Previous studies have demonstrated that the cell surface receptor Slamf1 (CD150) is requisite for optimal NADPH-oxidase (Nox2) dependent reactive oxygen species (ROS) production by phagocytes in response to Gram- bacteria. By contrast, Slamf8 (CD353) is a negative regulator of ROS in response to Gram+ and Gram- bacteria. Employing in vivo migration after skin sensitization, induction of peritonitis, and repopulation of the small intestine demonstrates that in vivo migration of Slamf1-/- dendritic cells and macrophages is reduced, as compared to wt mice. By contrast, in vivo migration of Slamf8-/- dendritic cells, macrophages and neutrophils is accelerated. These opposing effects of Slamf1 and Slamf8 are cell-intrinsic as judged by in vitro migration in transwell chambers in response to CCL19, CCL21 or CSF-1. Importantly, inhibiting ROS production of Slamf8-/- macrophages by diphenyleneiodonium chloride blocks this in vitro migration. We conclude that Slamf1 and Slamf8 govern ROS–dependent innate immune responses of myeloid cells, thus modulating migration of these cells during inflammation in an opposing manner.

Type 1 diabetes (T1D) is characterized by pancreatic islet infiltration by autoreactive immune cells and a near-total loss of β-cells1. Restoration of insulin-producing β-cells coupled with immunomodulation to suppress the autoimmune attack has emerged as a potential approach to counter T1D2–4. Here we report that enhancing β-cell mass early in life, in two models of female NOD mice, results in immunomodulation of T-cells, reduced islet infiltration and lower β-cell apoptosis, that together protect them from developing T1D. The animals displayed altered β-cell antigens, and islet transplantation studies showed prolonged graft survival in the NOD-LIRKO model. Adoptive transfer of splenocytes from the NOD-LIRKOs prevented development of diabetes in pre-diabetic NOD mice. A significant increase in the splenic CD4+CD25+FoxP3+ regulatory T-cell (Treg) population was observed to underlie the protected phenotype since Treg depletion rendered NOD-LIRKO mice diabetic. The increase in Tregs coupled with activation of TGF-β/SMAD3 signaling pathway in pathogenic T-cells favored reduced ability to kill β-cells. These data support a previously unidentified observation that initiating β-cell proliferation, alone, prior to islet infiltration by immune cells alters the identity of β-cells, decreases pathologic self-reactivity of effector cells and increases Tregs to prevent progression of T1D.

Whereas the SLAMF-associated protein (SAP) is involved in differentiation of T follicular helper (Tfh) cells and antibody responses, the precise requirements of SLAMF receptors in humoral immune responses are incompletely understood. By analyzing mice with targeted disruptions of the Slamf1, Slamf5, and Slamf6 genes, we found that both T-dependent and T-independent antibody responses were twofold higher compared to those in single knockout mice. These data suggest a suppressive synergy of SLAMF1, SLAMF5, and SLAMF6 in humoral immunity, which contrasts the decreased antibody responses resulting from a defective GC reaction in the absence of the adapter SAP. In adoptive co-transfer assays, both [Slamf1 + 5 + 6]−− B and T cells were capable of inducing enhanced antibody responses, but more pronounced enhancement was observed after adoptive transfer of [Slamf1 + 5 + 6]−− B cells compared to that of [Slamf1 + 5 + 6]−− T cells. In support of [Slamf1 + 5 + 6]−− B cell intrinsic activity, [Slamf1 + 5 + 6]−− mice also mounted significantly higher antibody responses to T-independent type 2 antigen. Furthermore, treatment of mice with anti-SLAMF6 monoclonal antibody results in severe inhibition of the development of Tfh cells and GC B cells, confirming a suppressive effect of SLAMF6. Taken together, these results establish SLAMF1, SLAMF5, and SLAMF6 as important negative regulators of humoral immune response, consistent with the notion that SLAM family receptors have dual functions in immune responses.

