CN110483603B

CN110483603B - Fluoro dithiophosphate compound and preparation method and application thereof

Info

Publication number: CN110483603B
Application number: CN201910847244.6A
Authority: CN
Inventors: 李子婧; 杨鸿章; 刘欢欢; 牟钊彪; 王潮
Original assignee: Xiamen University
Current assignee: Xiamen University
Priority date: 2019-09-09
Filing date: 2019-09-09
Publication date: 2021-03-26
Anticipated expiration: 2039-09-09
Also published as: CN110483603A

Abstract

The present invention relates to a fluorodithiophosphoric acid ester compound and a preparation method and application thereof. The fluorodithiophosphoric acid ester compound has a structure represented by formula (I). In the present invention, 1,3,2-dithiophosphalane is used as the ^18/19 F-labeling precursor for the first time, and a nucleophilic substitution-elimination low reaction energy barrier labeling strategy is used to construct ultra-fast and one-step fluorodithiophene Phosphoric acid ester compounds; the fluorodithiophosphoric acid ester compounds of the present invention can be prepared in an aqueous solution, while the labeling conditions are mild and the reaction time is short, and a 97% yield can be achieved under the condition of 20 ° C for 30 seconds. , no need to dry F- ^, high specific activity, convenient purification, good stability of the labeled product in vitro and in vivo, and has broad application in the field of positron medicine for the preparation of phosphate analog molecular probes, heat-sensitive and solvent-sensitive peptides or proteins and other biomolecules application prospects.

Description

Fluoro dithiophosphate compound and preparation method and application thereof

The technical field is as follows:

the invention relates to synthesis and labeling of a novel fluoro phosphorodithioate compound, in particular to application of the compound in preparation of a positron emission developer.

Background art:

positron Emission Tomography (PET) is an imaging technology capable of displaying biomolecular metabolism and activities of various receptors and nerve mediators on a living body, and is widely used in diagnosis and differential diagnosis of various diseases such as tumors, nervous systems and cardiovascular systems, disease judgment, treatment effect evaluation, organ function research, new drug development and the like.

Among the nuclides commonly used in PET imaging,¹⁸f is a positron nuclide which is most widely applied, and has excellent chemical properties and nuclide characteristics: the Van der Waals radius of the fluorine atoms is similar to that of the hydrogen atoms, and the influence on the biological properties of the labeled compound is small;¹⁸the F half-life is appropriate (109.8 minutes), allowing for multi-step labeling reactions and delayed PET imaging; 97% beta⁺Decay, maximum energy of ray is 0.635MeV, and the imaging space resolution can be improved with less radiation damage to normal tissues. These advantages are that¹⁸The research of the F labeling method and the development of PET probes become a research hotspot at present.

In recent years, on non-carbon atoms¹⁸The F-labeling method is gradually developed and used for the preparation of PET probes. The marking method has good selectivity and high Radiochemical yield (RCYs), and compared with the method for marking fluorine on carbon atoms, the marking condition is relatively mild. However, of such non-carbon atoms¹⁸The F labeling method has the following disadvantages: 1) the labeling is carried out in a specific pH environment; 2) the labeled precursor has a large molecular weight; 3) in vivo defluorination phenomena, etc.

Because the dissociation energy of the P-F bond is equivalent to that of C-F (H at 25℃)₂P-F bond dissociation energy 461.5 + -10.5 kjmol^-1，CH₃-F bond dissociation energy 460. + -. 8.4kjmol-1), thus constructing P-¹⁸F replaces the conventional C-¹⁸The F labeling method has great application prospect. In the 20 th century, 60 s, synthesis by solid phase reaction at 800 ℃ was reported¹⁸F and³²p binuclear element labeled Na³²PO₂ ¹⁸F. In 2005, the passage of phosphorus oxychloride at room temperature was reported¹⁸F]F^-Construction of P-containing compounds by halogen metathesis¹⁸F, radiochemical yield as high as 96%, unfortunately P^-18F showed poor stability. Thus, improving the stability of the P-F bond is critical for the labeling of P-F.

Disclosure of Invention

The invention realizes ultrafast speed by a nucleophilic substitution strategy by using 1,3, 2-phosphorodithioic cyclopentane compounds as precursorsRapid, gentle construction of P-¹⁸F or P-¹⁹And (4) a F bond. The electronegativity of P is reduced by means of sulfurization and anions are introduced into the inner salt to improve stability. The ultra-fast-mild labeling method is applied to various biological molecules such as polypeptide, nucleoside and the like¹⁸F, obtaining a batch of novel early disease diagnosis probes with simple preparation, high specific activity, good in vivo stability and high targeting according to the method.

The first purpose of the invention is to provide a fluoro dithiophosphate compound, which has a structure shown in a formula (I):

wherein R is selected from:

wherein m, n, o, p, q, r, s are all 0-7, X is selected from-H, -CN, -NO₂，-OCH₃，-OC₆H₅，-N(CH₃)₂or-CHO.

The fluoro dithiophosphate compound provided by the invention has extremely high stability in vivo, small volume and low molecular weight, can be used for marking and imitating phosphate groups in a way of being extremely similar to phosphate groups, and is also an excellent marked artificial limb, and can realize radioactive marking on various polypeptides, proteins and metal molecules under mild conditions.

As a particularly typical technical scheme of the invention, the fluorinated dithiophosphate compound has a structure of one of the following:

the two fluoro phosphorodithioate compounds are structurally simplest compared with other structures and represent two cases of non-conjugation and conjugation of substituents, and the properties of the labeling and the labeling reaction can be better studied by taking the two cases as examples. Both the conjugated structure and the non-conjugated structure exhibit higher labeling efficiency in pure organic solvents, but the non-conjugated structure exhibits higher labeling efficiency under aqueous conditions compared to the conjugated structure. Comprises¹⁹The fluoro-dithiophosphate compound of F can achieve similar effects by connecting other fluorescent structures or labeling forms.

