Phenylalanine-derived bithiazole compounds and uses thereof

Abstract

The present application relates to phenylalanine-derived bithiazole compounds and uses thereof, in particular to compounds of formula I wherein X and R are as defined herein, or a pharmaceutically acceptable salt thereof, pharmaceutical compositions comprising said compounds, and uses of said compounds in the manufacture of a medicament for the prevention or treatment of cancer, in particular pancreatic cancer.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry, and more specifically to phenylalanine-derived bithiazole compounds, pharmaceutical compositions comprising said compounds, and the use of said compounds in the preparation of medicaments for the prevention or treatment of diseases. Background Technology

[0002] Pancreatic cancer, known as the "king of cancers," has long been a major medical problem that plagues human health and urgently needs to be addressed. The pancreas is located in the upper right abdomen and is easily overlooked in early cancer screening. According to statistics, more than 80% of initial diagnoses are already at an advanced stage, and only 10% of patients are suitable for surgical resection. Even among pancreatic cancer patients who are suitable for surgery, the chance of postoperative metastasis is very high, and these recurrent patients often exhibit high resistance to radiotherapy and chemotherapy (Pancreatic cancer. Lancet, 2020, 395: 2008-2020.). In 2020, data from the WHO Global Cancer Report (GLOBOCAN 2020) showed that pancreatic cancer ranked 14th in new cases of all malignant tumors worldwide (up to 500,000 cases) and 7th in deaths (up to 470,000 cases) (CA Cancer J Clin, 2021, 71(3), 209-249.). my country is also a high-incidence area for pancreatic cancer, and the number of patients is showing an increasing trend year by year. (China Cancer., Analysis and Prediction of Pancreatic Cancer Incidence and Mortality Trends in China from 2000 to 2030 Based on GBD Data, 2022).

[0003] Currently, although there are many types of anti-tumor drugs, such as classic cytotoxic drugs, anti-hormonal drugs, and the emerging molecularly targeted drugs, which are divided into small molecule chemical drugs and biological agents (Semin Cancer Biol, 2021, 68:123-131.), drugs for treating pancreatic cancer are still very scarce. Pancreatic cancer responds poorly to most chemotherapy drugs, and surgical resection is currently the only chance of a cure. Early detection is difficult, and late-stage treatment is less effective. 80-85% of patients are already in the late stage when the disease is discovered, making surgical treatment impossible. Therefore, chemotherapy plays an important role (J Surg Oncol, 2017, 116(1):25-34.). Gemcitabine is the main first-line drug used clinically for early-stage pancreatic cancer that can be treated surgically and for late-stage pancreatic cancer that cannot be directly treated surgically. In addition, derivatives of traditional anticancer drugs such as albumin-bound paclitaxel, oxaliplatin, liposomal irinotecan hydrochloride, FOLFIRINOX, and tegafur are also widely used in the treatment of pancreatic cancer. In recent years, as the molecular mechanisms of pancreatic cancer have been continuously elucidated, new targeted drugs such as adherent kinase, PARP inhibitors, lysine oxidase inhibitors, and chemokine CC-motif receptor 2 (CCR2) inhibitors have also made some progress (Chinese Journal of Cancer, New Advances in Pancreatic Cancer Research and Treatment in 2021, 2022, 32, 1-12.).

[0004] However, there is still a great need to discover and develop new compounds for treating cancer, especially pancreatic cancer. Summary of the Invention

[0005] The inventors, while undertaking and completing the Shandong Provincial Natural Science Foundation project "Efficient Construction of Bithiazole / Oxazole Structural Units and Development and Application in Marthiaapeptide-like Anticancer Marine Cyclic Peptides (Project No.: ZR2020MB105)," discovered that the lead compound Boc-D-alanine bithiazole-tryptamine compound is effective against HCT116 colon cancer cell line (IC). 50 =2.31μM), gastric cancer cell line MKN45 (IC 50 =10.02μM), non-small cell lung cancer cell line A549 (IC50) 50 =5.71μM) and drug-resistant oral cancer cell line KBV (IC 50 =3.31μM) all showed a certain inhibitory effect (ZL202011573589.6).

