Eradicating malignant tumors by vaccine-elicited host immunity remains a major medical challenge. To date, correlates of immune protection remain unknown for malignant mesothelioma. In this study, we demonstrated that antigen-specific CD8+ T-cell immune response correlates with the elimination of malignant mesothelioma by a model PD-1–based DNA vaccine. Unlike the nonprotective tumor antigen WT1-based DNA vaccines, the model vaccine showed complete and long-lasting protection against lethal mesothelioma challenge in immunocompetent BALB/c mice. Furthermore, it remained highly immunogenic in tumor-bearing animals and led to therapeutic cure of preexisting mesothelioma. T-cell depletion and adoptive transfer experiments revealed that vaccine-elicited CD8+ T cells conferred to the protective efficacy in a dose-dependent way. Also, these CD8+ T cells functioned by releasing inflammatory IFNγ and TNFα in the vicinity of target cells as well as by initiating TRAIL-directed tumor cell apoptosis. Importantly, repeated DNA vaccinations, a major advantage over live-vectored vaccines with issues of preexisting immunity, achieve an active functional state, not only preventing the rise of exhausted PD-1+ and Tim-3+ CD8+ T cells but also suppressing tumor-induced myeloid-derived suppressive cells and Treg cells, with the frequency of antigen-specific CD8+ T cells inversely correlating with tumor mass. Our results provide new insights into quantitative and qualitative requirements of vaccine-elicited functional CD8+ T cells in cancer prevention and immunotherapy. Cancer Res; 74(21); 1–12. ©2014 AACR.

Eradicating malignant tumor by vaccine-elicited host immunity remains a major medical challenge. To date, correlates of immune protection remain unknown for malignant mesothelioma. In this study, we demonstrated that antigen-specific CD8+ T cell immune response correlates with the elimination of malignant mesothelioma by a model PD-1-based DNA vaccine. Unlike the non-protective tumor antigen WT1-based DNA vaccines, the model vaccine showed complete and long-lasting protection against lethal mesothelioma challenge in immunocompetent BALB/c mice. Furthermore, it remained highly immunogenic in tumor-bearing animals and led to therapeutic cure of pre-existing mesothelioma. T cell depletion and adoptive transfer experiments revealed that vaccine-elicited CD8+ T cells conferred to the protective efficacy in a dose-dependent way. Also, these CD8+ T cells functioned by releasing inflammatory IFN-γ and TNF-α in the vicinity of target cells as well as by initiating TRAIL-directed tumor cell apoptosis. Importantly, repeated DNA vaccinations, a major advantage over live-vectored vaccines with issues of pre-existing immunity, achieve an active functional state not only preventing the rise of exhausted PD-1+ and Tim-3+ CD8+ T cells but also suppressing tumor-induced MDSCs and Treg cells, with the frequency of antigen-specific CD8+ T cells inversely correlating with tumor mass. Our results provide new insights into quantitative and qualitative requirements of vaccine-elicited functional CD8+ T cells in cancer prevention and immunotherapy.

Mutations in the epidermal growth factor receptor (EGFR) are the most common actionable genetic abnormalities yet discovered in lung cancer. However, targeting these mutations with kinase inhibitors is not curative in advanced disease and has yet to demonstrate an impact on potentially curable, early-stage disease, with some data suggesting adverse outcomes. Here, we report that treatment of EGFR-mutated lung cancer cell lines with erlotinib, while showing robust cell death, enriches the ALDH+ stem-like cells through EGFR-dependent activation of Notch3. In addition, we demonstrate that erlotinib treatment increases the clonogenicity of lung cancer cells in a sphere-forming assay, suggesting increased stem-like cell potential. We demonstrate that inhibition of EGFR kinase activity leads to activation of Notch transcriptional targets in a γ secretase inhibitor-sensitive manner and causes Notch activation, leading to an increase in ALDH high+ cells. We also find a kinase-dependent physical association between the Notch3 and EGFR receptors and tyrosine phosphorylation of Notch3. This could explain the worsened survival observed in some studies of erlotinib treatment at early-stage disease, and suggests that specific dual targeting might overcome this adverse effect. Cancer Res; 74(19); 1–13. ©2014 AACR.

Although it is now widely appreciated that antitumor immunity is critical to impede tumor growth and progression, there remain significant gaps in knowledge about the mechanisms used by tumors to escape immune control. In tumor cells, we hypothesized that one mechanism of immune escape used by tumors involves the synthesis and extracellular shedding of gangliosides, a class of biologically active cell surface glycosphingolipids with known immunosuppressive properties. In this study, we report that tumor cells engineered to be ganglioside deficient exhibit impaired tumorigenicity, supporting a link between ganglioside-dependent immune escape and tumor outgrowth. Notably, we documented a dramatic reduction in the numbers and function of tumor-infiltrating myeloid-derived suppressor cells (MDSC) in ganglioside-deficient tumors, in contrast with the large MDSC infiltrates seen in ganglioside-rich littermate control tumors. Transient ganglioside reconstitution of the tumor cell inoculum was sufficient to increase MDSC infiltration, supporting a direct connection between ganglioside production by tumor cells and the recruitment of immunosuppressive MDSC into the tumor microenvironment. Our results reveal a novel mechanism of immune escape that supports tumor growth, with broad implications given that many human tumors produce and shed high levels of gangliosides. Cancer Res; 74(19); 1–9. ©2014 AACR.

SWI/SNF chromatin remodeling complexes regulate critical cellular processes, including cell-cycle control, programmed cell death, differentiation, genomic instability, and DNA repair. Inactivation of this class of chromatin remodeling complex has been associated with a variety of malignancies, including lung, ovarian, renal, liver, and pediatric cancers. In particular, approximately 10% of primary human lung non–small cell lung cancers (NSCLC) display attenuations in the BRG1 ATPase, a core factor in SWI/SNF complexes. To evaluate the role of BRG1 attenuation in NSCLC development, we examined the effect of BRG1 silencing in primary and established human NSCLC cells. BRG1 loss altered cellular morphology and increased tumorigenic potential. Gene expression analyses showed reduced expression of genes known to be associated with progression of human NSCLC. We demonstrated that BRG1 losses in NSCLC cells were associated with variations in chromatin structure, including differences in nucleosome positioning and occupancy surrounding transcriptional start sites of disease-relevant genes. Our results offer direct evidence that BRG1 attenuation contributes to NSCLC aggressiveness by altering nucleosome positioning at a wide range of genes, including key cancer-associated genes. Cancer Res; 74(22); 1–13. ©2014 AACR.