A recombinant polypeptide with broad-spectrum inhibition of novel coronavirus and its application

Abstract

The present invention discloses a recombinant polypeptide that broadly inhibits the novel coronavirus and its application, relating to the fields of synthetic biology and medicine. The recombinant polypeptide is composed of a polypeptide derived from ACE2 and a polypeptide derived from the novel coronavirus HR2, connected by a linker. The polypeptide derived from ACE2 has six amino acid mutations (AKT / VEW and NHE / DIQ) introduced to enhance its interaction with RBD. The recombinant polypeptide of the present invention has high-efficiency and broad-spectrum activity against the novel coronavirus and can be used to prepare and develop drugs against the novel coronavirus. The recombinant polypeptide provided by the present invention can effectively inhibit the novel coronavirus and its various mutants and can be used to prepare pharmaceutical compositions for the prevention or treatment of coronavirus. It has the potential to be developed into a broad-spectrum anti-coronavirus drug to minimize the prevalence of existing coronaviruses and possible new and re-emerging coronaviruses in the future.

Description

Technical Field

[0001] The present invention relates to the fields of synthetic biology and medicine, and specifically to a recombinant polypeptide that broadly inhibits the novel coronavirus and its application. Background Art

[0002] The virus particle is spherical, with spike structures arranged on its surface composed of four structural proteins: spike protein (S), nucleocapsid protein (N), membrane protein (M), and envelope protein (E). Among them, the trimeric S glycoprotein directly determines the host range and infection efficiency through conformational dynamic equilibrium and molecular interaction mechanisms.

[0003] The coronavirus spike protein plays a crucial role in viral invasion of target cells. The S protein comprises two functional subunits, S1 and S2. The receptor-binding domain (RBD) of the S1 subunit is responsible for recognizing and binding to the host cell receptor, angiotensin-converting enzyme-2 (ACE2), triggering protease cleavage and activation. Subsequently, the S2 subunit undergoes a series of conformational changes, prompting the heptad repeat sequence HR1 to form a trimeric coiled-coil structure, exposing its hydrophobic pocket. This allows the heptad repeat sequence to bind to HR2 to form a six-helix bundle (6-HB), ultimately driving fusion of the virus with the host cell membrane. Therefore, the RBD and HR1 regions are ideal targets for the development of viral entry inhibitors.

[0004] However, during the evolution of coronaviruses, the spike protein RBD and N-terminal domain (NTD) undergo frequent mutations. These mutations not only enhance the virus's immune evasion capabilities, allowing it to evade the host immune system, but may also lead to changes in its transmissibility and pathogenicity. Despite this, certain hidden sites within the spike protein trimer remain relatively conserved, remaining unchanged across different mutant strains. These highly conserved sites provide potential targets for the development of antiviral drugs.

[0005] Therefore, the development of broad-spectrum antiviral drugs that can effectively combat the virus and its mutants has become a key to preventing and treating infectious disease outbreaks. For example, patent application publication number CN115925826A discloses a peptide that has a strong inhibitory effect on both the original strain of the novel coronavirus and its various variants; another example is patent application publication number CN114437184A, which discloses a peptide that has a strong inhibitory effect on both the original strain of the novel coronavirus and its various variants. As well as the inventors' previous research (Bi W, Tang K, Chen G, et al. An enhanced broad-spectrum peptide inhibits Omicron variants in vivo. Cell Rep Med. Published online February 3, 2024. doi:10.1016 / j.xcrm.2024.101418), the peptide A1 derived from the ACE2 receptor was fused with the peptide HR2m derived from the highly conserved region HR2 of SARS-CoV-2. The results showed that it can broadly inhibit the infection of the new coronavirus, but the interaction between the peptide A1 and RBD is relatively weak. Summary of the Invention

[0006] Based on the shortcomings of the existing basis, the present invention provides a recombinant polypeptide with a broad spectrum of inhibition of the new coronavirus and its application. On the basis of ACE2 polypeptide A1, six amino acid mutations (AKT / VEW and NHE / DIQ) are further introduced to enhance its interaction with RBD, thereby enhancing its antiviral activity.

