Variations in disease severity outcomes for Influenza live viral challenges in man: Meta-

14 Objectives: Live Influenza challenges in man are valuable models for testing the efficacy of 15 vaccines and antiviral agents. Volunteers are treated with an investigational agent and their 16 clinical outcomes post-challenge are compared to those of Placebo treated volunteers. Despite 17 using a common protocol, recruitment criteria and a similar dose of the same challenge strain, 18 we noticed differences in disease severity outcomes between Placebo groups from different 19 studies. We investigated whether these differences were significant and, if so, whether any 20 pattern was identifiable and its possible causes. 21 Methods: We compared the clinical outcomes post-challenge in Placebo groups from five 22 clinical studies carried out between 2008 and 2013. Correlations between pre-challenge 23 heterosubtypic cellular response (IFN−γ) and post-challenge clinical outcomes were also 24 investigated in one study. 25 Results: Placebo groups from studies carried out between 2009 and 2010 attained significantly 26 reduced (p<0.05) symptom scores post-challenge than Placebo groups from studies carried out 27 in either 2008 or 2013. Also, in a 2010 study, the frequency of high influenza heterosubtypic 28 cellular responders pre-vaccination was significantly lower in the test group (FLU-v) than in the 29 Placebo group (p=0.04). Moreover, the Placebo group’s increased pre-existing heterosubtypic 30 cellular response correlated with reductions in symptom score and viral shedding post31 challenge (p≤0.023). Only post-vaccination did the Test group display an equivalent correlation. 32 Conclusions: The last influenza pandemic coincides with a significant reduction in disease 33 severity outcomes. This reduction also appears to correlate with increased pre-existing 34 influenza heterosubtypic cellular responses. 35 on A uust 7, 2017 by gest httpcvi.asm . rg/ D ow nladed fom INTRODUCTION 36 Live influenza challenges in man are valuable models for testing the efficacy of vaccines and 37 antiviral agents. Their basis is simple: a group of volunteers is treated with an investigational 38 agent and their clinical outcomes post-challenge are compared to those of a group of Placebo 39 treated volunteers. Their logistics, in contrast, are complex. 40 Influenza infection elicits a range of immune responses. One such response is the production of 41 strain-specific neutralising antibodies that confer immunity against infection by the same strain 42 (1). As a result, a key volunteer exclusion criterion in challenge studies is the detection of pre43 existing neutralising antibodies (HAI>10) to the challenge strain. Another such response is the 44 generation of antiviral cellular immune responses. Despite existing evidence as to their 45 protective role during infection (2-4), pre-existing cellular immune responses to the challenge 46 strain are not normally assessed during volunteer recruitment. 47 We have developed a novel vaccine (FLU-v) that elicits broad influenza heterosubtypic cellular 48 responses without inducing any significant antibody response (5,6, 7). In humans, FLU-v was 49 found to be safe, well tolerated and, in a live challenge study, to induce a vaccine specific 50 cellular response whose magnitude correlated with reductions in symptom score and viral 51 shedding (7). No such correlations were seen in the Placebo group, but we did notice that both 52 viral shedding and symptom score post-challenge were much lower (50%) in our Placebo group 53 than in the Placebo group from a previous study. To establish the significance of these 54 differences, we compared the Placebo group outcomes of several other live influenza challenge 55 studies. All these studies, although involving different placebo agents, were carried out by the 56 same clinical group by the same clinical group (Retroscreen Ltd), using the same recruitment 57 on A uust 7, 2017 by gest httpcvi.asm . rg/ D ow nladed fom criteria, the same viral strain and dose and the same method for determining post-challenge 58 clinical and virological outcomes. This meta-analysis revealed an experiment of nature that we 59 believe provides interesting insights in the potential of the cellular immune system in 60 controlling Influenza infection. 61