CN114989166B - Tumor KRAS G12C mutation targeting positron tracer agent, preparation method and application - Google Patents
Tumor KRAS G12C mutation targeting positron tracer agent, preparation method and application Download PDFInfo
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Abstract
The invention relates to a tumor KRAS G12C mutation targeting positron tracer agent, a preparation method and application. The existing KRAS mutation detection means cannot completely meet the target requirement of accurate diagnosis and treatment. The invention provides tumor KRAS G12C mutation targeting compounds 18 F/ 19 F-QZLO containing 18 The F-QZLO positron tracer is used for nuclide diagnosis of KRAS G12C mutation positive tumors, 19 F-QZLO was used as a stabilizing standard. The invention marks the radionuclide on the AMG510 through glycol or polyethylene glycol chains 18 F, constructing a PET molecular image probe, serving as a tumor KRAS G12C mutation targeting positron tracer agent and providing a stable standard substance 19 F-QZLO has proper physicochemical and radiological properties and ideal biological characteristics, can be used for PET imaging of KRAS G12C mutant tumors, and is helpful for specific diagnosis of the tumors, selection of targeted therapeutic drugs and efficacy evaluation.
Description
Technical Field
The invention relates to a positron tracer for positron emission computed tomography (Positron Emission Computed Tomography, PET), in particular to a tumor murine sarcoma virus oncogene (kirsten rat sarcoma viral oncogene, KRAS) G12C mutation targeting positron tracer, a preparation method and application thereof.
Background
Lung cancer is a serious disease that severely threatens human health, with morbidity and mortality leading to all tumors. In recent years, the therapeutic effect of EGFR mutant lung cancer is revolutionarily improved by molecular targeted drugs represented by EGFR-tyrosine kinase inhibitors (Epidermal growth factor receptor-tyrosine kinase inhibitor, EGFR-TKI), and the Progression-free survival (PFS) is greatly increased to 8.4-13.1 months from 4.6-6.7 months of traditional chemoradiotherapy. However, clinical studies have also shown that 29% -63% of EGFR mutant patients do not respond to EGFR-TKI treatment. Therefore, it is difficult to accurately evaluate and predict the treatment effect of EGFR-TKI of lung cancer by simply relying on EGFR mutation screening, and a new strategy for EGFR-TKI curative effect prediction needs to be established.
KRAS mutation, one of the most common genetic mutations in lung cancer, can significantly reduce or even completely eliminate the therapeutic response of lung cancer patients to EGFR-TKI, and is an important efficacy predictive marker. Based on the important role of KRAS mutation in lung cancer molecular targeted therapy decisions and prognosis, several international organizations such as the american society for clinical oncology (American Society of Clinical Oncology, ASCO) recommend KRAS mutation screening as a routinely accepted clinical molecular examination for lung cancer patients; the national cancer network (National Comprehensive Cancer Network, NCCN) guidelines indicate that "KRAS mutation or not determines whether patients still have to detect other molecular therapeutic targets". Therefore, accurate detection of KRAS mutation is a significant clinical need for accurate diagnosis and treatment of lung cancer. However, in the existing KRAS mutation detection means in clinic, invasive puncture biopsy is limited by the limitation of a puncture part and cannot accurately reflect the heterogeneity of KRAS, and the problem of low sensitivity exists in circulating tumor DNA detection, so that the two problems cannot completely meet the target of accurate diagnosis and treatment. The molecular image can be used for real-time, quantitative and visual imaging of living biological molecules.
In recent years, KRAS mutation targeting molecules AMG510 and ARS1620 and the like are sequentially introduced, the former has been approved by FDA and marketed, and the KRAS mutation targeting molecules AMG510 and ARS1620 can inhibit the activity of KRAS G12C protein by irreversibly binding to the KRAS mutation targeting molecules AMG and ARS1620, so that a brand-new development opportunity is provided for molecular imaging of KRAS mutation. Patients with objective remission rate (Objective Response Rate, ORR, patient proportion with tumor volume reduction of > 30%) of 36% (95% CI: 28-45), 81% (95% CI: 73-87) achieved disease control (achieving complete remission, partial remission and patient proportion with stable disease condition exceeding 3 months) with once daily oral administration of 960mg AMG510 treatment. Median duration of remission (Duration of response, doR) was 10 months.
