An antibacterial and antitumor synergist and its synthesis method and application

Abstract

The present invention discloses an antibacterial and antitumor synergist, a synthesis method and an application thereof. In view of the problem of drug resistance of antibacterial and antitumor drugs, the current mainstream strategy is still to re-develop new drugs for drug resistance-related targets; this strategy is time-consuming and labor-intensive, and cannot solve the drug resistance problem in the short term. The present invention discloses a class of compounds that can effectively increase the drug sensitivity of bacteria and tumor cells to antibiotics and anticancer drugs, and systematically optimizes the structure thereof, providing new candidate drug molecules for the development of clinical antibacterial and antitumor adjuvants.

Description

Technical Field

[0001] The invention belongs to pharmaceutical technology, and specifically relates to the synthesis and application of an antibacterial and antitumor synergist. Background Art

[0002] In recent years, due to the abuse of antibiotics, many bacteria have developed serious drug resistance, such as methicillin-resistant Staphylococcus aureus. It is estimated that by 2050, 10 million people will die from drug-resistant bacterial infections each year. The existing solution to the problem of drug resistance is combined drug therapy. However, this method only treats the symptoms and not the root cause. Once the target protein at the antibiotic binding site on the bacterial cell membrane mutates, the bacteria mutate, and combined drug therapy will also produce drug resistance, which will lead to the emergence of new multidrug-resistant bacteria or even super bacteria. Another hot topic of current research is peptide drugs. Researchers extract peptides from natural animals and plants and verify their antibacterial activity. However, this method currently has the problems of difficult administration, inability to mass produce, and high price, and is not suitable for clinical use. Similarly, anti-tumor drug resistance is the main reason for the failure and recurrence of clinical tumor treatment. One of the main reasons for the emergence of anti-tumor drug resistance is the dose-limiting toxicity of anti-tumor drugs, which leads to the limitation of clinical drug doses and the inability to effectively eliminate tumors in a short period of time, thus leading to the emergence of tumor resistance. An effective way to solve the problem of antibacterial and antitumor drug resistance is to develop corresponding sensitizing adjuvants to increase the sensitivity of antibiotics and chemotherapy drugs, thereby reducing the incidence of drug resistance and increasing the sensitivity of resistant bacteria / tumor cells to the corresponding drugs. In response to the problem of antibacterial and antitumor drug resistance, the current mainstream strategy is still to re-develop new drugs for resistance-related targets. This strategy is time-consuming and labor-intensive and cannot solve the problem of drug resistance in the short term. Summary of the invention

[0003] The present invention discloses a class of compounds that can effectively increase the drug sensitivity of bacteria and tumor cells to antibiotics and chemotherapeutic drugs, and systematically optimizes the structure thereof, providing a new candidate drug molecule for the research and development of clinical antibacterial and antitumor adjuvants.

[0004] The present invention adopts the following technical solution:

[0005] An antibacterial and antitumor synergist has the following chemical structure:

[0006]

[0007] Where R1 is -NO2, -CN, -SO2R5, -CF3, -N +(R5)3 or -COR5; R2 and R3 are independently selected from H or alkyl; R4 is an azocyclic or substituted azocyclic, azobisbicyclic, an alkoxy or substituted alkoxy, an amino or substituted amino, a phenolic or substituted phenolic, a sulfone or substituted sulfone, a mercapto or substituted mercapto; X is N or N + O - ; Y is O, S, Se or NR6, C(R6)2; R5 is hydrogen or C1-C6 alkyl; R6 is H or alkyl. The compound of the parent core structure has no antibacterial or antitumor effect by itself, but when used in combination with antibiotics and antitumor drugs, it can increase the drug sensitivity of bacteria and tumor cells.

[0008] In the above technical scheme, the substituent in R4 is a C1-C6 alkyl group, a nitrogen heterocyclic group, an ester group, a phenyl group or a substituted phenyl group. In the substituted phenyl group, the substituent is a C1-C3 alkyl group; the nitrogen-oxygen biheterocyclic group is a morpholinyl group; the nitrogen heterocyclic group is a piperazinyl group or an imidazolyl group; in the substituted nitrogen heterocyclic group, the substituent is on the nitrogen.