The second object of the present invention is to provide a precursor compound of the above-mentioned fluorodithiophosphate compound, which has a structure represented by formula (ii):

wherein R is defined as above for the fluoro dithiophosphate compound.

The invention takes the structure shown in the formula (II) for the first time, namely 1,3, 2-dithiophospholane as the fluoro dithiophosphate compound^18/19F marks the precursor, and a positron nuclide probe is constructed by a nucleophilic substitution marking strategy, so that the method has obvious advantages in the aspect of marking conditions. Compared with the traditional marking method, the marking method has the advantages of higher marking rate, shorter marking time and better marking water resistance, and can still maintain certain marking efficiency under the water content of 20 percent.

The third object of the present invention is to provide a method for preparing the precursor compound, which specifically comprises the following steps:

(1) carrying out addition reaction on the compound a and 1, 2-ethanedithiol to obtain a compound b;

(2) performing nucleophilic substitution reaction on the compound b and alcohol to obtain a compound c;

(3) performing addition reaction on the compound c and sulfur powder to obtain a precursor compound d;

wherein R is defined as above for the fluoro dithiophosphate compound.

Preferably, step 1) uses benzene as a solvent, preferably anhydrous benzene.

Preferably, both step 2) and step 3) use dichloromethane as solvent.

As a more desirable embodiment, the preparation of the precursor compound comprises the steps of:

1) placing the compound a in a reaction bottle, adding anhydrous benzene under the protection of argon, dropwise adding 1, 2-ethanedithiol into the reaction bottle under the condition of ice-water bath, reacting for more than 10 minutes, reacting for 1-3 hours at room temperature, filtering, and performing reduced pressure spin drying;

2) placing the compound b and 5-ethylthio tetrazole in a reaction bottle, adding anhydrous dichloromethane, adding an alcohol compound, reacting at room temperature for 2-3 hours, and performing reduced pressure spin drying;

3) and (3) placing the compound c in a reaction bottle, adding anhydrous dichloromethane, adding excessive sulfur powder, reacting overnight, performing reduced pressure spin-drying, adding a small amount of ethyl acetate, filtering to remove excessive sulfur powder, and performing column chromatography purification to obtain a precursor compound d.

Further, the reaction time period in step 3) is preferably 24 hours.

The fourth object of the present invention is to provide a method for preparing the above fluorinated dithiophosphate compound:

the precursor compound is used as a raw material, and the fluoro dithiophosphate compound is prepared by a nucleophilic substitution marking strategy;

or the precursor compound is prepared by the preparation method, and then the fluoro dithiophosphate compound is prepared by a nucleophilic substitution marking strategy.

Preferably, the label is a nucleophilic substitution label¹⁹The strategy of F is as follows: the precursor compound and tetrabutylammonium fluoride are used as raw materials.

In particular, nucleophilic substitution labels¹⁹The strategy of F is as follows: placing the compound shown in the formula (II) in a reaction bottle, adding anhydrous tetrahydrofuran, adding tetrabutylammonium fluoride, reacting for 2 minutes at room temperature, performing reduced pressure spin-drying, and purifying by column chromatography to obtain the compound¹⁹F labeled fluoro dithiophosphate compounds.

Preferably, the label is a nucleophilic substitution label¹⁸The strategy of F is as follows: under the condition of pure organic phase, the precursor compound of the formula (II) is used as raw material warp¹⁸F]F^-By nucleophilic substitution¹⁸F label, or directly using the aqueous solution of fluoride ion and the precursor compound of formula (II)¹⁸F]F^-By nucleophilic substitution¹⁸And F, labeling, wherein the reaction temperature in the nucleophilic substitution process is 20-80 ℃, and the reaction time is 10-300 seconds.

Preferably, the reaction temperature in the nucleophilic substitution process is 20-37 ℃ and the reaction time is 10-30 seconds.

More preferably, the reaction temperature during nucleophilic substitution is 20 ℃ and the reaction time is 30 seconds.

The fluoro dithiophosphate compound can be directly prepared in the aqueous solution of fluoride ions and can be simultaneously prepared with the existing fluoro dithiophosphate compound¹⁸Compared with the F labeling method, the reaction conditions are milder, the reaction time is greatly shortened to 30 seconds, the labeling process is simpler, and the labeling efficiency is improved to a great extent.

In particular, the present invention provides 2 different nucleophilic substitutions depending on the labeling conditions¹⁸F, marking strategy:

the method comprises the following steps: under the condition of pure organic phase, the precursor compound of the formula (II) is used as raw material warp [ 2 ]¹⁸F]F^-By nucleophilic substitution¹⁸And F, marking.

With 8mg of 4, 7, 13, 16, 21, 24-hexaoxa-1, 10-diazabicyclo [8.8.8 ]]Twenty sixAlkane (K2.2.2) and 1mg K₂CO₃The aqueous acetonitrile solution of (1), (2)¹⁸F]F^-Enriching QMA-Sep-Pak column, azeotropic dewatering with acetonitrile, and adding K₂CO₃，K2.2.2，[¹⁸F]F^-The mixture (2) and 0.5mg of the labeled precursor compound of formula (II) are dissolved in 100. mu.L of acetonitrile solvent and reacted at 20 to 80 ℃ for 10 to 300 seconds. The reaction was stopped, and about 10mL of water was added to dilute the reaction system, and the activated Sep-Pak C18 column was used to collect the filtrate in vial No. 1 (mainly, non-reacted¹⁸F]F^-) Then, the column was washed with 10mL of water, and the filtrate was collected into No. 2 bottle (ensuring that no reaction was involved [ [ solution ] ])¹⁸F]F^-Thoroughly rinsing and cleaning), blowing and drying a Sep-Pak C18 column by using nitrogen, washing a C18 column by using 2mL of acetonitrile, collecting filtrate into a No. 3 bottle, blowing and concentrating the nitrogen, separating and purifying by HPLC, and blowing and drying the acetonitrile in the solution by using nitrogen to obtain the product¹⁸F-labeled end product of formula (I), with greater than 99% radiochemical purity of the label.

