Long-acting cat omega 2 interferon recombinant protein FeIFN-omega 2-Fc and application

Abstract

The invention belongs to the field of interferon gene engineering, and discloses a long-acting cat omega2 interferon recombinant protein FeIFN-omega2-Fc and application, and the FeIFN-omega2-Fc provided by the invention has the characteristics of high activity, obvious antiviral effect and long half-life period. The applicant carries out effective fusion and mutation on the basis of omega2 interferon, and finally obtains the recombinant protein FeIFN-omega2-Fc. The protein obviously activates downstream signal factors of interferon, has the effect of resisting various cat viruses, shows good safety and curative effect when cat herpes treatment experiments are carried out on cats, and compared with FeIFN-omega2, the half-life period of recombinant interferon is obviously prolonged. The FeIFN-omega2-Fc has a good antiviral effect, can effectively prevent and treat cat viral diseases, has a wide application prospect, and particularly has a relatively good popularization potential in clinical treatment.

Description

Technical Field

[0001] The invention belongs to the field of interferon genetic engineering, and specifically relates to a long-acting feline ω2 interferon recombinant protein FeIFN-ω2-Fc and an application thereof. The FeIFN-ω2-Fc provided by the invention has the characteristics of high activity, antiviral effect and long half-life. Background Art

[0002] The popularity of cats as family pets has promoted the rapid development of the pet industry, but feline viral diseases, such as feline parvovirus (FPV), feline calicivirus (FCV), and feline herpesvirus (FHV), still pose a serious threat to cat health. Although existing vaccines and drugs are helpful for some viral diseases, existing treatments are still insufficient when dealing with new or emerging viruses. Therefore, it is particularly important to develop new complementary treatment strategies.

[0003] Interferon (IFN) is an immunomodulatory factor that can enhance the host's immune response and inhibit viral replication, and is widely used in antiviral and anti-tumor fields. The interferon ω (IFN-ω) gene was first discovered in humans in 1985, and has gradually been studied and applied in other animal species. Feline interferon ω (FeIFN-ω) has also received widespread attention as a natural antiviral protein. FeIFN-ω contains 13 subtypes, among which FeIFN-ω2 exhibits higher antiviral activity. Compared with FeIFN-α, FeIFN-ω2 shows stronger antiviral efficacy in inhibiting the replication of FH V and FCV.

[0004] However, the clinical application of FeIFN-ω2 is limited by its pharmacokinetic properties, especially its short half-life, which requires frequent subcutaneous administration during treatment, further increasing the treatment burden and stress response of cats. To overcome this limitation, researchers have tried to improve the half-life of FeIFN-ω2 in various ways. For example, PEG modification and serum albumin modification can prolong the half-life of FeIFN-ω2, but are usually accompanied by a decrease in antiviral activity, and the preparation process is complicated and costly. In addition, traditional FeIFN-ω2 production methods, such as Escherichia coli or yeast cell expression, have problems with incomplete protein folding and incomplete glycosylation modification, which affect its biological activity.

[0005] Chinese hamster ovary (CHO) cells are a mammalian cell system widely used in biopharmaceutical production. They can provide glycosylation modifications consistent with mammalian cells, thereby significantly improving the stability and biological activity of proteins. In addition, the CHO system can effectively reduce the production of endotoxins and improve the safety of clinical applications. Moreover, Fc fusion proteins can not only retain the biological activity of functional protein molecules, but also have antibody properties, which can significantly increase the half-life. Studies have shown that the use of CHO cells to express human IFN-ω and combine it with Fc fusion technology, and mutate specific sites, can significantly prolong the half-life of human IFN-ω and maintain its antiviral activity. Although Fc fusion technology has made some progress in the application of human IFN, research on feline interferon (FeIFN-ω2) fusion with Fc and homodimers is still relatively limited, and many recombinant interferon proteins currently lack effective in vitro and in vivo antiviral effect verification. Summary of the invention

[0006] The present invention aims to solve the above problems and provide a long-acting and stable feline ω2 interferon recombinant protein FeIFN-ω2-Fc, the sequence of which is shown in SEQ ID NO. 3. The recombinant protein FeIFN-ω2-Fc provided by the present invention significantly improves the activity, antiviral effect and half-life of interferon, providing a new idea for the research and development of antiviral drugs.

[0007] Another object of the present invention is to provide the use of the recombinant protein FeIFN-ω2-Fc in the preparation of a drug for treating or preventing feline viral infection.

