A nanoantibody targeting CD38, a pharmaceutical composition and its application

Abstract

The present invention provides a nano-antibody targeting CD38, a pharmaceutical composition, and its application, relating to the field of biomedicine technology. The nano-antibody targeting CD38 is any one of a first antibody and a second antibody, and both include three complementary determining regions; the sequences of the complementary determining regions CDR1 to 3 of the first antibody are shown in SEQ.ID NO.1, SEQ.ID NO.3, and SEQ.ID NO.5; the sequences of the complementary determining regions CDR1 to 3 of the second antibody are shown in SEQ.ID NO.2, SEQ.ID NO.4, and SEQ.ID NO.6. The nano-antibody has a high affinity for both purified and cell surface CD38 proteins, providing an effective new method for targeting CD38 for anti-tumor intervention; and molecular probes and other detection reagents can be prepared based on the nano-antibody, providing a new approach for tumor-specific imaging and diagnosis by specifically binding to CD38 on the tumor surface.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and more specifically to a CD38-targeting nanoantibody, a pharmaceutical composition, and applications thereof. Background Art

[0002] Targeted cancer therapy is a method of treating tumors by specifically targeting molecules or signaling pathways that play a crucial role in tumor development and progression. Targeted therapies tailored to the specific molecular phenotype of a tumor are increasingly becoming the preferred approach for cancer treatment. Targeted cancer drugs can be broadly categorized into two main categories: monoclonal antibodies and small molecule compounds. With the advancement of research, new targets continue to emerge, leading to numerous breakthroughs in anti-cancer drug development, with the number of targeted cancer drugs increasing at a rate of 5 to 6 per year. However, the effectiveness of targeted therapies is limited by the heterogeneity of tumor cells, resulting in significant differences in the response of different patients to the same drug. Furthermore, tumor cells can develop drug resistance through various mechanisms, resulting in reduced therapeutic efficacy. Therefore, the current development of targeted drugs remains insufficient, limiting the widespread application of existing treatments and preventing the vast majority of patients from benefiting from them. Therefore, there is an urgent need to discover new molecular targets and develop more effective molecularly targeted therapy strategies to improve the precision and effectiveness of treatment and reduce the treatment burden on patients.

[0003] CD38 (Cluster of Differentiation 38) is a type II transmembrane glycoprotein with a molecular weight of approximately 46kD. In 1980, CD38 was first discovered by scientists such as EL Reinherz and SF Schlossman as a specific marker for T cell activation. The CD38 protein sequence includes a short cytoplasmic tail at the N-terminus, a single transmembrane domain, and a longer extracellular region at the C-terminus. Current studies have shown that CD38 protein is a bifunctional extracellular enzyme with cyclase and hydrolase activities, and is involved in nucleotide metabolism. CD38 uses NAD + It serves as a substrate to form nucleotide metabolites such as cyclic ADP ribose (cADPR), which is an effective second messenger that regulates Ca2+ in the cytoplasm. 2+ Mobilization of CD38 and activation of signaling pathways that control various biological processes. Studies have also found that CD38 can degrade ATP and NAD + , cADPR, AMP, thereby producing adenosine, inducing an inhibitory immune microenvironment. In short, the activity of CD38 is crucial for maintaining NAD + The dynamic balance of substances such as vitamin C and niacinamide is very important.

[0004] Based on the analysis of TCGA data, CD38 is highly expressed in some non-solid and solid tumors. The former include chronic B-cell leukemia, multiple myeloma, mantle cell lymphoma, etc., and the latter include lung cancer, gastric cancer, melanoma, glioma, esophageal cancer, cervical cancer, etc. Although CD38 is expressed in a variety of human tissues, its expression level in other normal organs is extremely low or low. Moreover, after the primary cancer with high expression of CD38 metastasizes, the tumor tissue in the metastatic lesion still maintains a high level of CD38 expression. Many studies have shown that CD38 has great potential as a new target for the treatment of malignant tumors and their metastases, and is a very promising target protein molecule.

[0005] At present, some research progress has been made in the development of antibodies targeting CD38. Currently, monoclonal antibodies targeting CD38 that have been used in clinical treatment include daratumumab, isatuximab and MOR202. Researchers have analyzed the co-crystal structure of these antibodies and CD38; constructed fluorescent antibodies based on these antibodies, and developed a method for rapid quantification of cell surface CD38 using flow cytometry; in addition, immunotoxins targeting CD38 have been constructed, showing a highly selective killing effect on multiple myeloma (MM) cells, EC 50 It reaches the pM level, has good efficacy and high specificity and safety.

