Radionuclide-labeled polymer as well as preparation method and application thereof

Abstract

The invention discloses a radionuclide-labeled polymer as well as a preparation method and application thereof. Specifically, the invention provides a radionuclide labeled polymer which has the following structure: G-L1-L2-P, wherein G is a radionuclide-containing group and comprises a chelating group and a radionuclide, the chelating group is chelated with the radionuclide, and L1 and L2 are independently chemical bonds or linking groups; p is a temperature-sensitive structural unit. The radionuclide-labeled polymer can be used as a tracer of hydrogel, can be combined with the hydrogel under the condition that the property of the hydrogel is not changed, has good compatibility and stability, and further can be used for tracing implanted hydrogel and continuously tracking the position and state of the implanted hydrogel in real time.

Description

Technical Field

[0001] The invention relates to a radionuclide-labeled polymer, a preparation method and application thereof. Background Art

[0002] With the rapid development of biomedical technology, biocompatible hydrogels have been widely used in many biomedical fields such as filling, repair, drug delivery, interventional embolization, etc. For example, hydrogels have been used as advanced vascular embolization materials in the treatment of tumor diseases with rich blood supply, such as hepatocellular carcinoma.

[0003] There is no method to continuously and in real time track the state and position of hydrogels after implantation in vivo, which makes it extremely difficult to evaluate the performance of implantable hydrogels in vivo. Although existing angiography can image vascular embolism to a certain extent, the imaging time is limited and cannot continuously observe the state and position of implantable hydrogels, and cannot meet the imaging needs of hydrogel materials with longer implantation cycles. Summary of the invention

[0004] In view of the problem that the existing hydrogel materials cannot meet the imaging requirements of a longer implantation period, the present invention provides a radionuclide-labeled polymer, a preparation method and an application thereof. The radionuclide-labeled polymer provided by the present invention can be used as a tracer for hydrogels, does not change the properties of the hydrogel itself, has good compatibility with the hydrogel, has good binding stability, can maintain a high radioactivity purity in vivo and in vitro for more than 11 days, can be used as a tracer for hydrogel implantation in vivo, is used to continuously and in real time track the position and state of the hydrogel after implantation in vivo, and can be used for in vivo performance evaluation of implantable hydrogels.

[0005] The present invention provides a radionuclide labeled polymer having the following structure: G-L1-L2-P;

[0006] Wherein, G is a group chelated with a radionuclide, which includes a chelating group and a radionuclide, and the chelating group chelates with the radionuclide;

[0007] L1 and L2 are independently a chemical bond or a linking group;

[0008] P is a temperature-sensitive structural unit.

[0009] In one embodiment, G consists of a chelating group and a radionuclide.

[0010] In one embodiment, the radionuclide is 89 Zr;

[0011] The chelating group is a chelating group formed by a bifunctional chelating agent.

[0012] In one embodiment, L1 is a chemical bond or a connecting group represented by formula (I-1):

[0013] (I-1)

[0014] In formula (I-1), x and y are independently selected from integers of 0 to 3;

[0015] R1 is , C 1-5 The alkylene or chemical bonds;

[0016] R e for , substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 heteroarylene;

[0017] R d C 1-5 The alkylene or chemical bonds;

[0018] R b is hydrogen, hydroxyl or C 1-5 alkyl;

[0019] X is O or S; e represents the connection site between G and L1, i represents the connection site between R1 and R e , f represents the connection site between L1 and L2;

[0020] The L2 is a chemical bond, such as a group represented by formula (II-1) or (II-2):

[0021] (II-1)

[0022] (II-2),

[0023] Wherein, in formula (II-1), z is 0, 1, 2 or 3;

[0024] In formula (II-2), R2 and R3 are independently hydrogen, C 1-5 Alkyl, substituted or unsubstituted C 6-10 Aryl, or substituted or unsubstituted C 5-10 Heteroaryl, amide or thiol; f represents the connection site between L1 and L2, and h represents the connection site between L2 and P;

[0025] u is 0, 1, 2, or 3;

[0026] Each R 1-1 , R e-1 and R d-1 Independent for C1-5 of alkylene.

[0027] In one embodiment, P is a chitosan structural unit, an acrylamide structural unit or a group represented by formula (III-1):

[0028]

[0029] (III-1),

[0030] Wherein, in formula (III-1), a and c are independently selected from the range of 2-130; b is in the range of 10-80, and R is selected from hydrogen and C 1-5 alkyl.

[0031] In the present invention, in the compound represented by formula (I), each C 1-5 The alkylene group is methylene, , , or .

[0032] In the present invention, each C 6-10 Arylene is phenylene or naphthylene, for example phenylene.

[0033] In the present invention, each C 5-10 The heteroaryl group is preferably C 5-10 Heteroaryl, wherein the heteroatom is independently selected from one or two of N, O and S, and the number of the heteroatoms is independently 1, 2 or 3, preferably the heteroatom is N or O, and the number of the heteroatoms is 1 or 2, such as pyridine, pyrimidine or pyran.

[0034] In the present invention, each C 1-5 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl or sec-butyl, for example methyl, ethyl or isopropyl.

[0035] In the present invention, each substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 Heteroarylene, or substituted or unsubstituted C 6-10 Aryl, or substituted or unsubstituted C 5-10 The substituents in the heteroaryl group are independently C 1-3 alkyl, halogen, hydroxyl or cyano; said C 1-3 The alkyl group is methyl, ethyl or propyl;

[0036] The halogen is F, Cl, Br or I.

[0037] The chelating group may be a conventional chelating group in the art, preferably a chelating group formed by a bifunctional chelating agent, for example, the bifunctional chelating agent is DOTA, NOTA, NODA, NODAGA, DOTP, TETA, ATSM, PTSM, EDTA, EC, HBEDCC, DTPA, SBAD, BAPEN, DFO or TACN.

[0038] In one embodiment, the chelating group is , or .

[0039] In one embodiment, L1 is a linking group represented by formula (I-2) or (I-3):

[0040] (I-2),

[0041] (I-3)

[0042] Among them, in formula (I-2), R1 is or C 1-5 alkylene, i represents the connection with Re; R e is substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 Heteroarylene; wherein e represents connection with G, and f represents connection with L2;

[0043] In formula (I-3), each n is independently selected from an integer in the range of 0-10, R b It is hydrogen or hydroxyl.

[0044] In one embodiment, in formula (I-2), R1 or methylene i represents R e Connection site.

[0045] In one embodiment, in formula (I-2), R e is substituted or unsubstituted C 6-10 Arylene groups, such as phenylene groups;

[0046] In one embodiment, in formula (I-2), R1 is methylene or , and R e is phenyl, i represents e connect.

[0047] In one embodiment, in formula (I-3), each n is independently 1, 2, 3, 4 or 5, R b It is hydroxyl.

[0048] In one scheme, L1 is , or , e indicates connection with G, and f indicates connection with L2.

[0049] In one embodiment, in formula (II-1), z is 2.

[0050] In one embodiment, in formula (II-2), R2 and R3 are both methyl.

[0051] In one embodiment, in formula (II-2), u is 0.

[0052] In one embodiment, in formula (II-2), R2 and R3 are both methyl groups, and u is 0.

[0053] In one scheme, L2 is (For example )or .

[0054] In one embodiment, in formula (III-1), a and c are independently integers in the range of 80-130, and b is an integer in the range of 30-60; for example, a or c is 80 or 101, and b is 27 or 56.

[0055] In one embodiment, the acrylamide structural unit includes a group represented by formula (III-2):

[0056] (III-2)

[0057] In formula (III-2), R c is hydrogen, -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2, wherein m is an integer between 0 and 5, R a are independently -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2 is a divalent group formed by losing one hydrogen;

[0058] p or q is independently an integer in the range of 20-200.

[0059] In one embodiment, in formula (III-2), R c -CH-(CH3)2 or -(CH2) m -NH2, wherein m is 1, 2, 3, 4 or 5.

[0060] In one embodiment, in formula (III-2), R c It is -CH-(CH3)2 or -(CH2)2-NH2.

[0061] In one embodiment, in formula (III-2), Ra -(CH2) m -NH2 loses hydrogen to form a divalent group, wherein m is an integer between 0 and 5, for example -(CH2)2-NH-.

