Biology subjects

Stephanopoulos, G.

Publications and source records attributed to Stephanopoulos, G..

6 recordsLinked to original sources

bioRxiv2026

Protein S-acylation dynamics provide metabolic plasticity to acute myeloid leukemia cells

Though cancer cells altered metabolism has been recognized for a century, the clinical success of metabolic targeting remains limited due to metabolic plasticity. Here, we use acute myeloid leukemia (AML) as a model to investigate this adaptability through combinatorial metabolic compound screening. Synthetic lethality emerged when AML cells were simultaneously treated with a glutaminase inhibitor and TOFA, a hypolipidemic agent. Sensitivity to this combination was also seen in primary patient samples and in other cancer types, while healthy hematopoietic progenitors were not affected. Unexpectedly, we discovered that TOFA acts through a non-canonical inhibition of protein S-acyltransferases. Protein S-acylation in AML cells specifically requires 16-to-18 carbon long fatty acids and is essential to maintain mitochondrial respiration upon glutaminolysis inhibition. Healthy cells in contrast have high intrinsic metabolic flexibility independent of S-acylation. Our results expose a unique mechanism of metabolic plasticity in cancer that could be targeted to enhance metabolic anti-cancer therapies.

cancer biology↗

bioRxiv2024

NADPH composite index analysis quantifies the relationship between compartmentalized NADPH dynamics and growth rates in cancer cells

NADPH, a highly compartmentalized electron donor in mammalian cells, plays essential roles in cell metabolism. However, little is known about how cytosolic and mitochondrial NADPH dynamics relate to cancer cell growth rates in response to varying nutrient conditions. To address this issue, we present NADPH composite index analysis, which quantifies the relationship between compartmentalized NADPH dynamics and growth rates using genetically encoded NADPH sensors, automated image analysis pipeline, and correlation analysis. Through this analysis, we demonstrated that compartmentalized NADPH dynamics patterns were cancer cell-type dependent. Specifically, cytosolic and mitochondrial NADPH dynamics of MDA-MB-231 decreased in response to serine deprivation, while those of HCT-116 increased in response to serine or glutamine deprivation. Furthermore, by introducing a fractional contribution parameter, we correlated cytosolic and mitochondrial NADPH dynamics to growth rates. Using this parameter, we identified cancer cell lines whose growth rates were selectively inhibited by targeting cytosolic or mitochondrial NADPH metabolism. Mechanistically, we identified citrate transporter as a key mitochondrial transporter that maintains compartmentalized NADPH dynamics and growth rates. Altogether, our results present a significant advance in quantifying the relationship between compartmentalized NADPH dynamics and cancer cell growth rates, highlighting a potential of targeting compartmentalized NADPH metabolism for selective cancer cell growth inhibitions.

systems biology↗

bioRxiv2023

Drug screening in human physiologic medium identifies uric acid as an inhibitor of rigosertib efficacy

The non-physiological nutrient levels found in traditional culture media have been shown to affect numerous aspects of cancer cell physiology, including how cells respond to certain therapeutic agents. Here, we comprehensively evaluated how physiological nutrient levels impact therapeutic response by performing drug screening in human plasma-like medium (HPLM). We observed dramatic nutrient-dependent changes in sensitivity to a variety of FDA-approved and clinically trialed compounds, including rigosertib, an experimental cancer therapeutic that has recently failed in phase 3 clinical trials. Mechanistically, we found that the ability of rigosertib to destabilize microtubules is strongly inhibited by the purine metabolism waste product uric acid, which is uniquely abundant in humans relative to traditional in vitro and in vivo cancer models. Structural modelling studies suggest that uric acid interacts with the tubulin-rigosertib complex and may act as an uncompetitive inhibitor of rigosertib. These results offer a possible explanation for the failure of rigosertib in clinical trials and demonstrate the utility of physiological media to achieve in vitro results that better represent human therapeutic responses.