Signaling lymphocytic activation molecule (SLAM)-associated protein (SAP) plays an essential role in the immune system mediating the function of several members of the SLAM family (SLAMF) of receptors, whose expression is essential for T, NK, and B-cell responses. Additionally, the expression of SAP in double-positive thymocytes is mandatory for natural killer T (NKT) cells and, in mouse, for innate CD8+ T cell development. To date, only two members of the SLAMF of receptors, Slamf1 and Slamf6, have been shown to positively cooperate during NKT cell differentiation in mouse. However, it is less clear whether other members of this family may also participate in the development of these innate T cells. Here, we show that Slamf[1 + 6]−/− and Slamf[1 + 5 + 6]−/−B6 mice have ~70% reduction of NKT cells compared to wild-type B6 mice. Unexpectedly, the proportion of innate CD8+ T cells slightly increased in the Slamf[1 + 5 + 6]−/−, but not in the Slamf[1 + 6]−/− strain, suggesting that Slamf5 may function as a negative regulator of innate CD8+ T cell development. Accordingly, Slamf5−/− B6 mice showed an exclusive expansion of innate CD8+ T cells, but not NKT cells. Interestingly, the SAP-independent Slamf7−/− strain showed an expansion of both splenic innate CD8+ T cells and thymic NKT cells. On the other hand, and similar to what was recently shown in Slamf3−/− BALB/c mice, the proportions of thymic promyelocytic leukemia zinc finger (PLZFhi) NKT cells and innate CD8+ T cells significantly increased in the SAP-independent Slamf8−/− BALB/c strain. In summary, these results show that NKT and innate CD8+ T cell development can be regulated in a SAP-dependent and -independent fashion by SLAMF receptors, in which Slamf1, Slamf6, and Slamf8 affect development of NKT cells, and that Slamf5, Slamf7, and Slamf8 affect the development of innate CD8+ T cells.

The tumor microenvironment in leukemia and solid tumors induces a shift of activated CD8 + cytotoxic T cells to an exhausted state, characterized by loss of proliferative capacity and impaired immunologic synapse formation. Efficient strategies and targets need to be identified to overcome T-cell exhaustion and further improve overall responses in the clinic. Here, we took advantage of the E-TCL1 chronic lymphocytic leukemia (CLL) and B16 melanoma mouse models to assess the role of the homophilic cell surface receptor SLAMF6 as an immune checkpoint regulator. The transfer of SLAMF6 + E-TCL1 cells into SLAMF6-/recipients, in contrast to wild-type (WT) recipients, significantly induced expansion of a PD-1 + subpopulation among CD3 + CD44 + CD8 + T cells, which had impaired cytotoxic functions. Conversely, administering anti-SLAMF6 significantly reduced the leukemic burden in E-TCL1 recipient WT mice concomitantly with a loss of PD-1 + CD3 + CD44 + CD8 + T cells with significantly increased effector functions. Anti-SLAMF6 significantly reduced leukemic burden in the peritoneal cavity, a niche where antibody-dependent cellular cytotoxicity (ADCC) is impaired, possibly through activation of CD8 + T cells. Targeting of SLAMF6 not only impacted tumor growth in B cell-related leukemia and lymphomas but also nonhematopoietic tumors like B16 melanoma, where SLAMF6 is not expressed. In vitro exhausted CD8 + T cells showed increased degranulation when anti-human SLAMF6 was added in culture. Taken together, anti-SLAMF6 both effectively corrected CD8 + Tcell dysfunction and had a direct effect on tumor progression. The outcomes of our studies suggest that targeting SLAMF6 is a potential therapeutic strategy.

The signaling lymphocyte activation molecule family [SLAMF] of cell surface receptors partakes in both the development of several immunocyte lineages and innate and adaptive immune responses in humans and mice. For instance, the homophilic molecule SLAMF6 (CD352) is in part involved in natural killer T cell development, but also modulates T follicular helper cell and germinal B cell interactions. Here we report that upon transplantation of a well-defined aggressive murine B220+CD5+ Chronic Lymphocytic Leukemia (CLL) cell clone, TCL1-192, into SCID mice one injection of a monoclonal antibody directed against SLAMF6 (αSlamf6) abrogates tumor progression in the spleen, bone marrow and blood. Similarly, progression of a murine B cell lymphoma, LMP2A/λMyc, was also eliminated by αSlamf6. But, surprisingly, αSLAMF6 neither eliminated TCL1-192 nor LMP2A/λMyc cells, which resided in the peritoneal cavity or omentum. This appeared to be dependent upon the tumor environment, which affected the frequency of sub-populations of the TCL1-192 clone or the inability of peritoneal macrophages to induce Antibody Dependent Cellular Cytotoxicity (ADCC). However, co-administering αSlamf6 with the Bruton tyrosine kinase (Btk) inhibitor, ibrutinib, synergized to efficiently eliminate the tumor cells in the spleen, bone marrow, liver and the peritoneal cavity. Because an anti-human SLAMF6 mAb efficiently killed human CLL cells in vitro and in vivo, we propose that a combination of αSlamf6 with ibrutinib should be considered as a novel therapeutic approach for CLL and other B cell tumors.