[0008] In order to achieve the above object, the present invention adopts the following technical measures:

[0009] The applicant connected the HRV 3C restriction site to the C-terminus of the original sequence of FeIFN-ω2, followed by a GGGGS flexible connecting peptide, and then added an IgG Fc fragment to enhance its stability and half-life, and replaced the 250th S at the amino acid terminal with Y, the 252nd S with T, and the 254th T with E to further increase its half-life; the amino acid sequence of the recombinant protein FeIFN-ω2-Fc finally obtained is shown in SEQ ID NO.3.

[0010] The protection scope of the present invention also includes:

[0011] The gene encoding the recombinant protein described in SEQ ID NO.3.

[0012] An expression cassette, a recombinant vector, a recombinant microorganism or an in vitro recombinant cell having the above coding gene.

[0013] The use of the above recombinant protein, recombinant protein encoding gene or expression box with the above encoding gene, recombinant vector, recombinant microorganism or in vitro recombinant cell in the preparation of feline ω2 interferon recombinant protein.

[0014] Use of the above recombinant protein, recombinant protein encoding gene or expression cassette having the above encoding gene, recombinant vector, recombinant microorganism or in vitro recombinant cell in the preparation of drugs for treating or preventing viral infection.

[0015] In the above-mentioned application, the virus is a mammalian virus.

[0016] In the above application, preferably, the virus is a feline virus;

[0017] In the above application, preferably, the cat virus includes: feline herpes virus, feline calicivirus and / or feline panleukopenia virus.

[0018] In the above application, preferably, the virus also includes vesicular stomatitis virus.

[0019] The encoding gene of the mutant described in SEQ ID NO.3 is preferably shown in SEQ ID NO.4.

[0020] In the above-mentioned applications or methods, preferably, the in vitro recombinant cells are in vitro recombinant CHO cells.

[0021] Compared with the prior art, the present invention has the following positive effects:

[0022] The recombinant interferon FeIFN-ω2-Fc provided by the present invention solves the problems of low biological activity and short half-life of existing cat ω2 interferon. It shows a more superior half-life than the research related to the long-term modification of cat ω2 interferon in the existing literature, and the antiviral effect of FeIFN-ω2-Fc is verified in animal experiments for the first time. The experimental results show that FeIFN-ω2-Fc protein can significantly reduce the viral titer of feline herpes virus (feline herpesvirus, FHV) and effectively alleviate the clinical symptoms caused by viral infection. These advantages make the FeIFN-ω2-Fc of the present invention have a stronger effect and a broader application prospect in antiviral treatment than the prior art.

[0023] The recombinant protein adopts CHO electrotransfection expression system, the expression product has high purity, high protein expression, good stability, simple production process, and is suitable for large-scale industrial production. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Figure 1 The results of SDS-PAGE of three ω2 interferon recombinant proteins;

[0025] Among them, M: DNA molecular mass standard; 1: ω2 recombinant interferon (FeIFN-ω2) expression product; 2: ω2-Fc recombinant interferon (FeIFN-ω2-Fc) expression product; 3: ω2-Dimer recombinant interferon (FeIFN-ω2-Dimer) expression product.

[0026] Figure 2 The activation of four downstream signaling molecules by three recombinant interferons.

[0027] Figure 3 The three recombinant interferons are effective against different feline viruses.

[0028] Figure 4 To investigate the trend of body weight and temperature changes in a cat safety experiment.

[0029] Figure 5 This is the trend of body weight and temperature changes in cats undergoing therapeutic experiments.

[0030] Figure 6 The results of pathological biopsies of therapeutic experiments in cats;

[0031] Among them: the infection group (positive control) showed that the normal structure of lung tissue disappeared, alveolar epithelial cells were necrotic, and a large number of inflammatory cells (mainly neutrophils) were infiltrated in the alveolar cavity and lung interstitium; the treatment group showed that the alveolar wall was slightly thickened, and other lung tissue structures were basically normal, and no obvious histopathological changes were observed; the control group (negative control) showed that the lung tissue had a normal and complete histological structure.

[0032] Figure 7 These are the results of virus content testing in oral, nasal and anal swabs for cat therapeutic experiments.

[0033] Figure 8 The results of interferon half-life determination. DETAILED DESCRIPTION

[0034] The following specific implementation cases are intended to illustrate the content of the present invention, but are not intended to limit the scope of the present invention. Any modification or replacement of the method, steps or conditions of the present invention without departing from the spirit and essence of the present invention shall be regarded as part of the present invention.