The second method comprises the following steps: directly takes the fluorinion water solution and the precursor compound of the formula (II) as raw materials¹⁸F]F^-By nucleophilic substitution¹⁸And F, marking.

Comprises¹⁸F]F^-The aqueous solution of fluoride ion (10. mu.L) and 0.5mg of the labeled precursor compound of formula (II) are dissolved in 100. mu.L of acetonitrile solvent and reacted at 20 to 80 ℃ for 10 to 300 seconds. The reaction was stopped, and about 10mL of water was added to dilute the reaction system, which was then passed through a Sep-Pak C18 column, and the filtrate was collected into bottle No. 1 (mainly, non-reacted product [, ]¹⁸F]F^-) Then, the column was washed with 10mL of water, and the filtrate was collected into No. 2 bottle (ensuring that no reaction was involved [ [ solution ] ])¹⁸F]F^-Thoroughly rinsing and cleaning), blowing and drying a Sep-Pak C18 column by using nitrogen, washing the Sep-Pak C18 column by using 2mL of acetonitrile, collecting filtrate into a No. 3 bottle, blowing and concentrating the nitrogen, separating and purifying by HPLC (high performance liquid chromatography), and blowing and drying the acetonitrile in the solution by using nitrogen to obtain the product¹⁸F-labeled end product of formula (I), with greater than 99% radiochemical purity of the label.

The above preparation method provided by the present invention, HPLC purification operation, is a conventional technical means in the art, and the present invention is not particularly limited thereto.

The above preparation methods, the relative amounts of raw materials and solvents, and the operations such as reduced pressure spin-drying and column chromatography purification provided by the present invention are all conventional technical means in the field, and the present invention is not particularly limited thereto.

The fifth purpose of the invention is to provide the application of the fluorinated dithiophosphate compound or the precursor compound thereof in positron emission tomography; the preferred application in the preparation of the positron emission imaging agent comprises the preparation of phosphate analogue probes and positron drugs of heat-sensitive and solvent-sensitive biomolecules (preferably polypeptides or proteins and the like).

Preferably, the phosphoanalogs are phosphotyrosine and adenosine monophosphate.

Preferably, the polypeptide is c (rgdfk) [ cyclo (Arg-Gly-Asp-dpe-Lys) ], octreotide, PSMA ligand and/or aptamer.

The sixth purpose of the present invention is to provide an application of the above fluorinated dithiophosphate compound as a prosthetic limb (linker) for labeling and modifying protein, polypeptide or metal nanoparticles under mild conditions:

wherein R is as defined above for the fluorinated dithiophosphate compound, and R²Is a protein, polypeptide or metal nanoparticle.

The labeling means may be a form of disulfide bond formation or coordinate bond formation with a metal.

The invention at least realizes the following beneficial effects:

(1) the invention improves the in vivo stability of the fluorophosphate ester by reducing the electronegativity of P through a sulfurization method and introducing anions to form an inner salt so as to improve the stability.

(2) 1,3, 2-dithiophospholane is used as a labeling precursor for the first time, and a nucleophilic substitution labeling strategy is adopted, so that the labeling condition is mild, the labeling can be carried out at about 20 ℃, the specific activity is high, the labeling speed is very high, and only 30 seconds are needed.

(3) For the first time, for positron emission imaging agents, the innovative use of fluorodithiophosphates¹⁸And F, marking.

(4) The novel functional diversity of the fluorodithiophosphate structures, which closely resemble the phosphate structures, can be used to mimic and label phosphate analogs.

Drawings

FIG. 1 is the compound [ 2 ] prepared in example 2¹⁸F]4 is shown in¹⁸HPLC results of F-labeled fluorodithiophosphates;

FIG. 2 is the compound prepared in example 5¹⁸F]7 is shown in¹⁸HPLC results of F-labeled fluorodithiophosphates;

FIG. 3 is the compound [ 2 ] prepared in example 13¹⁸F]26 to¹⁸HPLC results of F-labeled fluorodithiophosphates;

FIG. 4 is the compound [ 2 ] prepared in example 15¹⁸F]29, HPLC results of 18F-labeled fluorodithiophosphates;

FIG. 5 shows the compound [ 2 ] in Experimental example 1¹⁸F]4 and [ 2 ]¹⁸F]FIG. 7 is a graph showing the relationship between the time and radiochemical yield, where the abscissa is the time (unit: seconds) and the ordinate is the yield (unit: percent);

FIG. 6 is a graph showing the labeling time and radiochemical yield of the conventional labeling method for the control group in Experimental example 1;

FIG. 7 shows a compound [ 2 ] in Experimental example 2¹⁸F]4 and [ 2 ]¹⁸F]7 a schematic diagram of the relationship between the temperature and water content (1-acetonitrile ratio) and radiochemical yield, the abscissa being the acetonitrile ratio (unit: percent) and the ordinate being the yield (unit: percent);

FIG. 8 is a graph showing the relationship between the temperature and water content of a conventional labeling method and the radiochemical yield of a control in Experimental example 2;

FIG. 9 shows Compound [ 2 ] provided in Experimental example 3¹⁸F]4, a schematic diagram of a positron emission dynamic 60-minute imaging result in a normal Balb/C mouse;

