The Human papillomavirus 16 (HPV16) E7 oncoprotein is a promising candidate for development of anti-cancer therapeutic vaccine. We have prepared the expression construct carrying mutagenized E7 oncoprotein fused to the C-terminus of Tobacco mosaic virus (TMV) coat protein via 15 amino acids β-sheet linker. The fusion protein was expressed in Escherichia coli MC 1061 cells. We have obtained high level expression, but most of the protein remained in insoluble inclusion bodies. To increase the ratio of soluble protein various molecular chaperones (TF, DnaK-DnaJ-GrpE, GroEL-GroES) were used. The immunological reactivity of expressed recombinant protein was evaluated with anti-E7 and anti-TMV antibodies. The distribution of expressed product during ultracentrifugation on sucrose gradient was studied.
- MeSH
- elektroforéza v polyakrylamidovém gelu MeSH
- Escherichia coli genetika MeSH
- exprese genu MeSH
- infekce papilomavirem virologie MeSH
- klonování DNA metody MeSH
- králíci MeSH
- lidé MeSH
- lidský papilomavirus 16 chemie genetika imunologie MeSH
- molekulární chaperony chemie genetika MeSH
- mutageneze MeSH
- myši MeSH
- Papillomavirus E7 - proteiny chemie genetika imunologie MeSH
- protilátky imunologie MeSH
- rekombinantní fúzní proteiny chemie genetika imunologie MeSH
- rozpustnost MeSH
- virové plášťové proteiny chemie genetika imunologie MeSH
- virus tabákové mozaiky chemie genetika imunologie MeSH
- zvířata MeSH
- Check Tag
- králíci MeSH
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Downregulation of MHC class I expression on the cell surface is a common mechanism by which tumour cells, including cervical carcinoma, can escape the T cell-mediated anti-tumour immunity. This downregulation represents an obstacle for the efficacy of anti-tumour vaccines. In this study, we investigated the efficacy of prophylactic peptide and peptide-pulsed dendritic cell-based vaccines in a murine model of experimental MHC class I-deficient tumours (TC-1/A9), expressing E6/E7 oncogenes derived from HPV16, and compared the efficacy of particular vaccination settings to anti-tumour protection against parental MHC class I-positive TC-1 tumours. Peptide vaccine based on the 'short' peptide E749-57 harbouring solely the CTL epitope and co-administered to the C57BL/6 mice with CpG oligodeoxynucleotide (CpG ODN) 1826 was effective against MHC class I-positive but not -deficient tumours, while the 'longer' peptide E744-62 (peptide 8Q, harbouring CTL and Th epitopes)-based vaccines were also effective against MHC class I-deficient tumours. We have compared the adjuvant efficacies of two CpG ODN, CpG ODN 1826 and CpG ODN 1585. The 8Q peptide immunisation combined with CpG ODN 1585 inhibited growth of the TC-1/A9 tumours more effectively as compared to CpG ODN 1826. Further, we investigated the efficacy of cellular vaccines based on ex vivo cultured dendritic cells pulsed with either E749-57 or E744-62 peptides and matured with CpG ODN 1826. Unlike in the peptide immunisation setting, treatment with dendritic cells pulsed with a 'short' peptide resulted in the tumour growth inhibition, albeit weaker as compared to the immunisation with the longer peptide. Our data demonstrate that peptide and dendritic cell-based vaccines can be designed to elicit protective immunity against MHC class I-deficient tumours.
- MeSH
- adjuvantní radioterapie metody MeSH
- CpG ostrůvky MeSH
- dendritické buňky cytologie MeSH
- epitopy chemie MeSH
- geny MHC třídy I MeSH
- lidé MeSH
- myši MeSH
- oligonukleotidy genetika MeSH
- Papillomavirus E7 - proteiny chemie MeSH
- peptidy chemie MeSH
- protinádorové vakcíny chemie MeSH
- průtoková cytometrie MeSH
- regulace genové exprese MeSH
- subjednotkové vakcíny genetika MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- myši MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, N.I.H., Extramural MeSH