[0035] Unless otherwise specified, the technical means used in the embodiments are conventional techniques well known to those skilled in the art.

[0036] Embodiment 1:

[0037] Design and acquisition of three recombinant interferons:

[0038] 1.1 Design of three recombinant interferon nucleotide sequences:

[0039] The gene sequence and amino acid sequence of FeIFN-ω2 (GenBank accession number: NP_001095910.1) and the sequence of cat IgG Fc (GenBank accession number: ATI97569.1) were searched and obtained from NCBI.

[0040] Design three interferon amino acid sequences:

[0041] (1) FeIFN-ω2: The HRV 3C restriction site was connected to the C-terminus of the original sequence of FeIFN-ω2, followed by a GG GGS flexible linker peptide, a Twin-Strep-tag and a 10×His tag; the amino acid sequence of FeIFN-ω2 interferon is shown in SEQ ID NO.1, and the nucleotide sequence encoding it is shown in SEQ ID NO.2.

[0042] (2) FeIFN-ω2-Fc: The stability and half-life of FeIFN-ω2 were enhanced by connecting the HRV 3C restriction site to the C-terminus of the original sequence of FeIFN-ω2, followed by a GGGGS flexible linker peptide, and then adding an IgG Fc fragment. The 250th S at the amino acid terminal was replaced with Y, the 252nd S was replaced with T, and the 254th T was replaced with E to further increase its half-life. The amino acid sequence of FeIFN-ω2-Fc interferon is shown in SEQ ID NO.3, and the nucleotide sequence encoding it is shown in SEQ ID NO.4.

[0043] (3) FeIFN-ω2-Dimer: The optimal position for removing the signal peptide was predicted on the website http: / / www.cbs.dtu.dk / services / SecretomeP / , thereby optimizing the dimer structure of FeIFN-ω2-Dimer interferon. A flexible linker was added after the original sequence of FeIFN-ω2, and then the ω2 interferon molecule without the signal peptide was added to form a dimer structure; the amino acid sequence of FeIFN-ω2-Dimer interferon is shown in SEQ ID NO.5, and the nucleotide sequence encoding it is shown in SEQ ID NO.6.

[0044] The above recombinant interferon can be directly synthesized commercially, or obtained by microbial expression. The present invention obtains it by commercial synthesis and CHO eukaryotic expression.

[0045] 1.2 Synthesis of gene sequences and plasmid construction

[0046] DNAman software was used to analyze restriction enzyme sites. According to the results of the restriction enzyme site analysis and the multiple cloning sites on the transfer vector pCMV, restriction enzyme sites that did not exist in the target gene sequence were added to both ends of the target gene. According to the analysis results, restriction enzyme sites HindIII and Kozak sequences were added to the 5' end, and EcoRI restriction sites were added to the 3' end. The determined gene sequence was handed over to Qingke Biotechnology Co., Ltd. for synthesis, CHO system codon optimization (the optimized sequences are shown in SEQID NO. 2, 4, and 6, respectively), and inserted into the pCMV vector to form plasmids pCMV-FeIFN-ω2, pCMV-FeIFN-ω2-Fc, and pCMV-FeIFN-ω2-Dimer.

[0047] 1.3 Expression of three interferons in CHO

[0048] Under sterile conditions, an Xcell gene pulser (Bio-Rad) was used for electrotransfection. The perforation voltage was set to 300 V, the capacitance was 900 μF single pulse, and the resistance was infinite. A disposable 4 mm electroporation cup (Bio-Rad) was added with 40 μg of plasmid DNA and 0.7 mL of CHO cell suspension (1.5 × 10 7 cells / mL). The plasmids were introduced into CHO cells using electroporation. After transfection, the cells in the electroporation cup were transferred to a triangular culture flask, 30 mL of basal culture medium was added, and the cells were cultured in a shaker at 36-37°C and 5% CO2 for 24 h. Subsequently, the cells were collected by low-speed centrifugation and replaced with glutamine-free basal culture medium containing 50 μM MSX.

[0049] 1.4 Purification of three interferon proteins

[0050] The transfected CHO cells were inoculated (0.5×10 6 cells / mL) in a 2L Erlenmeyer flask containing 400mL CD CHO culture medium, tighten the vented bottle cap, and culture in a shaking incubator at 36-37°C and 5% CO2 for about 3 days. Check cell activity and density every day. After 3 days, when the density reaches 4×10 6 Cells / mL were fed regularly, and after 7-8 days of culture, the culture was terminated when the proportion of live cells dropped to 80%-90%.

