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Article History

Received: 19 June 2025

Accepted: 27 August 2025

First Online: 8 September 2025

Declarations

:

: Animal experiments in this study were approved by the local Ethics Committee of Animal Experiments authorities (LANUV, North Rhine-Westphalia, Germany) under license number 81-02.04.2019-A491. All mice were maintained according to FELASA recommendations and in compliance with the European Union and German guidelines. Mice were monitored biweekly for tumor progression using MRI imaging, and animals were euthanized once a sufficient tumor burden was reached, as per predefined criteria (e.g., weight loss > 20%, severe tumor ulceration, or breathing difficulties). Euthanasia was conducted in compliance with the license 8102.04.2019-A491 approved by the LANUV authority (North-Rhine-Westphalia, Germany) and adhered to FELASA, EU, and German guidelines. The euthanasia method involved terminal anesthesia followed by transcardial perfusion with 10 ml sterile PBS to clear blood from the lungs and minimize contamination of tissue samples. The specific anesthesia used for imaging and terminal procedures was isoflurane (2.5% in oxygen) or intraperitoneal injection of ketamine (100 mg/kg) combined with xylazine (20 mg/kg). Anesthesia depth was confirmed by the absence of response to toe pinch before proceeding with perfusion and tissue harvesting. This approach was chosen to ensure minimal suffering, rapid unconsciousness, and high-quality tissue preservation suitable for downstream analyses such as IMC and genomic studies.

: Not applicable.

: H.C.R. received consulting and lecture fees from Abbvie, AstraZeneca, Vertex, and Merck. H.C.R. received research funding from AstraZeneca and Gilead Pharmaceuticals. H.C.R. is a co-founder of CDL Therapeutics GmbH. The remaining authors declare no competing financial interest.