News from … Part I

Hartmann Lab, DKFZ, Heidelberg, Germany

Therapy-induced metabolic remodeling in HER2-negative breast cancer
Breast cancer remains a leading cause of cancer-related mortality among women. Although it is often diagnosed early, up to 35% of HER2-negative cases relapse as metastatic disease, with median overall survival ranging from 2 years (TNBC) to 5 years (hormone receptor-positive). Several new treatments have been tested in early-stage disease to reduce metastatic relapse, including antiangiogenic agents such as bevacizumab. Bevacizumab can normalize the vasculature and stroma in about one-third of cases, leading to a significant metabolic shift in cancer cells from primarily glycolytic to mitochondrial energy production. This shift can be exploited therapeutically by combining antiangiogenics with mitochondrial inhibitors, known as “metabolic synthetic lethality.” Using spatially resolved single-cell proteomics, we investigated samples from a phase 0/I clinical study (NCT02806817) that combined bevacizumab with ME-344, a mitochondrial complex I inhibitor. We show that bevacizumab reshapes vascular and epithelial metabolism and reorganizes immune cell interactions, thereby increasing epithelial reliance on oxidative phosphorylation. ME-344 then leverages this state to cause mitochondrial dysfunction, reprogram cellular metabolism, and induce oxidative damage in epithelial cells. This is accompanied by an accumulation of iron-rich CD163+ macrophages in certain niches, along with activation and exhaustion of CD8+ T cells. These findings highlight the importance of spatial context in evaluating targeted therapies and suggest that mitochondrial and iron metabolism may offer new opportunities to modulate tumor immunity in combination treatment strategies.

Department of Dermatology, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany

Cross talk between skin and joints impairs bone homeostasis in psoriatic arthritis

Psoriasis (PsO) and psoriatic arthritis (PsA) involve interconnected pathologies of the skin and joints. While Innate Lymphoid Cells (ILCs) are implicated in the “skin-joint axis,” the mechanisms driving the transition from cutaneous to joint inflammation remain unclear. This study utilizes Imaging Mass Cytometry (IMC) to spatially profile immune populations in PsO and PsA lesional skin.

The high-dimensionality of our 33-antibody panel allowed for the simultaneous assessment of broad lineage and functional markers on a single tissue section. This multiplexing capability enabled the precise resolution of heterogeneous populations—including rare ILCs, Tissue-Resident Memory T cells (TRMs), and distinct monocyte and T-cell subsets—while preserving their spatial context within the tissue architecture.

In a pilot analysis of two patient samples, we observed that the chemokine CXCL16 was predominantly expressed in the epidermis and monocytes. Furthermore, preliminary comparisons suggested distinct immunophenotypes: the PsO sample exhibited a trend toward stronger expression of both CXCL16 and its receptor CXCR6, as well as a higher frequency of TRMs, compared to the PsA sample.

These initial observations suggest that the CXCL16/CXCR6 axis may be more prominent in the migration of the immune cells in PsA patients. While validation in a larger cohort is required, this spatially resolved approach demonstrates the feasibility of using IMC to uncover disease-driving factors. Expanding this study will be crucial for identifying biomarkers predictive of the migration of skinderived immune cells to joint tissues.

Department of Medicine 3 – Rheumatology and Immunology, Friedrich-Alexander-Universität Erlangen-Nürnberg and Uniklinikum Erlangen, Erlangen, Germany.

Deutsches Zentrum für Immuntherapie (DZI), Friedrich-Alexander-Universität Erlangen-Nürnberg and Uniklinikum Erlangen

Lymphatic tissue remodelling following CD19 CAR T-cell therapy assessed by imaging mass cytometry
Anti-CD19 chimeric antigen receptor (CAR) T-cell therapy can induce long-term drug-free remission in patients with autoimmune diseases (AIDs). Two previous proof-of-concept studies based on sequential lymph node biopsies demonstrated a consistent and complete B-cell depletion in lymphatic tissue after CD19 CAR T-cell treatment, as assessed by conventional immunohistochemistry, suggesting a potential mechanism underlying an “immune reset” in AIDs.^1,2

In this study, we applied imaging mass cytometry (IMC) to deconvolute cellular heterogeneity at single-cell resolution, analyzing 1,328,358 cells to investigate changes in spatial distribution and cellular neighborhoods before and after CAR T-cell therapy, in comparison with healthy controls. In parallel, lymph node samples from patients treated with the standard-of-care B-cell–depleting agent rituximab were analyzed.

