CN115925687A - EGFR (epidermal growth factor receptor) -targeting compound, PET (polyethylene terephthalate) molecular probe and preparation method and application thereof

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Abstract

Description

Claims

CN115925687A

Publication number: CN115925687A
Application number: CN202211707648.3A
Authority: CN
Inventors: 邵丹; 程佑; 李学飞
Original assignee: Guangdong General Hospital
Current assignee: Guangdong General Hospital
Priority date: 2022-12-29
Filing date: 2022-12-29
Publication date: 2023-04-07
Anticipated expiration: 2042-12-29
Also published as: CN115925687B

The invention discloses a compound targeting EGFR, a PET molecular probe, a preparation method and an application thereof, wherein the structure of the compound is shown as the formula (I):

the EGFR-targeting compound and the PET molecular probe can target EGFR-T790M mutation, and have the advantages of strong specificity, high sensitivity, popularization and application and the like.

EGFR (epidermal growth factor receptor) -targeting compound, PET (polyethylene terephthalate) molecular probe and preparation method and application thereof

Technical Field

The invention belongs to the field of radiochemistry, and particularly relates to an EGFR (epidermal growth factor receptor) -targeted compound, a PET (polyethylene terephthalate) molecular probe, and a preparation method and application thereof.

Background

The patient is injected with a low dose of radioactive molecular probe capable of targeting a specific tumor marker, and positron emission tomography (PET-CT) can provide information such as the position, the size, the shape and the like of the tumor in a noninvasive, real-time, dynamic and high-sensitivity manner. The development of EGFR-TKIs therapeutic drugs promotes the development of PET molecular probes based on the mother nucleus structure of EGFR-TKIs therapeutic drugs, and the PET molecular probes can be mainly classified into ¹¹ C、 ¹⁸ F and ⁶⁸ three major classes of Ga labels. At present, the number of the current day, ¹¹ c-labeled third-generation EGFR-TKIs oxitinib (Osimetinib) is a probe which can be used for detecting EGFR-T790M genes and entering clinical experiments. The existing EGFR PET molecular probe has the following defects: (1) Poor specificity, inability to distinguish between common and rare mutations of EGFR (T790M); (2) The sensitivity is low, and the tumor tissue is difficult to sensitively and accurately identify because the normal tissue has higher radiopharmaceutical uptake degree; (3) The ability of penetrating the blood brain barrier can not be rapidly and well realized, and the diagnosis and the evaluation of the curative effect of the patient with brain metastasis NSCLC are difficult to be realized; (4) ¹¹ C used due to its short half-life (20.4 min) ¹¹ The C-labeled PET imaging agent can only be produced in the PET center equipped with an accelerator, which limits its clinical spread.

Therefore, the development of the PET molecular probe which has strong specificity and high sensitivity and can be popularized and applied and targets the EGFR-T790M mutation has important practical significance.

Disclosure of Invention

The present invention is directed to solving at least one of the problems of the prior art described above. Therefore, the invention provides a compound and a PET molecular probe targeting EGFR, which can target EGFR-T790M mutation and have the advantages of strong specificity, high sensitivity, popularization and application and the like.

The invention also provides a preparation method of the PET molecular probe.

The invention also provides an application of the compound or the PET molecular probe.

According to one aspect of the present invention, there is provided a compound targeting EGFR, or a pharmaceutically acceptable salt thereof, the compound having the structure according to formula (I):

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according to one aspect of the present invention, there is provided a PET molecular probe targeting EGFR, wherein the PET molecular probe is structured as the compound of claim 1, or a pharmaceutically acceptable salt thereof ¹⁸ The specific structure of the F labeled molecule is shown as the formula (II):

according to another aspect of the present invention, there is provided a method for preparing the PET molecular probe, comprising the steps of:

s1: obtaining an active structure substituted by a p-toluenesulfonate group shown as a formula (III);

s2: the active structure and ¹⁸ F-KF reaction to obtain the PET molecular probe;

the reaction equation is as follows:

in the present invention, fluoroethyl is carried out using the above method ¹⁸ F labeling, using p-toluenesulfonate-substituted active structure (OTs-substituted F) as a drug labeling precursor, and directly using a one-step method to label ¹⁸ F is introduced onto the parent nucleus structure. The method has high reaction efficiency, and can effectively obtain ¹⁸ F labeled molecule, is suitable for hospital and clinical use.

