CN112451527B

CN112451527B - Application of aminopyrimidine compounds

Info

Publication number: CN112451527B
Application number: CN202011402371.4A
Authority: CN
Inventors: 张培龙; 兰文丽
Original assignee: Beijing Anshi Biotechnology Co ltd
Current assignee: Beijing Anshi Biotechnology Co ltd
Priority date: 2020-12-02
Filing date: 2020-12-02
Publication date: 2022-08-16
Anticipated expiration: 2040-12-02
Also published as: CN112451527A

Abstract

The invention belongs to the technical field of biomedicine, and particularly relates to application of aminopyrimidine compounds, in particular to novel application of aminopyrimidine compounds or pharmaceutically acceptable forms thereof in medicaments for treating diseases caused by one or more insertion or deletion mutations in an exon 20 structure domain of EGFR or HER 2.

Description

Application of aminopyrimidine compounds

Technical Field

The invention relates to an amino-piperidine compound capable of regulating activity of ErbB family kinase, in particular to a novel application of the amino-piperidine compound or a pharmaceutically acceptable form thereof in medicines for treating diseases caused by one or more insertion or deletion mutations in exon 20 domain of EGFR or HER 2.

Background

EGFR and HER2 are receptor tyrosine kinases ErbB receptor family members. Dimerization of ErbB receptors results in phosphorylation of certain amino acid residues in the intracellular domain and activation of multiple signaling mechanisms. Dysregulation of ErbB family signaling can lead to cell proliferation, invasion, metastasis and angiogenesis. Currently, many drugs targeting EGFR or HER2 have been used in cancers such as lung cancer, head and neck cancer, and breast cancer.

About 30-40% of non-small cell lung cancer patients of asian descent have EGFR mutations. EGFR mutations occur predominantly at exons 8-21, which encode the kinase domain, with 90% of EGFR mutations resulting from either a 19 exon deletion or a 21 exon L858R mutation. The EGFR 20 exon insertion mutation accounts for about 4-9.2% of patients with EGFR mutation, and most EGFR 20 exon insertion occurs in the coding region between amino acids 767-774.

EGFR 20 exon insertion mutation is insensitive to clinical approved reversible or irreversible EGFR kinase inhibitors such as gefitinib, erlotinib, afatinib and oxitinib, and a novel kinase inhibitor with higher drug effect and selectivity is clinically needed. In other words, there is a need in the art for compounds that exhibit high inhibition of EGFR 20 exon, while exhibiting relatively low inhibition of wild-type EGFR, thereby reducing clinical adverse events (e.g., skin rash and/or diarrhea) associated with inhibition of wild-type EGFR while exerting anti-cancer efficacy.

HER 220 exon insertion mutation accounts for about 2-4% of non-small cell lung cancer, wherein A775_ G776insYVMA accounts for more than 70%. Like EGFR 20 exon, clinically effective drugs targeting HER 220 exon insertion mutations are also lacking.

Disclosure of Invention

In order to solve the problems, the invention provides a series of amino-melidines compounds which have high inhibitory activity on insertion mutation of EGFR 20 exon and HER 220 exon.

In one aspect, the present invention provides a novel use of the following compounds or pharmaceutically acceptable salts thereof for the manufacture of a medicament for the treatment of diseases caused by one or more insertion or deletion mutations in the exon 20 domain of EGFR or HER 2.

In another aspect, the invention provides the use as described above, wherein the cancer is lung cancer, colorectal cancer, pancreatic cancer, head and neck cancer, breast cancer, ovarian cancer, uterine cancer or gastric cancer.

In another aspect, the invention provides the use as described above, wherein the cancer is non-small cell lung cancer.

Detailed Description

Therapeutic uses of the compounds herein

The present invention provides a compound or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the treatment of cancer associated with one or more insertion or deletion mutations in the exon 20 domain of EGFR or HER 2.

"pharmaceutically acceptable salt" refers to conventional acid addition salts or base addition salts that retain the biological effectiveness and properties of the compound, which are formed from suitable non-toxic organic or inorganic acids or organic or inorganic bases. Examples of acid addition salts include those derived from inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, sulfamic acid, phosphoric acid and nitric acid, and those derived from organic acids such as p-toluenesulfonic acid, salicylic acid, methanesulfonic acid, oxalic acid, succinic acid, citric acid, maleic acid, lactic acid, fumaric acid, and the like. Examples of base addition salts include salts derived from ammonium, potassium, sodium and quaternary ammonium hydroxides, such as tetramethylammonium hydroxide. Chemical modification of a pharmaceutical compound (i.e., drug) into a salt is a technique well known to pharmaceutical chemists to achieve improved physical and chemical stability, hygroscopicity, flowability, and solubility of compounds.