Disclosure of Invention
The invention aims to provide a tumor KRAS G12C mutation targeting positron tracer agent, a preparation method and application thereof, and the tumor KRAS G12C mutation targeting positron tracer agent is subjected to radionuclide based on AMG510 18 F labeling, and constructing a positron tracer of a targeting tumor KRAS G12C to solve the problem that the existing KRAS mutation detection means cannot completely meet the accurate diagnosis and treatment target.
In order to achieve the above purpose, the technical scheme adopted by the invention is as follows:
providing a tumor KRAS G12C mutation targeting compound which is 18 F/ 19 F-QZLO has the chemical structure:
where n=0, 1,2,3,4,5 or 6.
In another aspect, there is provided a positron tracer comprising a tumor KRAS G12C mutation targeting compound as described, said positron tracer comprising said compound being 18 F-QZLO has the chemical structure:
where n=0, 1,2,3,4,5 or 6.
Further, the positron tracer further comprises a solvent.
Further, the solvent is PBS containing ethanol or injection water or physiological saline solution.
In another aspect, there is provided the use of a positron tracer as described for nuclide diagnosis of KRAS G12C mutation positive tumors.
In another aspect, there is provided a method of preparing a positron tracer as described, the method comprising:
dissolving a compound 3, namely AMG510, in dimethyl sulfoxide, adding an acetonitrile solution of a compound 1, namely ethylene glycol di-p-toluenesulfonate, adding anhydrous potassium carbonate and potassium iodide for reaction, separating a product by a high performance liquid chromatography, and freeze-drying overnight to obtain a marked precursor compound 5, namely QZLO-OTs;
preparation of the compositions containing the described compositions by automated synthesis modules 18 Positron tracer of F-QZLO.
Further, the preparation of the composition containing the said components by means of an automated synthesis module 18 The positron tracer of the F-QZLO specifically comprises the following steps:
dissolving anhydrous potassium carbonate in water to obtain a solution A, dissolving aminopolyether in acetonitrile to obtain a solution B, and mixing the solution A and the solution B to obtain a reagent No. 1; dissolving a compound 5, namely QZLO-OTs, in anhydrous acetonitrile to obtain a No. 2 reagent;
prepared by cyclotron 18 F]The fluoride ion is passed through and adsorbed on the QMA anion exchange column, and then adsorbed on the QMA anion exchange column 18 F]F - Eluting with reagent No. 1 into a reaction bottle of a synthesis module, heating and maintaining under the protection of high-purity nitrogen or helium to remove water, and cooling;
adding the reagent No. 2 into the reaction bottle, heating and maintaining to obtain a mixture C, diluting the mixture C with injection water after the mixture C is cooled to room temperature to obtain a mixture D, passing the mixture D through a C18 reversed phase solid phase extraction column, and eluting adsorbates on the C18 reversed phase solid phase extraction column into a syringe sequentially by using absolute ethyl alcohol and injection water to obtain a mixed solution E;
acetonitrile is taken as a mobile phase, and the mixed solution E is separated by the preparation liquid chromatography of a synthesizer;
passing the separated product through C18 reversed phase solid phase extraction column, washing with injection water, eluting adsorbate on the C18 reversed phase solid phase extraction column with absolute ethyl alcohol and injection water sequentially, and filtering with a sterilizing filter to obtain the final product 18 Positron tracer of F-QZLO.
In another aspect, there is provided the use of a tumor KRAS G12C mutation targeting compound as described, said compound being 19 F-QZLO has the chemical structure:
wherein n=0, 1,2,3,4,5 or 6;
the said 19 F-QZLO was used as a stabilizing standard.
