Degradable spherical nucleic acid for delivering protein drug across blood-brain barrier as well as preparation method and application of degradable spherical nucleic acid

Abstract

The invention belongs to the field of research and development of tumor drugs, and discloses a degradable spherical nucleic acid for delivering a protein drug across a blood-brain barrier, aiming at the urgent problems that the protein drug is difficult to cross the blood-brain barrier, poor in targeting property and the like when being delivered to brain tumors, and the invention discloses a degradable spherical nucleic acid for delivering a protein drug across the blood-brain barrier. The surface of the inner core is connected with a nucleic acid aptamer through a covalent bond to form a spherical nucleic acid structure. Protein drugs are delivered through the spherical nucleic acid structure, and benefited from the scavenger receptor mediated cell transfer effect, the main barrier, namely the blood-brain barrier, in brain delivery can be effectively overcome; meanwhile, due to packaging of the protein medicine, the protein medicine is prevented from being released in blood in advance, the possibility that the protein medicine is cleared away in advance is reduced, and the risk that the protein medicine causes adverse immune response is also reduced. Moreover, according to the degradable spherical nucleic acid for delivering the protein medicine across the blood brain barrier, the delivery variety of the spherical nucleic acid is expanded, the retention risk of the spherical nucleic acid is reduced through the idea of responsive degradation, and the degradable spherical nucleic acid has a good application prospect.

Description

Technical Field

[0001] The present invention belongs to the field of nano drug carriers, and particularly relates to a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier, its preparation method and application. Background Art

[0002] The blood-brain barrier refers to the barrier between the plasma and brain cells formed by the cerebral capillary wall and glial cells and the barrier between the plasma and cerebrospinal fluid formed by the choroid plexus. While protecting the biological brain tissue from the influence of various harmful substances, it also greatly reduces the efficiency of drug delivery to the brain to play its role. According to statistics, more than 98% of small molecule drugs and almost all macromolecular therapeutic agents cannot cross the blood-brain barrier. Among them, protein drugs are particularly difficult to deliver to the brain due to their large molecular weight, easy clearance, and problems of immunogenic exposure.

[0003] Spherical nucleic acid is a structure that has received attention in recent years. Its characteristic is a three-dimensional structure formed by high-density and highly oriented nucleic acids. Thanks to this structure, it can cross the blood-brain barrier through the transcytosis of scavenger receptors and improve the delivery efficiency. The structure of spherical nucleic acid mainly consists of a core and a high-density nucleic acid shell, and the types of its cores are mainly: inorganic nanoparticle cores, organic micelle cores, semiconductor quantum dots, etc. Most of these cores do not have the property of biodegradation. The existing spherical nucleic acid structures are mainly used to deliver nucleic acid drugs. Some spherical nucleic acid designs can deliver small molecule drugs or fluorescent probes, but there are still certain difficulties in the safe delivery of protein drugs. For example, it is difficult to load protein drugs due to their large molecular weight, the carrier needs to effectively protect protein drugs, the existence of biological barriers, and it is difficult to achieve precise release at the target site, etc. This limits the application of existing spherical nucleic acids as protein drug delivery carriers. Summary of the Invention

[0004] The technical problem to be solved by the present invention is to provide a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier in view of the above deficiencies in the prior art, which has the advantages of biocompatibility, efficient crossing of the blood-brain barrier, and responsive degradation, and can effectively deliver protein drugs to brain tumors.

[0005] The technical solution adopted by the present invention to solve the above problems is as follows:

[0006] A degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier is a spherical nucleic acid structure formed by using a degradable nanosilica sphere wrapped with protein drugs as the core, and nucleic acid aptamers are covalently connected to the surface of the core.

[0007] According to the above scheme, after the degradable nanosilica spheres encapsulating the protein drug are modified by amination, they are covalently linked to the nucleic acid aptamer with a carboxyl group. Further, about 500 - 2000 nucleic acids are encapsulated in each degradable nanosilica sphere, achieving high-density connection of nucleic acids and forming a spherical nucleic acid structure.

