Implications of Standardized Uptake Values of Oral Squamous Cell Carcinoma in PET-CT on Prognosis, Tumor Characteristics and Mitochondrial DNA Heteroplasmy

Under aerobic conditions, some cancers switch to glycolysis to cover their energy requirements. Taking advantage of this process, functional imaging techniques such as PET-CT can be used to detect and assess tumorous tissues. The aim of this study was to investigate standardized uptake values and mitochondrial DNA mutations in oral squamous cell carcinoma. A cohort of 57 patients underwent 18[F]FDG-PET-CT and standardized uptake values were collected. In 15 patients, data on mitochondrial DNA mutations of the tumor were available. Kaplan–Meier curves were calculated, and correlation analyses as well as univariate Cox proportional hazard models were performed. Using ROC analysis to determine a statistical threshold for SUVmax in PET investigations, a cut-off value was determined at 9.765 MB/mL. Survival analysis for SUVmax in these groups showed a Hazard Ratio of 4 (95% CI 1.7–9) in the high SUVmax group with 5-year survival rates of 23.5% (p = 0.00042). For SUVmax and clinicopathological tumor features, significant correlations were found. A tendency towards higher mtDNA heteroplasmy levels in high SUVmax groups could be observed. We were able to confirm the prognostic value of SUVmax in OSCC, showing higher survival rates at lower SUVmax levels. Correlations between SUVmax and distinct tumor characteristics were highly significant, providing evidence that SUVmax may act as a reliable diagnostic parameter. Correlation analysis of mtDNA mutations suggests an influence on metabolic activity in OSCC

Differential Utilization of Dietary Fatty Acids in Benign and Malignant Cells of the Prostate

<div><p>Tumor cells adapt via metabolic reprogramming to meet elevated energy demands due to continuous proliferation, for example by switching to alternative energy sources. Nutrients such as glucose, fatty acids, ketone bodies and amino acids may be utilized as preferred substrates to fulfill increased energy requirements. In this study we investigated the metabolic characteristics of benign and cancer cells of the prostate with respect to their utilization of medium chain (MCTs) and long chain triglycerides (LCTs) under standard and glucose-starved culture conditions by assessing cell viability, glycolytic activity, mitochondrial respiration, the expression of genes encoding key metabolic enzymes as well as mitochondrial mass and mtDNA content. We report that BE prostate cells (RWPE-1) have a higher competence to utilize fatty acids as energy source than PCa cells (LNCaP, ABL, PC3) as shown not only by increased cell viability upon fatty acid supplementation but also by an increased ß-oxidation of fatty acids, although the base-line respiration was 2-fold higher in prostate cancer cells. Moreover, BE RWPE-1 cells were found to compensate for glucose starvation in the presence of fatty acids. Of notice, these findings were confirmed <i>in vivo</i> by showing that PCa tissue has a lower capacity in oxidizing fatty acids than benign prostate. Collectively, these metabolic differences between benign and prostate cancer cells and especially their differential utilization of fatty acids could be exploited to establish novel diagnostic and therapeutic strategies.</p></div

Effects of the ketone body 3-hydroxy butyrate (3-OHB) under glucose starvation.

<p>(A) Overview of cellular ketone body metabolism. Upon starvation, FAs are metabolized to ketone bodies in the liver, representing an important compensatory energy source for the cells. Oxidoreductase 3-hydroxybutyrate dehydrogenase (BDH) mediates the first step of ketone body degradation from 3-hydroxybutyrate into aceto-acetate, which is subsequently converted into two molecules of acetyl-CoA for TCA cycle. (B) Expression of BDH1, BDH2, and AACS was determined in BE (RWPE-1) and PCa cells (LNCaP, ABL, PC3) by qPCR and depicted as mean expression values relative to the housekeeping gene HMBS. (C and D) Effects of 3-OHB on cell viability under glucose starvation. Cells were cultured in 6-well plates in triplicates for 24 h prior to reduction of glucose concentrations to (C) 0.5 g/L or (D) 0.25 g/L in the absence (mock) or presence of 5 mM 3-OHB. Cell viability was assessed after 72 h using WST-1 assay. Values were normalized to vehicle control (mock) under standard growth conditions (1g/L glucose), which were set at 1.0. All results are expressed as mean values (±SEM) of three independent experiments. Significance is indicated (*, P < 0,05; **, P < 0.01; ***, P < 0.001).</p

Differential OXPHOS capacities of BE and PCa cells.

<p>BE RWPE-1 and PCa cells (LNCaP, ABL, PC3) were cultured under standard culture conditions (1g/L glucose) for 48 h. (A) Base-line respiration (ROUTINE state) was measured in intact cells and expressed as O2 flow per cells, IO2 (pmol.s-1.10–6 cells). (B and D) Substrate control factor was assessed in permeabilized cells (B) or tissue (D) and indicates the relative increase of respiration measured after subsequent titration of octanoyl-carnitine, glutamate, pyruvate and succinate in the ADP-stimulated (OXPHOS) state. (C and E) Coupling control ratios were assessed in permeabilized cells (C) and tissues (E), respectively. The L/P ratio provides a degree for coupling efficiency and the P/E ratio embodies the relative limitation of OXPHOS capacity exerted by the phosphorylation system. All results are expressed as mean value (±SEM) of three independent experiments for cultured cell lines (n = 3) and six paired BE and cancer tissue samples (n = 6). Significance is indicated (*, P < 0.05; **, P < 0.01; ***, P < 0.001)</p

BE prostate epithelial cells utilize fatty acids as preferential energy source.

<p>(A) Simplified diagram of fatty acid (FA) catabolism. FAs such as MCTs and LCTs enter the cell via FA protein transporter such as CD36 (FAT, fatty acid translocase) or via direct diffusion, respectively. In the cell, they are converted to fatty acyl CoA by fatty acyl-CoA synthetase (FACS) and further transported into the mitochondria, a step which requires carnitine palmitoyltransferase 1 (CPT1) for the transport of LCTs across the inner mitochondrial membrane. Within the mitochondria, FAs are metabolized through the FA β-oxidation pathway resulting in the production of acetyl-CoA for the TCA cycle. (B) Effects of various FAs (MCTs, LCTs, and MCTs/LCTs) on the viability of BE (RWPE-1) and PCa (LNCaP, ABL, PC3) cells were evaluated by WST-1 assay. Cells were seeded in 96 well plates in triplicates and treated with 200 μM of the indicated FAs or vehicle (mock) for 72 h. Supernatants from the same experiment were subjected to assess (C) glucose consumption and (D) lactate production via enzymatic assays as described in material and methods. All values are normalized to vehicle control (mock), which was set at 1.0. Results are expressed as mean values (±SEM) from three independent experiments. Statistical significance is indicated (*, P < 0.05; **, P < 0.01).</p

Subject characteristics.

<p>Subject characteristics.</p

Composition of different oils employed in this study.

<p>DMSO was used as solvent to add the different oils to culture the media (20 mM stock solutions).</p><p>Composition of different oils employed in this study.</p

Primer sequences for mtDNA copy number assessment.

<p>Primer sequences for mtDNA copy number assessment.</p