Conjugates with radiation-responsive groups and uses thereof

Abstract

The present disclosure provides a conjugate having a radiation responsive group and its use, wherein the conjugate has a general formula of P-L-D, wherein P is a radiation responsive group containing an aromatic group or a heteroaromatic group; L does not exist or is a divalent linking group; and D is an imaging agent or a therapeutic agent. Under irradiation, the radiation responsive group P in the conjugate undergoes a chemical reaction, so that the radiation responsive group P or the radiation responsive group P and the linking group L (if present) fall off from the conjugate together, releasing the active molecule D.

Description

Technical Field

[0001] The present disclosure relates to the fields of medical treatment and imaging, and in particular to a conjugate containing a radiation responsive group and a therapeutic agent or an imaging agent and uses thereof. Background Art

[0002] Activating prodrugs by physiological stimulation or external stimulation has a very promising prospect in disease treatment and has attracted widespread attention. Because the prodrug itself is low-toxic or non-toxic, but through certain stimulations, such as light, heat, pH changes and enzymatic reactions, the chemical structure of the prodrug can be changed to restore it to an active structure. However, these stimulation methods have their limitations, especially prodrugs with photosensitivity. Photosensitive prodrugs restore drug activity through photochemical removal reactions under ultraviolet visible light, but the penetration ability of ultraviolet visible light is limited (several millimeters), and it is not possible to penetrate tissues well, so it is not possible to effectively activate prodrug molecules deep in the tissue to achieve the purpose of treatment. Therefore, it is still necessary to develop a prodrug that can be activated in vivo, such as deep in the tissue, by a non-enzymatic method.

[0003] On the other hand, in special environments with hydroxyl radicals, it may be necessary to develop a probe that can sensitively respond to hydroxyl radicals. Hydroxyl radicals can be from mammals (including humans). Alternatively, hydroxyl radicals can be produced by irradiating aqueous solutions or by chemical reactions such as the decomposition of hydrogen peroxide. In some cases, in order to improve the imaging performance of body tissues and organs in an environment where hydroxyl radicals are produced by irradiation, it is necessary to develop a probe with radiation response performance. In other cases, in order to detect hydroxyl radicals in solutions, such as the concentration of hydroxyl radicals in vivo or during oxidation treatment of wastewater by Fenton's reagent, it may also be necessary to develop a probe that can sensitively respond to free radicals. Summary of the invention

[0004] One aspect of the present disclosure provides a conjugate of general formula (I),

[0005] P——L——D (I)

[0006] Wherein P is a radiation-responsive group containing an aromatic group or a heteroaromatic group; L does not exist or is a divalent linking group; and D is an imaging agent or a therapeutic agent. Under irradiation, the radiation-responsive group P in the conjugate undergoes a chemical reaction, so that the radiation-responsive group P or the radiation-responsive group P and the linking group L (if present) fall off from the conjugate together, releasing the active molecule D.

[0007] In one embodiment of the present disclosure, the radiation responsive group is a group that can chemically react with the hydroxyl radicals generated by radiation and thus fall off from the conjugate, or the radiation responsive group is a group that can chemically react with the hydroxyl radicals generated by radiation and thus fall off from the conjugate together with the linker L.

[0008] In the conjugate provided by at least one embodiment of the present disclosure, the aryl or heteroaryl group is, for example, selected from the following group: phenyl, naphthyl, anthracenyl, phenanthrenyl, biphenyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, tetrazinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, Azolyl, iso Azolyl, Oxazolyl, triazolyl, furazanyl, thiazolyl, isothiazolyl, indolyl, benzothiazolyl, benzodioxolyl, benzo oxazolyl, benzothienyl, quinolyl, tetrahydroisoquinolyl, isoquinolyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, chromonyl, coumarinyl, benzopyranyl, cinnolinyl, quinoxalinyl, indazolyl, pyrrolopyridinyl, furopyridinyl, dihydroisoindolyl, tetrahydroquinolyl, carbazolyl, dibenzofuranyl, dibenzothienyl, benzindolyl, phenanthrolinyl, acridinyl, phenanthridinyl and xanthenyl; wherein the aryl or heteroaryl is optionally substituted with one or more substituents selected from the group consisting of: halogen wherein R1 and R2 are independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C6 alkyl, C4-C6 alkyl, C5-C6 alkyl, C6-C6 alkyl, C7-C6 alkyl, C8-C6 alkyl, C9-C6 alkyl, C10-C6 alkyl, C11-C6 alkyl, C12-C6 alkyl, C13-C6 alkyl, C14-C6 alkyl, C15-C6 alkyl, C16-C6 alkyl, C17-C6 alkyl, C18-C6 alkyl, C19-C6 alkyl, C21-C6 alkyl, C22-C6 alkyl, C23-C6 alkyl, C27-C6 alkyl, C28-C6 alkyl, C29-C6 alkyl, C3-C6 alkyl, C4-C6 alkyl, C5-C6 alkyl, C6-C6 alkyl, C7-C6 alkyl, C8-C6 alkyl, C9-C6 alkyl, C1 10 Cycloalkyl, C6-C 20 The alkyl, alkenyl, cycloalkyl, aryl and heteroaryl groups are optionally substituted with one or more halogen, hydroxyl, mercapto, -CN, nitro, -N3, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C 10 Cycloalkyl, C6-C 20 The moiety is substituted with aryl or heteroaryl having 5 to 20 ring atoms.

[0009] For example, in the conjugate provided in at least one embodiment of the present disclosure, the aryl or heteroaryl group is selected from phenyl, pyrrolyl, furyl, thienyl, pyrazolyl and imidazolyl, and the aryl or heteroaryl group is optionally substituted with one or more amino, amine, -NHCOCH3, phenyl, methoxy, ethoxy, methyl, ethyl, hydroxyl, alkenyl, F, Cl, Br, I, -N3 or methyl.

[0010] In a preferred embodiment of the present disclosure, the aryl or heteroaryl group has no oxo, -C(=O)-, -C(=S)-, -CN, -SO3H, or -CF3 directly connected thereto.

[0011] In a preferred embodiment of the present disclosure, the aryl group is an optionally substituted resorcinol group.

[0012] In the conjugate provided in at least one embodiment of the present disclosure, L is, for example, selected from -SS-, linear C1-C6-alkylene-, branched C3-C6-alkylene-, -R3-XC(=O)-Y-, -R3-XC(=S)-Y-, -R3-X-, -XC(=O)-Y-, -XC(=S)-Y- or -CH2-XC(=Z)-Y-CH2CH2-EC(=J)-F-, wherein R3 is selected from C1-C3 alkylene or -CH2-C2 alkenyl-; X, Y, E and F are independently selected from O, NH or S; Z and J are independently selected from O or S; L may be optionally substituted with hydroxyl, thiol, NH2, F, Cl, Br, I, -N3, methyl.

[0013] In a preferred embodiment of the present disclosure, L is -CH2-X-(C=O)-Y-, -CH2-X-(C=S)-Y-, -CH2-X- or -X-(C=O)-Y-, wherein X and Y are independently selected from O, NH, S. In a more preferred embodiment of the present disclosure, L is -CH2-O-(C=O)-, -CH2-O-(C=O)-NH- or -O-(C=O)-NH-.

[0014] In the conjugate provided in at least one embodiment of the present disclosure, D is a fluorescent imaging agent or a therapeutic agent for treating cancer.

[0015] Another aspect of the present disclosure also provides use of the above conjugate in a method of treatment or imaging.

[0016] In one embodiment of the present disclosure, the conjugate is used for qualitative or quantitative detection of hydroxyl radicals. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] In order to more clearly illustrate the technical solutions of the embodiments of the present disclosure, the drawings of the embodiments will be briefly introduced below. Obviously, the drawings in the following description only relate to some embodiments of the present disclosure, but are not intended to limit the present invention.

[0018] Figure 1a The release of Fmoc-lys by 10 Gy of γ-irradiation of compound 13 in different solutions is shown;

[0019] Figure 1b The UV absorption peak of compound 14 after 1000 Gy of γ-irradiation and incubation at 37°C is shown;

[0020] Figure 2a The fluorescence emission spectrum of the conjugate Fqf-OH under irradiation dose gradient is shown;

[0021] Figure 2b The relationship between the fluorescence intensity of the conjugate Fqf-OH and the irradiation dose is shown;

[0022] Figure 2c The fluorescence response of the conjugate Fqf-OH to different ROS / RNS is shown;

[0023] Figure 3 Confocal imaging studies of the release of fluorescent molecules from the conjugate Fqf-OH in vitro after γ-irradiation are shown;

[0024] Figure 4 shows the in vivo fluorescent molecular release of the conjugate Fqf-OH; and

[0025] Figure 5 Compound 27 is shown as a prodrug molecule that releases a drug molecule after gamma irradiation. DETAILED DESCRIPTION

[0026] In order to make the purpose, technical solution and advantages of the embodiments of the present disclosure clearer, the technical solution of the embodiments of the present disclosure will be clearly and completely described below in conjunction with the drawings of the embodiments of the present disclosure. Obviously, the described embodiments are part of the embodiments of the present disclosure, not all of the embodiments. Based on the described embodiments of the present disclosure, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0027] The present invention may be implemented in other specific forms without departing from the basic attributes of the present invention. It should be understood that, without conflict, any and all embodiments of the present invention may be combined with the technical features of any other embodiment or multiple other embodiments to obtain other embodiments. The present invention includes such combinations to obtain other embodiments.

[0028] Unless otherwise defined, technical or scientific terms used in the present disclosure should have the common meanings understood by one of ordinary skill in the art to which the present invention belongs.

[0029] The description of the present disclosure should be interpreted as being consistent with the laws and principles of chemical bonding. In some cases, a hydrogen atom may be removed in order to accommodate a substituent at a given position. For example, the active molecule D may remove a hydrogen atom in the conjugate and be directly connected to the radiation responsive group or connected to the radiation responsive group through a linker L.

[0030] The words “including”, “comprising” or “comprising” and the like used in the present disclosure mean that the elements preceding the words include the elements listed after the words and their equivalents, but not excluding other elements.

