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

Listeria-derived recombinant proteins are processed by a unique MHC class I processing pathway (100.10)

Abstract Studies examining antigen processing in infected cells suggest that the efficiency of peptide-MHC class I (pMHC) production can differ by as much as 100 to 1000-fold depending on the source of the protein from which the peptide determinant is derived. For recombinant proteins expressed from vaccinia virus (rVV), ~3000 protein molecules must be degraded to generate one surface pMHC complex. By contrast, the measured efficiency of pMHC production from native Listeria monocytogenes proteins is ~1 surface pMHC complex per 3.5 to 35 protein molecules degraded. What accounts for this 100 to 1000-fold difference in efficiency remains unresolved. One possible explanation is that processing efficiency is a function of the peptide and/or protein studied. It is also possible that exogenously derived proteins, such as those secreted by Listeria, enter a different processing pathway than host-cell derived proteins. To better understand these differences in processing efficiency, we engineered Listeria strains to express recombinant proteins similar to those used for the rVV efficiency studies. We found that surface pMHC production was 20-fold more efficient from recombinant proteins expressed from Listeria than from a rVV-derived recombinant protein. These data suggest proteins expressed from Listeria follow a distinct and highly efficient processing pathway relative to host cell derived proteins.</jats:p

Proteasomes Get By with Lots of Help from Their Friends

AbstractProteasomes can't do it all. It was previously known that aminopeptidases frequently degrade proteasome-generated peptides. Now it appears that another protease, tripeptidyl peptidase II (TPP II), plays a critical role in cleaving proteasomal produced peptides into shorter peptides that can then be degraded by aminopeptidases

A vaccinia virus expression system to study antigen processing of multiple recombinant proteins (130.43)

Abstract A major obstacle in designing effective vaccines is our limited knowledge of the mechanisms involved in eliciting a protective cell-mediated immune response. In recent years, research has focused on the development of recombinant vaccines, in which antigenic peptides derived from a specific pathogen are delivered to the cells via live vectors. For a vaccine to be effective in an out-bred population it must generate as many antigenic determinants as possible. One method that can be used to generate multiple peptide determinants is to express two or more recombinant proteins from a single live vaccine vector. To this end, we developed a recombinant vaccinia virus system that expresses two full-length influenza virus proteins; nucleoprotein (NP) and acidic polymerase (PA). In addition to the NP and PA proteins, our expression system is designed to produce yellow and red fluorescent proteins, which allow us to monitor, in a quantitative manner, recombinant protein expression both in vitro and in vivo. Using this novel expression system we have found that the route of immunization affects the immunodominance hierarchy of the host response to not only vector-derived peptides but also to recombinant peptides.</jats:p