[0006] Nanobodies, also known as single-domain antibodies, are the smallest antigen-binding fragments obtained from heavy-chain antibodies naturally present in the blood of camelids. Compared with traditional antibodies, nanobodies have advantages such as smaller size (about 15kD), good stability and solubility, easy modification, low production cost, and the ability to recognize hidden or uncommon antigenic sites. Although nanobodies targeting CD38 have made some progress in recent years, new targeting antibodies or their combinations are still needed to make them more selective, more efficient, less toxic and more convenient for patients to continue to improve the clinical outcomes of patients with related tumors. Summary of the Invention

[0007] The present invention provides a nanobody targeting CD38, wherein the nanobody targeting CD38 is any one of the following two antibodies:

[0008] primary and secondary antibodies;

[0009] The first antibody and the second antibody each include three complementarity determining regions;

[0010] Wherein, the sequence of the complementary determining region CDR1 of the first antibody is shown as SEQ.ID NO.1, the sequence of CDR2 is shown as SEQ.ID NO.3, and the sequence of CDR3 is shown as SEQ.ID NO.5;

[0011] The sequence of the complementary determining region CDR1 of the second antibody is shown in SEQ.ID NO.2, the sequence of CDR2 is shown in SEQ.ID NO.4, and the sequence of CDR3 is shown in SEQ.ID NO.6.

[0012] Preferably, the amino acid sequence of the first antibody is shown as SEQ.ID NO.7.

[0013] Preferably, the amino acid sequence of the second antibody is shown as SEQ.ID NO.8.

[0014] In addition, the present invention also provides a pharmaceutical composition comprising the CD38-targeting nanobody or its antigen-binding fragment as described above.

[0015] In addition, the present invention also provides a use of a nanoantibody targeting CD38 as described above in the preparation of a drug for diagnosing and / or treating tumors, wherein the tumor includes one or more of epidermal tissue tumors, connective tissue tumors, lymphatic and hematopoietic system tumors, organ epithelial cell tumors, reproductive system and urinary system tumors, digestive system tumors, nervous system tumors, and endocrine system tumors.

[0016] Preferably, the tumor is a respiratory system tumor among epithelial cell tumors of the organ.

[0017] Preferably, the tumor is non-small cell lung cancer among the respiratory system tumors.

[0018] In addition, the present invention also provides the use of the nanoantibody targeting CD38 as described above in the preparation of a product for detecting tumors.

[0019] Preferably, the product for detecting tumors includes at least one of a detection reagent, a detection kit and a detection device.

[0020] The present invention provides a nanobody targeting CD38, a pharmaceutical composition, and its application, wherein the nanobody targeting CD38 is any one of the following two antibodies: a first antibody and a second antibody; the first antibody and the second antibody both include three complementary determining regions; wherein the sequence of CDR1 of the complementary determining region of the first antibody is shown as SEQ.ID NO.1, the sequence of CDR2 is shown as SEQ.ID NO.3, and the sequence of CDR3 is shown as SEQ.ID NO.5; the sequence of CDR1 of the complementary determining region of the second antibody is shown as SEQ.ID NO.2, the sequence of CDR2 is shown as SEQ.ID NO.4, and the sequence of CDR3 is shown as SEQ.ID NO.6. The present invention obtains two nanobodies with high affinity to CD38 protein based on screening of phage nanobody library, and verifies their binding to CD38 protein. The two nanobodies targeting CD38 provided in the present invention have high affinity to both purified and cell surface CD38 protein. The specific treatment and / or delivery strategies developed based on these two nanobodies (including derivative antibodies modified by additions, subtractions, or substitutions to the original sequences of these two nanobodies) provide an effective new approach for targeting CD38 for anti-tumor intervention. Furthermore, molecular probes and other detection reagents can be prepared based on these nanobodies, which specifically bind to CD38 on the tumor surface, providing a new approach for tumor-specific imaging and diagnosis. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] Figure 1 The immunofluorescence staining results of CD38 protein on PC9 cells in Example 1 of the present invention are as follows;

[0022] Figure 2 The immunofluorescence staining results of CD38 protein on A549 cells in Example 1 of the present invention are as follows;

[0023] Figure 3 The Western Blot results of CD38 expression on PC9 and A549 cells in Example 1 of the present invention are as follows;