[0031] The “imaging” used in the present disclosure is not limited to imaging, but also includes detection based on luminescence intensity, such as qualitative or quantitative detection of hydroxyl radicals based on the intensity of near-infrared excited fluorescence.

[0032] The radiation source of the present disclosure may be α, β, or γ rays generated by the decay of radionuclides. X-rays, γ rays, high-energy electrons, protons, heavy ions generated by external radiation sources, and α particles generated by boron neutron capture therapy (BNCT) and other possible external or internal radiation may also be applicable to the present disclosure.

[0033] High-energy radiation (such as X-rays and gamma rays) can be used as external stimuli to cause chemical reactions in prodrug molecules or imaging agent precursors to release drug molecules or imaging agents. Due to the high penetrating power of the radiation and the high spatiotemporal resolution of the radiation, prodrug molecules or imaging agent precursors can be very effectively activated by radiotherapy equipment. For example, X-ray irradiation is used as an external trigger for activating prodrugs. Since the chemical reactions triggered by radiation can be controlled in space and time, the area, time and dose of such prodrugs converted into their active forms can be precisely controlled.

[0034] However, the biggest problem with using high-energy radiation as an activation method is that the chemical reactions triggered by high-energy radiation are usually lacking in selectivity. When removing the protecting groups in the prodrug molecules or imaging agent precursors, the structures of the drug molecules or imaging agents may be destroyed at the same time. Therefore, the problem that needs to be solved urgently is to find a group with high reactivity, high reaction rate and high selectivity to radiation as the protecting group of the drug molecules or imaging agents.

[0035] The term "conjugate" used herein refers to a compound obtained by coupling an active molecule such as a drug or imaging agent with a radiation-responsive group through a linker or without a linker, wherein after irradiation with, for example, X-rays or gamma rays, the radiation-responsive group and the linker (if present) are separated from the active molecule to release the active molecule. Please note that although the conjugate contains a radiation-responsive group, it does not mean that the conjugate must be used in a radiation environment. For example, in the oxidation treatment of wastewater by the Fenton reagent method, the free radicals generated by hydrogen peroxide may also separate the radiation-responsive group and the linker (if present) from the active molecule to release the active molecule.

[0036] The term "C1-C6 alkyl" as used herein refers to a straight or branched alkane chain containing 1-6 carbon atoms. Representative examples of C1-C6 alkyl include, but are not limited to, methyl (C1), ethyl (C2), n-propyl (C3), isopropyl (C3), n-butyl (C4), tert-butyl (C4), sec-butyl (C4), isobutyl (C4), n-pentyl (C5), 3-pentyl (C5), neopentyl (C5), 3-methyl-2-butane (C5), tert-pentyl (C5) and n-hexyl (C6), etc. The term "lower alkyl" refers to a straight or branched alkyl group having 1 to 4 carbon atoms. "Substituted alkyl" refers to an alkyl group substituted with one or more substituents, preferably 1 to 4 substituents, at any available point of attachment. The term "haloalkyl" refers to an alkyl group having one or more halogen substituents, including, but not limited to, groups such as -CH2Br, -CH2I, -CH2Cl, -CH2F, -CHF2 and -CF3.

[0037] As used herein, the term "alkylene" refers to a divalent hydrocarbon group as described above for "alkyl" but with two points of attachment. For example, a methylene group is a -CH2- group and an ethylene group is a -CH2-CH2- group.

[0038] As used herein, the terms "alkoxy" and "alkylthio" refer to an alkyl group as described above attached via an oxygen bond (-O-) or a sulfur bond (-S-), respectively. The terms "substituted alkoxy" and "substituted alkylthio" refer to a substituted alkyl group attached via an oxygen bond or a sulfur bond, respectively. "Lower alkoxy" is a group OR in which R is a lower alkyl group (an alkyl group containing 1 to 4 carbon atoms).

[0039] As used herein, the term "halogen" refers to fluorine, chlorine, iodine or bromine.

[0040] As used herein, the term "optionally substituted" means either unsubstituted or having at least one non-hydrogen substituent that does not destroy the activity possessed by the unsubstituted analog.

[0041] Radiation responsive group P

[0042] The term "radiation responsive group" as used herein refers to a group that can undergo a chemical change and thus fall off from a molecule after irradiation, such as X-rays or gamma rays. Please note that although the conjugate contains a radiation responsive group, it does not mean that the conjugate must be used in a radiation environment. For example, in the oxidation treatment of wastewater by the Fenton reagent method, the free radicals generated by hydrogen peroxide may also separate the radiation responsive group and the linker (if present) from the active molecule to release the active molecule.

[0043] The radiation responsive group P of the present disclosure contains an aryl group or a heteroaryl group. In one embodiment, the radiation responsive group P is an optionally substituted aryl group or a heteroaryl group, that is, the aryl group or the heteroaryl group may be unsubstituted or substituted.

[0044] In one embodiment, the hydroxyl radicals generated by irradiation cause the radiation responsive group P to undergo a chemical reaction, so that the radiation responsive group P or the radiation responsive group P and the linker L (if present) are detached from the conjugate together, releasing the active molecule D.

[0045] The term "aryl" refers to a hydrocarbon molecule having 6 to 20 carbon atoms (C6-C 20 ) is a monovalent aromatic hydrocarbon group. Typical aryl groups include, but are not limited to, phenyl, naphthyl, anthracenyl, phenanthrenyl, biphenyl, and the like.

[0046] The term "heteroaryl" refers to a monovalent aromatic group as a 5- or 6-membered ring and a fused ring system comprising 5 to 20 atoms (wherein at least one ring is aromatic), which contains one or more heteroatoms independently selected from nitrogen, oxygen and sulfur, if the ring contains multiple oxygen atoms, these oxygen atoms are not directly adjacent.

[0047] Exemplary monocyclic heteroaryl groups include pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, tetrazinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, Azolyl, iso Azolyl, Oxazolyl, triazolyl, furazanyl, thiazolyl, isothiazolyl and the like.

[0048] Exemplary bicyclic heteroaryl groups include indolyl, benzothiazolyl, benzodioxolyl, benzo oxazolyl, benzothiophenyl, quinolyl, tetrahydroisoquinolyl, isoquinolyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, chromonyl, coumarinyl, benzopyranyl, cinnolinyl, quinoxalinyl, indazolyl, pyrrolopyridinyl, furopyridinyl, dihydroisoindolyl, tetrahydroquinolyl and the like.

[0049] Exemplary tricyclic heteroaryl groups include carbazolyl, dibenzofuranyl, dibenzothiophenyl, benzindolyl, phenanthrolinyl, acridinyl, phenanthridinyl, xanthene, and the like.

[0050] In one embodiment, the aryl or heteroaryl group has no substituents other than the linker L to which it is attached.

[0051] In another embodiment, the aryl or heteroaryl is substituted with one or more substituents, including but not limited to halogen, -R1, -NR1R2, -CN, -NO2, -N3, -OR1, -SR1, -NHCOR1, -O-COR1, -CH=CR1R2, -C(=O)-R1, -C(=O)-OR1, -C(=O)-Cl, -C(=O)-NH2, -C(=O)-NH-R1 and -C(=O)-NR1R2, wherein R1 and R2 are independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C 10 Cycloalkyl, C6-C 20 The alkyl, alkenyl, cycloalkyl, aryl and heteroaryl groups described for the substituents are optionally substituted with one or more halogen, hydroxyl, mercapto, -NH2, -CN, -NO2, -N3, -NHCOH, -OC(=O)H, -C(=O)H, -C(=O)-OH, -C(=O)-Cl, -C(=O)-NH2, -C(=O)-NH-CH3, -C(=O)-CH3, -C(=O)-OCH3, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C 10 Cycloalkyl, C6-C 20 The moiety is substituted with aryl or heteroaryl having 5 to 20 ring atoms.

[0052] In a preferred embodiment, the aryl or heteroaryl group is substituted with one or more substituents, including hydroxyl, alkoxy (e.g., methoxy, ethoxy, propoxy, butoxy), amino, methylamino, dimethylamino. It is believed that these substituents have electron donating properties and can increase the electron cloud density of the aryl or heteroaryl group, which is conducive to the radiation-responsive group to undergo a chemical reaction under irradiation and thus fall off from the conjugate.

[0053] In a preferred embodiment, the aryl or heteroaryl group is substituted by one or more substituents, and the substituents include halogen, such as fluorine, chlorine, bromine, iodine, and more preferably include iodine.

[0054] In a preferred embodiment, the aryl or heteroaryl group is substituted with one or more substituents, and the substituents include -N3.

[0055] In a preferred embodiment, the aryl or heteroaryl group has no oxo (i.e., =O), -C(=O)-, -C(=S)-, -CN, -SO3H, -CF3 directly connected thereto. It is believed that these groups have strong electron-withdrawing properties, which will reduce the electron cloud density of the aryl or heteroaryl group, and are not conducive to the chemical reaction of the radiation-responsive group under irradiation.

[0056] For example, the radiation responsive group may be selected from:

[0057]

[0058] Where X is the connection site. From the position of X, it can be seen that the radiation responsive group P can be directly connected to L or D through an aryl or heteroaryl group, or can be connected to L or D through a divalent group on the aryl or heteroaryl group, such as a methylene group.

[0059] Divalent linking group L

[0060] In one embodiment, there is no divalent linking group L in the conjugate of the present disclosure, that is, the conjugate of general formula (I) is actually a structure of P-D. In this case, the radiation-responsive group P falls off from the conjugate after irradiation, releasing the active molecule D. For example, the hydroxyl radicals generated by irradiating the aqueous solution cause the radiation-responsive group P to undergo a chemical reaction, so that the radiation-responsive group P falls off from the conjugate, releasing the active molecule D.