[0007] The specific technical solutions of the present invention are as follows:

[0008] The present invention provides a recombinant polypeptide that broadly inhibits the novel coronavirus, wherein the recombinant polypeptide comprises a polypeptide A3M6 derived from ACE2 and a polypeptide HR2 derived from the novel coronavirus HR2;

[0009] The amino acid sequence of the polypeptide A3M6 is N-terminal-IEEQVEWFLDKFDIQAEDLFYQSSLGGSKGDFR-C-terminal,

[0010] The amino acid sequence of the polypeptide HR2 is N-terminal-DVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQ-C-terminal.

[0011] Preferably, the polypeptide A3M6 and the polypeptide HR2 are connected via a linker; the linker is a flexible linker or a rigid linker;

[0012] The amino acid sequence of the flexible linker is (GGGGS) n ,

[0013] The amino acid sequence of the rigid linker is (MALEK) n ,

[0014] Wherein, n=3-10, and n is an integer.

[0015] Further preferably, the n=7, the recombinant polypeptide is named A3M6L35HR2 (FL), and its inhibitory activity against SARS-CoV-2 is 4.9 nM. The results show that after introducing 6 amino acid mutations into the ACE2-derived polypeptide and fusing it with the HR2 polypeptide through a flexible linker, its inhibitory activity is significantly improved. And the IC of A3M6L35HR2 (FL) for inhibiting SARS-CoV-2 is 4.9 nM. 50 A1L35HR2 (IC 50 =157 nM) and HR2 (IC 50 =157 nM) peptide by 7-fold and 32-fold.

[0016] The present invention also provides a nucleic acid molecule encoding the recombinant polypeptide.

[0017] Preferably, the gene sequence of A3M6L35HR2 (FL) is shown as SEQ ID NO.8.

[0018] The present invention also provides a recombinant vector comprising the nucleic acid molecule.

[0019] The present invention also provides a host cell comprising the recombinant vector.

[0020] The present invention also provides a method for preparing the polypeptide, which includes designing and synthesizing the coding gene of the recombinant polypeptide, constructing a recombinant expression vector of the dual-target polypeptide, transforming it into Escherichia coli for expanded culture and induced expression, and separating and purifying it.

[0021] The present invention also provides the use of the recombinant polypeptide, the nucleic acid molecule, the recombinant vector or the host cell in the preparation of a drug for preventing or treating diseases caused by coronavirus infection.

[0022] In the above application, the disease caused by the coronavirus infection is a respiratory system infection. The respiratory system infection can be a respiratory tract infection and / or a lung infection.

[0023] In the above application, the coronavirus is the original strain of the new coronavirus and / or the mutant strain of the new coronavirus; the mutant strain of the new coronavirus is D614G, Beta, Delta and / or Omicron series mutant strains.

[0024] In a specific embodiment of the present invention, the Omicron series variants include XBB, BF7, BA.2, EG.5.1, BA.2.86 and KP.2, but are not limited thereto.

[0025] In the embodiment of the present invention, the results showed that A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) all had good inhibitory activity against different mutant strains of SARS-CoV-2, and A3M6L35HR2 (FL) had the best inhibitory activity against different mutant strains of SARS-CoV-2, with an IC 50 Between 1.7-16.2 nM.

[0026] The present invention also provides a pharmaceutical composition, which contains the recombinant polypeptide, the nucleic acid molecule, the recombinant vector or the host cell, and a pharmaceutically acceptable carrier.

[0027] Preferably, the pharmaceutical composition is in the form of a nasal spray formulation, an oral formulation, or a parenteral formulation;

[0028] Further preferably, the oral preparation is selected from tablets, capsules, granules, suspensions and pills;

[0029] Further preferably, the parenteral preparation is an injectable or bolus preparation;

[0030] Preferably, the pharmaceutical composition is a vaccine composition.

