-
摘要: 单纯疱疹病毒1型（herpes simplex virus type 1，HSV-1）是一种能够在各类人群中携带和传播并能引起包括口唇疱疹、荚膜炎、角膜炎和病毒性脑炎等疾病的重要病原体。虽然已有多种类型的HSV-1疫苗处于研发的不同阶段，但仍没有商业化的疫苗上市销售。临床上使用的特异性抗HSV-1药物如阿昔洛韦、伐昔洛韦和喷昔洛韦等也面临严重的抗药性威胁，开发新的特异性抗HSV-1药物是当前所面临的主要任务之一。siRNA是一种长度为20～25 核苷酸的双链RNA，通过在转录后水平上沉默基因表达发挥干扰作用。siRNA作为一种新的、有潜力的抗病毒药物备受关注，发展也较为迅速。综述近年来siRNA在抗HSV-1方面的研究进展，包括靶向HSV-1关键基因和HSV-1互作的宿主细胞基因的siRNA设计、递送和靶向策略。Abstract: HSV-1 is an important pathogen that can be carried and transmitted in various populations and can cause diseases including herpes labialis, capsulatus, keratitis and viral encephalitis. Although there are several types of HSV-1 vaccines in various stages of development, there is still no commercially available vaccine on the market. The specific anti-HSV-1 drugs used in clinical practice, such as acyclovir, valaciclovir and peniclovir, are also facing the serious threat of resistance. The development of new specific anti-HSV-1 drugs is one of the main tasks currently faced. siRNA is a double-stranded RNA with a length of 20-25 nucleotides that plays an interfering role by silencing gene expression at the post-transcriptional level. As a new and potential antiviral drug, siRNA has attracted much attention and developed rapidly. In this paper, we review the recent progress of siRNA in anti-HSV-1 research, including the design, delivery and targeting strategies of siRNA targeting key HSV-1 genes and HSV-1 interacting host cell genes.
-
Key words:
- siRNA /
- HSV-1 /
- Research Progress
-
单纯疱疹病毒1型（herpes simplex virus type 1，HSV-1）在世界范围内普遍流行，引起临床上常见的口唇疱疹 （herpes labialis，HL）并反复发作，由于病毒感染后终身携带以及抗病毒药物的耐用性、副作用等问题而导致疾病难以完全治疗。1995年小干扰RNA（small interfering RNA，siRNA）的发现为治疗HSV-1感染提供了基因层面的新策略，人工合成的siRNA通过特异性沉默效应靶向与HSV-1增殖有关的自身或宿主基因，进而影响病毒增殖。研究表明，基于RNA干扰（RNA interference，RNAi）的抗HSV-1疗法可能发展为一种有效的临床治疗手段。
1. HSV-1生物学
HSV-1是一种广泛存在于世界范围内的常见病毒，具有普遍性、潜伏感染和年龄分布的特征，其感染率在不同地区和人群中大有不同[1−2]。全球每年大约14 000例新生儿感染HSV，其中HSV-1比例占到了
4000 例，尤其在美洲、欧洲和西太平洋地区，HSV-1 的病例数比HSV-2多[3]。即使HSV-1的血清阳性率逐年降低，但是在青年人中HSV-1的血清阳性率仍达到了57.7%[4]。据世界卫生组织（World Health Organization，WHO）最新数据显示，全世界0～49岁中约有67%的人群受到了HSV-1的感染。HSV-1的普遍流行使世界医疗负担加重，在全球范围内都有着重要的健康影响，自新型冠状病毒（Corona Virus Disease 2019，COVID-19）爆发以来，有报道称感染或接种 COVID-19疫苗诱发了 HSV-1 的再激活现象[5]。HSV-1大多数的感染是通过口唇接触传播而引起不同程度的HL。感染宿主外周组织后，一部分病毒便会通过逆行轴突感染神经元，其最大特征是进行潜伏性感染，人三叉神经节是HSV-1主要的潜伏部位，一旦潜伏建立病毒便难以被清除。HSV-1感染通常表现为轻度的疱疹症状，但在免疫系统受损或其他因素影响下，可能导致严重的并发症[6]。HSV-1感染会引起严重和罕见的疾病，如失明和脑炎[7]。反复的脑感染也可积累性的造成神经元受损，如阿尔兹海默症（Alzheimer's disease，AD）[8]。尽管目前市面上具有有效的传统抗HSV-1药物可以减轻症状，例如阿昔洛韦（Acyclovir）和喷西洛韦（Penciliovir），该类核苷类药物虽能抑制HSV-1的复制，但在长期的治疗中，存在耐药性和副作用等问题[9−12]。多款用于预防HSV-1感染的疫苗也大多数处于不同的研发阶段，还没有批准上市的产品可供人们使用[13]。因此，寻找新的治疗策略显得尤为重要。
2. RNAi技术
近年来，RNA干扰（RNA interference，RNAi）已经被广泛用于各类口腔疾病、变异体调控以及病毒治疗等方面[14−16]。通过序列特异性方式可沉默与疾病相关基因，使小干扰RNA（small interfering RNA，siRNA）成为一种有前途的治疗方式。siRNA是人工合成的外源双链反义RNA（double-stranded antisense RNA，dsRNA），是一类独特的DNA转录物，其大小为21～23个核苷酸，具有与mRNA完全互补的特性。siRNA通过RNA干扰与同源靶基因结合促进mRNA降解，在转录后的表达水平上进行高效特异性沉默。1995年，RNAi在秀丽隐干线虫中被证明参与基因调控表达[17]；随后Fire等[18]首次揭示RNAi现象并因此获得了诺贝尔奖，2001年，在果蝇模型中Elbashir等[19]表明具有21nt至22nt的RNA是实现RNAi的重要序列特异性结构，siRNA具有突出的3′ 末端和磷酸化的5′ 端，其实现高效切割的位点位于其所结合区域的中心附近。更重要的是，合成的siRNA在哺乳动物细胞中可实现转录后基因特异性沉默[20]，该项研究推动了siRNA在哺乳动物中的应用奠定了基础。