[0061] In one embodiment, a divalent linker L is present in the conjugate of the present disclosure, and the divalent linker L couples the radiation-responsive group P and the active molecule D together, and after irradiation, the divalent linker L and the radiation-responsive group P fall off from the conjugate together, releasing the active molecule D. For example, the hydroxyl radicals generated by irradiating the aqueous solution cause the radiation-responsive group P to undergo a chemical reaction, so that the radiation-responsive group P and the divalent linker L to which it is connected fall off from the conjugate together, releasing the active molecule D.

[0062] In one embodiment, the linking group L is selected from -SS-, linear C1-C6-alkylene-, branched C3-C6-alkylene-, -R3-XC(=O)-Y-, -R3-XC(=S)-Y-, -R3-X-, -XC(=O)-Y-, -XC(=S)-Y- or -CH2-XC(=Z)-Y-CH2CH2-EC(=J)-F-, wherein R3 is selected from C1-C3 alkylene or -CH2-C2 alkenyl-; X, Y, E and F are independently selected from O, NH or S; Z and J are independently selected from O or S; the linking group L may be optionally substituted with hydroxyl, thiol, NH2, F, Cl, Br, I, -N3, methyl.

[0063] In a preferred embodiment, L is -CH2-X-(C=O)-Y-, -CH2-X-(C=S)-Y-, -CH2-X- or -X-(C=O)-Y-, wherein X and Y are independently selected from O, NH, S. In a more preferred embodiment, L is -CH2-O-(C=O)-, -CH2-O-(C=O)-NH- or -O-(C=O)-NH-.

[0064] Active molecule D

[0065] The term "active molecule" as used herein refers to a molecule with activity released from the conjugate, which is derived from D in the conjugate PLD. In one embodiment, the active molecule is a therapeutic agent or an imaging agent. In a preferred embodiment, the therapeutic agent or imaging agent does not have an aromatic ring or heteroaromatic ring with a rich electron cloud. For example, although the therapeutic agent or imaging agent has an aromatic ring or a heteroaromatic ring, it has a strong electron-withdrawing group such as a quaternary ammonium ion, -CN, -COOH, -CHO, -SO3H, -CF3, etc. on the aromatic ring or the heteroaromatic ring, so that the therapeutic agent or imaging agent is relatively stable under irradiation.

[0066] It should be noted that in the conjugate PLD, the radiation reaction activity of the radiation response group P (i.e., the activity of chemical reaction under irradiation) is significantly higher than that of the linker L and the active molecule D. For example, when the radiation response group P and the active molecule D both have an aromatic ring or a heteroaromatic ring, the radiation reaction activity of P is significantly higher than that of D, and the reasons may be one or more of the following: the electron cloud density of the aromatic ring or the heteroaromatic ring of P is higher than that of the aromatic ring or the heteroaromatic ring of D; the presence of a large substituent on the ring of D causes steric hindrance; the presence of a large group around the ring of D causes steric hindrance; the presence of one or more strong electron donating groups on the ring of P; the presence of a weak electron donating group or one or more electron withdrawing groups on the ring of D.

[0067] Therapeutic agents

[0068] The therapeutic agent in the conjugate PLD can be any therapeutic agent that can be coupled to PL or P, as long as it can be separated from PL or P and release a therapeutically active molecule under the action of radiation. In the present disclosure, the released therapeutically active molecule is generally the same structure as the therapeutic agent D in the conjugate PLD. In some embodiments, the released therapeutically active molecule is slightly different in structure from the therapeutic agent D in the conjugate PLD, that is, the therapeutic agent molecule undergoes a chemical reaction under the action of radiation but does not significantly affect its therapeutic activity.

[0069] Suitable molecules that can be used as therapeutic agents include, but are not limited to, polypeptides, oligopeptides, peptidomimetics, amino acids, enzyme inhibitors, hormones, toxins, antibiotics, anti-inflammatory substances, and the like.

[0070] In a preferred embodiment, the active molecule is a therapeutic agent for treating cancer.

[0071] When the active molecule is a therapeutic agent for treating cancer, the combined treatment of radiotherapy and chemotherapy can be achieved by means of this radiotherapy-activated prodrug, that is, while radiation is treating cancer, irradiation also activates the release of chemical anti-tumor drugs to achieve chemotherapy.

[0072] In one embodiment, the present disclosure uses a drug containing a primary amine or a secondary amine as a therapeutic agent. In a preferred embodiment, the present disclosure uses an anticancer drug containing a primary amine or a secondary amine as a therapeutic agent. Drugs containing primary or secondary amines include, for example, ibrutinib, acalabrutinib, zabutinib, doxorubicin, mitomycin-C, mitomycin-A, daunorubicin, aminopterin, actinomycin, bleomycin, 9-aminocamptothecin, N8-acetyl spermidine, 1-(2-chloroethyl)-1,2-dimethylmethanesulfonyl hydrazide, yunnanmycin, gemcitabine, cytarabine, dolastatin, dacarbazine, 5-fluorouracil and their derivatives. Drugs containing primary or secondary amines also include amino derivatives of drugs that do not naturally contain amino groups. In other words, drugs that do not originally contain amino groups can be chemically modified to have amino groups, and then coupled to radiation-responsive groups or to radiation-responsive groups and linkers by the primary or secondary amine coupling method described in the present disclosure.

[0073] In a preferred embodiment, the therapeutic agent is monomethyl auristatin E.

[0074] In one embodiment, the present disclosure uses a hydroxyl-containing drug as a therapeutic agent. In a preferred embodiment, the present disclosure uses a hydroxyl-containing anticancer drug as a therapeutic agent. Hydroxyl-containing therapeutic agents include, for example, paclitaxel, docetaxel, gemcitabine, cytarabine, etc. Hydroxyl-containing therapeutic agents also include hydroxyl derivatives of drugs that do not naturally contain hydroxyl groups. In other words, drugs that do not originally contain hydroxyl groups can be chemically modified to have hydroxyl groups, and then coupled to a radiation-responsive group or to a radiation-responsive group and a linker through the hydroxyl coupling method described in the present disclosure.

[0075] In one embodiment, the present disclosure uses a thiol-containing drug as a therapeutic agent. In a preferred embodiment, the present disclosure uses a thiol-containing anticancer drug as a therapeutic agent. Examples of thiol-containing therapeutic agents include 6-mercaptopurine, etc. The thiol-containing therapeutic agent also includes thiol derivatives of drugs that do not naturally contain thiol groups. In other words, a drug that does not originally contain a thiol group can be chemically modified to have a thiol group, and then coupled to a radiation-responsive group or to a radiation-responsive group and a linker through the thiol coupling method described in the present disclosure.

[0076] Imaging Agents

[0077] The imaging agent in the conjugate PLD can be any imaging agent that can be coupled to PL or P, as long as it can be separated from PL or P under the action of radiation and release a molecule that can be imaged. In the present disclosure, the released molecule that can be imaged is generally the same structure as the imaging agent in the conjugate PLD. In some embodiments, the released molecule that can be imaged is slightly different in structure from the imaging agent in the conjugate PLD, that is, the imaging agent molecule undergoes a chemical reaction under the action of radiation but does not significantly affect its imaging performance.

[0078] Imaging agents that can be used in the present disclosure include, but are not limited to, dyes, PET imaging agents (e.g. 18 F-FDG), radiolabeled reagents, etc.

[0079] Dyes such as rhodamine dyes, cyanine dyes, fluorescein dyes, etc. Representative examples of rhodamine dyes include, but are not limited to, 5-carboxytetramethylrhodamine (5-TAMRA), rhodamine B, rhodamine 6G, TRITC, Texas Red, rhodamine 123, sulforhodamine 101, etc. Examples of fluorescein dyes include, but are not limited to, fluorescein, fluorescein maleimide, 5-aminofluorescein, 6-aminofluorescein, fluorescein isocyanate (FITC), NHS-fluorescein, etc.

[0080] In some embodiments, radiolabeled agents can be used as imaging agents of the present teachings. In some embodiments, the rhodamine dye or fluorescein dye can be isotopically labeled. Examples of isotopes suitable for inclusion in the conjugate include hydrogen (e.g. 2 H and 3 H), carbon (e.g. 11 C. 13 C and 14 C), chlorine (e.g. 36 Cl), fluorine (e.g. 18 F), iodine (e.g. 123 I and 125 I), nitrogen (e.g. 13 N and 15 N), oxygen (e.g. 15 O. 17 O and 18 O), phosphorus (e.g. 32 P) and sulfur (e.g. 35 S) isotopes.

[0081] Isotopically labeled conjugates can be prepared by conventional techniques known to those skilled in the art, using an appropriate isotopically labeled reagent in place of the unlabeled reagent.

[0082] If the imaging agent has a hydroxyl group, a thiol group or an amino group, it can be coupled to the radiation response group or to the radiation response group and the linker by the hydroxyl group, thiol group or amino group coupling method described in the present disclosure. It is also possible to chemically modify the imaging agent that does not originally contain a hydroxyl group, thiol group or amino group to have a hydroxyl group, thiol group or amino group, and then couple it to the radiation response group or to the radiation response group and the linker by the hydroxyl group, thiol group or amino group coupling method described in the present disclosure.

[0083] In some embodiments, the conjugate PLD wherein D is an imaging agent is used as an imaging agent in an irradiated environment.

[0084] In some embodiments, the conjugate PLD in which D is an imaging agent is not used in an irradiation environment, but is used in an environment that can generate hydroxyl radicals, for example, it can be used as a fluorescent detector to detect the concentration of hydroxyl radicals in an aqueous solution when treating wastewater by oxidation using the Fenton reagent method. In this case, the conjugate PLD reacts with the hydroxyl radicals in the solution to release the imaging agent D, and the fluorescence intensity is detected using the fluorescence characteristics of the imaging agent, thereby calculating the concentration of hydroxyl radicals in the solution.

[0085] Coupling reaction

[0086] In the conjugate PLD, P and D are coupled via a linker L, or there is no linker L but P and D are directly coupled, that is, the conjugate is actually P—D.

[0087] The disclosed compounds can be prepared by a variety of methods known to those skilled in the art of organic synthesis. During the course of synthesis, it may be necessary to modify the groups using protecting groups to mitigate (especially eliminate) undesirable side reactions at the protected sites.

