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Systematic Lupus Erythematosus (SLE) is an autoimmune syndrome of unclear etiology. While T and B cell abnormalities contribute to disease pathogenesis, recent work suggests that inflammatory M1-like macrophages also play a role. Previous work showed that the TLR2/1 agonist PAM3CSK4 (PAM3) could stimulate normal human monocytes to preferentially differentiate into immunosuppressive M2-like rather than inflammatory M1-like macrophages. This raised the possibility of PAM3 being used to normalize the M1:M2 ratio in SLE. Consistent with that possibility, monocytes from lupus patients differentiated into M2-like macrophages when treated with PAM3 in vitro. Furthermore, lupus-prone NZB x NZW F1 mice responded similarly to weekly PAM3 treatment. Normalization of the M2 macrophage frequency was associated with delayed disease progression, decreased autoantibody and inflammatory cytokine synthesis, reduced proteinuria and prolonged survival in NZB x NZW F1 mice. The ability of PAM3 to bias monocyte differentiation in favor of immunosuppressive macrophages may represent a novel approach to the therapy of SLE.

Systematic Lupus Erythematosus (SLE) is an autoimmune disease characterized by the overproduction of autoantibodies and the chronic inflammation of multiple organs. Early studies suggested that T and/or B cell abnormalities were critical to disease pathogenesis in lupus patients. More recent work shows that abnormalities in the innate immunity play a contributory if not central role. In that context, we find that SLE patients have significantly more CD14lowCD16+ monocytes compared to healthy controls.CD14lowCD16+ monocytes are a subset known as non-classical monocytes and display inflammatory characteristics upon activation. The frequency of inflammatory macrophages correlates with disease severity and is associated with reduced clearance of apoptotic cells and the overproduction of inflammatory cytokines. We previously found that the TLR2/1 agonist PAM3CSK4 (PAM3) can inhibit the generation of M1- while promoting the generation of immunosuppressive M2-like macrophages. This work examines whether PAM3 can shift the M1:M2 macrophage ratio to restore balance in lupus patients. Even though phenotypically, PAM3 did not seem to induce as many as CD163+ (a functional marker of M2) macrophages like M-CSF did, there was no significant difference in the endocytosis ability of the CD163+ macrophages induced by PAM3 compared M-CSF induced ones. The in vivo activity of PAM3 was investigated in the NZB/W murine model of lupus. Preliminary results indicate that, PAM3 injection increases the survival and decreases proteinuria in mice sera. These findings suggest that PAM3 may promote the polarization of SLE monocytes into immunosuppressive macrophages and thus could represent a novel approach to the therapy of this autoimmune disease.

Introduction TLR7/8 agonist (3M-052) and TLR9 agonist (CpG ODN) both trigger innate immune responses supporting the induction of tumor-specific immunity. Previous studies showed that this specific combination of TLR agonists synergistically promoted the elimination of large established tumors (>500 mm3). This effect was optimized when delivered directly into the tumor, leading to a conversion from immunosuppressive to inflammatory milieu. The current work explores the effect of a combination of a localized TLR agonists therapy and a systemic chemotherapy on lung metastases. Methods 66Cl4-luc cells (a luciferase-expressing variant of the 4T1 murine mammary carcinoma line) were orthotopically implanted into the fourth mammary fat pad of syngeneic Balb/c mice. The combination therapy was initiated when the primary tumor exceeded 300 mm3 and pulmonary metastases were present. Results Treatment delivered into both the primary tumor and the lungs, when associated with systemic chemotherapy significantly reduced tumor growth and prolonged survival. Discussion The combined effect of the two TLR agonists was further enhanced when associated with classic chemotherapy, providing a significant decrease of tumor growth at both sites. These results provide new leads to explore when considering lung metastases treatment.

Resolvin E1 (RvE1) is a potent proresolving mediator of inflammation derived from omega-3 eicosapentaenoic acid that acts locally to stop leukocyte recruitment and promote resolution. RvE1 displays potent counter-regulatory and tissue-protective actions in vitro and in vivo. Periodontal disease is a local inflammatory disease initiated by bacteria characterized by neutrophil-mediated tissue injury followed by development of a chronic immune lesion. In this study, we report the treatment of established periodontitis using RvE1 as a monotherapy in rabbits compared with structurally related lipids PGE2 and leukotriene B4. PGE2 and leukotriene B4 each enhanced development of periodontitis and worsened the severity of disease. Promotion of resolution of inflammation as a therapeutic target with RvE1 resulted in complete restoration of the local lesion, and reduction in the systemic inflammatory markers C-reactive protein and IL-1β. This report is the first to show that resolution of inflammation by a naturally occurring endogenous lipid mediator results in complete regeneration of pathologically lost tissues, including bone.

The TLR7/8 agonist 3M-052 and the TLR9 agonist CpG ODN both trigger innate immune responses that support the induction of tumor-specific immunity. Previous studies showed that intra-tumoral delivery of this agonist combination was synergistic and led to the eradication of large primary tumors and the establishment of long-term protective immunity in murine tumor models. We explored the effect of delivering 3M-052 plus CpG ODN to the lungs of mice bearing breast-derived lung metastases. 66Cl4-Luc cells (a luciferase-expressing variant of the 4T1 murine mammary carcinoma) were orthotopically implanted in the fourth mammary fat pad of Balb/c mice. Progression of the primary tumor and lung metastases was monitored by bioluminescence. TLR agonist treatment by intratracheal instillation was initiated after mets were detected in the lungs. Results show that intra-pulmonary treatment with the combination of TLR7/8 and TLR9 agonists slowed the growth of the lung metastases compared to either agonist alone. This treatment decreased pulmonary tumor burden and significantly prolonged survival when compared to control (untreated) animals. In contrast, pulmonary therapy had no significant effect on the primary tumor. Results from ongoing studies examining the effect of higher/more frequent lung delivery combined with direct treatment of the primary tumor will be presented.