[0031] In terms of recombinant polypeptide design, simply expressing the polypeptides in series will affect each other's structure and may mask the target of interaction with the virus. Therefore, simply expressing the polypeptides in series cannot guarantee that each polypeptide can form the correct structure, or even cannot be expressed. On the one hand, the present invention connects the two polypeptides in series by adding different linkers between them, so that the two polypeptides do not interfere with each other in structure. Most importantly, the introduction of six amino acid mutations in the ACE2 polypeptide further ensures that the recombinant polypeptides do not affect each other in spatial structure. At the same time, the introduction of six amino acid mutations further enhances the interaction between the polypeptide and the virus, thereby maximizing the efficient and broad-spectrum inhibition of novel coronavirus infection.

[0032] The present invention obtains a recombinant polypeptide through biosynthesis in the microorganism Escherichia coli. The polypeptide has high efficacy and broad-spectrum anti-novel coronavirus activity and has the advantages of simple process, low cost, high yield, and easy promotion. Chemical solid-phase synthesis of polypeptides is complex and time-consuming, and it is difficult to obtain such polypeptides with a large number of repetitive amino acid sequences.

[0033] Beneficial effects of the present invention:

[0034] The recombinant polypeptides described herein include a polypeptide derived from the ACE2 receptor and a polypeptide of the novel coronavirus HR2, both of which are expressed as a fusion via a flexible or rigid linker. The recombinant polypeptides provided by the present invention can effectively inhibit the novel coronavirus and its various mutants, and can be used to prepare pharmaceutical compositions for the prevention or treatment of coronaviruses, with the potential to be developed into broad-spectrum anti-coronavirus drugs. BRIEF DESCRIPTION OF THE DRAWINGS

[0035] Figure 1 Design (A) and identification (B) of recombinant polypeptides;

[0036] Figure 2 Size exclusion chromatography (SEC) analysis of recombinant peptides; AC represents the SEC analysis of three peptides, A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2), respectively;

[0037] Figure 3 is the inhibitory activity of the recombinant polypeptide against SARS-CoV-2; wherein, A is the inhibitory activity of A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) against SARS-CoV-2 pseudovirus; B is the inhibitory activity of A3M6L35HR2 (FL), A3M6, HR2 polypeptide and A3M6+HR2 mixture against SARS-CoV-2;

[0038] Figure 4 The effect of the recombinant polypeptide on the activity of target cells;

[0039] Figure 5 represents the inhibitory activity of the recombinant polypeptide against different mutant strains of SARS-CoV-2; AH represents the inhibitory activity of different mutant strains;

[0040] Figure 6 The recombinant polypeptide inhibits the cell-cell fusion activity mediated by the SARS-CoV-2 D614G S protein; A is a representative diagram of cell-cell fusion; B is a statistical diagram of the cell-cell fusion inhibitory activity;

[0041] Figure 7 for the interaction of the recombinant peptide with SARS-CoV-2 HR1;

[0042] Figure 8is the secondary structure of the recombinant polypeptide and its complex; wherein A is the secondary structure of A3M6, HR1 and the mixture of A3M6+HR1; B is the secondary structure of HR2, HR1 and the mixture of HR2+HR1; C is the secondary structure of A3M6L35HR2 (FL), HR1 and the mixture of A3M6L35HR2 (FL)+HR1. DETAILED DESCRIPTION

[0043] Example 1

[0044] 1. Design of recombinant peptides

[0045] The recombinant polypeptide of the present invention is composed of polypeptides derived from ACE2 and SARS-CoV-2 HR2 through a flexible or rigid linker, and is composed of the following polypeptide structures from the N-terminus to the C-terminus: polypeptide A3M6 derived from ACE2 (sequence as Figure 1 A in FIG), a flexible linker (GGGGS) n or a rigid linker (MALEK) n and a polypeptide HR2 derived from HR2 (sequence as shown in FIG). Figure 1 ), wherein the linker may comprise n tandem repeat sequences, and n may be in the range of 3-10.

[0046] In this example, n=7 was selected as an example, wherein the recombinant polypeptide connected by a flexible linker was named A3M6L35HR2 (FL), and the recombinant polypeptide connected by a rigid linker was named A3M6L35HR2 (HL2). The existing A1L35HR2 polypeptide was used for comparison. The specific amino acid sequence is as follows: Figure 1 As shown in A.