Examples

The present disclosure is further described in conjunction with the following examples, which are included to explain and illustrate the disclosure, and not to limit the disclosure.

Preparation examples

The compounds 1 to 74 of the invention and the comparative compounds 75 and 76 were obtained according to the test method of Chinese patent CN 108707139.

Active examples

In the experiment, a celltiter-glo (CTG) method is adopted to measure the inhibition effect of the compounds on the growth of 7 strains of cells (Ba/F3 EGFR-H773-V774ins _ NPH, Ba/F3 EGFR-D770-N771ins _ SVD, Ba/F3 EGFR-V769_ D770insASV and Ba/F3 EGFR-WT), and the 50% inhibition concentration IC50 is calculated. The test used AZD9291 as a positive control compound.

1 design of the experiment

Compounds were assayed on selected cells and vehicle controls were set for a total of 9 concentrations, 3 replicates per concentration.

2 reagents and consumables

1) Fetal bovine serum FBS (GBICO)

2)

Luminescent Cell Viability Assay(Promega)

3) 96-hole transparent flat-bottom black wall plate

3 apparatus

SpectraMax multi-mark microplate detector, PerkinElmer, 2104-0010A;

CO ₂ incubator, Thermo Scientific, Model 3100 Series;

biological safety cabinet, Thermo Scientific, Model 1300Series a 2;

inverted microscope, Olympus, CKX41 SF;

4 Experimental methods

Cell culture and inoculation

1) Cells in the logarithmic growth phase were harvested and counted using a cell counter. Cell viability was checked by trypan blue exclusion to ensure that the viability of each cell line was above 90%.

2) Adjusting the cell concentration; 90 μ L of cell suspension was added to 96-well plates, respectively.

3) Cells in 96-well plates were incubated overnight at 37 ℃ with 5% CO2 and 95% humidity.

T0 reading board

1) 10 μ L of PBS per well was added to T0 plates seeded with cells.

2) The CTG reagents were thawed and the cell plates were equilibrated to room temperature for 30 minutes.

3) An equal volume of CTG solution was added to each well.

4) Cells were lysed by shaking on an orbital shaker for 5 minutes.

5) The cell plate was left at room temperature for 20 minutes to stabilize the luminescence signal.

6) The cold light value of T0 was read.

Dosing

1) A10 mM stock solution was prepared by adding the corresponding volume of DMSO to the corresponding dry compound powder according to the test drug table.

2) A 1000 x, 3.16 fold gradient dilution of the drug solution was prepared as referenced in appendix I.

3) The 1000 Xgradient diluted drug solution was diluted 100 times with PBS to make 10 times drug solution, the highest concentration was 100 μ M, 9 concentrations, 3.16 times dilution, 10 μ L drug solution was added to each well of 96-well plate inoculated with cells, three multiple wells were set for each drug concentration, and the final concentration of DMSO was 0.1%.

4) The cells in the dosed 96-well plate were incubated for a further 72 hours at 37 ℃ under 5% CO2 and 95% humidity, after which they were subjected to CTG analysis.

Terminal reading board

1) The CTG reagents were thawed and the cell plates were equilibrated to room temperature for 30 minutes.

2) An equal volume of CTG solution was added to each well.

3) Cells were lysed by shaking on an orbital shaker for 5 minutes.

4) The cell plate was left at room temperature for 20 minutes to stabilize the luminescence signal.

5) And reading the cold light value.

Data processing

Data were analyzed using GraphPad Prism 5.0 software, fitted to the data using non-linear sigmoidal regression to derive a dose-effect curve, and IC was calculated therefrom ₅₀ The value is obtained.

Cell survival rate (%) ═ (Lum) _{Drug to be tested} -Lum _{Culture fluid control} )/(Lum _{Cell controls} -Lum _{Culture fluid control} )×100％.

Results of the experiment

The data of the assay are shown in Table 1, wherein Ba/F3 EGFR WT assay is not added with EGF.

TABLE 1

And (4) test conclusion: compounds 1-74 had significantly lower IC50 values at EGFR 20 exon and HER 220 exon insertion mutations than control compounds 75, 76 and AZD 9291. Compounds 1-74 were effective in inhibiting cell proliferation of EGFR 20 exon and HER 220 exon insertion mutations, and exhibited superior cellular activities to 75, 76 and AZD 9291.

Claims

1. Use of a compound selected from the group consisting of:

2. the use of claim 1, wherein the disease comprises a tumor.

3. The use according to claim 1, wherein the disease comprises lung cancer, colorectal cancer, pancreatic cancer, head and neck cancer, breast cancer, ovarian cancer, uterine cancer or gastric cancer.

4. The use according to claim 1, wherein the disease is non-small cell lung cancer.