[0009] Specifically, the substituted nitrogen heterocyclic group has one of the following chemical structural formulas:

[0010]

[0011] R8 is a C1-C6 alkyl group, an ester group, a phenyl group or a substituted phenyl group, wherein the substituent in the substituted phenyl group is a C1-C3 alkyl group; the number of carbon atoms in the ester group is 3-6; and R9 is a C1-C6 alkyl group.

[0012] In the above technical solution, when R1 or R4 is a group with a positive charge, the synergist has a conventional anion coordination, and the anion can be a trifluoromethanesulfonic acid group, a halogen, and the like.

[0013] The present invention discloses a method for preparing the above-mentioned antibacterial and antitumor synergist. In the presence of an organic base, a 7-chloro-substituted benzoheterocyclic core compound is reacted with a substituted compound (corresponding amine, alcohol, phenol, thiophenol compound) to prepare the above-mentioned antibacterial and antitumor synergist. Preferably, the reaction is carried out in nitrogen, and the molar ratio of the 7-chloro-substituted benzoheterocyclic core compound to the amine compound is (1-1.5):1. The reaction is carried out at room temperature for 3-12 hours. The structure of the 7-chloro-substituted benzoheterocyclic core compound is as follows:

[0014]

[0015] The substituents therein are as described above.

[0016] The present invention discloses the use of the antibacterial and antitumor synergist in improving the therapeutic effect of antibacterial and / or antitumor drugs; or the use of the antibacterial and antitumor synergist in preparing drugs for improving the therapeutic effect of antibacterial and / or antitumor drugs.

[0017] The present invention discloses the use of the antibacterial and antitumor synergist in improving the therapeutic effect of photosensitizer drugs or in preparing drugs for improving the therapeutic effect of photosensitizer drugs. Photosensitizer drugs are used for antibacterial or tumor treatment.

[0018] The invention discloses the application of the antibacterial and antitumor synergist in the preparation of antibacterial and / or antitumor drugs.

[0019] The current mainstream strategy for the problem of drug resistance of antibacterial and antitumor drugs is to re-develop new drugs for drug resistance-related targets; this strategy is time-consuming and labor-intensive and cannot solve the drug resistance problem in the short term. The present invention discloses a class of compounds that can effectively increase the drug sensitivity of bacteria and tumor cells to antibiotics and anticancer drugs, and systematically optimizes their structure, providing new candidate drug molecules for the development of clinical antibacterial and antitumor adjuvants. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 are the chemical structural formulas of compounds 1 to 33 of the present invention.

[0021] Figure 2 Schematic diagram of the preparation of compound 2 of the present invention.

[0022] Figure 3 is the bacterial survival rate (Staphylococcus aureus) after synergistic use of compound 2 and gentamicin.

[0023] Figure 4 It is the bacterial survival rate after adding sensitizer and gentamicin to Pseudomonas aeruginosa, methicillin-resistant Staphylococcus aureus, Escherichia coli and Acinetobacter baumannii (clinical isolates).

[0024] Figure 5 It is the bacterial survival rate after the synergistic use of sensitizers and different antibiotics.

[0025] Figure 6 The bacterial survival rate (Staphylococcus aureus) after synergistic use of compound 2 and photosensitizer Ce6 under 655nm laser irradiation.

[0026] Figure 7 The bacterial survival rate under 655nm laser irradiation after synergistic use of compound 2 and photosensitizer Ce6.

[0027] Figure 8 is the cell survival rate after compound 2 was combined with camptothecin.

[0028] Fig. 9 It is the cell survival rate after compound 2 is used in combination with doxorubicin and CA-4.

[0029] Fig.10is the survival rate of mice after synergistic administration of compound 2 and gentamicin.

[0030] Fig.11 This is a statistical chart of colony counts in mouse peritoneal fluid and internal organs. DETAILED DESCRIPTION

[0031] The present invention discloses a sensitizer for antibiotics and anticancer drugs, thereby solving the problem of bacterial resistance to antibiotics and tumor cell resistance to anticancer drugs. Specifically, the present invention synthesizes compounds 1 to 33 (chemical structure shown in Figure 1 ), its activity in increasing the drug sensitivity of bacteria and tumor cells was evaluated in vitro, and its ability to increase the drug sensitivity of bacteria to antibiotics was verified in vivo.