cancer biology↗

bioRxiv2022

Non-targeted metabolomics identifies erythronate accumulation in cancer cells

Using a non-targeted isotope-assisted metabolomics approach, we identified erythronate as a metabolite that accumulates in several human cancer cell lines. Erythronate has been reported to be a detoxification product derived from off-target glycolytic metabolism. We provide data supporting a possible alternative route to erythronate production involving the dephosphorylation of the pentose phosphate pathway intermediate erythrose-4-phosphate to form erythrose, followed by the oxidation of erythrose by an aldehyde dehydrogenase. Finally, we detected increased erythronate concentrations in tumors relative to adjacent normal tissues from lung cancer patients. These findings suggest the accumulation of erythronate to be an example of metabolic reprogramming in cancer cells, raising the possibility that elevated level of erythronate may serve as a biomarker of certain types of cancer.

biochemistry↗

bioRxiv2022

Oncogenic metabolic rewiring independent of proliferative control in human mammary epithelial cells

The use of isotopic tracers and metabolic flux analysis (MFA) has unveiled a number of metabolic pathways differentially activated in cancer cells. To support efforts to design effective metabolic therapies for cancer, we sought to distinguish metabolic behavior in cancer versus normal cells growing at the same rate. To this end, we performed 13C-isotope tracing and MFA in human mammary epithelial cells (HMECs) harboring different combinations of oncogenes. By introducing a new quantity termed metabolic flux intensity, defined as pathway flux divided by specific growth rate, we showed that metabolism is dually controlled by proliferation and oncogenotypes. 13C-MFA further revealed that oxidative pentose phosphate pathway (oxPPP), malate dehydrogenase (MDH) and isocitrate dehydrogenase (IDH) were most enhanced in cancerous HMECs. Drug targeting of these pathways selectively reduced growth in the tumorigenic HMEC line. Our study provides direct evidence that metabolism of cancer cells is different than that of normal proliferating cells.

biochemistry↗

bioRxiv2020

Constructing an ethanol utilization pathway in Escherichia coli to produce acetyl-CoA derived compounds

Engineering microbes to utilize non-conventional substrates could create short and efficient pathways to convert substrate into product. In this study, we designed and constructed a two-step heterologous ethanol utilization pathway (EUP) in Escherichia coli by using acetaldehyde dehydrogenase (encoded by ada) from Dickeya zeae and alcohol dehydrogenase (encoded by adh2) from Saccharomyces cerevisiae. This EUP can convert ethanol into acetyl-CoA without ATP consumption, and generate two molecules of NADH per molecule of ethanol. We optimized the expression of these two genes and found that ethanol consumption could be improved by expressing them in a specific order (ada-adh2) with a constitutive promoter (PgyrA). The engineered E. coli strain with EUP consumed approximately 8 g/L of ethanol in 96 hours when it was used as sole carbon source. Subsequently, we combined EUP with the biosynthesis of polyhydroxybutyrate (PHB), a biodegradable polymer derived from acetyl-CoA. The engineered E. coli strain carrying EUP and PHB biosynthetic pathway produced 1.1 g/L of PHB from 10 g/L of ethanol and 1 g/L of aspartate family amino acids in 96 hours. We also engineered E. coli strain to produced 24 mg/L of prenol from 10 g/L of ethanol in 48 hours, supporting the feasibility of converting ethanol into different classes of acetyl-CoA derived compounds. HighlightsO_LIEngineered Escherichia coli strains to grow on ethanol as sole carbon source C_LIO_LIDemonstrated that ethanol was converted into acetyl-CoA (AcCoA) through two pathways (acetaldehyde-acetate-AcCoA and acetaldehyde-AcCoA) C_LIO_LIConverted ethanol into two acetyl-CoA derived products with low structural similarity (polyhydroxybutyrate and prenol) C_LIO_LIDiscovered that supplementation of the aspartate family amino acids can substantially improve cell growth on ethanol C_LI

bioengineering↗