35; 0.2% CoatingReagent #3; and 50 Mm EDTA.

[0047] 2. Preparation of compounds.

[0048] 1) Prepare the test compound to 50 times the highest test concentration using 100% DMSO. Transfer 100 µL of the compound dilution into the well plate.

[0049] 2) Add 100 µL of 100% DMSO to two wells and designate this plate as the original plate. Transfer 10 µL of the compound from the original plate to a new 96-well plate and designate this plate as the intermediate plate. Add 90 µL of 1x kinase base buffer to each well of the intermediate plate and place the plate on a shaker to mix the compound solution with the 1x kinase base buffer thoroughly. Then, take 5 µL of the mixture from each well of the intermediate plate and transfer it to a 384-well plate to form replicates. Designate this plate as the detection plate.

[0050] 3. Enzyme reaction.

[0051] 1) Prepare a 2.5x enzyme solution and add the kinase to a 1x kinase base buffer.

[0052] 2) Prepare a 2.5x peptide buffer and add the FAM-labeled peptide and ATP to a 1x kinase base buffer.

[0053] 3) Transfer the 2.5x enzyme solution to the test plate. Each well of the test plate contains 5 µL of compound solution containing 10% DMSO, followed by 10 µL of 2.5x enzyme solution, and incubate at 25 °C for 10 min.

[0054] 4) Add 2.5x peptide solution to each well of the detection plate, incubate at 28 °C for an appropriate time, and then add 25 µL of stop buffer to terminate the enzyme reaction.

[0055] 4. Read and record the raw data for each well, and perform the corresponding transformations on the raw data.

[0056] 1) Inhibition rate = (maximum value - compound conversion value) / (maximum value - minimum value) * 100, where the maximum value is the data of the DMSO control group, and the minimum value is the blank value without enzyme addition. 2) Calculate the half-maximal inhibitory concentration (IC50). 50 The values ​​were plotted with log[drug concentration] on the x-axis and inhibition rate on the y-axis. A dose-response curve was fitted in Graphpad Prism 5 to obtain the drug concentration at 50% inhibition, which is the IC50 of this compound at the kinase level. 50value.

[0057] The table below (Table 4) provides the average IC50 values ​​for the compounds in Table 1 with respect to JAK3 and TAK1. 50 Range, where "A" represents IC 50 Values ​​less than 1 nM, "B" indicates IC 50 Values ​​between 1 nM and 10 nM, where "C" indicates IC 50 Values ​​between 10 nM and 100 nM, where "D" indicates IC 50 The value is between 100 nM and 1000 nM.

[0058] Table 10 Example 3: In vitro liver microsomal enzyme metabolic stability test The compounds were reacted with human liver microsomes in an NADPH incubation system. The reaction was terminated at 0, 5, 15, 30, 45, 60, and 90 min. The supernatant was extracted by low-temperature centrifugation, and the residual amount of the compounds in the reaction solution was analyzed by LC-MS / MS. The half-life (t) of the corresponding compounds was calculated by plotting the drug-time curve. 1 / 2 = 0.693 / curve slope) and clearance rate (Cl int =0.693 / t 1 / 2 ) Table 11 Example 4: In vivo anti-atopic dermatitis treatment study of representative compounds Animals: Six-week-old female BALB / c mice were acclimatized for one week. Hair removal cream was applied to a 4 cm section of the mouse's back. 2 Remove the hair and weigh the mice at their initial weight.

[0059] Sensitization phase of AD model: 2,4-dinitrochlorobenzene (DNCB) powder was diluted to 1% in acetone and olive oil (3:1, v:v). On days 1 and 4, 200 µL and 20 µL of 1% DNCB were applied to the back and left and right ear skin of mice, respectively, while normal mice were treated with the corresponding volume of solvent.

[0060] The attack phase of the AD model: Starting from day 7, 0.4% DNCB was applied to the back and left and right ears of mice every 2 days to maintain AD symptoms, while normal mice were applied with the corresponding volume of solvent.

[0061] Treatment grouping: On day 7, the dorsal skin fraction and ear thickness of each mouse were assessed using the scoring method, and mice were randomly assigned to groups based on the average values. In addition to the normal control group and the model group, the other treatment groups were rituximab (30 mg / kg, PO), tovatinib (30 mg / kg, PO), compound (2a 30 mg / kg, PO), compound (2a 30 mg / kg, IP), and compound (2a 50 mg / kg, PO), respectively, for a total of 21 days. Mouse weight and skin lesion manifestations were recorded every 3 days during this period.

[0062] Histopathological studies: Skin from the back and ears of mice was fixed in 4% formalin solution, embedded in paraffin, sectioned, and stained with hematoxylin and eosin (H&E) to study epidermal thickness. Toluidine blue (TB) staining was used to detect cell infiltration and mast cell number.