2.1 siRNA的作用机制
通常情况下，dsRNA通过跨膜蛋白SID-1转移至细胞质，在细胞质中被Dicer核糖核酸酶III （dicer ribonucleaseIII，RNaseIII）切割成更小的dsRNA分子，被称为siRNA，它包含一条正义链和一条反义链[21]。siRNA反义链是具有生物学活性的链，成熟的siRNA与RNA沉默诱导复合体（RNA silencing induction complex，RISC）结合并激活该复合物，随后RISC的核酸内切酶（argonaute 2，AGO2）切割siRNA的正义链，而将反义链运输到细胞质中与mRNA结合，由于靶mRNA的序列与siRNA反义链是完全互补的，导致siRNA高效特异性沉默靶基因[22−23]。DNA转录成dsRNA，通过SID-1转移至细胞质，在其中被Dicer切割成初级双链siRNA，成熟的siRNA与RISC结合形成复合物，通过反义链与靶mRNA序列结合导致mRNA降解。见图1。
3. 靶向HSV-1编码基因的siRNA
近年来，siRNA作为一种新的抗HSV-1策略得到了广泛的研究。siRNA可以靶向HSV-1的基因发挥直接抗病毒效应。同时，靶向与HSV-1互作的宿主基因能够实现间接抗病毒效应。
3.1 靶向HSV-1编码衣壳蛋白基因的siRNA
人工合成的siRNA是潜在的HSV-1基因治疗药物，由于其特异靶向性、低毒和高效而成为人们广泛关注的焦点[24]。Jin等[25]设计多个siRNA分别靶向HSV-1的UL18、UL19、UL26、UL26.5、UL35和UL38核衣壳蛋白编码基因，研究结果显示UL18和UL19效果显著。此外，他们还通过电镜定位实验表明，联合靶向病毒衣壳蛋白VP 23和VP 5能更显著影响HSV-1复制。
3.2 靶向HSV-1编码α基因的siRNA
感染细胞蛋白0（infected cell protein 0，ICP0）是单纯疱疹病毒在裂解性和潜伏性感染期间的重要调节因子，在缺乏ICP0蛋白的病毒中，发现宿主细胞能够抑制病毒基因组的转录。Agbaria等[26]将siRNA做成靶向HSV-1 ICP0蛋白的脂质体LipDOPE-siHSV（DOPE是内体逃逸辅助脂质）并转染受感染细胞，研究表明ICP0的表达水平呈剂量依赖性下降。使用siHSV脂质体治疗后，ICP0的表达水平降低产生的影响在病毒斑块减少实验和3D表皮模型中都得到了验证。
3.3 靶向HSV-1编码β基因的siRNA
UL29编码单链DNA结合蛋白ICP8，不但在病毒DNA复制中起到重要作用还表现出抑制来自亲本基因组的转录和刺激晚期基因转录[27-28]。通过复制缺陷型5型人腺病毒（adenovirus type 5，Adv 5）作为载体介导的shRNA，以UL29为靶标，转导后有效降低UL29的蛋白水平并在体外显著抑制HSV-1复制[29]。HSV-1 DNA聚合酶是病毒复制的一个关键酶，临床上使用的大多数核苷类药物都以其作为靶点。HSV-1核糖核苷酸还原酶（ribonucleotide reductase，RR）基因作为一种β基因，在病毒DNA复制中起重要的调节作用。UL39和UL40是编码RR基因同源二聚体的关键蛋白，Silva等[30]通过siRNA特异性地沉默UL39和UL40表达，导致HSV-1的增殖被抑制，其中一条siRNA有效抑制了99%的病毒增殖。此外，Zhe等[31−34]研究表明使用siRNA靶向视网膜上皮细胞中的HSV-1 ICP4蛋白和UL39（编码ICP6）可有效减少HSV-1增殖。
3.4 靶向HSV-1编码γ基因的siRNA
UL28蛋白是切割和包装HSV-1 DNA所需的七种病毒蛋白之一[35]，Adv5作为载体介导的shRNA，以UL28为靶标，转导后有效降低了UL28的蛋白表达水并在体外显著抑制HSV-1复制[29]。通过酶促反应制备的靶向UL54、UL29和UL27的siRNA在不同程度上抑制了病毒靶基因mRNA表达水平。然而，靶向UL29的siRNA最具潜力[36]。在小鼠角膜模型实验中，靶向UL29的siRNA能抑制病毒增殖，减轻了病毒的感染症状并增加了小鼠存活的时间[37]。此外，siRNA-UL29对TK缺陷型HSV-1治疗也有效[38]。酶法合成的2'-氟修饰Dicer底物siRNA（2′ -F-siRNA）靶向HSV-1 UL29基因，对RNA聚合酶A（RNA polymerase A，RNase A）具有高抗性而不易被降解的优势，与未修饰的siRNA相比其抗HSV-1效力增加了100倍[39]。VP16是病毒立即早期（immediate early，IE）的重要转录基因，Zhang等[40]设计了靶向VP16（由UL48基因编码）的siRNA-1和靶向DNA聚合酶的siRNA-4，表明2种siRNA都显著抑制HSV-1复制。
HSV-1包膜蛋白能与宿主细胞表明受体结合，促进病毒与宿主细胞之间的相互作用。Zhu等[41]通过转染靶向gD（US6）的特异性siRNA到Vero细胞中，gD的敲低导致HSV-1空斑数和病毒蛋白的表达和复制效率降低。转染靶向gE（US8）特异性siRNA到HaCaT细胞中，相比之下，用siRNA转染宿主细胞中WT HSV-1感染产生类似于gE突变病毒的噬斑，与靶向gD的siRNA有类似作用[42]。
综上所述，siRNA靶向HSV-1关键基因的研究为开发新型抗HSV-1药物提供了重要的理论基础和实验依据，为治疗HSV-1感染提供了新的思路和途径。靶向HSV-1编码基因的siRNA，见表1。
表 1 靶向HSV-1编码基因的siRNATable 1. Summary of siRNAs targeting HSV-1-encoded genes靶基因 siRNA 正向 （5′-3′） 反向 （5′-3′） 干扰效率（%） 参考文献 UL18 siUL18-1 GCACCGUUAACCUUCGCAATT UUGCGAAGGUUAACGGUGCTT 86.78 [25] siUL18-2 GUCCUUAACAUGGUUUACUTT AGUAAACCAUGUUAAGGACTT 60.00 siUL18-3 CCAUCAUCCUUACGCUAAUTT AUUAGCGUAAGGAUGAUGGTT 87.25 siUL18-4 CCCGUUAUACGCUAUCCCUAA AGGGAUAGCGUAUAACGGGGG 65.00 UL19 siUL19-1 CCAGCGACGUACAGUUUAATT UUAAACUGUACGUCGCUGGCG 79.71 siUL19-2 CUUUUGUGCCGAUGCATT UGCAUCGGCAACAACAAAGTT 81.52 siUL19-3 CGACCGACGUCAACUACUUTT AAGUAGUUGACGUCGGUCGTT 81.52 siUL19-4 CCAGCGACGUACAGUUUAATT UUAAACUGUACGUCGCUGGTT 78.86 UL26 siUL26-1 CCGUUAACAACAUGAUGCUTT AGCAUCAUGUUGGUUAACGGCG 40.00 siUL26-2 CCGAUUUGUUCGUCUCUCATT UGAGAGACGAACAAAUCGGCG 81.21 siUL26-3 CUGUUGUACCUGAUCACCAAC UGGUGAUCAGGUACAACAGGC NC siUL26-4 CCGUUAACAACAUGAUGCUGC AGCAUCAUGUUGGUUAACGGCG 10.00 UL26.5 siUL26.5 CCGAUUUGUUCGUCUCUCAUU UUGGCUAAACAAGCAGAGAGU 52.12 UL28 shRNAUL28 GATCCGCAGGTGCAGACCTATG