[0008] According to the above scheme, the nucleic acid aptamer is a nucleic acid aptamer capable of targeting tumor cells and has a carboxyl group at the 5'-end. Preferably, it is the nucleic acid aptamer AS1411, and the sequence is 5'-COOH-GGTGGTGGTGGTTGTGGTGGTGGTGG-3'.

[0009] The preparation method of the degradable spherical nucleic acid for trans-blood-brain-barrier delivery of protein drugs includes the following steps:

[0010] 1) After fully mixing the aqueous solution of the protein drug with tetraethyl orthosilicate (TEOS) and bis-[3-(triethoxysilyl)propyl]-tetrasulfide (BTEPDS), slowly drop it into the organic phase solution to form a microemulsion system, and carry out a hydrolysis reaction under the action of ammonia water to obtain degradable nanosilica spheres encapsulating the protein drug;

[0011] 2) Disperse the degradable nanosilica spheres encapsulating the protein drug obtained in step 1) in absolute ethanol, add 3-aminopropyltriethoxysilane (APTES) for surface amination modification to obtain aminated degradable nanosilica spheres;

[0012] 3) Dissolve the nucleic acid aptamer with a carboxyl group in a buffer solution, add N-hydroxysuccinimide (NHS) and 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC) to activate its carboxyl group to obtain an activated nucleic acid aptamer solution;

[0013] 4) Disperse the aminated degradable silica spheres obtained in step 2) in absolute ethanol, add the activated nucleic acid aptamer solution obtained in step 3), and after reacting for 6 - 12 h, obtain degradable spherical nucleic acids encapsulating the protein drug, that is, the degradable spherical nucleic acids for trans-blood-brain-barrier delivery of protein drugs.

[0014] According to the above scheme, in step 1), the concentration of the aqueous solution of the protein drug is 1 - 4 mg / mL; the volume ratio of the aqueous solution of the protein drug to TEOS, BTEPDS, and ammonia water is (5 - 7):(0.6 - 1):(1 - 1.4):1; among them, the concentration of ammonia water is 20 - 35%.

[0015] According to the above scheme, in step 1), the organic phase solution is composed of Triton X-100, hexanol, and hexane mixed in a volume ratio of 1:(0.8 - 1.2):(3 - 5), and the volume ratio of the organic phase solution to the aqueous solution of the protein drug is (30 - 45):1.

[0016] According to the above - mentioned scheme, in step 1), the conditions for the hydrolysis reaction are: room temperature, and the reaction time is 8 - 15 h.

[0017] According to the above - mentioned scheme, in step 2), the concentration of the degradable nanosilica spheres encapsulating the protein drug in absolute ethanol is 0.2 - 1 mg / mL; the mass ratio of the degradable nanosilica spheres encapsulating the protein drug to APTES is 1:(4 - 6).

[0018] According to the above - mentioned scheme, in step 2), the conditions for the amination modification are: room temperature, and the reaction time is 6 - 10 h.

[0019] According to the above - mentioned scheme, in step 3), the pH of the buffer solution is in the range of 4.5 - 7.2. NHS and EDC are respectively prepared into solutions with a concentration of 5 - 15 mM in advance using the buffer solution, and the nucleic acid aptamer with a carboxyl group is prepared into a solution with a concentration of 50 - 150 μM in advance using the buffer solution. During activation, the final concentrations of NHS and EDC, and the nucleic acid aptamer with a carboxyl group in the activation system of this step are 2 - 5 mM, 2 - 5 mM, and 5 - 40 μM respectively. The activation is carried out at room temperature, and the activation time is 10 - 30 min. Among them, the buffer solution can be PBS, HEPES buffer solution, etc.

[0020] According to the above - mentioned scheme, in step 4), the concentration of the aminated degradable silica spheres in absolute ethanol is 0.5 - 1.5 mg / mL; 5 - 15 μL of the activated nucleic acid aptamer solution is added to each milligram of the aminated degradable silica spheres, and the concentration of the activated nucleic acid aptamer solution is preferably 20 - 50 μM.

