Single-agent immunotherapy has achieved limited clinical benefit to date in patients suffering from pancreatic ductal adenocarcinoma (PDAC). This may be due to the presence of a uniquely immunosuppressive tumor microenvironment (TME). Critical obstacles to immunotherapy in PDAC tumors include a high number of tumor-associated immunosuppressive cells and a uniquely desmoplastic stroma that acts as a barrier to T-cell infiltration. We have identified hyperactivated focal adhesion kinase (FAK) activity in neoplastic PDAC cells as a significant regulator of the fibrotic and immunosuppressive TME. We found that FAK activity was elevated in human PDAC tissues and correlates with high levels of fibrosis and poor CD8+ cytotoxic T-cell infiltration. Single-agent FAK inhibition using the selective FAK inhibitor VS-4718 significantly limited tumor progression, resulting in a doubling of survival in the p48-Cre/LSL-KrasG12D/p53Flox/+ (KPC) mouse model of human PDAC. This delay in tumor progression was associated with dramatically reduced tumor fibrosis, and decreased numbers of tumor-infiltrating immunosuppressive cells. We also found that FAK inhibition rendered the previously unresponsive KPC mouse model responsive to T cell immunotherapy and PD-1 antagonists. These data suggest that FAK inhibition increases immune surveillance by overcoming the fibrotic and immunosuppressive PDAC TME and renders tumors responsive to immunotherapy.

SUMMARY Tumor-associated macrophages (TAMs) are essential components of the cancer microenvironment and play critical roles in the regulation of tumor progression. Optimal therapeutic intervention requires in-depth understanding of the sources that sustain macrophages in malignant tissues. In this study, we investigated the ontogeny of TAMs in murine pancreatic ductal adenocarcinoma (PDAC) models. We identified both inflammatory monocytes and tissue-resident macrophages as sources of TAMs. Unexpectedly, significant portions of pancreas-resident macrophages originated from embryonic development and expanded through in situ proliferation during tumor progression. Whereas monocyte-derived TAMs played more potent roles in antigen presentation, embryonically derived TAMs exhibited a pro-fibrotic transcriptional profile, indicative of their role in producing and remodeling extracellular matrix molecules. Collectively, these findings uncover the heterogeneity of TAM origin and functions, and could provide therapeutic insight for PDAC treatment.

SUMMARY The mammalian immune system discriminates between modes of cell death, with necrosis often resulting in inflammation and adaptive immunity, while apoptosis tends to be anti-inflammatory, promoting immune tolerance. In many systems immune tolerance can be established through cross presentation of antigens derived from apoptotic cells via the MHC class I pathway to CD8+ T cells. We have examined the features of apoptosis responsible for tolerance to cell-mediated immune responses in vivo, specifically the roles of caspases and the mitochondria. Our results show that caspase activation targets the mitochondria to produce ROS, which are critical to tolerance induction by apoptotic cells. ROS oxidizes the potential danger signal HMGB1 released from dying cells, thereby neutralizing its stimulatory activity and promoting tolerance. Apoptotic cells failed to induce tolerance and instead stimulated immune responses when caspase-dependent ROS activity was prohibited by scavenging or by mutation of a mitochondrial caspase target, p75 NDUSF1. Similarly blocking sites of oxidation in HMGB1 prevented tolerance induction by apoptotic cells. These results suggest that caspase orchestrated mitochondrial events determine the impact of apoptotic cells on the immune response.

We examined the relationship between cell death and tolerance induction following antigen injection into the anterior chamber of the eye. Our data show that when inflammatory cells undergo apoptosis following infection with HSV-1, tolerance to the virus was observed. In contrast, when cell death was absent due to defects in Fas or FasL, immune tolerance was not observed. Further studies revealed that cell death and tolerance required that the lymphoid cells be Fas+ and the eye be FasL+. Additionally, we show that while Fas/FasL-mediated apoptosis occurred in the eye, it was apoptotic cell death that was critical for tolerance induction. Our results further demonstrate immune privilege is not a passive process involving physical barriers, but is an active process that employs an important natural mechanism to induce cell death and immune tolerance.

Crucial transitions in cancer-including tumor initiation, local expansion, metastasis, and therapeutic resistance-involve complex interactions between cells within the dynamic tumor ecosystem. Transformative single-cell genomics technologies and spatial multiplex in situ methods now provide an opportunity to interrogate this complexity at unprecedented resolution. The Human Tumor Atlas Network (HTAN), part of the National Cancer Institute (NCI) Cancer Moonshot Initiative, will establish a clinical, experimental, computational, and organizational framework to generate informative and accessible three-dimensional atlases of cancer transitions for a diverse set of tumor types. This effort complements both ongoing efforts to map healthy organs and previous largescale cancer genomics approaches focused on bulk sequencing at a single point in time. Generating single-cell, multiparametric, longitudinal atlases and integrating them with clinical outcomes should help identify novel predictive biomarkers and features as well as therapeutically relevant cell types, cell states, and cellular interactions across transitions. The resulting tumor atlases should have a profound impact on our understanding of cancer biology and have the potential to improve cancer detection, prevention, and therapeutic discovery for better precision-medicine treatments of cancer patients and those at risk for cancer.Cancer forms and progresses through a series of critical transitions-from pre-malignant to malignant states, from locally contained to metastatic disease, and from treatment-responsive to treatment-resistant tumors (Figure 1). Although specifics differ across tumor types and patients, all transitions involve complex dynamic interactions between diverse pre-malignant, malignant, and non-malignant cells (e.g., stroma cells and immune cells), often organized in specific patterns within the tumor