TGTTTTCAAGAGAACACATAGG
TCTGCACCTGCTTTTTTGGAAANC DAY1 50.00 [29] DAY2 70.00 DAY3 60.00 DAY4 40.00 UL29 shRNAUL29 GATCCGCAATCAATTCCAACCG
GTGCTTCAAGAGAGCACCGGTT
GGAATTGATTGCTTTTTTGGAAANC DAY1 60.00 DAY2 80.00 DAY3 60.00 DAY4 50.00 UL30 siRNA-4 GGUACAACAUCAUCAACUUTT AAGUUGAUGAUGUUGUACCTT 6 h 60.00 [40] 12 h 80.00 UL35 siUL35-1 CACGCAAACAACACGUUUATT UAACGUGUUGUUGCGUGGG 50.00 [25] siUL35-2 GCCACCAAUAACUCUCAGUTT ACUGAGAGUUAUUGGUGGCCA 55.95 siUL35-3 CUCUCAGUUUAUCAUGGAUTT AUCCAUGAUAAACUGAGAGTT 22.00 siUL35-4 GUUUGUCGUCGAGAACCUTT AGGUUCUCGAACGACAAACGG 30.00 UL38 siUL38-1 GGCCUAGUGUCGUUUAACUTT AGUUAAACGACACUAGGCCCG NC [25] siUL38-2 GGAUCACCAAACCGAUUCATT UGAAUCGUGUUGGUGAUCCCGG 10.00 siUL38-3 GCGUUUCUGUACCUGGUAUTT AUACCAGGUACAGAAACGCCG 82.48 siUL38-4 GUUGUGUGUACGUGAUCAATT UUGAUCACGUACACACAACAC NC UL39
siRNA1 CUGCACCAUGAUCAUCGACdTdT GUCGAUGAUCAUGGUGCAGdTdT 29.63 [33] siRNA2 AUCGGCCCUGAAGUAUGAGdTdT CUCAUACUUCAGGGCCGAUUG 27.07 siRNA3 GCGCUGCGACAAUAUCUUCdTdT GAAGAUAUUGUCGCAGCGCUG NC siRNA4 CCAUAGCCAAUCCAUGACCdTdT GGUCAUGGAUUGGCUAUGGUC NC UL40 siRNA-1 GACGACCUGGUUACGGAAAdTdT UUUCCGUAACCAGGUCGUCGG 2.73 [30] siRNA-2 AAUGCAUCGAAGUCGUACAdTdT UGUACGACUUCGAUGCAUUCC 69.83 siRNA-3 UCACCUGCCAGUCAAACGAdTdT UCGUUUGACUGGCAGGUGACC 67.94 siRNA-4 AAAUUGGUGUGUUUGUCGGUG AAAUUGGUGUGUUUGUCGGUG 81.31 ICP4 siRNA GCAACAGCAGCUCCUUCAUdTdT dTdTCGUUGUCGUCGAGGAAGUA 12 h 69.00 24 h 95.00 VP16 siRNA-1 GGUACUUUAUGGUGUUGAUTT AUCAACACCAUAAAGUACCTT NC [31，40] 4. 靶向与HSV-1互作宿主基因的siRNA
病毒是一种专性细胞寄生生物，在病毒复制增殖的过程中，许多的宿主细胞基因参与了病毒的生命活动过程。HSV-1在增殖的各个阶段都与宿主基因发生着广泛的相互作用，靶向与HSV-1互作宿主基因的siRNA，同样可以发挥抗HSV-1的功能，这也是设计siRNA药物的策略之一。
4.1 靶向宿主细胞表观调控通路的siRNA
表观遗传调控在HSV-1的感染过程中发挥重要的作用[43]。在HSV-1感染的IE期，病毒基因组能招募组蛋白去甲基化酶1（histone demethylase1，LSD1）促进病毒基因的转录，利用siRNA敲低LSD 1基因的表达有效地抑制HSV-1的裂解复制和潜伏再激活感染[44]。HSV-1感染后显著上调锌指转录因子胰岛素瘤相关1（zinc finger transcription factor insulinoma-associated 1，INSM1）的表达，而用siRNA敲低INSM1基因的表达后抑制了HSV-1的复制[45]。
增殖细胞核抗原（proliferating cell nuclear antigen，PCNA）是调控HSV-1复制及组蛋白沉积的重要细胞因子[46]。用siRNA敲低PCNA的表达后，HSV-1的滴度有所下降，表明PCNA对病毒复制有促进作用[47]。
蔗糖非发酵蛋白2同源物（sucrose nonfermenting protein 2 homolog，SNF2H）是染色质重塑酶的ISWI家族的成员，在HSV-1 复制区室（HSV-1 replication chamber，RC）中表达最丰富。特异性siRNA 敲低SNF 2H mRNA表达水平导致病毒复制减少，表明SNF2H能够促进HSV-1的增殖[48]。
CoREST是一种辅助性抑制蛋白，与抑制性转录因子REST和组蛋白去乙酰化酶（HDACs）1或2形成复合物来抑制细胞基因表达。先前的研究表明，ICP0可以与CoREST结合并将HDAC1从CoREST/REST/去甲基化酶1（LSD1）抑制复合物中排斥出去，靶向CoREST/REST的siRNA导致了病毒α蛋白（ICP4、ICP0、ICP22和ICP27）和β蛋白ICP8的积累均显著减少[49]。
4.2 靶向宿主细胞内吞信号通路的siRNA
内吞网络是细胞进行吞噬、运输和分解细胞外物质的复杂系统，细胞环境中的出芽过程由4种转运复合物（transport complex，ESCRT）和一些蛋白介导，在维持内吞网络完整性方面起着重要作用[50]。其中，ALIX和TSG101是研究的最为深入的ESCRT的互作蛋白，而且HSV-1 编码蛋白的中也存在潜在的ALIX和TSG101结合序列基序，对病毒包膜的形成具有一定作用。Pawliczek等[51]通过siRNA敲低ALIX和TSG101后发现，HSV-1增殖但并不需要ALIX和TSG101[52]。
细胞囊泡内吞过程需要ESCRT-III组分CHMP4C参与，先前被证明对HSV-1 包膜内吞网络循环的完整性并且是不可缺少的。Russell等[53]转染特异性siRNA敲低CHMP4C表达后能显著降低HSV-1的滴度。
