InVivoMAb anti-mouse CD4

Checkpoint blockade with antibodies specific for cytotoxic T lymphocyte-associated protein (CTLA)-4 or programmed cell death 1 (PDCD1; also known as PD-1) elicits durable tumor regression in metastatic cancer, but these dramatic responses are confined to a minority of patients. This suboptimal outcome is probably due in part to the complex network of immunosuppressive pathways present in advanced tumors, which are unlikely to be overcome by intervention at a single signaling checkpoint. Here we describe a combination immunotherapy that recruits a variety of innate and adaptive immune cells to eliminate large tumor burdens in syngeneic tumor models and a genetically engineered mouse model of melanoma; to our knowledge tumors of this size have not previously been curable by treatments relying on endogenous immunity. Maximal antitumor efficacy required four components: a tumor-antigen-targeting antibody, a recombinant interleukin-2 with an extended half-life, anti-PD-1 and a powerful T cell vaccine. Depletion experiments revealed that CD8+ T cells, cross-presenting dendritic cells and several other innate immune cell subsets were required for tumor regression. Effective treatment induced infiltration of immune cells and production of inflammatory cytokines in the tumor, enhanced antibody-mediated tumor antigen uptake and promoted antigen spreading. These results demonstrate the capacity of an elicited endogenous immune response to destroy large, established tumors and elucidate essential characteristics of combination immunotherapies that are capable of curing a majority of tumors in experimental settings typically viewed as intractable.

Using several tumor models, we demonstrate that mice deficient in Bcl11b in T cells, although having reduced numbers of T cells in the peripheral lymphoid organs, developed significantly less tumors compared with wild-type mice. Bcl11b(-/-) CD4(+) T cells, with elevated TNF-alpha levels, but not the Bcl11b(-/-) CD8(+) T cells, were required for the reduced tumor burden, as were NK1.1(+) cells, found in increased numbers in Bcl11b(F/F)/CD4-Cre mice. Among NK1.1(+) cells, the NK cell population was predominant in number and was the only population displaying elevated granzyme B levels and increased degranulation, although not increased proliferation. Although the number of myeloid-derived suppressor cells was increased in the lungs with metastatic tumors of Bcl11b(F/F)/CD4-Cre mice, their arginase-1 levels were severely reduced. The increase in NK cell and myeloid-derived suppressor cell numbers was associated with increased bone marrow and splenic hematopoiesis. Finally, the reduced tumor burden, increased numbers of NK cells in the lung, and increased hematopoiesis in Bcl11b(F/F)/CD4-Cre mice were all dependent on TNF-alpha. Moreover, TNF-alpha treatment of wild-type mice also reduced the tumor burden and increased hematopoiesis and the numbers and activity of NK cells in the lung. In vitro treatment with TNF-alpha of lineage-negative hematopoietic progenitors increased NK and myeloid differentiation, further supporting a role of TNF-alpha in promoting hematopoiesis. These studies reveal a novel role for TNF-alpha in the antitumor immune response, specifically in stimulating hematopoiesis and increasing the numbers and activity of NK cells.

The costimulatory B7-1 (CD80)/B7-2 (CD86) molecules, along with T-cell receptor stimulation, together facilitate T-cell activation. This explains why in vivo B7 costimulation neutralization efficiently silences a variety of human autoimmune disorders. Paradoxically, however, B7 blockade also potently moderates accumulation of immune-suppressive regulatory T cells (Tregs) essential for protection against multiorgan systemic autoimmunity. Here we show that B7 deprivation in mice overrides the necessity for Tregs in averting systemic autoimmunity and inflammation in extraintestinal tissues, whereas peripherally induced Tregs retained in the absence of B7 selectively mitigate intestinal inflammation caused by Th17 effector CD4(+) T cells. The need for additional immune suppression in the intestine reflects commensal microbe-driven T-cell activation through the accessory costimulation molecules ICOSL and OX40L. Eradication of commensal enteric bacteria mitigates intestinal inflammation and IL-17 production triggered by Treg depletion in B7-deficient mice, whereas re-establishing intestinal colonization with Candida albicans primes expansion of Th17 cells with commensal specificity. Thus, neutralizing B7 costimulation uncovers an essential role for Tregs in selectively averting intestinal inflammation by Th17 CD4(+) T cells with commensal microbe specificity.

The differentiation and protective capacity of Plasmodium-specific T cells are regulated by both positive and negative signals during malaria, but the molecular and cellular details remain poorly defined. Here we show that malaria patients and Plasmodium-infected rodents exhibit atypical expression of the co-stimulatory receptor OX40 on CD4 T cells and that therapeutic enhancement of OX40 signaling enhances helper CD4 T cell activity, humoral immunity, and parasite clearance in rodents. However, these beneficial effects of OX40 signaling are abrogated following coordinate blockade of PD-1 co-inhibitory pathways, which are also upregulated during malaria and associated with elevated parasitemia. Co-administration of biologics blocking PD-1 and promoting OX40 signaling induces excessive interferon-gamma that directly limits helper T cell-mediated support of humoral immunity and decreases parasite control. Our results show that targeting OX40 can enhance Plasmodium control and that crosstalk between co-inhibitory and co-stimulatory pathways in pathogen-specific CD4 T cells can impact pathogen clearance.