FIG. 10 is the compound [ 2 ] provided in experimental example 3¹⁸F]4 time-activity curve in normal Balb/C mice, where the abscissa is time (unit: min) and the ordinate is the percentage of radioactivity injected per gram of tissue (unit:% ID/g);

FIG. 11 is the compound [ 2 ] provided in experimental example 4¹⁸F]7 in normal Balb/C mouse body positron emission dynamic 60 minutes imaging result;

FIG. 12 is the compound [ 2 ] provided in experimental example 4¹⁸F]7 time-activity curves in normal Balb/C mice, where the abscissa is time (unit: min) and the ordinate is the percentage of radioactivity injected per gram of tissue (unit:% ID/g);

FIG. 13 is the compound [ 2 ] provided in experimental example 5¹⁸F]Positron emission 15 minute blood pool imaging of 7-labeled human serum albumin (HAS) in normal Balb/C mice;

FIG. 14 is the compound [ 2 ] provided in experimental example 6¹⁸F]7, emitting positive electron of the marked nano palladium sheet in 4T1 female Balb/C mouse for 120 min tumor imaging result;

FIG. 15 is the compound [ 2 ] provided in experimental example 7¹⁸F]29 positron emission in MDA-MB-453 subcutaneous tumor nude mice for 15 min tumor imaging.

Detailed Description

The following examples are intended to illustrate the invention but are not intended to limit the scope of the invention.

Example 1

The present embodiment provides a¹⁹The preparation process and the specific steps of the F-labeled fluoro dithiophosphate compound are as follows:

synthesis procedure for Compound 1:

10mmol of dichloro (diethylamino) phosphine is placed in a reaction bottle, 40mL of anhydrous benzene is added under the protection of argon, 10mmol of 1, 2-ethanedithiol is dropwise added into the reaction bottle under the condition of ice-water bath for reaction for 30 minutes, the reaction is carried out for 2 hours at room temperature, and then filtration and reduced pressure spin drying are carried out.

Nuclear magnetic data for compound 1:

³¹P NMR(162MHz,CDCl₃):δ106.98.

synthesis procedure for Compound 2:

placing 5mol of the compound 1 and 6mmol of 5-ethylthio tetrazole in a reaction bottle, adding 40mL of anhydrous dichloromethane, adding 5mmol of 3-butyn-1-ol, reacting at room temperature for 3 hours, filtering, and performing reduced pressure spin drying.

Nuclear magnetic data for compound 2:

³¹P NMR(162MHz,CDCl₃):δ148.93.

synthesis procedure for Compound 3:

placing 5mmol of compound 2 in a reaction bottle, adding 20mL of anhydrous dichloromethane, adding excessive sulfur powder, reacting overnight, performing reduced pressure spin-drying, adding a small amount of ethyl acetate, filtering to remove excessive sulfur powder, and purifying by column chromatography.

Nuclear magnetic data for compound 3:

¹H NMR(400MHz,CDCl₃):δ2.07(t,1H,J＝27.5Hz),2.68(m,2H),3.69(m,4H),4.27(m,2H).¹³C NMR(101MHz,CDCl₃)δ79.59(s),70.34(s),65.65(d,J＝8.8Hz),41.44(s),20.42(d,J＝9.1Hz).³¹P NMR(161.9MHz,CDCl₃):δ125.84.

synthesis procedure for Compound 4:

placing 1mmol of compound 3 into a reaction bottle, adding 5mL of anhydrous tetrahydrofuran, adding 2mmol of tetrabutylammonium fluoride, reacting for 3 hours at room temperature, performing reduced pressure spin drying, and performing column chromatography purification to obtain¹⁹F-labeled fluoro phosphorodithioate 4.

Nuclear magnetic data for compound 4:

¹H NMR(400MHz,CDCl₃):δ1.04(t,12H),1.51(m,8H),1.69(m,8H),1.99(t,1H,J＝9.7Hz),2.66(m,2H),3.35(t,8H,J＝52.9Hz),4.25(m,2H).¹³C NMR(101MHz,CDCl₃):δ80.73(s),69.48(s),64.30(d,J＝7.9Hz),59.03(s),24.16(s),20.60(d,J＝8.3Hz),19.76(s),13.69(s).³¹P NMR(162MHz,CDCl₃):δ122.84(s)；116.03(s).¹⁹F NMR(376MHz,CDCl₃):δ-5.81(s),-8.75(s).

example 2

The embodiment provides a method for positron emission tomography¹⁸The synthesis steps of the F-labeled fluoro dithiophosphate compound and the compound 3 are the same as those in the example 1, and the specific labeling steps are as follows:

the method comprises the following steps: with 8mg K2.2.2 and 1mg K₂CO₃The aqueous acetonitrile solution of (1), (2)¹⁸F]F^-Enriching QMA-Sep-Pak column, azeotropic dewatering with acetonitrile, and adding K₂CO₃，K2.2.2，[¹⁸F]F^-The mixture of (4) and 0.5mg of Compound 3 were dissolved in 100. mu.L of acetonitrile solvent and reacted at 20 ℃ for 30 seconds. The reaction was stopped, and about 10mL of water was added to dilute the reaction system, which was then passed through a Sep-Pak C18 column, and the filtrate was collected into bottle No. 1 (mainly, non-reacted product [, ]¹⁸F]F^-) Then, the column was washed with 10mL of water, and the filtrate was collected into No. 2 bottle (ensuring that no reaction was involved [ [ solution ] ])¹⁸F]F^-Thoroughly rinsing and cleaning), blowing and drying a Sep-Pak C18 column by using nitrogen, washing a C18 column by using 2mL of acetonitrile, collecting filtrate into a No. 3 bottle, blowing and concentrating the nitrogen, separating and purifying by HPLC, and blowing and drying the acetonitrile in the solution by using nitrogen to obtain the product¹⁸F-labeled fluorodithiophosphate compound¹⁸F]4, as shown in FIG. 1, the tag radiochemical purity is greater than 99%.