In another aspect, there is provided a method for preparing a tumor KRAS G12C mutation targeting compound as described above, which is 19 F-QZLO, said 19 The preparation method of the F-QZLO comprises the following steps:
adding the compound 1, namely glycol di-p-toluenesulfonate and tetrabutylammonium fluoride, into acetonitrile, adding anhydrous potassium carbonate and potassium iodide for reaction, separating a product by a high performance liquid chromatography, and freeze-drying overnight to obtain a compound 2;
dissolving compound 3, AMG510, in dimethyl sulfoxide, adding acetonitrile solution of compound 2, adding anhydrous potassium carbonate and potassium iodide for reaction, separating product by high performance liquid chromatography, and lyophilizing overnight to obtain stable standard compound 4 19 F-QZLO。
Compared with the prior art, the invention has the following beneficial effects:
the invention marks the AMG510 with the radionuclide 18 F, structureThe PET molecular imaging probe is established and used as a tumor KRAS G12C mutation targeting positron tracer agent, and experiments show that the PET molecular imaging probe has proper physicochemical and radiological properties and relatively ideal biological characteristics, can be used for PET imaging of KRAS G12C mutation tumors, and is beneficial to specific diagnosis of the tumors, selection of targeted therapeutic drugs and efficacy evaluation.
The invention selects radionuclides 18 F marks AMG 510. Because of the carbon element in the molecular skeleton, radionuclides are commonly used 11 C is labeled to minimally alter the molecular structure to ensure its affinity and selectivity for the receptor, but 11 C has short half-life (T) 1/2 =20.3 min), an online accelerator is required, which is not beneficial to clinical popularization; while 68 The metal nuclide labels such as Ga and the like need to modify chelating groups such as DOTA and the like, and the molecular structure is greatly changed, so that the affinity and the selectivity of the metal nuclide labels to a receptor are affected; thus, in contrast to these species, 18 f has a suitable half-life (T 1/2 =109.8 min), so that the tracer has sufficient time in the labeling preparation and clinical use processes, and the radionuclide is labeled at the tail end of the AMG510 after the glycol or polyethylene glycol chain is connected to the hydroxyl end of the AMG 18 The F has small influence on molecular structure, can ensure the affinity and selectivity of the F to a receptor, and simultaneously, the polyethylene glycol chain can reduce the fat-solubility of the compound, thereby being beneficial to clinical application.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings that are required in the embodiments or the description of the prior art will be briefly described, it being obvious that the drawings in the following description are only some embodiments of the invention, and that other embodiments of the drawings can be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is an HRMS of compound 2 prepared in example 1 of the present invention.
FIG. 2 is an HRMS plot of compound 4 prepared in example 1 of the present invention.
FIG. 3 is an HRMS plot of compound 5 prepared in example 1 of the present invention.
FIG. 4 is a schematic illustration of the preparation of example 1 of the present invention 18 F-QZLO Allinone module labeling schematic diagram.
FIG. 5 is a schematic illustration of the process of example 1 of the present invention 18 F-QZLO radioactive high performance liquid chromatography (Radio-HPLC) diagram.
FIG. 6 is a schematic diagram of the process of example 1 of the present invention 18 F-QZLO in vitro stability results.
FIG. 7 is a schematic illustration of the process of example 1 of the present invention 18 F-QZLO distribution results in normal mice.
FIG. 8 is a photograph of an example 1 of the present invention 18 F-QZLO in KRAS G12C mutant tumor-bearing mice PET imaging results map.
Detailed Description
In order that the invention may be readily understood, a more complete description of the invention will be rendered by reference to the appended drawings. The drawings illustrate preferred embodiments of the invention. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
It should be noted that like reference numerals and letters refer to like items, and thus once an item is defined in one embodiment, no further definition or explanation thereof is necessary in subsequent embodiments. The particular implementation is described in terms of steps in some embodiments for clarity and accuracy of presentation and should not be construed as limiting the sequence. In addition, in the examples, the chemical used in the steps is an existing material or a commercially available product.
