Targeted Metabolomic Profiles Are Strongly Correlated With Metabolic Alterations In Patients With Sickle Cell/Beta Thalassemia Disease

The complex pathophysiology of Sickle Cell Disease (SCD) makes unlikely that a single therapeutic agent will prevent or reverse all SCD complications. Metabolomic analysis might help in the characterization of the endogenous and exogenous effects of potential new treatments. Metabolites are small molecules that are chemically transformed during metabolism and provide a functional readout of cellular state. Metabolites serve as direct signatures of biochemical activity and are therefore easier to correlate with phenotype. The metabolome is typically defined as the collection of small molecules produced by cells and offers a window for interrogating how mechanistic biochemistry relates to cellular phenotype. There are very few reports providing comprehensive measurements of metabolites present in blood and almost no reports on metabolites changes associated with SCD. In this context we aimed to detect and to quantify targeted metabolites’ abnormalities in patients with Sickle Cell/beta thalassemia disease (HbS/β^Thal) and their implication in pathways that might be of interest to prevent vaso-occlusion and/or to monitor the effects of new therapies on SCD.

Twenty adult patients with HbS/β^Thal were enrolled in the study, while 20 apparently healthy individuals matched for age served as controls. Targeted metabolome analyses based on aminoacids and carnitines were performed after extraction from dry blood spots (DBSs) on filter paper using High Performance Liquid Chromatography followed by tandem Mass Spectrometry (LC/MS/MS), with derivatization (AB SCIEX 5500 triple quadrupole and QTRAP® LC/MS/MS Systems, Framingham, MA, USA) with reagents provided by Chromsystems Instruments & Chemicals, Germany. The injection volume was 10 µL and the analysis lasted ca. 2 min.

The main results of the study showed that: a) fifty metabolites were separated in patients and controls samples, b) mapping the results of analyses, patients with HbS/β^Thal compared to controls had 17 metabolites with significantly lower concentration, 10 metabolites with comparable concentration and 23 metabolites with significantly higher concentration, c) L-arginine and L-ornithine concentrations were significantly lower in patients HbS/β^Thal compared to controls, 9.3±2.6 vs 14.7±3.7 µmoles/L, (p<0.01), and 116.0±15.0 vs 211.2±19.5 µmoles/L, (p<0.001), respectively, while L-citrulline was lower in patients HbS/β^Thal compared to controls but no significantly 21.8±2.5 vs 25.1±2.5 µmoles/L, (p>0.06) and d) carnitine, acetylcarnitine and propionylcarnitine correlated significantly positive with reticulocyte production index (p<0.001).

Our findings showed significant alterations in whole blood metabolome of patients with HbS/β^Thal. Also we demonstrated the very important metabolic abnormality of Nitric Oxide biosynthesis pathway due to the low concentration of the aminoacids serving of substrates in this cycle and disturbances in carnitine and acylcarnitines homeostasis. These abnormalities in the metabolome reflected the hemolysis, inflammatory process and pulmonary hypertension observed in these patients.

No relevant conflicts of interest to declare.