WO2019029081A1 - 干扰素κ在制备抗囊膜病毒药物方面的应用 - Google Patents

干扰素κ在制备抗囊膜病毒药物方面的应用 Download PDF

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WO2019029081A1
WO2019029081A1 PCT/CN2017/116350 CN2017116350W WO2019029081A1 WO 2019029081 A1 WO2019029081 A1 WO 2019029081A1 CN 2017116350 W CN2017116350 W CN 2017116350W WO 2019029081 A1 WO2019029081 A1 WO 2019029081A1
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徐建青
张晓燕
傅卫辉
陈健
何涌泉
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上海市公共卫生临床中心
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    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
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    • A61K38/19—Cytokines; Lymphokines; Interferons
    • A61K38/21—Interferons [IFN]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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  • the invention belongs to the field of biomedical technology, and particularly relates to the application of interferon-kappa (IFN- ⁇ ) in the preparation of anti-enveloped virus drugs.
  • IFN- ⁇ interferon-kappa
  • IFN-I type I interferon
  • HBV anti-hepatitis B virus
  • hepatitis C virus A therapeutic drug for HCV infection, IFN- ⁇ is used to treat multiple sclerosis (MS).
  • IFN- ⁇ can activate the expression of antiviral factors by interacting with the IFN receptor (IFNR) 1/2.
  • IFN- ⁇ The mechanism of IFN- ⁇ may be that IFN- ⁇ binds to the receptor and stimulates phosphorylation of tyrosine kinase 2 (Tyk2) and Janus kinase 1 (Jak1), further leading to signaling and transcriptional activator 1/2 (STAT1/2). Phosphorylation and dimerization, up-regulation of the expression of the interferon-stimulated gene (ISG). These genes regulate a wide range of cellular responses, including antiviral effects, anti-tumor effects, enhanced NK cell activity, and activation of adaptive immune responses.
  • Tyk2 tyrosine kinase 2
  • Jak1 Janus kinase 1
  • STAT1/2 signaling and transcriptional activator 1/2
  • ISG interferon-stimulated gene
  • IFN- ⁇ is selectively expressed in epithelial keratinocytes after viral infection or double-stranded RNA stimulation, or treatment with IFN- ⁇ and IFN- ⁇ , activating protein kinase R (PKR), 2,5-oligo Adenylate synthase (OAS), and interferon-induced GTP-binding protein (MxA) and other antiviral gene expression, thereby inhibiting the replication of encephalomyocarditis virus (ECMV) and human papillomavirus (HPV).
  • PSR protein kinase R
  • OFAS 2,5-oligo Adenylate synthase
  • MxA interferon-induced GTP-binding protein
  • ECMV and HPV are non-enveloped viruses, their replication mechanism is inconsistent with enveloped virus, and non-enveloped viruses generally enter cells through receptor-mediated endocytosis, while enveloped viruses pass through the cell membrane or endosomal membrane. Fusion, the viral capsid and nucleic acid are introduced into the cell for replication, and therefore, it cannot be concluded whether IFN- ⁇ can also inhibit the replication of the envelope virus.
  • influenza virus and Zika virus caused by the influenza virus and Zika virus in the envelope virus have caused significant public health security risks and also placed a heavy burden on social and economic development. There is still no effective vaccine to prevent infection. It has been reported in the literature that influenza virus can block the IFN-I signaling pathway by binding to IFN upstream and downstream regulatory factors, thereby escaping the antiviral effect of IFN-I, while Zika virus can block STAT1 and STAT2 phosphorylation. Escape the antiviral effect of IFN-I, other antiviral small molecule drugs can effectively improve the prognosis of patients, but there are virus resistance mutations under drug selection pressure, and some primary infections caused by drug-resistant virus transmission are difficult to select. risks of. Therefore, there is an urgent need to develop new antiviral strategies and therapeutic drugs.
  • the present invention mainly finds that IFN- ⁇ inhibits the replication of various envelope viruses, including influenza virus and Zika virus, and exhibits broad-spectrum antiviral effects.