[0021] Application of the degradable spherical nucleic acid for delivering protein drugs across the blood - brain barrier in the delivery of brain glioma - related protein drugs. In the present invention, the protein drugs mainly include serum albumin, nuclease, antigen polypeptide, interleukin, tumor necrosis factor, interferon, rituximab, trastuzumab, etc. Specifically, the present invention utilizes the special physiological environment inside tumor cells to achieve the responsive degradation of nanosilica spheres, and the protein drugs play a role inside tumor cells.

[0022] The technical concept of the present invention is as follows:

[0023] First, the present invention encapsulates protein drugs in degradable nanosilica spheres through the microemulsion method. The reaction is carried out under mild conditions of normal temperature and stirring, avoiding the inactivation of protein drugs. Then, it is modified to have a large number of amino groups on its surface, and this modification process will not affect its degradation. Furthermore, a large number of nucleic acid aptamers are connected to its surface through covalent bonds to form a high-density nucleic acid outer layer. Although a small amount of nucleic acid aptamers can help it target tumors, the high-density nucleic acid modification can form a spherical nucleic acid structure, which can cross the blood-brain barrier through the transcytosis pathway mediated by scavenger receptors and deliver protein drugs to tumor cells, having the advantages of biocompatibility and efficient crossing of the blood-brain barrier.

[0024] The concentration of glutathione (GSH) in tumor cells is more than 1000 times that of extracellular glutathione concentration and more than 4 times that of normal cells. When the degradable spherical nucleic acid structure reaches tumor cells, it degrades under the action of high-concentration glutathione in tumor cells and releases the internal protein drugs (high-concentration glutathione will break the disulfide bonds from BTEPDS), thus achieving responsive degradation in tumor cells and further realizing the safe targeted delivery of protein drugs.

[0025] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0026] First, the present invention delivers protein drugs through a spherical nucleic acid structure. Thanks to the transcytosis mediated by scavenger receptors, it can effectively overcome the blood-brain barrier, which is the main obstacle in brain delivery. At the same time, the encapsulation of protein drugs avoids their premature release in the blood, reduces the possibility of their premature clearance, and also reduces the risk of adverse immune reactions caused by protein drugs.

[0027] Second, from the perspective of spherical nucleic acids, traditional spherical nucleic acids often use non-degradable nanoparticles as the core, such as gold nanoparticles, or form the core through the self-assembly of polymer micelles. This not only limits the types of drugs that can be delivered as a delivery carrier but also greatly increases the risk of their retention in the brain. The degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier proposed by the present invention not only expands the delivery types of spherical nucleic acids but also reduces its retention risk through the concept of responsive degradation, having good application prospects. Description of the Drawings

[0028] Figure 1 Transmission electron micrograph of the degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier prepared in the example.

[0029] Figure 2 High-resolution electron micrograph and mapping diagram of the degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier prepared in the example.

[0030] Figure 3 XPS energy spectrum of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs prepared in the example.

[0031] Figure 4 Zeta potential changes of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs prepared in the example and during the modification process.

[0032] Figure 5 Degradation performance of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs prepared in the example. Among them, a is the transmission electron micrograph of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs after being treated with GSH for different times; b is the curve of the change in the protein concentration measured in the supernatant after the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs encapsulating the protein is treated with GSH for different times.

[0033] Figure 6 Cross-blood-brain-barrier performance of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs prepared in the example in an in vitro model. Among them, a is the permeability of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs in the in vitro model; b is the situation of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs being taken up by mouse glioma cells after crossing the blood-brain barrier studied by confocal microscopy in the in vitro model.

[0034] Figure 7 Enrichment of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs prepared in the example in vivo. Detailed implementation mode

[0035] To better understand the present invention, the content of the present invention will be further clarified below in conjunction with the examples, but the present invention is not limited to the following examples only.