[0062] In one embodiment, in formula (III-2), p is an integer in the range of 100-150, and q is an integer in the range of 20-40; for example, when R c When it is -CH-(CH3)2, p is 110-125, q is an integer in the range of 20-25 or R c When it is -(CH2)2-NH2, p is an integer in the range of 120-135, and q is an integer in the range of 25-35.

[0063] In one embodiment, P is , , or .

[0064] In one embodiment, the P is derived from a polymer represented by formula (IV):

[0065]

[0066] (IV),

[0067] Wherein, in formula (IV), a, b, c and R are defined as above.

[0068] In one embodiment, in formula (IV), the average relative molecular weight of the polymer is 8000-13000 Da, such as 8600 Da-11500 Da, preferably 8600 Da or 11500 Da.

[0069] In some embodiments, the P is derived from a polymer represented by formula (V):

[0070] (V),

[0071] In formula (V), R c is hydrogen, -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2, wherein m is an integer within the range of 0-5, p is an integer within the range of 100-150, and q is an integer within the range of 20-40.

[0072] In one embodiment, the polymer satisfies the following embodiments (1) to (5):

[0073] Solution (1):

[0074] The chelating group is ;

[0075] The P is ; Wherein, R is a methyl group, a is an integer in the range of 75-110, and b is an integer in the range of 25-70;

[0076] The L1 is , where e indicates connection with G, and f indicates connection with L2;

[0077] The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P;

[0078] Solution (2):

[0079] The chelating group is ;

[0080] The P is , wherein R is methyl, a is an integer in the range of 75-110, and b is an integer in the range of 25-70;

[0081] The L1 is , where e indicates connection with G, and f indicates connection with L2;

[0082] The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P;

[0083] Solution (3):

[0084] The chelating group is ;

[0085] The P is , wherein R is methyl, a is an integer in the range of 75-110, and b is an integer in the range of 25-70;

[0086] The L1 is , where e indicates connection with G, and f indicates connection with L2;

[0087] The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P;

[0088] Solution (4):

[0089] The chelating group is ;

[0090] The P is or ;

[0091] The L1 is , where e indicates connection with G, and f indicates connection with L2;

[0092] The L2 is a chemical bond;

[0093] Solution (5):

[0094] The chelating group is ;

[0095] The P is or ;

[0096] The L1 is a chemical bond;

[0097] The L2 is a chemical bond.

[0098] In some embodiments of the present invention, the chemical bond is a covalent bond.

[0099] In a certain embodiment, the radionuclide-labeled polymer is any of the following structures:

[0100] , ,

[0101] , or , where a, b, c, p, q, R, R a and R c As mentioned above, for example

[0102] , ,

[0103] , or .

[0104] In some embodiments, the radionuclide-labeled polymer is any of the following structures:

[0105] ,

[0106] ,

[0107] ,

[0108] ,

[0109] ,

[0110] ,

[0111] ,

[0112] ,

[0113] or .

[0114] The present invention also provides a polymer as shown in formula A:

[0115] ;

[0116] Wherein, L1, L2, and P are defined as above, Q is a chelating group, and the chelating group is defined as above.

[0117] In one embodiment, the polymer as shown in formula A is any of the following structures:

[0118] , , , or

[0119] Wherein a, b, c, p, q, R, Rc and Ra are as described above, for example

[0120] , , , , , , , , or .

[0121] The present invention also provides a method for preparing a radionuclide-labeled polymer, which comprises the following steps: mixing and incubating the polymer represented by the above formula (A) with a solution containing a radionuclide to obtain a radionuclide-labeled polymer.

[0122] In one embodiment, in the preparation method, the radionuclide-labeled polymer is as described above.

[0123] In one embodiment, the preparation method comprises the following steps:

[0124] S1. Providing an oxalic acid solution containing radioactive metal ions to obtain a mixed solution 1;

[0125] S2. The mixed solution 1 was incubated with Na2CO3 solution at room temperature to obtain a mixed solution 2;

[0126] S3. Mixing the mixed solution 2 with the polymer represented by formula (A) and the buffer solution, incubating, and obtaining a product after the reaction is completed;

[0127] Wherein, in step S1., the oxalic acid solution solvent is water;

[0128] In step S2., the Na2CO3 solution solvent is water.

[0129] Preferably, the preparation method further comprises:

[0130] S0. Preparing the polymer to be labeled;

[0131] Wherein, step S0 includes the following steps:

[0132] S01. Synthesis of amino derivatives of temperature-sensitive polymers;

[0133] S02. The amino derivative is subjected to a chelating reaction with a chelating agent to generate a polymer represented by formula (A), wherein the polymer represented by formula (A) is a chelate of a polymer and a chelating group.

[0134] In one embodiment, S01 includes the following steps:

[0135] (1) reacting the amino-protected amino acid with the temperature-sensitive polymer in anhydrous dichloromethane in the presence of N,N'-dicyclohexylcarbodiimide and 4-dimethylaminopyridine to obtain a crude product;

[0136] (2) The crude product of step (1) is subjected to a deprotection reaction in anhydrous dichloromethane to obtain an amino derivative of a temperature-sensitive polymer.

[0137] In one embodiment, S02 includes the following steps:

[0138] (3) In a bicarbonate solution, the amino derivative in step (2) undergoes an addition reaction with a chelating agent to obtain a polymer A to be labeled;

[0139] Wherein, the amino-protected amino acid is Boc-protected valine;

[0140] The deprotection reaction is to remove Boc protection;

[0141] The bifunctional chelating agent is one or more of DFO and DOTA.

[0142] The present invention also provides a gel composition, which comprises the radionuclide-labeled polymer and the hydrogel preparation as described above.

[0143] The hydrogel preparation of the present application may be a conventional hydrogel preparation used in the art for in vivo vascular embolization, in vivo hemostasis, drug delivery, in vivo tissue filling or in vivo tissue repair, for example, preferably including a temperature-sensitive gelling agent and a gelling agent.

[0144] In a certain embodiment, the temperature-sensitive gelling agent is one or more of poloxamer, N-isopropylacrylamide polymer, chitosan or polyethylene glycol block copolymer.

[0145] In one embodiment, the gelling agent is one or more of sodium alginate, hydroxymethyl cellulose and CaCl2.

[0146] In one embodiment, the hydrogel formulation further comprises an imaging agent, such as iohexol.

[0147] In a certain embodiment, based on the total volume of the hydrogel preparation, the mass volume ratio of the temperature-sensitive gelling agent is 0.12-0.2g / mL, and the mass volume ratio of the gelling agent is 0.005-0.5g / mL, preferably the mass volume ratio of the temperature-sensitive gelling agent is 0.14-0.18g / mL, and the mass volume ratio of the gelling agent is 0.005-0.3g / mL. In a certain embodiment, the hydrogel preparation includes: 0.2-1g / mL iohexol, 0.02-0.1g / mL sodium alginate, 0.12-0.2g / mL poloxamer, 0.005-0.1g / mL hydroxymethyl cellulose or CaCl2, and water.

[0148] In one embodiment, the gelling agent is prepared by the following preparation method: sodium alginate and hydroxymethyl cellulose are dissolved in an aqueous solution of iohexol, heated to dissolve, and cooled to obtain; in the aqueous solution of iohexol, the mass ratio of iohexol to water is (1-5):3, for example, 2:3.

[0149] In a certain embodiment, the hydrogel preparation includes one or more of sodium alginate, poloxamer and hydroxymethyl cellulose. Preferably, the hydrogel preparation consists of iohexol, sodium alginate, poloxamer, hydroxymethyl cellulose and water.

[0150] In one embodiment, the mass volume ratio of the radionuclide labeled polymer to the hydrogel preparation is 0.01 mg / mL to 2 mg / mL, preferably 0.01 mg / mL to 1.5 mg / mL, more preferably 0.06 mg / mL to 1 mg / mL, such as 0.06 mg / mL.

[0151] In a certain embodiment, based on the volume of the gel composition, the gel composition consists of 0.06 mg / mL of the radionuclide-labeled polymer, 0.4 g / mL of iohexol, 0.02 g / mL of sodium alginate, 0.14 g / mL of poloxamer, 0.005 g / mL of hydroxymethylcellulose and the balance of water.

[0152] In a certain embodiment, the sol-gel transition temperature of the gel composition is 25-37°C, preferably 25.0-28.3°C, such as 25.5-27.5°C.