[0047] 2. Construction of recombinant vector pET28a-A3M6L35HR2 (FL)

[0048] The following primers were designed (the underlined parts represent Nco I and Xho I restriction enzyme cutting site):

[0049] Nco I A3M6 F:ggagatata CCATGG GCATTGAAGAACAAGTGGAATGGTTTCTGGATAAATTTGATATTCAAGCGGAAGATC,

[0050] 6 HIS Xho I HR2 R:ggtggtggtg CTCGAG CTGTTCATATTTGCCCAG;

[0051] Using the laboratory pET28a-A1L35HR2 vector as a template, the A3M6L35HR2 (FL) gene fragment was amplified by PCR technology, and the above gene fragment was connected to the NcoI / Xho The pET28a linear vector was obtained by double enzyme digestion with I to obtain the pET28a-A3M6L35HR2 (FL) recombinant vector.

[0052] pET28a-A3M6L35HR2 (HL2) was synthesized and recombined by GENEWIZ Biotechnology Co., Ltd.

[0053] 3. Expression and purification of A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2)

[0054] The plasmids encoding A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) genes (sequences shown in SEQ ID NOs. 7-9) were transformed into Escherichia coli BL21 (DE3), and single colonies were selected and cultured in LB medium. The bacterial solution was amplified at 37°C and 220 rpm to a bacterial concentration of OD 600 The p-value reached approximately 0.5. Induction was performed by adding 0.5 mM isopropyl-β-D-thiogalactopyranoside (IPTG) at 37°C for 5 h. Cells were harvested by centrifugation at 12,000 × g for 3 min, suspended in lysis buffer (10 mM imidazole containing 0.1% Triton), disrupted by sonication, and centrifuged at 12,000 × g for 10 min. The supernatant was bound to Ni-NTA at 4°C for 30 min. Contaminants were washed with 20 mM, 30 mM, 50 mM, and 100 mM imidazole, and the target protein was eluted with 250 mM imidazole. The purified recombinant peptide was analyzed by SDS-PAGE and immunoblotting.

[0055] like Figure 1 As shown in Figure B, the three polypeptides A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) all migrated to positions close to the expected molecular weight of the band.

[0056] 4. SEC Analysis

[0057] The crude recombinant peptide was further analyzed using a Superdex 75 Increase 10 / 300 GL column. The column was first connected to an AKTA instrument and washed with water for one column volume (CV). The column was then equilibrated with 1× PBS for one CV. The preliminarily purified peptide was filtered through a 0.22 μm membrane and then loaded onto a Superdex 75 Increase 10 / 300 GL column. Detection was performed at 280 nm at a flow rate of 0.5 ml / min.

[0058] like Figure 2 As shown in the figure, the peak volumes of the three polypeptides A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) are between 10 and 15 ml, indicating that their molecular weight is around 15 kDa.

[0059] Example 2

[0060] 1. Preparation of SARS-CoV-2 pseudovirus

[0061] 6.5 × 10 6 293T cells were plated in 10 cm cell culture plates to a cell density of approximately 80% before transfection. One hour before transfection, fresh culture medium was replaced. Transfection reagent (Transfection Reagentlipo2000, TR001), the luciferase reporter plasmid pNL4-3.Luc.RE, and the SARS-CoV-2 S protein-expressing plasmids pcDNA3.1-SARS-CoV-2-S and its mutants (Beta, Delta, XBB, BF7, BA.2, EG.5.1, BA.2.86, and KP.2) were diluted in serum-free DMEM. The diluted transfection reagent was added dropwise to the diluted plasmids at a 1:1 ratio, gently mixed, and incubated at room temperature for 20 minutes. The transfection working solution was added dropwise to the culture medium, and the cells were incubated at 37°C in a 5% CO2 incubator. Twelve hours after transfection, the culture medium was discarded and replaced with fresh cell culture medium. 48-72 h after transfection, the cell supernatant was collected, centrifuged at 3000 × g for 10 min, aliquoted, and stored at -80°C until use.