[0024] Figure 4 This is the Coomassie blue staining result of the CD38-targeting nanobody (fused with a His tag and an HA tag) expressed and purified in Example 2 of the present invention (size is approximately 16 kD);

[0025] Figure 5 Western Blot detection results of the nanobody targeting CD38 in Example 2 of the present invention (using HA and His tag antibodies);

[0026] Figure 6The results of the ELISA method for detecting the binding activity of the CD38-targeting nanobody to the CD38 protein in the second embodiment of the present invention (all results showed binding activity);

[0027] Figure 7 The affinity test result of the first antibody NbD5 in the CD38-targeting Nanobody based on the Biacore SPR platform in Example 2 of the present invention and the CD38 protein (38.26 nM);

[0028] Figure 8 This is the affinity test result (21.07 nM) of the second antibody NbD6 in the CD38-targeting Nanobody based on the Biacore SPR platform in Example 2 of the present invention to the CD38 protein;

[0029] Figure 9 The results of localization of CD38-targeting nanobodies in PC9 and A549 cells based on laser confocal microscopy in the second embodiment of the present invention (both nanobodies can be localized to CD38 on the cell surface);

[0030] Figure 10 The results of the binding of different concentrations of CD38-targeting nanobodies to CD38 antigen on PC9 cells in Example 2 of the present invention are shown;

[0031] Figure 11 These are the binding results of different concentrations of CD38-targeting Nanobodies to the CD38 antigen on A549 cells in Example 2 of the present invention.

[0032] The purpose, features and advantages of the present invention will be further described with reference to the accompanying drawings and in conjunction with the embodiments. DETAILED DESCRIPTION

[0033] In an embodiment of the present application, a nanobody targeting CD38 is provided, wherein the nanobody targeting CD38 is any one of the following two antibodies:

[0034] primary and secondary antibodies;

[0035] The first antibody and the second antibody each include three complementarity determining regions;

[0036] Wherein, the sequence of the complementary determining region CDR1 of the first antibody is shown as SEQ.ID NO.1, the sequence of CDR2 is shown as SEQ.ID NO.3, and the sequence of CDR3 is shown as SEQ.ID NO.5;

[0037] The sequence of the complementary determining region CDR1 of the second antibody is shown in SEQ.ID NO.2, the sequence of CDR2 is shown in SEQ.ID NO.4, and the sequence of CDR3 is shown in SEQ.ID NO.6.

[0038] As mentioned above, the complementary determining regions (CDRs) of the amino acid sequence are the most frequently variable and diverse parts of the antibody molecule. CDRs are located in the convergent region of the antibody structure and are primarily composed of three hypervariable regions (HVRs). The antigen-specific residues that interact with these regions are called variable regions (V).

[0039] The main function of CDRs is to directly interact with specific antigen molecules through the most diverse functional regions of the antibody molecule, thereby achieving antigen recognition and binding during the immune process. Therefore, CDRs are an important component that ensures the high specificity of antibody molecules.

[0040] Furthermore, the amino acid sequence of the first antibody is shown in SEQ.ID NO.7.

[0041] Furthermore, the amino acid sequence of the second antibody is shown in SEQ.ID NO.8.

[0042] Therefore, the nanoantibodies targeting CD38 provided by the present invention include a first antibody (named NbD5) and a second antibody (named NbD6); both antibodies have three complementary determining regions, namely CDR1, CDR2 and CDR3, whose corresponding sequence numbers are shown in Table 1, and the specific sequences are shown in Table 2.

[0043] Table 1. Correspondence between the complementary determining region sequences and amino acid sequences of nanobodies targeting CD38

[0044] Nanobodies targeting CD38 CDR1 CDR2 CDR3 Amino acid sequence Primary Antibody SEQ.ID NO.1 SEQ.ID NO.3 SEQ.ID NO.5 SEQ.ID NO.7 Secondary Antibody SEQ.ID NO.2 SEQ.ID NO.4 SEQ.ID NO.6 SEQ.ID NO.8

[0045] Table 2. Complementarity determining region sequences and amino acid sequences of the first and second antibodies in CD38-targeting nanobodies

[0046]

[0047] In addition, in the examples of the present application, a pharmaceutical composition is also provided, which contains the CD38-targeting nanobody or its antigen-binding fragment as described above.

[0048] In addition, the present invention also provides a use of a nanoantibody targeting CD38 as described above in the preparation of a drug for diagnosing and / or treating tumors, wherein the tumor includes one or more of epidermal tissue tumors, connective tissue tumors, lymphatic and hematopoietic system tumors, organ epithelial cell tumors, reproductive system and urinary system tumors, digestive system tumors, nervous system tumors, and endocrine system tumors.

