A reactive oxygen species-responsive local anesthetic drug based on quaternary ammonium lidocaine derivatives, its preparation and application

Abstract

This invention belongs to the field of pharmaceuticals, specifically relating to a reactive oxygen species-responsive local anesthetic drug based on a quaternary ammonium lidocaine derivative, its preparation, and its application. The drug of this invention can produce a long-lasting and selective anesthetic effect in vivo.

Description

Technical Field

[0001] This invention belongs to the field of pharmaceuticals, specifically relating to a reactive oxygen species-responsive local anesthetic drug based on a quaternary ammonium lidocaine derivative, its preparation and application. Background Technology

[0002] Compared with systemic analgesics, local anesthetics demonstrate significant advantages in relieving pain and reducing adverse reactions. However, their short-acting nature often limits their clinical application, thus necessitating the development of local anesthetics with long-acting, selective, and low-toxicity effects (Wang Q, Zhang Y, Liu J, et al. Quaternary lidocaine derivatives: past, present, and future[J]. Drug Design, Development and Therapy, 2021:195-207.). Quaternary lidocaine derivatives (QLDs) have been shown to possess these characteristics in electrophysiological studies and are therefore considered a very promising local anesthetic. QLDs are expected to become a hot research area for future local anesthetic drugs. QLDs began to attract researchers' attention in the 1970s. They have a similar chemical structure to commonly used lidocaine, but their molecules contain quaternary ammonium groups, which gives QLDs a longer duration of anesthesia and a more selective local anesthetic effect. QLDs have a wide range of applications, such as clinical medicine, surgery, and pain management. Because its mechanism of action is local anesthesia, it can effectively relieve pain and reduce the adverse reactions caused by general anesthetic drugs. At the same time, the long-lasting effect of QLDs can shorten the patient's recovery time after surgery and reduce unnecessary medical costs. Currently, the main types of QLDs include QX-222, QX-572, QX-314, and QX-OH. Among them, QX-314, as a widely studied local anesthetic, has advantages such as longer duration of action and selective sensory blockade compared to traditional local anesthetics (Lim TK, Macleod BA, Ries CR, et al. The quaternary lidocaine derivative, QX-314, produces long-lasting local anesthesia in animal models in vivo[J]. The Journal of the American Society of Anesthesiologists, 2007, 107:305-311.). However, QX-314 still has some limitations. Due to its very strong dose dependence, serious toxic side effects can occur when the dose is too high, therefore strict dosage control is required. Furthermore, the reduced lipophilicity of QX-314 makes it difficult for it to enter nerve cells and exert its effects. Therefore, overcoming these limitations is a top priority.

[0003] Quaternary ammonium lidocaine derivatives such as QX-314 and QX-222 possess the property of prolonged nerve conduction blockade. Among them, QX-314 has attracted widespread attention because, in addition to producing prolonged peripheral nerve blockade, it also exhibits sensory-selective analgesia when used in combination with capsaicin and chemopreservation enhancers. However, researchers used lidocaine (70 mM) as a positive control and saline as a negative control to investigate the adverse reactions of QX-314. They found that all mice injected with saline and lidocaine showed normal results. In contrast, in the QX-314 injection group, at a dose of 5 mM, 4 out of 6 mice (67%) experienced motor blockade and 5 out of 6 mice (83%) experienced sensory blockade, while at a dose of 10 mM, 5 out of 6 mice (83%) experienced sensory blockade, and 6 out of 6 mice (100%) showed a significant stimulus response. Notably, one of the six mice injected with a 5 mM dose (17%) died, and two of the six mice injected with a 10 mM dose (33%) died. These results suggest that while QX-314 can produce a durable nerve block, it is unlikely to be a suitable drug for human spinal anesthesia due to significant toxicity observed in mice (Schwarz SK, Cheung H MC, Ries CR, et al. Lumbar intrathecal administration of the quaternary lidocaine derivative, QX-314, produces irritation and death in mice[J]. The Journal of the American Society of Anesthesiologists, 2010, 113:438-444.). Furthermore, the study found that two of the four mice injected with 150 mM QX-314 developed nerve blocks that did not resolve during a week-long observation period. This indicates that high concentrations of QX-314 can lead to irreversible nerve blockade, posing a risk of neurotoxicity (Sagie I, Kohane DS. Prolonged sensory-selective nerve blockade[J]. Proceedings of the National Academy of Sciences, 2010, 107:3740-3745.). Systemic administration may even induce severe central nervous system and cardiotoxicity. Researchers observed signs of central nervous system toxicity (convulsions, ataxia, loss of righting reflex, and / or death) in mice by intravenously injecting different doses of lidocaine or QX-314 (dose range 7.5-30 mg / kg). Simultaneously, electrocardiograms were measured in mice to assess the cardiotoxicity of the drug.Statistical analysis of experimental data revealed that the ED50 for lidocaine's central nervous system toxicity was 19.5 mg / kg, while QX-314's ED50 was 10.7 mg / kg, lower than lidocaine's. Similarly, lidocaine's ED50 for cardiotoxicity was also higher than QX-314's. These results indicate that QX-314, at the same dose, has a significantly higher relative potency against central nervous system and cardiac toxicity than lidocaine, and QX-314 is not safer than lidocaine in terms of systemic toxicity. Although QX-314 may be a potentially clinically useful drug, its systemic toxicity compared to traditional local anesthetics offers no advantage in its application. Summary of the Invention