In some embodiments of the present invention, the specific steps of step S2 include: the active structure is subjected to, ¹⁸ And mixing the F-KF, alkali and a phase transfer catalyst for reaction to obtain the PET molecular probe.

In some embodiments of the invention, the base comprises K ₂ CO ₃ 、Na ₂ CO ₃ Me4NHCO3, etc.; the phase transfer catalyst comprises K _2.2.2 TEAB, etc.; the solvent includes DMSO, meCN, DMF, and the like.

In some preferred embodiments of the invention, the base is K ₂ CO ₃ 。

In some preferred embodiments of the invention, the phase transfer catalyst is K _2.2.2 。K _2.2.2 Is an aminopolyether (cryptate), aminopolyether (cryptand).

In some preferred embodiments of the invention, the solvent of the reaction is DMSO; the reaction temperature is 100-120 ℃; the reaction time is 10-20 minutes.

In the present invention, it is demonstrated by the experiments of examples that K is the base ₂ CO ₃ The phase transfer catalyst is K _2.2.2 And the solvent is DMSO, the temperature is 100-120 ℃, the time is 10-20 minutes, and the product can be obtained

In some embodiments of the present invention, the preparation method of the active structure in the step S1 is as follows:

according to a further aspect of the present invention, there is provided a use of the above compound or PET molecular probe, comprising at least one of:

i. preparation of medicaments for cancer diagnosis;

ii. Preparing a medicament for evaluating the curative effect of the medicament;

and iii, preparing a medicament for diagnosing and evaluating the curative effect of the brain metastasis NSCLC patient.

In some embodiments of the invention, the cancer is lung cancer.

In some embodiments of the invention, the medicament further comprises a pharmaceutically acceptable salt and/or an adjuvant.

In some preferred embodiments of the present invention, the excipient refers to an excipient that is conventional in the pharmaceutical field, such as: diluents, excipients such as water, etc., fillers such as starch, sucrose, etc.; binders such as cellulose derivatives, alginates, gelatin, and polyvinylpyrrolidone; humectants such as glycerol; disintegrating agents such as agar, calcium carbonate and sodium bicarbonate; absorption enhancers such as quaternary ammonium compounds; surfactants such as cetyl alcohol; adsorption carriers such as kaolin and bentonite clay; lubricants such as talc, calcium stearate and magnesium stearate, and polyethylene glycol, and the like. Other adjuvants such as sweetener, flavoring agent, etc. can also be added into the composition.

As used herein, the term "pharmaceutically acceptable" refers to those substances which are, within the scope of normal medical judgment, suitable for use in contact with the tissues of patients without undue toxicity, irritation, allergic response, and the like, commensurate with a reasonable benefit to risk ratio, and effective for their intended use.

In the present invention, the term "pharmaceutically acceptable salt" refers to a salt with a pharmaceutically acceptable non-toxic base or acid, including inorganic or organic bases and inorganic or organic acids. The salt of an inorganic base may be selected, for example, from: aluminum, ammonium, calcium, copper, iron, ferrous, lithium, magnesium, manganese, manganous, potassium, sodium, and zinc salts. Further, the salt of the pharmaceutically acceptable inorganic base may be selected from ammonium, calcium, magnesium, potassium and sodium salts. One or more crystal structures may be present in the solid salt, as well as in the form of hydrates. The pharmaceutically acceptable salts of organic non-toxic bases may be selected, for example, from: primary, secondary and tertiary amine salts, the substituted amines including naturally occurring substituted amines, cyclic amines and basic ion exchange resins such as arginine, betaine, caffeine, choline, N-dibenzylethylenediamine, diethylamine, 2-diethylaminoethanol, 2-dimethylaminoethanol, ethanolamine, ethylenediamine, N-ethylmorpholine, N-ethylpiperidine, glucosamine, histidine, hydrabamine, isopropylamine, lysine, methylglucamine, morpholine, piperazine, piperidine, polyamine resins, procaine, purines, theobromine, triethylamine, trimethylamine, tripropylamine and tromethamine.