[0006] In our in-depth research, we unexpectedly discovered a new skeletal thiazole compound that exhibits particularly excellent killing effects on pancreatic cancer cells.

[0007] On the one hand, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof.

[0008]

[0009] in:

[0010] -X- is selected from: C 1-6 -alkylene-;

[0011] R is selected from: -NR1R2, 9-10 membered bicyclic heteroaryl- and

[0012] R1, R2, R3, and R4 are each independently selected from H and C. 1-6 alkyl-.

[0013] In some implementations, R is selected from: -NR1R2, 9-10 membered bicyclic heteroaryl- and

[0014] In some implementations, R is selected from: -NR1R2, indole- and

[0015] In some implementation schemes, R is selected from or

[0016] In some implementations, -X- is selected from: C 1-3 -alkylene-; preferably -CH2-CH2-.

[0017] In one embodiment, the compound of formula I is selected from:

[0018]

[0019] In another aspect, the present invention provides pharmaceutical compositions comprising a compound of formula I or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable excipient.

[0020] On the other hand, the present invention provides a compound of formula I or a pharmaceutically acceptable salt thereof for treatment, particularly for treating cancer, preferably pancreatic cancer.

[0021] On the other hand, the present invention provides the use of a compound of formula I or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating cancer, preferably pancreatic cancer.

[0022] In another aspect, methods for synthesizing compounds of formula (I) are provided, and general synthetic schemes for synthesizing the compounds of the present invention are illustrated below.

[0023] In one embodiment, the present invention provides a method for preparing a compound of formula I, comprising the following steps:

[0024]

[0025] Compound 10 reacts with NH2-XR to produce compound I.

[0026] Where X and R are defined as in Equation I above.

[0027] In one embodiment, compound 10 is prepared by a method comprising the following steps:

[0028]

[0029] Step 1: React compound 3 with compound 4 to obtain compound 5;

[0030] Step 2: Hydrolyze compound 5 to obtain compound 6;

[0031] Step 3: React compound 6 with compound 4 to obtain compound 7;

[0032] Step 4: Hydrolyze compound 7 to obtain compound 8;

[0033] Step 6: React compound 8 with compound 4 to obtain compound 9; and

[0034] Step 6: Hydrolyze compound 9 to obtain compound 10.

[0035] For each reaction step, appropriate reaction conditions are known to those skilled in the art or can be conventionally determined, such as those shown in the examples. Unless otherwise specified, the starting materials and reagents used in the preparation of these compounds are generally commercially available or can be prepared by the methods described below, similar methods, or methods known in the art. If necessary, the starting materials and intermediates in the synthetic reaction process can be separated and purified using conventional techniques, including but not limited to filtration, distillation, crystallization, and chromatography. The materials can be characterized using conventional methods, including physical constants and spectroscopic data.

[0036] definition:

[0037] Unless otherwise specified, the various terms used in this specification and claims have the meanings shown below. Where a particular term or phrase is not specifically defined, it should not be considered uncertain or unclear, but rather should be appropriately understood in accordance with the context herein or its ordinary meaning in the art.

[0038] As used herein, the term "alkyl" refers to a fully saturated straight-chain or branched aliphatic hydrocarbon group. Specifically, alkyl groups can have 1 to 6, 1 to 5, 1 to 4, 1 to 3, or 1 to 2 carbon atoms. A suitable C... 1-6Examples of alkyl groups include, but are not limited to, methyl, ethyl, n-propyl, and isopropyl.

[0039] When the alkyl groups defined above become divalent groups, that is, when they have two single bonds attached to two other groups, they are called "alkylene" groups. Alkylenes can have 1 to 6, 1 to 5, 1 to 4, 1 to 3, or 1 to 2 carbon atoms. Non-limiting examples of alkylenes include methylene, ethylene, methylmethylene, or propylene.