4.3 靶向宿主细胞基因的其他siRNA
Nectin-1是HSV进入宿主细胞的受体蛋白，先前研究表明nectin-1在HaCaT角质形成细胞系中高度表达[54]。从小鼠角质形成细胞中敲除nectin-1导致HSV-1进入减少[55]，Sayers等[56−57]使用siRNA敲除 nTERT细胞的nectin-1和HVEM受体导致病毒进入减少。B5蛋白是HSV-1进入宿主的辅助受体，siRNA沉默B5蛋白表达后导致HSV-1空斑形成水平明显降低，ICP4、ICP0、ICP27和VP16蛋白表达也显著降低[58]。靶向与HSV-1互作宿主基因的siRNA见表2。
表 2 靶向宿主基因与HSV-1相互作用的siRNATable 2. Summary of siRNA targeting host genes interacting with HSV-1靶基因 siRNA 正向（5′-3′） 参考文献 INSM1 siINSM1 UCCGCAAGCUGCACUUCGATT [45] SNF2H siRNA5 GGAAUGGUAUACUCGGAUA [48] siRNA6 GGGCAAA UAGAUUCGAGUA siRNA7 GGAUUUACCAAUUGGAAUA siRNA8 GUUCUUUCCUCCACGUUUA REST siREST GUGAUACUGUAGAUUACAC [49] coREST sicoREST AAGAUUGUCCCGUUCUUGACU [59] TSG101 siTSG101 CCUCCAGUCUUCUCUCGUCTT [52] ALIX siALIX GCCGCUGGUGAAGUUCAUCTT 5. 小结
HSV-1是一种能在神经细胞中发生潜伏感染的病毒，这使得很难通过药物治疗或机体的免疫力彻底将其清除。siRNA在2001年被首次报道后，为治疗HSV-1带来了新希望[19]。通过siRNA靶向HSV-1基因和互作宿主基因以及与其他治疗手段的联合应用为治疗HSV-1感染及相关siRNA疫苗开发提供了新的策略。
siRNA的高效递送体系统也是值得进行深入研究的方向。目前在mRNA疫苗中常用的脂纳米颗粒 （lipid nanoparticles，LNP） 是非常成熟的递送系统，但其细胞毒性和激活天然免疫的副作用仍需进一步的改进。
尽管siRNA药物治疗病毒性疾病取得了一些进展，目前还没有基于RNAi的抗HSV-1疗法，其临床应用仍面临着挑战，需要进一步的研究和探索来解决相关的技术难题和安全性问题。
-
表 1 靶向HSV-1编码基因的siRNA
Table 1. Summary of siRNAs targeting HSV-1-encoded genes
靶基因 siRNA 正向 （5′-3′） 反向 （5′-3′） 干扰效率（%） 参考文献 UL18 siUL18-1 GCACCGUUAACCUUCGCAATT UUGCGAAGGUUAACGGUGCTT 86.78 [25] siUL18-2 GUCCUUAACAUGGUUUACUTT AGUAAACCAUGUUAAGGACTT 60.00 siUL18-3 CCAUCAUCCUUACGCUAAUTT AUUAGCGUAAGGAUGAUGGTT 87.25 siUL18-4 CCCGUUAUACGCUAUCCCUAA AGGGAUAGCGUAUAACGGGGG 65.00 UL19 siUL19-1 CCAGCGACGUACAGUUUAATT UUAAACUGUACGUCGCUGGCG 79.71 siUL19-2 CUUUUGUGCCGAUGCATT UGCAUCGGCAACAACAAAGTT 81.52 siUL19-3 CGACCGACGUCAACUACUUTT AAGUAGUUGACGUCGGUCGTT 81.52 siUL19-4 CCAGCGACGUACAGUUUAATT UUAAACUGUACGUCGCUGGTT 78.86 UL26 siUL26-1 CCGUUAACAACAUGAUGCUTT AGCAUCAUGUUGGUUAACGGCG 40.00 siUL26-2 CCGAUUUGUUCGUCUCUCATT UGAGAGACGAACAAAUCGGCG 81.21 siUL26-3 CUGUUGUACCUGAUCACCAAC UGGUGAUCAGGUACAACAGGC NC siUL26-4 CCGUUAACAACAUGAUGCUGC AGCAUCAUGUUGGUUAACGGCG 10.00 UL26.5 siUL26.5 CCGAUUUGUUCGUCUCUCAUU UUGGCUAAACAAGCAGAGAGU 52.12 UL28 shRNAUL28 GATCCGCAGGTGCAGACCTATG
TGTTTTCAAGAGAACACATAGG
TCTGCACCTGCTTTTTTGGAAANC DAY1 50.00 [29] DAY2 70.00 DAY3 60.00 DAY4 40.00 UL29 shRNAUL29 GATCCGCAATCAATTCCAACCG
GTGCTTCAAGAGAGCACCGGTT
GGAATTGATTGCTTTTTTGGAAANC DAY1 60.00 DAY2 80.00 DAY3 60.00 DAY4 50.00 UL30 siRNA-4 GGUACAACAUCAUCAACUUTT AAGUUGAUGAUGUUGUACCTT 6 h 60.00 [40] 12 h 80.00 UL35 siUL35-1 CACGCAAACAACACGUUUATT UAACGUGUUGUUGCGUGGG 50.00 [25] siUL35-2 GCCACCAAUAACUCUCAGUTT ACUGAGAGUUAUUGGUGGCCA 55.95 siUL35-3 CUCUCAGUUUAUCAUGGAUTT AUCCAUGAUAAACUGAGAGTT 22.00 siUL35-4 GUUUGUCGUCGAGAACCUTT AGGUUCUCGAACGACAAACGG 30.00 UL38 siUL38-1 GGCCUAGUGUCGUUUAACUTT AGUUAAACGACACUAGGCCCG NC [25] siUL38-2 GGAUCACCAAACCGAUUCATT UGAAUCGUGUUGGUGAUCCCGG 10.00 siUL38-3 GCGUUUCUGUACCUGGUAUTT AUACCAGGUACAGAAACGCCG 82.48 siUL38-4 GUUGUGUGUACGUGAUCAATT UUGAUCACGUACACACAACAC NC UL39