The invention has at least the following beneficial effects:

1. the EGFR-targeting compound or PET molecular probe is structurally optimized based on the third-generation EGFR-TKI capable of specifically recognizing EGFR-T790M gene mutation, and simultaneously, in order to enable the pharmacokinetic behavior of a developer to be close to that of a therapeutic drug, the chemical structure of the EGFR-targeting compound or PET molecular probe is slightly changed based on the structure of a mother nucleus;

2. the PET probes of the invention are related to other EGFR-TKI molecular probes (e.g. ¹¹ C-Osimertinib， ¹⁸ F-Afatinib and the like) with low background of lung, liver and spleen, and is beneficial to displaying focus;

3. can pass through the blood brain barrier, the indole ring N thereof ₁ The substituent isThe introduction of 2-fluoroethyl group and 2-fluoroethyl group will maintain their biological activity, while the introduction of fluorine atoms can be used for radioactive elements ¹⁸ And F is marked.

4. Is composed of ¹⁸ F label, half life is appropriate.

Drawings

The invention is further described with reference to the following figures and examples, in which:

FIG. 1 is a graph of a concentration standard curve of a standard substance in example 3 of the present invention;

FIG. 2 is a UV and radio HPLC chromatogram of the isolated and purified radiation product of example 3 of the present invention;

FIG. 3 is a comparison of the UV HPLC spectra of the radioactive product and the nonradioactive standard product of example 3 of the present invention;

FIG. 4 shows normal nude mice in example 4 of the present invention ¹⁸ F-fluoroethyltinib 120min Micro-PET/CT image display.

Detailed Description

The concept and technical effects of the present invention will be clearly and completely described below in conjunction with the embodiments to fully understand the objects, features and effects of the present invention. It is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments, and other embodiments obtained by those skilled in the art without inventive efforts are within the protection scope of the present invention based on the embodiments of the present invention. The test methods used in the examples are all conventional methods unless otherwise specified; the materials, reagents and the like used are commercially available reagents and materials unless otherwise specified.

Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein like reference numerals refer to the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are illustrative only for the purpose of explaining the present invention, and are not to be construed as limiting the present invention.

In the description of the present invention, the meaning of a plurality is one or more, the meaning of a plurality is two or more, and the above, below, exceeding, etc. are understood as excluding the present numbers, and the above, below, within, etc. are understood as including the present numbers. If there is a description of first and second for the purpose of distinguishing technical features only, this is not to be understood as indicating or implying a relative importance or implicitly indicating the number of technical features indicated or implicitly indicating the precedence of technical features indicated.

In the description of the present invention, reference to the description of the terms "one embodiment," "some embodiments," "an illustrative embodiment," "an example," "a specific example," or "some examples," etc., means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the present invention. In this specification, the schematic representations of the terms used above do not necessarily refer to the same embodiment or example. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.

Example 1: design and Synthesis of precursors and standards

Firstly, screening a drug structure of a specific targeting EGFR-T790M mutation, designing a series of fluorine-containing labeled PET molecular probes targeting the EGFR-T790M mutation based on the relationship between the drug structure and the activity from the core chemical structure of the third generation EGFR-TKIs therapeutic drug oxitinib, and selecting the PET molecular probes with better imaging effect ¹⁸ F-fluoroethyltinib is used as a PET molecular probe targeting EGFR. The structure of the PET probe targeting EGFR-T790M in the present application is shown below and designated herein ¹⁸ F-fluoroethyltinib:

taking the synthesis example of the standard product and the labeled precursor of the fluoroethyltinib, ¹⁸ the synthesis route of the probe standard sample of F-fluoroethyltinib labeled Precursor (Precursor) is shown below.

Synthesis of intermediate 2:

in a 500mL three-necked flask, 2-methoxy-4-fluoro-5-nitroaniline (5.00g, 26.86mmol,1.0 eq) and methylene chloride (200 mL) were added in this order. After the raw materials were completely dissolved, a solution of di-tert-butyl dicarbonate (6.16g, 28.20mmol, 1.05eq) in dichloromethane was slowly added dropwise over 1 hour using a dropping funnel. After the dropwise addition, the reaction was continued at room temperature for 7 hours with stirring. The reaction mixture was washed with saturated brine, dried and concentrated to give intermediate 2 (7.15 g, colorless oily viscous liquid), LCMS (EI, M/z): [ M + H ] + =287.3.