Example 3 (product Structure same as example 2)

The second method comprises the following steps: comprises¹⁸F]F^-The aqueous fluoride ion solution of (2) and 0.5mg of Compound 3 were dissolved in 100. mu.L of acetonitrile solvent and reacted at 20 ℃ for 30 seconds. The reaction was stopped, and about 10mL of water was added to dilute the reaction system, which was then passed through a Sep-Pak C18 column, and the filtrate was collected into bottle No. 1 (mainly, non-reacted product [, ]¹⁸F]F^-) Then the column was washed with 10ml of water and the filtrate was collectedTo bottle No. 2 (ensure that no reaction will take place [, ] [, ]¹⁸F]F^-Thoroughly rinsing and cleaning), blowing and drying a Sep-Pak C18 column by using nitrogen, washing the Sep-Pak C18 column by using 2mL of acetonitrile, collecting filtrate into a No. 3 bottle, blowing and concentrating the nitrogen, separating and purifying by HPLC (high performance liquid chromatography), and blowing and drying the acetonitrile in the solution by using nitrogen to obtain the product¹⁸F-labeled fluorodithiophosphate compound¹⁸F]And 4, the radiochemical purity of the marker is more than 99 percent.

Example 4

The embodiment provides¹⁹The preparation process and the specific steps of the F-labeled fluoro dithiophosphate compound are as follows:

the synthesis procedure of compound 7 was the same as that of compound 4 in example 1.

Nuclear magnetic data for compound 5:

³¹P NMR(162MHz,CDCl₃):δ146.55.

nuclear magnetic data for compound 6:

¹H NMR(400MHz,CD₃OD):δ1.43(m,12H,c₁＝7.6Hz,J₂＝7.3Hz)；2.68(m,2H)；3.69(m,4H)；4.27(m,4H).

nuclear magnetic data for compound 7:

¹H NMR(400MHz,MeOD):δ1.04(t,12H),1.43(m,8H),1.68(m,8H),3.26(t,8H,J＝17.7Hz),7.16(t,1H,J₁＝14.3Hz),7.30(m,4H).¹³C NMR(101MHz,MeOD):δ152.49(d,J＝10.3Hz),128.69(s),123.76(s),121.55(s),58.34(s),23.59(s),19.37(s),12.72(s).³¹P NMR(162MHz,MeOD):δ118.27(s),111.39(s).¹⁹F NMR(376MHz,MeOD):δ-2.68(s),-5.61(s).

example 5

The embodiment provides a method for positron emission tomography¹⁸The synthesis steps of F-labeled fluoro phosphorodithioate compound, compound 6, are the same as in example 4, and the labeling steps are as follows:

the method comprises the following steps: compound [ 2 ]¹⁸F]7 and the compound of example 2¹⁸F]The labeling step 4 is the same as that of the labeling step, and the result is shown in FIG. 2, Compound 2¹⁸F]The radiochemical purity of 7 is more than 99 percent.

Example 6 (product Structure same as example 5)

The second method comprises the following steps: compound [ 2 ]¹⁸F]7 and the compound of example 3¹⁸F]4 the labeling step is the same, and the radiochemical purity of the label is more than 99 percent.

Example 7

The synthesis procedure of compound 10 was the same as that of compound 4 in example 1.

Compound 8 nuclear magnetic data:

³¹P NMR(162MHz,CDCl3):δ146.55.

compound 9 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃):δ4.88(tt,J＝12.7,6.3Hz,1H),3.78–3.58(m,4H),1.83–1.67(m,1H),1.58(td,J＝13.5,6.6Hz,1H),1.50–1.35(m,5H).

¹³C NMR(101MHz,CDCl₃):δ78.07(d,J＝9.8Hz),41.49(d,J＝13.6Hz),39.46(d,J＝6.0Hz),21.46(d,J＝3.1Hz),18.55(s),13.90(s).

³¹P NMR(162MHz,CDCl₃):δ118.86.

compound 10 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃)δ4.80(td,J＝12.7,6.3Hz,1H),3.42–3.33(m,8H),1.75–1.68(m,8H),1.56–1.37(m,12H),1.34(d,J＝6.2Hz,3H),1.04(t,J＝7.3Hz,12H),0.93(t,J＝7.1Hz,3H).

¹³C NMR(101MHz,CDCl₃)δ74.56(s,1H),59.02(s,4H),39.97(s,1H),24.21(s,5H),21.69(s,2H),19.78(s,7H),18.77(s,2H),14.15(s,2H),13.73(s,7H).

³¹P NMR(162MHz,CDCl₃)δ122.71(s,1H),115.92(s,1H).

¹⁹F NMR(376MHz,CDCl₃)δ0.33(s,1H),-2.59(s,1H).

example 8

The synthesis procedure for compound 13 was the same as for compound 4 in example 1.

Compound 11 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ146.25.

compound 12 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ121.07.

compound 13 nuclear magnetic data:

³¹P NMR(162MHz,CD₃OD):δ117.27(s),110.39(s).

example 9

The synthesis procedure for compound 16 was the same as for compound 4 in example 1.

Compound 14 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ147.87.

compound 15 nuclear magnetic data:

¹³C NMR(101MHz,CDCl₃:δ79.44(d,J＝9.6Hz),41.42(s),33.29(s),25.11(s),23.63(d,J＝13.3Hz).

³¹P NMR(162MHz,CDCl₃):δ118.14.

compound 16 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃)δ4.62(s,1H),3.43–3.30(m,8H),2.05(d,J＝11.6Hz,2H),1.87–1.62(m,12H),1.58–1.41(m,12H),1.04(t,J＝7.3Hz,12H).

