(6r) Ultrasmall C’ Dots Activate Pro-Inflammatory Anti-Tumor Responses in the Microenvironment of PDGF-B Driven High Grade Gliomas

Research Interests: My research goals are directed toward developing Clinical Translational nanoscale technologies with particular emphasis on developing nanoscale biomaterials and immunoengineering systems for immuno-oncology, immunoengineering, and molecular imaging. My main research interest lies in immuno-oncology of solid tumors, with a focus on myeloid cells, particularly tumor-associated macrophages, dendritic cells, and myeloid-derived suppressor cells.A major question my research is investigating is, “What role do nanoparticles play in the tumor microenvironment to regulate innate and adaptive immune systems?” Thus, I am studying the impact of proteins and signals that promote the activation and migration of pro-inflammatory cells into tumor microenvironment and also affect the phenotype of immune cells. In addition, I study human antigen presenting cells (APCs), chemokines, and myeloid cells in linking innate and adaptive immunity. My focus has been on dynamic interactions of macrophages and dendritic cells with T cells during antigen presentation and the possibility of using nanoparticles in immuno-oncology to prime T cells for immune checkpoint inhibitors and CAR-T Cell Therapy.

Teaching Interests: My teaching interests lie in the interface of Immunoengineering, cancer immunotherapy, and nanomedicine. I plan to develop highly engaging and critically important courses such as: (1) nanoparticles in cancer immunotherapy. (2) Advanced topics in biomaterials and cancer therapy with a special focus on the relationships between the tumor microenvironment, nanoparticles, and the immune system. I will also arrange journal clubs in order to: Improve competence and confidence in leading discussion and resolving conflicts and improve and refine presentation and communication skills. Through the journal club, I will share research findings or the latest literature in the field.

Introduction: Therapeutic approaches targeting high-grade glioma have largely failed. One alternative strategy is to regulate cells in the tumor microenvironment (TME), such as tumor-associated macrophages and microglia (TAMs). TAMs account for as much as 30% of the tumor mass in mouse models of high-grade glioma and in brain tumor patients; their accumulation is associated with a higher glioma grade and poor patient prognosis. TAMs closely resemble the M2-polarized macrophages and have been shown to contribute to tumor initiation and maintenance, as well as influence anti-tumor autoimmunity via cytokine release and inflammatory recruitment to the TME. Tumors, in turn, can promote the polarization of monocytes into M2 TAMs by releasing factors, such as TGF-beta and M-CSF. We have previously showed that systemically administered sub 8-nm targeted Cornell prime dots (C’ dots) are preferentially internalized by cancer cells, rather than TAMs, and can induce ferroptosis under select conditions. However, the influence of these particle-based probes on immune cell populations (e.g., TAMs) within the TME is unclear. In this work, we hypothesize that non-targeted particles (i.e., PEG-C’ dots) may induce a pro-inflammatory immune response by promoting M1 macrophage polarization and depleting M2 macrophages.

Materials and Methods: Bone marrow derived macrophages (BMDMs) were isolated from nestin–Tv-a; Cdkn2a−/− transgenic mice (PDG) and treated over time with a range of PEG-C’ dot concentrations. Gene signature profiles and cytokine secretion profiles were evaluated in vitro by quantitative real-time PCR and luminex assays, respectively. Cell phenotyping, T cell priming, and phagocytosis rates were evaluating by flow cytometry and immunofluorescence assays. In vivo, PDGF-B driven high grade glioma mice (mGBM) were i.v.-injected with a single-dose PEG-C’ dots to assess for alterations in the TME, including (1) polarization of TAMs, (2) tumor cell phagocytosis, and (3) promotion of pro-inflammatory cytokine secretion profiles.

Results and Discussion: In vitro, exposure of macrophages to PEG-C’ dots led to upregulated M1- and downregulated M2 signatures over a 24-hour period. Macrophages also demonstrated increased pro-inflammatory cytokines and phagocytosis rates in comparison with untreated controls. In addition, PEG-C’ dot exposure led to proliferation and activation of T cells, as well as a significant increase in proliferation of antigen presenting cells. In vivo, PEG-C’ dots significantly enhanced pro-inflammatory responses in the TME. Multicolor flow cytometry of brain tumor tissue specimens revealed prominence of infiltrating macrophages within PEG-C’ dot-treated tumors when compared with untreated tumors and WT brains, accounting for about 50% of all live cells in the brains of treated animals. M1-like subtype (MHC-II high Ly6C-) expression levels were more prevalent in PEG-C’ dot-treated tumors, unlike M2-like subtype (MHC-II- Ly6C-) expression levels, which were lower in treated tumors. Concurrent Luminex assay studies revealed significantly suppressed expression of pro-tumoral cytokines. In addition, anti-inflammatory cells and angiogenesis marker levels were significantly higher in untreated groups, confirmed by CD206⁺ and CD31⁺ immunohistochemistry staining.

Conclusion: In a first-of-its-kind study, a distinct pro-inflammatory mechanism was induced in non-functionalized C’ dot-exposed immune cells (TAMs, T cells) and mGBM tumors. The results of this work suggest that anti-tumor responses may be augmented by induction of this pro-inflammatory phenotype, which might further be exploited as part of a combinatorial treatment regimen.