[0051] Cell debris was removed by high-speed centrifugation, and the supernatant was collected and filtered through a 0.45 μm filter membrane. The FeIFN-ω2-Fc filtrate was loaded onto a Protein A column pre-equilibrated with PBS (20 mM sodium phosphate, 140 mM NaCl, pH 7.4) and washed to A 280Baseline level, then eluted with sodium acetate buffer (0.1M sodium citrate, pH3.0), collected the eluate and adjusted the pH to 7.4-8.0 by 0.2M Tris-HCl. Similarly, FeIFN-ω2 and FeIFN-ω2-Dimer filtrates were loaded onto Strep purification columns pre-equilibrated with Strep equilibration solution (100mM Tris-HCL, 150mM NACl, 1mM EDT A, pH8.0) and washed to A 280 Baseline level, and then the bound substances were eluted with biotin buffer (50mM biotin in binding buffer).

[0052] Finally, the three proteins were concentrated using a 10kDa ultrafiltration tube and further purified by a Superdex 200Increase gel filtration column at a flow rate of 0.5mL / min. PBS was used as the eluent to finally obtain high-purity recombinant proteins. The purified protein solution was sterilized and filtered through a 0.22μm filter membrane, and stored in aliquots at -80°C. According to SDS-PAGE detection, the molecular weights of FeIFN-ω2, FeIFN-ω2-Fc, and FeIFN-ω2-Dimer were approximately 25kDa, 55kDa, and 45kDa, which were all in line with expectations ( Figure 1 ). The protein yields after purification in 400 mL of culture medium were: FeIFN-ω2: 17.9 mg, FeIFN-ω2-Fc: 91.3 mg, FeIFN-ω2-Dimer: 13.2 mg.

[0053] Embodiment 2:

[0054] Analysis of the effects of three purified recombinant interferons against vesicular stomatitis virus:

[0055] The in vitro biological activity of FeIFN series recombinant interferon proteins was detected by F81-VSV microcytopathic inhibition assay.

[0056] First, the F81 cell suspension was inoculated into a 96-well cell culture plate, and 100 μl was added to each well. After the cells adhered to the wall, the three recombinant interferons were diluted tenfold to different gradients with the assay culture medium (2% bovine serum RPMI), and added to the 96-well plate that had been plated with F81 cells, and 100 μl was added to each well. The culture medium was discarded, and 100 μl of VSV virus solution (dosage of 100 TCID) was added to each well. 50 ), continue culturing for 24 hours, and observe under an inverted fluorescence microscope. When 100% of the cells in each well of the virus control group show fluorescence, and the cells in the cell control group are still growing well without fluorescence, it indicates that the control system is qualified and a comprehensive observation can be made.

[0057] The test results showed that the in vitro antiviral titer of FeIFN-ω2 was 10 8.42 IU / mg, the in vitro antiviral potency of FeIFN-ω2-Fc is 10 7.05 IU / mg, the in vitro antiviral potency of FeIFN-ω2-Dimer is 10 6.74 IU / mg (Table 1).

[0058] In summary, the three recombinant interferons FeIFN-ω2, FeIFN-ω2-Fc and FeIFN-ω2-Dimer provided by the present invention have good in vitro antiviral activity and provide potential application value for the treatment of feline viral infections.

[0059] Table 1 Antiviral activity of target proteins in each group (F81-VSV)

[0060]

[0061] Embodiment 3:

[0062] Activation of downstream signaling molecules by three recombinant interferons

[0063] In order to further detect the antiviral activity of recombinant feline interferon at the mRNA level and explore the duration of its efficacy, four typical downstream signaling molecules of interferon were selected as detection genes, specifically interferon stimulated gene 15 (ISG15), myxovirus resistance 1 (Mx1), interferon-induced protein with tetratricopeptide repeat-1 (IFIT1) and interferon-induced protein with tetratricopeptide repeat-3 (IFIT3), and relative fluorescence quantitative PCR was performed to detect gene expression levels. The gene sequences of the four downstream factors of feline interferon were searched, and four gene primers were designed and synthesized (Table 2). F81 cell suspension was inoculated in 24-well plates and incubated with 500 μL for 10 3ng / mL of three interferons, and a control group without interferon incubation was set up, with 3 replicates for each group. Cell samples were collected at 12h, 24h, 36h, 48h, 60h, and 72h after incubation, and the cell RNA was extracted and reversed for quantitative detection. On F81 cells, the quantitative results showed that the three interferons can significantly activate the four classical interferon downstream signaling molecules, and this effect can last for at least 72h; its activation effect on downstream signaling molecules is mainly concentrated in the early stage after incubation (12-24h), and then it decreases rapidly. Among them, FeIFN-ω2 has the most significant activation effect on downstream signaling molecules ( Figure 2 ), FeIFN-ω2-Fc and FeIFN-ω2-Dimer activated downstream signaling molecules to varying degrees.