[0040] As used herein, the term "9-10 membered bicyclic heteroaryl" refers to an aromatic bicyclic system having 9-10 ring members and containing 1-3 (e.g., 1, 2, or 3) heteroatoms selected from N, O, and / or S. For example, a 9-10 membered bicyclic heteroaryl is a fused bicyclic heteroaryl. Representative heteroaryl groups include, but are not limited to, indole, isoindole, indazinyl, and indazole groups.

[0041] "Boc" refers to tert-butyloxycarbonyl, i.e.

[0042] As used in this article, the term "pharmaceutically acceptable excipient" is used.

[0043] As used herein, the term “pharmaceutical acceptable” means a molecular entity or composition that is approved or can be approved by the relevant authorities of each country, or is listed in a generally accepted pharmacopoeia for use in animals and more specifically in humans, or that will not produce adverse, allergic or other adverse reactions when administered appropriately to animals, such as humans.

[0044] As used herein, the term "pharmaceutically acceptable salt" means a salt of the compound of the present invention that is pharmaceutically acceptable and has the pharmacological activity required by the parent compound. Specifically, such salts are non-toxic and may be inorganic acid addition salts, organic acid addition salts, or base addition salts. Specifically, such salts include: (1) acid addition salts formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, etc.; or acid addition salts formed with organic acids such as acetic acid, propionic acid, hexanoic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, benzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid. Sulfonic acid, camphor sulfonic acid, glucohepanoic acid, 3-phenylpropionic acid, trimethylacetic acid, tert-butylacetic acid, lauryl sulfate, gluconic acid, glutamic acid, hydroxynaphthenic acid, salicylic acid, stearic acid, mucoconic acid, etc.; or (2) salts formed when acidic protons present in the parent compound are replaced by metal ions such as alkali metal ions, alkaline earth metal ions or aluminum ions, or when coordinated with organic bases such as ethanolamine, diethanolamine, triethanolamine, N-methylglucosamine, etc. Those skilled in the art understand the general principles and techniques for preparing pharmaceutical salts, such as those described in Berge et al., Pharm ScL, 66, 1-19. (1977).

[0045] As used herein, the term "pharmaceuticalally acceptable excipient" refers to an additive in a pharmaceutical composition other than the active ingredient, also known as an excipient, that is pharmacologically inactive, compatible with other components in the composition, and acceptable for administration to warm-blooded animals such as humans. It can be used as a carrier or medium for the compounds of this invention in the form of administration, examples including but not limited to cellulose and its derivatives (such as sodium carboxymethyl cellulose, cellulose acetate, etc.), gelatin, talc, solid lubricants (such as magnesium stearate), calcium sulfate, vegetable oils, polyols (such as propylene glycol, glycerin, mannitol, sorbitol, etc.), emulsifiers (such as Tween derivatives), wetting agents (such as sodium dodecyl sulfate), colorants, flavoring agents, stabilizers, antioxidants, preservatives, etc.

[0046] As used herein, "compounds of the present invention" includes compounds of formula I of the present invention or pharmaceutically acceptable salts thereof.

[0047] Beneficial effects:

[0048] First, the compounds of the present invention all showed significant killing effects on pancreatic cancer cell lines PANC-1, ASPC-1, and Miapaca-2.

[0049] Secondly, the compounds of the present invention use commercially available compounds as raw materials, and have the advantages of simple preparation methods, easy separation, and excellent killing effect on pancreatic cancer cell lines. Attached Figure Description

[0050] The accompanying drawings are provided to further understand the technical solutions of the present invention and constitute a part of the specification. They are used together with the specific embodiments of this application to explain the technical solutions of the present invention and do not constitute a limitation on the technical solutions of the present invention.

[0051] Figure 1 : The 1H NMR spectrum of compound 7.

[0052] Figure 2 : Carbon NMR spectrum of compound 7.