[0153] The present invention also provides a radionuclide-labeled polymer, which is prepared by the preparation method described above.

[0154] The present invention also provides a tracer, which comprises the radionuclide-labeled polymer as described above.

[0155] In a certain embodiment, the tracer is prepared by adding a polymer labeled with a radionuclide as described above to the hydrogel formulation as described above, and the tracer is a PET / CT imaging agent or a SPECT imaging agent, so that the in vivo distribution of the gel composition can be observed by PET / CT imaging.

[0156] In one embodiment, the tracer is used to evaluate the embolic properties of the gel composition as described above.

[0157] The present invention also provides an application of the gel composition as described above, wherein the application is application of the gel composition in preparing materials for in vivo vascular embolism, in vivo hemostasis or in vivo tissue repair.

[0158] In a certain embodiment, the application is for the preparation of an embolic agent, wherein the embolic agent may be an embolic agent for transcatheter arterial chemoembolization for treating tumors.

[0159] The present invention also provides a tracing method for an implantable hydrogel, which comprises the following steps:

[0160] (1) providing a hydrogel and adding a polymer labeled with a radionuclide as described above;

[0161] (2) implanted into the subject;

[0162] (3) Perform PET / CT imaging at multiple preset time points to check the imaging results and / or perform radioactivity counting.

[0163] The present application also provides a method for radioactively labeling a hydrogel preparation, comprising:

[0164] Providing a radionuclide-labeled polymer as described above;

[0165] The radionuclide labeled polymer is added to the hydrogel formulation as described above.

[0166] In one embodiment, the temperature-sensitive gelling agent and the radionuclide-labeled polymer have the same temperature-sensitive structural unit.

[0167] Preferably, when the temperature-sensitive gelling agent is poloxamer, P in the radionuclide-labeled polymer comprises a structure represented by formula (III-1);

[0168] When the temperature-sensitive gelling agent is an N-isopropylacrylamide polymer, P in the radionuclide-labeled polymer comprises a structure represented by formula (III-2).

[0169] In one embodiment, the subject is a human, a primate, or a non-primate mammal.

[0170] In one embodiment, the subject is an experimental animal model.

[0171] In addition to the foregoing, when used in the specification and claims of this application, unless otherwise specifically indicated, the following terms have the following meanings:

[0172] Herein, "temperature sensitivity" refers to the property of gelation from liquid or solidification from liquid to gel or solid as the temperature increases. "Hydrogel preparation" refers to a water-soluble or hydrophilic polymer or a composition containing such a polymer, and also includes a gel-state substance formed by the composition through certain chemical crosslinking or physical crosslinking; the hydrogel preparation may include a temperature-sensitive polymer, such as poloxamer or acrylamide polymers or other temperature-sensitive polymers known in the art, so the hydrogel preparation has temperature-responsive properties, such as conversion from sol state to gel state at a certain temperature; the hydrogel preparation can be used as a material for in vivo vascular embolism, in vivo hemostasis, drug delivery, in vivo tissue filling or in vivo tissue repair. The hydrogel preparation can be any hydrogel material for medical, pharmaceutical or cosmetic use.

[0173] As used herein, "phase transition temperature" refers to the temperature or temperature range at which a polymer undergoes a phase transition.

[0174] The term "polymer" refers to a molecule formed by the chemical combination of two or more oligomeric units. The chemical units are usually linked by covalent bonds. The two or more chemical units in a polymer can be all identical, in which case the polymer is called a homopolymer. They can also be a combination of different units, and these polymers are called copolymers.

[0175] The term "chemical bond" refers to the general term for the strong interaction force between two or more adjacent atoms (or ions) within a pure molecule or crystal, generally ionic bond, covalent bond, or metallic bond.

[0176] The term "linking group" refers to the part of an organic compound that has lost one or more atoms. It is a general term for atomic groups and radicals that have a connecting effect. A linking group usually contains a covalent bond, but a linking group is not a chemical bond.

[0177] The term "heteroaryl" refers to an aromatic system containing heteroatoms, wherein the number of the heteroatoms is 1, 2, 3 or 4, and the heteroatoms are selected from N, O or S. The heteroaryl groups include, but are not limited to, thiazolyl, thienyl, pyridyl and pyrimidinyl.

[0178] Without violating the common sense in the art, the above-mentioned preferred conditions can be arbitrarily combined to obtain the preferred embodiments of the present invention.

[0179] The reagents and raw materials used in the present invention are commercially available.

[0180] The beneficial effects of the present invention are as follows: the present invention provides a radionuclide-labeled polymer, which can be used for mixing with a thermosensitive hydrogel, does not change the physicochemical properties of the hydrogel itself, and can still maintain the original viscosity and thermosensitive curing properties of the hydrogel. The radionuclide-labeled signal can exist stably in the body for a long time, maintain a long half-life, and the continuous imaging period can reach more than 11 days. It can be used for in vivo tracing of embolic agents in hydrogels, and the distribution of hydrogel embolism in the body can be conveniently and continuously monitored without multiple angiography.

[0181] The radioactive nuclide-labeled polymer of the present invention is covalently linked to the polymer unit via a chelating group, and then chelates the radioactive nuclide via the chelating group, thereby having a better binding force with the hydrogel preparation. When used for radioactive labeling of implantable hydrogels, it has better stability and in vivo imaging time than non-covalent bonding methods such as direct addition of zirconium salt chelation or hydrogen bonding. BRIEF DESCRIPTION OF THE DRAWINGS

[0182] Figure 1 For B01 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer in vivo for 5 h;

[0183] Figure 2 For B01 dog 89 Coronal images of the dynamic in vivo coronal images of Zr-labeled poloxamer for 5 h;

[0184] Figure 3 For B01 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 1d in vivo;

[0185] Figure 4 For B01 dog 89 Coronal images of the dynamic in vivo Zr-labeled poloxamer 1d;

[0186] Figure 5 For B01 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 5d in vivo;

[0187] Figure 6 For B01 dog89 Dynamic coronal images of Zr-labeled poloxamer 5d in vivo;

[0188] Figure 7 For B01 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 7d in vivo;

[0189] Figure 8 For B01 dog 89 Dynamic coronal images of Zr-labeled poloxamer 7d in vivo;

[0190] Fig. 9 For B01 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 11d in vivo;

[0191] Fig.10 For B01 dog 89 Dynamic coronal images of Zr-labeled poloxamer 11d in vivo;

[0192] Fig.11 For B04 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer in vivo for 5 h;

[0193] Fig.12 For B04 dog 89 Dynamic coronal images of Zr-labeled poloxamer in vivo for 5 h;

[0194] Fig.13 For B04 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 1d in vivo;

[0195] Fig.14 For B04 dog 89 Coronal images of the dynamic in vivo Zr-labeled poloxamer 1d;

[0196] Fig.15 For B04 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 5d in vivo;

[0197] Fig.16 For B04 dog 89 Dynamic coronal images of Zr-labeled poloxamer 5d in vivo;

[0198] Fig.17 For B04 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 7d in vivo;

[0199] Fig.18 For B04 dog 89 Dynamic coronal images of Zr-labeled poloxamer 7d in vivo;

[0200] Fig.19For B04 dog 89 Dynamic cross-sectional images of Zr-labeled poloxamer 11d in vivo;

[0201] Fig. 20 For B04 dog 89 Dynamic coronal images of Zr-labeled poloxamer 11d in vivo;

[0202] Fig.21 for 89 Calibration curve between the detection values ​​of Zr activity meter and γ counter. DETAILED DESCRIPTION

[0203] The present invention is further described below by way of examples, but the present invention is not limited to the scope of the examples. The experimental methods in the following examples without specifying specific conditions are carried out according to conventional methods and conditions, or selected according to the product specifications.

[0204] Example:

[0205] Material:

[0206] Polymer 1:

[0207]

[0208] Manufacturer: BASF CORPORATION, model: Kolliphor® P 407Geismar.

[0209] Polymer 2:

[0210]

[0211] Manufacturer: BASF CORPORATION, model: Kolliphor® P 188 Geismar.

[0212] Chitosan: average relative molecular weight 5000Da, product number C434552, non-animal origin, purchased from Shanghai Aladdin Biochemical Technology Co., Ltd.

