Fusion protein with GLP and GIP dual activities and application thereof

Abstract

The invention relates to the field of medicine, in particular to fusion protein with GLP and GIP dual activity and application of the fusion protein. The GLP and GIP dual-activity protein in the fusion protein with the GLP and GIP dual-activity is obtained through specific design, and the GLP and GIP dual-activity protein is further fused at the N end of a heavy chain constant region and a light chain constant region of IgG4 or the N end of Fc to obtain the fusion protein with the GLP and GIP dual-activity. Research results show that the fusion protein with GLP and GIP dual activities has an obvious effect of reducing the weight of mice, and the fusion protein can be applied to drugs or products for slimming or treating fatty liver.

Description

Technical Field

[0001] The present invention relates to the field of medicine, and in particular to a fusion protein having dual activities of GLP and GIP and applications thereof. Background Art

[0002] Glucagon-like peptide (GLP) is a polypeptide expressed by the proglucagon gene, which exerts its biological effects by binding to the corresponding G protein-coupled receptors. GLP can significantly increase the first and second phase insulin secretion, reduce glucagon levels, and delay gastric emptying. Because its blood sugar lowering effect is blood sugar-dependent and has the effect of reducing body weight, GLP and its analogs are widely used in the treatment of type 2 diabetes and alleviate insulin resistance caused by obesity.

[0003] Glucose-dependent insulinotropic polypeptide (GIP) is a 42 amino acid gastrointestinal regulatory peptide that plays a physiological role in glucose homeostasis by stimulating insulin secretion from pancreatic β cells in the presence of glucose and protecting pancreatic β cells.

[0004] Although the GLP-1 / GIP dual-activity analog (Tirzepatide) is currently used for weight loss, its effect is still unsatisfactory. Therefore, there is an urgent need in the art to develop a drug that can significantly control the patient's weight. Summary of the invention

[0005] Based on the needs of the prior art, the present application provides a fusion protein having dual activities of GLP and GIP and its application.

[0006] First, the present application discloses a protein having dual activities of GLP and GIP, which is a self-designed GLP and GIP targeting polypeptide, and its amino acid sequence is shown in SEQ ID NO.1.

[0007] Furthermore, the present invention provides a fusion protein having dual activities of GLP and GIP, which further comprises Fc or immunoglobulin IgG4 on the basis of the above-mentioned protein; wherein the amino acid sequence of Fc is shown in SEQ ID NO.2.

[0008] Specifically, the amino acid sequence of the heavy chain constant region of IgG4 is shown in SEQ ID NO.3.

[0009] The amino acid sequence of the light chain constant region of IgG4 is shown in SEQ ID NO.4.

[0010] Specifically, a signal peptide is fused to the N-terminus of the GLP-GIP targeting polypeptide, and the signal peptide sequence is shown in SEQ ID NO.5.

[0011] The present invention provides a dual-active fusion protein of GLP and GIP, which is obtained by fusing the dual-active protein of GLP and GIP with IgG4 protein or Fc protein; More specifically, it is obtained by fusing the dual-active proteins of GLP and GIP to the N-terminus of Fc; it is obtained by fusing the dual-active proteins of GLP and GIP to the N-terminus of the IgG4 heavy chain constant region and the light chain constant region.

[0012] More specifically, the amino acid sequence of Fc is shown in SEQ ID NO.2; the amino acid sequence of the IgG4 heavy chain constant region is shown in SEQ ID NO.3; and the amino acid sequence of the IgG4 light chain constant region is shown in SEQ ID NO.4.

[0013] Furthermore, the N-termini of the Fc or IgG4 heavy chain constant region and the light chain constant region are fused to dual-active proteins of GLP and GIP via connecting peptides, respectively. Preferably, the amino acid sequence of the connecting peptide is as shown in SEQ ID NO.6; The amino acid sequence obtained by fusing the dual-active proteins of GLP and GIP to the N-terminus of Fc is shown in SEQ ID NO.7.

[0014] The amino acid sequences obtained by fusing the dual-active proteins of GLP and GIP to the N-termini of the heavy chain constant region and the light chain constant region of IgG4 are shown in SEQ ID NO.8 and SEQ ID NO.9, respectively.

[0015] Optionally, a signal peptide is fused to the N-terminus of the GLP and GIP dual-active protein; preferably, the amino acid sequence of the signal peptide is as shown in SEQ ID NO.5.

