Chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid as well as preparation method and application of chemical exchange saturation transfer magnetic resonance imaging probe

Abstract

The invention discloses a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronate, a preparation method and application, and relates to the technical field of chemical exchange saturation transfer magnetic resonance imaging. The key point of the technical scheme is as follows: the structure of the probe (Aen-FF-SSZ) consists of an alendronate group (Aen), a phenylalanine-phenylalanine (FF) self-assembly group and a salazosulfapyridine (SSZ) CEST MRI (Magnetic Resonance Imaging) unit. The CEST MRI molecular probe based on the alendronate is successfully developed, CEST MRI imaging and treatment of osteosarcoma are achieved, and the structure also has a wide clinical medical transformation prospect.

Description

Technical Field

[0001] The invention relates to the technical field of chemical exchange saturation transfer magnetic resonance imaging, and more specifically, to a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid, a preparation method and an application thereof. Background Art

[0002] At present, chemical exchange saturation transfer magnetic resonance imaging, as a "label-free", highly biocompatible, non-invasive, and deep tissue penetrating imaging method, has become a hot research direction in tumor diagnosis and treatment. Related studies have also verified that this imaging method has a certain clinical transformation capability. For example, chemical exchange saturation transfer imaging based on amide protons has become an important detection method for clinical brain tumors, inflammation, and neurodegenerative diseases. At the same time, the signal intensity in chemical exchange saturation transfer magnetic resonance imaging is affected by the physiological environment, such as temperature, pH value, etc. To a certain extent, the intensity of its signal reflects the relevant information of the physiological environment. There are related articles that apply this feature to cell transplantation and other detection aspects. Therefore, the study of contrast agents related to chemical exchange saturation transfer imaging has great prospects in achieving accurate diagnosis and building an integrated diagnosis and treatment platform.

[0003] Since the 1940s, bisphosphonate compounds have become commonly used bone protective agents in clinical practice due to their excellent bone targeting and bone adsorption capabilities. Today, they have been widely used in the design of various bone-targeted biological probes due to their strong adsorption capacity with hydroxyapatite in bone. After being adsorbed by bone, bisphosphonates will remain in it for a long time. When bone-related diseases (bone defects, osteosarcoma) occur, specifically activated osteoclasts will form acidic chambers to corrode bone. In the process of corroding bone, bisphosphonates adsorbed in the bone will be released and enriched in the acidic chambers. High concentrations of bisphosphonates will inhibit the related functions of osteoclasts and prevent further destruction of bone. Summary of the invention

[0004] The purpose of the present invention is to provide a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid, a preparation method and an application thereof, which are used for imaging of osteosarcoma.

[0005] The above technical objectives of the present invention are achieved through the following technical solutions: a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid, wherein the structure of the probe (Alen-FF-SSZ) consists of an alendronic acid group (Alen), a phenylalanine-phenylalanine (FF) self-assembly group and a sulfasalazine (SSZ) CEST MRI imaging unit, and its specific structure is as follows:

[0006]

[0007] By adopting the above technical solution, the probe achieves bone adsorption through the specific adsorption of the alendronic acid group and bone, and achieves shear release of the sulfasalazine group through the esterase overexpressed in the tumor tissue. The released sulfasalazine exchangeable proton group is exposed to turn on the CEST MRI signal.

[0008] The present invention further provides a method for preparing the above-mentioned chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid, which comprises the following five steps:

[0009] S1. Preparation of SSZ-COOH;

[0010] S2. Preparation of Trt-SH-FF-SSZ;

[0011] S3. Preparation of SH-FF-SSZ;

[0012] S4. Preparation of Alen-Male;

[0013] S5. Synthesis of Alen-FF-SSZ.

[0014] The present invention is further configured as follows: the specific steps for synthesizing SSZ-COOH in S1 are as follows:

[0015] (1) dissolving sulfasalazine in tetrahydrofuran, adding an excess of thionyl chloride thereto, stirring and reacting for one hour, and then spinning off the tetrahydrofuran and the excess thionyl chloride in the reaction container;

[0016] (2) Hydroxyacetic acid is dissolved in dichloromethane, and a small amount of triethylamine is added to mix and dissolve. The mixed solution is added dropwise to the product of the previous step after spin drying. After stirring for three hours, the product is separated and purified by column chromatography to obtain SSZ-COOH.

