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

Schizosaccharomyces pombe Dss1p Is a DNA Damage Checkpoint Protein That Recruits Rad24p, Cdc25p, and Rae1p to DNA Double-strand Breaks*

Schizosaccharomyces pombe Dss1p and its homologs function in multiple cellular processes including recombinational repair of DNA and nuclear export of messenger RNA. We found that Tap-tagged Rad24p, a member of the 14-3-3 class of proteins, co-purified Dss1p along with mitotic activator Cdc25p, messenger RNA export/cell cycle factor Rae1p, 19 S proteasomal factors, and recombination protein Rhp51p (a Rad51p homolog). Using chromatin immunoprecipitation, we found that Dss1p recruited Rad24p and Rae1p to the double-strand break (DSB) sites. Furthermore, Cdc25p also recruited to the DSB site, and its recruitment was dependent on Dss1p, Rad24p, and the protein kinase Chk1p. Following DSB, all nuclear Cdc25p was found to be chromatin-associated. We found that Dss1p and Rae1p have a DNA damage checkpoint function, and upon treatment with UV light Δdss1 cells entered mitosis prematurely with indistinguishable timing from Δrad24 cells. Taken together, these results suggest that Dss1p plays a critical role in linking repair and checkpoint factors to damaged DNA sites by specifically recruiting Rad24p and Cdc25p to the DSBs. We suggest that the sequestration of Cdc25p to DNA damage sites could provide a mechanism for S. pombe cells to arrest at G2/M boundary in response to DNA damage

Homolog of BRCA2-interacting Dss1p and Uap56p link Mlo3p and Rae1p for mRNA export in fission yeast

The breast cancer tumor suppressor BRCA2-interacting protein, DSS1, and its homologs are critical for DNA recombination in eukaryotic cells. We found that Dss1p, along with Mlo3p and Uap56p, Schizosaccharomyces pombe homologs of two messenger RNA (mRNA) export factors of the NXF–NXT pathway, is required for mRNA export in S. pombe. Previously, we showed that the nuclear pore-associated Rae1p is an essential mRNA export factor in S. pombe. Here, we show that Dss1p and Uap56p function by linking mRNA adapter Mlo3p to Rae1p for targeting mRNA–protein complex (mRNP) to the proteins of the nuclear pore complex (NPC). Dss1p preferentially recruits to genes in vivo and interacts with –FG (phenylalanine glycine) nucleoporins in vivo and in vitro. Thus, Dss1p may function at multiple steps of mRNA export, from mRNP biogenesis to their targeting and translocation through the NPC

Supplementary Data 7 from ETV4-Dependent Transcriptional Plasticity Maintains <i>MYC</i> Expression and Results in IMiD Resistance in Multiple Myeloma

Genes whose expression are associated with ETV4 in NDMM and in RRMM patients enrolled in the CoMMpass study.</p

Supplementary Data 3 from ETV4-Dependent Transcriptional Plasticity Maintains <i>MYC</i> Expression and Results in IMiD Resistance in Multiple Myeloma

KZF1 ChIP-seq peaks as determined with MACS2 relative to MM1S cells input.</p

Supplementary Data 6 from ETV4-Dependent Transcriptional Plasticity Maintains <i>MYC</i> Expression and Results in IMiD Resistance in Multiple Myeloma

ETV4 mRNA data and outcome from the POLLUX and MM-010 trials.</p

Supplementary Data 5 from ETV4-Dependent Transcriptional Plasticity Maintains <i>MYC</i> Expression and Results in IMiD Resistance in Multiple Myeloma

ETV4 ChIP-seq binding sites in RPMI8226 cells.</p

Supplementary Data 2 from ETV4-Dependent Transcriptional Plasticity Maintains <i>MYC</i> Expression and Results in IMiD Resistance in Multiple Myeloma

Differentially expressed genes and GSEA in OPM2 cells following shRNA knockdown of IKZF3.</p