[0036] Example 1

[0037] Preparation of the degradable spherical nucleic acid for cross-blood-brain-barrier delivery of protein drugs:

[0038] (1) Disperse 0.75 mg of bovine serum albumin (BSA) powder in 300 μL of deionized water, add 40 μL of TEOS and 60 μL of BTEPDS and mix well. Then, drop the above mixed solution into the organic phase solution composed of 1.8 mL of Triton X-100, 1.8 mL of n-hexanol and 7.5 mL of cyclohexane. During the dropping process, the organic phase solution needs to be stirred. After the mixed solution forms a uniform emulsion, add 50 μL of ammonia water to provide the alkaline environment required for the reaction (the concentration of ammonia water is 30%), react at room temperature for 12 h, then add acetone to break the emulsion, centrifuge to collect the precipitate and wash it to obtain the degradable nanosilica spheres encapsulating the protein drug.

[0039] (2) 7.5 mg of the degradable nanosilica spheres encapsulating the protein drug were dispersed in 7.5 mL of absolute ethanol. 40 μL of 3-aminopropyltriethoxysilane (APTES) was added, and after reacting for 8 h, the product was collected by centrifugation, washed, and freeze-dried to obtain amino-functionalized degradable nanosilica spheres.

[0040] (3) 1 mg of the amino-functionalized degradable nanosilica spheres was weighed and dispersed in 1 mL of absolute ethanol to obtain an amino-functionalized nanosilica sphere dispersion. Meanwhile, 10 μL of a solution of the nucleic acid aptamer AS1411 modified with a 5'-carboxyl group (100 μM, PBS, pH = 6.8) was added to 10 μL each of an EDC solution (10 mM, PBS, pH = 6.8) and an NHS solution (10 mM, PBS, pH = 6.8). After activation for 20 min, the activated nucleic acid aptamer solution was added to the above-mentioned amino-functionalized nanosilica sphere dispersion. After reacting for 8 h, the product was collected by centrifugation, which was the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier.

[0041] Characterization of the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier:

[0042] As Figure 1 Transmission electron microscope (TEM) images showed that the degradable spherical nucleic acid for delivering the protein drug prepared above had a spherical structure, and the size mainly concentrated in the range of 50 - 100 nm; as Figure 2 shown in the mapping diagram and Figure 3 from the XPS analysis, the elemental composition of the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier was mainly Si, C, O, N, S, etc., indirectly proving the successful preparation of the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier.

[0043] As Figure 4 The Zeta potential of the product during the synthesis of the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier was analyzed using a dynamic light scattering analyzer. As shown in the figure, the surface potential of the silica spheres encapsulating the protein after amino-functionalization increased, and with the successful modification of the nucleic acid, the surface potential decreased, which proved the successful modification of each step of the degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier.

[0044] As Figure 5 a The prepared degradable spherical nucleic acid for delivering the protein drug across the blood-brain barrier was dispersed in a 10 mM glutathione solution to make the concentration of the degradable spherical nucleic acid 1 mg / mL. After treating for different times (i.e., observing the phenomenon after stirring for different times after mixing), the morphological changes were observed by transmission electron microscopy. In the figure, it can be observed that the degradable spherical nucleic acid degraded within 24 h under the action of glutathione, and the morphology gradually became blurred.

[0045] Disperse the degradable spherical nucleic acid prepared above in a 10 mM glutathione solution so that the concentration of the degradable spherical nucleic acid is 1 mg / mL. After treatment for 12, 24, and 48 h respectively, collect the supernatant, and use a 30 kd ultrafiltration tube to remove the residual GSH and enrich the released protein. As Figure 5 shown in b, after treatment with glutathione, use a protein concentration kit to measure the protein concentration in the supernatant. First, combine Figure 5 b. It can be found that under the action of glutathione, as the morphology of the spherical nucleic acid gradually disintegrates, the protein encapsulated inside it can be successfully released. The results show that the degradable spherical nucleic acid for delivering protein drugs prepared in this example can effectively release the protein encapsulated inside it under the action of glutathione.

