Characterization of an anti-Human Papillomavirus (HPV) agent

抗人乳头瘤病毒 (HPV) 药物的表征

基本信息

批准号：
10618912
负责人：
Asok Antony
金额：
--
依托单位：
RLR VA MEDICAL CENTER
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2024-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10618912
关键词：
Affinity American Binding Proteins Biological Assay Biological Products Cancer Model Capsid Capsid Proteins Cells Development Dose Effectiveness Elements Episome Exhibits Folic Acid Folic Acid Deficiency Gel Generations Genital Genitalia Genitourinary system Genomics HPV-High Risk Heterogeneous-Nuclear Ribonucleoproteins Histology Human Papilloma Virus Vaccine Human Papilloma Virus-Related Malignant Neoplasm Human Papillomavirus Human papilloma virus infection Human papillomavirus 16 Human papillomavirus 18 Human papillomavirus 6 Implant In Vitro Individual Infection L2 viral capsid protein Legal patent Low risk HPV Malignant Neoplasms Messenger RNA Modeling New Agents Newly Diagnosed Nuclear Nucleotides Nude Mice Oncogenic Oropharyngeal Proteins Publishing RNA RNA-Protein Interaction Reporter Ribonucleoproteins Risk Role Safety Sexual Partners Sexual Transmission Standardization Teenagers Testing Therapeutic Time Tissue Donors Tissues Veterans Viral Virion Virus Xenograft Model carcinogenesis comparison control density experimental study high risk implantation in vivo in vivo Model innovation keratinocyte malignant oropharynx neoplasm mouse model mutant novel particle prevent protein expression response subcutaneous transmission process tumor viral transmission

项目摘要

Despite the advent of effective anti-Human Papillomavirus (HPV) vaccines, there are no biological agents to reliably prevent ~80 million Americans from transmitting their infectious HPV viral particles to sexual partners. Earlier we determined that the post-translational homocysteinylation of an mRNA-binding protein (heterogenous nuclear ribonucleoprotein-E1, hnRNP-E1) can transform hnRNP-E1 into a moiety with high affinity for a HPV16 57-nucleotide (nt) RNA cis-element under conditions of folate deficiency; this interaction led to a profound inhibition of both HPV16 L1 and L2 viral capsid proteins that are essential for HPV16- encapsidation (and infectivity). We have patented a powerful mutant of hnRNP-E1 [DomPos-E1(C293S)] that functions like homocysteinylated-hnRNP-E1 under folate-replete conditions. Because DomPos-E1(C293S)] has such a strong likelihood to eliminate HPV16 viral capsid proteins and thereby function as an anti-HPV agent, we wish to test its therapeutic potential both in vitro and in our novel HPV16-xenograft model in Beige Nude mice. In Specific Aim 1 we will compare effects of the interaction of DomPos-E1(C293S)-protein [relative to control wild-type(wt)-like-E1(G292A)-proteins] with HPV16 57-nt cis-element in eliminating HPV16 L1 and L2 viral capsid protein expression. We will also extend these studies to assess the interaction of DomPos-E1(C293S) with similar cis-elements from low risk HPV6 and 11 and high-risk types (HPV18, 31, 33, 45, 52, 58). Next, we will confirm the greater impact of DomPos-E1(C293S)- over control wt-like-E1(G292A)- expression in reducing HPV16 L1 and L2 after stable transduction into HPV16-harboring keratinocytes that are also transformed into HPV16-organotypic rafts; then we can evaluate the extent in reduction of infectious HPV16 viral particles in 18- day old rafts and whether there is any increase in genomic integration by amplified capsid-less HPV16 episomes. In Specific Aim 2, we will subcutaneously implant these DomPos-E1(C293S)- or control wt-like- E1(G292A))- expressing rafts in Beige Nude mice using our recently published model. This will allow us to assess the relative effects of DomPos-E1(C293S)- over control wt-like-E1(G292A) in reducing both HPV16 viral capsid proteins and infectious viral particles of HPV16 over the ensuing 8 weeks in vivo; evaluating if this reduces the capacity of the implanted HPV16-raft to auto-infect itself; determining changes in genomic integration by amplified capsid-less HPV16 episomes; and in prolonging the expected time of rafts to develop HPV16-cancers. In Specific Aim 3, we will determine if DomPos-E1(C293S) is significantly more effective than control wt- like-E1(G292A) in preventing transmission of HPV16 to adjacent tissue. We will adapt our in vivo model to assess HPV16-infectivity wherein the effectiveness of transmission of infectious HPV16 from one donor tissue to an uninfected recipient tissue is assessed over 8 weeks in Beige Nude mice. These studies will help determine if DomPos-E1(C293S) or its mutant derivatives can be moved forward as first-in-class agents to help reduce transmission of infectious HPV16 viral particles from an infected host.

