use-cerebral-open-flow-microperfusion-for-longitudinal-collection

Use of Cerebral Open-Flow Microperfusion for the Longitudinal Collection of Interstitial Fluid in an Animal Model of Glioblastoma

Diseases of the central nervous system (CNS) manifest through a complex, dynamic network of intercellular communication. Glioblastoma (GBM), the most ...

Northwestern University

Immunometabolism is a burgeoning field of research that investigates how immune cells harness nutrients to drive their growth and functions. Myeloid cells play a pivotal role in tumor biology, yet their metabolic influence on tumor growth and antitumor immune responses remains inadequately understood. This Review explores the metabolic landscape of tumor-associated macrophages, including the immunoregulatory roles of glucose, fatty acids, glutamine, and arginine, alongside the tools used to perturb their metabolism to promote antitumor immunity. The confounding role of metabolic inhibitors on our interpretation of myeloid metabolic phenotypes will also be discussed. A binary metabolic schema is currently used to describe macrophage immunological phenotypes, characterizing inflammatory M1 phenotypes, as supported by glycolysis, and immunosuppressive M2 phenotypes, as supported by oxidative phosphorylation. However, this classification likely underestimates the variety of states in vivo. Understanding these nuances will be critical when developing interventional metabolic strategies. Future research should focus on refining drug specificity and targeted delivery methods to maximize therapeutic efficacy.

Modulation of macrophage metabolism as an emerging immunotherapy strategy for cancer.

Sequential multiplex methodologies such as Akoya CODEX, Miltenyi MACSima, Rarecyte Orion, and others require modification of the antibodies by conjugation to an oligo or a specific fluorophore which means the use of off-the-shelf reagents is not possible. Modifications of these antibodies are typically performed via reduction chemistry and thus require verification and validation post-modification. Fixed panels are therefore developed due to various limitations including spectral overlap that creates spectral unmixing issues, steric hindrance, harsh antibody removal, and tissue degradation throughout the labeling. As such, a complex interrogation evaluating multiple study hypotheses and/or endpoints requires the development of sequential panels, reconstruction, and realignment of the tissue that necessitate a z-stack strategy. Standardized antibody panels are typically fixed and require substantial validation efforts to modify a single target and thus do not evolve with the pace of research interests. To increase the throughput of profiling cells within the human central nervous system (CNS), we developed and validated a CNS-specific library with an associated analysis platform using the newly developed Lunaphore COMET platform. The COMET is an automated staining/imaging instrument integrating a reagent deck for staining buffers and off-the-shelf label-free primary antibodies and fluorophore-labeled secondary antibodies, which feed into a circular plate holding up to 4 slides that are automatically imaged in microscope-operated control software. For this study, standard formalin fixed paraffin embedded histology slides are used. However, the COMET is capable of imaging fresh-frozen samples using specialized settings. Our methodologies address an unmet need in the neuroscience field while leveraging prior developmental efforts in the domain of immunology spatial profiling. Cataloging and validating a large series of antibodies on the COMET along with developing CNS autofluorescence management strategies while optimizing standard operating procedures have allowed for the visualization at the subcellular level. Forty analytes can be used to analyze one specimen which has clinical utility in cases in which the CNS can only be sampled by biopsy. CNS biopsies, depending on the anatomical location, can have limited available volume to a degree that requires prioritization and restriction to select analysis. In-depth bioinformatic imaging analysis can be done using standard bioinformatic tools and software such as Visiopharm. These results establish a general framework for imaging and quantifying cell populations and networks within the CNS while providing the scientific community with standard operating procedures.

High Dimensional Proteomic Multiplex Imaging of the Central Nervous System Using the COMET System.

Leptomeningeal disease (LMD) is a devastating manifestation of late-stage cancer which currently suffers from a lack of effective therapeutics. Unfortunately, a significant obstacle preventing the widespread development and testing of therapeutics for LMD is the lack of biologically accurate animal models. We provide overviews of six types of animal models of leptomeningeal metastasis from solid tumors: injection of tumor cells into the cerebrospinal fluid (CSF), blood, or brain parenchyma; subcutaneous or mammary fat pad injection of tumor cells; the LeptoM/LM-phenotype model; and genetic manipulation. We identify the pros and cons of each model and suggest broad areas of future research that could improve each model in terms of its similarity to human LMD.

Article	Year
Modulation of macrophage metabolism as an emerging immunotherapy strategy for cancer. The Journal of clinical investigation\| PubMed ID: 38226622	2024
High Dimensional Proteomic Multiplex Imaging of the Central Nervous System Using the COMET System. bioRxiv : the preprint server for biology\| PubMed ID: 40027731	2025
Beyond the brain: exploring the impact of animal models of leptomeningeal disease from solid tumors. Acta neuropathologica communications\| PubMed ID: 40383789	2025

Jillyn Turunen

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