[0017] The present invention is further configured as follows: the specific steps of preparing Trt-SH-FF-SSZ in S2 are as follows:

[0018] (1) FF-SSZ was synthesized using the standard solid phase synthesis (SPPS) protocol and the product was purified by high performance liquid chromatography;

[0019] (2) dissolving the product of the previous step with triethyl methyl mercaptan, N-(benzotriazol-1-yl)-N,N,N',N'-tetramethylurethane hexafluorophosphate (HBTU), and N-hydroxybenzotriazole in dimethylformamide (dimethylformamide) and N,N-diethylethylamine, and adding N,N-diethylethylamine;

[0020] (3) stirring the reaction overnight and monitoring the reaction using an analytical high performance liquid chromatography;

[0021] (4) After the raw materials are completely reacted, the product Trt-SH-FF-SSZ is separated using a preparative high performance liquid chromatograph.

[0022] The present invention is further configured as follows: the specific steps of preparing SH-FF-SSZ in S3 are as follows: dissolving the product prepared in S2 in a dichloromethane solution containing 95% trifluoroacetic acid, removing the triphenylmethyl group in the product of the previous step to obtain the compound SH-FF-SSZ, and removing excess trifluoroacetic acid and dichloromethane by rotary evaporation to obtain an orange-yellow product solid SH-FF-SSZ.

[0023] The present invention is further configured as follows: the specific steps of preparing Alen-Male in S4 are as follows:

[0024] (1) dissolving alendronate sodium in water and adjusting the pH to 9 so that alendronate is completely dissolved in the water;

[0025] (2) 3-maleimidopropionic acid hydroxysuccinimide ester is suspended in 1,4-dioxane, and the 3-maleimidopropionic acid hydroxysuccinimide ester suspension is slowly added dropwise to the sodium alendronate aqueous solution, and during the addition, the pH of the mixed solution is adjusted to between 9 and 10 using 1 mM NaOH solution. After the addition is completed, the mixture is placed on a shaker and shaken for 15 minutes;

[0026] (3) After shaking, adjust the pH to 7 using 1 mM hydrochloric acid solution;

[0027] (4) The reaction solution after adjusting the pH is extracted with dichloromethane to remove the remaining 3-maleimidopropionic acid hydroxysuccinimide ester in the reaction, and the aqueous phase after extraction is collected, which contains Alen-Male.

[0028] The present invention is further configured as follows: the specific steps of synthesizing Alen-FF-SSZ in S5 are as follows:

[0029] (1) Dissolve SH-FF-SSZ solid in pyridine solution, and slowly add the aqueous solution containing Alen-Male into it several times;

[0030] (2) After the addition is completed, the mixture is placed on a shaker to react for 1 h, and the formation of the product is detected by high performance liquid chromatography;

[0031] (3) After the amount of the final product no longer increases, the final product Alen-FF-SSZ is separated by high performance liquid chromatography.

[0032] The present invention further provides an application of alendronic acid-based chemical exchange saturation transfer magnetic resonance imaging probe in the preparation of a drug for inhibiting the growth of osteosarcoma.

[0033] The present invention further provides an application of a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid in the preparation of a drug for treating osteosarcoma.

[0034] The present invention further provides an application of a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid in realizing CEST MRI imaging of osteosarcoma tissue.

[0035] In summary, the present invention has the following beneficial effects:

[0036] 1. The present invention provides a synthetic route for a chemical exchange saturation transfer magnetic resonance imaging probe that can be opened by esterase cleavage in a living body. The raw materials for synthesizing the probe are easily available, the cost is low, and the synthesis is convenient.

[0037] 2. This product provides a new synthetic route for linking bisphosphonates to peptide chains, greatly improving the yield of the synthesis.

[0038] 3. The present invention provides the use of a chemical exchange saturation transfer magnetic resonance imaging probe, which can perform magnetic resonance imaging of osteosarcoma cells in situ and has significant advantages over small molecules.