AMG510 (motorasib) is a small molecule, specific, irreversible KRAS G12C inhibitor that binds irreversibly to KRAS G12C protein to inhibit its activity, interferes with the dissociation of GDP on KRAS G12C protein, and inhibits KRAS-mediated signaling by locking KRAS G12C protein in an inactive GDP-bound state. Positron emission computed tomography (Positron Emission Computed Tomography, PET) is a clinical examination imaging technique in the field of nuclear medicine, and involves a marker in the examination process, and after the marker is injected into a human body, the condition of life metabolism activity is reflected by the accumulation of the substance in metabolism, so that the purpose of diagnosis is achieved. At present, no report of radionuclide labeling on AMG510 is seen, and no radioactive diagnosis on KRAS G12C mutation positive tumors by using the AMG510 as a positron tracer for PET imaging is seen.
The invention connects glycol or polyethylene glycol chain on the hydroxyl end of AMG510 and marks the radionuclide 18 F, obtaining a tumor KRAS G12C mutation targeting compound, constructing a PET molecular image probe, and preparing a PET imaging positron tracer for targeting the tumor KRAS G12C, so that the preparation of the tracer based on AMG510 for PET examination has feasibility and is named as 18 F-Quinazolineone (quinazolinone), i.e 18 F-QZLO has the chemical structure:
comprises 18 The positron tracer of F-QZLO also comprises a solvent, wherein the solvent can be PBS containing ethanol or injection water or physiological saline solution, and the solvent is added in the corresponding steps in the preparation process. The positron tracer structure contains a ligand AMG510 targeting the KRAS G12C mutant protein and a PEG chain for radionuclide labeling, so that the positron tracer structure can be used for nuclide diagnosis of KRAS G12C mutant positive tumors.
Comprises 18 The preparation method of the positron tracer of the F-QZLO comprises the following steps:
dissolving a compound 3, namely AMG510, in dimethyl sulfoxide, adding an acetonitrile solution of a compound 1, namely ethylene glycol di-p-toluenesulfonate, adding anhydrous potassium carbonate and potassium iodide for reaction, separating a product by a high performance liquid chromatography, and freeze-drying overnight to obtain a marked precursor compound 5, namely QZLO-OTs;
preparation of the compositions containing the described compositions by automated synthesis modules 18 A positron tracer for F-QZLO comprising:
dissolving anhydrous potassium carbonate in water to obtain a solution A, dissolving aminopolyether in acetonitrile to obtain a solution B, and mixing the solution A and the solution B to obtain a reagent No. 1; dissolving a compound 5, namely QZLO-OTs, in anhydrous acetonitrile to obtain a No. 2 reagent;
prepared by cyclotron 18 F]The fluoride ion is passed through and adsorbed on the QMA anion exchange column, and then adsorbed on the QMA anion exchange column 18 F]F - Eluting with reagent No. 1 into a reaction bottle of a synthesis module, heating and maintaining under the protection of high-purity nitrogen or helium to remove water, and cooling;
adding the reagent No. 2 into the reaction bottle, heating and maintaining to obtain a mixture C, diluting the mixture C with injection water after the mixture C is cooled to room temperature to obtain a mixture D, passing the mixture D through a C18 reversed phase solid phase extraction column, and eluting adsorbates on the C18 reversed phase solid phase extraction column into a syringe sequentially by using absolute ethyl alcohol and injection water to obtain a mixed solution E;
acetonitrile is taken as a mobile phase, and the mixed solution E is separated by the preparation liquid chromatography of a synthesizer;
passing the separated product through C18 reversed phase solid phase extraction column, washing with injection water, eluting adsorbate on the C18 reversed phase solid phase extraction column with absolute ethyl alcohol and injection water sequentially, and filtering with a sterilizing filter to obtain the final product 18 Positron tracer of F-QZLO.
