Preparation method and application of fluorene carboxamide compound

Abstract

The invention relates to an N-((2-(2, 6-dioxopiperidine-3-yl)-1-oxoisoindoline-5-yl) methyl)-9, 9-dimethyl-9H-fluorene-2-formamide compound as shown in a formula (I) or a stereoisomer, a racemate, a deuterated substance, a solvate, a prodrug, a metabolite, a pharmaceutically acceptable salt or eutectic crystal thereof, as well as an intermediate and a preparation method of the N-((2-(2, 6-dioxopiperidine-3-yl)-1-oxoisoindoline-5-yl) methyl)-9, 9-dimethyl-9H-fluorene-2-formamide compound. The invention further discloses application of the compound in preparation of drugs for treating GSPT1 related diseases.

Description

Technical Field

[0001] This invention relates to the field of medicinal chemistry, specifically to methods for preparing fluorenylformamide compounds and their uses. Background Technology

[0002] Molecular glue degraders work by binding to CRBN and then recruiting protein substrates. After ubiquitination, they can be degraded by the proteasome. Unlike traditional protein inhibitors, the effect of molecular glue on proteins is irreversible. Therefore, compared with traditional protease inhibitors, molecular glue is more effective in cell function.

[0003] Currently, molecular adhesives in the clinical stage include MRT2359 from Monte Rosa and FD001 from Chengdu Fendi. Due to the accidental nature of the discovery of molecular adhesives, their rational design presents certain challenges.

[0004] This invention designs and synthesizes a compound that, compared to the clinical compound CC90009, provides a novel structure that can potently inhibit NCI-H1299 non-small cell lung cancer cells, offering more new structures for the discovery of molecular glues. Summary of the Invention

[0005] The present invention provides a compound of formula (I), its deuterated derivative, its stereoisomer, its tautomer, or a mixture thereof.

[0006] 1. The compound N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-9,9-dimethyl-9H-fluorene-2-carboxamide has the structure shown in formula (I):

[0007]

[0008] In the embodiments, the compound of formula I, its deuterated derivatives, its stereoisomers, its tautomers, or mixtures thereof, is characterized in that the compound is selected from, but is not limited to, the following compounds:

[0009]

[0010] Another aspect of the present invention is to provide the use of the above-described compounds in the preparation of medicaments for treating or preventing diseases related to GSPT1, CK1a, or IKZF1 / 2 / 3; preferably diseases related to GSPT1 dysfunction.

[0011] In some implementations, the disease associated with GSPT1 is cancer.

[0012] In some preferred embodiments, the cancer includes solid tumors or hematologic malignancies.

[0013] In some more preferred embodiments, the cancer is selected from, but not limited to: bladder cancer, bone cancer, brain cancer, breast cancer, cervical cancer, chest cancer, colon cancer, rectal cancer, colorectal cancer, endometrial cancer, esophageal cancer, eye cancer, head cancer, kidney cancer, liver cancer, lymph node cancer, lung cancer, upper respiratory tract and digestive tract cancers, cervical cancer, ovarian cancer, pancreatic cancer, prostate cancer, thyroid cancer, rectal cancer, skin cancer, neuroblastoma, glioblastoma, hemangiopericytoma, multiple brain metastases, and polymorphic glioma. Glioblastoma, glioblastoma, brainstem glioma, malignant brain tumors with poor prognosis, malignant glioma, recurrent malignant glioma, anaplastic astrocytoma, anaplastic oligodendroglioma, neuroendocrine tumors, Kaposi's sarcoma, malignant melanoma, malignant mesothelioma, malignant pleural effusion mesothelioma syndrome, peritoneal carcinoma, serous carcinoma, gynecological sarcoma, soft tissue sarcoma, scleroderma, leiomyosarcoma, fallopian tube carcinoma, leukemia, lymphoma, leukemia. Detailed Implementation

[0014] To make the present invention easier to understand, the present invention will be described in detail below with reference to embodiments. These embodiments are for illustrative purposes only and are not limited to the scope of application of the present invention.

[0015] Unless otherwise defined, the technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure pertains.

[0016] Terms and definitions:

[0017] The terminology used in this invention is explained below. If the meaning of a particular term differs from that commonly understood by those skilled in the art, the meaning as used in this invention shall prevail. If not defined in this invention, it shall have the meaning commonly understood by those skilled in the art. Unless otherwise stated, the terms used in this invention have the following meanings:

[0018] The compounds of this invention may contain an asymmetric center, and thus may produce enantiomers, diastereomers, and other stereoisomers. This invention is intended to include all such possible isomers, as well as their racemic and optically pure forms, whether or not they are specifically described herein. Optically active (+) and (-), (R)- and (S)-, or (D)- and (L)- isomers can be prepared using chiral synthesis or chiral reagents, or resolved using conventional techniques, such as chromatography and fractional crystallization. Conventional techniques for preparing / separating individual enantiomers include chiral synthesis from suitable optically pure precursors or resolution of racemic mixtures (or racemic mixtures of salts or derivatives) using, for example, chiral high-performance liquid chromatography (HPLC). When the compounds described herein contain an alkene double bond or other geometrically asymmetric center, unless otherwise stated, the compound is intended to include both E and Z geometric isomers. Similarly, all tautomers are also intended to be included.

