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Activating RAS missense mutations are among the most prevalent genomic alterations observed in human cancers and drive oncogenesis in the three most lethal tumor types. Emerging evidence suggests mutant KRAS (mKRAS) may be targeted immunologically, but mKRAS epitopes remain poorly defined. Here we employ a multi-omics approach to characterize HLA class I-restricted mKRAS epitopes. We provide proteomic evidence of mKRAS epitope processing and presentation by high prevalence HLA class I alleles. Select epitopes are immunogenic enabling mKRAS-specific TCRαβ isolation. TCR transfer to primary CD8+ T cells confers cytotoxicity against mKRAS tumor cell lines independent of histologic origin, and the kinetics of lytic activity correlates with mKRAS peptide-HLA class I complex abundance. Adoptive transfer of mKRAS-TCR engineered CD8+ T cells leads to tumor eradication in a xenograft model of metastatic lung cancer. This study validates mKRAS peptides as bona fide epitopes facilitating the development of immune therapies targeting this oncoprotein. KRAS is commonly mutated at codon 12 in several cancer types, offering a unique opportunity for the development of neoantigen-targeted immunotherapy. Here the authors present a pipeline for the prediction, identification and validation of HLA class-I restricted mutant KRAS G12 peptides, leading to the generation of mutant KRAS-specific T cell receptors for adoptive T cell immunotherapy. [ABSTRACT FROM AUTHOR]

Neovascularization and inflammation are independent biological processes but are linked in response to injury. The role of inflammation-dampening cytokines in the regulation of angiogenesis remains to be clarified. The purpose of this work was to test the hypothesis that IL-19 can induce angiogenesis in the absence of tissue hypoxia and to identify potential mechanisms. Using the aortic ring model of angiogenesis, we found significantly reduced sprouting capacity in aortic rings from IL-19−/− compared with wild-type mice. Using an in vivo assay, we found that IL-19−/− mice respond to vascular endothelial growth factor (VEGF) significantly less than wild-type mice and demonstrate decreased capillary formation in Matrigel plugs. IL-19 signals through the IL-20 receptor complex, and IL-19 induces IL-20 receptor subunit expression in aortic rings and cultured human vascular smooth muscle cells, but not endothelial cells, in a peroxisome proliferator-activated receptor-γ-dependent mechanism. IL-19 activates STAT3, and IL-19 angiogenic activity in aortic rings is STAT3-dependent. Using a quantitative RT-PCR screening assay, we determined that IL-19 has direct proangiogenic effects on aortic rings by inducing angiogenic gene expression. M2 macrophages participate in angiogenesis, and IL-19 has indirect angiogenic effects, as IL-19-stimulated bone marrow-derived macrophages secrete proangiogenic factors that induce greater sprouting of aortic rings than unstimulated controls. Using a quantitative RT-PCR screen, we determined that IL-19 induces expression of angiogenic cytokines in bone marrow-derived macrophages. Together, these data suggest that IL-19 can promote angiogenesis in the absence of hypoxia by at least two distinct mechanisms: 1) direct effects on vascular cells and 2) indirect effects by stimulation of macrophages. [ABSTRACT FROM AUTHOR]

Although it is evident that there is complex interplay among genetic and environmental factors contributing to systemic autoimmunity, the events inciting autoreactivity are incompletely understood. Previously we demonstrated that MRL-MpJ mice posses a genetic background susceptible to autoimmunity development under conditions of altered inhibitory signaling. To gain better understanding of the influence of exogenous factors on autoreactivity in susceptible individuals, young MRL-MpJ mice were challenged with a single injection of heterologous protein and evaluated for evidence of autoimmunity. We found that MRL-MpJ mice developed high titer serum reactivity to DNA within 1 week of protein administration reaching maximal levels within 1 month. Importantly, the level of autoimmunity was sustained for an extended period of time (6 months). This was accompanied by a substantial increase in germinal center B cell and plasma cell numbers. In contrast, control mice showed no change in autoreactivity or lymphocyte homeostasis. Autoimmunity was dependent on marginal zone B cells as their depletion reduced serum auto-reactivity after challenge, thus suggesting immune stimulation with heterologous proteins can precipitate loss of B cell tolerance and autoimmunity in genetically prone individuals. This model may provide an important tool to further investigate the mechanisms whereby environmental stimuli trigger autoimmune reactivity in susceptible hosts. [ABSTRACT FROM AUTHOR]

Vascular smooth muscle cell (VSMC) migration is an important cellular event in multiple vascular diseases, including atherosclerosis, restenosis, and transplant vasculopathy. Little is known regarding the effects of anti-inflammatory interleukins on VSMC migration. This study tested the hypothesis that an anti-inflammatory Th2 interleukin, interleukin-19 (IL-19), could decrease VSMC motility. IL-19 significantly decreased platelet-derived growth factor (PDGF)-stimulated VSMC chemotaxis in Boyden chambers and migration in scratch wound assays. IL-19 significantly decreased VSMC spreading in response to PDGF. To determine the molecular mechanism(s) for these cellular effects, we examined the effect of IL-19 on activation of proteins that regulate VSMC cytoskeletal dynamics and locomotion. IL-19 decreased PDGF-driven activation of several cytoskeletal regulatory proteins that play an important role in smooth muscle cell motility, including heat shock protein-27 (HSP27), myosin light chain (MLC), and cofilin. IL-19 decreased PDGF activation of the Rac1 and RhoA GTPases, important integrators of migratory signals. IL-19 was unable to inhibit VSMC migration nor was able to inhibit activation of cytoskeletal regulatory proteins in VSMC transduced with a constitutively active Rac1 mutant (RacV14), suggesting that IL-19 inhibits events proximal to Rac1 activation. Together, these data are the first to indicate that IL-19 can have important inhibitory effects on VSMC motility and activation of cytoskeletal regulatory proteins. This has important implications for the use of anti-inflammatory cytokines in the treatment of vascular occlusive disease. [ABSTRACT FROM AUTHOR]