Glycocalyx Targeting and Augmenting Cellular Immunity in Lung Cancer

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In lung cancer, tumor immunity is fueled by antigen presenting cells (APCs). We found that targeting glycans on dendritic immune cells (DCs, as “professional” APCs) boosts CD8+ T cell influx and tumor cytolysis, likely via augmented presentation of tumor antigens (Ag) on DCs. Augmenting continuous tumor Ag sensing via this “endogenous” tumor microenvironment (TME) system may improve the specificity and magnitude of anti-tumor immunity. In Merit work, we discovered how genetic targeting of unique DC glycans boosts anti-tumor T cell responses, at least in part via Ag presentation mechanisms. We here build on initial insights. An advantage to facilitating this process is that the “epicenter” for tumor Ag recognition (i.e., MHC/Ag - T-cell receptor axis) may respond to dynamic Ag changes (including new mutations) to sustain tumor-cytotoxic CD8+ T cells in the TME. Current immune-checkpoint therapy in advanced-stage lung cancer shows durable remissions that are limited (<25%), and autoimmune toxicity poses serious challenges. We hypothesize that altering glycan fine structure (proteoglycan under-sulfation or sialic acid inhibition) on the APC glycocalyx will boost Ag presentation and augment anti-tumor T cell responses. To study this, with novel inhibitor strategies, we propose Aims as follows: Study how targeting glycocalyx composition on key APCs from the TME augments anti-tumor T cell responses, and test how genetic under-sulfation and under-sialylation of the lung APC glycocalyx affects tumor growth inhibition and specific anti-tumor T-cell responses, including in vivo T cell proliferation and infiltration (Aim 1). This will involve altering the fine structure of heparan sulfate or inhibiting sialic acid glycans that repress APC effector functions in the TME. We will examine CD8+ T cell proliferation, activation, and tumor cytolysis in the setting of APC-targeted glycan inhibition in tumor models with Ag-responsive T cell reporting systems. Studies will include characterizing how mutation affects tolerance/exhaustion signatures in DCs purified from the lung TME. We also study lymphatic glycan targeting to transform lymphatic endothelium into a novel APC platform. Discover mechanisms by which glycan structural changes promote spatial and temporal display of MHC/Ag on tumor DCs, and study how this augments engagement with the T cell receptor (TcR) using model Ag and transgenic T cells (Aim 2). We will study how DC glycocalyx mutations in the fine structure of heparan sulfate and sialic acid affect Ag presentation by tumor-Ag sensitized DCs or ex-vivo marrow DCs pulsed with model Ag. We will also study how such mutations on model Ag systems affect TcR interactions with DC MHC-I/Ag using novel proximity ligation technology. Beyond these approaches, we will measure MHC-I/Ag internalization on the DC surface in the setting of glycocalyx targeting strategies. We will include parallel studies on tumor lymphatic endothelial cells as unique APCs in Ag-presentation/TcR engagement studies using similar methods. We will develop glycan-specific inhibitors that augment APC-mediated tumor-kill responses and tumor antigen responsiveness when applied to mouse as well as human lung carcinomas, and complement this with studies of real-time tumor regression using inhibitors in spontaneous lung cancer models (Aim 3). We will study anti- tumor T-cell responses using adoptive therapeutic approaches with glycan-targeted DCs, including antisense oligonucleotide (ASO) approaches to inhibit tumor-APC glycan fine structure, and examine anti-tumor CD8+ T cell responses in ASO targeted spontaneous lung cancer models. We include immune technology to study T cell responses to unique model tumor antigens as well as non-biased responses to high neo-antigen tumor loads in the system. We will also assess how APC glycocalyx targeting affects immune checkpoint pathways in tumor CD8+ T cells, and study how pairing inhibitors with low-level checkpoint blockade potentiates responses.