We have previously reported that adoptive transfer of tumor‐draining lymph node (TDLN) B cells confers tumor regression in a spontaneous pulmonary metastasis mouse model of breast cancer. In this study, we identified IL‐10‐producing cells within these B cells, and found that IL‐10 removal, either by using IL‐10^−/− TDLN B cells or by systemic neutralization of IL‐10, significantly augmented the therapeutic efficacy of adoptively transferred TDLN B cells. Depletion of IL‐10 in B‐cell adoptive transfers significantly increased CTLs and B‐cell activity of PBMCs and splenic cells in the recipient. Activated TDLN B cells express Fas ligand, which was further enhanced by coculture of these TDLN B cells with 4T1 tumor cells. Effector B cells killed tumor cells directly in vitro in an antigen specific and Fas ligand‐dependent manner. Trafficking of TDLN B cells in vivo suggested that they were recruited to the tumor and lung as well as secondary lymphoid organs. These findings further define the biological function of antitumor effector B cells, which may offer alternative cellular therapies to cancer.

Dendritic cell (DC)-based vaccine strategies aimed at targeting cancer stem–like cells (CSC) may be most efficacious if deployed in the adjuvant setting. In this study, we offer preclinical evidence that this is the case for a CSC-DC vaccine as tested in murine models of SCC7 squamous cell cancer and D5 melanoma. Vaccination of mice with an ALDHhigh SCC7 CSC-DC vaccine after surgical excision of established SCC7 tumors reduced local tumor relapse and prolonged host survival. This effect was augmented significantly by simultaneous administration of anti-PD-L1, an immune checkpoint inhibitor. In the minimal disease setting of D5 melanoma, treatment of mice with ALDHhigh CSC-DC vaccination inhibited primary tumor growth, reduced spontaneous lung metastases, and increased host survival. In this setting, CCR10 and its ligands were downregulated on ALDHhigh D5 CSCs and in lung tissues, respectively, after vaccination with ALDHhigh D5 CSC-DC. RNAi-mediated attenuation of CCR10 blocked tumor cell migration in vitro and metastasis in vivo. T cells harvested from mice vaccinated with ALDHhigh D5 CSC-DC selectively killed ALDHhigh D5 CSCs, with additional evidence of humoral immunologic engagement and a reduction in ALDHhigh cells in residual tumors. Overall, our results offered a preclinical proof of concept for the use of ALDHhigh CSC-DC vaccines in the adjuvant setting to more effectively limit local tumor recurrence and spontaneous pulmonary metastasis, as compared with traditional DC vaccines, with increased host survival further accentuated by simultaneous PD-L1 blockade. Cancer Res; 76(16); 4661–72. ©2016 AACR.

In antitumor immunity, dendritic cells (DCs) capture, process, and present tumor antigens to T cells, initiating a tumoricidal response. However, DCs are often dysfunctional due to their exposure to the tumor microenvironment (TME), leading to tumor escape from immune surveillance. Here, a vital role of microRNA-155 (miR-155) in regulating the function of DCs in breast cancer is reported. Host miR-155 deficiency enhanced breast cancer growth in mice, accompanied by reduced DCs in the tumors and draining lymph nodes. miR-155 deficiency in DCs impaired their maturation, migration ability, cytokine production, and the ability to activate T cells. We demonstrate that miR-155 regulates DC migration through epigenetic modulation of CCR7 expression. Moreover, IL-6 and IL-10, two cytokines abundant in the TME, are found to impair DC maturation by suppressing miR-155 expression. Furthermore, animal studies show that a lack of miR-155 diminishes the effectiveness of DC-based immunotherapy for breast cancer. In conclusion, these findings suggest that miR-155 is a master regulator of DC function in breast cancer, including maturation, cytokine secretion, migration toward lymph nodes, and activation of T-cells. These results suggest that boosting the expression of a single microRNA, miR-155, may significantly improve the efficacy of DC-based immunotherapies for breast cancer.

Purpose: To rapidly identify protein abundance changes in biopsy‐level fresh‐frozen hepatocellular carcinoma (HCC).Experimental design: The pressure‐cycling technology (PCT) is applied and sequential window acquisition of all theoretical mass spectra (SWATH‐MS) workflow is optimized to analyze 38 biopsy‐level tissue samples from 19 HCC patients. Each proteome is analyzed with 45 min LC gradient. MCM7 is validated using immunohistochemistry (IHC).Results: A total of 11 787 proteotypic peptides from 2579 SwissProt proteins are quantified with high confidence. The coefficient of variation (CV) of peptide yield using PCT is 32.9%, and the R² of peptide quantification is 0.9729. Five hundred forty‐one proteins showed significant abundance change between the tumor area and its adjacent benign area. From 24 upregulated pathways and 13 suppressed ones, enhanced biomolecule synthesis and suppressed small molecular metabolism in liver tumor tissues are observed. Protein changes based on α‐fetoprotein expression and hepatitis B virus infection are further analyzed. The data altogether highlight 16 promising tumor marker candidates. The upregulation of minichromosome maintenance complex component 7 (MCM7) is further observed in multiple HCC tumor tissues by IHC.Conclusions and clinical relevance: The practicality of rapid proteomic analysis of biopsy‐level fresh‐frozen HCC tissue samples with PCT–SWATH has been demonstrated and promising tumor marker candidates including MCM7 are identified.