And (3) synthesis of an intermediate 3:

in a 500mL three-necked flask, intermediate 2 (7.00g, 24.45mmol, 1.0eq), anhydrous DMF (200 mL) was added in this order, followed by N, N' -trimethyl-1,2-ethylenediamine, and the reaction was stirred at 80 ℃ for 12h. TLC plate material showed 1 disappearance, DMF was removed under reduced pressure, and the reaction solution was washed with saturated brine and Na ₂ SO ₄ The dried, concentrated under reduced pressure crude product was purified by silica gel column chromatography (eluent DCM: meOH =20]+＝369.4。

Synthesis of intermediate 4:

under nitrogen protection, in a 250mL autoclave, intermediate 3 (8.30g, 22.53mmol, 1.0eq), anhydrous methanol (100 mL) and then ammonium formate (4.19g, 67.58mmol, 3.0eq), pd/C (10%, 0.83 g) and magneton were added in this order. The reaction is stirred for 2 to 4 hours at the temperature of 80 ℃. TLC dot plate material showed disappearance of intermediate 3, after Pd/C filtration through celite, concentration under reduced pressure, and column chromatography on silica gel (eluent DCM: meOH =30 1) to afford intermediate 4 (7.53 g, colorless oily viscous liquid), LCMS (EI, M/z) [ M + H] ⁺ ＝339.4。

Synthesis of intermediate 5:

in a 500mL three-necked flask, intermediate 4 (7.20g, 21.27mmol, 1.0eq), and anhydrous DCM (150 mL) were sequentially added, followed by slowly adding a solution of acryloyl chloride (2.31g, 25.53mmol, 1.2eq) in DCM (50 mL) dropwise at 0 ℃. After the dropwise addition, the reaction solution was stirred at room temperatureStirring and reacting for 4h. TLC point plate material showed disappearance of intermediate 4, DCM was removed under reduced pressure, and the reaction was washed successively with saturated brine, saturated NaHCO3 solution, and then anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by silica gel column chromatography (eluent DCM: meOH = 20) to give intermediate 5 (8.27 g, colorless oily viscous liquid), LCMS (EI, M/z) [ M + H)]+＝393.5。

Synthesis of intermediate 6:

a500 mL three-necked flask was charged with a reaction mixture of intermediate 5 (7.90g, 20.13mmol, 1.0eq), anhydrous dioxane (100 mL), and 4M HCl in dioxane (100 mL) in this order, and the mixture was stirred at room temperature for 2 hours. TLC plates starting material showed disappearance of intermediate 5 and concentration under reduced pressure gave a white solid. The crude product was dissolved in 200mL DCM and then sequentially washed with saturated brine and saturated NaHCO ₃ Washing with the solution, followed by anhydrous Na ₂ SO ₄ And (5) drying. Concentration under reduced pressure gave intermediate 5 (6.55 g, white solid), LCMS (EI, M/z): M + H]+＝329.8。 ¹ H-NMR(400MHz,CDCl ₃ )δ9.57(s,1H),6.71-6.62(m,1H),6.48(s,1H),6.32-6.30(m,1H),5.80-5.77(m,1H),5.80-5.77(m,1H),4.65(s,2H),3.87(s,3H),3.45-3.40(m,4H),3.34-3.31(m,3H),2.84-2.82(d,J＝4.8Hz,6H),2.77(s,3H)。

Synthesis of intermediate 9:

indole (2.40g, 20.48mmol,1.0 eq), anhydrous THF (100 mL) were added sequentially in a 250mL three-necked flask at 0 ℃ under nitrogen, followed by the slow addition of a solution of methylmagnesium bromide in THF (1M, 51.2mL2.5eq) in DCM (50 mL) at 0 ℃. After completion of the addition of the Grignard reagent, a THF solution (20 mL) of the starting material 8 was slowly added dropwise, and then the reaction solution was stirred at room temperature for 8 hours. After disappearance of indole on the TLC plate, the reaction was quenched by slowly dropping saturated saline, and 300mL of ethyl acetate and 100mL of saturated saline were added to the reaction solution. The separated organic phase was washed with saturated brine, saturated NaHCO3 solution, and then with anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by column chromatography on silica gel (eluent PE: EA = 1:1) to give intermediate 9 (1.92 g, yellow powder), LCMS (EI, M/z): M + H]+＝230.6。