[0032] The present invention discloses a method for preparing the above-mentioned antibacterial and antitumor synergist. Under a nitrogen atmosphere, a compound (1.1 equiv) of a 7-chloro-substituted benzoheterocyclic core structure is dissolved in an appropriate amount of dichloromethane (DCM), and then the corresponding amine / alcohol / phenol (1.0 equiv) and the corresponding base (DIPEA / TEA, 1.5 equiv) are added to react at room temperature for 3-12 h. After the reaction is complete, the solvent is concentrated under reduced pressure, and the residue obtained is separated and purified by column chromatography to obtain the target compound.

[0033] Embodiment 1

[0034] Compound 1: N-methylpiperazine (45.63 g, 0.45 mmol) and N,N-diisopropylethylamine (88.3 mg, 0.68 mmol) were mixed and dissolved in dichloromethane (2 mL), and then 4-chloro-7-nitrobenzo-2-oxa-1,3-oxadiazole (0.10 g, 0.50 mmol) was dissolved in 2 mL of DCM and added dropwise to the reaction solution. The reaction was allowed to proceed at room temperature for 8 h. After the reaction was complete, N,N-diisopropylethylamine was washed with water, and extracted with DCM. The obtained organic phase was concentrated under reduced pressure to obtain a crude product, and then separated by silica gel column chromatography to obtain an orange-red solid with a yield of 83%. 1 H NMR (400 MHz, CDCl3) δ 8.44 (d, J = 8.9 Hz, 1H),6.32 (d, J = 8.9 Hz, 1H), 4.13 (s, 4H), 2.68 (s, 4H), 2.40 (s, 3H).

[0035] The synthesis of compound 2 can be found in Figure 2 , as follows:

[0036] (a) Synthesis of BocMP. 1-Methylpiperazine (0.5 g, 4.99 mmol) was dissolved in tetrahydrofuran (6 mL), and then di-tert-butyl dicarbonate (1.72 mL, 7.49 mmol) was added at 0°C, and the reaction was moved to room temperature for 2 hours. The obtained reaction solution was concentrated under reduced pressure, and then the residue was separated and purified by column chromatography to obtain the product BocMP with a yield of 92%. 1 H NMR (400 MHz, CDCl3) δ 3.43 (s, 4H), 2.34 (s, 4H), 2.29 (d, J = 1.1 Hz, 3H), 1.45(d, J = 1.0 Hz, 9H).

[0037] (b) Synthesis of DMP. 1-Boc-4-methylpiperazine (BocMP, 0.5 g, 4.99 mmol) was dissolved in dry dichloromethane (DCM, 10 mL), and then methyl trifluoromethanesulfonate (0.409 g, 2.49 mmol) was added dropwise, and the mixture was reacted at room temperature for 1 hour. Then, trifluoroacetic acid was added dropwise to the reaction solution, and the mixture was reacted at room temperature for 1 hour. After the reaction was complete, the reaction solution was concentrated under reduced pressure, and the residue was washed with 10 mL of methanol. The precipitated white solid was the product, with a yield of 86%. 1 H NMR (400 MHz, D2O) δ 3.79 (d, J = 4.9 Hz, 4H), 3.73 (s, 4H), 3.34 (s, 6H).

[0038] (c) Synthesis of compound 2. DMP (0.2 g, 0.76 mmol) and triethylamine (210.2 μL, 1.15 mmol) were mixed and dissolved in dichloromethane (10 mL), and then 4-chloro-7-nitrobenz-2-oxa-1,3-diazole (NBD-Cl, 0.181 g, 0.91 mmol) was dissolved in 2 mL of DCM and added dropwise to the reaction solution. The reaction was allowed to proceed at room temperature for 12 h. After the reaction was complete, the solvent was concentrated under reduced pressure and the residue was chromatographed on a basic alumina column to obtain a brown product 2 with a yield of 30%. 1 HNMR (400 MHz, D2O) δ 8.50 (d, J = 8.8 Hz, 1H), 6.62 (d, J = 8.8 Hz, 1H), 4.47(s, 4H), 3.79 (s, 4H), 3.35 (s, 6H).