The invention also provides another tumor KRAS G12C mutation targeting compound 19 F-Quinazolineone ( 19 F-QZLO) by 18 F, marking, which can be used as a stable standard substance, and has the chemical structure as follows:
19 the preparation method of the F-QZLO comprises the following steps:
adding the compound 1, namely glycol di-p-toluenesulfonate and tetrabutylammonium fluoride, into acetonitrile, adding anhydrous potassium carbonate and potassium iodide for reaction, separating a product by a high performance liquid chromatography, and freeze-drying overnight to obtain a compound 2;
dissolving compound 3, AMG510, in dimethyl sulfoxide, adding acetonitrile solution of compound 2, adding anhydrous potassium carbonate and potassium iodide, and reacting with high efficiencySeparating the product by liquid chromatography, and lyophilizing overnight to obtain stable standard compound 4 19 F-QZLO。
The general formula of the two tumor KRAS G12C mutation targeting compounds is as follows:
the structure comprises KRAS G12C targeting ligand AMG510, ethylene glycol or polyethylene glycol (PEG) chain and labeled nuclides 18 F/ 19 F. Where n=0, 1,2,3,4,5 or 6.
Both compounds can be prepared by the same synthetic route, 18 F/ 19 the synthetic route of F-QZLO is specifically as follows:
step 1: the compound 1 is glycol di-p-toluenesulfonate, and the compound 1 is subjected to a reaction condition "a" to obtain a compound 2, wherein the reaction condition "a" is as follows: TBAF, K 2 CO 3 ，ACN，KI，90℃。
n=0, 1,2,3,4,5 or 6.
The step 1 specifically comprises the following steps:
compound 1 was added to Acetonitrile (ACN) with tetrabutylammonium fluoride (TBAF) 1:1, and 3 equivalents of anhydrous potassium carbonate (K) 2 CO 3 ) And a trace of KI, stirred at 90℃for 12 hours. The product was isolated by High Performance Liquid Chromatography (HPLC) and freeze-dried overnight to give compound 2.
Step 2: compound 3 is AMG510, compound 3 yields compound 4 via reaction conditions "b" of: compound 2,K 2 CO 3 ，KI，DMSO，ACN，90℃。
n=0, 1,2,3,4,5 or 6.
The step 2 is specifically as follows:
compound 3 was dissolved in dimethyl sulfoxide (DMSO), 2-fold equivalent of acetonitrile solution of compound 2 was added, and 3-fold equivalent of anhydrous potassium carbonate and a trace of KI were added, and stirred at 90 ℃ for 12 hours. The product was isolated by HPLC and lyophilized overnight to give stable standard compound 4, designated 19 F-QZLO。
Step 3: compound 3AMG510, compound 3 gave compound 5 via reaction condition "c", which is: compounds 1, K 2 CO 3 ，KI，ACN，DMSO，90℃。
n=0, 1,2,3,4,5 or 6.
The step 3 is specifically as follows:
compound 3 was dissolved in dimethyl sulfoxide, 2-fold equivalent of acetonitrile solution of compound 1 was added, and 3-fold equivalent of anhydrous potassium carbonate and a trace of KI were added, followed by stirring at 90 ℃ for 12 hours. The product was isolated by HPLC and freeze-dried overnight to give labeled precursor compound 5, designated QZLO-OTs.
Step 4: compound 5 is obtained by reaction condition "d 18 F-QZLO, the reaction condition "d" is: [ 18 F]F - ， Kryptofix 2.2.2，K 2 CO 3 ，ACN，10min，100℃。
n=0, 1,2,3,4,5 or 6.
The step 4 is specifically as follows:
step 4.1: dissolving anhydrous potassium carbonate in water to obtain a solution A, dissolving aminopolyether (Kryptofix 2.2.2) in acetonitrile to obtain a solution B, and mixing the solution A and the solution B to obtain a reagent No. 1; dissolving the compound 5 in anhydrous acetonitrile to obtain a reagent No. 2;
step 4.2: prepared by cyclotron 18 F]The fluoride ion is introduced and adsorbed on the QMA anion exchange columnThen adsorbed on QMA anion exchange column 18 F]F - Eluting with reagent No. 1 into a reaction bottle of a synthesizer, heating to 60 ℃ under the protection of high-purity nitrogen or helium for 1.0min, heating to 85 ℃ for 2.0min, heating to 120 ℃ for 2.0min, and finally cooling to 60 ℃;
step 4.3: adding a reagent No. 2 into the reaction bottle, heating to 100 ℃ and keeping the temperature for 10.0min to obtain a mixture C, diluting the mixture C with injection water after the mixture C is cooled to room temperature to obtain a mixture D, passing the mixture D through a C18 reversed phase solid phase extraction column, and eluting adsorbates on the C18 reversed phase solid phase extraction column into an injector by using absolute ethyl alcohol and injection water in sequence to obtain a mixed solution E;
step 4.4: separating the mixed solution E by using 45% acetonitrile as a mobile phase through a preparation liquid chromatograph of a synthesizer;
step 4.5: diluting the separated product with injection water, passing through C18 reversed phase solid phase extraction column, washing with injection water, eluting adsorbate on the C18 reversed phase solid phase extraction column with absolute ethyl alcohol and injection water sequentially, and filtering with sterilizing filter to obtain the final product 18 Positron tracer of F-QZLO.
