Activin-A is a transforming growth factor-␤ (TGF-␤) superfamily member that plays a pivotal role in many developmental and reproductive processes. It is also involved in neuroprotection, apoptosis of tumor and some immune cells, wound healing, and cancer. Its role as an immuneregulating protein has not previously been described. Here we demonstrate for the first time that activin-A has potent autocrine effects on the capacity of human dendritic cells (DCs) to stimulate immune responses. Human monocyte-derived DCs IntroductionDendritic cells (DCs) form sentinel networks within the body sampling the microenvironment for pathogens, tissue injury, and inflammation via an array of pattern recognition receptors. [1][2][3] Pathogen encounter induces DC maturation, resulting in profound alterations in DC function. Antigen uptake is reduced, antigen processing is enhanced, and proinflammatory mediators are released. [4][5][6][7][8] The class, magnitude, and timing of cytokine or chemokine release are under exquisite control through both autocrine and paracrine signals as well as the signal strength and magnitude of the initiating stimulus. 8 DC cytokine and chemokine production can be induced by specific classes of stimuli. These include CD40 ligand (CD40L) and pathogen signals, such as toll-like receptor (TLR) agonists (eg, lipopolysaccharide [LPS] or intact bacteria). Appropriate release of cytokines, chemokines, and other soluble mediators by DCs and neighboring cells induces and moderates inflammation, recruits innate effectors, and regulates T-cell functions. [9][10][11][12] Many cytokines/chemokines produced by DCs at the epicenter of infection and inflammation, such as interleukin-6 (IL-6), 13,14 IL-8, 4,15,16 IL-10 17,18 as well as the potent T helper 1 (Th-1) cytokine, 19,20 have pleiotropic effects ranging from enhancing to inhibitory depending on the context and target cell type. However, uncontrolled cytokine/chemokine release within this microenvironment can also result in inappropriate T-and B-cell responses and subsequent immunopathology. [21][22][23] In this regard, the immune system has evolved to coordinately express mediators that attenuate exaggerated or inappropriate responses so as to minimize tissue damage and immunopathology (eg, prostaglandin E 2 (PGE 2 ), adenosine triphosphate (ATP), transforming growth factor-␤ [TGF-␤]). 24 Activin-A is a homodimer of activin-␤A subunits and was first described as a reproductive factor that accentuates the release of follicle-stimulating hormone. 25 It is a member of the TGF-␤ superfamily of cytokines and intimately shares with TGF-␤ the Smad intracellular signaling proteins. 26 The signaling, however, occurs through separate and distinct serine threonine kinase receptor subunits, and its release into the circulation during acute systemic inflammation differs from TGF-␤. 27 Activin-A signals through heteromeric receptor complexes consisting of both type I (ALK 2, 4, or 7) and type II (ActRIIA and ActRIIB) receptors. In addition, it is known to be pivotal in ...

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The ability of dendritic cells (DCs) to cross-present protein tumor antigens to cytotoxic T lymphocytes (CTLs) underpins the success of therapeutic cancer vaccines. We studied cross-presentation of the cancer/testis antigen, NY-ESO-1, and the melanoma differentiation antigen, Melan-A by human DC subsets. Monocyte-derived DCs (MoDCs) efficiently cross-presented human leukocyte associated (HLA)–A2-restricted epitopes from either a formulated NY-ESO-1/ISCOMATRIX vaccine or when either antigen was mixed with ISCOMATRIX adjuvant. HLA-A2 epitope generation required endosomal acidification and was proteasome-independent for NY-ESO-1 and proteasome-dependent for Melan-A. Both MoDCs and CD1c+ blood DCs cross-presented NY-ESO-1–specific HLA-A2157-165–, HLA-B760-72–, and HLA-Cw392-100–restricted epitopes when formulated as an NY-ESO-1/ISCOMATRIX vaccine, but this was limited when NY-ESO-1 and ISCOMATRIX adjuvant were added separately to the DC cultures. Finally, cross-presentation of NY-ESO-1157-165/HLA-A2, NY-ESO-160-72/HLA-B7, and NY-ESO-192-100/HLA-Cw3 epitopes was proteasome-dependent when formulated as immune complexes (ICs) but only proteasome-dependent for NY-ESO-160-72/HLA-B7–restricted cross-presentation facilitated by ISCOMATRIX adjuvant. We demonstrate, for the first time, proteasome-dependent and independent cross-presentation of HLA-A–, B–, and C–restricted epitopes within the same full-length tumor antigen by human DCs. Our findings identify important differences in the capacities of human DC subsets to cross-present clinically relevant, full-length tumor antigens and how vaccine formulation impacts CTL responses in vivo.

Natural killer (NK) cells are critical for innate tumor immunity due to their specialized ability to recognize and kill neoplastically transformed cells. However, NK cells require a specific set of cytokine-mediated signals to achieve optimal effector function. Th1-associated cytokines promote effector functions that are inhibited by the prototypic Th2 cytokine IL4 and the TGFb superfamily members TGFb1 and activin-A. Interestingly, the largest subgroup of the TGFb superfamily are the bone morphogenetic proteins (BMP), but the effects of BMP signaling on NK cell effector functions have not been evaluated. Here, we demonstrate that blood-circulating NK cells express type I and II BMP receptors, BMP-2 and BMP-6 ligands, and phosphorylated isoforms of Smad-1/-5/-8, which mediate BMP family member signaling. In opposition to the inhibitory effects of TGFb1 or activin-A, autocrine BMP signaling was supportive to NK cell function. Mechanistic investigations in cytokine and TLR-L-activated NK cells revealed that BMP signaling optimized IFNg and global cytokine and chemokine production, phenotypic activation and proliferation, and autologous dendritic cell activation and target cytotoxicity. Collectively, our findings identify a novel auto-activatory pathway that is essential for optimal NK cell effector function, one that might be therapeutically manipulated to help eradicate tumors. Cancer Res; 74(18); 5019-31. Ó2014 AACR.

<p>Supplemental Figure S3. Effects of dorsomorphin on NK cells phenotype and functionality.</p>

<p>Supplemental Figure S5. Effects of DMH1 on NK cells IFN-y production and killing capacity upon target cell engagement.</p>

<p>Supplemental Figure S4. Effects of DMH1 on NK cells IFN-y production and proliferation.</p>

<p>Supplemental Figure S3. Effects of dorsomorphin on NK cells phenotype and functionality.</p>