¹³C NMR(101MHz,CDCl₃)δ76.31(d,J＝8.6Hz),59.04(s),33.84(s),25.49(s),24.19(s),19.74(s),13.66(s).

³¹P NMR(162MHz,CDCl₃)δ120.87(s),113.48(s).

¹⁹F NMR(376MHz,CDCl₃)δ-1.67(s),-4.59(s).

example 10

The synthetic procedure for compound 19 was the same as compound 4 in example 1.

Compound 17 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ151.61.

compound 18 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃):δ7.32(s,1H),7.30(s,1H),7.28(s,1H),6.88(t,J＝17.1Hz,3H),3.93–3.52(m,7H).

¹³C NMR(101MHz,CDCl₃):δ160.49(s,),151.83–151.63(m),129.79(s),114.01(s),107.95(s),55.45–55.25(m),41.97(s).

³¹P NMR(162MHz,CDCl₃):δ118.31.

compound 19 nuclear magnetic data:

¹H NMR(400MHz,MeOD)δ7.19(d,J＝7.2Hz,2H),6.88(d,J＝9.0Hz,2H),3.81(s,3H),3.31–3.23(m,8H),1.70(dt,J＝15.8,7.9Hz,8H),1.44(dt,J＝14.6,7.4Hz,8H),1.05(t,J＝7.3Hz,12H).

¹³C NMR(101MHz,MeOD)δ156.32(s),146.04(d,J＝10.7Hz),122.29(s),113.60(s),58.19(s),54.63(s),23.47(s),19.32(s),12.58(s).

³¹P NMR(162MHz,MeOD)δ121.77(s),114.94(s).

¹⁹F NMR(376MHz,MeOD)δ-10.36(s),-13.30(s).

example 11

The synthetic procedure for compound 22 was the same as compound 4 in example 1.

Compound 20 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ148.12

compound 21 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃):δ7.49–7.33(m,5H),5.20(d,J＝10.9Hz,2H),3.78–3.59(m,4H).

¹³C NMR(101MHz,CDCl₃):δ128.59(d,J＝4.5Hz,2H),128.33(s,2H),69.75(s,1H),41.42(s,2H).

³¹P NMR(162MHz,CDCl₃):δ121.30.

compound 22 nuclear magnetic data:

¹H NMR(400MHz,MeOD)δ7.38(ddd,J＝23.9,21.9,7.3Hz,5H),5.13(d,J＝9.4Hz,2H),3.31–3.24(m,8H),1.76–1.63(m,8H),1.52–1.39(m,8H),1.05(t,J＝7.3Hz,12H).

¹³C NMR(101MHz,MeOD)δ137.75(d,J＝9.6Hz),127.90(s),127.37(d,J＝6.8Hz),67.95(s),58.18(s),23.47(s),19.32(s),12.59(s).

³¹P NMR(162MHz,MeOD)δ124.06(s),117.22(s).

¹⁹F NMR(376MHz,MeOD)δ-7.29(s),-10.26(s).

example 12

The synthetic procedure for compound 25 is the same as for compound 4 in example 1, which only has one more deprotection process for preparing compound 26 from compound 25, without affecting the utility of the compound itself, as follows:

1mmol of compound 25 was placed in a reaction flask, 5mL of THF was added, 5mL of 1mol/L HCl was added dropwise under ice bath conditions, and after 10 minutes the ice bath was removed and the reaction was allowed to proceed at room temperature for 2 hours. Under the ice-bath condition, NaOH with the concentration of 5mol/L is dropwise added to adjust the pH value to 14, and the reaction is carried out for 2 hours at room temperature. After the reaction was completed, the pH was adjusted to neutral by adding hydrochloric acid and spin-dried under reduced pressure and purified by column chromatography.

Compound 23 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ148.12

compound 24 nuclear magnetic data:

¹³C NMR(101MHz,MeOD)δ172.64(s),156.35(s),149.95(d,J＝13.0Hz),134.68(s),130.10(s,37H),121.71(s),79.29(s),55.02(s),51.17(s),41.67(s),27.31(s).

³¹P NMR(162MHz,CDCl₃):δ121.04.

compound 25 nuclear magnetic data:

¹H NMR(400MHz,MeOD)δ7.45(dd,J＝200.4,29.1Hz,4H),4.39–4.25(m,1H),3.72(s,2H),3.35(s,3H),3.35–3.25(m,8H),1.71(dt,J＝16.2,8.3Hz,8H),1.54–1.33(m,17H),1.06(t,J＝7.1Hz,12H).

compound 26 nuclear magnetic data:

¹H NMR(400MHz,D₂O)δ7.53–7.26(m,4H),4.03(dd,J＝8.1,5.1Hz,1H),3.39(dd,J＝14.4,5.1Hz,1H),3.25–3.07(m,1H).

example 13

Compound [ 2 ]¹⁸F]25 with the Compound [ 2 ] of example 2¹⁸F]4 same, this example only has one more step consisting of the compound [18F ]]25 preparation of the Compound [ 2 ]¹⁸F]26, the application of the compound per se is not influenced, and the deprotection process is as follows:

will be provided with¹⁸F-labeled Compound [ alpha ], [ alpha¹⁸F]25 placed in a reaction flask, 10. mu.L of HCl was added and reacted for 10 minutes. Blowing the mixture to dry by nitrogen at room temperature, dropwise adding NaOH with the concentration of 5mol/L to adjust the pH value to 14, reacting for 10 minutes at room temperature, adding hydrochloric acid to adjust the pH value to be neutral, and purifying by HPLC to obtain the product¹⁸F-labeled Compound [ alpha ], [ alpha¹⁸F]26, HPLC purification results are shown in FIG. 3, and the label is more than 99% pure.