Example 1:
preparation of the composition containing
I.e. having a tetrapolyethylene glycol chain 18 Positron tracer of F-QZLO:
step 1: compound 1, tetraethylene glycol bis-p-toluenesulfonate (1.05 g,2.09 mmol), tetrabutylammonium fluoride (TBAF, 0.55g,2.10 mmol), anhydrous potassium carbonate (K) 2 CO 3 0.89g,6.44 mmol) and a trace of KI were added to 2mL of acetonitrile and stirred at 90℃under reflux for 12 hours. The product was isolated by HPLC and freeze-dried overnight to give compound 2 (0.51 mg, 69.6%) as a yellow oil. HRMS (ESI-Tof), [ M+H ]] + :m/z calculated for C 15 H 24 FO 6 S + 351.1272,found 351.0597。
Step 2: will beCompound 3, AMG510 (3, 26.2mg,0.047 mmol), was dissolved in 0.2mL of anhydrous DMSO and a solution of 0.8mL of Compound 2 (27.6 mg,0.079 mmol) in acetonitrile was added to form a pale yellow clear solution. After anhydrous potassium carbonate (25.9 mg,0.19 mmol) was added, the solution was thickened to yellow, and the reaction was heated under reflux for 12 hours. Purifying by HPLC to obtain product, lyophilizing overnight to obtain white powder compound 4, i.e. stable standard product [ (] 19 F-QZLO,13.8mg，39.8％)。HRMS(ESI-Tof),[M+ H] + :m/z calculated for C 38 H 46 F 3 N 6 O 6 + 739.3425,found 739.1376。
Step 3: the same synthetic method as that of compound 4 was used for compound 1 (136.0 mg,0.28 mmol) and compound 3 (81.2 mg,0.14 mmol) to give labeled precursor compound 5 as a white solid, namely labeled precursor (QZLO-OTs, 43.0mg, 15.5%). HRMS (ESI-Tof), [ M+H ]] + :m/z calculated for C 45 H 52 F 2 N 6 O 9 S + 890.3485,found 891.1248。
Step 4: 18 F-QZLO was prepared by a Trasis Allinone automated synthesis module.