[0213] Iohexol: Manufacturer: Shanghai Stari Pharmaceutical Co., Ltd.

[0214] Sodium alginate: Manufacturer: Qingdao Mingyue Alginate Tissue Engineering Materials Co., Ltd.

[0215] 89 Zr:

[0216] Manufacturer: PerkinElmer, Inc.

[0217] Purity and content: After unpacking, the total activity concentration was detected to be 19.54mCi / mL.

[0218] Prescription composition: 89-Zr is dissolved in 1 mol / L oxalic acid solution.

[0219] Storage conditions: Store at room temperature in a 2 mL V-bottom screw-cap glass bottle in a lead container, and avoid ionizing radiation.

[0220] N-isopropylacrylamide monomer: from Shanghai Aladdin, product number I106818.

[0221] N-(3-aminopropyl) methacrylamide monomer (APAM monomer): Shanghai Aladdin product number N129096.

[0222] Beagle animal source: Nanjing Chaimen Biotechnology Co., Ltd.

[0223] Other reagents are commercially available unless otherwise specified.

[0224] Preparation Example 1 Radiolabeled Polymer 89 Preparation of Zr-DFO-P1-1

[0225] 1.1 Synthesis of thermosensitive polymer and DFO chelate

[0226] (1) Polymer amino activation

[0227] Boc-Val-OH (Boc-L-valine) 1.086 g, DCC (N,N'-dicyclohexylcarbodiimide) 1.91 g, DMAP (4-dimethylaminopyridine) 1.22 g, in a molar ratio of 1:2:2, were dissolved in 40 mL of ultra-dry anhydrous DCM (dichloromethane), reacted at room temperature under argon protection for half an hour, 5.75 g of polymer 1 (P1-1) was added, and the reaction was continued for 2 days. The DCM was concentrated by rotary evaporation, precipitated with ice ether, and the white precipitate was collected by filtration, re-dissolved with 3 mL of DCM, and precipitated with ice ether again. The precipitation and filtration were repeated three times, and the precipitate was re-dissolved in ultrapure water at a ratio of 1 mg:1 mL (heated), dialyzed for three days, and lyophilized to obtain 4.13 g of Boc-protected polymer P1-1-Val-Boc with a yield of 71.9%.

[0228] (2) Removal of Boc protection

[0229] Weigh 4 g of polymer P1-1-Val-Boc and dissolve it in 20 mL of DCM. Place it in an ice-water bath, add 45 mL of trifluoroacetic acid under argon protection, stir and react for 2 hours, evaporate the solvent, redissolve the product in chloroform, spin dry the product, dissolve it in hot water and dialyze it for three days, and freeze-dry to obtain 3.2 g of the deprotected polymer P1-1-Val with a yield of 80%.

[0230]

[0231] (3) Preparation of DFO chelate

[0232] 15.3 mg of polymer P1-1-Val was dissolved in 4 mL of 0.1 M NaHCO3 (sodium bicarbonate) solution, and p-SCN-Bn-DFO (CAS No.: 1222468-90-7) (2 mg; 200 μL DMSO) was added. The molar reaction ratio of polymer P1-1-Val to p-SCN-Bn-DFO was 1:2 and the mixture was slowly added dropwise. The mixture was reacted at room temperature for 1 h. After being spin-dried, it was dissolved in hot water and dialyzed for three days. After freeze-drying, a total of 10.2 mg of polymer DFO-P1-1 was obtained with a purity of 99.3% and a yield of 66.7%.

[0233] .

[0234] 1.2 89 Synthesis of Zr-labeled polymers

[0235]

[0236] 89 Zr-DFO-P1-1

[0237] (1) Prepare the marking solution

[0238] 89 Oxalic acid solution of Zr: Zirconium [ 89 Zr] was dispersed in 1 M oxalic acid solution (pH < 4) (PerkinElmer, Inc);

[0239] Preparation of 0.5 M HEPES (N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid) buffer: First, add 18 mL of ultrapure water to 20 mL of 1M HEPES buffer and check its pH value. If the pH value is <7.1, use 1M NaOH to adjust its pH value; if the pH value is >7.3, use 1M H2SO4 to adjust its pH value. When the pH value of the HEPES buffer is within the range of 7.1 to 7.3, continue to add a certain volume of ultrapure water to a total volume of 40 mL.

[0240] Preparation of polymer DFO-P1-1 solution (1 mg / mL): Accurately weigh 0.6 mg of polymer DFO-P1-1 and dissolve it in 600 µL of 10 mM pH 8.4 borate buffer solution.

[0241] (2) 89 Zr labeling reaction

[0242] A. Accurately measure 84 μL of 89 Zr oxalic acid solution into a glass reaction bottle;

[0243] B. Add 1 M oxalic acid solution to the glass reaction bottle to make the total volume in the reaction bottle 200 μL;

[0244] C. Use a pipette to accurately pipette 90 μL of 2M Na2CO3 solution into the reaction bottle and incubate at room temperature for 3 minutes;

[0245] D. While gently shaking the reaction bottle, dissolve 0.3 mL 0.5M HEPES (pH 7.1-7.3), 600 µL prepared polymer DFO-P1-1 solution (1 mg / mL), and then add 0.7 mL 0.5M HEPES to the reaction bottle in sequence (Note: the pH value of the reaction solution should be between 6.8 and 7.2 to obtain the best labeling efficiency. In addition, the metal needle of the syringe is prohibited in the solution preparation);

[0246] E. Incubate at room temperature for 1 hour while gently shaking the reaction bottle;

[0247] F. After the reaction is completed, use an ultrafiltration centrifuge tube to purify the reaction product to remove free 89 Zr (washing solution: 10 mM pH 8.4 borate buffer solution; 500 µL*2 times).

[0248] Preparation Example 2 Radiolabeled polymer 89 Preparation of Zr-DFO-P1-2

[0249]

[0250] Only polymer 1 (P1-1) in Preparation Example 1 was replaced by polymer 2 (P1-2), and the amount of polymer 2 was calculated according to the average relative molecular weight, and the reaction was carried out according to the steps of Preparation Example 1.

[0251] Preparation Example 3 Radiolabeled polymer 89 Preparation of Zr-DOTA-P1-1

[0252]

[0253] The reaction was carried out according to the procedure of Preparation Example 1 except that p-SCN-Bn-DFO (CAS No.: 1222468-90-7) in Preparation Example 1 was replaced with p-SCN-Bn-DOTA (CAS No.: 127985-74-4).

[0254] Preparation Example 4 Radiolabeled polymer 89 Preparation of Zr-DFO-P2

[0255] Chitosan was dissolved in 0.1M acetic acid solution, p-SCN-Bn-DFO (2mg; 200μL DMSO) was added, chitosan and p-SCN-Bn-DFO were reacted in a molar ratio of 1:2 and slowly added dropwise at 37°C for 4h, and then dissolved in hot water for dialysis for three days after spin drying. The product DFO-P2 was obtained after freeze drying. Then, the steps in step 1.2 of preparation example 1 were followed. 89 Radiolabeling of Zr 89 Zr-DFO-P2.

[0256] Preparation Example 5 Radiolabeled Polymer 89 Preparation of Zr-DOTA-P3-1

[0257] (1) Synthesis of Boc-protected N-isopropylacrylamide (NIPAM) polymer:

[0258] 3.69 g of N-isopropylacrylamide monomer (NIPAM), 1.76 g of N-hydroxysuccinimide (NHS) and 290 mg of azobisisobutyronitrile (AIBN) were dissolved in 10 mL of THF, and the mixture was reacted at 60 °C for 12 h to prepare pNIPAM-NHS, and then 80.6 g of N-Boc-ethylenediamine was added to react for 12 h, and then precipitated with 150 mL of ether, filtered, washed twice with ether, and dried to obtain pNIPAM-Boc with a yield of 83%;

[0259]

[0260] (2) Removal of Boc protection:

[0261] 2.4 g pNIPAM-Boc was dissolved in 50 mL trifluoroacetic acid aqueous solution (volume ratio of trifluoroacetic acid: water was 8:2), stirred at room temperature for 2 h, vacuum rotary dried, redissolved in water, purified by Sephadex G25 column, and freeze-dried to obtain pNIPAM with a yield of 90%;

[0262]

[0263] (3) DOTA activation: 48 mg DOTA was dissolved in 2 mL water, 9.0 mg 1-ethyl-(3-dimethylaminopropyl)carbodiimide (EDC) was dissolved in 0.5 mL water, the two aqueous solutions were mixed to form a reaction mixture, and the pH of the reaction mixture was adjusted to 5.0 with 0.1 M NaOH. Then 6.8 mg NHS was added, stirred in an ice bath, and reacted for 40 minutes to obtain DOTA-NHS;

[0264]

[0265] (4) Synthesis of pNIPAM-DOTA compound: 298 mg of pNIPAM (containing 0.03 mmol of amino group) was dissolved in a mixed solvent including 3 mmol of N,N-diisopropylethylamine (DIEA) and 6 mL of DMAC, heated to 45 °C, and 0.03 mmol of DOTA-NHS was added. The mixture was reacted for 3 hours and purified by centrifugal filtration. The molecular weight cutoff of the centrifugal filtration membrane was 3.5 kDa. After purification, the mixture was freeze-dried to obtain the chelated ligand product DOTA-P3-1.