[0004] To address the shortcomings of existing methods, the present invention aims to provide a reactive oxygen species-responsive local anesthetic drug based on a quaternary ammonium lidocaine derivative, as well as its preparation and application.

[0005] The technical solution adopted by this invention to solve its technical problem is:

[0006] In a first aspect, the present invention protects a compound S-QXOH, said compound having the following structure:

[0007]

[0008] When the above compounds come into contact with reactive oxygen species, the sulfur atoms in the molecules are oxidized by the reactive oxygen species, and the oxygen atoms of the reactive oxygen species further attack the ester bonds in the molecules, thereby breaking the ester bonds and releasing the local anesthetic QX-OH molecules to produce an anesthetic effect.

[0009]

[0010] Secondly, this invention protects the method for preparing the aforementioned compound, the synthetic route of which is as follows:

[0011]

[0012] The specific preparation method is as follows:

[0013] Step 1, Synthesis of compound QX-OH: Lidocaine and 2-bromoethanol were added to a reaction flask and heated to 100±5℃. After the reaction was completed, ethyl acetate was added to the reaction solution and stirred for 0.5±0.5h. The mixture was then filtered and washed to obtain compound QX-OH.

[0014] Step 2, Synthesis of compound Ace-QXOH: Under nitrogen protection, compound QX-OH was dissolved in N,N-dimethylformamide. Triethylamine was added to the reaction solution, and the mixture was stirred in an ice bath. After the reaction solution was cooled, acryloyl chloride was slowly added dropwise to the reaction solution. The reaction was allowed to proceed at room temperature. After the reaction was completed, ice water was added to the reaction solution to quench the reaction. The aqueous phase was extracted, the organic phase was collected, dried with anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified to obtain compound Ace-QXOH.

[0015] Step 3, Synthesis of compound S-QXOH: Compound Ace-QXOH was dissolved in acetonitrile, and acetylcysteine ​​and triethylamine were added to the reaction solution in sequence. The mixture was stirred at room temperature for 5 ± 5 min, the solvent was evaporated, and the crude product was purified to obtain compound S-QXOH.

[0016] Thirdly, this invention protects the use of the compounds described above in the preparation of local anesthetic drugs.

[0017] In a specific implementation plan, the anesthetic drug is a local anesthetic drug.

[0018] Preferably, the local anesthetic is a long-acting local anesthetic or a selective local anesthetic.

[0019] Fourthly, the present invention protects a pharmaceutical composition comprising the compounds described above.

[0020] In specific implementations, the pharmaceutical composition may further contain a pharmaceutically acceptable carrier or excipient.

[0021] Preferably, the pharmaceutically acceptable carrier or excipient is selected from sodium bicarbonate and physiological saline or water for injection.

[0022] In specific implementations, the pharmaceutical composition includes oral formulations, injectable formulations, transdermal formulations, or mucosal formulations.

[0023] This invention establishes a rat sciatic nerve block model. 0.6 mL of compound S-QXOH (30 mM) was injected into the sciatic nerve of rats. Ten minutes after injection, the rats exhibited normal pain sensation. Then, 0.3 mL of 3% H2O2 was injected into the sciatic nerve of rats. Ten minutes after injection of 3% H2O2, complete pain sensation blockade was achieved in the rats, and this blockade remained intact for 60 minutes. The experimental results demonstrate that the compound of this invention can induce prolonged sensory blockade in rats.

[0024] Beneficial effects

[0025] This invention provides a reactive oxygen species-responsive local anesthetic drug based on a quaternary ammonium lidocaine derivative, which can produce a long-lasting and low-toxicity anesthetic effect in vivo and is suitable for widespread application. Attached Figure Description

[0026] Figure 1 Results of sciatic nerve injection of compound S-QXOH (30mM) in 0.6mL; the 10min time point is the injection time of hydrogen peroxide.

[0027] Figure 2 To determine the survival rate of C2C12 mouse myoblasts after incubation with different concentrations of lidocaine, compound QX314, and S-QXOH.

[0028] Figure 3 To determine the survival rate of PC12 rat adrenal pheochromocytoma cells after incubation with different concentrations of lidocaine, compound QX314, and S-QXOH.