[0053] Figure 3 : The 1H NMR spectrum of compound 9.

[0054] Figure 4 : Carbon NMR spectrum of compound 9.

[0055] Figure 5 : The 1H NMR spectrum of compound 1.

[0056] Figure 6 : Carbon NMR spectrum of compound 1.

[0057] Figure 7 : The 1H NMR spectrum of compound 2.

[0058] Figure 8 : Carbon NMR spectrum of compound 2.

[0059] Figure 9 : The 1H NMR spectrum of compound 11.

[0060] Figure 10 : Carbon NMR spectrum of compound 11.

[0061] Figure 11 : The 1H NMR spectrum of compound 12.

[0062] Figure 12 : Carbon NMR spectrum of compound 12.

[0063] Figure 13 Half-maximal inhibitory concentration (IC50) of compound 11 against human pancreatic cancer cell lines PANC-1, ASPC-1, and Miapaca-2.

[0064] The compounds described in this specification are further illustrated in the following examples. These examples are given by way of illustration only and are not limiting. Detailed implementation method:

[0065] The technical solution of the present invention will be further described below with reference to specific embodiments; however, the scope of protection of the present invention is not limited to these embodiments. All changes or equivalent substitutions that do not depart from the concept of the present invention are included within the scope of protection of the present invention.

[0066] Experimental methods not specifically described in the following examples are generally performed under standard conditions for such reactions or under conditions recommended by the manufacturer.

[0067] The abbreviations used in this application have the meanings commonly understood in the art, unless otherwise clearly defined in the specification. The meanings of some of the abbreviations used in the specification are listed below:

[0068] FDPP: Pentafluorophenyl diphenyl phosphate

[0069] DIPEA: Diisopropylethylamine;

[0070] HATU: 2-(7-Azobenzotriazole)-N,N,N',N'-Tetramethylurea hexafluorophosphate;

[0071] DBU: 1,8-diazahexacyclic[5,4,0]undecene-7.

[0072] Example 1 – Preparation of Boc-D-phenylalanine-derived bisthiazole compound 1:

[0073]

[0074] Preparation of compound 5:

[0075] In a 250 ml round-bottom flask, 3.3 g of Boc-D-phenylalanine dissolved in 100 ml of dichloromethane was added. While stirring, 3.15 g of Et3N, 4.75 g of FDPP (pentafluorophenyl diphenyl phosphate, CAS: 138687-69-1), 1.5 g of dimethyl azidocysteine, and 8.18 g of PPh3 were added sequentially. The mixture was heated in an oil bath at 50 °C for 5 h. After removing the oil bath, the mixture was evaporated to dryness and passed through a column chromatography with a petroleum ether:ethyl acetate ratio of 6:1. After separating PPh3, the column was washed with ethyl acetate to obtain an oily mixture. This mixture was dissolved in 50 ml of dichloromethane, and 1.7 g of DBU and 2.2 g of BrCCl3 were added. The mixture was stirred at room temperature for 10 min, and TLC monitoring was maintained until the reaction was complete. The mixture was then evaporated to dryness at low temperature and passed through a column chromatography with a petroleum ether:ethyl acetate ratio of 3:1 to obtain 2 g of product (compound 5), with a yield of 71%. [α] D 20 +16.96 (c 2.5, CHCl3); 1 H NMR(400MHz, CDCl3)δ:1.39(s,9H),3.22-3.38(m,2H),3.96(s,3H),5.27(s,br,2H),7.08-7.10(m,2H),7.22-7.28(m,3H),8.06(s,1H); 13C NMR (100MHz, CDCl3) δ: 28.4, 41.7, 52.7, 54.0, 80.5, 127.2, 127.7, 128.8, 129.6, 136.3, 147.1, 155.1, 162.0, 173.5.

