Development of poliovirus-rotavirus chimeric vaccine by using poliovirus artificial interfering particle RNA

利用脊髓灰质炎病毒人工干扰颗粒RNA研制脊髓灰质炎病毒-轮状病毒嵌合疫苗

• 批准号: 04557025
• 负责人: TANIGUCHI Koki
• 金额: $ 12.48万
• 依托单位: SAPPORO MEDICAL UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Developmental Scientific Research (B)
• 财政年份: 1992
• 资助国家: 日本
• 起止时间: 1992 至 1994
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-04557025/
• 关键词: Rotavirus; Poliovirus; Vaccine; VP4 gene; DI particle; Cross-reactive neutralization epitope; キメラウイルス; 効反応性中和領域; 中和抗原; VP4; 交又反応中和エピトープ

We constructed polio-rota VP4 chimeric cDNA by inserting the DNA fragments encoding VP8' or VP5' into pSM1 (T7) 3 or pSM1 (T7) 2 vector having poliovirus artificial DI genome. In the HeLa cells transfected with polio-rota chimeric RNA obtained from pSM1 (T7) 3-VP5', pSM1 (T7) 3-VP8', or pSM1 (T7) 2-VP8'+VP5', replication of rotavirus-specific RNA could be detected. However, the expression of rotaviral protein was observed only in the cells transfected with pSM1 (T7) 3-VP8' or pSM1 (T7) 2-VP8'+VP5'. The antigenicity of the expressed VP5' might be weak in this system because of the conformational change in the from of the VP5' fused with polioviral protein. Rotaviral protein reactive with antiserum to VP4 could be detected in the cells at the fourth generation after transfection with pSM1(T7)3-VP8' and infection with infectious poiliovirus. By preparing of polio-rota chimeric virus with high titer and by using the transgenic mice with poliovirus receptor which have the potency of oral inoculation, the immunogenicity of the poliorota chimeric virus would be investigated.We determined the nucleotide and deduced amino acid sequences of the VP4 genes of the rotaviruses from various mammalian species including humans. Comparative analysis of the VP4 sequences showed that there are at least 17 kinds of VP4 genotypes and that species specificity and interspecies relatedness in VP4 geneotypes are present. It was also shown that some human rotaviruses have VP4 geneotypes related to those of animal rotaviruses. These findings should be taken into considerataion for rotavirus vaccine development.

我们将编码VP 8 '或VP 5'的DNA片段分别插入到含有脊髓灰质炎病毒人工DI基因组的pSM 1（T7）3或pSM 1（T7）2载体中，构建了脊髓灰质炎病毒VP 4嵌合cDNA。在用从pSM 1（T7）3-VP 5 '、pSM 1（T7）3-VP 8'或pSM 1（T7）2-VP 8 '+ VP 5'获得的脊髓灰质炎-轮状病毒嵌合RNA转染的HeLa细胞中，可以检测到轮状病毒特异性RNA的复制。而仅在转染pSM 1（T7）3-VP 8 '或pSM 1（T7）2-VP 8'+ VP 5 '的细胞中观察到轮状病毒蛋白的表达。在该系统中表达的VP 5 '抗原性较弱，这可能是由于VP 5'与脊髓灰质炎病毒蛋白融合后构象发生了变化。pSM 1（T7）3-VP 8 '转染第4代细胞后，用感染性脊髓灰质炎病毒感染，可检测到与VP 4抗血清反应的轮状病毒蛋白。通过制备高滴度的脊灰-轮状病毒嵌合体病毒，并利用具有口服免疫效力的脊灰病毒受体转基因小鼠，研究脊灰嵌合体病毒的免疫原性。对VP 4基因序列的比较分析表明，我国至少存在17种VP 4基因型，且存在种间亲缘关系和种间特异性。此外，一些人轮状病毒具有与动物轮状病毒相关的VP 4基因型。这些结果应考虑到在轮状病毒疫苗的开发。

项目成果

Kobayashi, N.: "Analysis of Gene Selection in Reassortant Formation between Canine Rotavirus K9 and Human Rotaviruses with Different Antigenic Specificities." Res.Virol.144. 361-370 (1993)

Kobayashi, N.：“具有不同抗原特异性的犬轮状病毒 K9 和人轮状病毒重配形成中的基因选择分析”。

Takagi, M.: "A Minor Prevalent Strain in a Severe Outbreak of Foal Diarrhea Associated with Serotype 3 Rotavirus." J.Vet.Med.Sci.55. 661-663 (1993)

Takagi, M.：“与血清型 3 轮状病毒相关的小马驹腹泻严重爆发中的一种次要流行菌株。”

Pongsuwanna, Y.: "Distinct Yearly Change of Serotype Distribution of Human Rotavirus in Thailand as Determined by ELISA and PCR." Epidemiol.Infect.111. 407-412 (1993)

Pongsuwanna, Y.：“通过 ELISA 和 PCR 测定泰国人类轮状病毒血清型分布的独特年度变化。”

Shozo Urasawa: "Antigenic and genetic alalyses of human rotaviruses in Chiang Mai,Thailand" Journal of Infectious Diseases. 166. 227-234 (1992)

Shozo Urasawa：“泰国清迈人类轮状病毒的抗原和遗传分析”《传染病杂志》。

Nobumichi Kobayashi: "Reactivity of anti-human rotavirus VP4 neutralizing monoclonal antibodies with animal rotaviruses and with unusual human rotaviruses having different P and G serotype." Res.Virol.144. 201-207 (1993)

Nobumichi Kobayashi：“抗人轮状病毒 VP4 中和单克隆抗体与动物轮状病毒和具有不同 P 和 G 血清型的不寻常人类轮状病毒的反应性。”