[0266] .

[0267] (5) Follow the steps 1.2 of Preparation Example 1 89 Zr radiolabeled to obtain 89 Zr-DOTA-P3-1.

[0268] .

[0269] In this preparation example, the N-isopropylacrylamide polymer (the product of step (2) of Preparation Example 5) has an average molecular weight Mn of 17400 g / mol as measured by size exclusion chromatography (SEC).

[0270] After DOTA was attached, the NMR spectra in D2O solution were recorded using a Varian AC400 NMR spectrometer (Varian NMRSystems, Palo Alto, CA). 1 H-NMR, based on the hydrogen spectrum, the molar ratio of the two repeating units was calculated to be about p:q=0.85:0.15.

[0271] Preparation Example 6 Radiolabeled Polymer 89 Preparation of Zr-DFO-P3-2

[0272] (1) Add 3.37 g of N-(3-aminopropyl)methacrylamide monomer (APAM monomer), add 164 mg of azobisisobutyronitrile (AIBN) as an initiator, and synthesize by free radical reaction in acetone-DMSO mixed solvent (V (acetone): V (DMSO) = 15:1). Polymerize at 55°C under vacuum conditions for 24 hours, and filter and recover the precipitate to obtain a crude product. The crude product is dissolved in a small amount of methanol, and 15 times the volume of excess acetone is added to precipitate, and vacuum dried to obtain polyaminopropylmethacrylamide. The average molecular weight Mn is 19200-21500 g / mol measured by size exclusion chromatography (SEC), Synchropak CATSEC-300 column.

[0273] (2) p-SCN-Bn-DFO (CAS No.: 1222468-90-7) (2 mg; 200 μL DMSO) was slowly added to 12 mg polyaminopropyl methacrylamide, and sodium hydroxide was added to adjust the pH to 6.0. The mixture was reacted at room temperature for 1 h, and then the mixture was dissolved in water and dialyzed for three days. The molecular weight cutoff of the centrifugal filter membrane was 10 kDa. After freeze-drying, a total of 9.7 mg of polymer was obtained, with a yield of 69.3%. The D2O solution was recorded using a Varian AC400 NMR spectrometer (Varian NMR Systems, Palo Alto, CA). 1 H-NMR, calculated based on the hydrogen spectrum, gave a molar ratio of the two repeating units of p:q=0.82:0.18.

[0274] (3) Follow the steps 1.2 of Preparation Example 1 89 Zr radiolabeled to obtain 89 Zr-DFO-P3-2.

[0275] .

[0276] Preparation Example 7 Radiolabeled Polymer 89 Preparation of Zr-DFO-LP1-1

[0277] The p-SCN-Bn-DFO in Preparation Example 1 was replaced with compound D-1 (CAS: 1623757-39-0), and P1-Val was reacted with D-1 to prepare a DFO-coupled polymer, and then the reaction was carried out according to step 1.2 of Preparation Example 1. 89 Zr marker.

[0278]

[0279] Preparation Example 8: Preparation 89 Zr-labeled hydrogel preparation

[0280] (1) Preparation 1

[0281] Preparation of iohexol aqueous solution: Accurately measure iohexol and water for injection in a mass ratio of 2 g:3 g, mix well, and prepare 40% iohexol solution.

[0282] Prepare a solution of sodium alginate and hydroxymethyl cellulose: weigh 2 g of sodium alginate and 0.5 g of hydroxymethyl cellulose (HPMC-100M), slowly add the sodium alginate and hydroxymethyl cellulose to 100 mL of 40% iohexol solution, slowly heat the room temperature to 35°C to promote the dissolution of the sodium alginate and hydroxymethyl cellulose, and let it cool to room temperature after they are completely dissolved.

[0283] Preparation of gel composition: accurately weigh 14 g of poloxamer 407, slowly add it to 100 mL of the prepared sodium alginate and hydroxymethyl cellulose solution, and let it stand at 4°C for 3-4 hours until it is completely dissolved to obtain preparation 1. The sol-gel phase transition temperature is 26.5°C. At 37°C, it gels within 32 seconds and turns into a solid within 50 seconds; the viscosity is 0.9 Pa.S (25°C), 1.2×10 3 Pa.S (30℃).

[0284] (2) Formulation 2: Embrace HES polyethylene glycol liquid embolic agent.

[0285] (3) Preparation 3: Peptide Pushen TM Temperature-sensitive liquid embolic agent (copolymer of N-isopropylacrylamide and N-n-propylacrylamide).

[0286] (4) Preparation of zirconium labeled gel composition:

[0287] Preparation 89 Zr polymer-labeled preparation: 1 mL of the 89 The Zr-labeled polymer solution (containing 0.6 mg of polymer) was mixed with 10 mL of Preparation 1, Preparation 2, or Preparation 3, stirred gently, and then refrigerated at 4-8°C.

[0288] (5) Preparation 89 Zr non-chelating agent labeled preparations: radioisotope 89 1 mL of the Zr oxalic acid solution was added to 10 mL of Preparation 1, Preparation 2 or Preparation 3 and mixed.

[0289] Test Example 1: Performance Test

[0290] 1.1 89 Radiochemical purity of Zr-labeled hydrogel preparations

[0291] A Mini-Scan thin-layer scanner (Eckert & Ziegler Radiopharma Inc., USA) was used with fast thin-layer chromatography paper (Agilent ITLC paper strips, 1 cm × 10 cm, USA) as the carrier. After spotting, the system was developed with 20 mM citric acid solution (pH 4.9-5.1). After the chromatography paper was dried, data was collected using a thin-layer scanner. After data collection was completed, images were obtained for data analysis.

[0292] The performance test results of the gel composition prepared according to the ratio of each polymer and hydrogel preparation prepared in the preparation example 8. (1) are as follows Table 1:

[0293] Table 1

[0294]

[0295] In this study, thin layer chromatography was used to evaluate the 89 The radiochemical purity of the Zr-labeled hydrogel preparation and the labeling stability at different time points. As shown in the table above, 89 The radiochemical purity of the Zr-labeled hydrogel preparations was maintained above 93% at different time stages, indicating that 89 The in vitro stability of the Zr-labeled hydrogel preparation was good. 89 The radiochemical purity of Zr-labeled polymers used in hydrogel preparations was greater than 93% after labeling, meeting the experimental requirements, and the radiochemical purity was still greater than 93% after 19 days of storage. 89 Zr in oxalic acid solution, 89 Zr ions chelate with the hydroxyl groups of sodium alginate and promote the cross-linking of sodium alginate to form a network structure. 89 Zr coating, to achieve 89 The chelate-free Zr coating has a higher labeling efficiency at the initial labeling time of 1 day, but the radiochemical purity decreases with the extension of storage time, indicating that 89 The content of Zr-labeled sodium alginate decreases, and the 89 Zr is separated from the marker, and the stability of the experimental group numbered h is poor. The polymer provided in this experimental example has better stability and is not easy to degrade, because the chelating group is chemically chelated with the polymer by covalent bonds, and the radioactive metal element is complexed by the chelating group, which reduces the release of free radioactive elements and prolongs the radioactive labeling and imaging time.

[0296] 1.2 89 Sol-gel rheological properties of Zr-labeled gel compositions

[0297] The tests were performed on an AR-1000 constant stress rheometer (Manufacturer: TA Instruments) with built-in temperature and gap calibration.

