Intravital and Whole-Organ Imaging Reveals Capture of Melanoma-Derived Antigen by Lymph Node Subcapsular Macrophages Leading to Widespread Deposition on Follicular Dendritic Cells

Federica Moalli,Steven T Proulx,Michael Detmar,Jens V Stein,Reto Schwendener,Christoph Schlapbach

doi:10.3389/fimmu.2015.00114

Abstract

Aberrant antigens expressed by tumor cells, such as in melanoma, are often associated with humoral immune responses, which may in turn influence tumor progression. Despite recent data showing the central role of adaptive immune responses on cancer spread or control, it remains poorly understood where and how tumor-derived antigen (TDA) induces a humoral immune response in tumor-bearing hosts. Based on our observation of TDA accumulation in B cell areas of lymph nodes (LNs) from melanoma patients, we developed a pre-metastatic B16.F10 melanoma model expressing a fluorescent fusion protein, tandem dimer tomato, as a surrogate TDA. Using intravital two-photon microscopy (2PM) and whole-mount 3D LN imaging of tumor-draining LNs in immunocompetent mice, we report an unexpectedly widespread accumulation of TDA on follicular dendritic cells (FDCs), which were dynamically scanned by circulating B cells. Furthermore, 2PM imaging identified macrophages located in the subcapsular sinus of tumor-draining LNs to capture subcellular TDA-containing particles arriving in afferent lymph. As a consequence, depletion of macrophages or genetic ablation of B cells and FDCs resulted in dramatically reduced TDA capture in tumor-draining LNs. In sum, we identified a major pathway for the induction of humoral responses in a melanoma model, which may be exploitable to manipulate anti-TDA antibody production during cancer immunotherapy.

Highlights

Melanoma, a potentially lethal skin cancer, is often associated with the induction of humoral responses, which are typically inversely correlated with the extent of the disease [1, 2]
IMMUNOFLUORESCENT ANALYSIS OF MelA CO-LOCALIZATION WITH B CELLS IN HUMAN tumor-draining lymph nodes (TDLNs) SECTIONS To establish whether tumor-derived antigen (TDA) deposition is observed in human lymphoid tissue, we analyzed paraffin TDLN sections of human melanoma patients by immunofluorescent staining, focusing on MelA as model melanoma antigen
TDA deposition is found in human TDLNs, where it is likely to contribute to the anti-IgG response observed in melanoma patients

Summary

Introduction

A potentially lethal skin cancer, is often associated with the induction of humoral responses, which are typically inversely correlated with the extent of the disease [1, 2]. There are two major factors underlying this surge of interest: first, comprehensive patient studies have shown that the presence of adaptive immune cells in tumors positively correlates with a beneficial prognosis [8, 9]. A recent study has shown that antibodies directed against both surface and cytoplasmic molecules contribute to efficient CD8+ T cell responses by forming immune complexes that are efficiently taken up by dendritic cells (DCs) for presentation to cytotoxic T cells [14]. Antibodies directed against intracellular tumor antigens are able to directly kill cancer cells [15]. Anti-TDA Ig may exert multiple effects on tumor control, beyond direct killing or antibody-dependent cytotoxicity

Methods

Results

Conclusion