[0039] 4. The compound structure of the present invention achieves adsorption to bone through the bisphosphonic acid structure, thereby increasing the retention time of the compound in the tumor tissue, thereby increasing the imaging signal intensity and the therapeutic effect. BRIEF DESCRIPTION OF THE DRAWINGS

[0040] Figure 1 It is the synthetic route of Alen-FF-SSZ;

[0041] Figure 2 This is the high-resolution mass spectrum of Alen-FF-SSZ;

[0042] Figure 3 is the H NMR spectrum of Alen-FF-SSZ;

[0043] Figure 4 is the Z spectrum of Alen-FF-SSZ in Example 2 when the CEST MRI signal is turned on in vitro;

[0044] Figure 5 is a statistical graph of the CEST MRI signal turn-on of Alen-FF-SSZ in vitro in Example 2;

[0045] Figure 6 This is the synthetic route of FF-SSZ;

[0046] Figure 7 This is the high-resolution mass spectrum of the FF-SSZ compound;

[0047] Figure 8 is the H NMR spectrum of FF-SSZ;

[0048] Fig. 9 This is the flow chart of the animal CEST MRI imaging experiment in Example 3;

[0049] Fig.10 This is a pseudo-color image of the CEST MRI imaging of the animal in Example 3;

[0050] Fig.11 is the CEST MRI signal statistics of Alen-FF-SSZ animal imaging in Example 3;

[0051] Fig.12 is the CEST MRI signal statistics of FF-SSZ animal imaging in Example 3;

[0052] Fig.13 It is the statistical graph of CEST MRI image signals of SSZ animals in Example 3;

[0053] Fig.14 is the CEST MRI signal statistics of the three compounds in Example 3 at 2 h;

[0054] Fig.15 is the flow chart of the animal treatment experiment in Example 4;

[0055] Fig.16 is the anatomical diagram of each group of tumors in Example 4;

[0056] Fig.17 is a statistical graph of tumor size measurements in each group in Example 4;

[0057] Fig.18 is a statistical graph of tumor weight in Example 4;

[0058] Fig.19 is a Micro-CT scan of the mouse leg bone in Example 4;

[0059] Fig. 20 is a statistical graph of the bone volume of the mouse leg bone in Example 4;

[0060] Fig.21 is a statistical graph of the ratio of bone volume to tissue volume of the mouse leg bone in Example 4;

[0061] Fig. 22 is a statistical graph of mouse weight in Example 4;

[0062] Fig.23 This is a graph of mouse blood biochemical measurements in Example 4. DETAILED DESCRIPTION

[0063] The following is combined with Figure 1-23 The present invention is described in further detail.

[0064] Example 1. Specific synthesis process of the probe

[0065] 1. Take 500 mg of sulfasalazine in a round-bottom flask, add 5 mL of tetrahydrofuran to completely dissolve it, and add 2 ml of thionyl chloride solution to it, and stir to react for 1 hour. After the reaction is completed, use a rotary evaporator to remove the tetrahydrofuran liquid and excess thionyl chloride in the flask. Further dissolve 200 mg of glycolic acid in dichloromethane solution, add 500 μL of triethylamine, and then drop the mixed liquid into the above round-bottom flask several times to completely dissolve the solid in the original flask, and stir to react for 3 hours. Use thin layer chromatography to detect the reaction until the raw material reacts. Use column chromatography separation means (methanol: dichloromethane = 1:10) to separate the product to obtain SSZ-COOH.

[0066] 2. The compound FF-SSZ was synthesized by the standard solid phase peptide synthesis (SPPS) protocol (Solid phase peptide synthesis. I. The synthesis of a tetrapeptide; J. Am. Chem. Soc. 1963, 85 (14), 2149–2154, DOI: 10.1021 / ja00897a025). The product obtained by solid phase synthesis was dissolved in dimethylformamide with triethyl methyl mercaptan, N-(benzotriazole-1-yl)-N,N,N',N'-tetramethylurethane hexafluorophosphate (HBTU), and N-hydroxybenzotriazole, and N,N-diethylethylamine was added. The reaction was stirred overnight and the reaction was detected by analytical high performance liquid chromatography. After the raw materials were completely reacted, the product Trt-SH-FF-SSZ was separated by preparative high performance liquid chromatography.

