阅读说明：本技术 作为kcnq增强剂的1-((2-(2,2,2-三氟乙氧基)吡啶-4-基)甲基)脲衍生物 (1- ((2- (2,2, 2-trifluoroethoxy) pyridin-4-yl) methyl) urea derivatives as KCNQ enhancers ) 是由 H.J.塞克雷斯 A.C.威廉姆斯 M.A.沃顿 于 2020-02-04 设计创作，主要内容包括：本申请涉及钾通道的小分子增强剂(例如Kv7增强剂——也称为KCNQ增强剂)、包含此类化合物的组合物,以及使用此类化合物治疗由运动神经元兴奋性变化引起的肌萎缩性侧索硬化和其它神经系统疾病的方法,所述疾病包括但不限于原发性侧索硬化、假性延髓麻痹、进行性延髓麻痹、进行性肌肉萎缩和癫痫。(The present application relates to small molecule enhancers of potassium channels (e.g., Kv7 enhancers, also known as KCNQ enhancers), compositions comprising such compounds, and methods of using such compounds to treat amyotrophic lateral sclerosis and other neurological diseases caused by motor neuron excitatory changes, including, but not limited to, primary lateral sclerosis, pseudobulbar palsy, progressive bulbar palsy, progressive muscle atrophy, and epilepsy.)

1. A compound of the formula:

wherein R1 is

And wherein R2 is H or OH.

2. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R1 is:

3. the compound of claim 2, or a pharmaceutically acceptable salt thereof, wherein R2 is OH.

4. A compound according to claim 3, or a pharmaceutically acceptable salt thereof, of the formula:

wherein the compound comprises a single enantiomer having an optical rotation of (+) in methanol.

5. A compound according to claim 3, or a pharmaceutically acceptable salt thereof, of the formula:

wherein the compound comprises a single enantiomer having (-) optical rotation in methanol.

6. The compound of claim 2, or a pharmaceutically acceptable salt thereof, wherein R2 is H.

7. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, wherein R1 is

8. The compound of claim 7, or a pharmaceutically acceptable salt thereof, wherein R2 is H.

9. The compound of claim 7, or a pharmaceutically acceptable salt thereof, wherein R2 is OH.

10. A compound according to claim 9, or a pharmaceutically acceptable salt thereof, of the formula:

11. a compound according to claim 9, or a pharmaceutically acceptable salt thereof, of the formula:

12. a pharmaceutical composition comprising a compound of any one of claims 1-11, or a pharmaceutically acceptable salt thereof, together with one or more pharmaceutically acceptable carriers, diluents, or excipients.

13. A method of treating a disease caused by motor neuron excitatory changes, comprising administering to a patient in need thereof an effective amount of a compound according to any one of claims 1-11, or a pharmaceutically acceptable salt thereof.

14. The method of claim 13, wherein the disease caused by a change in motor neuron excitability is ALS.

15. A method of treating a disease caused by motor neuron excitatory changes, comprising administering to a patient in need thereof the pharmaceutical composition of claim 12.

16. The method of claim 15, wherein the disease caused by a change in motor neuron excitability is ALS.

17. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for use in therapy.

18. A compound according to any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, for use in the treatment of a disease caused by a change in motor neuron excitability.

19. The compound of claim 18, or a pharmaceutically acceptable salt thereof, wherein the disease caused by a change in motoneuron excitability is ALS.

20. Use of a compound of any one of claims 1-11, or a pharmaceutically acceptable salt thereof, in the manufacture of a medicament for treating a disease caused by a change in motor neuron excitability.

21. The compound or pharmaceutically acceptable salt thereof for use according to claim 20, wherein the disease caused by a change in motor neuron excitability is ALS.

22. A process for preparing a pharmaceutical composition comprising mixing a compound of any one of claims 1-11, or a pharmaceutically acceptable salt thereof, with one or more pharmaceutically acceptable carriers, diluents, or excipients.

23. A compound, or a pharmaceutically acceptable salt thereof, of the formula:

24. a compound, or a pharmaceutically acceptable salt thereof, of the formula:

wherein the compound comprises a single enantiomer having an optical rotation of (+) in methanol.

25. A compound, or a pharmaceutically acceptable salt thereof, of the formula:

Stock solutions of 1-mg/mL of example 1 were prepared and serially diluted into pooled rat blood to prepare standards ranging from 1 to 10000 ng/mL. Blood was added to a blank DBS card to prepare a standard. A3 mm long DBS standard or sample was added to a 96 well plate followed by 180. mu.L of Internal Standard (IS) in 1:1ACN: MeOH. After shaking for 45min, the extract was diluted 2-fold with water and analyzed for drug concentration by LC/MSMS.