[0039] Compound 3: 2-chloro-1-methylpyridinium-1-iodide (0.1 g, 0.39 mmol) and triethylamine (108.8 μL, 0.78 mmol) were placed in a round-bottom flask, and then solvent DCM (10 mL) was added, and then NBD-piperazine (0.098 g, 0.39 mmol) was dissolved in 2 mL of DCM and added dropwise to the reaction solution, and reacted at room temperature for 12 h. After the reaction was complete, the solvent was concentrated under reduced pressure and removed, and the residue was chromatographed on a basic alumina column to obtain a reddish-brown product 3 with a yield of 54%. 1 H NMR (400MHz, DMSO- d 6 ) δ 8.64 (dd, J = 6.4, 1.5 Hz, 1H), 8.56 (d, J = 9.1 Hz, 1H),8.40 – 8.28 (m, 1H), 7.69 (d, J = 8.2 Hz, 1H), 7.61 – 7.47 (m, 1H), 6.74 (d, J = 9.2 Hz, 1H), 4.37 (s, 4H), 4.15 (s, 3H), 3.80 – 3.60 (m, 4H).

[0040] Compound 4: red solid, yield 78%. 1 H NMR (400 MHz, CDCl3) δ 8.44 (d, J = 8.6Hz, 1H), 7.73 (s, 1H), 7.30 (d, J = 5.9 Hz, 4H), 6.64 (d, J = 8.6 Hz, 1H),2.42 (s, 3H).

[0041] Compound 5: reddish brown solid, yield 69%. 1 H NMR (400 MHz, CDCl3) δ 8.44 (d, J = 9.1Hz, 1H), 6.13 (d, J = 9.1 Hz, 1H), 3.96 (s, 4H), 1.40 (t, J = 7.1 Hz, 6H).

[0042] Compound 6: red solid, yield 70%. 1H NMR (500 MHz, CDCl3) δ 9.14 (s, 1H), 8.17 (s, 1H), 6.12 (d, J = 7.5 Hz, 1H), 5.38 (d, J = 7.5 Hz, 1H), 4.36 (s, 3H).

[0043] Compound 7: reddish brown solid, yield 50%. 1 H NMR (500 MHz, CDCl3) δ 8.17 (d, J =15.0 Hz, 1H), 7.84 (d, J = 15.0 Hz, 1H), 7.60 (d, J = 15.0 Hz, 1H), 7.27 (d, J = 15.0 Hz, 1H), 7.15 (s, 1H).

[0044] Compound 8: Sodium ethoxide (68.20 mg, 1.00 mmol) was dissolved in ethanol (2 mL), and then 4-chloro-7-nitrobenz-2-oxa-1,3-oxadiazole (0.10 g, 0.50 mmol) was dissolved in 2 mL of ethanol, and then added dropwise to the reaction solution, and reacted at room temperature for 1 h. After the reaction was complete, the sodium ethoxide was washed with water, and extracted with DCM. The obtained organic phase was concentrated under reduced pressure to obtain a crude product, and then a yellow solid was obtained by silica gel column chromatography with a yield of 38%. 1 H NMR (400 MHz, CDCl3) δ 8.13 (d, J =9.8 Hz, 1H), 7.50 (d, J = 9.8 Hz, 1H), 4.48 (q, J = 7.0 Hz, 2H), 1.62 – 1.54(m, 3H).

[0045] Compound 9: orange-red solid, yield 55%. 1 H NMR (500 MHz, CDCl3) δ 7.50 (s, 1H), 6.63 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 1.18 (s, 1H).

[0046] Compound 10: reddish brown solid, yield 73%. 1H NMR (500 MHz, CDCl3) δ 7.47 (s, 1H), 6.91 (s, 1H), 3.99 (s, 2H), 3.86 (s, 2H), 3.76 (s, 4H), 3.32 (s, 6H).

[0047] Compound 11: yellow solid, yield 40%. 1 H NMR (500 MHz, CDCl3) δ 7.57 (s, 1H), 6.80 (s, 1H), 3.46 (s, 4H), 3.31 (s, 3H), 2.81 (s, 4H), 1.17 (s, 1H).

[0048] Compound 12: reddish brown solid, yield 50%. 1 H NMR (500 MHz, CDCl3) δ 7.41 (s, 1H), 6.80 (s, 1H), 3.87 (s, 2H), 3.80 (s, 2H), 3.73 (s, 4H), 3.31 (s, 3H), 3.02(s, 6H).