[0088] When the active molecule D contains hydroxyl, thiol and amino groups at the same time or contains multiple hydroxyl, thiol or amino groups, the other hydroxyl, thiol or amino groups can be protected first, and then coupled with the radiation response group P or with the radiation response group P and the linker L. Suitable hydroxyl protecting groups include trimethylsilyl (TMS), triethylsilyl (TES), tert-butyldimethylsilyl (TBDMS), tert-butyldiphenylsilyl (TBDPS), triisopropylsilyl (TIPS), etc. Suitable thiol protecting groups include trimethoxybenzyl (Tmob), diphenylmethyl, triphenylmethyl, tert-butyl, fluorenylmethyl (Fm), 2,4-dinitro-phenethyl (Dnpe), 9-fluorenylmethyleneoxycarbonyl (Fmoc), etc. Suitable amino protecting groups include trityl (Trt), 2,4-dimethoxybenzyl (Dmb), p-methoxybenzyl (PMB), phthaloyl (Pht), p-toluenesulfonyl (Tos), trifluoroacetyl (Tfa), o-(p-)nitrobenzenesulfonyl (Ns), pivaloyl, benzoyl, allyloxycarbonyl (Alloc), trimethylsilylethoxycarbonyl (Teoc), first (or second) oxycarbonyl, trifluoroacetyl, tert-butyloxycarbonyl (BOC), benzyloxycarbonyl (CBz) and 9-fluorenylmethyleneoxycarbonyl (Fmoc) etc. Those skilled in the art can easily determine whether the protection is needed. For a general description of protecting groups and their uses, see Greene and Wuts (Protective Groups In Organic Synthesis, Wiley-Interscience, 3rd Edition, 1999).

[0089] In one embodiment, the conjugate PLD contains a urea group, which can be synthesized by reacting a primary or secondary amine with triphosgene and then reacting with another amine molecule to synthesize urea. Commonly used solvents include: dichloromethane, tetrahydrofuran, etc. Commonly used bases include triethylamine, diisopropylethylamine, etc.

[0090]

[0091] In one embodiment, the conjugate PLD contains a urea group, which can be obtained by reacting a primary or secondary amine with a chloroformate such as p-nitrophenyl chloroformate under alkaline conditions (e.g., in the presence of diisopropylethylamine) to obtain the corresponding alkoxycarbonamide, which is then reacted with another amine molecule to synthesize urea. If a secondary amine is used to react with the chloroformate, it is usually carried out in DMAP-CH3CN.

[0092] In one embodiment, the conjugate PLD contains a thiourea group, which can be obtained by reacting a thio reagent (such as Lawsson's reagent) with urea prepared as above to give thiourea.

[0093] In one embodiment, the conjugate PLD contains a thiourea group, which can be synthesized into thiourea by reacting a primary or secondary amine with thiophosgene and then reacting with another amine molecule.

[0094] for The conjugate of the type (wherein X and Y are each independently O, S or NR', and R' is methyl or hydrogen) can be prepared by reacting R-OH, R-SH or R-NHR' (R' is methyl or hydrogen) with a chloroformate such as p-nitrophenyl chloroformate under alkaline conditions (e.g., in the presence of diisopropylethylamine), and then reacting with another molecule of R1-OH, R1-SH or R1-NHR' (R' is methyl or hydrogen). The preparation of such conjugates is illustrated as an example.

[0095]

[0096] The corresponding alcohol ROH (0.6 mmol, 1.0 eq) was dissolved in dichloromethane (4 mL, anhydrous) and cooled to 0°C. DIPEA (232 mg, 1.8 mmol, 3.0 eq) was added followed by compound 14 (503 mg, 1.5 mmol, 2.5 eq). The reaction mixture was warmed to room temperature over 1 hour, at which time it was quenched with water and extracted with dichloromethane (3 x 20 mL). The organic layers were combined, dried over anhydrous Na2SO4, and spin-dried under reduced pressure. The crude product was purified by silica gel chromatography using hexane / ethyl acetate (9:1).

[0097]

[0098] In a round-bottomed flask, R1OH (1mmol, 1.0 equivalent) and DIPEA (2mmol, 2.0 equivalent) were dissolved in 30mL THF and stirred for 2 minutes. Compound 15 (1mmol, 1.0 equivalent) was added and the mixture was stirred at room temperature for 1h. After the complete reaction was completed, the reaction solution was extracted with ethyl acetate and washed twice with water and saturated NaCl solution. The organic layers were combined, dried over anhydrous Na2SO4, and spin-dried under reduced pressure. The crude product was purified by silica gel chromatography.

[0099] Active molecule release experiment:

[0100] The compound is formulated into an aqueous solution and then exposed to different doses of radiation. The active molecules released from the solution are then quantitatively analyzed by HPLC-MS. The amount of active molecules released is plotted against the radiation dose to determine the release rate per unit dose.

[0101]

[0102] 10mM H2O2, t-BuOH or DMSO was added to the aqueous solution of compound 13 to change the yield of hydroxyl radicals under irradiation. Using 10μM potassium phosphate buffer of compound 13 as the control group, the yield of hydroxyl radicals was greatly increased in the solution with 10mM H2O2 added. Under this condition, the amount of Fmoc-lys released increased. In contrast, the amount of Fmoc-lys released was significantly reduced by adding 10mM t-BuOH and 10mM DMSO, two substances that can quench hydroxyl radicals, see Figure 1a . Figure 1a The graph shows the release of Fmoc-lys from compound 13 (10 μM, 0.02 M potassium phosphate buffer, pH=7.4, 0.1% DMF) after 10 Gy of γ-irradiation in four solutions (control group; containing 10 mM H2O2; containing 10 mM t-BuOH; containing 10 mM DMSO).

[0103] Without being bound by any particular theory, the applicant believes that high-energy rays cause water to undergo radiolysis to produce a large number of hydroxyl radicals, which then undergo electrophilic addition to the aromatic ring to generate phenol, which then releases the active ingredient Fmoc-lys through 1,4 or 1,6-elimination reaction (as shown in the following reaction formula).

[0104]

[0105] Compound 14 at high concentration (1 mM in pure water) was irradiated with γ-rays at 100 Gy / min ( 60 After irradiation with Co for 10 min and receiving a total dose of 1000 Gy, the inventors found a new UV absorption peak (see Figure 1b ), and its molecular weight indicated that a hydroxyl group was added to compound 14. After being stored at 37°C for 1 hour, the newly generated UV peak almost completely disappeared, while the mass spectrum peak of lysine was greatly enhanced. This indicates that the reaction is caused by the addition reaction of hydroxyl radicals with aromatic rings with high electron cloud density, followed by 1,4 or 1,6 elimination. In addition, in the irradiation study of compound 12, the inventors found the appearance of a removal product with the addition of a phenolic hydroxyl group through the single ion channel mode of the mass spectrometer.

[0106] Example

[0107] The starting material of embodiment is commercially available and / or can be prepared with the multiple methods known to the technical staff of organic synthesis field.The technical staff of organic synthesis field can suitably select reaction conditions (comprising solvent, reaction atmosphere, reaction temperature, duration of experiment and aftertreatment) in the following synthetic method.The technical staff of organic synthesis field will understand that the functional group present in each part of molecule should be compatible with proposed reagent and reaction.NMR uses BrukerAVANCE 400MHz spectrometer to record.High resolution mass spectrum uses Bruker Fourier Transform Ion Cyclotron resonance mass spectrometer to measure.The liquid chromatography-mass spectrometry used is Waters e2695 instrument, is equipped with Waters2995PDA and Waters Acquity QDA mass spectrometer.

[0108] 1. Exemplary Synthesis of PLD Conjugate with Fmoc-Lysine

[0109] Step 1: Synthesis of intermediate compound 15

[0110]

[0111] The corresponding alcohol R-OH (0.6 mmol, 1.0 equiv) was dissolved in anhydrous dichloromethane (4 mL) and cooled to 0°C. N,N-diisopropylethylamine (232 mg, 1.8 mmol, 3.0 equiv) was added, followed by p-nitrophenyl chloroformate (503 mg, 1.5 mmol, 2.5 equiv). The reaction mixture was warmed to room temperature over 1 hour, at which time it was quenched with water and extracted with dichloromethane (3×20 mL). The organic layers were combined, dried over anhydrous Na2SO4, and spin-dried under reduced pressure. The crude product was purified by silica gel chromatography using hexane / ethyl acetate (9:1) to give compound 15.

[0112] Step 2: Synthesis of conjugate PLD

[0113]

[0114] In a round-bottom flask, Fmoc-lysine (1 mmol, 1.0 equiv) and N,N-diisopropylethylamine (2 mmol, 2.0 equiv) were dissolved in 15 mL THF and 15 mL H2O and stirred for 2 min. Compound 15 (1 mmol, 1.0 equiv) was added and the mixture was stirred at room temperature for 1 h. After the reaction was completed, the reaction solution was extracted with ethyl acetate and washed twice with water and saturated NaCl solution. The organic layers were combined, dried over anhydrous Na2SO4, and spin-dried under reduced pressure. The crude product was purified by silica gel chromatography, eluting with hexane / ethyl acetate (4:1), ethyl acetate (100%), dichloromethane / methanol (25:1), and ethyl acetate / methanol (10:1) to obtain the conjugate PLD.

[0115] By selecting different ROH, the compounds in Table 1 were synthesized according to the above method. NMR and MS data of exemplary compounds:

[0116]

[0117] 1 H NMR (400MHz, DMSO-d6) δ12.58(s,1H),7.90(d,J=7.5Hz,2H),7.73(d,J=7.5Hz ,2H),7.62(d,J=8.1Hz,1H),7.42(t,J=7.5Hz,2H),7.33(t,J=7.5,2H),7.07( t,J=5.7Hz,1H),4.25–4.17(m,1H),3.96(q,J=7.1Hz,2H),3.90(m,1H),2.95( q,J=6.5Hz,2H),1.75–1.50(m,2H),1.43–1.27(m,4H),1.14(t,J=7.0Hz,3H).