[0298] The rheometer is equipped with a cone / plate geometry (40 mm diameter, 4° cone angle). The degassed sample is dispensed onto a temperature-controlled peltier plate, pre-equilibrated to 25°C. During the rheological test, the sample is covered with an aqueous solvent trap to prevent sample moisture loss. During the test, the temperature of the peltier plate is controlled within ±0.1°C of the preset temperature. Prior to the test, the geometric gap is calibrated. After loading an excess sample, the geometry is lowered to the predetermined gap to remove excess sample. Before starting the test, the sample is allowed to equilibrate for 30 seconds. Data is processed using TA data analysis software.

[0299] The viscosity tests of various hydrogel formulations were evaluated using flow rheology. To determine how the hydrogels behave under increasing shear stress, steady-state flow experiments were performed at 25°C. The viscosity of the samples was measured over a shear stress range of 1Pa to 100Pa. The thermal responsiveness of various hydrogel formulations was rheologically evaluated by oscillation measurements. A temperature scan of 10°C-40°C was performed on all hydrogel formulations. The sol-gel transition temperature is defined as the temperature at which gelation occurs. The gel point is defined as the temperature at which the storage modulus (G') is equal to the loss modulus (G").

[0300] Therefore, gelation is considered to have occurred when G'>G". The temperature was increased at a rate of 1°C / min, and the oscillation pressure and angular frequency remained constant.

[0301] The performance test results of the gel composition prepared by the polymer and hydrogel preparation prepared in the preparation example are shown in Table 2 below.

[0302] Table 2

[0303]

[0304] It can be seen that the addition of radiolabeled polymers has no significant effect on the viscosity and phase transition temperature of the liquid embolic agent, and does not affect the temperature-sensitive properties of the rheological properties of the hydrogel preparation itself. In particular, when the radiolabeled polymer and the temperature-sensitive material of the hydrogel preparation have the same or similar structural units, the radiolabeled polymer and the hydrogel preparation have excellent compatibility, can be more evenly and stably distributed in the hydrogel preparation, and have less effect on the viscosity and phase transition temperature of the hydrogel preparation itself.

[0305] Test Example 2 Animal in vivo test

[0306] This experiment observed the content of radioactive labeled polymers in the blood of animals, and used PET / CT tomography to collect signals within the window width range of 110keV±15%, and dynamically observed the radioactive labeled polymers prepared in Preparation Examples 1-7 of this application. 89Dynamic distribution of Zr-labeled polymers within a certain period of time after implantation into Beagle dogs according to the scheme shown in Table 3.

[0307] Beagle dogs were given general anesthesia (1.5% sodium pentobarbital, 20 mg / kg), and the femoral artery was bluntly separated. The femoral artery was punctured under direct vision, and a 5F arterial sheath was inserted. The femoral artery and arterial sheath were fixed with medical absorbable sutures. Under the guidance of a digital subtraction angiography (DSA), a 4F angiography catheter was inserted from the femoral artery to the abdominal aorta. A Y valve was connected to the end of the 4F angiography catheter. A 2.4 F microcatheter was selectively inserted into the common hepatic artery from the positive port of the Y valve. Gel was injected through the microcatheter to avoid gel reflux as much as possible. After embolization, the catheter and arterial sheath were pulled out, the femoral artery puncture site was ligated, and the muscles and skin were sutured layer by layer.

[0308] The hydrogel preparation prepared in the preparation example was used as an animal vascular embolic agent and implanted into Beagle dogs. The implantation scheme is shown in Table 3:

[0309] Table 3 Implantation plan

[0310]

[0311] 2.1 In the blood 89 Zr detection

[0312] The dosage of the embolic agent was 1 mL of the labeled gel for each animal. The specific activity of the gel was 280 μCi / mL, and the dosage for each dog was about 280 μCi / animal. Blood was collected from the animal's forelimb cephalic vein 5h, 1d, 3d, 5d, 7d, and 11d after hepatic artery embolization using a sodium heparin anticoagulation negative pressure blood collection tube. The blood volume was about 2 mL. After blood collection, the blood samples were tested and analyzed using a WIZARD γ counter. Establishment 89 The calibration curve of Zr activity meter detection value (μCi) and γ counter detection value (CPM) is used for the conversion of activity and CPM. The calibration curve is shown in Fig.21 .

[0313] The blood volume was estimated based on the dog's systemic circulating blood volume and blood leakage count results, animal dosage and body weight data. 89 Zr dose and calculate blood 89 The radioactivity of Zr accounts for the percentage of the total dose. The circulating blood volume of Beagle dogs is 85mL / kg. The total blood volume is calculated based on the animal's weight, and then the total blood exposure is calculated based on the total blood volume. The blood exposure percentage is calculated by comparing it with the total drug dose after attenuation. The percentage of blood exposure dose to total dose = body weight kg*85mL / kg*blood test value (μCi / g) / administered dose, and the blood test value is corrected to the time of administration.

[0314] Table 4 Animal blood 89Zr radioactivity

[0315]

[0316] Table 5 Animal blood 89 Zr radioactivity percentage

[0317]

[0318] As shown in Tables 4 and 5, on the first day of embolization, the blood of dogs B01-B14 89 The percentage of Zr radioactivity to the total dose was less than 7% and then gradually decreased. Seven days after administration, the radioactivity of Zr in the blood of dogs B01-B13 was 89 The radioactivity of Zr accounts for about 0.69%-2% of the total dose.

[0319] Overall, blood 89 The radioactivity of Zr accounts for a relatively low proportion, indicating that the gel formed by the test substance mainly remains in the embolization site in vivo, and also proves that the test substance has good stability after labeling and long-term placement. 89 After the hydrogel preparation of Zr-labeled polymer, no free Zr was observed in the blood. 89 Abnormal increase or decrease of Zr indicates better stability in the animal body and no obvious free state is observed.

[0320] The polymer provided in this test example is 89 Zr is chemically chelated with the polymer through the chelating group. 89 The Zr oxalate co-labeled preparations have better stability, are not easily degraded, and reduce the release of free radioactive elements.

[0321] B14 dog implanted with gel 89 Zr is directly mixed and coated without chemical chelation of thermosensitive polymer. The radioactivity in the blood within 5h-11d after implantation is significantly higher than that of other implants including 89 The radioactive signal in the blood was high within 1-5 days, indicating that radioactive elements were constantly released into the blood circulation. After 5 days, most of the embolization sites were 89 Zr has entered the blood, no more 89 Zr escapes, so the radioactivity in the blood decreases after 7 days. The radioactivity entering the blood circulation within 11 days accounts for 60% of the total dose, indicating that direct addition of ionized 89 Zr, the radioactive label has poor stability and is easily dissociated from the embolization site. The radioactive label cannot stably track the location of the hydrogel for a long time.

[0322] In implantation 89In animals of hydrogel preparations of Zr-labeled polymers, polymers of different structures as radioactive markers of hydrogel preparations all showed lower levels of blood radioactivity and showed better stability. In particular, when the hydrogel preparation and the radioactively labeled polymer have the same thermosensitive polymer units, the hydrogel preparation and the radioactively labeled polymer have more similar viscosities and phase transition temperatures, and the two can be better compatible and tightly combined, and the radioactive labeling shows excellent stability. For example, when a radioactively labeled polymer containing an alkyl ether structural unit is added to preparation 1 or 2 89 Zr-DFO-P1-1 or 89 When Zr-DFO-P1-2 was added, the total percentage of radioactive counts in the blood during the test period was significantly lower than that added to preparation 3; radiolabeled polymers containing acrylamide structural units 89 When Zr-DFO-P3-2 is added to Preparation 3 which uses acrylamide polymer as a temperature-sensitive component, it also exhibits significantly better stability than when it is added to Preparation 1.

[0323] Compared with animal B01 and animal B11, only the linker group of the radioactive polymer in the preparation is different, but the tracing effect of animal B01 is better, indicating that 89 Zr-DFO-P1-1 ratio 89 Zr-DFO-LP1-1 has a better effect, which may be due to 89 The chain length of the connecting group in Zr-DFO-LP1-1 is affected.

[0324] 2.2 In vivo dynamic imaging studies

[0325] Collection time: 5h, 1d, 5d, 7d, 11d, and 18d after embolization.

