Isotopics – December 2018 | Edition 8

As we continue to push the limits of biological analysis across multiple ’omics fields, we recognize that the tools for success are more important and more diverse than ever. CIL remains dedicated to multiomics research by developing new stable isotope products to better understand disease mechanisms and detect biomarkers with more confidence and accuracy than ever before. Our product offering consists of highly enriched, individual standards as well as complex multi-component mixes and kits for qualification, quantification, and quality control. The articles selected for this edition utilize a myriad of these products; please take a look for yourself.

Lysine Propionylation Boosts Proteome Sequence Coverage and Enables a “Silent SILAC” Labeling Strategy for Relative Protein Quantification

Schräder, C.U.; Moore, S.; Goodarzi, A.A.; et al.

Quantitative proteomic protocols incorporating stable isotopes tend to do so with metabolic or chemical processes and typically utilize trypsin. Although the utility of alternative proteases has been demonstrated, quantitative studies using these alternatives are effectively nonexistent. Here, Schriemer and colleagues developed a novel isotope labeling strategy to quantify metabolically labeled proteins on the MS2 level regardless of protease. This relies on SILAC-like metabolic labeling and TMT-like quantitative analysis, to deliver improved protein sequence coverage without sacrifice to protein identifications.

¹³C-Labeled Yeast as Internal Standard for LC-MS/MS and LC High Resolution MS-based Amino Acid Quantification in Human Plasma

Hermann, G.; Schwaiger, M.; Volejnik, P.; et al.

Isotopically labeled extracts can be powerful standards for analyte quantification in MS-based assays. Here, Hermann and colleagues evaluated the utility of an in vivo-produced metabolite yeast extract in the absolute quantification of 14 amino acids in human plasma (SRM 1950). The amino acid content of the U-¹³C yeast extract (ISO1) was determined using SRM 2389a. The calibrated ISO1 served as internal standards in the cross-platform validations and was found to deliver excellent accuracy without impact on other assay figures of merit.

Glutamine-derived 2-Hydroxyglutarate Is Associated with Disease Progression in Plasma Cell Malignancies

Gonsalves, W.I.; Ramakrishnan, V.; Hitosugi, T.; et al.

To better understand the clinical significance of 2-hydroxyglutarate (2-HG) in its progression of plasma cell malignancies, Gonsalves and colleagues investigated the production and flux of TCA cycle metabolites in various cell and tissue analyses. The quantitative assessments were facilitated by a stable isotope-resolved metabolomics (SIRM) approach that utilized U-¹³C standards and MS-based methodologies. This study revealed that 2-HG is produced from clonal plasma cells (PCs) and is detectable in the peripheral blood and bone marrow plasma of patients with PC disorders. Also observed was evidence for the exclusive production of 2-HG from glutamine metabolism that appears to be correlated with c-Myc upregulation. This is a clinically significant finding considering the functional link of 2-HG with multiple myeloma that has the potential to be exploited as a decision factor in therapy initiation.

Hepatic Ketogenic Insufficiency Reprograms Hepatic Glycogen Metabolism and the Lipidome

d'Avignon, D.A.; Puchalska, P.; Ercal, B.; et al. 

In this article, the authors use metabolomics and stable isotope tracers in animals to reveal an unexpected relationship between ketogenic insufficiency and hepatic glycogen metabolism. They find that when ketogenesis is impaired in mice by knocking down 3-hydroxymethylglutaryl-CoA synthase, hepatic glucose production increased by more than 60% in the fed state. This increase in glucose production was fueled by breaking down glycogen. Additionally, the authors show that bis(monoacylglycero)phosphates accumulate in the livers of fed animals with ketogenic insufficiency. These lipids, which also accumulated in mouse models of nonalcoholic fatty liver disease (NAFLD), may represent a generalized biomarker for NAFLD progression.

A Targeted Metabolomics Approach for Clinical Diagnosis of Inborn Errors of Metabolism

Jacob, M.; Malkawi, A.; Albast, N., et al.

In an effort to improve biological interpretation in clinical research, Rahman and colleagues from King Faisal Specialist Hospital and Research Center developed a targeted LC-MS/MS metabolomics method to screen for a broad collection of inborn errors of metabolism (IEM) in rat and human tissues. The validated assay utilizes stable isotope standards and was demonstrated to be sensitive and robust in differential metabolic profiling of common IEMs.

A Guide to ¹³C Metabolic Flux Analysis for the Cancer Biologist

Antoniewicz, M.R.

Here Maciek Antoniewicz reviews ¹³C metabolic flux analysis (MFA) for quantifying intracellular fluxes in cancer cell metabolism. Detailed are the fundamentals for experimental design, analytical measurement, and statistical analysis along with the merits and limitations of various tracer approaches.

SNMMI Image of the Year Is the Endocyte 177 Lu PSMA-617

Each year, SNMMI chooses an image that exemplifies the most promising advances in the field of nuclear medicine and molecular imaging. Positron emission tomography (PET) scans showing very positive results obtained in cancer patients after treatment with the radionuclide lutetium-177 (¹⁷⁷Lu) PSMA-617 prostate cancer therapy were selected by the Society of Nuclear Medicine and Molecular Imaging (SNMMI) as the SNMMI Image of the year. ABX (a CIL subsidiary) licensed this therapy last year to Endocyte.