Step 4.1: 3.0mg of potassium carbonate (K) 2 CO 3 ) Dissolving in 0.2mL of water to obtain solution A, 13.0mg of aminopolyether (Kryptofix 2.2.2, K) 2.2.2 ) Dissolving in 0.8mL Acetonitrile (ACN) to obtain solution B, and mixing the solution A and the solution B to obtain a reagent No. 1;
2.0mg of QZLO-OTs was dissolved in 1.0mL of anhydrous acetonitrile to obtain reagent No. 2. The QZLO-OTs are compounds with the following structures:
step 4.2: prepared by cyclotron 18 F]Fluoride ion ([ V) 18 F]F - 300mCi,2.5 mL) was passed through and adsorbed on a QMA anion exchange column, and then the adsorption on the QMA anion exchange column was performed 18 F]F - Eluting with reagent No. 1 into the reaction flask of the synthesizer, and then eluting with high purity nitrogen (N 2 ) Heating to 60 ℃ under protectionAnd maintaining for 1.0min, heating to 85deg.C and maintaining for 2.0min, then heating to 120deg.C and maintaining for 2.0min, and finally cooling to 60deg.C;
step 4.3: adding a No. 2 reagent into the reaction bottle, heating to 100 ℃ and keeping the temperature for 10.0min to obtain a mixture C, diluting the mixture C to 15 mL by using injection water (water for injection) after the mixture C is cooled to room temperature to obtain a mixture D, passing the mixture D through a C18 reversed-phase solid-phase extraction column, and eluting adsorbate on the C18 reversed-phase solid-phase extraction column into a syringe by using 2.0mL of absolute ethyl alcohol (anhydrous ethanol) and 2.0mL of injection water in sequence to obtain a mixed solution E;
step 4.4: separating the mixed solution E by a preparation liquid chromatograph of a synthesizer by taking 45% acetonitrile as a mobile phase and the flow rate of 4mL/min, and collecting the separated product with the retention time of 21-23 min on a gamma chromatograph in a rotary bottle (containing 25mL of injection water) in 50 mL;
step 4.5: the separated product is passed through a C18 reversed-phase solid phase extraction column, then washed by 10mL of injection water, and then the adsorbate on the reversed-phase solid phase extraction column is eluted by 1.5mL of absolute ethyl alcohol (anhydrous ethanol) and 13.5mL of injection water in turn, and then filtered by a sterilizing filter (Merck Millipore MILLEX-GV 0.22 μm SLGVR33 RB) to obtain the product for injection 18 F-QZLO solution (10% ethanol).
The preparation time of this example was about 90 minutes, with a total radiochemical yield of 21.53±1.46% (n=3, corrected for decay) and a radiochemical purity of greater than 99%.
The positron tracers prepared in this example are described in their performance assays as follows:
(1) High performance liquid chromatography (Radio-HPLC) identification:
HPLC conditions: the column was an octadecyl bonded silica gel reversed phase column (Inertsil ODS-SP,4. mm ×250mm,5 μm, shimadzu Corp.) with acetonitrile and water (v: v=50:50) as mobile phases, isocratic elution, flow rate 1mL/min. 18 The retention time of F-QZLO is 14.5min, the radiochemical purity is more than 99 percent, which is higher than that of the F-QZLO in pharmacopoeia 18 The F-deoxyglucose has a radiochemical purity of greater than 90% (China pharmacopoeia 2020 edition, two parts). The Radio-HPLC results are shown in FIG. 5.
(2) Checking:
the pH value is 5.0-8.0 (Chinese pharmacopoeia 2020 edition, two parts, appendix VI H). Bacterial endotoxin detection: taking proper amount of the product (i.e. filtering with a sterilizing filter to obtain injectable product 18 F]QZLO solution), after 60-fold dilution with water for bacterial endotoxin inspection, was tested according to standard methods (chinese pharmacopoeia 2020 edition, two parts, appendix XI E), the endotoxin content per 1mL of the product being less than 15EU. Sterile inspection: and detecting a proper amount of the product according to a standard method (Chinese pharmacopoeia 2020 edition, two parts and annex XI H), wherein the product meets the requirements.
(3) Radioactivity concentration: accurately measuring a certain volume of the product, placing the product in an activity meter to measure the activity, and calculating the radioactive concentration according to the volume of the sample and the activity thereof. The radioactive concentration of the product is more than 110MBq/mL.
(4) Validity period: and calculating for 6 hours from the calibration time.
(5) Fat-solubility: measured by a shaking flask method 18 The F-QZLO lipid solubility was log d=2.17±0.08.
(6) Normal mouse in vivo distribution: FIG. 7 is a diagram of 18 F-QZLO distribution results in normal mice. The results show that the data obtained from the above-mentioned method, 18 F-QZLO has similar intake value in each organ and is mainly discharged out of the body through intestinal metabolism. And can see 18 F-QZLO showed no significant in vivo defluorination, and bone uptake was about 4% ID/g after 2 hours of injection.