Synthesis of intermediate 10:

in a 250mL three-necked flask, intermediate 10 (1.10 g,2.84mmol,1.0 eq), anhydrous DMF (80 mL), potassium carbonate (0.96g, 6.97mmol,2.0 eq), potassium iodide (1.16 g,6.97mmol,2.0 eq), 2-bromoethoxy tert-butyldimethylsilane (1.0 g, 4.10 mmol,1.20 eq) were added in this order. The reaction was then stirred at 80 ℃ overnight. After disappearance of indole on TLC plate, DMF was removed by concentration under reduced pressure, and 300mL of ethyl acetate and 100mL of saturated aqueous sodium chloride were added to the concentrate. The separated organic phase was washed with saturated brine, saturated NaHCO3 solution and then with anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by column chromatography on silica gel (eluent PE: EA = 2:1) to give intermediate 10 (1.32 g, yellow solid), LCMS (EI, M/z): M + H]+＝390.0。

Synthesis of intermediate 11:

intermediate 10 (1.10 g,2.84mmol,1.0 eq), DCM (100 mL), 40% aqueous HF (10 mL) were added sequentially to a 250mL three-necked flask with stirring at 0 ℃. The reaction solution was then stirred at room temperature for 30 minutes. After TLC plate showed disappearance of intermediate 10, 200mL DCM,200mL saturated brine solution was added to the reaction. The separated organic phase is sequentially treated with 200mL of saturated saline and saturated NaHCO ₃ 200mL of the solution was washed, followed by anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by column chromatography on silica gel (eluent PE: EA = 2:1) to give intermediate 11 (0.76 g, yellow solid), LCMS (EI, M/z): M + H]+＝274.7。

Synthesis of intermediate 12:

intermediate 12 (0.70g, 1.64mmol, 1.0eq), dry isobutanol (50 mL), intermediate 6 (526.1mg, 1.80mmol, 1.10eq), p-toluenesulfonic acid (563.4mg, 3.27mmol, 2.0eq) were sequentially added to a 250mL three-necked flask with stirring at 0 ℃, and then a DCM solution (50 mL) of p-toluenesulfonyl chloride (0.66g, 3.45mmol, 1.5eq) was slowly added dropwise to the reaction solution for 30 minutes, and the reaction solution was stirred at room temperature for 8 hours after completion of dropwise addition. After TLC plate showed disappearance of intermediate 11, 200mL DCM,200mL saturated brine solution was added to the reaction. Separation ofThe organic phase was washed with 200mL of saturated brine, 200mL of 0.1M HCl solution, and then with anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by column chromatography on silica gel (eluent PE: EA = 2:1) to give intermediate 12 (0.93 g, white solid), LCMS (EI, M/z): M + H]+＝428.9。

¹⁸ Synthesis of F-fluoroethyltinib-labeled Precursor (Precursor):