[0049] Furthermore, the tumor is a respiratory system tumor among the organ epithelial cell tumors.

[0050] Furthermore, the tumor is non-small cell lung cancer among respiratory system tumors.

[0051] In addition, the present invention also provides a use of the above-mentioned CD38-targeting nanobody in the preparation of a detection product for detecting tumors.

[0052] Furthermore, the product for detecting tumors includes at least one of a detection reagent, a detection kit, and a detection device.

[0053] The detection reagents mentioned above can be reagents for detecting specific biomarkers or chemical components, and can be designed to bind to nanoantibodies to improve the specificity and sensitivity of detection.

[0054] As mentioned above, the test kit may contain all the components required to perform the test, such as but not limited to reagents, buffers, standards and sometimes test consumables. These kits are designed to simplify the test process and make it more user-friendly.

[0055] The detection device mentioned above can be a device for reading and analyzing the results of a detection reagent or kit, or a device that uses the aforementioned Nanobodies to detect tumors, or functions as a detection reagent or kit. These devices can be manual or automated, and can include software for data processing and result interpretation.

[0056] Example 1: Verification of CD38 protein expression

[0057] 1. Immunofluorescence staining

[0058] A549 and PC9 cells were revived and passaged three times before being seeded into 24-well plates with slides for subsequent experiments. The specific steps are as follows:

[0059] (1) Place cells on ice, wash three times with PBS, and fix with 4% paraformaldehyde for 10 min;

[0060] (2) Wash with PBS three times and block with 3% BSA for 1 hour;

[0061] (3) Dilute CD38 antibody 1:300 in blocking solution and incubate with cells at 4°C overnight;

[0062] (4) Wash three times with pre-cooled PBS, dilute the secondary antibody conjugated with Alexa 488 at a dilution of 1:1000 in blocking solution, and incubate with the cells in the dark for 1 hour;

[0063] (5) Wash the sections with pre-cooled PBS three times, seal the sections with DAPI-containing sealing medium, and examine the expression of CD38 protein using a confocal microscope.

[0064] 2. Western Blot

[0065] A549 and PC9 cells were revived and passaged three times before being seeded into 100 mm culture dishes for subsequent experiments. The specific steps are as follows:

[0066] (1) Digest with trypsin for 3-4 minutes, add 4-5 mL of PBS to dilute the trypsin to terminate the digestion, and collect the cells after centrifugation;

[0067] (2) Add 200 μL of Lysis Buffer (RIPA + 1× Protease inhibitor) to lyse A549 and PC9 cells;

[0068] (3) Ultrasonicate for 30 seconds (energy 25%, working time 3 seconds, interval 7 seconds), centrifuge at 14000 rpm / 20 min / 4°C, aspirate 90% of the supernatant (180 μL) into a new EP tube, and measure the protein concentration by BCA assay (prepare BCA assay for protein concentration at the same time, take new ordinary EP tubes equal to the number of samples, add 54 μL PBS + 6 μL sample);

[0069] (4) 12.5% ​​SDS-PAGE run, transfer to a 0.45 μm PVDF membrane (200 mA, 120 min), block with 5% BSA for 1 h, and incubate with the primary antibody overnight;

[0070] (5) Wash with TBST five times, 5 min each time, incubate with secondary antibody for 1 h, wash with TBST five times, 5 min each time, and develop color.

[0071] Experimental results:

[0072] In this example, the expression profile and prognosis of CD38 in multiple cancer types were analyzed using the TCGA database. This example focused on verifying the expression of CD38 in non-small cell lung cancer A549 cells and PC9 cells. In this example, CD38 expression on the surface of PC9 cells was detected by immunofluorescence staining (refer to Figure 1 ), CD38 is expressed on the surface of A549 (reference Figure 2). Western Blot was used for further verification. Figure 3 The results showed that CD38 was significantly expressed in both A549 and PC9 cells.