[0076] Preparation of compound 7:

[0077] Compound 5 was dissolved in 10 ml of methanol and 10 ml of THF, stirred until dissolved, and then 10 ml of 0.5 N LiOH aqueous solution was added. After 10 min, the reaction was monitored by thin-layer chromatography. After the reaction was complete, 30 ml of water was added to the system, and concentrated hydrochloric acid was diluted 10:1 to adjust the pH of the system to 2-3. The system was extracted three times with an equal volume of ethyl petroleum ether, washed once with saturated NaCl, and dried over anhydrous Na2SO4 to obtain compound 6 (2 g, 86.6%). The product from the previous step was dissolved in 50 ml of dichloromethane, and Et3N 4.44 g, FDPP 4.2 g, dimethyl azidocysteine ​​1.76 g, and PPh3 11.7 g were added sequentially while stirring. The mixture was heated in an oil bath at 50 °C for 5 h, then the oil bath was removed, the mixture was evaporated to dryness, and passed through a column. The ratio of petroleum ether to ethyl acetate was 6:1. After separating PPh3, the column was flushed with ethyl petroleum ether to obtain an oily mixture. The above mixture was dissolved in 30 ml of dichloromethane, and 0.83 ml of DBU and 0.54 ml of BrCCl3 were added. The reaction was monitored by thin-layer chromatography for ten minutes until completion. The product (compound 7) was obtained by rotary evaporation at low temperature and column chromatography (petroleum ether: ethyl acetate = 2:1), yielding 850 mg of product. The two-step yield was 34%. [α] D 20 +22.8 (c 1, CHCl3); 1 H NMR (500MHz, CDCl3) δ8.18 (s, 1H), 8.00 (s, 1H), 7.22 (m, J = 13.5, 6.8Hz, 3H), 7.11 (d,J=7.1Hz,2H),5.28(m,J=14.9Hz,2H),3.95(s,3H),3.31(s,2H),1.39(s,9H). 13 C NMR (125MHz, CDCl3) δ: 173.1, 163.5, 161.9, 155.0, 148.2, 147.5, 136.2, 129.5, 128.6, 128.1, 127.0, 117.3, 80.3, 53.7, 52.5, 41.4, 28.3.

[0078] Preparation of compound 9:

[0079] The above product was hydrolyzed using the same steps as before, yielding 750 mg of dithiazolium acid, with a yield of 91.1%. This product was dissolved in 30 mL of dichloromethane, and under stirring, 1.4 g of Et3N, 1.3 g of FDPP, 556 mg of dimethyl azidocysteine, and 3.6 g of PPh3 were added sequentially. The mixture was heated in an oil bath at 50 °C for 5 h, then the oil bath was removed, and the solution was evaporated to dryness and passed through a column in a petroleum ether:ethyl acetate ratio of 6:1. After separating PPh3, the column was washed with ethyl acetate to obtain an oily mixture. This mixture was dissolved in 15 mL of dichloromethane, and 0.265 mL of DBU and 0.172 mL of BrCCl3 were added. The reaction was monitored by TLC until completion. The solution was evaporated to dryness at low temperature and passed through a column in a petroleum ether:ethyl acetate ratio of 2:1 to give trithiazolium compound 9 (250 mg), with a two-step yield of 27%. [α] D 20 +50.4 (c 1, CHCl3); 1 H NMR(500MHz,d-DMSO)δ8.62(s,1H),8.42(s,1H),8.30(s,1H),7.88(d,J=8.5H z,1H),7.51–7.07(m,5H),5.75(s,2H),5.06(s,1H),3.88(s,3H),1.33(s,9H). 13 C NMR(500MHz,d-DMSO)δ:176.4,163.5,162.8,161.6,155.7,148.8,147.5,14 7.2,138.1,130.3,129.7,128.6,119.8,118.8,79.1,55.4,54.7,52.6,28.6.