siRNA1 CUGCACCAUGAUCAUCGACdTdT GUCGAUGAUCAUGGUGCAGdTdT 29.63 [33] siRNA2 AUCGGCCCUGAAGUAUGAGdTdT CUCAUACUUCAGGGCCGAUUG 27.07 siRNA3 GCGCUGCGACAAUAUCUUCdTdT GAAGAUAUUGUCGCAGCGCUG NC siRNA4 CCAUAGCCAAUCCAUGACCdTdT GGUCAUGGAUUGGCUAUGGUC NC UL40 siRNA-1 GACGACCUGGUUACGGAAAdTdT UUUCCGUAACCAGGUCGUCGG 2.73 [30] siRNA-2 AAUGCAUCGAAGUCGUACAdTdT UGUACGACUUCGAUGCAUUCC 69.83 siRNA-3 UCACCUGCCAGUCAAACGAdTdT UCGUUUGACUGGCAGGUGACC 67.94 siRNA-4 AAAUUGGUGUGUUUGUCGGUG AAAUUGGUGUGUUUGUCGGUG 81.31 ICP4 siRNA GCAACAGCAGCUCCUUCAUdTdT dTdTCGUUGUCGUCGAGGAAGUA 12 h 69.00 24 h 95.00 VP16 siRNA-1 GGUACUUUAUGGUGUUGAUTT AUCAACACCAUAAAGUACCTT NC [31，40] 
下载: 导出CSV
表 2 靶向宿主基因与HSV-1相互作用的siRNA
Table 2. Summary of siRNA targeting host genes interacting with HSV-1
靶基因 siRNA 正向（5′-3′） 参考文献 INSM1 siINSM1 UCCGCAAGCUGCACUUCGATT [45] SNF2H siRNA5 GGAAUGGUAUACUCGGAUA [48] siRNA6 GGGCAAA UAGAUUCGAGUA siRNA7 GGAUUUACCAAUUGGAAUA siRNA8 GUUCUUUCCUCCACGUUUA REST siREST GUGAUACUGUAGAUUACAC [49] coREST sicoREST AAGAUUGUCCCGUUCUUGACU [59] TSG101 siTSG101 CCUCCAGUCUUCUCUCGUCTT [52] ALIX siALIX GCCGCUGGUGAAGUUCAUCTT
下载: 导出CSV
-
[1] Looker K J,Magaret A S,May M T,et al. Global and regional estimates of prevalent and incident herpes simplex virus type 1 infections in 2012[J]. PLoS One,2015,10(10):e0140765. doi: 10.1371/journal.pone.0140765 [2] James C,Harfouche M,Welton N J,et al. Herpes simplex virus: Global infection prevalence and incidence estimates,2016[J]. Bull World Health Organ,2020,98(5):315-329. doi: 10.2471/BLT.19.237149 [3] Looker K J,Magaret A S,May M T,et al. First estimates of the global and regional incidence of neonatal herpes infection[J]. Lancet Glob Health,2017,5(3):e300-e309. doi: 10.1016/S2214-109X(16)30362-X [4] Xu F,Sternberg M R,Kottiri B J,et al. Trends in herpes simplex virus type 1 and type 2 seroprevalence in the United States[J]. Jama,2006,296(8):964-973. doi: 10.1001/jama.296.8.964 [5] Navarro-Bielsa A,Gracia-Cazana T,Aldea-Manrique B,et al. COVID-19 infection and vaccines: Potential triggers of Herpesviridae reactivation[J]. An Bras Dermatol,2023,98(3):347-354. doi: 10.1016/j.abd.2022.09.004 [6] Whitley RJRoizman B. Herpes simplex virus infections[J]. The Lancet,2001,357(9267):1513-1518. doi: 10.1016/S0140-6736(00)04638-9 [7] Marcocci M E,Napoletani G,Protto V,et al. Herpes simplex virus-1 in the brain: The dark side of a sneaky infection[J]. Trends Microbiol,2020,28(10):808-820. doi: 10.1016/j.tim.2020.03.003 [8] De Chiara G,Marcocci M E,Sgarbanti R,et al. Infectious agents and neurodegeneration[J]. Mol Neurobiol,2012,46(3):614-638. doi: 10.1007/s12035-012-8320-7 [9] De Clercq E. A 40-year journey in search of selective antiviral chemotherapy[J]. Annual Review of Pharmacology and Toxicology,2011,51(1):1-24. doi: 10.1146/annurev-pharmtox-010510-100228 [10] De Clercq E. Antivirals: Past,present and future[J]. Biochem Pharmacol,2013,85(6):727-744. doi: 10.1016/j.bcp.2012.12.011 [11] Burrel S,Boutolleau D,Azar G,et al. Phenotypic and genotypic characterization of acyclovir-resistant corneal HSV-1 isolates from immunocompetent patients with recurrent herpetic keratitis[J]. J Clin Virol,2013,58(1):321-324. doi: 10.1016/j.jcv.2013.05.001 [12] Sadowski L A,Upadhyay R,Greeley Z W,et al. Current drugs to treat infections with herpes simplex viruses-1 and -2[J]. Viruses,2021,13(7):1228. doi: 10.3390/v13071228 [13] Preda