[0063] Figure 1 This is a visualization of the therapeutic effect of compound 2a on a DNCB-induced AD mouse model, divided into a staining representative plot (Part A) and a quantitative statistical plot (Part B): Part A contains four parallel sub-figures, corresponding to "Skin H&E staining," "Skin TB staining," "Ear H&E staining," and "Ear TB staining," respectively. Each sub-figure shows a comparison of pathological sections from seven groups of experimental samples. The groups, from left to right, are: Normal group: healthy mice without AD induction and without drug administration; Model group: mice with AD induction and without drug administration; Positive drug control group 1 (ritle. 30 mg / kg PO): mice induced with AD and orally administered 30 mg / kg ritexitinib (a marketed JAK inhibitor); Positive drug control group 2 (tofa. 30 mg / kg PO): mice induced with AD and orally administered 30 mg / kg tolvatinib (a marketed JAK inhibitor); Compound 2a treatment group 1 (2a 30 mg / kg PO): mice induced with AD and orally administered 30 mg / kg compound 2a; Compound 2a treatment group 2 (2a 30 mg / kg IP): mice induced with AD and intraperitoneally injected with 30 mg / kg compound 2a; Compound 2a treatment group 3 (2a 50 mg / kg IP); PO): After AD induction, administer compound 2a orally at a dose of 50 mg / kg.

[0064] Normal group: The epidermis is thin and has a uniform structure, and there is no obvious inflammatory cell infiltration in the dermis; Model group: The epidermis is significantly thickened (about 2-3 times that of the normal group), the tissue is disordered, and a large number of inflammatory cells are visible in the dermis (consistent with AD pathological characteristics); Treatment group: All drug administration groups can reduce the thickness of the epidermis to varying degrees, among which the 2a 50mg / kg PO group and the 2a 30mg / kg IP group have the most significant effects, with the epidermal thickness approaching that of the normal group and the tissue integrity restored better, which is superior to the litexitinib and tolvatinib groups.

[0065] Normal group: Only a small number of scattered mast cells were observed in the skin tissue; Model group: A large number of mast cells were aggregated, indicating a severe inflammatory response; Treatment group: The number of mast cells in each treatment group was significantly reduced in 2a, especially in the 2a 50mg / kg PO group, where the mast cell density was close to that of the normal group. Although there was improvement in the litexitinib and tovatinib groups, the effect was weaker than that in the 2a high-dose and intraperitoneal injection groups.

[0066] Normal group: The ear epidermis is extremely thin and the structure is clear; Model group: The ear epidermis is significantly thickened, and even hyperkeratosis and tissue edema are observed; Treatment group: All treatment groups in 2a can effectively reduce the thickness of the ear epidermis. The 30mg / kg IP group and 50mg / kg PO group in 2a have the best effects, and the epidermal structure is restored to a regular state, which is better than the positive drug control group.

[0067] Normal group: Very few mast cells in ear tissue; Model group: Mast cells are densely distributed, indicating severe ear inflammation; Treatment group: The number of mast cells in the 2a treatment group was significantly reduced, with the 2a 50mg / kg PO group showing the most significant improvement, while the mast cell clearance effect in the positive drug control group was relatively weak.

[0068] Part B contains the quantitative statistical results of the pathological indicators in Part A, including four independent bar charts, corresponding to “skin epidermal thickness statistics”, “skin mast cell count statistics”, “ear epidermal thickness statistics”, and “ear mast cell count statistics”.

[0069] For the epidermal thickness statistical graph, the horizontal axis represents the experimental groups (from left to right: normal, model, rule 30 mg / kg PO, tofa 30 mg / kg PO, 2a 30 mg / kg PO, 2a 30 mg / kg IP, 2a 50 mg / kg PO); the vertical axis represents the quantitative value of epidermal thickness (units are arbitrary relative values, the larger the value, the thicker the epidermis). The ordinate values ​​of the model group were significantly higher than those of the normal group (###p<0.001, compared with the normal group), proving that the AD model was successfully constructed. The ordinate values ​​of all treatment groups were lower than those of the model group (***p<0.001, ****p<0.0001, compared with the model group). Among them, the ordinate values ​​of the 2a 50mg / kg PO group and the 2a 30mg / kg IP group were the lowest (close to the normal group). The 2a 30mg / kg PO group was superior to the litexitinib and tolvatinib groups. Compound 2a can reduce the thickness of the skin epidermis of AD mice in a dose-dependent (50mg / kg PO is better than 30mg / kg PO) and route-dependent (30mg / kg IP is better than 30mg / kg PO) manner. High-dose oral and intraperitoneal injection have the best effects.

[0070] For the statistical chart of mast cell count in the skin, the horizontal axis is exactly the same as the "Statistical Chart of Epidermal Thickness in the Skin"; the vertical axis is the quantitative value of mast cell count (the larger the value, the more severe the inflammation). The ordinate values ​​of the model group were significantly higher than those of the normal group (###p<0.001), indicating a large amount of mast cell infiltration. The ordinate values ​​of all treatment groups were significantly lower than those of the model group (***p<0.001), with the 2a 50mg / kg PO group having the lowest ordinate value, followed by the 2a 30mg / kg IP group, both of which were superior to the litexitinib and torvatinib groups. Compound 2a can effectively inhibit mast cell infiltration in the skin of AD mice, and its effect in clearing the core inflammatory markers is better than that of marketed positive control drugs.