[0016] The present invention also provides a nucleic acid encoding the dual-active fusion protein of GLP and GIP; The nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of Fc is shown in SEQ ID NO.10; The nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of the IgG4 heavy chain constant region is shown as SEQ ID NO.11, and the nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of the IgG4 light chain constant region is shown as SEQ ID NO.12.

[0017] The present invention also provides a recombinant expression vector of the encoding nucleic acid.

[0018] The present invention also provides a method for preparing a cell line that produces a fusion protein with dual activities of GLP and GIP, which is obtained by transfecting the encoding nucleic acid into mammalian cells, preferably, the mammal is a CHO cell; preferably, by constructing a recombinant expression vector containing the encoding nucleic acid, linearizing it and then transfecting it into mammalian cells.

[0019] The present invention also provides a cell strain producing a fusion protein with dual activities of GLP and GIP obtained by the method.

[0020] The present invention finally provides the use of the dual-active fusion protein of GLP and GIP and the cell line in preparing weight loss products or drugs for treating fatty liver.

[0021] Beneficial effects of the present invention: Based on the sequence analysis of GIP, GIP-1, GLP-1 / GIP dual-activity analogs (Tirzepatide, prepared by chemical synthesis), the present invention comprehensively considers the characteristics of GLP-1 and GIP sequences, selects amino acids suitable for cell expression, and innovatively designs a new polypeptide sequence (dual-target polypeptide) that co-targets GLP-1 and GIP. Further, it forms a fusion protein with dual activity of GLP and GIP with Fc or IgG4. The results of the study showed that compared with the control, the fusion protein with dual activity of GLP and GIP in the present application had a significant effect on reducing the weight of mice, the volume and weight of the mouse liver decreased, and the total cholesterol in the serum decreased significantly. It was also unexpectedly found that the effect of the fusion of GLP-GIP targeting polypeptide and Fc was more significant. The fusion protein with dual activity of GLP and GIP in the present application can be used in the research and development of drugs or products for weight loss and treatment of fatty liver. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 The body weight changes of mice treated with the fusion protein of the present invention.

[0023] Figure 2 Changes in liver volume of mice treated with fusion protein. Compared with the control group, "*" P<0.01, "**" P<0.05.

[0024] Figure 3 Changes in liver weight of mice treated with fusion protein. Compared with the control group, "*" P<0.01, "**" P<0.05, "***" P<0.001, "****" P<0.0001. DETAILED DESCRIPTION

[0025] The present invention is described below through specific implementation modes in order to better understand the present invention, but it does not constitute a limitation to the present invention.

[0026] 1. Construction of expression vector of GLP-GIP-IgG4 fusion protein (1) Design of GLP-GIP targeting polypeptide (dual-target polypeptide). The present invention newly designs a GLP-GIP targeting polypeptide (dual-target polypeptide) based on sequence analysis of GIP, GIP-1, GLP-1 / GIP dual-active analog (Tirzepatide), etc. The amino acid sequence of the GLP-GIP targeting polypeptide (dual-target polypeptide) is shown in SEQ ID NO. 1: HGEGTFTSDYSILLDKIAQKAFVNWLIAGGPSSGAPPPS.

[0027] (2) The GLP-GIP-Fc fusion protein of the present application: The GLP-GIP-Fc fusion protein is composed of a signal peptide, a GLP-GIP targeting polypeptide, a transfer peptide and Fc. The amino acid sequence of Fc is shown in SEQ ID NO: 2: AESKYGPPCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG.

[0028] The amino acid sequence of the signal peptide is shown in SEQ ID NO.5: METDTLLLWVLLLWVPGSTG; The amino acid sequence of the connecting peptide is shown in SEQ ID NO.6: GGGGSGGGGSGGGGS.

[0029] The amino acid sequence of the GLP-GIP-Fc fusion protein is shown in SEQ ID NO.7: METDTLLLWVLLLWVPGSTGHGEGTFTSDYSILLDKIAQKAFVNWLIAGGPSSGAPPPSGGGGSGGGGSGGGGSAESKYGPPCPPCPAPEAAGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG。

