NIAID-DVI: DENVax -- Jorge E Osorio

Blog Series: NIAID-DVI

Pre-Clinical and clinical development of recombinant live attenuated tetravalent dengue vaccine (DENVax)
Jorge E Osorio
Takeda Vaccines

So Takeda Vaccines took over Inviragen and have been developing a tetravalent, live attenuated dengue vaccine called DENVax. It consists of a molecularly characterized, attenuated DENV-2 strain and three chimeras. The chimeras all have the backbone of the attenuated DENV-2 strain, but the prM and E genes have been swapped out with DENV-1, DENV-3 and DENV-4.

DENVax design from Osorio et al., 2011.

Rapid Fire:

•  DENVax has been shown safe in phase 1 clinical trials.
• Vaccine was highly immunogenic in non-human primates
• T cell responses with both T cell subsets producing proinflammatory cytokines, IFN-gamma, TNF-alpha and IL-2 one month after priming
• CD4+ and CD8+ T cells target structural and non-structural proteins.
• It is now in phase 1b clinical trial
• A clinical trial that usually tests a new treatment in a small number of patients in order to determine the safety and appropriate dose of the new treatment. Phase 1b trials may also test the treatment in combination with other drugs or agents. (source)
• Also being tested in phase 2 'age-de-escalation' clinical trial in four endemic countries.
• Phase 2: A clinical trial that usually tests a new treatment in a small number of patients who have the illness that the drug is designed to treat. Phase 2 trials measure efficacy as well as safety. (source)
• Prelim data suggests DENVax induced significant neutralizing antibody responses to all four dengue viruses after one or two administrations.
• Does not induce changes in serum chemistry or hematology
• No wildtype dengue challenge has been done yet with respect to this virus
• DENVax warrents further evaluation in Phase 2b/Phase 3 efficacy studies
• Phase 3: A clinical trial that definitively tests a new treatment, usually in a large number of patients who have the illness that the drug is designed to treat. Phase 3 trials measure efficacy as well as safety as compared to the standard of care. (source)

 Mel's Thoughts:

• The backbone is an attentuated dengue 2 virus called PDK-53
• The phylogeneticist in me will probably tree that virus and compare it with other trees (if possible). It's a virus that's quite old, 1974 according to the accession on Genbank. Assuming I am looking at the right virus...viral strain S16803. This strain has also been passaged 53 times. Yowza! Great for attenuation...ok I get that but genetically I'm sure quite far away from the currently circulating diversity of dengue. Perhaps this doesn't matter...perhaps it does.
• In the 53 passages 3 mutations were recorded, near the 5' UTR, NS1 and NS3 genes (indicated by black triangles in figure above). Surprising given the viruses I've looked at like New Guinea C have several mutations relative to the Genbank reference. Specifically 9 non-synonymous mutations, 7 synonymous mutations, 1 gap and 2 bases which I had to call ambiguously because sanger and next generation sequencing data (454) kept conflicting and the overall site quality was low. So just surprising is all that after 53 passages this virus is mostly unchanged.
• Following on this vein of thought and given my lack of background in cell passaging, vaccine development and how to 'attenuate' something I sought out  my Post Doc mentor MAJ/Dr. Richard Jarman who usually welcomes to chance to take a break from military or admin stuff to talk research and augment my understanding on all things infectious. Some good points he brought up:
1. We have no idea what the passage history on the virus, New Guinea C, is (which I just cleaned up following NGS sequencing).
2. The reason the PDK53 might not have as many mutations, aside from the fact it's inherently different than New Guinea C, probably has to do with it's passage history.
3. Full passage histories are a necessity when interpreting the changes in different viruses especially vaccine attenuated ones.
4. Passage history matters...you've got different cell types that will exert different 'forces of nature' onto your virus hence promoting or extinguishing it's ability to genetically adapt.
5. Cell type examples where passaging can be done: Vero cells, HeLa, Primary Dog Kidney Cells (PDK), MDCK (also of canine origin), hey did you know there's a cell line for zebrafish? Not applicable to viral passsaging of dengue, but interesting nonetheless. I've seen work with dengue virus passaging in C6/36 cells mimicking the mosquito and dendritic (DC) cells mimicking the human. There have also been forays into macrophages...