1.14mL of MeOH H was used₂O (2:8) brain samples were homogenized. Standards were prepared by adding (spiking) stock solutions to a series of blank brain homogenates ranging from 5 to 50000 ng/mL. mu.L of standard or sample was transferred into a 96-well plate and 180. mu.L of Internal Standard (IS) in 1:1ACN: MeOH was added and mixed. The samples were centrifuged at 4000RPM for 10 minutes at 4 ℃. The supernatant was diluted 15-fold with water and analyzed by LC/MSMS.

Samples and standards were analyzed using a Sciex API 4000 triple quadrupole mass spectrometer (Sciex, Division of MDS inc., Toronto, Canada) in combination with a Shimadzu HPLC system (LC-IOAD, Shimadzu Corporation) and a Gilson 215 autosampler. Samples (0.01mL) were injected into a 5- μm Betasil C-18, 20X2.1mm Javelin (Thermo Electron Corp. Cat #70105-022106) HPLC column and eluted with a gradient. Chromatographic conditions include water/1 MNH of mobile phase A₄HCO₃(2000:10, v/v) and MeOH/1M N NH of mobile phase B₄HCO₃(2000:10, v/v) run with a gradient of 2.5-min, flow rate of 1.5 mL/min. Positive ion mode with turbo spray and ion source temperature of 740 ℃ were used for mass spectrometric detection. Quantification was performed using Multiple Reaction Monitoring (MRM) in the following transitions: quantification was performed using Multiple Reaction Monitoring (MRM) in the following transitions: example 1(m/z350.2 to m/z 233.0) and a simulated internal standard (m/z 263.1 to m/z 148.1). The linear regression plot of the peak area ratio of compound to internal standard compared to drug concentration was given quadratically at 1/x 2. Linear regression plot of peak area ratio of compound to internal standard versus drug concentration from 1/x²The square is obtained.

The simulated internal standard used was 2- (dimethylamino) -N-pentyl-3-phenyl-propionamide 2,2, 2-trifluoroacetic acid (1:1) and it has the following structure:

the analog internal standard was purchased from Syncom, a company from The Netherlands and addressed by Kadijk 3, 9747AT Groningen, The Netherlands.

After adding (spiking) the drug to these matrices and incubating for 4.5 hours at 37 ℃ with orbital shaking, the binding of the drug to rat plasma proteins and brain homogenates was determined using an in vitro dialysis method. The assay was performed using a HT dialysis microbalance device and dialysis membrane tape (MWCO 12-14 k). Samples with time 0 were taken after the protein matrix and samples were taken from the protein and buffer sides of the membrane after 4.5 hours of incubation. The parent was quantified by LC-MSMS at time points of 0 and 45 minutes. The fraction of unbound was calculated by dividing the concentration on the buffer side by the concentration on the protein side. Percent recovery can also be calculated by dividing the sum of the buffer and protein compartments by the time 0 concentration after 4.5 hours. Unbound compound concentration was calculated using total concentration x fraction unbound.

As a result: the effect of example 1 on the absolute threshold is shown in Table 4

TABLE 4

CMAP is a compound muscle action potential

(for more information on this assay, see R.Sittl et al, "The Kv7 porous catalyst formulations of secreted axos in isolated rat neural," Journal of The personal neural systems 15: 63-72 (2010), "M.Kovalchuk et al," Acute Effects of canal and recovery in tissues With intestinal around, synthetic bacterial crystals: ARanhybrid, Double-Medium, Placebo-Controlled, Crossful, "Tribute 7 day received 3/7 day 2018; online 13 day 2018 publication of biological molecules: Microphal 2018: Micropter, III, J.P.A.: U.S. Pat. No. 11 & 7. J.S.: U.S.: 1. Pat. No. 7: 7. A. The first publication of mineral substances of molecular substances, U.S.: 1. Pat. No. 7: 7. A.S.: U.S.: 1, U.S.: 1. A.S.: this application, U.S.: 1. A.S.: this publication of patent application No. 7, 3. A.S. 7, 3. publication of biological samples, 3. A.S. Pat. No. 3, et al.

There were significant differences in time and time-treatment interactions (two-way RM ANOVA; time effect F (26, 312) ═ 13.18, p ═ 0.0001; time-treatment interaction F (78, 312) ═ 2.888, p < 0.0001. Bonferroni multiple comparison tests showed that 30mg/kg of example 1 significantly increased the absolute threshold (decreased excitability) compared to vehicle XE-991 was able to reverse this increase (increased excitability).