[0049] Compound 13: reddish brown solid, yield 50%. 1 H NMR (500 MHz, CDCl3) δ 7.31 (s, 1H), 6.38 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 1.17 (s, 1H).

[0050] Compound 14: yellow solid, yield 45%. 1 H NMR (500 MHz, CDCl3) δ 7.35 (s, 1H), 6.66 (s, 1H), 3.97 (s, 2H), 3.80 (s, 2H), 3.64 (s, 4H), 3.22 (s, 6H).

[0051] Compound 15: red solid, yield 45%. 1 H NMR (500 MHz, CDCl3) δ 7.24 (s, 1H), 6.80 (s, 1H), 3.78 (s, 9H), 3.46 (s, 4H), 2.80 (s, 4H), 1.16 (s, 1H).

[0052] Compound 16: brown solid, yield 60%. 1H NMR (500 MHz, CDCl3) δ 7.32 (s, 1H), 6.81 (s, 1H), 3.91 (s, 2H), 3.80 (d, J = 3.0 Hz, 9H), 3.33 (s, 4H), 3.01 (s, 6H).

[0053] Compound 17: yellow solid, yield 75%. 1 H NMR (500 MHz, CDCl3) δ 7.86 (s, 1H), 6.56 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 2.62 (s, 3H), 1.16 (s, 1H).

[0054] Compound 18: reddish brown solid, yield 50%. 1 H NMR (500 MHz, CDCl3) δ 7.81 (s, 1H), 6.56 (s, 1H), 3.88 (d, J = 13.1 Hz, 4H), 3.55 (s, 4H), 3.04 (s, 6H), 2.62 (s, 3H).

[0055] Compound 19: reddish brown solid, yield 40%. 1 H NMR (500 MHz, CDCl3) δ 7.48 (s, 1H), 6.61 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 1.15 (s, 1H).

[0056] Compound 20: yellow solid, yield 45%. 1 H NMR (500 MHz, CDCl3) δ 7.47 (s, 1H), 6.59 (s, 1H), 3.84 (d, J = 14.3 Hz, 4H), 3.65 (s, 4H), 3.19 (s, 6H).

[0057] Compound 21: yellow-brown solid, yield 45%. 1 H NMR (500 MHz, CDCl3) δ 8.18 (s, 1H), 6.61 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 1.15 (s, 1H).

[0058] Compound 22: reddish brown solid, yield 60%.1 H NMR (300 MHz, DMSO- d 6 ) δ 8.62 (d, J =8.8 Hz, 1H), 6.90 (d, J = 8.7 Hz, 1H), 4.36 (s, 4H), 3.72 (s, 4H), 3.31 (s, 6H). 13 C NMR (101 MHz, DMSO- d 6 ) δ 152.4, 151.8, 147.8, 131.8, 131.3, 106.0,59.6, 50.3, 42.6.

[0059] Compound 23: red solid, yield 50%. 1 H NMR (500 MHz, CDCl3) δ 7.99 (s, 1H), 6.99 (s, 1H), 3.46 (s, 4H), 2.81 (s, 4H), 1.23 (s, 1H).

[0060] Compound 24: yellow-brown solid, yield 70%. 1 H NMR (300 MHz, DMSO- d 6 ) δ 8.34 (d, J =8.9 Hz, 1H), 6.91 (d, J = 9.3 Hz, 1H), 4.61 (s, 4H), 3.72 (s, 4H), 3.30 (s, 6H). 13 C NMR (101 MHz, DMSO- d 6 ) δ 146.1, 135.4, 129.5, 126.9, 121.9, 104.8,59.4, 50.2, 41.8.

[0061] Compound 25: yellow-brown solid, yield 70%. 1 H NMR (500 MHz, CDCl3) δ 9.74 (s, 1H), 6.97 (s, 1H), 3.37 (s, 4H), 2.81 (s, 4H), 1.25 (s, 6H), 1.19 (s, 1H).

[0062] Compound 26: reddish brown solid, yield 45%.1 H NMR (500 MHz, CDCl3) δ 9.76 (s, 1H), 7.06 (s, 1H), 3.88 (s, 4H), 3.75 (s, 2H), 3.66 (s, 2H), 3.11 (s, 6H), 1.26 (s, 6H).