[0046] Example 2

[0047] A preparation method of a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier, the specific steps are as follows:

[0048] (1) Replace the BSA powder with an equal mass of FITC-labeled bovine serum albumin (FITC-BSA) powder, and the others are the same as in Example 1;

[0049] (2) The same as in Example 1;

[0050] (3) For five groups of experiments, the volumes of the nucleic acid aptamer AS1411 solution (100 uM) modified with carboxyl at the 5' end are 10, 5, 2.5, 0.5, and 0 uL respectively, and the others are the same as in Example 1. The final products are degradable spherical nucleic acids for delivering protein drugs with different nucleic acid modification amounts. By comparing the total number of successfully modified nucleic acid chains measured by fluorescence method with the total number of nanoparticles obtained by nanoparticle tracking analysis, the modification amount of each nanosilica sphere is calculated to be approximately 2000, 1000, 500, 100, and 0 nucleic acid aptamers in turn, and they are named FB-SNA@2000, FB-SNA@1000, FB-SNA@500, FB-SNA@100, and FB-Si@0 in turn.

[0051] Study on the performance of the degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier:

[0052] Culture mouse glioma cells GL261 in the lower chamber and mouse cerebrovascular endothelial cells bend.3 in the upper chamber to construct a Transwell model. Add 50 ug of degradable spherical nucleic acids with different nucleic acid modification amounts to the upper chamber respectively. After transferring the upper chamber to the cell-free lower chamber and incubating for 8 h, take the liquid in the lower chamber and measure the transmittance of the degradable spherical nucleic acid by fluorescence intensity, as Figure 6As shown in Figure a, in an in vitro Transwell model, the modification of the nucleic acid aptamer increased the efficiency of the nano-silica spheres crossing the blood-brain barrier from approximately 10% to approximately 30%, demonstrating that the modification of the nucleic acid aptamer significantly improved its permeability through the in vitro blood-brain barrier model.

[0053] The mouse glioma cell line GL261 was cultured in the lower chamber, and the mouse cerebral vascular endothelial cell line bend.3 was cultured in the upper chamber to construct a Transwell model. 50 μg of degradable spherical nucleic acids with different nucleic acid modification amounts were added to the upper chamber respectively. After incubation for 8 h, a confocal microscope was used to observe the uptake of the degradable spherical nucleic acids by the tumor cells GL261 in the lower chamber. As Figure 6 shown in Figure b, with the increase in the nucleic acid modification amount, the green fluorescence entering the tumor cells was significantly increased compared with the control group. When the nucleic acid modification amount reached 1000 / nano-silica sphere, the change in green fluorescence was not significant with the increase in the nucleic acid modification amount, indicating that the modification of the nucleic acid aptamer enabled the spherical nucleic acids to effectively cross the in vitro blood-brain barrier model and be effectively taken up by the tumor cells. The nucleic acid modification amount is preferably 500 - 2000.

[0054] In addition, degradable spherical nucleic acids with different nucleic acid modification amounts were dispersed in PBS buffer (pH = 7.4) and injected into mice with glioma through the tail vein (100 mg / kg, calculated based on the mass of the spherical nucleic acids). After 12 h, the mice were dissected, and the fluorescence intensity was analyzed using in vivo imaging in region 1. As Figure 7 shown, a large amount of fluorescence was observed to be enriched at the tumor site, demonstrating that the degradable spherical nucleic acids for delivering protein drugs across the blood-brain barrier in vivo can also effectively cross the blood-brain barrier and target the tumor site.

[0055] The above are only the preferred embodiments of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the inventive concept of the present invention, several improvements and modifications can be made, and these all belong to the protection scope of the present invention.

Claims

1. A degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier, characterized in that: The degradable nano-silicon spheres encapsulating protein drugs are used as the inner core, and nucleic acid aptamers are covalently linked to the inner core surface to form a spherical nucleic acid structure; the nucleic acid aptamers are nucleic acid aptamers that can target tumor cells.