尽管有效的抗人乳头瘤病毒（HPV）疫苗出现了，但没有生物学剂可靠地防止约8000万美国人将其感染性的HPV病毒颗粒传播给性伴侣。早些时候，我们确定mRNA结合蛋白的翻译后同藻化（异源核核糖核蛋白-E1，HNRNP-E1）可以将HNRNP-E1转变为具有较高的部分在叶酸缺乏条件下，HPV16 57-核苷酸（NT）RNA CIS元素的亲和力；这种相互作用导致对HPV16-L1和L2病毒capsid蛋白的深刻抑制作用，这对于HPV16-至关重要封装（和感染力）。我们已经为HNRNP-E1的强大突变体提供了专利[DOMPOS-E1（C293S）] 在叶酸复合条件下，诸如同型凝固性HNRNP-E1之类的功能。因为DOMPOS-E1（C293S）具有消除HPV16病毒衣壳蛋白的可能性很强，从而充当抗HPV剂，我们希望在米色裸鼠中在体外和我们新颖的HPV16- Xenograpt模型中测试其治疗潜力。在特定目标1中，我们将比较DOMPOS-E1（C293S） - 蛋白质的相互作用[相对于用HPV16 57-NT顺式元素对照野生型（WT）-E1（G292A） - 蛋白质]消除HPV16 L1和L2 病毒式衣壳蛋白表达。我们还将扩展这些研究以评估DOMPOS-E1（C293S）的相互作用来自低风险HPV6和11和高风险类型的类似顺式元素（HPV18、31、33、45、52、58）。接下来，我们将确认DOMPOS-E1（C293S）的更大影响 - 对控制WT-E1（G292A） - 在还原中的表达 HPV16 L1和L2稳定转导为HPV16-HARBORING角质形成细胞，这些角质形成细胞也被转化为 HPV16-有机型筏；然后，我们可以评估18-- Day Old Rafts以及通过扩增无帽capsid的HPV16插发体的基因组整合是否有所增加。在特定的目标2中，我们将皮下植入这些DOMPOS-E1（C293S）或对照WT Like- E1（G292A）） - 使用我们最近发表的模型在米色裸鼠中表达筏。这将使我们能够评估 DOMPOS-E1（C293S）的相对影响 - 对控制WT-E1（G292A）在减少两个HPV16病毒capsid中的相对影响随后在体内的8周内，HPV16的蛋白质和传染性病毒颗粒；评估这是否减少了植入的HPV16-RAFT的容量自身自动感染；确定基因组整合的变化放大无钩状的HPV16插图；并延长了木筏发展HPV16-Canters的预期时间。在特定的目标3中，我们将确定DOMPOS-E1（C293S）是否比对照WT-明显更有效类似-E1（G292a）防止HPV16传播到相邻组织。我们将调整我们的体内模型来评估 HPV16感染性，其中传染性HPV16从一个供体组织传播到一个在米色裸鼠中，在8周内评估未感染的受体组织。这些研究将有助于确定DOMPOS-E1（C293S）或其突变衍生物是否可以向前移动作为一流的药物，可以帮助减少感染宿主的传染性HPV16病毒颗粒的传播。