[0063] Compound 27: red solid, yield 50%. 1 H NMR (300 MHz, D2O) δ 8.33 (d, J = 8.6 Hz,1H), 6.23 (d, J = 8.6Hz), 3.40 (s, 4H), 3.12 (s,4H).

[0064] Compound 28: orange-red solid, yield 55% 1 H NMR (400 MHz, D2O) δ 8.41 (d, J = 41.1Hz, 1H), 6.64 (d, J = 8.6 Hz, 1H), 3.78 (s, 4H), 3.72 (s, J = 5.0 Hz, 4H), 3.29 (s, 6H).

[0065] Compound 29: reddish brown solid, yield 62%. 1 H NMR (400 MHz, CDCl3) δ 8.32 (d, J = 8.5Hz, 1H), 6.28 (d, J = 8.5 Hz, 1H), 4.22- 4.15 (m, 2H), 3.79-3.68 (m,4H), 3.42-3.29 (m, 4H), 1.29 (t, J = 7.1 Hz, 3H).

[0066] Compound 30: red solid, yield 94%. 1 H NMR (400 MHz, CDCl3) δ 8.01 (d, J = 9.0Hz, 1H), 5.99 (d, J = 9.1 Hz, 1H), 3.87 (d, J = 6.8 Hz, 4H), 1.37 (t, J = 7.0Hz, 6H).

[0067] Compound 31: reddish brown solid, yield 64%. 1 H NMR (500 MHz, CDCl3) δ 8.17 (s, 1H), 7.19 (s, 1H), 3.92 (s, 3H).

[0068] Compound 32: reddish brown solid, yield 50%. 1 H NMR (300 MHz, CDCl3) δ 8.33 (d, J = 8.5Hz, 1H), 6.27 (d, J = 8.5 Hz, 1H), 3.93 (s, 4H), 3.48 – 3.33 (m, 4H).

[0069] Compound 33: reddish brown solid, yield 69%. 1 H NMR (300 MHz, CDCl3) δ 8.59 (s, 1H), 8.30 (d, J = 6.6 Hz, 1H), 7.38-7.09 (m, 4H), 6.36 (d, J = 7.1 Hz, 1H), 2.42 (s, 3H).

[0070] Example 2 General detection of synergistic effects of antibacterial and antitumor synergists combined with antibiotics

[0071] The plate count method was used to verify whether the combination of synergists and antibiotics could produce a synergistic effect to reduce bacterial resistance. Bacteria (Gram-positive bacteria Staphylococcus aureus) were added to the bacterial culture medium MHB containing 0.5 mM cysteine ​​(Cys) and incubated at 37°C until the bacterial concentration reached 2×10 7 CFu / ml, then dilute the bacteria 100 times with 0.9% sterile NaCl aqueous solution, add 0.1098μg / mL antibiotic (gentamicin), and then add synergist; then incubate the sample at 37℃, dilute the sample 1000 times at different times (0, 0.5, 1, 2h) and take 50μL on LB solid medium, incubate at 37℃ for 16h, calculate the number of colonies, compare the number of surviving bacteria, and determine the bacterial survival rate. No antibiotics or synergist is the blank control (WT).

[0072]

[0073] CFu refers to colony forming unit. Take compound 2 as an enhancer as an example: at 0h, the colonies of all groups were on the same horizontal line, and as time changed, it was found that the bacterial survival rate of the experimental group that only added the synergist compound 2 did not change significantly compared with the blank control group with only bacteria, indicating that the synergist compound 2 itself cannot inhibit bacteria. The bacterial survival rate of the experimental group added with gentamicin was significantly lower than that of the blank control group, indicating the antibacterial effect of gentamicin. In the experimental group that added gentamicin and the synergist, the bacterial survival rate was even lower than that of the group that only added gentamicin, indicating that the sensitizer and gentamicin produced a synergistic effect and could kill more bacteria. See Figure 3 , in the figure, Gm is gentamicin and S2 is compound 2.

[0074] Similarly, the sensitizing effects of other synergist compounds 1-33 on gentamicin are shown in Table 1.

[0075] Table 1 The sensitizing effect of synergist compounds on gentamicin

[0076]

[0077] As can be seen from Table 1, the compounds all have different antibiotic sensitization effects, among which compound 2 has the most obvious effect.

