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The four core members of the Interleukin-12 (IL-12) family of cytokines, IL-12, IL-23, IL-27 and IL-35 are heterodimers which share α- and β-cytokine subunits. All four cytokines are immune modulators and have been proposed to play divergent roles in inflammatory arthritis. In recent years additional combinations of α- and β-cytokine subunits belonging to the IL-12 family have been proposed to form novel cytokines such as IL-39. However, the actual extent of the combinatorial potential of the cytokine subunits in the human IL-12 family is not known. Here, we identify several combinations of subunits that form secreted heterodimeric assemblies based on a systematic orthogonal approach. The heterodimers are detected in the conditioned media harvested from mammalian cell cultures transfected with unfused pairs of cytokine subunits. While certain previously reported subunit combinations could not be recapitulated, our approach showed robustly that all four of the canonical members could be secreted. Furthermore, we provide evidence for the interaction between Cytokine Receptor Like Factor 1 (CRLF1) and Interleukin-12 subunit alpha (p35). Similar to IL-27 and IL-35 this novel heterodimer is not abundantly secreted rendering isolation from the conditioned medium very challenging, unlike IL-12 and IL-23. Our findings set the stage for fine-tuning approaches towards the biochemical reconstitution of IL-12 family cytokines for biochemical, cellular, and structural studies., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Immunotherapy has shown promising treatment effects for a variety of cancers. However, the immune treatment efficiency for solid tumors is limited owing to insufficient infiltration of immune cells into solid tumors. The conversion of tumor-supportive macrophages to tumor-suppressive macrophages, inducing the functional reversal of macrophages, is an effective method and contributes to a subsequent antitumor response. The current challenge in the field is the poor distribution and systemic side effects associated with the use of cytokines. As a solution to this issue, we designed and synthesized microenvironment-responsive nanoparticles (P) with IL-12 payload (IL-12⊂P1). These nanoparticles could promote the systemic administration and release of IL-12 in the tumor microenvironment, and the locally responsive property of IL-12⊂P1 could subsequently re-educate tumor-associated macrophages (TAMs). In particular, our results illustrated the great therapeutic effects derived from the functional conversion of macrophages. Our strategy was to design a microenvironment-responsive material for local macrophage modification to overcome the physiological barrier of solid tumors. The shifting of macrophage phenotypes via IL-12⊂P1 achieved immunomodulation in the microenvironment for cancer therapy, with negligible cytotoxicity. We expect that the functional regulation of TAMs by pH-responsive nanomaterials is a promising therapeutic approach for cancer immunotherapy., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

Biomedical device-associated infection is one of the most common and problematic complications faced by millions of patients worldwide. The current antibiotic therapy strategies face challenges, the most serious of which is antibiotic resistance. Studies have shown that the systemic level of interleukin 12 (IL-12) decreases following major injuries resulting in decreased cell-mediated immune response. Here we report the development of IL-12 nanoscale coatings using electrostatic layer-by-layer self-assembly nanotechnology. We found that IL-12 nanoscale coatings at the implant/tissue interface substantially decrease infections in vivo, and IL-12 nanoscale coatings are advantageous over traditional treatments. This approach could be a revolutionary step toward preventing device-associated infections using a non-antibiotic approach.

The active molecule in Coley's Toxin is not tumor necrosis factor (TNF) or endotoxin (LPS), but interleukin-12 (IL-12). IL-12 holds the key to improved anti-tumor immuns response.

Toll-like receptors (TLR) are involved in innate immunity by recognizing various bacterial components. We have previously reported that an active substance of ARS-2 purified from the culture medium of Chlorella vulgaris was a glycoprotein with a molecular weight of 63,100 amu and that this glycoprotein expressed antitumor activity, with the protein moiety in ARS-2 being necessary for this antitumor activity. Here, we show that ARS-2 stimulated spleen-adherent cells from C3H/HeJ lacking functional TLR4 to produce interleukin-12 (IL-12) p40, whereas such cytokine production was significantly impaired in ARS-2-stimulated spleen-adherent cells from TLR2 knockout mice. The overexpression of mouse TLR2 (mTLR2) and mouse CD14 (mCD14) conferred the ARS-2 inducibility of nuclear factor-kappaB activation to human HEK 293 cells. These results suggest that TLR2 signaling is at least partly involved in the antitumor activity of the water-soluble antitumor glycoprotein from C. vulgaris.

Spirulina platensis is a cyanobacterial species that is surmised to potentiate the immune system leading to suppression of cancer development and viral infection. Here, we identified the molecular mechanism of the human immune potentiating capacity of Spirulina by analyzing blood cells of volunteers with pre and post oral administration of hot water extract of Spirulina. NK functions represented by IFN gamma production and cytolysis were enhanced after administration of Spirulina in >50% subjects. IFN gamma was produced in an IL-12/IL-18-dependent fashion. In vitro stimulation of blood cells with BCG cell wall skeleton (CWS) allowed more potent IL-12 p40 production in cells from volunteers given Spirulina than in cells without pre-exposure to Spirulina. As BCG-CWS serves as a ligand for Toll-like receptor (TLR) 2 and 4 to raise the maturation stage of monocytes/macrophages, Spirulina may be involved in the signaling responses through Toll in blood cells even when orally administered. These observations indicated that in humans Spirulina acts directly on myeloid lineages and either directly or indirectly on NK cells. The presence of co-operative IL-12 and IL-18 is critically important for NK-mediated IFN gamma production.

The feasibility of using oligodeoxynucleotides (ODN) containing unmethylated CpG motifs as parenteral adjuvants for subunit vaccines against RSV was tested in BALB/c mice. Compared with immunization with natural F protein adsorbed to aluminum hydroxide (F/AlOH) adjuvant alone, coadministration of F/AlOH with CpG ODN resulted in statistically significant increases in serum neutralization titers, an enhanced generation of splenic antigen-dependent killer cell precursors, and accelerated clearance of infectious virus from lungs 4 days after challenge. The statistically significant increases in serum IFNgamma and anti-F protein IgG2a titers, and significantly diminished pulmonary IL-5 and eosinophilia after challenge indicated that CpG ODN enhanced the ability of F/AlOH to elicit type 1 immune responses. F protein-specific serum IgE titers were also reduced. Further analysis of pulmonary inflammatory cells demonstrated an expansion of CD8(+) T cells, relative to the CD4(+) T cell compartment. The potency of CpG ODN was not adversely affected in gene knockout mice devoid of the p35 chain of the IL-12 heterodimer. Taken together, the results suggest a novel formulation for naïve recipients of F protein-based subunit vaccines that does not result in a type 2 phenotype.