[0080] Preparation of compound 1:

[0081] Compound 9 was hydrolyzed, and 200 mg was dissolved in 25 mL of dichloromethane. Then, 74 mg of diisopropylethylamine, 217 mg of HATU, and 50 mg of N,N-dimethylethylamine were added sequentially. The reaction was detected by TLC. After completion, the mixture was quenched with saturated ammonium chloride, washed once with 0.5 N NaOH solution, washed once with saturated brine, dried over anhydrous sodium sulfate for 10 min, and then evaporated to dryness and passed through a column (dichloromethane:methanol = 15:1) to obtain compound 1 (200 mg, yield 90%). [α] D 20 +0.8 (c 0.5, CHCl3); 1H NMR (500MHz, CDCl3) δ8.22(s,1H),8.13(s,1H),8.07(s,1H),7.88(t,J=5.0Hz,1H),7.47-7.30(m,3H),7.25(d,J=7.0Hz,2H) ,5.52(br,s,1H),5.44(br,s,1H),3.69(q,J=6.0Hz,2H),3.50-3.41(m,2H),2.69(t,J=6.0Hz,2H),2.43(s,6H),1.53(s,9H). 13 C NMR(125MHz, CDCl3)δ:173.2,163.3,162.5,161.2,155.1,150.9,149.4,148.5,136.3 ,129.5,128.6,127.0,123.6,117.4,116.89,80.3,58.3,53.8,45.4,41.4,37.0,28.3.

[0082] Example 2 – Preparation of Boc-D-phenylalanine-derived bisthiazole compound 2:

[0083]

[0084] Using similar condensation conditions as compound 1, N,N-dimethylethylamine was replaced with 2-(1H-indol-3-yl)ethyl-1-amine to prepare target compound 2. 1 H NMR(500MHz,d-DMSO)δ:8.55(t,J=6.0Hz,1H),8.36-8.20(m,3H),7.89(d,J=8.5Hz,1H),7.64(d,J=8.0Hz,1H),7.32(dt,J=15.0,J=8.0Hz,6H),7.2 2(s,2H),7.08(t,J=7.5Hz,1H),6.99(t,J=7.5Hz,1H),3.61(dd,J=14.0, J=7.0Hz,2H),3.00(t,J=7.5Hz,2H),1.33(s,9H),1.21(d,J=10.5Hz,3H). 13C NMR(125MHz,d-DMSO)δ:176.4,163.6,162.1,160.8,155.7,151.5,149.2,147.5,138.2,136.8,129.7,128.7,1 27.7,126.9,124.7,123.1,121.5,119.0,118.9,118.7,112.2,111.9,79.1,54.7,38.7,29.5,28.6,25.7,22.6.

[0085] Example 3 – Preparation of Boc-D-phenylalanine-derived bithiazole compound 11:

[0086]

[0087] Using condensation conditions similar to those used in the preparation of compound 1, but replacing N,N-dimethylethylamine with ethylenediamine, target compound 11 was prepared. 1 H NMR(400MHz,d-DMSO)δ:8.51(t,J=5.6Hz,1H),8.39(s,1H),8.33(s,1H),8.31(s,1H),7.90(d,J=10.8Hz,1H),7.27-7.32(m,4H),7.21-7.24(m, 1H),7.22(s,2H),5.03-5.09(m,1H),3.58(dd,J=12.0,J=6.0Hz,2H),3. 35-3.39(m,1H),3.05-3.11(m,1H),3.02(t,J=6.0Hz,2H),1.33(s,9H). 13 C NMR(100MHz,d-DMSO)δ:176.0,163.3,161.7,161.2,155.3,150.6,148.7,147.1 ,137.8,129.6,128.3,126.5,125.3,118.7,78.7,67.1,36.8,28.2,26.4,25.2.