to a 250mL three-necked flask were added, while stirring at room temperature, intermediate 12 (0.70g, 1.64mmol, 1.0eq), dry isobutanol (70 mL), intermediate 6 (526.1mg, 1.80mmol, 1.10eq), p-toluenesulfonic acid (563.4 mg,3.27mmol, 2.0eq) in this order, and the reaction mixture was stirred at 70 ℃ for 12 hours. After TLC plate showing disappearance of intermediate 12, isobutanol was removed by concentration under reduced pressure, and the crude product was dissolved in 200mL of ethyl acetate and then dissolved in 200mL of saturated brine and then in 200mL of saturated NaHCO ₃ 200mL of the solution was washed, followed by anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was separated and purified by silica gel column chromatography (eluent PE: EA = 1:1), ¹⁸ f-fluoroethyltinib labeled precursor (0.64 g, light yellow powder), LCMS (EI, M/z): M + H]+＝684.3。 ¹ H-NMR(400MHz,CD ₃ OD)δ10.55(s,1H),10.43(s,1H),9.78(s,1H),9.30(s,1H),9.17(s,1H),8.49(s,1H),8.25(m,3H),8.09(s,1H),7.98(s,1H),7.78(s,1H),7.65(t,J＝12Hz,2H),7.69-7.41(m,1H),6.85(s,1H),6.0(s,4H),5.60(s,1H),4.97(m,4H),4.44(s,1H),4.39(d,J＝16Hz,1H),4.02(s,2H),3.71(s,2H),3.42(s,2H),3.30(s,5H),3.01(s,2H)； ¹³ C-NMR(400MHz,CD ₃ OD)δ165.2,163.7,160.6,158.4,146.7,138.8,138.5,134.8,133.5,130.6,128.6,128.1,127.3,127.2,127.1,123.4,122.5,122.1,115.2,113.6,111.3,108.9,105.2,58.1,56.7,55.7,54.9,45.7,43.8,43.7,21.5。

The synthetic route of the fluoroethyltinib standard substance is as follows:

synthesis of intermediate 13:

stirring at room temperatureNext, intermediate 9 (0.90g, 3.92mmol,1.0 eq), anhydrous DMF (50 mL), 2-fluoroiodoethane (2.05g, 11.76mmol,3.0 eq), anhydrous potassium carbonate (1.08g, 7.84mmol,2.0 eq) were added in this order to a 250mL three-necked flask, and the reaction mixture was stirred at 70 ℃ for 12 hours. TLC plate shows that intermediate 9 disappeared, after concentrating under reduced pressure to remove DMF, the concentrated crude product was redissolved with 200mL of ethyl acetate, and then 200mL of saturated brine solution was added. After separation by shaking, the organic phase was separated and washed with 200mL of saturated brine and 200mL of 0.1M HCl solution in this order, followed by anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was purified by silica gel column chromatography (eluent PE: EA = 2:1) to give intermediate 13 (0.86 g, white powder), LCMS (EI, M/z): M + H]+＝276.7。

Synthesis of fluoroethyltinib standard:

to a 250mL three-necked flask were added, while stirring at room temperature, intermediate 13 (0.76g, 2.76mmol, 1.0eq), dry isobutanol (80 mL), intermediate 6 (0.90g, 3.06mmol, 1.10eq), p-toluenesulfonic acid (0.95g, 5.51mmol, 2.0eq) in this order, and the reaction mixture was stirred at 80 ℃ for 18 hours. After TLC plate showing disappearance of intermediate 13, isobutanol was removed by concentration under reduced pressure, and the crude product was dissolved in 200mL of ethyl acetate and then dissolved in 200mL of saturated brine and then in 200mL of saturated NaHCO ₃ 200mL of the solution was washed, followed by anhydrous Na ₂ SO ₄ And (5) drying. The crude product after concentration under reduced pressure was separated and purified by silica gel column chromatography (eluent PE: EA = 1:1), ¹⁸ f-fluoroethyltinib standard (1.32 g, white powder), LCMS (EI, M/z): M + H]+＝532.6。 ¹ H-NMR(400MHz,CD ₃ OD)δ9.50(s,1H),8.63(s,1H),8.19(d,J＝4.0Hz,1H),8.07(s,1H),7.30(s,1H),7.14(s,2H),7.07(d,J＝4.0Hz,1H),6.77(s,1H),6.56-6.50(m,1H),6.32(d,J＝12.0Hz,1H),5.72(d,J＝8.0Hz,1H),5.03(s,2H),4.62(d,J＝20.0Hz,2H),4.38(d,J＝40Hz,2H),3.78(s,3H),2.78(s,2H),2.51(s,3H),2.09(s,6H)； ¹³ C-NMR(400MHz,CD ₃ OD)δ165.0,163.7,160.6,158.4,139.0,138.5,133.6,128.7,127.3,127.1,126.8,123.4,122.5,122.4,115.1,113.7,111.5,108.9,105.0,84.2,82.9,58.0,56.7,55.7,49.9,47.7(d,J＝17.0Hz),45.7,43.7； ¹⁹ F-NMR(375MHz,CD ₃ OD)δ220.9(m,1F)。

Example 2: probe needle ¹⁸ F radiolabelling method optimisation

This example developed a high yield, high specific activity probe ¹⁸ F labelling method for fluoroethyl ¹⁸ F labeling, in this example, the active structure substituted by p-toluenesulfonate (OTs substituted for F) is used as a drug labeling precursor, and the labeling method performed by reaction temperature, additive, solvent, reaction time and the like is searched to improve the radiolabeling yield.