[0030] The nucleotide sequence of the GLP-GIP-Fc fusion protein is shown as SEQ ID NO.10 below: ATGGAAACCGATACCCTCTTGCTTTGGGTACTGCTCCTGTGGGTACCTGGCAGCACCGGTCACGGTGAGGGAACATTCACCTCTGATTACTCCATCTTGCTGGATAAGATCGCTCAGAAGGCCTTCGTGAATTGGTTGATCGCAGGTGGACCTAGTTCTGGAGCTCCACCACCAAGTGGGGGCGGTGGATCTGGGGGTGGAGGTTCTGGCGGTGGGGGGTCCGCTGAAAGCAAGTATGGACCTCCTTGCCCACCCTGTCCTGCACCTGAGGCTGCCGGAGGGCCCAGTGTGTTCTTGTTCCCACCAAAACCCAAGGATACCCTCATGATCAGCCGGACCCCAGAGGTAACCTGCGTGGTCGTGGACGTGAGCCAAGAGGACCCAGAAGTACAATTCAATTGGTATGTGGATGGAGTGGAGGTTCATAACGCAAAAACAAAGCCCCGCGAGGAACAGTTCAACTCCACATACCGCGTGGTCAGCGTTCTGACTGTACTGCACCAGGACTGGCTCAACGGAAAGGAGTACAAATGCAAAGTTTCTAACAAGGGTCTGCCATCCAGCATCGAAAAAACTATCTCCAAAGCAAAGGGGCAGCCTCGTGAACCTCAGGTATACACTCTCCCACCCAGCCAGGAGGAAATGACCAAAAATCAGGTATCTCTCACATGTCTGGTCAAGGGATTCTACCCATCCGACATTGCCGTGGAGTGGGAGTCAAATGGCCAGCCCGAAAATAACTATAAGACCACCCCACCCGTCTTGGACAGCGATGGCAGCTTTTTCCTGTACTCACGCCTGACAGTGGATAAGAGCCGTTGGCAGGAGGGTAACGTATTTTCTTGCAGCGTCATGCATGAAGCCCTGCATAATCATTACACCCAGAAGTCTCTGAGCCTGTCCCTGGGC。

[0031] (3) Design different GLP-GIP-lgG4 fusion proteins.

[0032] (a) GLP-GIP-IgG4 fusion protein: The heavy chain sequence GLP-GIP-Hc is composed of a signal peptide, a GLP-GIP targeting polypeptide, a transfer peptide and an IgG4 protein Hc.

[0033] Among them, the amino acid sequence of the heavy chain constant region of IgG4 is as SEQ ID Shown in NO.3: PSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPAALGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGV EVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG.

[0034] The amino acid sequence of the heavy chain sequence GLP-GIP-Hc is shown in SEQ ID NO.8: METDTLLLWVLLLWVPGSTGHGEGTFTSDYSILLDKIAQKAFVNWLIAGGPSSGAPPPSGGGGSGGGGSGGGGSPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPAALGGPSVFLFPPKPKD TLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQV YTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLG.

[0035] The nucleotide sequence of the heavy chain sequence GLP-GIP-Hc is based on artificially optimized CHO cell preferred codons, as shown in SEQ ID NO.11:

[0036] The light chain sequence GLP-GIP-Lc is composed of a signal peptide, a GLP-GIP targeting polypeptide, a transfer peptide and an IgG4 protein Lc.

[0037] Among them, the amino acid sequence of the light chain constant region of IgG4 is shown in SEQ ID NO.4: APSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC.

[0038] The amino acid sequence of the light chain sequence GLP-GIP-Lc is shown in SEQ ID NO.9: METDTLLLWVLLLWVPGSTGHGEGTFTSDYSILLDKIAQKAFVNWLIAGGPSSGAPPPSGGGGSGGGGSGGGGSAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC.

[0039] The nucleotide sequence of the light chain sequence GLP-GIP-Lc is artificially optimized by the CHO cell preferred codon, as shown in SEQ ID NO.12: .

[0040] (b) Construction method of GLP-lgG4 (refer to CN202410464079.7) The genes encoding the above fusion proteins are all artificially optimized CHO cell preferred codons, and the full-length sequences are obtained by chemical synthesis. The above-synthesized GLP-GIP-Hc is cloned into the HindIII / EcoRI site of pKS001 to obtain the vector pKS001-GLP-GIP-Hc; the above-synthesized GLP-GIP-Lc is cloned into the XbaI / NotI site of pKS001-GLP-GIP-Hc to obtain the IgG4 expression vector pKS-GLP-GIP-IgG4. The above-synthesized GLP-GIP -Fc is cloned into the HindIII / EcoRI site of pKS001 to obtain the vector pKS001-GLP-GIP-Fc. The construction method of GLP-lgG4 refers to patent CN202410464079.7. The relevant experimental means use conventional experimental means of molecular biology.