  • the present invention will focus on the inhibitory function of IFN- ⁇ on envelope virus replication.
  • the technical problem solved by the present invention is that the IFN- ⁇ is used to stimulate the host's resistance, thereby inhibiting the infection and replication of the envelope virus, and can be used for preparing an anti-enveloped virus drug.
  • the technical solution adopted by the present invention is:
  • an interferon ⁇ for use in the preparation of an anti-enveloped virus drug
  • the source of IFN- ⁇ comprising human, murine and other mammalian sources
  • the nucleotide sequence of the interferon kappa encoding gene is SEQ ID No
  • the amino acid sequence is shown in SEQ ID No. 2.
  • the envelope virus includes, but is not limited to, influenza virus (H7N9, PR8 and H9N2) and Zika virus; antiviral drugs prepared by using IFN- ⁇ .
  • Prophylactic administration before infection can be used to prevent the severity of viral infection; it can also be administered as a therapeutic drug after viral infection to reduce the severity of viral infection; it can also be given continuously or intermittently from before infection to infection. medicine.
  • the present invention has the beneficial effects that the present invention finds a new use of IFN- ⁇ , which can inhibit envelope virus, such as influenza virus (including H1N1, H7N9 and H9N2), and the like.
  • envelope virus such as influenza virus (including H1N1, H7N9 and H9N2), and the like.
  • Zika virus has important clinical application value for the prevention and treatment of new infectious diseases such as highly pathogenic influenza virus and Zika virus.
  • Figure 1 shows the construction of an IFN- ⁇ eukaryotic expression vector and the expression of IFN- ⁇ protein.
  • a pSV1.0-IFN- ⁇ eukaryotic expression vector plasmid construction map;
  • b IFN- ⁇ protein was successfully expressed in 293T cells and cell supernatant.
  • FIG. 2 shows that IFN- ⁇ inhibits replication of influenza virus PR8, H9N2 and H7N9 in the human non-small cell lung cancer epithelial cell line A549.
  • a Western blot (WB) results show that overexpression of IFN- ⁇ can completely inhibit the expression of PR8 (H1N1) influenza virus nucleoprotein NP and M1 protein in A549 cells;
  • b Western blot (WB) results show overexpression of IFN - ⁇ can completely inhibit the expression of H9N2 influenza virus nucleoprotein NP and M1 protein in A549 cells;
  • c Western blot (WB) results show that overexpression of IFN- ⁇ can inhibit H7N9 influenza virus nucleoprotein NP and M1 protein in A549 cells Expression in.
  • FIG. 3 shows that IFN- ⁇ inhibits Zika virus replication in glial cell U-251.
  • WB Western blot
  • a Western blot
  • WB Western blot
  • b Western blot
  • results showed IFN- ⁇ overexpression Plasmid (left) and cell supernatant (right) inhibited the expression of Zika virus non-structural proteins (NS2b, NS3, NS5) in U-251 cells
  • c immunofluorescence statistical analysis showed overexpression of IFN- ⁇ plasmid Inhibition of Zika virus replication in U-251 cells
  • d immunofluorescence statistical analysis showed that 293T cell supernatant overexpressing IFN- ⁇ could inhibit the replication of Zika virus in U-251 cells.
  • Figure 4 shows that IFN- ⁇ upregulates RNA and protein expression levels of IFITM3.
  • a qPCR detection of IFN- ⁇ induced up-regulation of IFITM3 RNA;
  • b Western blot (WB) detection of IFN- ⁇ significantly up-regulated IFITM3 protein expression.
  • NC is an abbreviation of Native Control and represents the control group.
  • the present invention clones IFN- ⁇ from the human genome, the nucleotide sequence of the coding gene is shown in SEQ ID No. 1, and the full-length amino acid sequence is shown in SEQ ID No. 2, and the IFN- ⁇ of the present invention. It belongs to the type I interferon family, which has only 30% homology with IFN- ⁇ and IFN- ⁇ .