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After optimizing the alkali, the Phase Transfer Catalyst (PTC), the solvent, the temperature and the reaction time, the best marking condition is found by experiments and is 200mCi ¹⁸ F start, K ₂ CO ₃ As alkali, K _2.2.2 As phase transfer catalyst, DMSO as solvent, reacting the reaction mixture at 110 deg.C for 15min to obtain optimal labeling yield of 32.6%, and radioactive purity>99% and the specific activity is 373 GBq/. Mu.M. Therefore, OTs are excellent ¹⁸ F nucleophilic substitution of the leaving group, in which optimized conditions fluoroethyl is carried out ¹⁸ The F-tag would not be a problem.

Example 3: structure confirmation and purification of targeting EGFR compound and PET molecular probe

The chemical structures and purities of the targeted EGFR compound and the PET molecular probe were confirmed by high performance liquid chromatography HPLC. After separation by preparative HPLC, the probes that were successfully heat-labeled should be tested for chemical purity and radiochemical purity of the labeled product by analytical HPLC and radioactive TLC, respectively, while calculating specific activity using the standard curve of uv absorption of the standard sample.

3.1 analytical HPLC analytical method:

a chromatographic column: c18 150mm 4.0mm 5 μm;

column temperature: 25 ℃; sample introduction amount: 20 mu l of the mixture; flow rate: 1ml/min;

a detector: UV (. Lamda.214 nm) + B-FC-3200 (gamma ray detector);

mobile phase: a:0.1% aqueous trifluoroacetic acid (V/V); b:0.1% trifluoroacetic acid in acetonitrile (V/V);

the specific analytical methods are shown in table 1 below:

TABLE 1 HPLC analytical methods

The target compound is isolated by the above-mentioned separation and purification operation.

3.2 concentration standard curve of standard substance, as shown in figure 1:

the corresponding ultraviolet absorption areas of the standard samples at a series of concentrations (0.5ppm, 2.5ppm,5.0ppm and 7.5ppm) were tested to obtain a standard concentration curve, and the linear relationship between the sample concentration and the peak area was verified as shown in the following table 2:

TABLE 2 sample concentration-Peak area Linear relationship verification

Concentration of	0.5	2.5	5.0	7.5
					Peak area	25789	144860	283748	442066

The relation between the ultraviolet absorption area and the concentration is y =59069x-4778.4, and the correlation R ² =0.9993. The molecular weight of the standard is MW =531.64g/M, assuming that the amount of radioactivity in M (mL) volume of the sample to be tested isn (mCi), at a concentration c (μ g/mL), so that the specific activity of the radioactive product is:

3.3 ultraviolet and radioactive HPLC spectrograms of the separated and purified radioactive product are shown in FIG. 2, the ultraviolet retention time of the target product is 9.035min, and the ultraviolet purity is 100% (upper graph in FIG. 2); the retention time of the emission peak was 9.082min, and the emission purity was 100% (lower panel in FIG. 2). The UV peak and the emission peak overlap, indicating that the UV signal and the emission signal are the same compound.

3.4 comparing the ultraviolet HPLC spectra of the radioactive product and the nonradioactive standard product, as shown in FIG. 3, the ultraviolet retention time of the nonradioactive standard product is consistent with that of the radioactive product, indicating that the isolated and purified product is the radioactive element labeled standard product.

Example 4: in vivo development experiment of PET

Preparing normal Nude mice BALB/c-Nude, injecting it ¹⁸ F PET molecular Probe (in the above example) ¹⁸ F-fluoroethtinib) 4MBq, and imaging the nude mice using the small animal Micro-PET/CT within 120min, and recording the developed images for 30s, 5min, 15min, 30min, 60min, 90min and 120min, respectively, as shown in fig. 4.