[0073] Example 2: Screening and validation of nanobodies against the extracellular domain of CD38 protein

[0074] 1. Nanobody screening

[0075] The natural alpaca-derived phage display nanoantibody library was screened using the immunotube method. The selected phage display library had a capacity of 2×10 9 The screening steps are as follows:

[0076] (1) The target protein was coated on an immunotube at a concentration of 50 μg / mL and three rounds of enrichment screening were performed;

[0077] (2) The third round of phage eluate was used for plating, and 96 monoclonal clones were randomly selected for ELISA verification. The ELISA reading was 3 times greater than the corresponding BSA reading and the reading was greater than 0.5 as the positive standard;

[0078] (3) The positive monoclones identified by phage ELISA twice were sent to the company for sequencing to confirm the sequence information;

[0079] (4) designing and synthesizing the screened nanobodies based on the sequencing information, and expressing and purifying them through Escherichia coli;

[0080] (5) The affinity of the nanobodies was preliminarily identified using ELISA affinity experiments, and the nanobodies with better affinity were selected. After expression and purification, the affinity constants were determined by surface plasmon resonance (SPR).

[0081] 2. Purification and expression of nanobodies

[0082] The Nanobody gene sequence was cloned into the pCold-II vector, with a 6×His tag fused to its C-terminus. A hemagglutinin HA tag was also fused for subsequent detection. The expression and purification steps are as follows:

[0083] (1) To prevent the formation of inclusion bodies and protein degradation, 0.2 mM IPTG was used for induction at 16°C;

[0084] (2) Induce a large amount of expression according to the pre-experimental induction conditions, and break the bacteria with a high-pressure sterilizer at 1300W;

[0085] (3) Centrifugation at 12,000 g for 45 min at 4°C, and incubate the supernatant with Ni-NTA filler at 4°C for 1 h.

[0086] (4) After purification on a Ni-NTA column, the eluate was eluted with varying imidazole gradients, with each gradient approximately 5 mL. Purity was determined to be >95% by SDS-PAGE. After concentration to a volume of approximately 3 mL, BCA quantitative analysis revealed a concentration of 5.4 mg / mL for the nanoantibody NbD5 and 3.97 mg / mL for NbD6.

[0087] 3. ELISA experiment of nanoantibodies

[0088] The ELISA plate was coated with CD38 protein overnight and blocked, and then various concentrations of HA-tagged nanoantibodies were added and incubated at room temperature for 1 hour. The plate was rinsed with PBST three times and incubated with anti-HA antibody at room temperature for 1 hour. The signal was amplified by horseradish peroxidase (HRP)-labeled anti-HA antibody and developed with TMB. At the same time, irrelevant nanoantibody controls and irrelevant protein antigen blank controls were performed.

[0089] 4. Surface Plasmon Resonance Experiment

[0090] This experiment was used to verify the direct interaction between in vitro expressed and purified nanobodies and in vitro purified antigenic proteins and to calculate the equilibrium constant between the two. The purified antigenic protein (CD38) was immobilized on a chip, and different concentrations of nanobodies (NbD5 and NbD6) were sequentially added to analyze their affinity for the antigenic protein. The reaction signal was recorded over 300 seconds, and kinetic curves were generated to calculate relevant parameters.

[0091] 5. Targeted detection of nanoantibodies on cells

[0092] PC9 and A549 cells were revived and passaged three times before being seeded into 96-well plates for subsequent experiments. The specific steps are as follows:

[0093] (1) Place cells on ice, wash three times with PBS, and fix with 4% paraformaldehyde for 10 min;

[0094] (2) Wash with PBS three times and block with 5% BSA for 1 h;

[0095] (3) Nanobodies were diluted with 1% BSA to different concentration gradients (0-1000 nM) and incubated with cells for 1 h;

[0096] (4) Wash three times with pre-cooled PBST, dilute anti-HA antibody 1:1000 with 1% BSA, and incubate with cells for 1 h;

[0097] (5) Wash three times with pre-cooled PBST, dilute Alexa 488-conjugated secondary antibody 1:1000 with 1% BSA, and incubate with cells in the dark for 1 h;

[0098] (6) Wash with pre-cooled PBST three times, and detect the binding of nanoantibodies on the cell surface using an automatic electrophoresis fluorescence immunoassay.

[0099] Experimental results:

[0100] (1) First, a phage nanoantibody library was screened against CD38, and two positive clones were preliminarily identified, which were named NbD5 (first antibody) and NbD6 (second antibody) (the specific sequences and complementary determining regions (CDRs) of NbD5 and NbD6 are shown in Tables 1 and 2).

[0101] (2) Expression and purification of nanobodies (first antibody NbD5 and second antibody NbD6) and control nanobody C9, detection and confirmation of the correctness of the antibody protein ( Figure 4 and Figure 5 ).Depend on Figure 4 It can be seen that the Coomassie Brilliant Blue staining results showed that by expressing and purifying the nanobody protein fused with the His tag and the HA tag, high-purity nanobodies (NbD5 and NbD6) were obtained, with a size of about 16 kD; Figure 5 It can be seen that Western Blot detection using HA and His tag antibodies confirmed that the nanobody protein was expressed correctly.