We included 15 patients treated with CD19 CAR T cells and 7 patients treated with rituximab, all with paired baseline and 60-day follow-up inguinal needle biopsies, as well as 5 disease-free lymph node samples obtained during surgery from different anatomical sites, serving as healthy controls. In addition, for 8 of the 15 CAR T-cell–treated patients, lymph node samples collected one year after treatment were available.

Stromal, endothelial, B, T and myeloid cell compartments were identified, and immune subpopulations were further characterized, with a particular focus on the follicular network, including germinal center B cells (CD23⁺Bcl6⁺), follicular dendritic cells (CD23⁺CD21⁺) and T follicular helper cells (CD3⁺PD-1⁺Bcl6⁺). IMC confirmed complete B-cell depletion following CAR T-cell therapy, accompanied by a profound abrogation of the follicular architecture, and demonstrated a full restoration of normal follicular organization one year after treatment. Notably, early follow-up biopsies revealed, through neighborhood analysis, repopulation of the ablated follicular areas by stromal and myeloid cells, together with an increased presence of Foxp3⁺ cells.

By employing IMC, we provide an in-depth characterization of the lymph node immune landscape and its dynamic remodeling following CD19 CAR T-cell therapy. Our findings not only corroborate previous evidence of profound and durable B-cell depletion, but also uncover previously unappreciated alterations in T-cell and myeloid cell responses within a fully B-cell-depleted environment.

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BIH Cytometry Core Facility, BIH at Charite – Universitätsmedizin Berlin, Berlin, Germany
New adventures – changing from Hyperion ‘Tangerine’ to XTi ‘Blanche’

This talk will highlight the challenges – and positive surprises – we have encountered at the BIH CCF during our transition from the CyTOF2/Helios/Hyperion system to the new CyTOF XT/XTi platform. A particular focus will be on the core facility’s perspective, including our efforts to set up workflows for service requests, sample intake, sample measurements, and data management.

Medical University of Vienna, Vienna Austria

Integrative spatial proteomics by Imaging Mass Cytometry and MALDI-MSI uncovers tumor- and microenvironment-driven heterogeneity in breast cancer
Breast cancer (BC) remains a global health challenge, impacting a significant portion of the female population. Understanding proteomic heterogeneity within BC is crucial for improving diagnostic accuracy and therapeutic efficacy. Imaging Mass Cytometry (IMC) of tumor xenograft models provides high spatial resolution and targeted protein analysis within BC tissue architecture but is limited in
multiplexing. In contrast, Imaging Mass Spectrometry (IMS) enables broader molecular profiling with higher multiplexing capabilities, but limited in spatial resolution and confidence of identification.
Human BC cell lines (MCF-7, SKBR-3, MDA-MB-231) were inoculated into female athymic BALB/c-nude mice. Excised tumors were embedded in paraffin, or were snap-frozen in liquid nitrogen. One tissue was allocated for IMC and an adjacent one for IMS. An optimized IMC panel, selected based on literature review and a bulk proteomics experiment, revealed clear heterogeneity between BC subtypes and within tissues, demonstrating inter- and intra-tumoral heterogeneity. IMS explored the broader proteomic landscape and identified more candidate proteins. Bulk tissue LC-MS/MS identified approximately 17,000 proteins, enabling validation of IMS signals.
Integration of IMC and IMS datasets allowed spatial and molecular correlation. Segmentation maps aligned with histological assessments and distinguished tumor, stroma, and necrotic regions. Unsupervised U-MAP clustering revealed distinct molecular regions with unique proteomic profiles.
Combining IMC and IMS provided a comprehensive characterization of BC heterogeneity, identifying spatial proteomic variation not detectable by conventional methods. As next steps we aim to combine these translational research results with in vivo PET/MRI data to develop non-invasive diagnostic tools and enhance personalized therapeutic strategies.

Klinik für Innere Medizin II, Universitätsklinikum Freiburg, Freiburg, Germany

Heavy metal based research in Freiburg: Update 2026
This talk will give a brief overview about recent developments in the mass cytometry unit Freiburg and will discuss an approach to understand comprehensive signaling network changes in primary human exhausted T cells at the single-cell level.