[0019] "Stereoisomer" refers to a stable isomer that has a vertical asymmetric plane due to having at least one chiral element (including chiral center, chiral axis, chiral surface, etc.), thereby enabling plane-polarized light to rotate; this disclosure considers various stereoisomers and mixtures thereof, and includes "enantiomers".

[0020] "Tautomer" (or "tautomer form") refers to structural isomers with different energies that can be interconverted through a low energy barrier. For example, proton tautomers (or proton transfer tautomers) include (but are not limited to) interconversions through proton transfer, such as keto-enol isomerization, imine-enamine isomerization, amide-imine alcohol isomerization, etc. This disclosure includes tautomers of any of the compounds described.

[0021] Furthermore, the compounds of the present invention may contain one or more isotopic forms, namely, hydrogen isotopes of D or T, or isotopes of any atom, such as all isotopes of C and N. Any isotopic forms of C, N, or H in compounds with optional structures of the present invention are covered within the scope of protection of the present invention. The terms "comprising," "including," and "containing," and their equivalents, should be understood in an open, non-exclusive sense, meaning "including but not limited to," implying that in addition to the listed elements, components, and steps, other unspecified elements, components, and steps may also be covered.

[0022] The raw materials or intermediates used in this article can be prepared by or in accordance with methods known in the art or by the methods disclosed herein, or can be purchased from commercial entities.

[0023] In this invention, column chromatography generally uses Yantai Huanghai silica gel 200-300 mesh silica gel as the carrier.

[0024] Unless otherwise specified in this invention, the solutions mentioned in the reactions of this invention are aqueous solutions.

[0025] In this invention, "room temperature" refers to a temperature between 10°C and 25°C. Attached Figure Description

[0026] Figure 1 The antitumor activity of NCI-H1299 was demonstrated in CC90009 (IC50 > 10 μM) and Example 01 (IC50 = 31.9 nM). Detailed Implementation

[0028] The present invention will be further described in detail below with reference to the embodiments, but this is not intended to limit the present invention. Any equivalent substitutions made in the art based on the disclosure of the present invention shall fall within the protection scope of the present invention.

[0029] The structure of the compound was determined by mass spectrometry (MS) or nuclear magnetic resonance (NMR). 1 It was determined by HNMR.

[0030] Nuclear magnetic resonance (NMR) 1 HNMR shift (δ) is given in parts per million (ppm); nuclear magnetic resonance (NMR) 1 The ¹H NMR measurements were performed using a Bruker AVANCE-600 NMR spectrometer. The solvent was deuterated dimethyl sulfoxide (DMSO-d6), and the internal standard was tetramethylsilane (TMS). Chemical shifts were expressed as 10⁻¹⁰ NMR values. -6 (ppm) is given as the unit.

[0031] Mass spectrometry (MS) measurements were performed using a FINNIGAN LCQAd (ESI) mass spectrometer (manufacturer: Spectrum Technology, model: Spectrum 5210).

[0032] Thin-layer silicone uses Yantai Huanghai HSGF254 or Qingdao GF254 silicone sheets.

[0033] Column chromatography typically uses Yantai Huanghai silica gel with a mesh size of 200-300 as the carrier.

[0034] Unless otherwise specified in this invention, the solution mentioned in the reaction of this invention is an aqueous solution.

[0035] In this invention, "room temperature" refers to a temperature between 10°C and 25°C.

[0036] Example 01: Preparation of N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-9,9-dimethyl-9H-fluorene-2-carboxamide (Example 01)

[0037]

[0038] The preparation route is as follows:

[0039]

[0040] 9,9-Dimethyl-9H-fluorene-2-carboxylic acid (38 mg, 0.16 mmol), 3-(5-(aminomethyl)-1-oxoisoindolin-2-yl)piperidine-2,6-dione hydrochloride (50 mg, 0.16 mmol), HATU (92 mg, 0.24 mmol), TEA (49 mg, 0.48 mmol), and DMF (5 mL) were added to a round-bottom flask and stirred at room temperature for 5 h. The reaction was monitored by LCMS until the starting material was completely reacted. 20 mL of water was added to the reaction flask, and a large amount of white solid precipitated out. The solid was filtered to obtain the white target compound (60 mg, yield = 76%, purity = 94%).