[0041] 2. Construction of a cell line with stable expression of fusion protein After linearization of the recombinant expression plasmid by PvuI, it was transfected into CHO-K1 cells. For the specific transfection process, please refer to the instruction manual of Lipofectamine2000 (Invitrogen). 48 hours after transfection, the cells began to be cultured in the screening medium. The screening medium used was Hycell CHO medium containing 500μg / ml G418 and 6mM Glutamine. The screening medium was replaced every 3 days. After transfection, the cell viability first decreased and then increased under the action of the drug. When the cell viability rose to more than 90%, the transfected cells were plated on 96-well plates for monoclonal screening by limiting dilution method. The cell density of the plate was 0.5 / well. After the cells were statically cultured in the incubator for 14 days, the monoclonal cell expression supernatant was detected by ELISA. According to the test results, the 10 monoclonal cell lines with the highest expression level were screened out for each transfected cell for gradual expansion culture. After 3 days of culture in a 6-well plate, the cells were counted and the protein expression level was detected. The 3 cell lines with the highest protein expression level were selected for expansion culture in a T125 shake flask. After 3 days of culture, the cells were counted and the protein expression level was detected. The cells with the highest expression level were selected as stable cell lines.

[0042] 3. Expression and purification of fusion protein The high-expressing cells were cultured in serum-free CD OptiCHO and the culture supernatant was collected after a certain period of time. The fusion protein was purified by the following Protein A affinity chromatography method: Chromatographic column: AT Protein A Diamond affinity chromatography medium, column volume CV: 5 ml, flow rate: 5 ml / min, pressure limit: ≤0.3 MPa.

[0043] Pretreatment: Rinse with purified water for at least 5 CV.

[0044] Equilibration: First elute with elution buffer for 1 CV, then fully equilibrate with binding buffer for at least 10 CV before loading.

[0045] Loading: Load the filtered CHO suspension cell culture onto the equilibrated chromatography column, collect the load flow-through, and sample 100 μl for electrophoresis detection.

[0046] Elution: After loading, increase the flow rate to 5 ml / min and elute with binding buffer for at least 6 CV until the UV baseline is flat.

[0047] Elution: Use 100% elution buffer for elution until the UV baseline is flat, collect the eluted components in separate tubes, and add an appropriate amount of neutralization buffer to each collection tube according to the collection volume and mix well.

[0048] 4. Therapeutic efficacy test of fusion protein In this experiment, C57BL / 6JNifdc-DIO male mice induced by high-fat diet for 10 weeks were selected, and the drug was administered subcutaneously in this model mouse for 5 weeks with a dosing frequency of 2 times per week.

[0049] The fusion proteins used in this experiment include the GLP-GIP-IgG4 fusion protein (GLP-GIP-lgG4) of this application, the GLP-GIP-Fc fusion protein (GLP-GIP-Fc) and the patented GLP-IgG4 fusion protein. The fusion proteins used were diluted to the required concentration using a diluent (30mM PB, 0.04% Tween 80, 3% mannitol, pH7.0) to obtain each fusion protein solution. 25 SPF-grade C57BL / 6JNifdc-DIO male mice weighing 30-40g were induced for 10 weeks using Research Diet-D12492 feed and purchased from Biocytogen Jiangsu Gene Biotechnology Co., Ltd. and raised by Beijing Huilin Zegu Biotechnology Co., Ltd. 35 mice were randomly divided into 6 experimental groups and 1 blank control group. The mice in the 6 experimental groups were injected with 75μg of GLP-GIP-IgG4, 225μg of GLP-GIP-IgG4, 75μg of GLP-GIP-Fc, 225μg of GLP-GIP-Fc, 75μg of GLP-IgG4, and 225μg of GLP-IgG4, respectively. The fusion protein solution of the 6 experimental groups was 100μl. The blank control group was injected with 100μl of normal saline. Injection procedure: Inject twice a week for 5 consecutive weeks. The mice were weighed before each injection. After 5 weeks, the mice were killed, the mouse livers were removed, and the volume and weight of the mouse livers were measured. The results are shown in Figure 1-3 shown.

[0050] Depend on Figure 1 The results showed that, compared with the blank control group, the GLP-GIP-Fc 75μg and GLP-GIP -Fc225μg groups had a weight loss of nearly 20% and 23.9% respectively after five weeks; GLP-GIP-IgG4 had a slightly better weight loss effect on mice than GLP-IgG4. Figure 2~3 As shown in the results, compared with the blank control group, the liver volume and weight of the GLP-GIP-IgG4 225μg, GLP-GIP-Fc 75μg and GLP-GIP -Fc 225μg, GLP-IgG4 225μg groups decreased, and the decrease in liver volume and weight in the GLP-GIP -Fc 75μg and GLP-GIP -Fc 225μg groups was the most obvious.

