A preliminary randomized double blind placebo-controlled trial of intravenous immunoglobulin for Japanese encephalitis in Nepal

Abstract

Background: Japanese encephalitis (JE) virus (JEV) is a mosquito-borne flavivirus found across Asia that is closely related to West Nile virus. There is no known antiviral treatment for any flavivirus. Results from in vitro studies and animal models suggest intravenous immunoglobulin (IVIG) containing virus-specific neutralizing antibody may be effective in improving outcome in viral encephalitis. IVIG's anti-inflammatory properties may also be beneficial.

Methodology/principal findings: We performed a pilot feasibility randomized double-blind placebo-controlled trial of IVIG containing anti-JEV neutralizing antibody (ImmunoRel, 400mg/kg/day for 5 days) in children with suspected JE at two sites in Nepal; we also examined the effect on serum neutralizing antibody titre and cytokine profiles. 22 children were recruited, 13 of whom had confirmed JE; 11 received IVIG and 11 placebo, with no protocol violations. One child (IVIG group) died during treatment and two (placebo) subsequently following hospital discharge. Overall, there was no difference in outcome between treatment groups at discharge or follow up. Passive transfer of anti-JEV antibody was seen in JEV negative children. JEV positive children treated with IVIG had JEV-specific neutralizing antibody titres approximately 16 times higher than those treated with placebo (p=0.2), which was more than could be explained by passive transfer alone. IL-4 and IL-6 were higher in the IVIG group.

Conclusions/significance: A trial of IVIG for JE in Nepal is feasible. IVIG may augment the development of neutralizing antibodies in JEV positive patients. IVIG appears an appealing option for JE treatment that warrants further study.

Trial registration: ClinicalTrials.gov NCT01856205.

Fig 1. Flow diagram of study participants’ recruitment and follow-up.

All children enrolled, fitting the trial criteria, who were alive at discharge were attempted to be followed-up (n = 21). Twenty-one families were successfully contacted. Among these families, two children had died.

Fig 2. Anti-JEV neutralizing antibody in commercially available IVIG.

Mean and standard deviation of reciprocal 50% plaque reduction neutralization titres (PRNT₅₀) in vero cells using P3 wild type strain of JEV are shown for a: Serum control from JEV vaccinated individual, b: Vigam (USA), c: Bharat (India) batch 1, d: Hualan (China) batch 1, e: Bharat (India) batch 2, f: Hualan (China) batch 2, g: Sichuan (China), h: Reliance (India).

Fig 3. Difference in neutralizing antibody titres to JEV in children with acute encephalitis syndrome treated with IVIG or placebo.

Median and inter-quartile range of the difference in JEV PRNT₅₀ titres pre and post treatment is presented. Patients are grouped according to treatment. Difference in tires was assessed via Wilcoxon-Mann-Whitney test. Note: Two patients who received IVIG were not included in this analysis because of insufficient sample to undertake PRNT measurements.

Fig 4. Interleukin (IL)-4 and IL-6 abundance in children with acute encephalitis syndrome receiving intravenous immunoglobulin (IVIG) and placebo.

Median and inter-quartile range of change in cytokine abundance (pg/ml) pre and post treatment is presented for IL-4 and IL-6 separately. Cytokine abundance increased for both IL-4 and IL-6. This increase was significant for IL-4 (p = 0.043 and p = 0.068 for IL-4 and IL-6 respectively). Difference in abundance was assessed via Wilcoxon-Mann-Whitney test. Note: Four patients (three who received IVIG and one who received saline) were not included in this analysis because of insufficient sample to undertake the ELISA.