[0326] Acquisition method: nuclear medicine tomography + positioning CT. The animal is placed under the Discovery VCT instrument after general anesthesia. A whole-body CT scan is performed first to assist in the identification of tissues and organs, and then a whole-body nuclear medicine signal scan is performed from the head to the bottom of the lower limbs.

[0327] PET-CT imaging data processing: The coronal and cross-sectional images of the PET-CT imaging results of dogs B01-B14 were saved, and the region of interest (ROI) was outlined (outlined the main organs with nuclear medicine signals) based on the imaging images at the two positions and the nuclear medicine images, and the corresponding area was circled, and the percentage of the count of the outlined area to the whole body count was recorded.

[0328] The results are as follows Figure 1-Figure 20 and as shown in Table 6.

[0329] Figure 1 , Figure 3 , Figure 5 , Figure 7 , Fig. 9 These are the cross-sectional images of dog B01 at 0.5h, 1d, 5d, 7d, and 11d after implantation of the radiolabeled hydrogel preparation; Fig.11 , Fig.13 , Fig.15 , Fig.17 , Fig.19 These are cross-sectional images of dog B04 at 0.5h, 1d, 5d, 7d, and 11d after implantation of the radiolabeled hydrogel preparation. In the cross-sectional images, the upper left corner of each image is the CT localization map, the upper right corner is the nuclear medicine imaging image of the localization position, the lower left corner is the cross-sectional nuclear medicine image enrichment area image, and the lower right corner is the whole body nuclear medicine signal image (this image does not change with the change of the CT localization map). All regions are located in the same position - the liver.

[0330] Figure 2 , Figure 4 , Figure 6 , Figure 8 , Fig.10 These are the coronal images of dog B01 at 0.5h, 1d, 5d, 7d, and 11d after implantation of the radiolabeled hydrogel preparation; Fig.12 , Fig.14 , Fig.16 , Fig.18 , Fig. 20 These are transverse images of dog B04 at 0.5h, 1d, 5d, 7d, and 11d after implantation of the radiolabeled hydrogel preparation. Coronal images, the left corner of the image is the CT localization map, the second left is the nuclear medicine imaging map of the localization position, the third left is the coronal nuclear medicine image enrichment area image, and the fourth left is the whole body nuclear medicine image. All regions are located in the same position - the liver.

[0331] As can be seen from the images, the radioactive signal of the animal implanted with supplementation scheme A (B01) was mainly distributed in the liver at the beginning of 5 hours after implantation, with a small amount distributed in the intestine and bladder. During the period of 1-11 days after implantation, the radioactive signal was still mainly distributed in the liver, with only a very small amount distributed in other organs. Compared with B01, the radioactive signal of the animal implanted with supplementation scheme D (B04) was mainly distributed in the liver, but the signal distribution in the lungs, intestines, kidneys and brain was significantly more than that of B01.

[0332] Table 6 below records the average percentage of the total body counts of five enriched areas with higher counts, namely, liver, lung, intestine, kidney, and brain of five dogs.

[0333] Table 6 Percentage of organ counts in the total body counts of each animal (%)

[0334]

[0335] As shown in Table 6, the proportion of each animal drug in the liver on the day of administration was more than 80%, indicating that the test substance was embolized in the liver on the day of administration, and most of the radioactive signals were at the embolization site. The test substance had a high labeling efficiency and almost no degradation. During the 1-5 days of administration, the proportion of liver in the liver decreased. This may be because some of the test substances were embolized near the liver portal. The radioactive labeled polymers that failed to fully bind to the hydrogel during the administration stage were washed away by the blood flow and migrated to other tissues such as the intestine, lung and kidney for enrichment during this time. Therefore, a slight distribution can be seen in the intestine, lung and kidney on days 5-7, and the proportion of radioactive counts showed a slightly increasing trend; however, the proportion of liver counts still accounted for more than 80%, which can still significantly mark the location of administration embolism and does not affect the evaluation of the embolism state. From the 11th to the 18th day after intervention, and from the 5th to the 18th day after administration, the proportion of the test substance in the liver gradually increased to 100%, and the signals of the intestinal wall and other tissues and organs were significantly reduced. By the 18th day, the radioactive elements in other organs had been metabolized out of the body and decayed to no signal. Only the liver showed a signal, indicating that the test substance remained in the embolization site in the body, and 89 The Zr-labeled polymers showed good long-term stability in vivo.

[0336] Throughout the experimental period, 89 Zr-labeled polymers are mainly enriched in the liver and the average proportion is between 80% and 100%, indicating that most of the radioactive nuclides are tightly bound to the implant and have not detached. They can be significantly distinguished from the free weak signals and are excellent tracers for implantable hydrogels.

[0337] Specifically, the radioactive signal in the liver of animal B01 implanted with supplementation scheme A was significantly higher starting from 5 hours, and the liver showed a high radioactive intensity throughout the monitoring period of 1-18 days, all of which were above 80% or 84%, while the radioactive signals in other organs such as lungs, intestines, kidneys, and brain were very low, indicating that in animal B01, the radioactive labeled polymer was tightly bound to preparation 1 and embolized in the liver, and the embolization preparation had good stability. At the same time, the radioactive labeled polymer also maintained good stability and rarely detached from the embolization site. Animal B02 implanted with supplementation scheme B and animal B05 implanted with supplementation scheme E were also able to maintain more than 80% of the radioactive count in the liver throughout the monitoring period. In comparison, animal B04 implanted with supplementation scheme D had a relatively low radioactive signal concentration in the liver, which was 80.9% from the initial stage after implantation, indicating that after the addition scheme D was implanted in the body, a small amount of radioactive signals were freed into the blood and circulated to other organs, but at the 18th day, the radioactivity of other organs other than the liver of animal B04 was also metabolized out of the body. It can be seen that the polymer based on polymer P1 and the linking group 89Zr-DFO-P1-1 reacts directly with chitosan amino groups to form polymers 89 The Zr-DFO-P2 structure has stronger stability and a more stable radioactive tracer effect in hydrogel preparations.

[0338] Although the embodiments of the present application have been shown and described, those skilled in the art will appreciate that various changes, modifications, substitutions and variations may be made to the embodiments without departing from the principles and spirit of the present application, and that the scope of the present application is defined by the claims and their equivalents.

Claims

1. A radionuclide labeled polymer having the following structure: G-L1-L2-P; in, G is a group chelated with a radionuclide, which includes a chelating group and a radionuclide, wherein the chelating group chelates with the radionuclide; L1 and L2 are independently a chemical bond or a linking group; P is a temperature-sensitive structural unit.

2. The radionuclide-labeled polymer according to claim 1, characterized in that: The L1 is a chemical bond or a connecting group represented by formula (I-1): （I-1）, In formula (I-1), x and y are independently selected from integers of 0 to 3; R1 is , C 1-5 The alkylene or chemical bonds; R e for , substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 heteroarylene; R d C 1-5 The alkylene or chemical bonds; R b is hydrogen, hydroxyl or C 1-5 alkyl; X is O or S; e represents the connection site between G and L1, i represents the connection site between R1 and R e , f represents the connection site of L1 and L2; and / or, The L2 is a chemical bond, such as a group represented by formula (II-1) or (II-2): (II-1), (II-2), Wherein, in formula (II-1), z is 0, 1, 2 or 3; In formula (II-2), R2 and R3 are independently hydrogen, C 1-5 Alkyl, substituted or unsubstituted C 6-10 Aryl, or substituted or unsubstituted C 5-10 Heteroaryl, amide or thiol; f represents the connection site between L1 and L2, and h represents the connection site between L2 and P; u is 0, 1, 2, or 3; Each R 1-1 , R e-1 and R d-1 Independent for C 1-5 of alkylene.

3. The radionuclide-labeled polymer according to claim 2, characterized in that It satisfies one or more of the following conditions (1)-(5): (1) Each C 1-5 The alkylene group is methylene, , , or ; (2) Each C 6-10 Arylene is phenylene or naphthylene; (3) Each C 5-10 The heteroaryl group is preferably C 5-10 Heteroaryl, wherein the heteroatom is independently selected from one or two of N, O and S, and the number of heteroatoms is independently 1, 2 or 3, preferably the heteroatom is N or O, and the number of heteroatoms is 1 or 2; (4) Each C 1-5 Alkyl is methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl or sec-butyl; (5) Each substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 Heteroarylene, or substituted or unsubstituted C 6-10 Aryl, or substituted or unsubstituted C 5-10 The substituents in the heteroaryl group are independently C 1-3 alkyl, halogen, hydroxyl or cyano; The C 1-3 The alkyl group is methyl, ethyl or propyl; The halogen is F, Cl, Br or I.