M,Manolescu L S C,Chivu R D. Advances in alpha herpes viruses vaccines for human[J]. Vaccines (Basel),2023,11(6):1094. doi: 10.3390/vaccines11061094 [14] Pushparaj P N,Aarthi J J,Manikandan J,et al. siRNA,miRNA,and shRNA: In vivo applications[J]. J Dent Res,2008,87(11):992-1003. doi: 10.1177/154405910808701109 [15] Hu B,Zhong L,Weng Y,et al. Therapeutic siRNA: State of the art[J]. Signal Transduct Target Ther,2020,5(1):101. doi: 10.1038/s41392-020-0207-x [16] Tan F L,Yin J Q. RNAi,a new therapeutic strategy against viral infection[J]. Cell Res,2004,14(6):460-466. doi: 10.1038/sj.cr.7290248 [17] Guo S,Kemphues K J. par-1,a gene required for establishing polarity in C. elegans embryos,encodes a putative Ser/Thr kinase that is asymmetrically distributed[J]. Cell,1995,81(4):611-620. doi: 10.1016/0092-8674(95)90082-9 [18] Fire A,Xu S,Montgomery M K,et al. Potent and specific genetic interference by double-stranded RNA in Caenorhabditis elegans[J]. Nature,1998,391(6669):806-811. doi: 10.1038/35888 [19] Elbashir S M,Lendeckel W,Tuschl T. RNA interference is mediated by 21- and 22-nucleotide RNAs[J]. Genes Dev,2001,15(2):188-200. doi: 10.1101/gad.862301 [20] Elbashir S M,Harborth J,Lendeckel W,et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells[J]. Nature,2001,411(6836):494-498. doi: 10.1038/35078107 [21] Hammond S M,Bernstein E,Beach D,et al. An RNA-directed nuclease mediates post-transcriptional gene silencing in Drosophila cells[J]. Nature,2000,404(6775):293-296. doi: 10.1038/35005107 [22] Saurabh S,Vidyarthi A S,Prasad D. RNA interference: concept to reality in crop improvement[J]. Planta,2014,239(3):543-564. doi: 10.1007/s00425-013-2019-5 [23] Carthew R W,Sontheimer E J. Origins and mechanisms of miRNAs and siRNAs[J]. Cell,2009,136(4):642-655. doi: 10.1016/j.cell.2009.01.035 [24] Griffiths S J,Haas J. siRNA screening for genes involved in HSV-1 replication[J]. Bio Protoc,2014,4(16):e1209. [25] Jin F,Li S,Zheng K,et al. Silencing herpes simplex virus type 1 capsid protein encoding genes by siRNA: A promising antiviral therapeutic approach[J]. PLoS One,2014,9(5):e96623. doi: 10.1371/journal.pone.0096623 [26] Jbara-Agbaria D,Blondzik S,Burger-Kentischer A,et al. Liposomal siRNA formulations for the treatment of herpes simplex virus-1: In vitro characterization of physicochemical properties and activity,and in vivo biodistribution and toxicity studies[J]. Pharmaceutics,2022,14(3):633. doi: 10.3390/pharmaceutics14030633 [27] Taylor T J,Knipe D M. Proteomics of herpes simplex virus replication compartments: association of cellular DNA replication,repair,recombination,and chromatin remodeling proteins with ICP8[J]. J Virol,2004,78(11):5856-5866. doi: 10.1128/JVI.78.11.5856-5866.2004 [28] Bryant K F,Yan Z,Dreyfus D H,et al. Identification of a divalent metal cation binding site in herpes simplex virus 1 (HSV-1) ICP8 required for HSV replication[J]. J Virol,2012,86(12):6825-6834. doi: 10.1128/JVI.00374-12 [29] Song B,Liu X,Wang Q,et al. Adenovirus-mediated shRNA interference against HSV-1 replication in vitro[J]. J Neurovirol,2016,22(6):799-807. doi: 