[0102] (3) Verified by ELISA, such as Figure 6 As shown, it was confirmed that nanobodies NbD5 (first antibody) and NbD6 (second antibody) had binding activity to recombinant CD38 protein.

[0103] (4) Using the Biacore SPR platform, the affinity constants of NbD5 and NbD6 clones to recombinant CD38 protein were detected and determined. The results showed that the affinity of NbD5 (primary antibody) and NbD6 (secondary antibody) to CD38 protein were 38.26 nM (refer to Figure 7 ) and 21.07nM (reference Figure 8 ).

[0104] (5) Immunofluorescence and Cell ELISA were used to detect whether the nanobody can bind to the CD38 protein on the cell membrane surface. Figure 9 Under laser confocal microscopy, nanoantibodies NbD5 (primary antibody, concentration of 1 μM) and NbD6 (secondary antibody, concentration of 1 μM) can be localized on CD38 on the surface of PC9 cells and A549 cells, where the left column is the primary antibody NbD5 and the right column is the secondary antibody NbD6; Figure 10 Cell ELISA was used to detect the binding of different concentrations of the first antibody NbD5, the second antibody NbD6 and the control nanobody C9 to the CD38 antigen on PC9 cells; Figure 11 Cell ELISA was used to detect the binding of different concentrations of the first antibody NbD5, the second antibody NbD6, and the control nanobody C9 to the CD38 antigen on A549 cells. The results showed that the nanobodies NbD5 and NbD6 have specific recognition ability for the CD38 protein on the surface of PC9 and A549 cell membranes.

[0105] In summary, the two CD38-targeting Nanobodies provided in this invention exhibit high affinity for both purified and cell-surface CD38 proteins. Specific therapeutic and / or delivery strategies developed based on these two Nanobodies (including derivatives modified by additions, subtractions, or substitutions to the original Nanobodies) provide an effective new approach for CD38-targeted anti-tumor intervention. Furthermore, these Nanobodies can be used as a basis for the preparation of molecular probes and other detection reagents that, by specifically binding to CD38 on the tumor surface, provide a new approach for tumor-specific imaging and diagnosis.

[0106] In the description of this specification, the description with reference to the terms "one embodiment", "some embodiments", "example", "specific example", or "some examples" means that the specific features, structures, materials or characteristics described in conjunction with the embodiment or example are included in at least one embodiment or example of the present application. In this specification, the schematic representations of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials or characteristics described can be combined in any one or more embodiments or examples in a suitable manner. In addition, those skilled in the art can combine and combine different embodiments or examples described in this specification and features of different embodiments or examples without contradiction.

[0107] Although the embodiments of the present application have been shown and described above, it can be understood that the above embodiments are exemplary and cannot be understood as limitations on the present application. Ordinary technicians in this field can change, modify, replace and modify the above embodiments within the scope of the present application.

Claims

1. A nanobody targeting CD38, characterized in that The nanobody targeting CD38 is any one of the following two antibodies: primary and secondary antibodies; The first antibody and the second antibody each include three complementarity determining regions; Wherein, the sequence of the complementary determining region CDR1 of the first antibody is shown as SEQ.ID NO.1, the sequence of CDR2 is shown as SEQ.ID NO.3, and the sequence of CDR3 is shown as SEQ.ID NO.5; The sequence of the complementary determining region CDR1 of the second antibody is shown in SEQ.ID NO.2, the sequence of CDR2 is shown in SEQ.ID NO.4, and the sequence of CDR3 is shown in SEQ.ID NO.

6.

2. The CD38-targeting nanobody according to claim 1, wherein The amino acid sequence of the first antibody is shown in SEQ.ID NO.

7.

3. The CD38-targeting nanobody according to claim 1, wherein The amino acid sequence of the second antibody is shown in SEQ.ID NO.

8.

4. A pharmaceutical composition, characterized in that It contains the CD38-targeting nanobody or its antigen-binding fragment according to any one of claims 1 to 3.

5. Use of a CD38-targeting nanobody according to any one of claims 1 to 3 in the preparation of a product for detecting CD38.

6. The use according to claim 5, characterized in that The product for detecting CD38 includes at least one of a detection reagent, a detection kit and a detection device.

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