Metabolic Fingerprinting Links Oncogenic PIK3CA with Enhanced Arachidonic Acid-Derived Eicosanoids.

Oncogenic transformation is associated with profound changes in cellular metabolism, but whether tracking these can improve disease stratification or influence therapy decision-making is largely unknown. Using the iKnife to sample the aerosol of cauterized specimens, we demonstrate a new mode of real-time diagnosis, coupling metabolic phenotype to mutant PIK3CA genotype. Oncogenic PIK3CA results in an increase in arachidonic acid and a concomitant overproduction of eicosanoids, acting to promote cell proliferation beyond a cell-autonomous manner. Mechanistically, mutant PIK3CA drives a multimodal signaling network involving mTORC2-PKCζ-mediated activation of the calcium-dependent phospholipase A2 (cPLA2). Notably, inhibiting cPLA2 synergizes with fatty acid-free diet to restore immunogenicity and selectively reduce mutant PIK3CA-induced tumorigenicity. Besides highlighting the potential for metabolic phenotyping in stratified medicine, this study reveals an important role for activated PI3K signaling in regulating arachidonic acid metabolism, uncovering a targetable metabolic vulnerability that largely depends on dietary fat restriction. VIDEO ABSTRACT.

Metabolism and disposition of zamicastat in rats.

The metabolism and disposition of zamicastat, a reversible dopamine ß-hydroxylase (DßH) inhibitor, developed for treatment of Pulmonary Arterial Hypertension (PAH), were investigated in rats after oral and intravenous administration of [14C]-zamicastat.Zamicastat was rapidly absorbed and widely distributed to peripheral tissues, with total radioactivity almost completely recovered 168 h post-dose. Its main route of excretion was via faeces, whilst urine and expired air had minor roles.Maximum plasma concentration of zamicastat-related radioactivity occurred in the first hours, remaining quantifiable up to 144 h. The unchanged zamicastat plasma peak was 2 h post-dose and declined to low levels over 24 h.Zamicastat metabolism occurs largely during the first 8 h with only one metabolite identified in the latest time-point (96 h), the isothiocyanic acid/thiocyanic acid (tautomeric forms). Zamicastat metabolic pathway involved multiple reactions comprising desulphurisation, oxidative desulphurisation, N-debenzylation followed by further oxidation or N-acetylation, and the unexpected multistep metabolic pathway leading to isothiocyanic acid/thiocyanic acid.

Extracellular vesicles as a promising source of lipid biomarkers for breast cancer detection in blood plasma.

Extracellular vesicles (EVs), including exosomes and microvesicles, mediate intercellular communication in cancer, from development to metastasis. EV-based liquid biopsy is a promising strategy for cancer diagnosis as EVs can be found in cancer patients' body fluids. In this study, the lipid composition of breast cancer-derived EVs was studied as well as the potential of blood plasma EVs for the identification of lipid biomarkers for breast cancer detection. Initially, an untargeted lipidomic analysis was carried out for a panel of cancerous and non-cancerous mammary epithelial cells and their secreted EVs. We found that breast cancer-derived EVs are enriched in sphingolipids and glycerophospholipids compared to their parental cells. The initial in vitro study showed that EVs and their parental cells can be correctly classified (100% accuracy) between cancerous and non-cancerous, as well as into their respective breast cancer subtypes, based on their lipid composition. Subsequently, an untargeted lipidomic analysis was carried out for blood plasma EVs from women diagnosed with breast cancer (primary or progressive metastatic breast cancer) as well as healthy women. Correspondingly, when blood plasma EVs were analysed, breast cancer patients and healthy women were correctly classified with an overall accuracy of 93.1%, based on the EVs' lipid composition. Similarly, the analysis of patients with primary breast cancer and healthy women showed an overall accuracy of 95% for their correct classification. Furthermore, primary and metastatic breast cancers were correctly classified with an overall accuracy of 89.5%. This reveals that the blood plasma EVs' lipids may be a promising source of biomarkers for detection of breast cancer. Additionally, this study demonstrates the usefulness of untargeted lipidomics in the study of EV lipid composition and EV-associated biomarker discovery studies. This is a proof-of-concept study and a starting point for further analysis on the identification of EV-based biomarkers for breast cancer.

LXR activation by GW3965 alters fat tissue distribution and adipose tissue inflammation in ob/ob female mice.

To investigate the role of liver X receptor (LXR) in adipose tissue metabolism during obesity, ob/ob mice were treated for 5 weeks with the synthetic LXR agonist GW3965. MRI analysis revealed that pharmacological activation of LXR modified fat distribution by decreasing visceral (VS) fat and inversely increasing subcutaneous (SC) fat storage without affecting whole body fat content. This was concordant with opposite regulation by GW3965 of the lipolytic markers hormone-sensitive lipase (HSL) and adipose triglyceride lipase (ATGL) in the two fat depots; moreover, the expression of genes involved in lipogenesis was significantly induced in SC fat. Lipidomic analysis suggested that changes in lipid composition in response to GW3965 also varied between VS and SC fat. In both depots, the observed alteration in lipid composition indicated an overall change toward less lipotoxic lipids. Flow cytometry analysis showed decreased immune cell infiltration in adipose tissue of ob/ob mice in response to GW3965 treatment, which in VS fat mainly affected the macrophage population and in SC fat the lymphocyte population. In line with this, the expression and secretion of proinflammatory markers was decreased in both fat deposits with GW3965 treatment.