10.1007/s13365-016-0453-4 [30] Silva A P,Lopes J F,Paula V S. RNA interference inhibits herpes simplex virus type 1 isolated from saliva samples and mucocutaneous lesions[J]. Braz J Infect Dis,2014,18(4):441-444. doi: 10.1016/j.bjid.2014.01.011 [31] Duan F,Ni S,Nie Y,et al. Small interfering RNA targeting for infected-cell polypeptide 4 inhibits herpes simplex virus type 1 replication in retinal pigment epithelial cells[J]. Clinical & Experimental Ophthalmology,2012,40(2):195-204. [32] Liu Y T,Song B,Wang Y L,et al. [SiRNA targeting ICP4 attenuates HSV-1 replication][J]. Bing Du Xue Bao,2010,26(3):163-169. [33] Zhe R,Mei-Ying Z,Kitazato K,et al. Effect of siRNA on HSV-1 plaque formation and relative expression levels of UL39 mRNA[J]. Arch Virol,2008,153(7):1401-1406. doi: 10.1007/s00705-008-0110-1 [34] Ren Z,Li S,Wang Q L,et al. Effect of siRNAs on HSV-1 plaque formation and relative expression levels of RR mRNA[J]. Virol Sin,2011,26(1):40-46. doi: 10.1007/s12250-011-3162-9 [35] Heming J D,Conway J F,Homa F L. Herpesvirus capsid assembly and DNA packaging[J]. Adv Anat Embryol Cell Biol,2017,223:119-142. [36] Paavilainen H,Lehtinen J,Romanovskaya A,et al. Inhibition of clinical pathogenic herpes simplex virus 1 strains with enzymatically created siRNA pools[J]. J Med Virol,2016,88(12):2196-2205. doi: 10.1002/jmv.24578 [37] Paavilainen H,Lehtinen J,Romanovskaya A,et al. Topical treatment of herpes simplex virus infection with enzymatically created siRNA swarm[J]. Antivir Ther,2017,22(7):631-637. doi: 10.3851/IMP3153 [38] Kalke K,Lehtinen J,Gnjatovic J,et al. Herpes simplex virus type 1 clinical isolates respond to UL29-targeted siRNA swarm treatment independent of their acyclovir sensitivity[J]. Viruses,2020,12(12):1434. doi: 10.3390/v12121434 [39] Levanova A A,Kalke K M,Lund L M,et al. Enzymatically synthesized 2'-fluoro-modified Dicer-substrate siRNA swarms against herpes simplex virus demonstrate enhanced antiviral efficacy and low cytotoxicity[J]. Antiviral Res,2020,182:104916. doi: 10.1016/j.antiviral.2020.104916 [40] Zhang Y Q,Lai W,Li H,et al. Inhibition of herpes simplex virus type 1 by small interfering RNA[J]. Clin Exp Dermatol,2008,33(1):56-61. [41] Zhu Q C,Ren Z,Zhang C L,et al. Silencing HSV1 gD expression in cultured cells by RNA interference[J]. Bing Du Xue Bao,2007,23(1):22-27. [42] Bhuyan P K,Kariko K,Capodici J,et al. Short interfering RNA-mediated inhibition of herpes simplex virus type 1 gene expression and function during infection of human keratinocytes[J]. J Virol,2004,78(19):10276-10281. doi: 10.1128/JVI.78.19.10276-10281.2004 [43] 吴长静,邹雨芳,黄新伟. HSV1感染中的表观遗传调控机制研究进展[J]. 