  • IFN- ⁇ In order to study the function of IFN- ⁇ , we constructed an eukaryotic expression vector of IFN- ⁇ and expressed the mature secreted protein in eukaryotic cells in vitro, and then detected the replication of IFN- ⁇ protein against influenza virus and Zika virus. influences.
  • IFN- ⁇ eukaryotic expression plasmid using the eukaryotic expression vector pSV1.0. The construction method was as follows: the cDNA generated by reverse transcription of RNA extracted from A549 cells was used as a template, and the corresponding primers were used for PCR amplification.
  • the target gene was isolated on a 1% agarose gel, and the gel was recovered.
  • the PCR fragment was recovered by the Sanprep column DNA gel recovery kit, and the IFN- ⁇ recovery product and the pSV1.0 vector were used. Digestion of BamH I and Xba I was digested, and the fragment was ligated with the vector at 4 ° C overnight using T4 DNA ligase. The ligated product was transformed into E. coli TOP10 on day 2, in which kanamycin was contained. The plates were grown overnight. On the third day, single colonies were randomly picked for PCR identification, and positive clones were selected and identified by double enzyme digestion.
  • Example 2 IFN- ⁇ inhibits replication of influenza virus H7N9, PR8 and H9N2
  • the lung epithelial cell line A549 is derived from human non-small cell lung cancer epithelial cells and is the main cell model for studying influenza virus infection.
  • this example uses a 12-well plate.
  • the pSV1.0-IFN- ⁇ overexpression plasmid and the control plasmid pSV1.0-GFP were transfected into A549 cell line.
  • three influenza virus PR8 (Fig. 2a) and H9N2 diluted in 100 ⁇ L of serum-free medium were added respectively.
  • Fig. 2b the average number of particles (MOI) per virus infected with each cell was 1.
  • the primary antibodies used were Anti-Mx1 mAb (ab22396, 1:1000), anti-NP (bs-4976R, 1:1000), anti-IFN- ⁇ (H00056832-M01, 1:1000), and anti- ⁇ . -Actin (sc-69879, 1:1000).
  • the secondary antibodies were HRP-labeled goat anti-mouse (zb-2305, 1:3000) and HRP-labeled goat anti-rabbit (zb-2301, 1:3000).
  • Example 3 IFN- ⁇ inhibits the replication of Zika virus
  • Astrocyte U-251 cells are one of the main target cells of Zika virus infection.
  • MOI multiplicity of infection
  • Zika virus non-structural proteins NS2b, NS3, NS5
  • the Zika virus E protein was immunofluorescently stained, and the proportion of positive cells infected with Zika virus was statistically analyzed (Fig. 3d).
  • the dilution ratio of all primary antibodies was 1:1000, and the dilution ratio of HRP-labeled goat anti-rabbit secondary antibody was 1:3000.
  • the immunofluorescence primary antibody was a murine anti-flavivirus envelope protein antibody (D1-4G2-4-15, 1:200), and the secondary antibody was Alexa Fluor 568-labeled donkey anti-mouse IgG (ab175472, 1:1000).
  • IFN- ⁇ can significantly inhibit the replication of Zika virus.
  • IFN- ⁇ upregulates the RNA level of the antiviral gene IFITM3 (Fig. 4a) and protein levels (Fig. 4b).
  • the IFITM3 protein can significantly inhibit the replication of Zika virus.