[0062] 2. Pseudovirus inhibition test

[0063] Caco-2 cells were plated at 10 per well 4Cells were plated in a 96-well cell culture plate and cultured overnight to allow the cells to adhere to the wall. A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) peptides were diluted to the required experimental concentration (starting concentration 1 μM) and a 4-fold serial dilution was performed. Positive (SARS-CoV-2 pseudovirus + Caco-2 cells) and negative (Caco-2 cells) control wells were set at the same time. The SARS-CoV-2 pseudovirus was taken out of the -80°C refrigerator, dissolved and mixed, and then added to the diluted peptide drug and incubated in a 37°C incubator for 30 min. The mixture of peptide drug and SARS-CoV-2 pseudovirus was transferred to Caco-2 cells and cultured at 37°C overnight. The peptide drug and pseudovirus were discarded, and an equal volume of fresh cell culture medium was added and cultured for another two days. The luciferase enzyme activity (unit is relative light unit, RLU) was detected according to the method of the luciferase reporter gene assay kit, and the IC value of the peptide was calculated based on the result. 50 The inhibitory effect of peptide drugs on viruses was further evaluated.

[0064] The results are as follows Figure 3 As shown in Figure A, the inhibitory activities of A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) against SARS-CoV-2 were 35, 4.9 and 241 nM, respectively. This result shows that after introducing 6 amino acid mutations into the ACE2-derived peptide and fusing it with the HR2 peptide through a flexible linker, its inhibitory activity was significantly improved. The inventors then compared the inhibitory activity of A3M6L35HR2 (FL), A3M6, HR2 peptide and A3M6+HR2 mixture against SARS-CoV-2. The results showed that the A3M6 peptide alone had no significant inhibitory activity, while A3M6L35HR2 (FL) inhibited SARS-CoV-2 with an IC of 1. 50 is 4.8 nM, which is approximately the same as HR2 (IC 50 =157 nM) peptide (32 times Figure 3 B in ). Figure 5 A to Figure 5 The results showed that A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) all had inhibitory activity against different mutant strains of SARS-CoV-2, and A3M6L35HR2 (FL) had the best inhibitory activity against different mutant strains of SARS-CoV-2, with its IC 50 The range is 1.7-16.2 nM. The above results indicate that A3M6L35HR2 (FL) has broad-spectrum anti-SARS-CoV-2 activity.

[0065] 3. Safety Analysis of A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) Peptides

[0066] A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) peptides were diluted 4-fold in a 96-well plate, with 100 μl per well added to target Caco-2 cells. After incubation at 37°C in a CO2 incubator for 48 h, 5 μl of CCK-8 was added to each well and cultured for another 2 h. The OD450 absorbance was measured, and cell viability was calculated based on the absorbance.

[0067] The results are as follows Figure 4 As shown in the figure, cells treated with different concentrations of A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) showed no obvious cytotoxicity. When A1L35HR2, A3M6L35HR2 (FL) and A3M6L35HR2 (HL2) were incubated with cells at a high concentration of 20 μM, the viability of Caco-2 cells was above 90%, which was higher than the IC50 of A1L35HR2 for inhibiting SARS-CoV-2. 50 It is about 83 times higher, indicating that it has good safety in inhibiting SARS-CoV-2 at the cellular level.

[0068] 4. Cell-cell fusion inhibition assay

[0069] Caco-2 target cells were cultured at 10 4 The cells were plated in 96-well plates and cultured at 37°C overnight. Effector cells (293T / EGFP / D614G cells) expressing EGFP and SARS-CoV-2 D614G S protein were cultured at a rate of 2×10 4 After incubating each well with varying concentrations of A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) at 37°C for 30 minutes, the effector cells and peptide inhibitor mixture were transferred to target Caco-2 cells. After incubation at 37°C in a CO2 incubator for 4-5 hours, the cells were examined for fusion under a fluorescence microscope. If fusion was evident, the cells were fixed with 4% paraformaldehyde. Finally, the fusion inhibitory activity of the peptide was calculated based on the fusion status of each well.

[0070] The results are as follows Figure 6 As shown, A1L35HR2, A3M6L35HR2 (FL), and A3M6L35HR2 (HL2) inhibited S protein-mediated cell-cell fusion at 66, 7, and 180 nM, respectively.

