New Trends in Tumor Virology: Annecy Jan 19, 2010 Prophylactic HPV vaccines: next generations Lutz Gissmann l.gissmann@dkfz.de
Conflict of interest LG is a consultant to GSK and Sanofi Pasteur MSD and, due to existing IP, receives royalties from sales of Gardasil and Cervarix TM 2 Gissmann
1 st generation prophylactic vaccines: HPV L1 VLPs N wildtyp L1 C recombinant baculovirus, yeast 5x 72x monomer capsomer 100nm neutralizing antibodies capsid (VLP) 3 Gissmann
Prophylactic HPV vaccines Gardasil (MSD) Cervarix (GSK) composition HPV 6/11/16/18 HPV 16/18 adjuvant AAHS 1 AS04 2 scheme i.m. 0,2,6 months i.m. 0,1,6 months availability June 2006 October 2007 1 AAHS: Amorphous Aluminum Hydroxyphosphate 2 ASO4: Aluminum hydroxide + 3-deacetylated monophosphoryl Lipid A (MPL): detoxified lipopolysaccharide prepared from Salmonella minnesota 4 Gissmann
Strengths and weaknesses of the existing HPV vaccines are safe and highly efficient in clinical trials have the potential to significantly reduce the burden of cervical and other HPV-related cancers don t prevent all cases of cervical cancer and precursors continuation of screening don t treat existing infections or lesions are not particularly suited for use in areas of highest need cost, stability, multiple and invasive immunization 5 Gissmann
Future HPV Vaccines: Basic and Applied Research conducting research accompanying the introduction of the HPV vaccine exploiting existing vaccine in off-label trials modifying existing vaccines supplementing prophylactic vaccines developing HPV vaccines for other targets 6 Gissmann
Future HPV Vaccines: Basic and Applied Research conducting research accompanying the introduction of the HPV vaccine exploiting existing vaccine in off-label trials modifying existing vaccines supplementing prophylactic vaccines developing HPV vaccines for other targets 7 Gissmann
Future HPV Vaccines: Basic and Applied Research conducting research accompanying the introduction of the HPV vaccine exploiting existing vaccine in off-label trials modifying existing vaccines supplementing prophylactic vaccines developing HPV vaccines for other targets 8 Gissmann
Research accompanying the introduction of the HPV vaccine unresolved issues need to be address in population-based trials duration of protection immune correlate for protection replacement by uncovered HPV types efficacy in real life reduction in cancer incidence comparison of the different products analysis of bonus effects (protection against other tumors) 9 Gissmann
Future HPV Vaccines: Basic and Applied Research conducting research accompanying the introduction of the HPV vaccine exploiting existing vaccine in off-label trials modifying existing vaccines supplementing prophylactic vaccines developing HPV vaccines for different targets 10 Gissmann
Modifying the existing vaccines Special need for developing countries multivalent long-protecting without boosting immunogenic by non-invasive application 11 Gissmann
conserved N-terminus of PV L2 induces cross-neutralizing yet low titer abs 12 Gissmann Buck et et al 2008 J Virol 82:5190
Multimerization of L2 epitopes displayed by thioredoxin 13 Gissmann
Multimerized HPV 16 L2 epitope (20-38) induces high-titer neutralizing abs 14 Gissmann
Needle-less single-dose immunization: recombinant Adeno-Associated Virus (AAV) AAVs are apathogenic in humans AAVs induce persistent infections 15 Gissmann
HPV 16 L1 raav induces L1-specific serum antibodies after i.n. immunization 16 Gissmann Kuck et al. 2006
Single-dose immunization (intranasal) induces long-lasting immune response 17 Gissmann Kuck et al. 2006
Modifying the existing vaccines Special need for developing countries AFFORDABLE AND ROBUST multivalent long-protecting without boosting immunogenic by non-invasive application 18 Gissmann
HPV 16 L1 capsomeres versus VLPs: costly, >stable, = immunogenic N D 1-10 C E. coli 5x monomer capsomer 19 Gissmann
GST renders L1 soluble: purification of immunogenic L1 capsomeres Thrombin cleavage site GST HPV 16 L1 L1 GST lysate + ATP, MgCl 2 + 3,5 M urea centrifugation supernatant dialysis overnight gel filtration Binding to Glutathione- sepharose beads Thrombin cleavage Elution L1 100nm 20 Gissmann
assembly of capsomeres into VLPs ph = 5.4 1M NaCl D 1-10 175, 428 Cys Ser D409-431 (Dh4) T=7 T=7, T=4? capsomeres 21 Gissmann
The ability of L1 capsomeres to assemble into VLPs correlates with immunogenicity Dilution in PBS 37 C Stable particles immunogenicity Instable particles Capsomeres Capsomeres 22 Gissmann
HPV 16 L1-specific IgG titer Immunogenicity of HPV 16 L1 capsomeres correlates with VLP formation HPV16 L1-specific antibody titers 3.4 10 07 1.0 10 06 3.3 10 04 1.0 10 03 3.2 10 01 1.0 10 00 L1DN10DC29 L1D N10 dcys DC29-E7 L1DN10Dh4 DC29-E7 E7-L1D N10 Dh4DC29 CVLPs E7 pep 23 Gissmann
Immunogenicity of HPV 16 L1 capsomeres can be enhanced by adjuvants 6 rhesus macaques/group, 10 g HPV 16 capsomeres +/- adjuvant @ week 0 and 8 24 Gissmann
Future HPV Vaccines: Basic and Applied Research conducting research accompanying the introduction of the HPV vaccine exploiting existing vaccine in off-label trials modifying existing vaccines supplementing prophylactic vaccines developing HPV vaccines for different targets 25 Gissmann
Supplementing prophylactic vaccines therapeutic vaccines treatment of precursors combi vaccines with prophylactic and therapeutic features post-exposure vaccination 26 Gissmann
HPV Vaccines: different options for attack HPV tumor cell E6/E7 epithelial cell kill APC E6/E7 E6/7-specific CTL preventing infection (disease) neutralizing antibodies curing disease E6/E7-specific CTLs 27 Gissmann
Supplementing prophylactic vaccines therapeutic vaccines treatment of precursors combi vaccines with prophylactic and therapeutic features post-exposure vaccination 28 Gissmann
HPV-specific vaccines: windows of attack Prophylaxis Therapy pre- post-exposure INFECTION persistence HSIL cancer age [y] 15 20 50 + + +? antibodies +? + + + T cells (Th1) 29 Gissmann
HPV chimeric capsomeres L1 E7/E6 5x monomer capsomere prophylactic and therapeutic features in mice? treating persistent infections and preventing reinfection in a post-exposure scenario? suitable for developing countries: phase I clinical trial 30 Gissmann
Regression of HPV 16 E7 pos tumors 31 Gissmann
Summary The current HPV vaccines, although of very good performance, are far from being perfect. Several lines of research are needed to fully exploit the potential of the existing vaccines and to generate vaccines for different applications either following the existing concept (VLP) or developing novel strategies. 32 Gissmann
Acknowledgements Thanx Birgit Aengeneyndt Corinna Klein Yvonne Rubio Lysann Schädlich Tilo Senger Martin Müller Jürgen Kleinschmidt Robert L. Garcea Massimo Tommasino 33 Gissmann
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