[0088] Example 4 – Preparation of Boc-D-phenylalanine-derived bithiazole compound 12:

[0089]

[0090] Using condensation conditions similar to those used in the preparation of compound 1, N,N-dimethylethylamine was replaced with 2-(2-aminoethyl)-6-(dimethylamino)-1H-benzo[de]isoquinoline-1,3(2H)-dione to prepare target compound 12. 1H NMR (500MHz, d-DMSO) δ: 8.59 (dd, J = 7.5Hz, J = 2.0Hz, 1H), 8.49 (d, J = 7.5Hz, 1H), 8.44 (dd, J = 7. 5Hz,J=1.5Hz,1H),8.10-8.14(m,2H),8.03(s,1H),7.96(s,1H),7.65(dt,J=8.5,J=1.0Hz,1H), 7.23-7.29(m,4H),7.12-7.16(m,2H),7.10(d,J=7.5Hz,1H),5.35(s,br,1H),5.21-5.28(m,1H) ,4.56(t,J=6.0Hz,2H),3.88(q,J=5.5Hz,2H),3.37(t,J=5.5Hz,2H),3.11(s,6H),1.44(s,9H). 13 C NMR(100MHz, CDCl3)δ:165.3,164.8,163.5,162.3,161.7,157.4,151.0,149.9,148.8,133.2,131.7,131.5,130.7 ,129.8,128.9,127.3,125.5,125.1,123.4,123.1,117.5,117.0,114.8,113.5,77.5,54.7,45.0,39.5,39.3,28.5.

[0091] Example 5 – Pharmacological Test

[0092] The inhibitory activity of Boc-D-phenylalanine-derived thiazole compounds 1, 2, 11, and 12 on human pancreatic cancer cell lines PANC-1 (purchased from Nanjing Kebai Biotechnology Co., Ltd.), ASPC-1 (purchased from Nanjing Kebai Biotechnology Co., Ltd.), and Miapaca-2 (purchased from Nanjing Kebai Biotechnology Co., Ltd.) was evaluated.

[0093] The half-maximal inhibitory concentration (IC50) of the compound against human pancreatic tumor cell lines was determined using the MTT assay (3-(4,5-dimethylthiazol-2)-2,5-diphenyltetrazol bromide). 50 Detection principle: MTT can penetrate the cell membrane and enter the cell. Succinate dehydrogenase in the mitochondria of living cells reduces exogenous MTT to water-insoluble, blue-purple needle-shaped formazan crystals, which are deposited in the cell. The crystals can be dissolved by dimethyl sulfoxide (DMSO), and the absorbance reflects cell viability. Half-maximal inhibitory concentration (IC50) 50 The drug concentration is defined as the concentration at which 50% of tumor cells survive. A standard curve of cell growth inhibition rate is created based on the measured optical density (OD value), and the corresponding drug concentration is then obtained from the curve.

[0094] The determination method is as follows:

[0095] 1. Seed cells at a density of 2000 cells / well in 96-well plates and incubate in a CO2 incubator for 24 hours;

[0096] 2. Continuously dilute the analyte (stock solution 10mM) to 10μM, 5μM, 2.5μM, 1.25μM, 0.627μM, 0.3125, 0.1562, 0.078, 0.039, 0.020, and 0.010μM;

[0097] 3. Set up three sets of parallel wells for each concentration, add the test compound to the seeded cells, and continue culturing for 72 hours;

[0098] 4. Remove the 96-well plate, add 20 μl of MTT solution to each well, and incubate in an incubator for 2 hours;

[0099] 5. Discard the supernatant, add 150 μl DMSO to each well, shake well for 15 min, and wait for the formazan to completely dissolve;

[0100] 6. Measure the absorbance at 570 nm.

[0101] Three parallel experiments were conducted, and the average value of the results was taken. The measured IC 50 The values ​​are shown in Table 1.

