title: Plastid expression of a double-pentameric vaccine candidate containing human papillomavirus-16 L1 antigen fused with LTB as adjuvant: transplastomic plants show pleiotropic phenotypes.
abstract: Human papillomavirus (HPV) causes cervical cancer in women worldwide, which is currently prevented by vaccines based on virus-like particles (VLPs). However, these vaccines have certain limitations in their availability to developing countries, largely due to elevated costs. Concerning the highest burden of disease in resource-poor countries, development of an improved mucosal and cost-effective vaccine is a necessity. As an alternative to VLPs, capsomeres have been shown to be highly immunogenic and can be used as vaccine candidate. Furthermore, coupling of an adjuvant like Escherichia coli heat-labile enterotoxin subunit B (LTB) to an antigen can increase its immunogenicity and reduce the costs related to separate co-administration of adjuvants. Our study demonstrates the expression of two pentameric proteins: the modified HPV-16 L1 (L1_2xCysM) and LTB as a fusion protein in tobacco chloroplasts. Homoplasmy of the transplastomic plants was confirmed by Southern blotting. Western blot analysis showed that the LTB-L1 fusion protein was properly expressed in the plastids and the recombinant protein was estimated to accumulate up to 2% of total soluble protein. Proper folding and display of conformational epitopes for both LTB and L1 in the fusion protein was confirmed by GM1-ganglioside binding assay and antigen capture ELISA, respectively. However, all transplastomic lines showed chlorosis, male sterility and growth retardation, which persisted in the ensuing four generations studied. Nevertheless, plants reached maturity and produced seeds by pollination with wild-type plants. Taken together, these results pave the way for the possible development of a low-cost adjuvant-coupled vaccine with potentially improved immunogenicity against cervical cancer.
authors: Department of Applied Plant Sciences and Plant Biotechnology (DAPP), University of Natural Resources and Applied Life Sciences (BOKU), Vienna, Austria.
authors: Mohammad T
authors: MT
authors: Waheed
authors: Nadja
authors: N
authors: Thönes
authors: Martin
authors: M
authors: Müller
authors: Syed W
authors: SW
authors: Hassan
authors: Johanna
authors: J
authors: Gottschamel
authors: Elke
authors: E
authors: Lössl
authors: Hans-Peter
authors: HP
authors: Kaul
authors: Andreas G
authors: AG
authors: Lössl
mesh_terms: D000276:Adjuvants, Immunologic / Q000235:genetics; D000956:Antigens, Viral / Q000235:genetics; D001427:Bacterial Toxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D036022:Capsid Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D002736:Chloroplasts / Q000235:genetics / Q000378:metabolism; D004768:Enterotoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D000939:Epitopes; D029968:Escherichia coli Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005260:Female; D005677:G(M1) Ganglioside / Q000378:metabolism; D006801:Humans; D009856:Oncogene Proteins, Viral / Q000737:chemistry / Q000235:genetics / Q000276:immunology*; D053918:Papillomavirus Vaccines / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D010641:Phenotype; D051479:Plant Infertility / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics; D018087:Plastids / Q000235:genetics; D054817:Pollination; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D012639:Seeds / Q000502:physiology; D014026:Nicotiana / Q000235:genetics / Q000254:growth & development / Q000378:metabolism; D002583:Uterine Cervical Neoplasms / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D000956:Antigens, Viral; D001427:Bacterial Toxins; D036022:Capsid Proteins; D004768:Enterotoxins; D000939:Epitopes; D029968:Escherichia coli Proteins; D009856:Oncogene Proteins, Viral; D053918:Papillomavirus Vaccines; D011993:Recombinant Fusion Proteins; D005677:G(M1) Ganglioside; C063317:L1 protein, Human papillomavirus type 16; C027538:heat-labile enterotoxin, E coli