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Abstract

本发明公开了干扰素κ（IFN-κ）在制备抗囊膜病毒药物中的应用。所述IFN-κ编码基因的核苷酸序列如SEQ ID NO:1所示，氨基酸序列如SEQ ID NO:2所示，所述囊膜病毒包括但不限于流感和寨卡病毒。本发明发现IFN-κ诱导抗病毒蛋白IFITM3的表达，进而抑制流感病毒和寨卡病毒的感染和复制。

Description

干扰素κ在制备抗囊膜病毒药物方面的应用 技术领域
本发明属生物医学技术领域，具体涉及干扰素-kappa(IFN-κ)在制备抗囊膜病毒药物方面的应用。
背景技术
I型干扰素(IFN-I)家族的多位成员，作为抗病毒药物候选，已经完成临床的药物试验，如重组IFN-α2被应用于抗乙型肝炎病毒(HBV)和丙型肝炎病毒(HCV)感染的治疗药物，IFN-β被应用于治疗多发性硬化症(MS)。I型干扰素家族的另外一位相对较新的成员，于2001年报告的IFN-κ可通过与IFN受体(IFNR)1/2相互作用，激活抗病毒因子的表达。IFN-κ作用机制可能为IFN-κ与受体结合后，刺激酪氨酸激酶2(Tyk2)和Janus激酶1(Jak1)磷酸化，进一步导致信号传导及转录激活因子1/2(STAT1/2)的磷酸化和二聚化，上调干扰素刺激基因(ISG)的表达。这些基因调节广泛的细胞反应，包括抗病毒效应，抗肿瘤效应，增强NK细胞活性以及激活适应性免疫反应。研究表明，在病毒感染或双链RNA刺激，或用IFN-γ和IFN-β处理后，IFN-κ选择性地表达在上皮角质细胞中，活化蛋白激酶R(PKR)，2，5-寡腺苷酸合成酶(OAS)，和干扰素诱导的GTP结合蛋白(MxA)等抗病毒基因表达，从而抑制脑心肌炎病毒(ECMV)和人类乳头瘤病毒(HPV)的复制。然而，由于ECMV和HPV为无囊膜病毒，其复制机理与囊膜病毒并不一致，无囊膜病毒一般通过受体介导的内吞作用进入细胞，而囊膜病毒通过与细胞膜或内体膜融合，将病毒衣壳和核酸导入细胞中进行复制，因此，无法断定IFN-κ是否同样可以抑制囊膜病毒的复制。
囊膜病毒中的流感病毒和寨卡病毒(ZIKV)导致的流感和寨卡疫情造成了重大的公共卫生安全危险，也给社会经济发展造成沉重的负担，目前仍然缺乏有效的疫苗预防感染。已有文献报道，流感病毒可通过与IFN上、下游调节因子结合阻断IFN-I信号通路，从而逃逸IFN-I的抗病毒作用，而寨卡病毒则可通过阻断STAT1和STAT2磷酸化来逃脱IFN-I的抗病毒作用，其他抗病毒小分子药物能有效的改善患者预后，但存在药物选择压力下病毒耐药性突变、以及耐药性病毒传播而导致的部分初次感染患者用药选择困难的风险。因此，迫切需要开发新型抗病毒策略和治疗药物。
本发明主要发现IFN-κ可抑制多种囊膜病毒的复制，包括流感病毒、寨卡病毒，显示出广 谱抗病毒效果。本发明将着重阐述IFN-κ对囊膜病毒复制的抑制功能。
发明内容
本发明所解决的技术问题是：利用IFN-κ激发宿主的抵抗力，从而抑制囊膜病毒的感染和复制，可用于制备抗囊膜病毒药物。
为了解决上述技术问题，本发明所采用的技术方案是：
提供一种干扰素κ在制备抗囊膜病毒药物方面的应用，IFN-κ的来源包括人源、鼠源及其他哺乳动物来源；所述干扰素κ编码基因的核苷酸序列如SEQ ID No.1所示，氨基酸序列如SEQ ID No.2所示.所述的囊膜病毒包括但不仅限于流感病毒(H7N9，PR8和H9N2)和寨卡病毒；利用IFN-κ所制备的抗病毒药物可在感染前预防性给药用以预防病毒感染的严重程度；也可在病毒感染后作为治疗性药物给药以降低病毒感染的严重程度；也可自病毒感染前至感染后连续或间隔给药。