[0078] Different antibiotics were further selected for combination with sensitizer compound 2 to verify whether it was effective against Gram-negative bacteria and drug-resistant bacteria. Figure 4 As shown in the figure, Gm is gentamicin, Po B is polymyxin B, and S2 is compound 2: Pseudomonas aeruginosa (CICC 23683) and methicillin-resistant Staphylococcus aureus (clinical isolate) were selected, and the sensitizing effect of sensitizer 2 was verified. From the results, the bacterial survival rate of the experimental group with only the synergist compound 2 was not significantly changed compared with the blank control group with only bacteria, indicating that the synergist compound 2 itself cannot inhibit bacteria. The bacterial survival rate of the experimental group with the addition of gentamicin was significantly lower than that of the blank control group, indicating the antibacterial effect of gentamicin. In the experimental group with the addition of gentamicin and the synergist, the bacterial survival rate was lower than that of the group with only gentamicin, indicating that the sensitizer and gentamicin had a synergistic effect and could inhibit more bacteria. At the same time, it was verified whether compound 2 would have a synergistic effect with other antibiotics. Ciprofloxacin, vancomycin, ampicillin and erythromycin were selected to cooperate with compound 2, and the experimental results are shown in the following figure. Figure 5 As shown, these antibiotics all have a certain sensitization effect after being used in combination with sensitizer 2, where CI is ciprofloxacin, Amp is ampicillin, Vca is vancomycin, and Em is erythromycin. S2 is compound 2. The above results are shown in Table 2.

[0079]

[0080] Example 3: Synergists enhance the killing effect of singlet oxygen on bacteria

[0081] The plate count method was used to verify whether the synergist enhanced the singlet oxygen killing effect on bacteria. The bacteria were incubated on a shaker at 37°C and 200 rpm until the bacterial concentration reached 4×10 7 CFu / ml, and then the bacteria were diluted 100 times with 0.9% sterile NaCl aqueous solution, and 1μM Ce6 photosensitizer and 25μM sensitizer were added; after incubation on a shaker for 10 minutes, the laser (energy density of 60 J·cm -2 ) and then the reaction solution was diluted 100 times, spread on LB solid medium, and incubated at 37°C for 16 h. The bacterial survival rate was determined by counting the number of colonies and comparing the number of surviving bacteria.

[0082] Take compound 2 as an example: in the non-light group, the bacterial survival rate of each group almost reached 100%, indicating that the photosensitizer Ce6 and the synergist compound 2 themselves are not toxic to bacteria. When 655 was added to the photosensitizer group, it showed that the addition of the synergist enhanced the bactericidal effect of singlet oxygen. In the experimental group with only the sensitizer, the light did not inhibit the bacteria, indicating that the sensitizer itself has no bactericidal effect, but the effect is produced by the synergistic effect of the sensitizer and singlet oxygen. The experimental results are as follows Figure 6 As shown in the figure, Ce6 is a photosensitizer and S2 is compound 2.

[0083] Similar results were also observed in Escherichia coli and methicillin-resistant Staphylococcus aureus. Figure 7 As shown in the figure, Ce6 is a photosensitizer and S2 is compound 2.

[0084] Example 4: Synergistic effect of the combination of synergist and anticancer drug

[0085] A549 cells were seeded in a 96-well plate (8000 cells / well, 100 μL per well), and then cultured in a 37°C incubator for 24 hours. Drugs were then added (the control group was added with different concentrations of anticancer drugs or compound 2, and the experimental group was added with the synergistic combination of anticancer drugs and synergists). The plates were then cultured in a 37°C incubator for 12 hours. The drugs were aspirated and washed with PBS. 10% CCK8 solution was then added and the plates were cultured in an incubator for 30 minutes to 1 hour. The absorbance at 450 nm was measured by an enzyme reader to compare the differences between the groups. No drugs and compound 2 were used as blank controls.

[0086]

[0087] from Figure 8From the results, after the combination of camptothecin and compound 2, the survival rate of tumor cell A549 is lower than that of camptothecin alone, while compound 2 alone has no inhibitory effect on tumor cells. This phenomenon illustrates the synergistic anti-tumor activity of compound 2. In the figure, CPT is camptothecin and NBD-S2 is compound 2. Tumor cell A549 and anti-tumor drug doxorubicin were further selected to verify the anti-tumor sensitization effect of compounds 1-33. The experimental results are shown in Table 3. From the table, all compounds have sensitization effects, among which compound 2 has the best sensitization effect.