[0071] For the ear skin thickness chart, the horizontal axis is: [as described in the above statistics] Figure 1 The vertical axis represents the quantified value of ear skin thickness (a larger value indicates more severe ear inflammation). The vertical axis value of the model group was significantly higher than that of the normal group (###p<0.001), indicating significant ear skin thickening. The vertical axis values ​​of all treatment groups in 2a were significantly lower than those of the model group (***p<0.001). Among them, the 30mg / kg IP group and the 50mg / kg PO group of 2a showed the most significant effects, with vertical axis values ​​close to those of the normal group. The litexitinib and torvatinib groups showed smaller improvements. Compound 2a showed outstanding relief of ear inflammation in AD mice and could effectively reverse the pathological changes of ear skin thickening.

[0072] For the statistical chart of mast cell count in the ear, the horizontal axis is: compared to the above statistics. Figure 1The vertical axis represents the quantitative value of mast cell count in the ear (a larger value indicates more severe ear inflammation); the trend shows that the vertical axis value of the model group is significantly higher than that of the normal group (###p<0.001), indicating a high density of mast cells in the ear; the vertical axis values ​​of all treatment groups in -2a are significantly lower than those of the model group (***p<0.001), with the lowest values ​​in the 2a 50mg / kg PO group and the 30mg / kg IP group, and the inhibitory effects of litexitinib and torvatinib groups are weaker; compound 2a can effectively clear mast cells in the ear tissue of AD mice, inhibit inflammation in multiple sites, and its effect is superior to that of marketed drugs.

[0073] In summary, the pyrrolopyrimidine derivatives disclosed in this application utilize pyrrolopyrimidine as the core skeleton. R1 (substituted / unsubstituted phenyl or 5-6 membered heterocyclic group containing O / N / S heteroatoms) regulates the molecular electronic properties and hydrophobic interactions. X (containing N / O heterocyclic group or oxygen-linked alicyclic group) fixes the spatial conformation of the core skeleton and the central tertiary amine. Y1 (C2-C4 alkenyl / alkynyl, halogen-substituted C1-C4 alkyl) and Y2 (substituted / unsubstituted C1-C6 alkyl) synergistically optimize target binding specificity and in vivo physicochemical properties. The central tertiary amine acts as a connecting hub, anchoring the binding site through electrostatic interactions or hydrogen bonds with target acidic amino acid residues, while simultaneously regulating molecular water solubility and metabolic stability. These groups synergistically endow the compounds with specific binding ability to the target kinase. Experimentally, in vitro biochemical experiments confirmed that all compounds exhibited potent inhibitory activity against JAK3 (IC50). 50 <1 nM), it also exhibits good inhibitory effects on TAK1 and ITK, and demonstrates excellent metabolic stability in liver microsomes (half-lives mostly >40 minutes). In vivo studies on atopic dermatitis show that the representative compound 2a can significantly alleviate DNCB-induced AD symptoms in mice by reducing epidermal thickness and mast cell infiltration, with effects superior to marketed drugs litexitinib and torvatinib. In summary, this class of compounds has a rationally designed structure, strong targeting, significant efficacy, and stable metabolism. By targeting one, two, or three key kinases among JAK3, TAK1, and ITK, it provides high-quality candidate molecules for developing innovative drugs to treat various diseases (such as atopic dermatitis, rheumatoid arthritis, and related hematologic malignancies) driven by abnormalities of these kinases or coexisting in the same individual, demonstrating unique therapeutic potential and clinical application prospects.

[0074] Although embodiments of the present invention have been disclosed above, they are not limited to the applications listed in the specification and embodiments. They can be applied to various fields suitable for the present invention. For those skilled in the art, other modifications can be easily made. Therefore, without departing from the general concept defined by the claims and their equivalents, the present invention is not limited to the specific details and embodiments shown and described herein.

Claims

1. A pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof, characterized in that, The general formula of the pyrrolopyrimidine derivative is shown below; Wherein, R1 is a 5- or 6-membered heterocyclic group of substituted or unsubstituted phenyl or heteroatoms of oxygen, nitrogen and sulfur, and the substituent is selected from halogen atom, C1-C6 alkyl, C1-C3 alkoxy, cyano, nitro; X is selected from , , , , , , , , , , ; Y1 is a C2-C5 alkenyl, C2-C5 alkynyl, or halogen-substituted C1-C4 alkyl; Y2 is a substituted or unsubstituted C1-C6 alkyl group, and the substituent is selected from halogen atoms and C1-C3 alkoxy groups.

2. The pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof as described in claim 1, characterized in that, The structural formula of the pyrrolopyrimidine derivative is selected from: 。 3. The use of the pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof as described in claim 1, characterized in that, Used to prepare drugs that inhibit at least one of JAK3, TAK1, and ITK.

4. The use of the pyrrolopyrimidine derivative or a pharmaceutically acceptable salt thereof as described in claim 1, characterized in that, Used to prepare drugs for treating diseases caused by JAK3, TAK1, and ITK.

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