The nine SLAM family (Slamf) receptors are positive or negative regulators of adaptive and innate immune responses, and of several autoimmune diseases. Here we report that the transfer of Slamf6−/− B6 CD4+ T cells into co-isogenic bm12 mice causes SLE-like autoimmunity with elevated levels of autoantibodies. In addition, significantly higher percentages of Tfh cells and IFN-γ-producing CD4+ cells, as well as GC B cells were observed. Interestingly, the expression of the Slamf6-H1 isoform in Slamf6−/− CD4+ T cells did not induce this lupus-like phenotype. By contrast, Slamf1−/− or Slamf5−/− CD4+ T cells caused the same pathology as WT CD4+ T cells. As the transfer of Slamf [1+6]−/− or Slamf [1+5+6]−/− CD4+ T cells induced WT levels of autoantibodies, the presence of Slamf1 was requisite for the induction of increased levels of autoantibodies by Slamf6−/− CD4+ T cells. We conclude that Slamf6 functions as an inhibitory receptor that controls autoimmune responses.

Effective immune responses develop after a well-orchestrated series of events that include recognition, immune cell interactions and activation/inhibition of signaling pathways. The signaling lymphocyte activation molecule family (SLAMF) of cell surface receptors, which consists of nine transmembrane proteins (SLAMF1-9) expressed at different levels, are involved in viral and bacterial recognition, serve as co-stimulatory molecules at immune synapses, and modulate myeloid and lymphocyte development. SLAMF receptors are homophilic receptors, with the exception of SLAMF2 and SLAMF4, and are only expressed on hematopoietic cells. Their adaptors, SLAM associated protein (SAP) and Ewing's sarcoma-associated transcript 2 (EAT-2), bind to the cytoplasmic tails and control the functions and magnitude of SLAMF receptor signaling. In this review, we focus on the current knowledge on the role of SLAMF6 receptor in regulating immune functions and recent findings describing how SLAMF6 can be exploited as a target in human malignancies. The SLAM family of immune cell surface receptors is a member of the CD2 subfamily of the immunoglobulin (Ig) superfamily consisting of nine members, SLAMF1-9 [1-4]. SLAMF receptors are type I transmembrane glycoproteins comprised of an extracellular membrane containing an N-terminal V-Ig domain followed by a C2-Ig domain in the extracellular region (this set is duplicated in SLAMF3), a transmembrane region, and an intracellular cytoplasmic tail containing tyrosine based switch motifs (ITSM). Notable exceptions to this structure include SLAMF2, which has a glycosyl-phosphatidyl-inositol (GPI) membrane anchor and like SLAMF8 and SLAMF9 lack ITSM motifs [5-8]. Binding of SLAM associated adaptors; SAP and EAT-2, to cytoplasmic tails of various SLAMFs regulate their function on different immune cells. Expression of SLAMFs and their adaptors is restricted to hematopoietic cells. In addition, the gene loci are located on chromosome 1 in both mice and humans, except SAP, which is located on the X chromosome [9](Figure 1). All SLAMFs are homophilic receptors aside from SLAMF2 and SLAMF4, which bind each other [10-12]. The determination of the SLAMF3, SLAMF5 and SLAMF6 crystal structures revealed in-trans interactions through their IgV domains (SLAMF3 unpublished data, generously donated by Profs. Steve Almo and Stanley Nathenson, Albert Einstein College of Medicine (Figure 2) [13, 14]. Engagement of SLAMF ACCEPTED MANUSCRIPT A C C E P T E D M A N U S C R I P T receptors on immune cells (e.g. APC -T cell) trigger inhibitory or activating signals that modulate cellular responses. Within these homophilic and heterophilic interactions, the binding affinities for each SLAMF varies (SLAMF3 nM, SLAMF5 sub-M, SLAMF6 ~ 2M, SLAMF2/4 ~4M, SLAMF1 ~200 M) which likely contributes to functional differences within the family of receptors [12-15]. In addition to being self-ligands, SLAMF1 also serves as an entry receptor for Measles virus [16, 17] while SLAMF1, SLAMF2 and SLAMF6 have been demonstrated to interact...