[0088] Table 3 Results of synergistic use of compounds and doxorubicin

[0089]

[0090] Further experiments were conducted with compound 2 to illustrate the synergistic effect on anti-tumor drugs, and anti-tumor drugs such as doxorubicin and CA-4 were selected. The experimental results are as follows Fig. 9 As shown in the figure, Dox is doxorubicin and NBD-S2 is compound 2. After compound 2 is used in combination with doxorubicin or CA-4, its killing effect on tumor cells is increased, verifying the anti-tumor synergistic effect of sensitizer compound 2. HepG2, 4T1, and Hela cells were selected for the above experiments, and it was seen that after the chemotherapy drugs were used in combination with the sensitizer compound 2, the sensitivity of tumor cells to anti-tumor drugs was increased. The experimental results are shown in Table 4.

[0091]

[0092] Example 5 Compound 2 and the antibiotic gentamicin synergistically treat peritonitis infection in mice

[0093] Establishment of peritonitis model: The bacteria incubated overnight were centrifuged at 7000 rpm for 5 minutes, and then the supernatant was discarded and resuspended in PBS. This was repeated 3 times and the obtained bacterial solution was used for later use. Then, 15 g of sterile blab / c mice were taken and 200 μL of 1*10 9 The infection status of mice was observed by measuring the CFu / mL of methicillin-resistant Staphylococcus aureus (MRSA).

[0094] The method of cure rate experiment was established: 32 sterile balb / c mice weighing about 15 g were randomly divided into four cages, and each mouse was intraperitoneally injected with 200 μL 1*10 9 CFu / mL of MRSA. Two hours after infection, mice were injected with 20 mg / kg of Gm and / or 60 mg / kg of compound 2, and the control group was injected with PBS. The living conditions of the mice were then observed for seven days. Fig.10From the above data, all mice in the PBS group and the compound 2 treatment group died within 3 days after modeling. In the antibiotic administration group, only two mice survived, while in the compound 2 and antibiotic combination group, five mice survived, indicating that antibiotics and compound 2 have a synergistic effect in this model. In the figure, Gm is gentamicin and NBD-S2 is compound 2.

[0095] Organ infection statistics were established by taking 32 sterile balb / c mice weighing about 15 g and randomly dividing them into four groups. Each group was intraperitoneally injected with 200 μL of 1*10 9 CFu / mL of MRSA. Two hours after the injection, 20mg / kg of Gm, 60mg / kg of compound 2, compound 2+Gm, and PBS were injected respectively. 12h after the administration, the mice were drowned and dissected, 1mL PBS was injected into the abdominal cavity to dissolve the bacteria in the abdominal cavity, and then sucked out and placed in a 1.5mL EP tube, and then the heart, liver, spleen, lungs, and kidneys of the mice were taken out. Then homogenized, the homogenized liquid was diluted and plated, placed in a 37℃ incubator for 16h, and then counted.

[0096] from Fig.11 It can be seen that the colony count in the synergistic group was significantly less than that in the group with only antibiotics. These results strongly prove the synergistic bactericidal effect between antibiotics and compound 2. In the figure, Gm is gentamicin and NBD-S2 is compound 2.

[0097] The present invention develops corresponding adjuvant molecules to increase the drug sensitivity of bacteria and tumor cells to drugs, and particularly discloses that compounds with a 7-nitro-1,2,3-benzodiazepine mother core structure can increase the drug sensitivity of bacteria and tumor cells in vitro and in vivo. There is no related report in the prior art on the ability of such skeleton compounds to increase the drug sensitivity of bacteria and tumor cells.

Claims

1. Use of an antibacterial and antitumor synergist in the preparation of a drug for improving the therapeutic effect of antibacterial and / or antitumor drugs, characterized in that: The antibacterial and antitumor synergist is one of the following chemical formulas: ； The antibacterial drug is one of gentamicin, polymyxin B, ciprofloxacin, ampicillin, vancomycin, and erythromycin; the antitumor drug is one of camptothecin, doxorubicin, and CA-4.

2. Use of the antibacterial and antitumor synergist of claim 1 in the preparation of a drug for improving the therapeutic effect of the photosensitizer drug Ce6 photosensitizer.

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