Targeted Desorption Electrospray Ionization Mass Spectrometry Imaging for Drug Distribution, Toxicity, and Tissue Classification Studies.

With increased use of mass spectrometry imaging (MSI) in support of pharmaceutical research and development, there are opportunities to develop analytical pipelines that incorporate exploratory high-performance analysis with higher capacity and faster targeted MSI. Therefore, to enable faster MSI data acquisition we present analyte-targeted desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) utilizing a triple-quadrupole (TQ) mass analyzer. The evaluated platform configuration provided superior sensitivity compared to a conventional time-of-flight (TOF) mass analyzer and thus holds the potential to generate data applicable to pharmaceutical research and development. The platform was successfully operated with sampling rates up to 10 scans/s, comparing positively to the 1 scan/s commonly used on comparable DESI-TOF setups. The higher scan rate enabled investigation of the desorption/ionization processes of endogenous lipid species such as phosphatidylcholines and a co-administered cassette of four orally dosed drugs-erlotininb, moxifloxacin, olanzapine, and terfenadine. This was used to enable understanding of the impact of the desorption/ionization processes in order to optimize the operational parameters, resulting in improved compound coverage for olanzapine and the main olanzapine metabolite, hydroxy-olanzapine, in brain tissue sections compared to DESI-TOF analysis or matrix-assisted laser desorption/ionization (MALDI) platforms. The approach allowed reducing the amount of recorded information, thus reducing the size of datasets from up to 150 GB per experiment down to several hundred MB. The improved performance was demonstrated in case studies investigating the suitability of this approach for mapping drug distribution, spatially resolved profiling of drug-induced nephrotoxicity, and molecular-histological tissue classification of ovarian tumors specimens.

Enhanced triacylglycerol catabolism by carboxylesterase 1 promotes aggressive colorectal carcinoma.

The ability to adapt to low-nutrient microenvironments is essential for tumor cell survival and progression in solid cancers, such as colorectal carcinoma (CRC). Signaling by the NF-κB transcription factor pathway associates with advanced disease stages and shorter survival in patients with CRC. NF-κB has been shown to drive tumor-promoting inflammation, cancer cell survival, and intestinal epithelial cell (IEC) dedifferentiation in mouse models of CRC. However, whether NF-κB affects the metabolic adaptations that fuel aggressive disease in patients with CRC is unknown. Here, we identified carboxylesterase 1 (CES1) as an essential NF-κB-regulated lipase linking obesity-associated inflammation with fat metabolism and adaptation to energy stress in aggressive CRC. CES1 promoted CRC cell survival via cell-autonomous mechanisms that fuel fatty acid oxidation (FAO) and prevent the toxic build-up of triacylglycerols. We found that elevated CES1 expression correlated with worse outcomes in overweight patients with CRC. Accordingly, NF-κB drove CES1 expression in CRC consensus molecular subtype 4 (CMS4), which is associated with obesity, stemness, and inflammation. CES1 was also upregulated by gene amplifications of its transcriptional regulator HNF4A in CMS2 tumors, reinforcing its clinical relevance as a driver of CRC. This subtype-based distribution and unfavorable prognostic correlation distinguished CES1 from other intracellular triacylglycerol lipases and suggest CES1 could provide a route to treat aggressive CRC.

Epithelial ovarian carcinoma diagnosis by desorption electrospray ionization mass spectrometry imaging.

Ovarian cancer is highly prevalent among European women, and is the leading cause of gynaecological cancer death. Current histopathological diagnoses of tumour severity are based on interpretation of, for example, immunohistochemical staining. Desorption electrospray mass spectrometry imaging (DESI-MSI) generates spatially resolved metabolic profiles of tissues and supports an objective investigation of tumour biology. In this study, various ovarian tissue types were analysed by DESI-MSI and co-registered with their corresponding haematoxylin and eosin (H&E) stained images. The mass spectral data reveal tissue type-dependent lipid profiles which are consistent across the n = 110 samples (n = 107 patients) used in this study. Multivariate statistical methods were used to classify samples and identify molecular features discriminating between tissue types. Three main groups of samples (epithelial ovarian carcinoma, borderline ovarian tumours, normal ovarian stroma) were compared as were the carcinoma histotypes (serous, endometrioid, clear cell). Classification rates >84% were achieved for all analyses, and variables differing statistically between groups were determined and putatively identified. The changes noted in various lipid types help to provide a context in terms of tumour biochemistry. The classification of unseen samples demonstrates the capability of DESI-MSI to characterise ovarian samples and to overcome existing limitations in classical histopathology.