[0029] Figure 4 The staining results of muscle samples after injection of compounds QX314 and S-QXOH.

[0030] Figure 5 The results of sciatic nerve staining after injection of compound S-QXOH. Detailed Implementation

[0031] The present invention will be further described in detail below with reference to the embodiments. Reagents or instruments used without a specified manufacturer are considered to be conventional products that can be purchased on the market.

[0032] Example 1: Synthesis of compound S-QXOH

[0033]

[0034] F1: Synthesis of compound QX-OH. Lidocaine (5 g, 21.3 mmol) and 2-bromoethanol (1.5 mL, 21.3 mmol) were added to a reaction flask and heated to 100 °C. After the reaction was complete, 500 mL of ethyl acetate was added to the reaction solution and stirred for 0.5 h. The mixture was filtered and washed to obtain 7.3 g of QX-OH as a white solid, with a yield of 96%.

[0035] 1 H NMR (300MHz, Methanol-d4) δ7.14–7.07(m,3H),4.52(s,2H),4.07–3.98(m,2H),3.85–3.65(m,6H),2.24(s,6H),1.39(t,J=7.2Hz,6H). 13C NMR (75MHz, Methanol-d4) δ164.02,136.74,134.13,129.22,128.83,61.53,58.44,56.89,56.69,18.67,8.31.

[0036] F2: Synthesis of compound Ace-QXOH. Under nitrogen protection, QX-OH (5 g, 14 mmol) was dissolved in 30 mL of N,N-dimethylformamide. Triethylamine (2.9 mL, 21 mmol) was added to the reaction solution, and the mixture was stirred in an ice bath. After cooling, acryloyl chloride (1.36 mL, 16.8 mmol) was slowly added dropwise to the reaction solution, and the ice bath was removed, allowing the reaction to proceed at room temperature. After the reaction was complete, ice water was added to quench the reaction, and the aqueous phase was extracted with dichloromethane. The organic phase was collected, dried over anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified by column chromatography to give 3.5 g of compound Ace-QXOH, with a yield of 75.3%.

[0037] 1 H NMR (300MHz, DMSO-d6) δ10.20(s,1H),7.10(d,J=1.8Hz,3H),6.38(dd,J=17.2,1.7Hz,1H),6.23(dd,J=17.2,10.2Hz,1H),6.03(dd,J= 10.2,1.7Hz,1H),4.60(t,J=4.9Hz,2H),4.48(s,2H),3.97(t,J=5.0Hz,2H),3.65(q,J=7.1Hz,4H),2.16(s,6H),1.33(t,J=7.1Hz,6H).

[0038] F3: Synthesis of compound S-QXOH. Compound Ace-QXOH (0.5 g, 1.5 mmol) was dissolved in 5 mL of acetonitrile. Acetylcysteine ​​(0.24 g, 1.8 mmol) and triethylamine (0.31 mL, 2.2 mmol) were added sequentially to the reaction solution, and the mixture was stirred at room temperature for 5 min. The solvent was evaporated, and the crude product was purified by column chromatography to give 0.7 g of compound S-QXOH, with a yield of 94%.

[0039] 1H NMR(300MHz, Methanol-d4)δ7.08(d,J=3.8Hz,3H),4.59–4.54(m,1H),4.50(s,2H),4.38(dd,J=6.7,4.5Hz,1H),3.98(t,J=5.1Hz,2H),3 .76(q,J=7.2Hz,4H),3.18–2.99(m,2H),2.86–2.79(m,2H),2.69(dd,J=7.2,5.7Hz,2H),2.22(s,6H),1.94(s,3H),1.41(t,J=7.2Hz,6H).

[0040] Example 2: In vivo activity evaluation experiment

[0041] A rat sciatic nerve block model was established, and the hot plate test was used to determine whether sensory block occurred in the rats. In the control group, rats injected with 0.6 mL of 3% hydrogen peroxide solution showed no sensory or motor block at 10 and 30 min. When 0.6 mL of compound S-QXOH (30 mM) was injected into the sciatic nerve of rats, no sensory block occurred 10 min later. However, when 0.3 mL of 3% hydrogen peroxide solution was injected 10 min after the injection of 0.6 mL of compound S-QXOH (30 mM), complete sensory nerve block occurred 10 min later, and the sensory block persisted for 60 min. These experimental results demonstrate that the compound of this invention can induce prolonged sensory block in rats.

[0042] Example 3: Cytotoxicity Experiment

[0043] The toxicity of S-QXOH to muscle cells and nerve cells was evaluated using C2C12 mouse myoblasts (American Type Culture Collection (ATCC), Manassas, VA, USA) and PC12 rat adrenal pheochromocytoma cells (ATCC, Manassas, VA, USA), respectively.