[0067] 3. Dissolve the Trt-SH-FF-SSZ obtained in the previous step in a dichloromethane solution containing 95% trifluoroacetic acid, stir and react for 3 hours, and then use a rotary evaporator to remove trifluoroacetic acid and dichloromethane to obtain compound SH-FF-SSZ

[0068] 4. For the alendronic acid part, first dissolve alendronic acid sodium in water and adjust the pH to 9 so that alendronic acid is completely dissolved in water. Suspend 3-maleimidopropionic acid hydroxysuccinimide ester in 1,4-dioxane. Slowly drop the 3-maleimidopropionic acid hydroxysuccinimide ester suspension into the alendronic acid sodium aqueous solution. During the addition, use 1mM NaOH solution to adjust the pH of the mixed solution to between 9-10. After the addition is completed, place it on a shaker and shake for 15 minutes. After shaking, use 1mM hydrochloric acid solution to adjust the pH to 7. Extract the reaction solution with adjusted pH with dichloromethane to remove the remaining 3-maleimidopropionic acid hydroxysuccinimide ester in the reaction, and collect the extracted aqueous phase, which contains Alen-Male

[0069] 5. Dissolve the obtained SH-FF-SSZ in pyridine solution, and slowly add the aqueous solution containing Alen-Male several times. After the addition is completed, place it on a shaker to react for 1 hour, and use high performance liquid chromatography mass spectrometry to detect the formation of the product. When the amount of the final product no longer increases, use high performance liquid chromatography to separate the final product Alen-FF-SSZ. (M+Na=1232.1313), 1 HNMR(400MHz,DMSO)δ11.14(s,1H),8.49–8.37(m,3H),8.28–8.00(m,8H),7.88–7.82(m,1H), 7.48–7.16(m,12H),6.94(t,J=6.4Hz,1H),4.81(s,2H),4.67(td,J=8.9,4.5Hz,1H),4.55(q, J=7.9Hz,1H),4.05(q,J=4.3Hz,1H),3.64(t,J=7.7Hz,4H),3.08(p,J=6.7Hz,4H),2.87(m,5H ), 2.38 (t, J = 7.7Hz, 2H), 2.28 (t, J = 7.4Hz, 2H), 1.67 (p, J = 7.3Hz, 2H), 1.33 (q, J = 6.1Hz, 2H).

[0070] The synthetic route of the compound is Figure 1 As shown, Figure 2 This is the high-resolution mass spectrum of the final product. Figure 3 This is the hydrogen nuclear magnetic resonance spectrum of Alen-FF-SSZ.

[0071] Example 2. In vitro signal activation test example of the probe

[0072] 10 mM Alen-FF-SSZ and 500 U / mL esterase were dissolved in phosphate buffered saline (PBS, 10 mM, pH 7.4) containing 5% DMSO (v / v) and incubated overnight, and the CEST MRI signal of the mixed solution was measured. Figure 4-5 As shown, the results showed that the CEST MRI signal of Alen-FF-SSZ was significantly turned on after incubation with esterase.

[0073] Example 3. An example of using the probe in chemical exchange saturation transfer magnetic resonance imaging.

[0074] In order to reflect the advantages of the Alen group in Alen-FF-SSZ, a control compound: FF-SSZ was synthesized, and its structure is shown below:

[0075]

[0076] Figure 6 The synthetic route of FF-SSZ is: Figure 7 This is the high-resolution mass spectrum of FF-SSZ. Figure 8 This is the H NMR spectrum of FF-SSZ.

[0077] will contain 1×10 6 A suspension of human osteosarcoma cells 143B was injected into the tibial bone marrow cavity of 4-week-old Balb / c nude mice (each mouse weighed about 18-20 g) to construct an orthotopic osteosarcoma model in nude mice. 3 ), chemical exchange saturation transfer magnetic resonance imaging (CEST MRI, n = 4) was performed on the mice. After anesthesia, the mice were pre-scanned before drug injection; then, Alen-FF-SSZ (121 mg / kg) was injected orthotopically into the tumor tissue, and dynamic imaging was continued. The total scanning time was 8 hours. The experimental process is as follows Fig. 9 After scanning, the PLOF method was used to process the acquired signal data and perform graphical analysis. In the same way, FF-SSZ (75 mg / kg) and sulfasalazine solution (39.9 mg / kg) were injected into the orthotopic osteosarcoma model mice, and the scans were performed and the data processing was completed.