昆明医科大学学报,2024,45(1):172-178. doi: 10.12259/j.issn.2095-610X.S20240129 [44] Liang Y,Vogel J L,Narayanan A,et al. Inhibition of the histone demethylase LSD1 blocks alpha-herpesvirus lytic replication and reactivation from latency[J]. Nat Med,2009,15(11):1312-1317. doi: 10.1038/nm.2051 [45] Kamakura M,Goshima F,Luo C,et al. Herpes simplex virus induces the marked up-regulation of the zinc finger transcriptional factor INSM1,which modulates the expression and localization of the immediate early protein ICP0[J]. Virol J,2011,8:257. doi: 10.1186/1743-422X-8-257 [46] Olivo J F,Guille F,Lobel B. Microscopic hematuria. Semiologic value in urology. Management of microscopic hematuria[J]. J Urol (Paris),1989,95(8):453-458. [47] Sanders I,Boyer MF,Fraser N W. Early nucleosome deposition on,and replication of,HSV DNA requires cell factor PCNA[J]. J Neurovirol,2015,21(4):358-369. doi: 10.1007/s13365-015-0321-7 [48] Bryant K F,Colgrove R C,Knipe D M. Cellular SNF2H chromatin-remodeling factor promotes herpes simplex virus 1 immediate-early gene expression and replication[J]. MBio,2011,2(1):e00330-10. [49] Zhou G,Te D,Roizman B. The CoREST/REST repressor is both necessary and inimical for expression of herpes simplex virus genes[J]. mBio,2010,2(1):e00313-10. [50] Mccullough J,Colf LA,Sundquist W I. Membrane fission reactions of the mammalian ESCRT pathway[J]. Annu Rev Biochem,2013,82:663-692. doi: 10.1146/annurev-biochem-072909-101058 [51] Pawliczek T,Crump C M. Herpes simplex virus type 1 production requires a functional ESCRT-III complex but is independent of TSG101 and ALIX expression[J]. J Virol,2009,83(21):11254-11264. doi: 10.1128/JVI.00574-09 [52] Barnes J,Wilson D W. The ESCRT-II subunit EAP20/VPS25 and the bro1 domain proteins HD-PTP and BROX are individually dispensable for herpes simplex virus 1 replication[J]. J Virol,2020,94(4):e01641-19. [53] Russell T,Samolej J,Hollinshead M,et al. Novel role for ESCRT-III component CHMP4C in the integrity of the endocytic network utilized for herpes simplex virus envelopment[J]. mBio,2021,12(3):e02183-20. [54] Huber M T,Wisner T W,Hegde N R,et al. Herpes simplex virus with highly reduced gD levels can efficiently enter and spread between human keratinocytes[J]. J Virol,2001,75(21):10309-10318. doi: 10.1128/JVI.75.21.10309-10318.2001 [55] Petermann P,Thier K,Rahn E,et al. Entry mechanisms of herpes simplex virus 1 into murine epidermis: Involvement of nectin-1 and herpesvirus entry mediator as cellular receptors[J]. J Virol,2015,89(1):262-274. doi: 10.1128/JVI.02917-14 [56] Sayers C L,Elliott G. Herpes simplex virus 1 enters human keratinocytes by a nectin-1-dependent,rapid plasma membrane fusion pathway that functions at low temperature[J]. J Virol,2016,90(22):10379-10389. doi: 10.1128/JVI.01582-16 [57] Tiwari V,Oh M J,Kovacs M,et al. Role for nectin-1 in herpes simplex virus 1 entry and spread in human retinal pigment epithelial cells[J]. FEBS J,2008,275(21):5272-5285. doi: 10.1111/j.1742-4658.2008.06655.x [58] Cheshenko N,Trepanier J B,Segarra T J,et al. HSV usurps eukaryotic initiation factor 3 subunit M for viral protein translation: novel prevention target[J]. PLoS One,2010,5(7):e11829. doi: 10.1371/journal.pone.0011829 [59] Gu H,Liang Y,Mandel G,et al. Components of the REST/CoREST/histone deacetylase repressor complex are disrupted,modified,and translocated in HSV-1-infected cells[J]. Proc Natl Acad Sci U S A,2005,102(21):7571-7576. doi: 10.1073/pnas.0502658102 -
下载:
点击查看大图
计量
- 文章访问数: 766
- HTML全文浏览量: 200
- PDF下载量: 60
- 被引次数: 0