本发明的有益效果在于：与现有技术相比，本发明发现了IFN-κ的新用途，本发明所述IFN-κ能抑制囊膜病毒，如流感病毒(包括H1N1，H7N9和H9N2)和寨卡病毒的复制，对于预防与治疗高致病流感病毒与寨卡病毒等新发突发传染病有重要的临床应用价值。
附图说明
图1显示了IFN-κ真核表达载体的构建及IFN-κ蛋白的表达。a，pSV1.0-IFN-κ真核表达载体质粒构建图谱；b，IFN-κ蛋白在293T细胞和细胞上清中成功表达。
图2显示了IFN-κ抑制流感病毒PR8，H9N2和H7N9在人非小细胞肺癌上皮细胞系A549中复制。a，蛋白质免疫印迹(WB)结果显示过表达IFN-κ能完全抑制PR8(H1N1)流感病毒核蛋白NP和M1蛋白在A549细胞中的表达；b，蛋白质免疫印迹(WB)结果显示过表达IFN-κ能完全抑制H9N2流感病毒核蛋白NP和M1蛋白在A549细胞中的表达；c，蛋白质免疫印迹(WB)结果显示过表达IFN-κ能抑制H7N9流感病毒核蛋白NP和M1蛋白在A549细胞中的表达。
图3显示了IFN-κ抑制寨卡病毒在神经胶质细胞U-251中复制。a，蛋白质免疫印迹(WB)显示转染了IFN-κ质粒的U-251细胞和293T细胞上清都表达大量的IFN-κ蛋白；b，蛋白质免疫印迹(WB)结果显示IFN-κ过表达质粒(左)和细胞上清(右)能抑制寨卡病毒非结构蛋白(NS2b，NS3，NS5)在U-251细胞中的表达；c，免疫荧光统计分析结果显示过表达IFN-κ 质粒能抑制寨卡病毒在U-251细胞中复制；d，免疫荧光统计分析结果显示过表达IFN-κ的293T细胞上清能抑制寨卡病毒在U-251细胞中复制。
图4显示了IFN-κ上调IFITM3的RNA和蛋白表达水平。a，qPCR检测IFN-κ诱导IFITM3RNA水平上调；b，蛋白质免疫印迹(WB)检测IFN-κ显著上调IFITM3的蛋白表达。NC是Native Control的缩写，代表的是对照组。
具体实施方式
实施例一：pSV1.0-IFN-κ过表达质粒的构建
本发明从人基因组中克隆获得了IFN-κ，其编码基因的核苷酸序列如SEQ ID No.1所示，全长氨基酸序列如SEQ ID No.2所示，本发明所述IFN-κ属于I型干扰素家族，其与IFN-α和IFN-β仅有30％的同源性。
为了研究IFN-κ的功能，我们构建了IFN-κ的真核表达载体，并在体外细胞系中真核表达出成熟的分泌蛋白，然后检测IFN-κ蛋白对流感病毒和寨卡病毒复制的影响。首先，我们采用真核表达载体pSV1.0构建IFN-κ真核表达质粒，构建方式如下：以A549细胞抽提的RNA反转录生成的cDNA为模板，采用相应的引物进行PCR扩增。
PCR反应程序：
Figure PCTCN2017116350-appb-000001
扩增结束后，在1％的琼脂糖凝胶中分离目的基因，切胶回收，采用Sanprep柱式DNA胶回收试剂盒进行PCR片断回收，将IFN-κ回收产物与pSV1.0载体都采用内切酶BamH I、Xba I双酶切，酶切后再采用T4 DNA连接酶将片段和载体4℃连接过夜，第2天将连接产物转化至大肠杆菌E.coli TOP10，在含卡那霉素的培养板上过夜生长。第3天，随机挑取单菌落进行PCR鉴定，挑选阳性克隆再进行双酶切鉴定。再经过测序，突变位点校正，验证全部序列正确后，成功克隆出IFN-κ基因，质粒构建图谱如图1a。我们进一步检测了IFN-κ能否 表达且被分泌到细胞上清中，我们收集了pSV1.0-IFN-κ质粒转染的细胞和上清，进行蛋白质免疫印迹(WB)鉴定，发现IFN-κ在细胞和上清中都有表达，而pSV1.0对照转染的细胞和上清不能检测到IFN-κ蛋白的表达(图1b)。
