Background: BRAFV600E acts as an ATP-dependent cytosolic kinase. BRAFV600E inhibitors are widely available, but resistance to them is widely reported in the clinic. Lipid metabolism (fatty acids) is fundamental for energy and to control cell stress. Whether and how BRAFV600E impacts lipid metabolism regulation in papillary thyroid carcinoma (PTC) is still unknown. Acetyl-CoA carboxylase (ACC) is a rate-limiting enzyme for de novo lipid synthesis and inhibition of fatty acid oxidation (FAO). ACC1 and ACC2 genes encode distinct isoforms of ACC. The aim of our study was to determine the relationship between BRAFV600E and ACC in PTC. Methods: We performed RNA-seq and DNA copy number analyses in PTC and normal thyroid (NT) in The Cancer Genome Atlas samples. Validations were performed by using assays on PTC-derived cell lines of differing BRAF status and a xenograft mouse model derived from a heterozygous BRAFWT/V600E PTC-derived cell line with knockdown (sh) of ACC1 or ACC2. Results:ACC2 mRNA expression was significantly downregulated in BRAFV600E-PTC vs. BRAFWT-PTC or NT clinical samples. ACC2 protein levels were downregulated in BRAFV600E-PTC cell lines vs. the BRAFWT/WT PTC cell line. Vemurafenib increased ACC2 (and to a lesser extent ACC1) mRNA levels in PTC-derived cell lines in a BRAFV600E allelic dose-dependent manner. BRAFV600E inhibition increased de novo lipid synthesis rates, and decreased FAO due to oxygen consumption rate (OCR), and extracellular acidification rate (ECAR), after addition of palmitate. Only shACC2 significantly increased OCR rates due to FAO, while it decreased ECAR in BRAFV600E PTC-derived cells vs. controls. BRAFV600E inhibition synergized with shACC2 to increase intracellular reactive oxygen species production, leading to increased cell proliferation and, ultimately, vemurafenib resistance. Mice implanted with a BRAFWT/V600E PTC-derived cell line with shACC2 showed significantly increased tumor growth after vemurafenib treatment, while vehicle-treated controls, or shGFP control cells treated with vemurafenib showed stable tumor growth. Conclusions: These findings suggest a potential link between BRAFV600E and lipid metabolism regulation in PTC. BRAFV600E downregulates ACC2 levels, which deregulates de novo lipid synthesis, FAO due to OCR, and ECAR rates. ShACC2 may contribute to vemurafenib resistance and increased tumor growth. ACC2 rescue may represent a novel molecular strategy for overcoming resistance to BRAFV600E inhibitors in refractory PTC.
Publications by Year: 2021
2021
INTRODUCTION: Ubiquinol (reduced coenzyme Q10) is essential for adequate aerobic metabolism. The objective of this trial was to determine whether ubiquinol administration in patients resuscitated from cardiac arrest could increase physiological coenzyme Q10 levels, improve oxygen consumption, and reduce neurological biomarkers of injury.
MATERIALS AND METHODS: This was a randomized, double-blind, placebo-controlled trial in patients successfully resuscitated from cardiac arrest. Patients were randomized to receive enteral ubiquinol (300 mg) or placebo every 12 h for up to 7 days. The primary endpoint was total coenzyme Q10 plasma levels at 24 h after enrollment. Secondary endpoints included neuron specific enolase, S100B, lactate, cellular and global oxygen consumption, neurological status, and in-hospital mortality.
RESULTS: Forty-three patients were included in the modified intention-to-treat analysis. Median coenzyme Q10 levels were significantly higher in the ubiquinol group as compared to the placebo group at 24 h (441 [IQR, 215-510] ηg/mL vs. 113 [IQR, 94-208] ηg/mL, P < 0.001). Similar results were observed at 48 and 72 h. There were no differences between the two groups in any of the secondary endpoints. Median neuron specific enolase levels were not different between the two groups at 24 h (16.8 [IQR, 9.5-19.8] ηg/mL vs. 8.2 [IQR, 4.3-19.1] ηg/mL, P = 0.61).
CONCLUSIONS: Administration of enteral ubiquinol increased plasma coenzyme Q10 levels in post-cardiac arrest patients as compared to placebo. There were no differences in neurological biomarkers and oxygen consumption between the two groups.
UNLABELLED: Lower oxygen consumption is associated with worse survival in septic shock and in other forms of critical illness. No treatment that increases oxygen extraction, a key determinant of oxygen consumption, has been found. Thiamine is required for aerobic metabolism, and deficiency is common in the critically ill.
OBJECTIVES: We evaluated the effect of thiamine on oxygen consumption in patients requiring mechanical ventilation for an acute illness.
DESIGN: Phase II, randomized, double-blind, and placebo-controlled trial.
SETTING AND PARTICIPANTS: ICUs in a tertiary care hospital in the United States. Patients admitted to the ICU and requiring mechanical ventilation were screened for enrollment.
INTERVENTIONS: After enrollment, baseline measurement of oxygen consumption and baseline laboratories including lactate, central venous oxygen saturation, and pyruvate dehydrogenase, a single dose of 200 mg IV thiamine or placebo was administered. Oxygen consumption was then monitored for 6 additional hours and repeat laboratories were drawn at the end of the protocol.
MAIN OUTCOMES AND MEASURES: The primary outcome was the change in oxygen consumption. Analysis was done using linear regression with a first-order autoregressive variance-covariance structure to account for repeated measures within subjects. Secondary outcomes included change in lactate, central venous oxygen saturation, and pyruvate dehydrogenase quantity and activity.
RESULTS: Sixty-seven patients were enrolled. After excluding 11 patients due to inadequate quantity or quality of oxygen consumption data, 56 patients were included. There was no difference in change in oxygen consumption in the 6 hours after study drug. Results for secondary outcomes were similarly negative. In the prespecified subgroup of 18 thiamine deficient patients, there was a difference in the two oxygen consumption curves (p = 0.006), although no difference in median oxygen consumption or area under the curve.
CONCLUSIONS AND RELEVANCE: A single dose of IV thiamine did not alter oxygen consumption in patients requiring mechanical ventilation for acute illness.
UNLABELLED: mRNA vaccines can be developed and produced quickly, making them attractive for immediate outbreak responses. Furthermore, clinical trials have demonstrated rapid protection following mRNA vaccination. We sought to investigate how quickly mRNA vaccines elicit antibody responses compared to other vaccine modalities. We first examined immune kinetics of mRNA and DNA vaccines expressing SARS-CoV-2 spike in mice. We observed rapid induction of antigen-specific binding and neutralizing antibodies by day 5 following mRNA, but not DNA, immunization. The mRNA vaccine also induced increased levels of IL-5, IL-6 and MCP-1. We then evaluated immune kinetics of an HIV-1 mRNA vaccine in comparison to DNA, protein, and rhesus adenovirus 52 (RhAd52) vaccines with the same HIV-1 envelope antigen in mice. Induction of envelope-specific antibodies was observed by day 5 following mRNA vaccination, whereas antibodies were detected by day 7-14 following DNA, protein, and RhAd52 vaccination. Eliciting rapid humoral immunity may be an advantageous property of mRNA vaccines for controlling infectious disease outbreaks.
IMPORTANCE: mRNA vaccines can be developed and produced in record time. Here we demonstrate induction of rapid antibody responses by mRNA vaccines encoding two different viral antigens by day 5 following immunization in mice. The rapid immune kinetics of mRNA vaccines can be an advantageous property that makes them well suited for rapid control of infectious disease outbreaks.