Theme 9: Infection and innate immunity

Supervisors:

  A/Professor Kumar Visvanathan
  T: +61 3 9594 5548
E: Kumar.Visvanathan@med.monash.edu.au

1. Possible role of chemokine antagonists for the treatment of malaria

Malaria kills millions worldwide, but therapeutic choices are becoming increasingly limited by resistance. Developing new classes of antimalarials from bench to bedside is a slow and costly process. Using existing drugs, created for other purposes, is likely to significantly accelerate that process since safety and pharmacokinetic and pharmacodynamic data are already extant. The fact that they haven’t been previously used as antimalarials also lessens the risk of drug resistance. Vivax malaria is an important human pathogen causing up to 435 million human infections per annum. Most of these cases occur in the Asia Pacific region, and despite the reputation of vivax malaria as benign cause considerable morbidity and mortality. Chloroquine resistance is an increasing problem in P vivax infections especially in New Guinea. P knowlesii is an additional emerging infection in SE Asia which may also be vulnerable to DARC receptor blockade.

Vivax merozoites invade red blood cells using the ubiquitous chemokine receptor DARC (Duffy antigen receptor for chemokines). Maraviroc and other CCR5 antagonists have been developed to block HIV attachment to another specific chemokine receptor. These CCR5 antagonists are small molecules designed to mimic the attachment of chemokines to their specific receptor. However, DARC is a non-specific site of attachment by chemokines with a similar chemokine attachment site for some of the same chemokines that attach to CCR5. Currently no data exist regarding the attachment of chemokine receptor antagonists to DARC.

2. Vitamin D in chronic viral infections

A one year programme to examine the concept of vitamin D supplementation as an adjuvant for vaccination with hepatitis B vaccine undertaken in a laboratory with expertise in immunology of innate immunity and hepatitis B.

We propose to vaccinate 50 normal volunteers with hepatitis B (HBV) vaccine according to the manufacturer's recommendations. Vaccinees will also receive at the time of vaccination 1.0 mL of a oral Vitamin D emulsion in olive oil containing 50,000 IU Vitamin D or a matching oral placebo (olive oil). HBV vaccine was chosen because of its inclusion in the EPI and the strong serologic correlation with clinical protection form infection. The second dose of the vaccine will be administered at  one month and the third at 6 months, these doses will not be accompanied by Vitamin D.

Blood and saliva samples will be obtained at the time of the first HBV vaccine dose, at one month post dose 1, and one month following the 6 month vaccination (dose 3). After obtaining written consent, whole blood will be collected. Serum will be separated and stored at –70C for further analysis including anti-HBs and liver enzyme (AST and ALT) and Vitamin  D levels. Participants will be randomized into intervention and placebo groups without reference to the initial vitamin D level because we wish to investigate the potential adjuvant effect that would be translatable to a population setting where individual vitamin D levels would not be available.

Analysis: We will assess both systemic and mucosal responses to the vaccine. To assess the mucosal response to the vaccine, saliva samples will be obtained by use of a sterile sponge swab. Specific salivary levels of IgA and IgG antibody against the HBV vaccine will be determined by ELISA.  Arbitrary values will calculated by using weighted probit regression analysis and an internal serum standard.

Peripheral blood mononuclear cells (PBMCs) will be separated. Cells are then subjected to stimulation with HBs antigen were 5 ug/mL for 16 hours and intracellular cytokine analysis performed on CD4 positive T cells looking for IFN-g and IL-4. Serum samples will be  analyzed for anti-HBs antibody by using AxSYM AUSAB_ (Abbott Diagnostics Division,Wiesbaden, Germany). Excess PBMCs will be used to examine markers of innate immunity including Toll like receptor expression and function. All of the techniques mentioned are currently established in our laboratory. Pairwise comparisons between placebo and intervention groups will be performed at each time point of the systemic and mucosal antibody titres,and also the fraction attaining an antibody level over 10. We will also perform an analysis stratified for baseline vitamin D levels as a categorical variable comparing those with moderate-severe vitamin D deficiency (25OHD<50nmol/l) to those with normal vitamin D levels. The vitamin D supplement half-life of 2-3 months implies that its effect will have abated by the 6 month dose of the vaccine. We will be able to test therefore whether the single dose used in our protocol affects long term antibody responses or not.