实施例二：IFN-κ抑制流感病毒H7N9，PR8和H9N2的复制
肺上皮细胞系A549来源于人非小细胞肺癌上皮细胞，是用于研究流感病毒感染的主要细胞模型，为了验证IFN-κ对于不同亚型流感病毒感染的作用，本实施例采用12孔板在A549细胞系中转染pSV1.0-IFN-κ过表达质粒和对照质粒pSV1.0-GFP，24h后，分别加入100μL不含血清的培养基稀释的三种流感病毒PR8(图2a)、H9N2(图2b)、以及H7N9(图2c)，平均每个细胞感染病毒的颗粒数(MOI)均为1。在CO 2孵箱中继续孵育2h后弃去病毒液，采用PBS洗2遍，加入DMEM完全培养基继续培养48h后收取细胞，通过蛋白质免疫印迹法分析IFN-κ表达和流感病毒核蛋白NP和M1的表达。所用的一抗分别为Anti-Mx1 mAb(ab22396，1∶1000)，anti-NP(bs-4976R，1∶1000)，anti-IFN-κ(H00056832-M01，1∶1000)，与anti-β-Actin(sc-69879，1∶1000)。二抗分别为HRP标记山羊抗小鼠(zb-2305，1∶3000)，HRP标记山羊抗兔(zb-2301，1∶3000)。
结果显示，转染对照载体pSV1.0-GFP 24h后，PR8能感染A549细胞，表达流感病毒NP和M1蛋白，而当转染IFN-κ表达载体pSV1.0-IFN-κ 24h后，A549细胞表达IFN-κ蛋白，然而流感病毒NP和M1蛋白不表达，说明IFN-κ能抑制流感病毒蛋白NP和M1的表达(图2a)。另外，采用同样的实验方法证实，IFN-κ也能抑制流感病毒H9N2和H7N9病毒蛋白NP和M1的表达，显示了广谱抗流感病毒复制的效果(图2b-2c)。
实施例三：IFN-κ抑制寨卡病毒的复制
星形胶质细胞U-251细胞，是寨卡病毒感染主要靶细胞之一。为了验证IFN-κ对寨卡病毒感染的作用，采用U-251细胞转染对照质粒pSV1.0和pSV1.0-IFN-κ质粒，转染36h后，收取部分孔的细胞，检测细胞中IFN-κ蛋白表达水平(图3a-左)。其余细胞感染寨卡病毒，感染复数(MOI＝2)，37℃感染2h，PBS洗2次，更换新鲜的含有2％FBS的DMEM培养基继续培养。寨卡病毒感染36h后，收取细胞，一部分细胞用于蛋白质免疫印迹(WB)分析寨卡病毒非结构蛋白(NS2b，NS3，NS5)在U-251细胞中的表达水平(图3b-左)。另一部分细胞采用免疫荧光染色技术，分析寨卡病毒包膜(E)蛋白表达水平，三次重复用于统计分析寨卡病毒感染的阳性细胞比例(图3c)。为了在蛋白水平验证IFN-κ蛋白对寨卡病毒复制的作用，我们在293T细胞中转染对照质粒pSV1.0和pSV1.0-IFN-κ质粒，36h后其细胞上清中 IFN-κ的表达如图3a-右，将400μL细胞上清加入预先铺好的U-251细胞中，37℃孵育3h，移除400μL上清，再加入寨卡病毒感染U-251细胞，MOI＝2，37℃感染2h，更换新鲜的含有2％FBS的DMEM培养基。寨卡病毒感染36h后，收取细胞，蛋白质免疫印迹(WB)分析寨卡病毒非结构蛋白(NS2b，NS3，NS5)在U-251细胞中的表达水平(图3b-右)。免疫荧光染色寨卡病毒E蛋白，统计分析寨卡病毒感染的阳性细胞比例(图3d)。所有一抗稀释比例为1∶1000，HRP标记山羊抗兔二抗稀释比例为1∶3000。免疫荧光一抗为鼠抗黄病毒包膜蛋白抗体(D1-4G2-4-15，1∶200)，二抗为Alexa Fluor 568标记的驴抗鼠IgG(ab175472，1∶1000)。