[0041] ESI-MS: m / z [M+H] + =493.75;

[0042] 1 H NMR(600MHz,DMSO-d6)δ10.98(s,1H),9.16(t,J=5.9Hz,1H),8.10(s,1H),7.93(s,2H) ,7.91–7.87(m,1H),7.71(d,J=7.8Hz,1H),7.61–7.56(m,2H),7.50(d,J=7.9Hz,1H),7 .39-7.35(m,2H),5.15-5.04(m,1H),4.63(d,J=5.8Hz,2H),4.48-4.26(m,2H),2.92–2 .87(m,1H),2.60(d,J=16.7Hz,1H),2.43-2.32(m,1H),2.03-1.94(m,1H),1.47(s,6H).

[0043] Biological Experiment

[0044] Experiment 1: Cell Experiment (NCI-H1299 Cell Proliferation Inhibition Assay)

[0045] NCI-H1299 cells were placed in McCoys-5A (Gibco) medium (containing 10% fetal bovine serum; ExCell bio) with the addition of 1% penicillin and streptomycin, and cultured in a cell culture incubator at 37°C, 5% CO2, and saturated humidity. After confluence, the cells were passaged into 96-well plates (5000 cells / well) and cultured overnight. Then, compounds at concentrations of 0.04 nM, 0.15 nM, 0.61 nM, 2.44 nM, 9.77 nM, 39.1 nM, 156.3 nM, 625 nM, 2500 nM, and 10000 nM were added to the cells. 0.1% DMSO and culture medium were used as solvents and negative controls, respectively. Each sample was tested in duplicate. After mixing the compounds thoroughly, the cells were cultured at 37°C and 5% CO2 for 72 hours. After treatment, the 96-well plate was placed at room temperature for 30 minutes, and then 50 μL of [unspecified substance] was added. (Promega) was added and vortexed for 2 min, then incubated at room temperature in the dark for 10 min. The 96-well plate was then placed in a SpectraMax Paradigm (Molecular Devices) to measure the Luminance signal. Cell viability inhibition rate (%) = 100 - (RLUcompound - RLUblank) / (RLUcontrol - RLUblank) * 100%, where Control was the 0.1% DMSO treatment group and blank was the culture medium treatment group. The IC50 of the compound was calculated using the nonlinear regression function "Dose-Response-Inhibition" in Graphpad Prism 8.0. 50 The dose-response relationship curve was plotted, and the experimental results are shown in Table 1.

[0046] Table 1: Results of cell proliferation inhibition experiments using exemplary compounds

[0047]

[0048] The experimental results show that the compound of this invention has a good inhibitory effect on NCI-H1299 cells, which is significantly better than the positive control CC90009. The experimental results are as follows: Figure 1 As shown.

[0049] The above embodiments are merely illustrative of implementation methods of the present invention. However, the present invention is not limited to the above embodiments. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the protection scope of the present invention.

[0050] Figure 1The NCI-H1299 antitumor activity graphs of CC90009 (IC50>10uM) and Example 01 (IC50=31.9nM) show that the antitumor activity of Example 01 is significantly better than that of the positive compound CC90009.

Claims

1. The compound N-((2-(2,6-dioxopiperidin-3-yl)-1-oxoisoindoline-5-yl)methyl)-9,9-dimethyl-9H-fluorene-2-carboxamide has the structure shown in formula (I):

2. Use of the compound of claim 1, its deuterated form, its stereoisomer, its tautomer, or mixture thereof in the preparation of a medicament for the treatment or prevention of diseases associated with or causing IKZF1, IKZF2, CK1a, or GSPT1.

3. In the use according to claim 2, the disease associated with IKZF1, IKZF2, CK1a, or GSPT1 is cancer; preferably, the cancer includes solid tumors or hematologic malignancies; more preferably, the cancer is selected from bladder cancer, bone cancer, brain cancer, breast cancer, cervical cancer, chest cancer, colon cancer, rectal cancer, colorectal cancer, endometrial cancer, esophageal cancer, eye cancer, head cancer, kidney cancer, liver cancer, lymph node cancer, lung cancer, upper respiratory tract and digestive tract cancers, cervical cancer, ovarian cancer, pancreatic cancer, prostate cancer, thyroid cancer, rectal cancer, skin cancer, and neurogenic cancer. Cell tumors, glioblastoma, hemangiopericytoma, multiple brain metastases, glioblastoma multiforme, glioblastoma, brainstem glioma, malignant brain tumors with poor prognosis, malignant glioma, recurrent malignant glioma, anaplastic astrocytoma, anaplastic oligodendroglioma, neuroendocrine tumors, Kaposi's sarcoma, malignant melanoma, malignant mesothelioma, malignant pleural effusion mesothelioma syndrome, peritoneal carcinoma, serous carcinoma, gynecological sarcoma, soft tissue sarcoma, scleroderma, leiomyosarcoma, fallopian tube carcinoma, leiomyosarcoma, leukemia, lymphoma, leukemia.

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