[0044] C2C12 cells were seeded at a density of 1,000 cells per well in DMEM containing 20% ​​FBS and 1% penicillin-streptomycin, and then incubated for 10–14 days in DMEM containing 2% horse serum and 1% penicillin-streptomycin to differentiate into myotubules. PC12 cells were seeded at a density of 4,000 cells per well in DMEM containing 2.5% FBS, 12.5% ​​horse serum, and 1% penicillin-streptomycin, and then incubated together for 7 days in DMEM containing 1% horse serum, 50 ng / mL nerve growth factor, and 1% penicillin-streptomycin. The cytotoxicity of S-QXOH was assessed using the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazole (MTS) colorimetric assay. C2C12 and PC12 cells were treated with different concentrations of compounds S-QXOH and QX-314, respectively, with lidocaine (a commonly used local anesthetic) as a positive control. After 24 hours of incubation, 40 μL of MTS solution was added to each well, and the cells were incubated for another 4 hours. Finally, [the cells were then...]. Synergy TM Absorbance was measured at 490 nm using an Mx microplate reader (BioTek Inc., VT, USA). Viability is expressed as the percentage of untreated cells.

[0045] The experimental results revealed that ( Figures 2-3 Cells incubated with compound S-QXOH showed significantly higher survival rates than those incubated with compound QX-314 and the positive control lidocaine. This indicates that compound S-QXOH has significantly lower cytotoxicity than compounds QX-314 and lidocaine.

[0046] Example 4: Muscle and Nerve Tissue Toxicity Experiment

[0047] Rats were injected with 0.6 mL of compound S-QXOH (30 mM) and 0.6 mL of compound QX-314 (30 mM) for four and fourteen days, respectively, and then euthanized by inhalation of carbon dioxide to assess the inflammatory and toxic effects of compounds S-QXOH and QX-314. The sciatic nerve and adjacent connective tissue closest to the nerve block injection site were obtained using sterilized surgical instruments. Muscle samples were fixed in 10% neutral buffered formalin and embedded in paraffin. After sectioning, the muscle tissue was stained with hematoxylin and eosin using standard techniques. Each section was examined blinded using an optical microscope. The rat sciatic nerve was fixed in Karnovsky's KII solution, treated accordingly, embedded in resin, and stained with toluidine blue. For the objectivity of the experimental results, the experimenter evaluated them blinded using an optical microscope. The experimental results show that ( Figures 4-5Compound S-QXOH has lower muscle toxicity than compound QX-314, and compound S-QXOH has no neurotoxicity.

[0048] The scope of protection of this invention is not limited to the above embodiments. Variations and advantages that can be conceived by those skilled in the art without departing from the spirit and scope of the inventive concept are included in this invention and are protected by the appended claims.

Claims

1. A compound having the following structure: 。 2. The method for preparing the compound according to claim 1, characterized in that, Its synthetic route is as follows: The specific preparation method is as follows: Step 1, Synthesis of compound QX-OH: Lidocaine and 2-bromoethanol were added to a reaction flask and heated to 100±5 °C. After the reaction was completed, ethyl acetate was added to the reaction solution and stirred for 0.5±0.5 h. The mixture was then filtered and washed to obtain compound QX-OH. Step 2, Synthesis of compound Ace-QXOH: Under nitrogen protection, compound QX-OH was dissolved in N,N-dimethylformamide. Triethylamine was added to the reaction solution, and the mixture was stirred in an ice bath. After the reaction solution was cooled, acryloyl chloride was slowly added dropwise to the reaction solution. The reaction was allowed to proceed at room temperature. After the reaction was completed, ice water was added to the reaction solution to quench the reaction. The aqueous phase was extracted, the organic phase was collected, dried with anhydrous sodium sulfate, filtered, and concentrated. The crude product was purified to obtain compound Ace-QXOH. Step 3, Synthesis of compound S-QXOH: Compound Ace-QXOH was dissolved in acetonitrile, and acetylcysteine ​​and triethylamine were added to the reaction solution in sequence. The mixture was stirred at room temperature for 5 ± 5 min, the solvent was evaporated, and the crude product was purified to obtain compound S-QXOH.

3. The use of the compound of claim 1 in the preparation of local anesthetic drugs.

4. The application according to claim 3, characterized in that, The local anesthetic drug is a long-acting local anesthetic drug or a selective local anesthetic drug.

5. A pharmaceutical composition, characterized in that, The pharmaceutical composition comprises the compound of claim 1.

6. The pharmaceutical composition according to claim 5, characterized in that, The pharmaceutical composition also contains pharmaceutically acceptable excipients.

7. The pharmaceutical composition according to claim 6, characterized in that, The pharmaceutically acceptable excipients are sodium bicarbonate, physiological saline, or water for injection.

8. The pharmaceutical composition according to any one of claims 5-7, characterized in that, The pharmaceutical composition is an oral preparation, an injectable preparation, a transdermal preparation, or a mucosal preparation.

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