[0118] 13 C NMR (101 MHz, DMSO-d6) δ 174.47, 156.70, 156.61, 144.31, 144.27, 141.18, 141.17, 128.10, 127.53, 125.76, 125.74, 120.58, 66.05, 59.88, 54.28, 47.12, 30.90, 29.49, 23.40, 15.16. HRMS (ESI): calculated value C 24 H 29 N2O6([M+H] + ):441.2020, measured value:441.2019.

[0119]

[0120] 1H NMR (400MHz, DMSO-d6) δ12.60(s,1H),7.90(d,J=7.5Hz,2H),7.73(d,J=7.5Hz,2H),7.62(d,J=7.9Hz,1H),7.41(q,J=7.7Hz,2H ),7.40–7.23(m,8H),5.00(s,2H),4.32–4.18(m,3H),3.90(m,1H),2.98(q,J=6.5Hz,2H)1.75–1.50(m,2H),1.43–1.27(m,4H),.

[0121] 13 C NMR(101MHz,DMSO-d6)δ174.49,156.62,156.54,144.30,144.26,141.17,137.72,128.81,128.20 ,128.10,127.53,125.75,120.58,66.04,65.58,54.29,47.11,40.51,39.85,30.90,29.45,23.41.

[0122] HRMS (ESI): Calculated value C 29 H 31 N2O6([M+H] + ):503.2177, measured value:503.2166.

[0123]

[0124] 1 H NMR (400MHz, DMSO-d6) δ12.58(s,1H),7.89(d,J=7.5Hz,2H),7.73(d,J=7.5Hz,2H),7.63 (d,J=8.0Hz,1H),7.42(t,J=7.5Hz,2H),7.33(t,J=7.5Hz,2H),7.29(d,J=5.7Hz,1H),6.5 0(d,J=2.3Hz,2H),6.42(t,J=2.3Hz,1H),4.94(s,2H),4.29(d,J=7.9Hz,2H),4.25–4.21 (m,1H),3.91(m,1H),3.72(s,6H),3.00(q,J=6.5Hz,2H),1.75–1.56(m,2H),1.39(m,4H).

[0125] 13C NMR(101MHz,DMSO-d6)δ174.45,160.91,156.63,156.48,144.30,141.17,140.10,128.10,127.53 ,125.76,120.57,105.74,99.81,66.05,65.40,55.62,54.24,47.12,40.51,30.86,29.44,23.41.

[0126] HRMS (ESI): Calculated value C 31 H 35 N2O8([M+H] + ):563.2388, measured value:563.2388.

[0127]

[0128] 1 H NMR (400MHz, DMSO-d6) δ12.59 (s, 1H), 9.26 (s, 2H), 7.90 (d, J = 7.5Hz, 2H), 7.73 (d, J=7.4Hz,2H),7.65(d,J=7.9Hz,1H),7.41(t,J=7.4Hz,2H),7.33(t,J=7.4,2H),7. 25(t,J=5.7Hz,1H),6.15(d,J=2.2Hz,2H),6.08(t,J=2.3Hz,1H),4.82(s,2H),4.3 5–4.18(m,3H),3.91(m,1H),2.98(q,J=6.5Hz,2H),1.77–1.54(m,2H),1.38(m,4H).

[0129] 13 C NMR(101MHz,DMSO-d6)δ174.47,158.81,156.64,156.56,144.30,141.16,139.69,128.11,127 .54,125.77,120.58,105.94,102.18,66.04,65.54,54.25,47.11,39.83,30.85,29.48,23.43.

[0130] HRMS (ESI): Calculated value C 29 H 31 N2O8([M+H] + ):535.2075, measured value:535.20734.

[0131] 2. Experiment on the release of active molecules from the PLD conjugate with Fmoc-lysine as D

[0132] The compound was dissolved in physiological saline at room temperature to prepare a 10 μM aqueous solution, and then irradiated using 60Co as a γ-ray source (or a radiotherapy instrument as an X-ray source) (all at 4 Gy / min). After irradiation for different times (2.5 min, 5 min, 10 min, 15 min, 20 min, 25 min), the Fmoc-Lys produced in the solution was quantitatively analyzed by HPLC-MS. The amount of released Fmoc-lys was then plotted against the radiation dose to determine the release rate.

[0133] Table 1: Release rate of conjugate PLD under 4Gy / min X-ray or γ-ray

[0134]

[0135]

[0136]

[0137] 3. Preparation of Fqf-OH

[0138]

[0139] Synthesis scheme of Fqf-OH

[0140] Reagents and conditions: (a) K2CO3, CH3I, DMF, rt, 12h; (b) 2N NaOH, MeOH, rt, 10h; (c) Tf2O, pyridine, CH2Cl2, 0℃ to room temperature; (d) Pd2(dba)3·CHCl3, xantphos, Cs2CO3, dibenzophenone imine, toluene, 105℃, 12h; (e) 1N HCl, THF, rt, 30min; (f) triphosgene, TEA, THF, rt, 19h; (g) iPrNEt2,3,5-bis(tert-butyldimethylsilyloxy)phenyl]methanol, CH2Cl2, rt, 1h; (h) TBAF, THF, rt, 10h.

[0141]

[0142] Step a, under N2 atmosphere, K2CO3 (9.88 g, 71.50 mmol) was slowly added to a solution of fluorescein (10 g, 28.60 mmol) in N,N-dimethylformamide (90 mL), and the reaction mixture was stirred at room temperature for 1 h. Then, iodomethane (12.18 g, 85.80 mmol) was added to the reaction mixture, and the mixture was stirred at room temperature for 12 h. The reaction was monitored by TLC, and after the reaction was completed, ice water was added to the reaction mixture and stirred at 0 ° C for 30 minutes. The yellow solid was filtered and washed with water to completely wash off K2CO3. The solid was dried to obtain the desired compound (10.30 g) in quantitative yield.

[0143] 1 H NMR (400MHz, DMSO-d6) δ8.21 (dd, J=8.0, 1.1Hz, 1H), 7.87 (td, J=7.5, 1.4Hz, 1 H),7.78(td,J=7.5,1.2Hz,1H),7.50(dd,J=7.5,1.1Hz,1H),7.23(d,J=2.7Hz, 1H),6.89(dd,J=8.7,2.3Hz,1H),6.84(d,J=9.1Hz,1H),6.80(d,J=9.6Hz,1H) ,6.39(dd,J=9.6,1.8Hz,1H),6.24(d,J=1.8Hz,1H),3.91(s,3H),3.58(s,3H).

[0144] 13 C NMR(100MHz,DMSO-d6)δ184.38,165.72,164.42,158.89,154.10,150.58,134.43,133.73,131.24 ,130.89,130.58,130.03,129.91,129.38,117.17,114.82,114.10,105.12,101.11,56.82,52.84.

[0145] HRMS(ESI + ): m / z calculated value C 22 H 17 O5[M+H] + :361.1076, measured value: 361.1072.

[0146]

[0147] Step b, slowly add 2N NaOH (50mL) aqueous solution to the methanol (150mL) solution of compound 30 (10.30g, 28.60mmol). The reaction mixture is stirred at room temperature for 10 hours. Methanol is evaporated under reduced pressure, and the water layer is washed twice with ether. The water layer is then acidified with 6N HCl. The solid is filtered, washed with water to completely remove HCl to reach pH=7, and dried to obtain the desired compound 31 (9.45g) in quantitative yield.

[0148] 1 H NMR (400MHz, DMSO-d6) δ10.16(s,1H),8.00(d,J=7.3Hz,1H),7.79(td,J=7.5,1.2Hz,1H),7.72(td,J=7.5,0.9Hz,1H),7. 27(d,J=7.8Hz,1H),6.93(d,J=2.7Hz,1H),6.74-6.67(m,2H),6.64(d,J=8.7Hz,1H),6.57(d,J=1.4Hz,2H),3.81(s,3H).

[0149] 13 C NMR(100MHz,DMSO-d6)δ169.20,161.56,160.11,153.00,152.41,152.30,152.22,136.17,130.67,129.63 ,129.48,126.58,125.20,124.54,113.34,112.42,111.53,109.98,102.73,101.31,83.31,65.44,56.18.

[0150] HRMS(ESI + ):m / z C 21 H 15 O5 Calcd. [M+H] + :347.0920, measured value: 347.0916.

[0151]

[0152] Step c, adds pyridine (0.91g, 12mmol) to anhydrous dichloromethane (12mL) solution of compound 31 (1.00g, 2.89mmol), and the reaction mixture is stirred at room temperature for 20min. The reaction mixture is cooled to 0 ℃, and trifluoromethanesulfonic anhydride (1.63g, 5.79mmol) is slowly added to the reaction mixture within 30 minutes. The reaction mixture is warmed to room temperature and stirred for 3 hours. The reaction is quenched with water and extracted with dichloromethane. The organic layer is washed with 1N HCl solution, water and brine successively. The combined organic layer is dried over anhydrous sodium sulfate, and volatiles are removed under reduced pressure. The crude residue is purified by flash chromatography to obtain compound 32 (0.98g) with a yield of 71%.

[0153] 1 H NMR (400 MHz, CHLOROFORM-d) δ 8.05 (d, J = 7.3 Hz, 1H), 7.71 (td, J = 7.5, 1.2 Hz, 1H), 7.66 (td, J = 7.4, 1.2 Hz, 1H), 7.25 (d, J = 2.4 Hz, 1H), 7.18 (d, J = 7.5, 1H), 7.00–6.87 (m, 2H), 6.80 (d, J = 2.5 Hz, 1H), 6.76–6.63 (m, 2H), 3.85 (s, 3H).

[0154] 13 C NMR (100 MHz, chloroform-d) δ 168.96, 161.67, 152.63, 152.06, 151.89, 149.98, 135.38, 130.18, 130.09, 129.02, 126.30, 125.34, 123.88, 120.27, 119.82, 116.71, 112.47, 110.54, 110.49, 100.93, 81.64, 55.67.