A moment to digress: I am not going to elaborate on macrophages because my background is limited but I must say everytime I hear 'macrophages' I think cell that eats things (ie. phagocyte) and then I get transported back to the first SciFi film my parents ever let me watch...from 1966 Fantastic Voyage. In that movie a guy gets eaten by a macrophage...

I couldn't find the clip of the guy getting nom'd by a macrophage...so here's the trailer, at about 1:45-1:50 little macrophages start attacking the girl...oh 1966 SciFi

Ok big digression aside...back to thoughts related to the meeting...

• Also of interest to me is the origin of the backbone which is Thailand according to Genbank. Do you think it will matter? American and Asian dengue viruses have been shown to be genetically distinct and actually Asian viruses have been shown to be potentially more virulent so I'm wondering if one will 'cover' the other's 'diversity'. [1, 2, 3] And I realize of course that genetics aren't the only factor in vaccine development; however that's my expertise so that's where I am coming from. The immune response of the individual matters and can 'shape' genetic diversity of the viral population (see next bullet)...if the in-going diversity is nearly '40 years old'...when it encounters 'newer' viral populations currently in circulation how will it respond? Can a old dog learn new tricks?
• And with respect to all vaccines...I'm always curious about the quasispecies component. I will dedicate a separate blog post to this issue. Now all these vaccine candidates are supposedly grown in culture and if they 'behave' like a dengue virus then they should theoretically produce quasispecies 'hopefully' that should cover the spectrum of dengue circulating. Yet I've seen no publications to this affect...and it interests (and concerns me).

So the DENVax literature buzz:

•  Osorio JE,et al. 2011. Development of DENVax: A chimeric dengue-2 PDK-53-based tetravalent vaccine for protection against dengue fever. Vaccine 29: 7251–7260.
•  Osorio JE et al., 2011. Efficacy of a Tetravalent Chimeric Dengue Vaccine (DENVax) in Cynomolgus Macaques. AJTMH 84: 978.
• Brewoo JN et al., 2012. Immunogenicity and efficacy of chimeric dengue vaccine (DENVax) formulations in interferon-deficient AG129 mice. Vaccine 30: 1513.

Some other nice background articles:

• Safety and Immunogenicity Study to Assess DENVax, a Live Attenuated Tetravalent Vaccine for Prevention of Dengue Fever
• Inviragen to initiate second stage of DENVax Phase 2 study in dengue
• Ospina NC et al. 2010. Prolonged Co-circulation of Two Distinct Dengue Virus Type 3 Lineages in the Hyperendemic Area of Medellín, Colombia. AJTMH 83: 672. 
• Mladinich KM et al. 2012. Dengue virus-specific CD4⁺ and CD8⁺ T lymphocytes target NS1, NS3 and NS5 in infected Indian rhesus macaques. Immunogenetics 64:111

Jorge has also been busy involved with efforts for a Chikungunya Vaccine:

•  Plante K et al., 2011. Novel Chikungunya Vaccine Candidate with an IRES-Based Attenuation and Host Range Alteration Mechanisms. PLoS Pathogens 7:e1002142
•  Chu H et al., 2013. Deciphering the protective role of adaptive immunity to CHIKV/IRES a novel candidate vaccine against Chikungunya in the A129 mouse model. Vaccine 31: 3353.
• Weaver SC et al., 2012. Chikungunya virus and prospects for a vaccine. Expert Reviews of Vaccines 11: 1087

References (linked in text as 1, 2, 3):

1. Cologna et al., 2005. Selection for Virulent Dengue Viruses Occurs in Humans and Mosquitoes. JVI 79: 853
2. Hang VT, et al. 2010. Emergence of the Asian 1 Genotype of Dengue Virus Serotype 2 in Viet Nam: In Vivo Fitness Advantage and Lineage Replacement in South-East Asia. PLoS Negl Trop Dis 4(7): e757. doi:10.1371/journal.pntd.0000757
3. Rico-Hesse, R. 2007. Dengue virus evolution and virulence models. Clin Infect Dis 44: 1462

 Update 9/24/2013: For those wanting more information about phase I, II, and III trials there's a nice blog post on Chimera's blog.