Dissecting innate immune responses to plasmodia and their impact on disease and vaccination

LEIF ERIK SANDER (Charité) in partnership with Ian Cockburn (ANU)

Deciphering innate immune sensing mechanisms of malaria would yield therapeutic targets and aid the development of targeted vaccination strategies. Various plasmodial pathogen-associated molecular patterns (PAMPs) have been described [1], among which glycosylphosphatidolinositol anchors, hemozoin, plasmodial DNA, and RNA have been studied in greater detail in mice [2-5]. In contrast, only limited data regarding innate sensing pathways of plasmodia in humans, and their functions during disease and vaccination exist. The Sander lab has recently identified bacterial RNA as a so-called viability-associated PAMP (vita-PAMP), which alerts the immune system to the presence of viable microorganisms [6, 7]. Unpublished data show that human antigen presenting cells (APC) distinguish between live and dead bacteria via TLR8-mediated recognition of bacterial RNA. Bacterial RNA triggers inflammatory cytokines including TNF, IL-1b and IL-12, and strongly skews T helper- and B cell responses [6; Ugolini et al., unpublished]. We will investigate the mechanisms and downstream immunological effects of innate immune recognition of viable plasmodia and plasmodial RNA.

Work package 1 (Leif Erik Sander & Florian Kurth): Assess human innate immune responses to blood stage plasmodia. Stimulate human APC, including monocytes, dendritic cells, and the THP-1 cell line, with live and heat inactivated blood stage P. falciparum (Pf) and purified PfRNA (Pf culture together with Frank Mockenhaupt). The innate immune response to Pf will be fully mapped in vitro by measurement of APC maturation, cytokine production and transcriptome analysis. The response will be compared to APC from malaria patients ex vivo. Sample processing, cell culture and analyses will be subject of two MD theses.

Work package 2 (Leif Erik Sander & Florian Kurth): Investigate the human sensing mechanisms of Pf infection. The group will use lentiviral knockdowns and, if necessary, focused lentiviral screens to decipher sensing pathways for plasmodial PAMPs. They will investigate potential correlations of gene polymorphisms with malaria susceptibility (together with Frank Mockenhaupt). Key findings will be translated to a murine system using P. berghei (Pb) to investigate if similar sensing mechanisms apply to Pb in murine APC and if innate sensing can be improved by transgenic expression of bacterial flagellin (together with Kai Matuschewski), which strongly enhances immunogenicity of tumor cells. For this part the group will also use plasmodial liver stages, which are frequently used for vaccination.

Work package 3 (Ian Cockburn): Targeting innate sensing to improve malaria vaccines in vivo. The group will then translate key findings to an experimental vaccination system by use of live or heat inactivated transgenic Pb expressing Pf model antigens (e.g. circumsporozoite protein) +/- flagellin (together with Kai Matuschewski) and assess vaccine responses in mice. The project will make use of established vaccination systems and the immunological tool sets available in the Cockburn laboratory (e.g. circumsporozoite protein-specific T-cell receptor and B-cell receptor transgenic mice).

Interlinkages: Florian Kurth (Charité), Frank Mockenhaupt (Charité), Kai Matuschewski (HUB)

References:

(1) Gazzineli, R.T. et al. (2014) Nat. Rev. Immunol. 14: 744-757

(2) Jani, D. et al. (2008) PLoS Pathog. 4: e1000053

(3) Parroche, P. et al. (2007) Proc. Natl, Acad. Sci. USA 104: 1919-1924

(4) Sharma, S. et al. (2011) Immunity 35: 194-207

(5) Liehl, P. et al.  (2014) Nat. Med. 20: 47-53

(6) Sander, L.E. et al. (2011) Nature 474: 385-389

(7) Blander, J.M. and Sander, L.E. (2012) Nat. Rev. Immunol. 12: 215-255