As shown in fig. 4, the lower graph is a PET-CT fusion graph at different time instants, and the upper graph is a corresponding PET imaging graph; the distribution of the PET molecular probes is shown, and the uptake distribution in organs of normal nude mice is low. From the imaging results, it can be seen that the molecular probes are relatively different from other EGFR-TKI molecular probes, such as ¹¹ C-Osimertinib、 ¹⁸ F-Afatinib, et al, herein ¹⁸ F-fluoroethyltinib has low background of lung, liver and spleen, and is favorable for displaying focus.

The invention has at least the following beneficial effects:

1) The treatment effect of NSCLC patients is closely related to the EGFR gene phenotype of tumor tissues, and the EGFR gene phenotype sensitive to the treatment of specific drugs is accurately screened, so that the treatment effect of the drugs is improved. The conventional methods such as blood detection, tumor puncture PCR gene analysis and the like have the limitations of low detection sensitivity, tumor heterogeneity and the like. The noninvasive PET image can overcome the limitation of the conventional method, improve the detection sensitivity, simplify the screening process and be beneficial to determining the personalized treatment scheme.

2) The molecular structure design of the PET probe developed by the invention is based on EGFR-TKIs therapeutic drugs, namely axitinib and amitinib, which are already on the market, and the PET probe can specifically identify EGFR-T790M mutation, and the safety and the relationship between the therapeutic effectiveness and the gene phenotype of EGFR are clinically verified. Only short half-life radionuclides can be used compared to oxitinib and amitinib ¹¹ C label (t 1/2= 20min), the probe developed by the invention has better biological activity and can be used ¹⁸ The F mark (t 1/2= 110min) is beneficial to popularization and application in a PET center without an accelerator, and has great advantages; and the excellent imaging effect is verified by experiments, the background of the compound is lower in lung, liver and spleen, and the compound is beneficial to displaying focus.

3) The molecular structure of the PET probe developed by the invention is highly consistent with that of the therapeutic drug, so that the pharmacokinetic behaviors of the PET probe and the therapeutic drug in vivo are very close to each other, the absorption distribution of the probe in vivo, which can reflect the therapeutic drug to the maximum extent, can be used for screening EGFR-T790M treatment sensitive patients and monitoring the therapeutic effect of the drug treatment of the patients, and if the drug resistance occurs, the feedback can be timely carried out, thereby being beneficial to optimizing the treatment scheme.

The embodiments of the present invention have been described in detail with reference to the drawings, but the present invention is not limited to the embodiments, and various changes can be made within the knowledge of those skilled in the art without departing from the gist of the present invention. Furthermore, the embodiments of the present invention and the features of the embodiments may be combined with each other without conflict.

95 gradient down to 20

5 gradient up to 80

20 gradient up to 95

80 gradient down to 5

1. A compound targeting EGFR, or a pharmaceutically acceptable salt thereof, wherein the compound has the structure according to formula (I):

2. an EGFR-targeting PET molecular probe having the structure of the compound of claim 1, or a pharmaceutically acceptable salt thereof ¹⁸ The specific structure of the F labeled molecule is shown as the formula (II):

3. a method for preparing a PET molecular probe according to claim 2, comprising the steps of:

s1: obtaining an active structure substituted by p-toluenesulfonate shown as a formula (III);

the reaction equation is as follows:

4. the preparation method according to claim 3, wherein the specific steps of the step S2 comprise: the active structure is, ¹⁸ And mixing the F-KF, alkali and a phase transfer catalyst, and then reacting to obtain the PET molecular probe.

5. The method according to claim 4, wherein the base is K ₂ CO ₃ 。

6. The method of claim 4, wherein the phase transfer catalyst is K _2.2.2 。

7. The method according to claim 4, wherein the solvent for the reaction is DMSO;

the reaction temperature is 100-120 ℃;

the reaction time is 10-20 minutes.

8. The method according to claim 3, wherein the active structure in step S1 is prepared as follows:

9. use of a compound according to claim 1 or a PET molecular probe according to claim 2, comprising at least one of:

i. preparation of medicine for cancer diagnosis;

10. Use according to claim 9, wherein the cancer is lung cancer.