Example 14

The synthesis procedure for compound 29 was the same as in example 12:

compound 26 nuclear magnetic data:

³¹P NMR(162MHz,CDCl₃):δ148.54

compound 27 nuclear magnetic data:

¹H NMR(400MHz,CDCl₃)δ8.37(s,1H),8.07(s,1H),6.66(s,2H),6.20(s,1H),5.48(s,1H),5.10(s,1H),4.60(s,1H),4.31(s,2H),3.52(ddd,J＝20.0,13.7,7.8Hz,4H),1.62(s,3H),1.41(s,3H).

³¹P NMR(162MHz,CDCl₃):δ121.34.

compound 28 nuclear magnetic data:

compound 29 nuclear magnetic data:

¹H NMR(400MHz,DMSO)δ8.71(s,1H),8.30(s,1H),7.93(s,2H),5.99(s,1H),4.68–4.56(m,1H),4.28–4.02(m,5H),3.45(d,J＝14.4Hz,1H).

example 15

Compound [ 2 ]¹⁸F]The synthesis procedure for 29 was the same as in example 13, and the deprotection procedure was as follows:¹⁸f-labeled [ alpha ], [ alpha¹⁸F]28 placing in a reaction flask, adding 10 μ L TFA, reacting for 10 min, adding NaOH to adjust pH to neutral, and purifying by HPLC¹⁸F-labeled Compound [ alpha ], [ alpha¹⁸F]29, results are shown in FIG. 4, with greater than 99% tag amplification.

In order to further verify the invention¹⁸The invention also provides the following experimental examples of the application effect of the F-labeled fluoro dithiophosphate compound:

experimental example 1

Subject: compound [ 2 ]¹⁸F]4 and [ 2 ]¹⁸F]7 of¹⁸F time of labelling as a function of yield.

Control subjects: the labeling time and radiochemical yield of conventional labeling methods.

The experimental method comprises the following steps:

adding no carrier¹⁸F]F^-And (4) carrying out azeotropic drying for 3 times. About 0.2nmol of the precursor is dissolved in 100. mu.L of acetonitrile and added to the solution containing the azeotropically dried [ 2 ]¹⁸F]F^-Penicillin bottle. The mixture was incubated at room temperature for 10-300 seconds and the radiochemical yield was determined by radio-HPLC. As shown in FIG. 5, the yield reached 90% or more at 10 seconds and reached a maximum of 97% to 98% at 30 seconds. The labeling time and yield of the conventional labeling method of the control group are shown in FIG. 6, the reaction time is usually 5-30 minutes, and the yield is 7% -90%. The labeling time and radiochemical yield of conventional labeling methods are derived from Angew. chem. int. Ed.2019,58, 2580-2605 (Chemistry for Positron Emission biology: Recent Advances in¹¹C^-,18F^-,13N^-,and ¹⁵O^-Labeling Reactions)。

Experimental example 2

Subject: compound [ 2 ]¹⁸F]4 and [ 2 ]¹⁸F]7 of¹⁸F, the relation between the temperature and the water content and the yield.

Control subjects: conventional labeling methods label temperature, water content, and yield.

The experimental method comprises the following steps:

adding no carrier¹⁸F]F^-An aqueous solution. About 0.2nmol of the precursor is dissolved in 100. mu.L of acetonitrile and an appropriate amount of carrier-free additive is added directly¹⁸F]F^-Aqueous solution (adjusted according to the water content ratio [ ] [, ]¹⁸F]F^-The amount of aqueous solution). The mixture was incubated at the indicated temperature for 30 seconds and the radiochemical yield was determined by radio-HPLC. As shown in FIG. 7, the reaction in the pure organic phase can obtain high radiochemical yield (97%) at room temperature and still has satisfactory radiochemical yield under the condition of 0-20% of water content. The reaction conditions and yields of the conventional labeling methods of the control group are shown in FIG. 8, and the conventional methods often require heating at high temperature and cannot be carried out under aqueous conditions¹⁸F-tag, which needs to be reacted in a pure organic phase. The relationship between the labeling temperature, the water content and the yield of the conventional labeling method is derived from Angew. chem. int. Ed.2019,58, 2580-2605 (Chemistry for Positron Emission Tomography: Recent Advances in¹¹C^-,¹⁸F^-,13N^-,and¹⁵O-Labeling Reactions)。

Experimental example 3

Subject: normal Balb/C mice.

Experimental reagent:

experimental groups: example 3 prepared [ alpha ], [¹⁸F]4 is shown in¹⁸F-labeled fluorodithiophosphates.

The experimental method comprises the following steps:

the product of example 3 is prepared¹⁸F]4 about 100 muL/100 muCi per mouse is injected into normal male Balb/C mice (weight 25g-30g) through tail vein, then 60 minutes of positron emission imaging is carried out, then 15 minutes, 30 minutes and 60 minutes of radioactivity uptake in bone and bladder are calculated respectively, and the experiment is repeated three times, and the experimental result is shown in figure 9 and figure 10.

The results of both FIG. 9 and FIG. 10 illustrate¹⁸F-labeled fluorodithiophosphate analog [ 2 ]¹⁸F]4 in vivo, with time, without significant bone uptake, indicating that it is stable in vivo, not susceptible to defluorination, and rapidly cleared by metabolic organs.

Experimental example 4

Subject: [¹⁸F]7 distribution in normal Balb/C mice.

Experimental reagent:

experimental groups: example 5 prepared [ alpha ]¹⁸F]7 is shown in¹⁸F-labeled fluorodithiophosphates.

The experimental method comprises the following steps:

prepared as in example 5¹⁸F]7 at a dose of about 100. mu.L/100. mu. Ci per mouse, the injection is performed into normal male Balb/C mice (weight 25g-30g) via tail vein, and then positron emission imaging is performed for 60 minutes, and then the uptake of radioactivity into bone and liver is calculated for 15 minutes, 30 minutes and 60 minutes respectively, and the experiment is repeated three times. The experimental results are shown in fig. 11 and 12.