[0155] HRMS(ESI + ):m / z C 22 H 14 Calculated value for F3O7S [M+H] + :479.0412, measured value: 479.0410.

[0156]

[0157] Steps d and e, before the reaction, all glassware are dried in an oven. Pd2(dba)3·CHCl3(0.21g, 0.24mmol) and xantphos(0.18g, 0.31mmol) are added to a solution of compound 32(0.98g, 2.04mmol) in toluene (20mL). The reaction mixture is purged with nitrogen, and cesium carbonate (0.93g, 2.86mmol) and benzophenone imine (0.44g, 2.45mmol) are then added thereto. The reaction mixture is heated at 105°C for 12 hours under a nitrogen atmosphere. After the reaction is completed by TLC, the reaction mixture is filtered through a short diatomaceous earth pad, which is then washed with dichloromethane. The filtrate is concentrated under reduced pressure, and the crude residue can be used for the next step reaction without further purification. The crude compound is dissolved in tetrahydrofuran (30mL), and 3mL 1N HCl is then added. The reaction mixture is stirred at room temperature for 30 minutes, and extracted with ethyl acetate. The organic layer was dried over anhydrous sodium sulfate, and the volatiles were removed under reduced pressure. The crude residue was purified by flash column chromatography with an eluting solvent (PE:EA=7:3) to give Compound 34 (0.58 g) in 82% yield.

[0158] 1 H NMR (400MHz, DMSO-d6) δ7.98(d,J=7.8Hz,1H),7.79(td,J=7.5,0.9Hz,1H),7.71(td,J=7.5,0.9Hz,1H),7.25(d,J=7.3Hz,1H),6.91(d,J=2.3Hz,1H),6 .68(dd,J=8.9,2.5Hz,1H),6.60(d,J=8.7Hz,1H),6.45(d,J=2.3Hz,1H),6. 38(d,J=8.2Hz,1H),6.34(dd,J=8.5,2.1Hz,1H),5.66(s,2H),3.81(s,3H)).

[0159] 13 C NMR(100MHz,DMSO-d6)δ169.29,161.43,153.03,152.66,152.50,151.88,136.00,130.49,129.37 ,129.02,126.92,125.04,124.52,112.06,111.87,111.79,105.82,101.27,99.60,84.20,56.15.

[0160] HRMS(ESI + ):m / z C 21 H 16Calculated value of NO4 [M+H] + :346.1079, measured value: 346.1071.

[0161]

[0162] Step f, before the reaction, all glassware are dried in an oven. In the reactor, compound 34 (0.83g, 2.40mmol) is dissolved in 16mL anhydrous tetrahydrofuran. Triphosgene (0.713g, 2.403mmol) is dissolved in 16mL anhydrous tetrahydrofuran and then injected into the reactor. Then the 2.92mL triethylamine (21.63mmol) of distillation is added. White fumes (gaseous phosgene) are formed immediately. The reactor is sealed, and the solution is vigorously stirred at room temperature for 19 hours. The mixture is concentrated under reduced pressure. The crude residue is quickly filtered, and the solid is washed with anhydrous tetrahydrofuran. Removal of solvent under reduced pressure obtains compound 6, which is brown oil (quantitative), which can be used without further purification.

[0163]

[0164] Step g, under ice-water bath, to a solution of compound 35 (0.05 g, 0.14 mmol) in dichloromethane (4 mL) was added a solution of triethylamine (0.037 g, 0.29 mmol) in dichloromethane (2 mL), and then 3,5-bis (tert-butyldimethylsilyloxy) phenyl] methanol (0.063 g, 0.17 mmol) was added. The reaction mixture was stirred at room temperature for 1 h. The solvent was evaporated under reduced pressure, and the residue was purified by flash column chromatography to obtain compound 36 (0.075 g) in a yield of 72%.

[0165] 1 H NMR (400 MHz, chloroform-d) δ 8.02 (dt, J = 7.3, 1.1 Hz, 1H), 7.66 (td, J = 7.5, 1.2 Hz, 1H), 7.61 (td, J = 7.5, 1.2 Hz, 1H), 7.55 (s, 1H), 7.25 (d, J = 2.4 Hz, 1H), 7.18 (d, J = 7.5, 1H) ,7.00–6.87(m,2H),6.80(d,J=2.5Hz,1H),6.76–6.63(m,2H),6.49(d,J=2.2Hz,2 H), 6.30 (t, J = 2.2Hz, 1H), 5.08 (s, 2H), 3.84 (s, 3H), 0.97 (s, 18H), 0.22 (s, 12H);

[0166] 13C NMR (100 MHz, chloroform-d) δ 161.38, 156.73, 152.45, 151.98, 139.86, 137.55, 135.03, 129.72, 129.02, 128.80, 125.05, 123.92, 113.17, 111.92, 111.78, 111.02, 100.84, 77.23, 67.04, 55.60, 25.68, 18.22, −4.36.

[0167] HRMS(ESI + ):m / z C 41 H 50 Calculated value of NO8Si2 [M+H] + :740.3069, measured value: 740.3072.

[0168]

[0169] Step h, at 0°C, a 1.0 M solution of tetrabutylammonium fluoride (TBAF) (230 μL, 0.23 mmol) was added dropwise to a solution of 36 (75 mg, 0.1 mmol). The mixture was stirred at room temperature for 10 hours. The solution was poured into water,

[0170] The residue was purified by silica gel flash column chromatography to give 37 (35.8 mg, 0.07 mmol, 70%) as an orange solid.

[0171] 1 H NMR (400MHz, acetonitrile-d3) δ8.09(s,1H),7.66(td,J=7.5,1.2Hz,1H),7.61(td,J=7.5,1.2Hz,1H),7.55(s,1H),7.25(d,J=2.4Hz,1H),7.18(d,J=7 .5,1H),7.00–6.87(m,2H),6.80(d,J=2.5Hz,1H),6.76–6.63(m,2H),6.49(d,J=2.2Hz,2H),6.30(t,J=2.2Hz,1H),5.08(s,2H),3.83(s,3H).

[0172] 13C NMR (100 MHz, acetonitrile-d3) δ 169.55, 162.23, 158.71, 153.88, 152.90, 152.18, 141.76, 139.72, 135.87, 130.56, 129.65, 129.23, 127.00, 125.27, 124.43, 114.87, 113.92, 112.28, 111.73, 106.65, 106.00, 102.44, 101.33, 66.68, 55.98.

[0173] HRMS(ESI + ):m / z C 29 H 22 Calculated value of NO8 [M+H] + :512.1340, measured value: 512.1339.

[0174] 4. In vitro fluorescent molecular release of Fqf-OH

[0175] In order to demonstrate the broad-spectrum feasibility of the small molecule release strategy under irradiation conditions and to facilitate detection, the inventors introduced a highly reactive resorcinol structure into the fluorescent molecule. Under the action of the hydroxyl radicals generated by irradiation, the fluorescent precursor molecule Fqf-OH undergoes a removal reaction, thereby releasing the fluorescent molecule.

[0176]

[0177] Fqf-OH was dissolved in 0.02 M phosphate buffer (pH = 7.4) to prepare a 10 μM solution (0.1% DMF as a cosolvent), and then exposed to 1 Gy / min of gamma rays ( 60 Co). By controlling the irradiation time, the total dose of the solution was irradiated in a gradient of 10Gy, and the doses were 0Gy, 10Gy, 20Gy, 30Gy, 40Gy, 50Gy, 60Gy, and 70Gy. The fluorescence intensity (measured at 518nm, with an excitation wavelength of 477nm) gradually increased with the increase of the dose ( Figure 2a ) and has a good linear relationship ( Figure 2b ). In order to verify that Fqf-OH has high selectivity for hydroxyl radicals, Fqf-OH was exposed to different RNS and ROS. It was found that only under the action of hydroxyl radicals could fluorescent molecules be released, and its reaction ability was more than 30 times that of other ROS and RNS ( Figure 2c ). Fqf-OH is stable to various metal ions such as sodium, potassium, calcium, iron, zinc ions and amino acids. Moreover, Fqf-OH is stable inside cells, and adding substances that exist at high concentrations inside cells does not restore the fluorescence activity.

[0178] 5. Intracellular fluorescent molecule release experiment of Fqf-OH

[0179] After verifying the high selectivity and high reactivity of Fqf-OH for the production of hydroxyl radicals under irradiation, it was introduced into the cell experiment to verify the release of Fqf-OH in cells. Before introducing it into the cell experiment, its toxicity was tested, and the cell survival rate was greater than 95% at 10μM Fqf-OH.

[0180] After culturing HeLa cells in the confocal dish for 24 h, the medium was removed, the cells were washed three times with HBSS, and then incubated with 10 μM Fqf-OH in HBSS solution (pH = 7.4, 0.1% DMF) for 30 min before being exposed to 60 After irradiation with Co source γ-ray at 1 Gy / min for 1 min and 5 Gy / min for 1, 2, and 4 min, confocal imaging with an excitation wavelength of 488 nm was performed. Figure 3 The amount of released fluorescent molecules increases with the increase of dose. After deducting the cell autofluorescence background, the fluorescence intensity increased by 2.6 times at a dose of 1Gy, and increased by 6.6, 12, and 18 times at 5Gy, 10Gy, and 20Gy, respectively ( Figure 3 The lower bar graph shows the fluorescence intensity of the three cells with Fqf-OH obtained by confocal imaging). The same experimental results can be obtained on 4T1 cells and MC38 ( Figure 3 ).

[0181] The above cell experiments show that precursor molecules can release fluorescent molecules in most cells through gamma rays.

[0182] 6. In vivo fluorescent molecule release

[0183] In order to verify the release of fluorescent precursor molecules in mouse tumors, we used the hospital's radiotherapy equipment. Since the radiotherapy equipment produces X-rays, we first proved that the release effect of fluorescent precursor molecules under X-rays and gamma rays is the same at the same dose. 20μL of 200μM (1% DMF as a cosolvent) PBS solution of fluorescent precursor Fqf-OH was locally injected into 4T1 tumor-bearing mice, and the mice were irradiated with 4Gy half an hour later. In order to simulate the tumor treatment experiment, we repeated this process 3 times ( Figure 4 After the mice were killed, the tumors were removed and sliced ​​for fluorescence imaging. It was found that the experimental group injected with fluorescent precursor molecules had a very obvious increase in fluorescence signal compared with the control group injected with saline ( Figure 4 ), after deducting the tissue autofluorescence background, the fluorescence intensity increased by 2.5 times after treatment. This experiment shows that fluorescent precursor molecules can release fluorescent molecules in vivo through X-ray irradiation.