2. The degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 1, characterized in that: The degradable nano-silicon spheres encapsulating the protein drug are modified by amino group and then covalently bonded with the nucleic acid aptamer with carboxyl group.

3. The degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 1, characterized in that: The size of the spherical nucleic acid is 50-120nm; each degradable nano-silicon sphere contains 500-2000 nucleic acids.

4. The degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 1, characterized in that: The nucleic acid aptamer is AS1411 with a carboxyl group at the 5' end; the protein drugs include serum albumin, nuclease, antigenic polypeptide, interleukin, tumor necrosis factor, interferon, rituximab, and trastuzumab.

5. The method for preparing a degradable spherical nucleic acid for delivering a protein drug across the blood-brain barrier according to any one of claims 1 to 4, characterized in that: The following steps are involved: 1) After the protein drug aqueous solution is fully mixed with ethyl orthosilicate and bis-[3-(triethoxysilyl)propyl]-tetrasulfide, it is slowly added dropwise to the organic phase solution to form a microemulsion system, and a hydrolysis reaction occurs under the action of ammonia water to obtain degradable nano-silicon spheres encapsulating the protein drug; 2) dispersing the degradable nano-silicon spheres encapsulating the protein drug obtained in step 1) in alcohol, adding 3-aminopropyltriethoxysilane to carry out surface amino modification, and obtaining amino-modified degradable nano-silicon spheres; 3) dissolving the nucleic acid aptamer with carboxyl group in a buffer solution, adding NHS and EDC to activate its carboxyl group, and obtaining an activated nucleic acid aptamer solution; 4) dispersing the amino-modified degradable silica spheres obtained in step 2) in alcohol, adding the activated nucleic acid aptamer solution obtained in step 3), and reacting to obtain degradable spherical nucleic acids encapsulating protein drugs, i.e., degradable spherical nucleic acids for delivering protein drugs across the blood-brain barrier.

6. The method for preparing a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 5, characterized in that: In step 1), the concentration of the protein drug aqueous solution is 1-4 mg / mL, and the volume ratio of the protein drug aqueous solution to ethyl silicate, bis-[3-(triethoxysilyl)propyl]-tetrasulfide and ammonia water is (5-7):(0.6-1):(1-1.4):

1.

7. The method for preparing a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 5, characterized in that: In step 1), the organic phase solution is mixed by Triton X-100, hexanol and hexane in a volume ratio of 1: (0.8-1.2): (3-5), and the volume ratio of the organic phase solution to the protein drug aqueous solution is (30-45): 1; in step 2), the concentration of the degradable nano-silicon spheres encapsulating the protein drug in alcohol is 0.5-1.5 mg / mL, and the mass ratio of the degradable nano-silicon spheres encapsulating the protein drug and 3-aminopropyltriethoxysilane is 1: (4-6).

8. The method for preparing a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 5, characterized in that: In step 1), the conditions for the hydrolysis reaction are: room temperature, reaction time 8-15h; in step 2), the reaction conditions for the amino modification are: room temperature, reaction time 6-10h; in step 3), activation is carried out at room temperature, the activation time is 10-30min; in step 4), room temperature, the reaction time is 6-12h.

9. The method for preparing a degradable spherical nucleic acid for delivering protein drugs across the blood-brain barrier according to claim 5, characterized in that: In step 3), the pH of the buffer solution is in the range of 4.5-7.2, and the final concentrations of NHS, EDC and the nucleic acid aptamer with a carboxyl group in the activation system of this step are 2-5mM, 2-5mM and 5-40uM respectively; in step 4), the concentration of the amino-degradable silica spheres in alcohol is 0.5-1.5mg / mL; 5-15uL of the activated nucleic acid aptamer solution is added per milligram of the amino-degradable silica spheres, and the concentration of the activated nucleic acid aptamer solution is 20-50uM.

10. Use of the degradable spherical nucleic acid according to claim 1 in tumor protein drug delivery.

Images