The results of both FIG. 11 and FIG. 12 illustrate¹⁸F-labeled fluorodithiophosphate analog [ 2 ]¹⁸F]7 become longer in vivo over time and no significant bone uptake indicates that it is stable in vivo, is not susceptible to defluorination, and can be rapidly cleared by metabolic organs.

Experimental example 5

Subject: [¹⁸F]7-labeled human serum albumin (HAS) was visualized in the blood pool of normal Balb/C mice.

Experimental reagent:

example 5 prepared [ alpha ]¹⁸F]7 is shown in¹⁸F-labelled fluorodithiophosphate and HAS.

The experimental method comprises the following steps:

the product of example 5 is prepared¹⁸F]7 was reacted with 2mg of human serum albumin (HAS) at room temperature for 30 minutes using pure water as a solvent (200. mu.L) to give [ 2 ], [ solution ]¹⁸F]7 labeled human serum albumin at a dose of about 100. mu.L/100. mu. Ci per mouse¹⁸F]7-labelled HAS injectionNormal male Balb/C mice (body weight 25-30g) were followed by 60 min positron emission imaging and the experiment was repeated three times. The results of the experiment are shown in FIG. 13.

Experimental example 6

Subject: compound [ 2 ]¹⁸F]Tumor imaging of 7-labeled nano-palladium sheets in 4T1 subcutaneous tumor female Balb/C mice.

Experimental reagent:

example 5 the prepared Compound¹⁸F]7 is shown in¹⁸F marked fluoro phosphorodithioate and nano palladium sheet with size of 40 nm.

The experimental method comprises the following steps:

the product of example 5 is prepared¹⁸F]Dissolving 7 and 200mg of 40nm nano palladium sheet in 100 μ L of water, and reacting at room temperature for 15 minutes to obtain the product¹⁸F]7-labeled nano palladium sheet. The dosage of [ 2 ] in a dose of about 100. mu.L/100. mu. Ci per mouse¹⁸F]7 tail veins are injected into 4T1 subcutaneous tumor female Balb/C mice (weight is 25g-30g), then 120 minutes of positron emission imaging is carried out, and the experiment is repeated three times. The results of the experiment are shown in FIG. 14.

Experimental example 7

Subject: compound [ 2 ]¹⁸F]29 tumors were imaged in nude mice with MDA-MB-453 subcutaneous tumors.

Experimental reagent:

example 5 prepared [ alpha ]¹⁸F]29 shown in¹⁸F-labeled fluorodithiophosphates.

The experimental method comprises the following steps:

the product of example 5 is prepared¹⁸F]29 into 4T1 subcutaneous female Balb/C mice (body weight of about 20 g) at a dose of about 100. mu.L/100. mu. Ci per mouse by tail vein injection, followed by dynamic positron emission imaging for 60 minutes, and the experiment was repeated three times. The results of the experiment are shown in FIG. 15.

Claims

1. fluorodithiophosphoric acid ester compound is characterized in that, has the structure shown in formula (I):

where R is selected from:

Wherein m, n, o, p, q, r, s are all 0-7; X is selected from -H, -CN, -NO ₂ , -OCH ₃ , -OC ₆ H ₅ , -N(CH ₃ ) ₂ or -CHO.

2. the preparation method of the described fluorodithiophosphoric acid ester compound of claim 1, is characterized in that: described preparation method comprises the steps:

The fluorodithiophosphoric acid ester compound is prepared by nucleophilic substitution reaction between the compound represented by the formula (II) and [ ¹⁸ F]F ^- ;

Wherein, R refers to the same as claimed in claim 1.

3. the preparation method of fluorodithiophosphoric acid ester compound according to claim 2 is characterized in that, the concrete steps ^of the nucleophilic substitution reaction of compound shown in formula (II) and [ ¹⁸ F]F- are as follows:

Under the condition of pure organic phase, the above-mentioned precursor compound formula (II) is used as the raw material to carry out ¹⁸ F labeling by [ ¹⁸ F]F ^- nucleophilic substitution, or the fluoride ion aqueous solution and the precursor compound formula (II) are directly used as the raw material for 18 F labeling. [ ¹⁸ F]F ^- nucleophilic substitution for ¹⁸ F labeling.

4. The preparation method of fluorodithiophosphoric acid ester compound according to claim 3, characterized in that, in the nucleophilic substitution reaction process, the reaction temperature is 20～80 DEG C, and the reaction time is 10～300 seconds .

5 . The preparation method of fluorodithiophosphoric acid ester compound according to claim 4 , wherein, in the nucleophilic substitution reaction process, the reaction temperature is 20～37° C., and the reaction time is 10～30 seconds. 6 . .

6. The application of the fluorodithiophosphoric acid ester compound according to claim 1 in the preparation of a positron emission imaging agent.

7. The use according to claim 6, wherein the positron emission imaging agent is a phosphate analog probe.

8. The application according to claim 7, wherein the phosphoric acid analogs are phosphotyrosine and adenosine monophosphate.

9. The application of the fluorodithiophosphate compound of claim 1 in labeling polypeptides and proteins for preparing positron emission imaging agents.

10. The use according to claim 9, wherein the polypeptide is c(RGDfK) [loop (Arg-Gly-Asp-dPhe-Lys)], octreotide, PSMA ligand and/or aptamer.

11. The fluorinated phosphorodithioate compound of claim 1 marks and modifies proteins, polypeptides or metal nanoparticles as prostheses under mild conditions, and the specific marking methods are as follows:

Wherein, R refers to the same as described in claim 1, and R ² is a protein, a polypeptide or a metal nanoparticle.