[0184] 7. Synthesis of PLD Conjugate with Monomethyl Auristatin E

[0185] The synthesis of the conjugate PLD is illustrated below using compound 27 as an example, wherein P is a resorcinol group, L is -CH2-OC(=O)-, and D is monomethyl auristatin E (MMAE for short).

[0186]

[0187] Intermediate compound 24

[0188] Compound 23 (5 mmol) was dissolved in dichloromethane (50 mL), and N, N-diisopropylethylamine (15 mmol, 3 equiv) was added, and the mixture was stirred vigorously at room temperature for ten minutes. Tert-butyldimethylsilyl chloride (10.5 mmol, 2.1 equiv) was added under an ice-water bath, and after 30 minutes, the mixture was returned to room temperature and reacted for another 2 hours. The reaction solution was diluted with water and extracted three times with ethyl acetate. The organic phases were combined and washed with hydrochloric acid solution at pH = 3, saturated sodium bicarbonate solution, water, and saturated brine in sequence. After being spin-dried, petroleum ether / ethyl acetate system (10:1) was used as eluent and compound 24 was obtained by purification on a silica gel column with a yield of 95%.

[0189] Intermediate compound 25

[0190] Compound 24 (4.5 mmol) was dissolved in ultra-dry tetrahydrofuran (50 mL), and LiAlH4 (4.5 mmol, 1 equiv) was added in batches under an ice-water bath. After 30 minutes, it was returned to room temperature and reacted for another 2 hours. Water was slowly added dropwise to the reaction solution under an ice-water bath to quench the reaction. The filtrate was filtered through diatomaceous earth, diluted with water, and extracted three times with ethyl acetate. The organic phases were combined and washed with hydrochloric acid solution at pH = 3, saturated sodium bicarbonate solution, water, and saturated brine in sequence. After being spin-dried, petroleum ether / ethyl acetate system (4:1) was used as eluent and compound 25 was purified by silica gel column with a yield of 98%.

[0191] Intermediate compound 26

[0192] Compound 25 (4 mmol) was dissolved in DCM (120 mL), and N,N-diisopropylethylamine (6 mmol, 1.5 equiv) was added and stirred for 10 minutes. Then p-nitrophenyl chloroformate (4.8 mmol, 1.2 equiv) was added. After 2 hours, the reaction solution was diluted with water and extracted three times with ethyl acetate. The organic phases were combined and washed with hydrochloric acid solution at pH = 3, saturated sodium bicarbonate aqueous solution, water, and saturated brine in sequence. After being spin-dried, petroleum ether / ethyl acetate system (4:1) was used as eluent and purified by silica gel column to obtain compound 26 with a yield of 81%.

[0193] Compound 27

[0194] Compound 26 (0.6 mmol) was dissolved in DCM (20 mL), 1-hydroxybenzotriazole (0.7 mmol, 1.2 equiv) and N,N-diisopropylethylamine (1.2 mmol, 2 equiv) were added, and the mixture was stirred for 10 minutes. MMAE (0.6 mmol, 1.0 equiv) was then added. The reaction mixture was allowed to react overnight. The reaction solution was diluted with water and extracted three times with ethyl acetate. The organic phases were combined and washed with a pH=3 hydrochloric acid solution, a saturated sodium bicarbonate aqueous solution, water, and a saturated saline solution. After being spun dry, the crude product was dissolved in THF (20 mL), and 1.4 mL (1.4 mmol, 1.2 equiv) of tetrabutylammonium fluoride in tetrahydrofuran solution was added. The mixture was reacted for 2 hours, spun dry, and a dichloromethane / methanol system (20:1) was used as an eluent. Compound 27 was purified by a silica gel column with a yield of 58%. High-resolution mass spectrometry: C 47 H 74 N5O 11 [M+H] + Calculated value 884.5379, measured value 884.5365.

[0195] 8. Experiment on the release of active molecules from compound 27

[0196]

[0197] Compound 27 was dissolved in physiological saline at room temperature to prepare a 10 μM aqueous solution. 60 Co was used as a γ-ray source (or a radiotherapy instrument as an X-ray source) for irradiation (all at 4 Gy / min). After irradiation for different time periods (2.5 min, 5 min, 10 min, 15 min, 20 min, and 25 min), the concentration of MMAE produced in the solution was quantitatively analyzed by HPLC-MS (see Figure 5The upper right corner of the figure shows the change in the mass spectrometry signal peak of the drug molecule after irradiation), and then the amount of released MMAE is plotted against the radiation dose. The drug molecule concentration has a good linear relationship with the dose, and the release rate is about 40nM / Gy (see Figure 5 ).

[0198] The 10 μM aqueous solution of compound 27 (prodrug molecule) that was not irradiated was diluted to 10 nM and then incubated with cells. It was found that the aqueous solution of compound 27 that was not irradiated had almost no cytotoxicity. 60 The cytotoxicity of Co after 10 Gy of γ-ray irradiation was restored to the same level as that of the drug molecule MMAE. From the relationship between cell survival and the logarithm of concentration ( Figure 5 As can be seen from the lower right corner of the figure, the cytotoxicity of the irradiated solution is significantly improved compared with the unirradiated prodrug molecules.

[0199] The embodiments of the present disclosure illustrate the feasibility of the strategy of releasing active molecules by irradiation in principle through fluorescent precursor molecules. Further, the embodiments of the present disclosure illustrate the feasibility of the strategy of releasing drugs by irradiation using MMAE drug molecules as a model. The embodiments of the present disclosure also illustrate that the concentration of active molecules released from the conjugate has a good linear relationship with the dose.

[0200] The above description is merely an exemplary embodiment of the present invention and is not intended to limit the protection scope of the present invention. The protection scope of the present invention is determined by the appended claims.

[0201] The present invention specifically relates to the following:

[0202] 1. A conjugate of general formula (I),

[0203] P——L——D (I)

[0204] Wherein P is a radiation responsive group containing an aromatic group or a heteroaromatic group; L is absent or is a divalent linking group; and D is an imaging agent or a therapeutic agent.

[0205] 2. A conjugate according to item 1, wherein the radiation responsive group is a group that can chemically react with the hydroxyl radicals generated by radiation and thereby fall off from the conjugate, or the radiation responsive group is a group that can chemically react with the hydroxyl radicals generated by radiation and thereby fall off from the conjugate together with the linker L.

[0206] 3. The conjugate according to item 1, wherein the aryl or heteroaryl group is selected from the group consisting of phenyl, naphthyl, anthracenyl, phenanthrenyl, biphenyl, pyridyl, pyridazinyl, pyrimidinyl, pyrazinyl, triazinyl, tetrazinyl, furanyl, thienyl, pyrrolyl, pyrazolyl, imidazolyl, triazolyl, tetrazolyl, Azolyl, iso Azolyl, Oxazolyl, triazolyl, furazanyl, thiazolyl, isothiazolyl, indolyl, benzothiazolyl, benzodioxolyl, benzo oxazolyl, benzothienyl, quinolyl, tetrahydroisoquinolyl, isoquinolyl, benzimidazolyl, benzopyranyl, indolizinyl, benzofuranyl, chromonyl, coumarinyl, benzopyranyl, cinnolinyl, quinoxalinyl, indazolyl, pyrrolopyridinyl, furopyridinyl, dihydroisoindolyl, tetrahydroquinolyl, carbazolyl, dibenzofuranyl, dibenzothienyl, benzindolyl, phenanthrolinyl, acridinyl, phenanthridinyl and xanthenyl;

[0207] wherein the aryl or heteroaryl is optionally substituted with one or more substituents selected from the group consisting of halogen, -R1, -NR1R2, -CN, -NO2, -N3, -OR1, -SR1, -NHCOR1, -O-COR1, -CH=CR1R2, -C(=O)-R1, -C(=O)-OR1, -C(=O)-Cl, -C(=O)-NH2, -C(=O)-NH-R1 and -C(=O)-NR1R2;

[0208] Wherein R1 and R2 are independently selected from H, C1-C6 alkyl, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C 10 Cycloalkyl, C6-C 20 The alkyl, alkenyl, cycloalkyl, aryl and heteroaryl groups are optionally substituted with one or more halogen, hydroxyl, mercapto, -CN, nitro, -N3, C1-C6 alkyl, C1-C6 alkoxy, C1-C6 haloalkyl, C2-C6 alkenyl, C3-C 10 Cycloalkyl, C6-C 20 The moiety is substituted with aryl or heteroaryl having 5 to 20 ring atoms.

[0209] 4. A conjugate according to claim 3, wherein the aryl or heteroaryl group is selected from phenyl, pyrrolyl, furanyl, thienyl, pyrazolyl and imidazolyl, and the aryl or heteroaryl group is optionally substituted with one or more amino, amine, -NHCOCH3, phenyl, methoxy, ethoxy, methyl, ethyl, hydroxyl, alkenyl, F, Cl, Br, I, -N3 or methyl.

[0210] 5. The conjugate according to item 3, wherein the aryl or heteroaryl group has no oxo, -C(=O)-, -C(=S)-, -CN, -SO3H, or -CF3 directly attached thereto.

[0211] 6. The conjugate according to item 5, wherein the aryl group is an optionally substituted resorcinol group.