[0102] Pharmacological tests showed that the Boc-D-phenylalanine-derived thiazolium compounds 1, 2, 11, and 12 of this invention exhibited significant antitumor activity against human pancreatic cancer cell lines PANC-1, ASPC-1, and Miapaca-2. Their IC50 values... 50 The lowest values ​​reached 0.02 μM, 0.20 μM, and 0.27 μM, respectively. These compounds have good drug development potential. In particular, compound 11 showed nearly 500-fold increased killing activity against pancreatic cancer cell line PANC-1 compared to the previously reported patent compound (patent number: ZL202011573589.6). Compound 11 also showed excellent inhibitory effects against ASPC-1. The naphthalimide derivative 12 showed an IC50 value against pancreatic cancer cell line Miapaca-2. 50 The value can also reach 0.27 μM.

[0103] Table 1

[0104]

[0105] Among them, Boc-D-Ala-trithiazol-tryptamine (control) is compound 12 reported in patent ZL202011573589.6.

[0106] The Boc-D-phenylalanine-derived bithiazole compounds prepared in this invention, based on structural observation and experimental verification results, may be a G-quadruplex targeting inhibitor. The G-quadruplex is a non-classical secondary structure formed by single-stranded DNA or RNA sequences rich in guanine (G). It is a square planar structure formed by four guanine bases linked by Hoogsteen hydrogen bonds. G-quadruplexes were first discovered in telomeres, and later found in the promoter regions of various proto-oncogenes, such as c-KIT and c-MYC, indicating their important transcriptional regulatory role in proto-oncogenes (Biochem Biophys ResCommun, 2020, 531(1):45-50). Experiments have shown that the content of G-quadruplexes in tumor tissues is significantly higher than in normal tissues, making it an important target for distinguishing normal tissues from tumor tissues. G-quadruplex ligands can bind to and stabilize G-quadruplexes, preventing them from being unwound for further transcription or translation, thereby inhibiting the expression of proto-oncogenes and killing tumor cells (Nucleic Acids Res. 2018, 46, 5355–5365.).

[0107] The above experimental results describe in detail the preferred embodiments of the present invention. However, the present invention is not limited to the specific details of the above embodiments. Within the scope of the present invention, various equivalent transformations can be made to the technical solutions of the present invention, and all such transformations fall within the protection scope of the present invention. To avoid unnecessary repetition, the present invention will not describe the various possible combinations of structures separately; as long as they do not violate the spirit of the present invention, they should also be considered as the content disclosed by the present invention.

Claims

1. A compound of formula I or a pharmaceutically acceptable salt thereof, in: -X- is selected from: C 1-6 -alkylene-; R is selected from: -NR1R2, 9-10 membered bicyclic heteroaryl groups containing 1-3 heteroatoms selected from N, O and / or S, and... ;and R1, R2, R3, and R4 are each independently selected from H and C. 1-6 alkyl-.

2. The compound of formula I according to claim 1, or a pharmaceutically acceptable salt thereof, Wherein R is selected from: -NR1R2, 9-10 membered bicyclic heteroaryl groups containing 1-3 heteroatoms selected from N, O and / or S, and R1, R2, R3 and R4 are as defined in claim 1.

3. The compound of formula I according to claim 2, or a pharmaceutically acceptable salt thereof, Where R is selected from: -NR1R2, indole- and R1, R2, R3 and R4 are as defined in claim 1.

4. The compound of formula I according to claim 2, or a pharmaceutically acceptable salt thereof, Where R is selected from , , or .

5. The compound of formula I according to any one of claims 1-4, or a pharmaceutically acceptable salt thereof. Where -X- is selected from C 1-3 -alkylene-.

6. The compound of formula I according to claim 5, or a pharmaceutically acceptable salt thereof, Where -X- stands for -CH2-CH2-.

7. The compound of formula I according to claim 1, or a pharmaceutically acceptable salt thereof, wherein the compound of formula I is selected from: 。 8. A pharmaceutical composition comprising a compound of formula I according to any one of claims 1-7 or a pharmaceutically acceptable salt thereof, and optionally a pharmaceutically acceptable excipient.

9. Use of a compound of formula I according to any one of claims 1-7 or a pharmaceutically acceptable salt thereof in the preparation of a medicament for treating cancer, wherein said cancer is pancreatic cancer.

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