[0064] Table 2 Primer sequences of multiple downstream factors of feline interferon

[0065]

[0066] Embodiment 4:

[0067] Effects of three recombinant interferons against different feline viruses (FHV, FCV, FPV)

[0068] To further evaluate the inhibitory effects of recombinant feline interferon FeIFN-ω2, FeIFN-ω2-Fc and FeIFN-ω2-Dimer on FHV, FCV and FPV at the cellular level, the TCID 50 ) method to detect the virus titer.

[0069] First, F81 cells were digested, and cell suspension was added to each well of a 24-well plate. After the cells adhered to the wall, 500 μL of three interferons of different dilutions were added to each well and incubated for 24 h. After the incubation, the 10% fetal bovine serum RPMI medium was discarded, and 0.1 MOI of FHV or FCV diluted in 2% fetal bovine serum RPMI medium was added. After adsorption for 2 h, the virus solution was discarded, and 500 μL of 2% fetal bovine serum RPMI medium was added again. Samples were collected at 12 h, 24 h, 36 h, and 48 h after infection, and the virus supernatant was harvested to detect its virus titer (TCID 50 ), three replicates were set for each group, and the group without interferon was used as the control.

[0070] FPV was inoculated with virus at the same time. FPV diluted with RPMI medium containing 2% fetal bovine serum at 0.1 MOI was mixed with three interferons of different doses and added to a 24-well plate. Then, the cell suspension was added. Samples were collected at 24h, 48h, 72h, and 96h after inoculation. The virus supernatant was harvested and its virus titer (TCID 50), three replicates were set for each group, and the group without interferon was used as the control.

[0071] TCID 50 The results showed that the three interferons could significantly inhibit the titers of the three cat viruses, and the effects were more significant at 24h and 48h. Among them, FeIFN-ω2 and FeIFN-ω2-Fc were better than FeIFN-ω2-Dimer in anti-FHV virus; FeIFN-ω2-Fc was better than FeIFN-ω2 and FeIFN-ω2-Dimer in anti-FPV virus; FeIFN-ω2 was better than FeIFN-ω2-Fc and FeIFN-ω2-Dimer in anti-FCV virus ( Figure 3 ).

[0072] Overall, the antiviral effect of FeIFN-ω2-Dimer is not as good as the other two interferon recombinant proteins. Considering the protein activity, antiviral effect and the advantages of extended half-life in vivo, FeIFN-ω2-Fc was finally selected as the candidate interferon for subsequent animal experiments.

[0073] Embodiment 5:

[0074] Animal safety experiment of FeIFN-ω2-Fc

[0075] To verify the animal safety of FeIFN-ω2-Fc, 3-month-old healthy cats weighing about 1 kg were selected (5 / group, half male and half female) and injected subcutaneously with FeIFN-ω2-Fc at 5 times the therapeutic dose (2 million IU / mL / kg) through the neck, twice a day, with an interval of 12 hours, for 7 consecutive days. The control group was subcutaneously injected with the same volume of PBS, and the regimen was the same as that of the experimental group. During the administration period, the body temperature, weight and clinical symptoms of the animals were continuously monitored and observed until the 14th day. The experimental results showed that no abnormal symptoms were observed in the animals in the experimental and control groups, and their body weight showed a fluctuating upward trend ( Figure 4 ), no obvious lesions were found in the autopsy, indicating that FeIFN-ω2-Fc has good animal safety.

[0076] Embodiment 6:

[0077] Animal therapeutic experiments of FeIFN-ω2-Fc

[0078] Fifteen healthy cats (half male and half female) weighing about 1 kg and aged 3 months were selected and divided into a treatment group (5), an infection group (5), and a control group (5). Except for the control group, each cat was infected with 0.5 mL of FHV virus (1.0×10 6 TCID 50 / mL). The treatment group started to take medicine after the onset of symptoms (in this embodiment, the medicine was taken on the 5th day after infection), and FeIFN-ω2-Fc was injected subcutaneously in the neck at a dose of 400,000 IU / kg, once every 2 days, and continued to take medicine for 5 to 7 days. The control group was anesthetized and nasal drops of PBS were administered and an equal volume of PBS was injected subcutaneously, and the scheme was the same as above. During the administration period, clinical symptoms, body temperature, body weight and virus shedding were continuously monitored, and the FHV quantitative detection primers in Table 3 were used to detect the copy number of FHV in the mixed swabs of the mouth, nose and anus, and the detection was carried out for 14 days.

