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. 2023 Aug 14;5(8):1303–1318. doi: 10.1038/s42255-023-00857-0

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Extended Data Fig. 3 — (A) H&E of distal colon tissue with an Apc-deficient tumour [same H&E as shown in Fig. 2B and Extended Data Fig. 2A; N. Adj.: normal adjacent tissue; S: stroma; T: tumour tissue]. Tissues analysed for n = 4 animals; n = 1 shown. Scale bar: 1 mm. (B) specific accumulation of an ion with m/z 464.288 in the tumour epithelial compartment of APC tumours as detected by MALDI-MSI. The two top tentative parent ion identifications (that is glycocholic acid and LysoPE 17:1) obtained from a publicly available database (https://hmdb.ca) were experimentally validated. (C) A ¹³C-glycocholic acid (¹³C-GA) standard solution was infused to detect the parent ion (blue dot indicates ¹³C label) using Triple Quadrupole Mass Spectrometry, and (D) a Multiple Reaction Monitoring (MRM) method was developed to study the abundance of ¹³C-GA in tissues via its a diagnostic ion of 75 Da (blue dot indicates ¹³C label). Tumour-bearing APC mice were administered ¹³C-GA (75 mg/kg p.o.; n = 2) or vehicle (HPMC/Tween:DMSO; v-v 90:10; n = 1). After 8.5 hours, serum, liver and distal colon were harvested and processed for LC-MS. ¹³C-GA was detected in (E) liver, (F) serum, (G) normal adjacent colonic tissue and tumour tissues. No increased abundance of (G) ¹³C-GA or (H) ¹²C-GA was observed in tumour tissues compared with normal adjacent colonic tissue. Panels E-H show the mean of metabolic extractions of the multiple tissue fragments for each mouse, each dot represents data obtained from a single tissue fragment, or serum extract. Liquid extraction surface analysis tandem mass spectrometry was applied to study fragmentation of the m/z 464.288 precursor ion, which showed matched ions to the predicted fragmentation pattern of LysoPE 17:1, as well as to the (I) fragmentation of a commercially available standard further supporting the ID of this metabolite of interest. All highlighted m/z of fragments and precursor ions on the mass spectra are present in both standard and sample.

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