Deployable Laboratory Response to Influenza Pandemic; PCR Assay Field Trials and Comparison with Reference Methods

Background: The influenza A/H1N1/09 pandemic spread quickly during the Southern Hemisphere winter in 2009 and reached epidemic proportions within weeks of the official WHO alert. Vulnerable population groups included indigenous Australians and remote northern population centres visited by international travellers. At the height of the Australian epidemic a large number of troops converged on a training area in northern Australia for an international exercise, raising concerns about their potential exposure to the emerging influenza threat before, during and immediately after their arrival in the area. Influenza A/H1N1/09 became the dominant seasonal variant and returned to Australia during the Southern winter the following year. Methods: A duplex nucleic acid amplification assay was developed within weeks of the first WHO influenza pandemic alert, demonstrated in northwestern Australia shortly afterwards and deployed as part of the pathology support for a field hospital during a military exercise during the initial epidemic surge in June 2009. Results: The nucleic acid amplification assay was twice as sensitive as a point of care influenza immunoassay, as specific but a little less sensitive than the reference laboratory nucleic acid amplification assay. Repetition of the field assay with blinded clinical samples obtained during the 2010 winter influenza season demonstrated a 91.7% congruence with the reference laboratory method. Conclusions: Rapid in-house development of a deployable epidemic influenza assay allowed a flexible laboratory response, effective targeting of limited disease control resources in an austere military environment, and provided the public health laboratory service with a set of verification tools for resource-limited settings. The assay method was suitable for rapid deployment in time for the 2010 Northern winter

31st Annual Meeting and Associated Programs of the Society for Immunotherapy of Cancer (SITC 2016) : part two

Background The immunological escape of tumors represents one of the main ob- stacles to the treatment of malignancies. The blockade of PD-1 or CTLA-4 receptors represented a milestone in the history of immunotherapy. However, immune checkpoint inhibitors seem to be effective in specific cohorts of patients. It has been proposed that their efficacy relies on the presence of an immunological response. Thus, we hypothesized that disruption of the PD-L1/PD-1 axis would synergize with our oncolytic vaccine platform PeptiCRAd. Methods We used murine B16OVA in vivo tumor models and flow cytometry analysis to investigate the immunological background. Results First, we found that high-burden B16OVA tumors were refractory to combination immunotherapy. However, with a more aggressive schedule, tumors with a lower burden were more susceptible to the combination of PeptiCRAd and PD-L1 blockade. The therapy signifi- cantly increased the median survival of mice (Fig. 7). Interestingly, the reduced growth of contralaterally injected B16F10 cells sug- gested the presence of a long lasting immunological memory also against non-targeted antigens. Concerning the functional state of tumor infiltrating lymphocytes (TILs), we found that all the immune therapies would enhance the percentage of activated (PD-1pos TIM- 3neg) T lymphocytes and reduce the amount of exhausted (PD-1pos TIM-3pos) cells compared to placebo. As expected, we found that PeptiCRAd monotherapy could increase the number of antigen spe- cific CD8+ T cells compared to other treatments. However, only the combination with PD-L1 blockade could significantly increase the ra- tio between activated and exhausted pentamer positive cells (p= 0.0058), suggesting that by disrupting the PD-1/PD-L1 axis we could decrease the amount of dysfunctional antigen specific T cells. We ob- served that the anatomical location deeply influenced the state of CD4+ and CD8+ T lymphocytes. In fact, TIM-3 expression was in- creased by 2 fold on TILs compared to splenic and lymphoid T cells. In the CD8+ compartment, the expression of PD-1 on the surface seemed to be restricted to the tumor micro-environment, while CD4 + T cells had a high expression of PD-1 also in lymphoid organs. Interestingly, we found that the levels of PD-1 were significantly higher on CD8+ T cells than on CD4+ T cells into the tumor micro- environment (p < 0.0001). Conclusions In conclusion, we demonstrated that the efficacy of immune check- point inhibitors might be strongly enhanced by their combination with cancer vaccines. PeptiCRAd was able to increase the number of antigen-specific T cells and PD-L1 blockade prevented their exhaus- tion, resulting in long-lasting immunological memory and increased median survival

Flora Barbiensis : in usum seminarii fratrum

edidit Frid. Adam SchollerTitelkupfe

Performance of deployable assay for detection of influenza A/H1N1 2009 according to specimen type and viral RNA extraction method.

<p>POCT  =  point of care test.</p

Deployable molecular microbiology laboratory.

<p>Deployable molecular biology equipment used during the exercise. The layout corresponds to the two modules; left table – static field hospital used for real time PCR assays, right table – field portable. Only the hand-held magnetic bead extraction device from the field portable module was used in an attempt to detect influenza A by PCR assay during the military exercise.</p