空间分辨代谢组学能够在原位研究肿瘤代谢物。肿瘤的间质和内质异质性是与患者预后密切相关的关键因素。肾上腺皮质癌（ACC）是一种非常罕见的肿瘤，与存活率较低有关。其临床预后具有高度变异性，但至今我们尚未了解肿瘤代谢异质性的贡献。对肿瘤异质性的深入理解需要通过基于分子特征的肿瘤亚群体的鉴定来评估与肿瘤侵袭性相关的情况。本研究使用高质量质谱成像（MALDI傅里叶变换离子回旋共振质谱成像）的空间代谢组学，通过代谢物亚群体的全新发现和Simpson多样性指数来评估代谢异质性。在72名ACC患者中鉴定出肿瘤亚群体后，我们还与25个正常肾上腺皮质组织切片进行了比较，以鉴定它们的共同和独特的代谢亚群体。我们观察到ACC肿瘤的异质性变异性，并发现高代谢异质性与较差的临床预后有关。此外，我们还鉴定出了作为独立预后因素的肿瘤亚群体，并发现了4种相关的抗癌药物作用途径。我们的研究可能有助于全面了解ACC中肿瘤亚群体的生物学意义，并表明代谢异质性可能会影响化疗。

Spatially resolved metabolomics enables the investigation of tumoral metabolites in situ. Inter- and intratumor heterogeneity are key factors associated with patient outcomes. Adrenocortical carcinoma (ACC) is an exceedingly rare tumor associated with poor survival. Its clinical prognosis is highly variable, but the contributions of tumor metabolic heterogeneity have not been investigated thus far to our knowledge. An in-depth understanding of tumor heterogeneity requires molecular feature-based identification of tumor subpopulations associated with tumor aggressiveness. Here, using spatial metabolomics by high-mass resolution MALDI Fourier transform ion cyclotron resonance mass spectrometry imaging, we assessed metabolic heterogeneity by de novo discovery of metabolic subpopulations and Simpson's diversity index. After identification of tumor subpopulations in 72 patients with ACC, we additionally performed a comparison with 25 tissue sections of normal adrenal cortex to identify their common and unique metabolic subpopulations. We observed variability of ACC tumor heterogeneity and correlation of high metabolic heterogeneity with worse clinical outcome. Moreover, we identified tumor subpopulations that served as independent prognostic factors and, furthermore, discovered 4 associated anticancer drug action pathways. Our research may facilitate comprehensive understanding of the biological implications of tumor subpopulations in ACC and showed that metabolic heterogeneity might impact chemotherapy.