PLoS ONE (2013)  8(10): e78407.

A Glycolipid Adjuvant, 7DW8-5, Enhances CD8+ T Cell Responses Induced by an Adenovirus-Vectored Malaria Vaccine in Non-Human Primates.

Neal N. Padte, Mar Boente-Carrera, Chasity D. Andrews, Jenny McManus, Brooke F. Grasperge, Agegnehu Gettie, Jordana G. Coelho-dos-Reis, Xiangming Li, Douglass Wu, Joseph T. Bruder, Martha Sedegah, Noelle Patterson, Thomas L. Richie, Chi-Huey Wong, David D. Ho, Sandhya Vasan, Moriya Tsuji

 

ABSTRACT

A key strategy to a successful vaccine against malaria is to identify and develop new adjuvants that can enhance T cell responses and improve protective immunity. Upon co-administration with a rodent malaria vaccine in mice, 7DW8-5, a recently identified novel analog of α-galactosylceramide (α-GalCer), enhances the level of malaria specific protective immune responses more strongly than the parent compound. In this study, we sought to determine whether 7DW8-5 could provide a similar potent adjuvant effect on a candidate human malaria vaccine in the more relevant non-human primate (NHP) model, prior to committing to clinical development. The candidate human malaria vaccine, AdPfCA (NMRC-M3V-Ad-PfCA), consists of two non-replicating recombinant adenoviral (Ad) vectors, one expressing the circumsporozoite protein (CSP) and another expressing the apical membrane antigen-1 (AMA1) of Plasmodium falciparum. In several phase 1 clinical trials, AdPfCA was well tolerated and demonstrated immunogenicity for both humoral and cell-mediated responses. In the study described herein, 25 rhesus macaques received prime and boost intramuscular (IM) immunizations of AdPfCA alone or with an ascending dose of 7DW8-5. Our results indicate that 7DW8-5 is safe and well-tolerated and provides a significant enhancement (up to 9-fold) in malaria-specific CD8+ T-cell responses after both priming and boosting phases, supporting further clinical development.

PMID: 24205224

 

SUPPLEMENT

A highly effective vaccine against the malaria pandemic remains elusive.  The AdPfCA vaccine (NMRC-M3V-Ad-PfCA; is a cocktail of two replication-deficient, recombinant Ad serotype 5 vectors, encoding for the sporozoite/early hepatic stage antigen, circumsporozoite protein (PfCSP), and the sporozoite/hepatic stage/erythrocytic stage antigen, apical membrane antigen-1 (PfAMA1) of P. falciparum (3D7 strain). Despite the induction of strong PfCSP- and PfAMA1 mediated T-cell responses and modest humoral responses by AdPfCA in clinical trials, volunteers later challenged by the bite of P. falciparum-infected mosquitoes all became parasitemic (1,2).  We sought to improve the immune response to AdPfCA by combining it with a novel glycolipid-based adjuvant, 7DW8-5 which has the unique property of boosting CD8+ T cell responses in a mouse model (3).

7DW8-5 is a glycosphingolipid compound derivative of α-galactosyl ceramide (α-GalCer), which binds the CD1d receptor to activate the natural killer T (NKT) cell, an innate immune cell that non-specifically triggers downstream activation of dendritic cells and other innate and adaptive immune cells (3). Originally developed as an anti-cancer agent, α-GalCer was shown to act as an adjuvant to enhance immune responses to vaccination (4).  7DW8-5 was identified from a screening library of α-GalCer analogues as having stronger binding affinity to the CD1d receptor than α-GalCer, inducing stronger NKT cell activation in vitro and a more potent adjuvant effect on both DNA and adenoviral based vaccines in mice (3).  While most adjuvants that are currently licensed or in development focus on enhancing the humoral response to proteins, glycosphingolipids enhance the cellular response to DNA-based vaccines and viral vectors.  We therefore sought to determine whether 7DW8-5 would enhance the response to AdPfCA in nonhuman primates, which more closely mimic the NKT cell repertoire of humans (5).