(7) PET imaging of KRAS G12C mutant tumor-bearing mice: in H358 xenograft mice, 1 hour of dynamic PET/CT imaging was performed. As shown in FIG. 7, immediately after administration, tumors were observed, whose uptake was 3.98% ID/g, and increased slightly over time. This indicates 18 F-QZLO reaches the tumor site with blood immediately after tail vein injection and is taken up by irreversible binding to Cys 12. At the same time, the uptake of muscle also increased with time, the tumor to muscle (T/M) ratio reached a maximum at 5 minutes (T/m=2.27), and after 30 minutes the tumor to muscle (T/M) ratio remained essentially around 1.7, indicating 18 The distribution of F-QZLO reached equilibrium 30 minutes after intravenous injection. We can also see from the imaging results that, 18 F-QZLO has higher initial uptake in liver andthrough rapid metabolism and excretion of bile and intestinal tracts, the obtained product is matched with the in-vivo distribution result of normal mice.
FIG. 6 is a diagram of 18 In vitro stability results of F-QZLO are shown, and after incubation for 2h at room temperature 18 The radiochemical purity of F-QZLO in normal saline is still more than 99%, which shows that 18 F-QZLO has high physiological saline stability and is beneficial to clinical use.
The foregoing description of the invention has been presented for purposes of illustration and description, and is not intended to be limiting. Several simple deductions, modifications or substitutions may also be made by a person skilled in the art to which the invention pertains, based on the idea of the invention.
Claims (6)
1. The application of a tumor KRAS G12C mutation targeting compound in preparing a nuclide diagnosis positron tracer for KRAS G12C mutation positive tumors is characterized in that:
the compound is 18 F-QZLO has the chemical structure:
where n=3.
2. A positron tracer comprising a compound of claim 1, wherein:
the compound contained in the positron tracer is 18 F-QZLO has the chemical structure:
where n=3.
3. The positron tracer of claim 2, wherein:
the positron tracer further includes a solvent.
4. A positron tracer as claimed in claim 3 wherein:
the solvent is PBS containing ethanol or injection water or physiological saline solution.
5. The method for preparing the positron tracer as claimed in claim 4, wherein:
the preparation method comprises the following steps:
dissolving a compound 3, namely AMG510, in dimethyl sulfoxide, adding an acetonitrile solution of a compound 1, namely ethylene glycol di-p-toluenesulfonate, adding anhydrous potassium carbonate and potassium iodide for reaction, separating a product by a high performance liquid chromatography, and freeze-drying overnight to obtain a marked precursor compound 5, namely QZLO-OTs;
preparation of the compositions containing the described compositions by automated synthesis modules 18 Positron tracer of F-QZLO.
6. The method for producing a positron tracer as claimed in claim 5, wherein:
preparation of the compositions containing the described compositions by automated synthesis modules 18 The positron tracer of the F-QZLO specifically comprises the following steps:
dissolving anhydrous potassium carbonate in water to obtain a solution A, dissolving aminopolyether in acetonitrile to obtain a solution B, and mixing the solution A and the solution B to obtain a reagent No. 1; dissolving a compound 5, namely QZLO-OTs, in anhydrous acetonitrile to obtain a No. 2 reagent;
prepared by cyclotron 18 F]The fluoride ion is passed through and adsorbed on the QMA anion exchange column, and then adsorbed on the QMA anion exchange column 18 F]F - Eluting with reagent No. 1 into a reaction bottle of a synthesis module, heating and maintaining under the protection of high-purity nitrogen or helium to remove water, and cooling;
adding the reagent No. 2 into the reaction bottle, heating and maintaining to obtain a mixture C, diluting the mixture C with injection water after the mixture C is cooled to room temperature to obtain a mixture D, passing the mixture D through a C18 reversed phase solid phase extraction column, and eluting adsorbates on the C18 reversed phase solid phase extraction column into a syringe sequentially by using absolute ethyl alcohol and injection water to obtain a mixed solution E;
acetonitrile is taken as a mobile phase, and the mixed solution E is separated by the preparation liquid chromatography of a synthesizer;
passing the separated product through C18 reversed phase solid phase extraction column, washing with injection water, eluting adsorbate on the C18 reversed phase solid phase extraction column with absolute ethyl alcohol and injection water sequentially, and filtering with a sterilizing filter to obtain the final product 18 Positron tracer of F-QZLO.
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