[0078] The results are as follows Figure 10-14As shown. The data showed that the CEST signal of the Alen-FF-SSZ and FF-SSZ groups continued to increase from 0 to 2 hours (Alen-FF-SSZ: 1.02%; FF-SSZ: 0.6%) and reached a peak at 2 hours; while the sulfasalazine group showed the highest signal intensity (1.49%) at the beginning of the experiment. This result verifies the mechanism that the Alen-FF-SSZ and FF-SSZ groups release SSZ through esterase-mediated enzymatic cleavage in vivo, thereby triggering the activation of the CEST signal.

[0079] In addition, compared with the sulfasalazine group, the signals of the Alen-FF-SSZ and FF-SSZ groups showed longer duration, indicating that the nanostructured smaller molecules have better retention ability. Among the two groups of nanostructured probes, the Alen-FF-SSZ group showed higher signal intensity at 2 hours, proving that the bone-targeted adsorption of the Alen group can retain more compounds in the tumor area.

[0080] In summary, the experimental results show that Alen-FF-SSZ can achieve targeted diagnosis of osteosarcoma tissue through precise CEST imaging, demonstrating its great potential in tumor imaging.

[0081] Example 4. An example of using the probe in the treatment of osteosarcoma.

[0082] will contain 1×10 6 Human osteosarcoma cell 143B suspension was injected into the tibial bone marrow cavity of 4-week-old Balb / c nude mice (each mouse weighed about 18-20 g) to construct an orthotopic osteosarcoma model in nude mice. 3 ), the mice were randomly divided into 4 groups (n=5 in each group) for treatment experiments. The treatment process was as follows Fig.15 As shown. Each group of mice was injected intratumorally every two days with: 18.15mg / kg Alen-FF-SSZ, 11.25mg / kg FF-SSZ, 5.98mg / kg SSZ, and PBS, respectively, for 14 consecutive days, and the tumor size and mouse body weight were monitored. On the 14th day, euthanasia was performed, and the mice were dissected to collect tumors and major organs (heart, liver, spleen, lung, kidney), and blood samples were collected. After the dissection, the tumor was weighed and photographed. The experimental results are shown in Figure 16-18 The results showed that the Alen-FF-SSZ group had the most significant therapeutic effect, and the tumor size was significantly smaller than that of the other three groups. The FF-SSZ group also showed a certain therapeutic effect, but it was lower than the Alen-FF-SSZ group. This may be due to the bone adsorption effect brought by the Alen group and the characteristics of the nanoparticle size, which caused more compounds to remain in the tumor tissue, resulting in differences in therapeutic effects.

[0083] Subsequently, Micro-CT scans were performed on the dissected mouse legs to detect their bone conditions. The experimental results are as follows: Figure 19-21 The results showed that the bone volume and the ratio of bone volume to tissue volume of the tibia of mice in the Alen-FF-SSZ group were the highest, indicating that the leg bones of this group of mice were well preserved, further proving that Alen-FF-SSZ can not only inhibit the growth of osteosarcoma, but also effectively prevent cancer cells from eroding bones. In addition, Figure 22-23 The experimental results shown in the figure show that the weight of mice in the Alen-FF-SSZ group did not change significantly during the treatment, and there was no abnormality in the blood biochemical indicators after treatment, proving that Alen-FF-SSZ had no obvious side effects on mice. In summary, the experimental results show that Alen-FF-SSZ can effectively treat osteosarcoma and protect bone.

[0084] This specific embodiment is merely an explanation of the present invention and is not a limitation of the present invention. After reading this specification, those skilled in the art may make non-creative modifications to the present embodiment as needed. However, as long as they are within the scope of the claims of the present invention, they are protected by the patent law.

Claims

1. A chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid, characterized in that: The structure of the probe (Alen-FF-SSZ) consists of an alendronic acid group (Alen), a phenylalanine-phenylalanine (FF) self-assembly group and a sulfasalazine (SSZ) CEST MRI imaging unit, and its specific structure is shown below:

2. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 1, characterized in that: It includes the following five steps: S1. Preparation of SSZ-COOH; S2. Preparation of Trt-SH-FF-SSZ; S3. Preparation of SH-FF-SSZ; S4. Preparation of Alen-Male; S5. Synthesis of Alen-FF-SSZ.

3. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 2, characterized in that: The specific steps of synthesizing SSZ-COOH in S1 are as follows: (1) dissolving sulfasalazine in tetrahydrofuran, adding an excess of thionyl chloride thereto, stirring and reacting for one hour, and then spinning off the tetrahydrofuran and the excess thionyl chloride in the reaction container; (2) Hydroxyacetic acid is dissolved in dichloromethane, and a small amount of triethylamine is added to mix and dissolve. The mixed solution is added dropwise to the product of the previous step after spin drying. After stirring for three hours, the product is separated and purified by column chromatography to obtain SSZ-COOH.

4. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 3, characterized in that: The specific steps of preparing Trt-SH-FF-SSZ in S2 are as follows: (1) FF-SSZ was synthesized using the standard solid phase synthesis (SPPS) protocol and the product was purified by high performance liquid chromatography; (2) dissolving the product of the previous step with triethyl methyl mercaptan, N-(benzotriazol-1-yl)-N,N,N',N'-tetramethylurethane hexafluorophosphate (HBTU), and N-hydroxybenzotriazole in dimethylformamide (dimethylformamide) and N,N-diethylethylamine, and adding N,N-diethylethylamine; (3) stirring the reaction overnight and monitoring the reaction using an analytical high performance liquid chromatography; (4) After the raw materials are completely reacted, the product Trt-SH-FF-SSZ is separated using a preparative high performance liquid chromatograph.

5. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 4, characterized in that: The specific steps of preparing SH-FF-SSZ in S3 are as follows: dissolving the product prepared in S2 in a dichloromethane solution containing 95% trifluoroacetic acid, removing the triphenylmethyl group in the product of the previous step to obtain the compound SH-FF-SSZ, and removing excess trifluoroacetic acid and dichloromethane by rotary evaporation to obtain an orange-yellow product solid SH-FF-SSZ.

6. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 5, characterized in that: The specific steps of preparing Alen-Male in S4 are as follows: (1) dissolving alendronate sodium in water and adjusting the pH to 9 so that alendronate is completely dissolved in the water; (2) 3-maleimidopropionic acid hydroxysuccinimide ester is suspended in 1,4-dioxane, and the 3-maleimidopropionic acid hydroxysuccinimide ester suspension is slowly added dropwise to the sodium alendronate aqueous solution, and during the addition, the pH of the mixed solution is adjusted to between 9 and 10 using 1 mM NaOH solution. After the addition is completed, the mixture is placed on a shaker and shaken for 15 minutes; (3) After shaking, adjust the pH to 7 using 1 mM hydrochloric acid solution; (4) The reaction solution after adjusting the pH is extracted with dichloromethane to remove the remaining 3-maleimidopropionic acid hydroxysuccinimide ester in the reaction, and the aqueous phase after extraction is collected, which contains Alen-Male.

7. The method for preparing a chemical exchange saturation transfer magnetic resonance imaging probe based on alendronic acid according to claim 6, characterized in that: The specific steps of synthesizing Alen-FF-SSZ in S5 are as follows: (1) Dissolve SH-FF-SSZ solid in pyridine solution, and slowly add the aqueous solution containing Alen-Male into it several times; (2) After the addition is completed, the mixture is placed on a shaker to react for 1 h, and the formation of the product is detected by high performance liquid chromatography; (3) After the amount of the final product no longer increases, the final product Alen-FF-SSZ is separated by high performance liquid chromatography.

8. Use of the alendronic acid-based chemical exchange saturation transfer magnetic resonance imaging probe according to claim 1 in the preparation of a drug for inhibiting the growth of osteosarcoma.

9. Use of the alendronic acid-based chemical exchange saturation transfer magnetic resonance imaging probe according to claim 1 in the preparation of a drug for treating osteosarcoma.

10. Use of the alendronic acid-based chemical exchange saturation transfer magnetic resonance imaging probe according to claim 1 in realizing CEST MRI imaging of osteosarcoma tissue.

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