7DW8-5 was co-administered at varying doses (0 μg, 0.1 μg, 1 μg, 10 μg or 100 μg) with a fixed dose of AdPfCA intramuscularly to rhesus macaques pre-screened to normalize circulating NKT cell levels across groups, at week 0 and week 25.  As depicted in Figure 1, 7DW8-5 enhanced the gamma interferon ELISpot response to CSP and AMA-1 peptides after both the prime and boost over AdPfCA alone.  CD4+ and CD8+ T cell depletion assays revealed that the effect was primarily mediated through CD8+ T cells.  The adjuvant effect on the cellular response was more pronounced than on the humoral response.

 

fig1Figure 1:  Gamma Interferon ELISpot to CSP and AMA1 Antigens Post Vaccination with Increasing 7DW8-5 Adjuvant

 

To more closely understand the mechanism of the adjuvant effect, we studied the number of circulating dendritic cells after vaccination and found that circulating dendritic cells increased in a dose-dependent manner immediately after co-administration with 7DW8-5.  Using three different activation markers, we demonstrated that these dendritic cells were significantly more activated after exposure to vaccination with 7DW8-5 than the unadjuvanated vaccine.  Finally, as expected with NKT cell activation, circulating NKT cell numbers dropped immediately after vaccination with 7DW8-5 plus AdPfCA, but not AdPfCA alone, and then recovered quickly.  Taken together, these results indicate a specific mechanism for the adjuvant effect on the cellular immune response via indirect activation of dendritic cells through NKT cell activation.

In order to support future clinical development of 7DW8-5 as an adjuvant in humans, macaques were closely assessed both clinically and via monitoring of circulating cytokines to support the safety of this novel molecule in human clinical trials.  Mild erythema was noted post vaccination that increased in proportion to 7DW8-5 dose, but was overall transient and not accompanied by systemic reactogenicity, defined by changes in body temperature, heart rate, or respiratory rate.  Furthermore, analysis of circulating cytokines 24 hours post priming vaccination revealed that the changes in circulating cytokines assessed were minimal and that their peak and trough levels in all of the 7DW8-5 dose groups over 24 hours were not statistically different as compared to the peak and trough cytokine levels of the control group.

This first study of 7DW8-5 in rhesus macaques is important in demonstrating that it provides a potent adjuvant effect to the CD8+ T cell response to both antigens in AdPfCA, unlike the majority of adjuvants which focus their effect on increasing the humoral response to protein vaccines.  Furthermore, the reassuring safety profile in nonhuman primates paves the way to move forward with clinical trials in humans (6).  If 7DW8-5 proves to be an effective adjuvant in humans, this could be formulated with vaccines against other pathogens requiring a strong CD8+ T cell effector response for prevention or control.

 

REFERENCES

1.   Sedegah M, Tamminga C, McGrath S, House B, Ganeshan H et al.  (2011) Adenovirus 5-vectored P. falciparum vaccine expressing CSP and AMA1. Part A: safety and immunogenicity in seronegative adults. PLOS ONE 6: e24586.

2.  Tamminga C, Sedegah M, Regis D, Chuang I, Epstein JE et al. (2011) Adenovirus-5-vectored P. falciparum vaccine expressing CSP and AMA1. Part B: safety, immunogenicity and protective efficacy of the  CSP component. PLOS ONE 6: e25868.

3. Li X, Fujio M, Imamura M, Wu D, Vasan S et al. (2010) Design of a potent CD1d-binding NKT cell ligand as a vaccine adjuvant. Proc Natl Acad Sci U S A 107: 13010-13015.

4.  Gonzalez-Aseguinolaza G, Van Kaer L, Bergmann CC, Wilson JM, Schmieg J et al. (2002) Natural killer T cell ligand alphagalactosylceramide enhances protective immunity induced by malaria vaccines. J Exp Med 195: 617-624.

5. Padte NN, Boente-Carrera M, Andrews CD, McManus J, Grasperge BF, et al. (2013) A Glycolipid Adjuvant, 7DW8-5, Enhances CD8+ T Cell Responses Induced by an Adenovirus-Vectored Malaria Vaccine in Non-Human Primates . PLoS ONE 8(10): e78407.

6.  Padte NN, Li X, Tsuji M, Vasan S (2011) Clinical development of a novel CD1d-binding NKT cell ligand as a vaccine adjuvant. Clin Immunol 140: 142-151.

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