[0079] Table 3 Primer sequences for absolute quantitative detection of FHV

[0080]

[0081] The results showed that the symptoms of the treatment group were significantly relieved 4 to 5 days after administration, the weight loss rate slowed down, the body temperature rose first and then slowly dropped after administration; the body temperature of the control group was stable, and the body temperature of the infected group remained at a higher temperature and then dropped later. The body weight of the control group and the infected group showed an increasing and continuous decreasing trend, respectively ( Figure 5 ). The results of pathological sections showed that the lung inflammation in the treatment group was relieved to a certain extent compared with the infection group, and the degree of inflammatory cell infiltration decreased ( Figure 6 ). Quantitative detection of virus content in oral, nasal and anal swabs showed that the virus titer in the treatment group was significantly lower than that in the infection group after administration ( Figure 7 ), indicating that FeIFN-ω2-Fc interferon has a good anti-FHV effect in vivo.

[0082] Embodiment 7:

[0083] Determination of FeIFN-ω2-Fc half-life

[0084] Adult healthy cats weighing about 1kg (5 cats / group, half male and half female) were selected and injected with FeIFN-ω2 and FeIF N-ω2-Fc proteins, respectively, and interferon was injected subcutaneously at a dose of 1 mg / kg. Venous blood was collected at 1h, 2h, 4h, 8h, 16h, 24h, 48h, 72h, 96h, 120h, 144h, 168h, 192h and 216h after administration. The blood samples were coagulated at 4℃, centrifuged at 3000r / min for 5min, and the serum was separated and stored at -20℃ for testing. The concentration of interferon protein in serum at each time point was determined by cytopathic inhibition method, and the curve was fitted using DAS pharmacokinetic software to calculate the pharmacokinetic parameters. The experimental results showed that the half-life of recombinant FeIFN-ω2-Fc protein was significantly prolonged, which has obvious advantages over natural interferon.

[0085] Table 4 Half-life of different interferons

[0086]

[0087] Compared with the half-life of about 4.9h of natural FeIFN-ω2, the half-life of recombinant FeIFN-ω2-Fc interferon is significantly extended to 40.554h, which is about 7 times higher, and the FeIFN-ω2 interferon protein is undetectable in the animal body for about 48h, while the FeIFN-ω2-Fc interferon protein can still be detected in the animal body for 216h. The long-term results of the obtained recombinant interferon are significantly better than the widely reported cat interferon preparations. And the FeIFN-ω2-Fc interferon of the present invention was stored for 3 months and 6 months and observed that there was no protein precipitation, and the protein had no obvious degradation. The results show that the long-acting FeIFN-ω2-Fc protein of the present invention has high biological activity, good stability, long half-life and high antiviral activity, and its production process is simple, with high practical value and application prospects.

Claims

1. An artificially synthesized recombinant protein FeIFN-ω2-Fc, the amino acid sequence of the recombinant protein is shown in SEQ ID NO.

3.

2. The gene encoding the recombinant protein described in SEQ ID NO.

3.

3. An expression cassette, recombinant vector, recombinant microorganism or in vitro recombinant cell having the coding gene according to claim 2.

4. Use of the recombinant protein according to claim 1, the coding gene according to claim 2 or the expression cassette, recombinant vector, recombinant microorganism or in vitro recombinant cell having the above coding gene according to claim 3 in the preparation of feline ω2 interferon recombinant protein.

5. Use of the recombinant protein according to claim 1, the coding gene according to claim 2, or the expression cassette, recombinant vector, recombinant microorganism or in vitro recombinant cell having the above coding gene according to claim 3 in the preparation of a drug for treating or preventing viral infection.

6. The use according to claim 5, wherein the virus is a mammalian virus.

7. The use according to claim 5, wherein the virus is a feline virus.

8. The use according to claim 7, wherein the feline virus is: feline herpes virus, feline calicivirus and / or feline panleukopenia virus.

9. The use according to claim 5, wherein the virus is vesicular stomatitis virus. The isolated recombinant cell according to claim 3, which is an isolated recombinant CHO cell.

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