SUMMARY Anti-HER2/neu antibody therapy is reported to mediate tumor regression by interrupting oncogenic signals and/or inducing FcR-mediated cytotoxicity. Here, we demonstrate that the mechanisms of tumor regression by this therapy also require the adaptive immune response. Activation of innate immunity and T cells, initiated by antibody treatment, was necessary. Intriguingly, the addition of chemotherapeutic drugs, while capable of enhancing the reduction of tumor burden, could abrogate antibody-initiated immunity leading to decreased resistance to re-challenge or earlier relapse. Increased influx of both innate and adaptive immune cells into the tumor microenvironment by a selected immunotherapy further enhanced subsequent antibody-induced immunity, leading to increased tumor eradication and resistance to re-challenge. Therefore, this study proposes a model and strategy for anti-HER2/neu antibody-mediated tumor clearance.

Epidermal growth factor receptor (EGFR) over-signaling leads to more aggressive tumor growth. The antitumor effect of Cetuximab, an anti-EGFR antibody, depends on oncogenic-signal blockade leading to tumor cell apoptosis and antibody dependent cell-mediated cytotoxicity (ADCC). However, whether adaptive immunity plays a role in Cetuximab-mediated tumor inhibition is unclear, as current xenograft models lack adaptive immunity and human-EGFR-dependent mouse tumor cell lines are unavailable. Using a newly developed xenograft model with reconstituted immune cells, we demonstrate that the Cetuximab effect becomes more pronounced and reduces the EGFR(+) human tumor burden when adaptive immunity is present. To further study this in a mouse tumor model, we created a novel EGFR(+) mouse tumor cell line and demonstrated that Cetuximab-induced tumor regression depends on both innate and adaptive immunity components, including CD8(+) T cells, MyD88, and FcγR. To test whether strong innate signals inside tumor tissues amplifies the Cetuximab-mediated therapeutic effect, Cetuximab was conjugated to CpG. This conjugate is more potent than Cetuximab alone for complete tumor regression and resistance to tumor rechallenge. Furthermore, Cetuximab-CpG conjugates can activate tumor-reactive T cells for tumor regression by increasing dendritic cell (DC) cross-presentation. Therefore, this study establishes new models to evaluate immune responses induced by antibody-based treatment, defines molecular mechanisms, and provides new tumor-regression strategies.

Previous studies have shown a correlation between pre-transplant conditioning intensity, intestinal barrier loss, and graft-versus-host disease (GVHD) severity. However, since irradiation and other forms of pre-transplant conditioning have pleiotropic effects, the precise role of intestinal barrier loss in GVHD pathogenesis remains unclear. We developed GVHD models that allowed us to isolate the specific contributions of distinct pre-transplant variables. First, intestinal damage was required for the induction of minor mismatch (MHC-matched) GVHD, but was not necessary for major mismatch GVHD, demonstrating fundamental pathogenic distinctions between these forms of disease. Moreover, recipient NK cells prevented minor mismatch GVHD by limiting expansion and target organ infiltration of alloreactive T cells via a perforin-dependent mechanism, revealing a previously unrecognized immunoregulatory function of recipient NK cells in GVHD. Minor mismatch GVHD required MyD88-mediated TLR4 signaling on donor cells, and intestinal damage could be bypassed by parenteral LPS administration, indicating a critical role for the influx of bacterial components triggered by intestinal barrier loss. In all, the data demonstrate that pre-transplant conditioning plays a dual role in promoting minor mismatch GVHD by both depleting recipient NK cells and inducing intestinal barrier loss.

Purpose Targeting oncogenic receptors with antibodies has been thought to suppress tumor growth mainly by interrupting oncogenic signals. Recently, the essential role for adaptive immunity, and CD8+ T cells in particular, has been established as a major factor for anti-HER2/neu mediated tumor regression. However, the role of CD4+ T cells is still being defined. The purpose of this study was to explore whether and to what extent CD4+ T cells are involved in mediating the effects of anti-HER2/neu therapy. Experimental Design The role of CD4+ T cells was examined using a transplant model of the rat HER2/neu overexpressing cell line TUBO. Tumor bearing mice were treated with anti-neu therapy in conjunction with CD4 depletion or CD40L blockade. The effects of CD4 depletion on the anti-tumor response were examined by tumor growth analysis and ELISPOT. Results In addition to CD8+ T cells, CD4+ T cells are also essential for anti-neu antibody-mediated tumor regression, but B cells are not required. The role for CD4+ cells is necessary throughout anti-neu therapy and not limited to helping CD8+ T cells. Expression of IFNγ is necessary for anti-neu therapy and IFNγ induces MHC-II expression on TUBO cells promoting direct recognition by CD4+ T cells. Furthermore, intratumoral depletion of CD4+ T cells or blockade of the activating cell-surface protein CD40L inhibits the anti-tumor response. Conclusions This study reveals essential role of CD4+ T cell for anti-neu mediated tumor regression.