结果显示，过表达pSV1.0-IFN-κ质粒，利用蛋白质免疫印迹方法能检测到高表达的IFN-κ蛋白(图3a)，而IFN-κ的表达对寨卡病毒非结构蛋白NS2b，NS3和NS5的表达都有明显抑制作用(图3b)。采用免疫荧光技术发现，在寨卡病毒感染后，与对照组比较，IFN-κ过表达质粒抑制了92％的寨卡病毒包膜蛋白表达(图3c)。我们接下来利用真核表达系统293T细胞过表达IFN-κ蛋白，与对照上清相比，IFN-κ过表达的293T细胞上清抑制40％寨卡病毒包膜蛋白表达(图3d)。这些结果证实IFN-κ能显著抑制寨卡病毒的复制。另外，我们初步探究了IFN-κ抑制病毒复制的机制，发现IFN-κ能上调抗病毒基因IFITM3的RNA水平(图4a)和蛋白水平(图4b)。而IFITM3蛋白能显著抑制寨卡病毒的复制。
以上显示和描述了本发明的基本原理、主要特征和本发明的优点。本行业的技术人员应该了解，本发明不受上述实施例的限制，上述实施例和说明书中描述的只是说明本发明的原理，在不脱离本发明精神和范围的前提下，本发明还会有各种变化和改进，这些变化和改进都落入要求保护的本发明范围内。本发明要求保护范围由所附的权利要求书及其等效物界定。

Claims (11)

  1. 干扰素κ(IFN-κ)在制备抗囊膜病毒药物方面的应用。
  2. 根据权利要求1所述的应用，其特征在于，所述的IFN-κ来源包括人源、鼠源及其他哺乳动物来源。
  3. 根据权利要求1所述的应用，其特征在于，人IFN-κ编码基因的核苷酸序列如SEQ ID No.1所示，氨基酸序列如SEQ ID No.2所示。
  4. 根据权利要求1所述的应用，其特征在于，所述的制备抗囊膜病毒药物包括含有权利要求4所述基因序列的各种载体与权利要求4所述序列编码的蛋白。
  5. 权利要求5中所述的“含有权利要求4所述基因序列的各种载体”包括但不限于质粒载体、痘病毒载体、腺病毒载体、腺相关病毒载体、单纯病毒载体、CMV载体、细胞载体、细菌载体等。
  6. 权利要求6中所述的“权利要求4所述序列编码的蛋白”包括单独表达的蛋白与融合表达的蛋白，融合表达的方式包括但不限于与抗体的Fc序列融合、与不同信号肽的融合等。
  7. 权利要求5、6中所述的蛋白可以进一步修饰，包括但不限于PEG修饰。
  8. 权利要求4、5、6、7所述的抗囊膜病毒药物可单独使用，也可与其他抗病毒药物或技术联合使用。
  9. 根据权利要求1所述应用，其特征在于：所述的囊膜病毒包括但不仅限于流感病毒和寨卡病毒。
  10. 根据权利要求4所述应用，其特征在于：所述的流感病毒包括但不限于H7N9，PR8和H9N2。
  11. 根据权利要求1所述应用，其特征在于：利用IFN-κ所制备的抗病毒药物可在感染前预防性给药用以预防病毒感染的致病作用；也可在病毒感染后作为治疗性药物给药清除病毒；也可自病毒感染前至感染后连续或间隔给药。
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