4. The radionuclide-labeled polymer according to claim 2, characterized in that It satisfies one or more of the following conditions (1)-(4): (1) L1 is a linking group represented by formula (I-2) or (I-3): （I-2）， （I-3） Among them, in formula (I-2), R1 is or C 1-5 The alkylene group, i represents the e Connect; R e is substituted or unsubstituted C 6-10 Arylene, or substituted or unsubstituted C 5-10 Heteroarylene; wherein e represents connection with G, and f represents connection with L2; In formula (I-3), each n is independently selected from an integer in the range of 0-10, and each R b are independently hydrogen or hydroxy; (2) In formula (II-1), z is 2; (3) In formula (II-2), R2 and R3 are both methyl groups; (4) In formula (II-2), u is 0.

5. The radionuclide-labeled polymer according to any one of claims 1 to 4, characterized in that: The P is one or more of a chitosan structural unit, an acrylamide structural unit or a group as shown in formula (III-1): ， (III-1), Wherein, in formula (III-1), a and c are independently selected from the range of 2-130; b is in the range of 10-80, and R is selected from hydrogen and C 1-5 alkyl.

6. The radionuclide-labeled polymer according to claim 5, characterized in that Satisfy at least one of the following conditions (1)-(2): (1) In formula (III-1), a and c are independently integers in the range of 80-130, and b is an integer in the range of 30-60; (2) The acrylamide structural unit includes a group represented by formula (III-2): (III-2) In formula (III-2), R c is hydrogen, -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2, wherein m is an integer between 0 and 5, R a is -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2 is a divalent group formed by losing one hydrogen; p or q is independently an integer in the range of 20-200.

7. The radionuclide-labeled polymer according to any one of claims 1 to 4, characterized in that: The radionuclide is 89 Zr; The chelating group is a chelating group formed by a bifunctional chelating agent.

8. The radionuclide-labeled polymer according to claim 7, characterized in that The chelating agent is DOTA, NOTA, NODA, NODAGA, DOTP, TETA, ATSM, PTSM, EDTA, EC, HBEDCC, DTPA, SBAD, BAPEN, DFO or TACN.

9. The radionuclide-labeled polymer according to claim 1, characterized in that It meets one or more of the following conditions: (1) The chelating group is , or ; (2) L1 is , or , e indicates connection with G, and f indicates connection with L2; (3) L2 is or , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P; (4) P is , , or .

10. The radionuclide-labeled polymer according to claim 1, characterized in that: The polymer satisfies any one of the following options (1) to (5): Solution (1): The chelating group is ; The P is ; Wherein, R is a methyl group, a is an integer in the range of 75-110, and b is an integer in the range of 25-70; The L1 is , where e indicates connection with G, and f indicates connection with L2; The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P; Solution (2): The chelating group is ; The P is , wherein R is methyl, a is an integer in the range of 75-110, and b is an integer in the range of 25-70; The L1 is , where e indicates connection with G, and f indicates connection with L2; The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P; Solution (3): The chelating group is ; The P is , wherein R is methyl, a is an integer in the range of 75-110, and b is an integer in the range of 25-70; The L1 is , where e indicates connection with G, and f indicates connection with L2; The L2 is , f represents the connection site between L1 and L2, and h represents the connection site between L2 and P; Solution (4): The chelating group is ; The P is or ; The L1 is , where e indicates connection with G, and f indicates connection with L2; The L2 is a chemical bond; Solution (5): The chelating group is ; The P is or ; The L1 is a chemical bond; The L2 is a chemical bond.

11. The radionuclide-labeled polymer according to claim 1, characterized in that: It is any of the following structures: 、 、 、 or ； in, a and c are independently selected from the range of 2-130; b is in the range of 10-80, R is selected from hydrogen and C 1-6 alkyl; R c is hydrogen, -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2, wherein m is an integer between 0 and 5, R a is -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2 is a divalent group formed by losing one hydrogen; p or q is independently an integer in the range of 20-200.

12. The radionuclide-labeled polymer according to claim 11, characterized in that: It is any of the following structures: 、 、 、 、 、 、 、 , or .

13. A polymer represented by formula (A): ; in, The definitions of L1, L2, and P are the same as those of L1, L2, and P in any one of claims 1 to 11; Q is a chelating group, and the chelating group is the same as the definition of the chelating group in any one of claims 1 to 11.

14. The polymer according to claim 13, characterized in that The polymer shown in formula (A) is any of the following structures: , , , or , a and c are independently selected from the range of 2-130; b is in the range of 10-80, R is selected from hydrogen and C 1-6 alkyl; R c is hydrogen, -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2, wherein m is an integer between 0 and 5, R a is -CH-(CH3)2, -C-(CH3)3, or -(CH2) m -NH2 is a divalent group formed by losing a hydrogen. p or q is independently an integer in the range of 20-200.

15. The polymer according to claim 14, characterized in that The polymer represented by formula (A) is any of the following structures: , , , , , , , , or .

16. A method for preparing a radionuclide-labeled polymer, comprising the following steps: mixing and incubating a polymer represented by formula (A) as claimed in any one of claims 13 to 15 with a solution containing a radionuclide to obtain a radionuclide-labeled polymer.

17. The preparation method according to claim 16, characterized in that: The radionuclide-labeled polymer is as described in any one of claims 1 to 12.

18. A gel composition comprising the radionuclide-labeled polymer according to any one of claims 1 to 12 and a hydrogel preparation.

19. The gel composition according to claim 18, characterized in that: It satisfies one or more of the following conditions (1)-(4): (1) The hydrogel preparation comprises a temperature-sensitive gelling agent and a gelling agent; (2) The hydrogel preparation further comprises a developer; (3) The mass volume ratio of the radionuclide labeled polymer to the hydrogel preparation is 0.01 mg / mL to 2 mg / mL; (4) The sol-gel transition temperature of the gel composition is 25°C-37°C.

20. The gel composition according to claim 19, characterized in that: It satisfies one or more of the following conditions (1)-(3): (1) The temperature-sensitive gelling agent is one or more of poloxamer, acrylamide polymer, chitosan or polyethylene glycol block copolymer; (2) The gelling agent is one or more of sodium alginate, hydroxymethyl cellulose and CaCl2; (3) The developer is iohexol.

21. The gel composition according to claim 20, characterized in that: Based on the volume of the hydrogel preparation, the hydrogel preparation comprises: 0.2-1g / mL iohexol, 0.02-0.1g / mL sodium alginate, 0.12-0.2g / mL poloxamer, 0.005-0.1 g / mL hydroxymethylcellulose or CaCl2, and water; and / or In the gel composition, the mass volume ratio of the radionuclide-labeled polymer to the hydrogel preparation is 0.01 mg / mL to 0.5 mg / mL.

22. The gel composition according to claim 21, characterized in that Based on the volume of the hydrogel preparation, the gel composition consists of 0.06 mg / mL of the radionuclide-labeled polymer, 0.4 g / mL of iohexol, 0.02 g / mL of sodium alginate, 0.14 g / mL of poloxamer, 0.005 g / mL of hydroxymethylcellulose and the balance of water.

23. A tracer comprising a polymer labeled with a radionuclide as claimed in any one of claims 1 to 12.

24. Use of a polymer labeled with a radionuclide according to any one of claims 1 to 12 in the preparation of a tracer, wherein the tracer is used to trace the gel composition according to any one of claims 18 to 22.

25. The use according to claim 24, characterized in that The tracer is a PET / CT imaging agent or a SPECT imaging agent.

26. A use of the gel composition according to any one of claims 18 to 22, wherein the use is use of the gel composition in preparing materials for in vivo vascular embolism, in vivo hemostasis, drug delivery or in vivo tissue repair.

27. A method for radiolabeling a hydrogel formulation, comprising: Providing a radionuclide-labeled polymer as claimed in any one of claims 1 to 12; The radionuclide-labeled polymer is added to the hydrogel formulation as described in the gel composition of any one of claims 18 to 22.

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