[0051] In the above experiment, blood was collected after the mice were weighed for the last time and fasted for 16 hours, and the total cholesterol (TC) assay kit (oxidase method) and triglyceride (TG) assay kit (oxidase method) (Shenzhen Mindray Bio-Medical Electronics Co., Ltd.) were used to detect the total cholesterol and triglyceride levels in the serum of the experimental group and the blank control group mice, respectively. The test results of total cholesterol and triglyceride are shown in Table 1.

[0052] Table 1

[0053] As shown in the results of Table 1, compared with the blank control group, the total cholesterol content in the experimental groups of the present application GLP-GIP-Fc 75 μg and GLP-GIP-Fc 225 μg decreased significantly, with the former decreasing more significantly.

[0054] Finally, it should be noted that the above is only a preferred embodiment of the present invention and is not intended to limit the present invention. Although the present invention has been described in detail with reference to the aforementioned embodiments, those skilled in the art can still modify the technical solutions described in the aforementioned various embodiments or replace some of the technical features therein by equivalents. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present invention shall be included in the protection scope of the present invention.

Claims

1. A protein with dual activity of GLP and GIP, characterized in that: Its amino acid sequence is shown in SEQ ID NO.

1.

2. A fusion protein with dual activity of GLP and GIP, characterized in that: The dual-active protein of GLP and GIP as claimed in claim 1 is fused with IgG4 protein or Fc protein; More specifically, they are fused to the N-terminus of Fc The GLP and GIP dual-active protein as claimed in claim 1 is obtained; the GLP and GIP dual-active protein as claimed in claim 1 is fused to the N-terminus of the IgG4 heavy chain constant region and the light chain constant region respectively.

3. The dual-active fusion protein of GLP and GIP according to claim 2, characterized in that: The amino acid sequence of Fc is shown in SEQ ID NO.2; the amino acid sequence of the IgG4 heavy chain constant region is shown in SEQ ID NO.3; and the amino acid sequence of the IgG4 light chain constant region is shown in SEQ ID NO.

4.

4. The dual-active fusion protein of GLP and GIP according to claim 2, characterized in that: The N-termini of the Fc or IgG4 heavy chain constant region and the light chain constant region are fused to the dual-active proteins of GLP and GIP through connecting peptides, respectively. Preferably, the amino acid sequence of the connecting peptide is as shown in SEQ ID NO.6; The amino acid sequence obtained by fusing the dual-active protein of GLP and GIP as claimed in claim 1 to the N-terminus of Fc is shown in SEQ ID NO.7; Fusion of the N-terminal of the IgG4 heavy chain constant region and the light chain constant region The amino acid sequences of the GLP and GIP dual-activity protein according to claim 1 are shown in SEQ ID NO.8 and SEQ ID NO.9, respectively.

5. The dual-active fusion protein of GLP and GIP according to claim 2, characterized in that: A signal peptide is fused to the N-terminus of the GLP and GIP dual-active protein; preferably, the amino acid sequence of the signal peptide is as shown in SEQ ID NO.

5.

6. A nucleic acid encoding a dual-active fusion protein of GLP and GIP as described in any one of claims 2 to 5; The nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of Fc is shown in SEQ ID NO.10; The nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of the IgG4 heavy chain constant region is shown in SEQ ID NO.11, and the nucleotide sequence encoding the protein with dual activity of GLP and GIP fused to the N-terminus of the IgG4 light chain constant region is shown in SEQ ID NO.

12.

7. The recombinant expression vector encoding the nucleic acid according to claim 6.

8. A method for preparing a cell line producing a fusion protein having dual activities of GLP and GIP, characterized in that: The method is obtained by transfecting a mammalian cell with the encoding nucleic acid as claimed in claim 5 or 6, preferably, the mammal is a CHO cell; preferably, the method is obtained by constructing a recombinant expression vector containing the encoding nucleic acid as claimed in claim 6, and then transfecting the vector into a mammalian cell after linearization.

9. A cell line producing a fusion protein having dual activities of GLP and GIP obtained according to the method of claim 8.

10. Use of the dual-active fusion protein of GLP and GIP according to any one of claims 2 to 5, or the cell line according to claim 9 in the preparation of weight loss products or drugs for treating fatty liver.

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