[0212] 7. The conjugate according to any one of items 1 to 6, wherein

[0213] L is selected from -SS-, linear C1-C6-alkylene-, branched C3-C6-alkylene-, -R3-XC(=O)-Y-, -R3-XC(=S)-Y-, -R3-X-, -XC(=O)-Y-, -XC(=S)-Y- or -CH2-XC(=Z)-Y-CH2CH2-EC(=J)-F-, wherein R3 is selected from C1-C3 alkylene or -CH2-C2 alkenyl-; X, Y, E and F are independently selected from O, NH or S; Z and J are independently selected from O or S; L may be optionally substituted with hydroxyl, thiol, NH2, F, Cl, Br, I, -N3, methyl.

[0214] 8. The conjugate according to item 7, wherein L is -CH2-X-(C=O)-Y-, -CH2-X-(C=S)-Y-, -CH2-X- or -X-(C=O)-Y-, wherein X and Y are independently selected from O, NH, S.

[0215] 9. The conjugate according to item 8, wherein L is -CH2-O-(C=O)-, -CH2-O-(C=O)-NH- or -O-(C=O)-NH-.

[0216] 10. The conjugate according to item 1, wherein D is a fluorescent imaging agent or a therapeutic agent for treating cancer.

[0217] 11. The conjugate according to claim 10, wherein no aromatic or heteroaromatic group is present in the fluorescent imaging agent or the therapeutic agent for treating cancer.

[0218] 12. The conjugate according to claim 10, wherein the fluorescent imaging agent or the therapeutic agent for treating cancer contains an aromatic or heteroaromatic group, and the aromatic or heteroaromatic group contains an oxo, -C(=O)-, -C(=S)-, -CN, -SO3H or -CF3 group directly attached to the aromatic or heteroaromatic group.

[0219] 13. A conjugate according to claim 1, wherein the therapeutic agent is selected from one or more of the following: monomethyl auristatin E, monomethyl auristatin F, ibrutinib, acalabrutinib, zanubrutinib, doxorubicin, mitomycin-C, mitomycin-A, daunorubicin, aminopterin, actinomycin, bleomycin, 9-aminocamptothecin, N8-acetyl spermidine, 1-(2-chloroethyl)-1,2-dimethanesulfonyl hydrazide, yunnanmycin, gemcitabine, cytarabine, dolastatin, dacarbazine, 5-fluorouracil; paclitaxel, docetaxel, gemcitabine, cytarabine; 6-mercaptopurine.

[0220] 14. The conjugate of claim 13, wherein the therapeutic agent is monomethyl auristatin E, monomethyl auristatin F, or 5-fluorouracil.

[0221] 15. The conjugate according to claim 1, wherein the imaging agent is selected from rhodamine dyes, cyanine dyes, fluorescein dyes, PET imaging agents, and radiolabeled agents.

[0222] 16. The conjugate according to item 15, which is Fqf-OH.

[0223] 17. A method of treatment or imaging, comprising orally administering or injecting a conjugate of any one of items 1 to 16 into a patient, and then subjecting the patient to precise irradiation.

[0224] 18. A method according to claim 17, wherein the patient is a human or mammal suffering from cancer.

[0225] 19. Use of the conjugate of any one of items 1 to 16 in the qualitative or quantitative detection of hydroxyl radicals.

[0226] 20. A method for qualitatively or quantitatively detecting hydroxyl free radicals: comprising adding Fqf-OH to a solution to be tested and measuring the fluorescence intensity of the solution.

Claims

1. A conjugate of general formula (I), P——L——D(I) Wherein P is a radiation responsive group containing an aromatic or heteroaromatic group; L is absent or is a divalent linking group; and D is an imaging agent or a therapeutic agent; wherein the aryl or heteroaryl is selected from phenyl, pyrrolyl, furyl, thienyl, pyrazolyl, imidazolyl, Azolyl, iso Azolyl, thiazolyl, isothiazolyl, the aryl or heteroaryl being substituted with one or more alkoxy, C1-C6 alkyl, hydroxyl, C2-C6 alkenyl, F, Cl, Br, I or -N3; Wherein when L is a divalent linking group, L is selected from -SS-, linear C1-C6-alkylene-, branched C3-C6-alkylene-, -CH2-O-(C=O)-, -R3-XC(=O)-Y-, -R3-XC(=S)-Y-, -R3-X-, -XC(=O)-Y-, -XC(=S)-Y- or -CH2-XC(=Z)-Y-CH2CH2-EC(=J)-F-, wherein R3 is C1-C3 alkylene; X, Y, E and F are independently selected from O, NH or S; Z and J are independently selected from O or S; L may be optionally substituted with hydroxyl, thiol, NH2, F, Cl, Br, I, -N3, methyl.

2. The conjugate according to claim 1, wherein the radiation-responsive group is a group that can chemically react with hydroxyl radicals generated by radiation and thereby fall off from the conjugate, or the radiation-responsive group is a group that can chemically react with hydroxyl radicals generated by radiation and thereby fall off from the conjugate together with the linker L.

3. The conjugate of claim 1, wherein the aryl or heteroaryl group is selected from phenyl, pyrrolyl, furanyl, thienyl, pyrazolyl and imidazolyl. The conjugate according to claim 1 , wherein the alkoxy group is methoxy, ethoxy, propoxy or butoxy.

5. The conjugate according to claim 1, wherein the aryl or heteroaryl group has no oxo, -C(=O)-, -C(=S)-, -CN, -SO3H, or -CF3 directly attached thereto. The conjugate according to claim 1 , wherein the substituted aryl group is a resorcinol group.

7. The conjugate of claim 1, wherein the aryl or heteroaryl group is substituted with one or more alkoxy, C1-C6 alkyl, hydroxy, C2-C6 alkenyl, Br or I. The conjugate according to claim 1 , wherein the C1-C6 alkyl substituted on the aryl or heteroaryl is methyl or ethyl.

9. The conjugate according to claim 1, wherein the aryl or heteroaryl is selected from phenyl, pyrrolyl, furanyl, thienyl, pyrazolyl and imidazolyl, the aryl is substituted with one or more methoxy, hydroxyl, Br or I; the heteroaryl is substituted with one or more methyl.

10. The conjugate according to any one of claims 1 to 9, wherein L is -CH2-X-(C=O)-Y-, -CH2-X-(C=S)-Y-, -CH2-X- or -X-(C=O)-Y-, wherein X and Y are independently selected from O, NH, S.

11. The conjugate according to claim 10, wherein L is -CH2-O-(C=O)-NH- or -O-(C=O)-NH-. The conjugate according to claim 1 , wherein D is a fluorescent imaging agent or a therapeutic agent for treating cancer.

13. A conjugate of general formula (I), P——L——D(I) Wherein P is a radiation responsive group containing an aromatic or heteroaromatic group; L is absent or is a divalent linking group; and D is an imaging agent or a therapeutic agent; wherein the aryl or heteroaryl is selected from phenyl, pyrrolyl, furanyl, thienyl, pyrazolyl and imidazolyl, wherein the phenyl is substituted with one or more amino, amine, -NHCOR1, phenyl, alkoxy, C1-C6 alkyl, hydroxyl, C2-C6 alkenyl, F, Cl, Br, I or -N3; ​​the heteroaryl is optionally substituted with one or more amino, amine, -NHCOR1, phenyl, alkoxy, C1-C6 alkyl, hydroxyl, C2-C6 alkenyl, F, Cl, Br, I or -N3, Wherein R1 is C1-C6 alkyl; Wherein when L is a divalent linking group, L is selected from -SS-, linear C1-C6-alkylene-, branched C3-C6-alkylene-, -CH2-O-(C=O)-, -R3-XC(=O)-Y-, -R3-XC(=S)-Y-, -R3-X-, -XC(=O)-Y-, -XC(=S)-Y- or -CH2-XC(=Z)-Y-CH2CH2-EC(=J)-F-, wherein R3 is C1-C3 alkylene; X, Y, E and F are independently selected from O, NH or S; Z and J are independently selected from O or S; L may be optionally substituted with hydroxyl, thiol, NH2, F, Cl, Br, I, -N3, methyl.

14. The conjugate according to claim 13, wherein the radiation-responsive group is a group that can chemically react with hydroxyl radicals generated by radiation and thereby fall off from the conjugate, or the radiation-responsive group is a group that can chemically react with hydroxyl radicals generated by radiation and thereby fall off from the conjugate together with the linker L. The conjugate according to claim 13 , wherein the amine group is a methylamine group or a dimethylamine group. The conjugate of claim 13 , wherein the alkoxy group is methoxy, ethoxy, propoxy or butoxy.

17. The conjugate according to claim 13, wherein the aryl or heteroaryl group has no oxo, -C(=O)-, -C(=S)-, -CN, -SO3H, or -CF3 directly attached thereto.

18. The conjugate of claim 13, wherein the substituted aryl group is a resorcinol group.

19. The conjugate of claim 13, wherein the heteroaryl group is optionally substituted with one or more amino, amine, -NHCOCH3, phenyl, alkoxy, C1-C6 alkyl, hydroxy, C2-C6 alkenyl, Br or I; and the phenyl group is substituted with one or more amino, amine, -NHCOCH3, phenyl, alkoxy, C1-C6 alkyl, hydroxy, C2-C6 alkenyl, Br or I.

20. The conjugate according to claim 13, wherein the C1-C6 alkyl optionally substituted on the heteroaryl is methyl or ethyl. The conjugate according to claim 13 , wherein the C1-C6 alkyl substituted on the phenyl group is a methyl group or an ethyl group.

22. The conjugate of claim 13, wherein the aryl or heteroaryl is selected from phenyl, pyrrolyl, furanyl, thienyl, pyrazolyl and imidazolyl, the heteroaryl is optionally substituted with one or more methyl groups; the aryl is substituted with one or more methoxy groups, hydroxyl groups, Br or I.

23. The conjugate according to any one of claims 13 to 22, wherein L is -CH2-X-(C=O)-Y-, -CH2-X-(C=S)-Y-, -CH2-X- or -X-(C=O)-Y-, wherein X and Y are independently selected from O, NH, S.

24. The conjugate according to claim 23, wherein L is -CH2-O-(C=O)-NH- or -O-(C=O)-NH-.

25. The conjugate of claim 13, wherein D is a fluorescent imaging agent or a therapeutic agent for treating cancer.

Images