[0071] 5. Biofilm interferometry (BLI) affinity detection

[0072] The A3M6L35HR2 (FL) peptide was biotinylated according to the instructions for use with EZ-Link NHS-PEG12-Biotin at a molar ratio of 1:3 (peptide:biotin). Unreacted biotin was then removed by ultrafiltration through a 3 kDa filter. The biotinylated A3M6L35HR2 (FL) peptide was diluted to 5 μg / ml and the HR1 peptide to 200 nM using BLI buffer (0.02% Tween 20 in PBS). Two-fold serial dilutions were then used for detection. Data were curve-fitted using ForteBio software, and k values were calculated. on 、k off and K D and other related parameters.

[0073] like Figure 7 As shown, A3M6L35HR2 (FL) has a strong interaction with HR1 peptide, with an affinity of 0.03 nM.

[0074] 6. Circular dichroism analysis

[0075] Circular dichroism (CD) spectroscopy can be used to determine the secondary conformation of peptides and the changes in secondary conformation that occur after interactions between peptide samples. Peptides and peptide mixtures are diluted to 10 μM in PB buffer and the peptide complexes are incubated at 37°C for 30 minutes. Peptide samples are placed in 0.1 cm quartz cuvettes and CD values are measured at 25°C within the wavelength range of 180-260 nm.

[0076] like Figure 8 As shown, the single A3M6 and HR1 peptides exhibit random coils, while the single HR2 and A3M6L35HR2 (FL) peptides exhibit double negative peaks at 208 nm and 222 nm, indicating that both peptides possess a certain amount of α-helical conformation. Furthermore, when HR2 and A3M6L35HR2 (FL) are mixed with the HR1 peptide, the negative peaks at 208 nm and 222 nm of the peptide complexes significantly deepen, indicating that the complexes have a higher α-helical content. This indicates that the HR2 and A3M6L35HR2 (FL) peptides can interact with the HR1 peptide, inducing the formation of a large amount of α-helical conformation.

Claims

1. A recombinant polypeptide that broadly inhibits the novel coronavirus SARS-CoV-2, characterized in that: The recombinant polypeptide comprises the polypeptide A3M6 derived from ACE2 and the polypeptide HR2 derived from the HR2 region of the novel coronavirus SARS-CoV-2; The amino acid sequence of the polypeptide A3M6 is IEEQVEWFLDKFDIQAEDLFYQSSLGGSKGDFR, The amino acid sequence of the polypeptide HR2 is DVDLGDISGINASVVNIQKEIDRLNEVAKNLNESLIDLQELGKYEQ; The recombinant polypeptide comprises polypeptide A3M6, linker, and polypeptide HR2 from N-terminus to C-terminus; The linker is a flexible linker; The amino acid sequence of the flexible linker is (GGGGS) n , where n=7.

2. A nucleic acid molecule, characterized in that The nucleic acid molecule encodes the recombinant polypeptide according to claim 1.

3. A recombinant vector, characterized in that The recombinant vector comprises the nucleic acid molecule according to claim 2.

4. A host cell, characterized in that The host cell comprises the recombinant vector according to claim 3.

5. Use of the recombinant polypeptide according to claim 1, the nucleic acid molecule according to claim 2, the recombinant vector according to claim 3, or the host cell according to claim 4 in the preparation of a medicament for preventing or treating diseases caused by coronavirus infection; The coronavirus is the novel coronavirus SARS-CoV-2 and / or a variant strain of the novel coronavirus SARS-CoV-2; The novel coronavirus SARS-CoV-2 variant is at least one of the D614G, Beta, Delta and Omicron series variants; The Omicron series variants are XBB, BF7, BA.2, EG.5.1, BA.2.86 or KP.

2.

6. The use according to claim 5, characterized in that The disease caused by the coronavirus infection is a respiratory system infection.

7. A pharmaceutical composition, characterized in that The pharmaceutical composition contains the recombinant polypeptide according to claim 1, the nucleic acid molecule according to claim 2, the recombinant vector according to claim 3 or the host cell according to claim 4, and a pharmaceutically acceptable carrier.

Images