bims-mibica 2024-11-03 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2024–11–03
24 papers selected by
Kelsey Fisher-Wellman, Wake Forest University

Noncanonical TCA cycle fosters canonical TCA cycle and mitochondrial integrity in acute myeloid leukemia.
Pathogenic mitochondrial DNA mutations inhibit melanoma metastasis.
Glucose-dependent glycosphingolipid biosynthesis fuels CD8+ T cell function and tumor control.
High mitochondrial DNA levels accelerate lung adenocarcinoma progression.
Structure of the turnover-ready state of an ancestral respiratory complex I.
Assays to monitor mitochondrial translation.
Membrane potential stimulates ADP import and ATP export by the mitochondrial ADP/ATP carrier due to its positively charged binding site.
Site of breast cancer metastasis is independent of single nutrient levels.
Engineering mtDNA Deletions by Reconstituting End-Joining in Human Mitochondria.
Methylmalonic acid induces metabolic abnormalities and exhaustion in CD8+ T cells to suppress anti-tumor immunity.
CYP19A1 regulates chemoresistance in colorectal cancer through modulation of estrogen biosynthesis and mitochondrial function.
The Integrated Stress Response Suppresses PINK1-dependent Mitophagy by Preserving Mitochondrial Import Efficiency.
Copper drives remodeling of metabolic state and progression of clear cell renal cell carcinoma.
A hierarchical hepatic de novo lipogenesis substrate supply network utilizing pyruvate, acetate, and ketones.
Venetoclax Dose Escalation Rapidly Activates a BAFF/BCL-2 Survival Axis in Chronic Lymphocytic Leukemia.
Targeting Oxidative Phosphorylation with a Novel Thiophene Carboxamide Increases the Efficacy of Imatinib against Leukemic Stem Cells in Chronic Myeloid Leukemia.
Inhibition of OXPHOS induces metabolic rewiring and reduces hypoxia in murine tumor models.
TFAM as a sensor of UVC-induced mitochondrial DNA damage.
Quantitation of metabolic activity from isotope tracing data using automated methodology.
Enhancing Radiation-induced Reactive Oxygen Species Generation Through Mitochondrial Transplantation in Human Glioblastoma.
Metabolic regulation of the glioblastoma stem cell epitranscriptome by malate dehydrogenase 2.
RAS-mutant leukaemia stem cells drive clinical resistance to venetoclax.
Oxidative phosphorylation related gene COA6 is a novel indicator for the prognosis and immune response in lung adenocarcinoma.
Aberrant fragmentomic features of circulating cell-free mitochondrial DNA as novel biomarkers for multi-cancer detection.

Cancer Sci. 2024 Oct 31.

Noncanonical TCA cycle fosters canonical TCA cycle and mitochondrial integrity in acute myeloid leukemia.

Atsushi Watanabe, Chartsiam Tipgomut, Haruhito Totani, Kentaro Yoshimura, Tomohiko Iwano, Hamed Bashiri, Lee Hui Chua, Chong Yang, Toshio Suda.

  Cancer cells rely on mitochondrial oxidative phosphorylation (OXPHOS) and the noncanonical tricarboxylic acid (TCA) cycle. In this paper, we shed light on the vital role played by the noncanonical TCA cycle in a host-side concession to mitochondria, especially in highly energy-demanding malignant tumor cells. Inhibition of ATP-citrate lyase (ACLY), a key enzyme in the noncanonical TCA cycle, induced apoptosis by increasing reactive oxygen species levels and DNA damage while reducing mitochondrial membrane potential. The mitochondrial membrane citrate transporter inhibitor, CTPI2, synergistically enhanced these effects. ACLY inhibition reduced cytosolic citrate levels and CTPI2 lowered ACLY activity, suggesting that the noncanonical TCA cycle is sustained by a positive feedback mechanism. These inhibitions impaired ATP production, particularly through OXPHOS. Metabolomic analysis of mitochondrial and cytosolic fractions revealed reduced levels of glutathione pathway-related and TCA cycle-related metabolite, except fumarate, in mitochondria following noncanonical TCA cycle inhibition. Despite the efficient energy supply to the cell by mitochondria, this symbiosis poses challenges related to reactive oxygen species and mitochondrial maintenance. In conclusion, the noncanonical TCA cycle is indispensable for the canonical TCA cycle and mitochondrial integrity, contributing to mitochondrial domestication.

Keywords:  ATP‐citrate lyase; antimetabolites; apoptosis; cancer metabolism; cell lines; hematopoietic organ; mitochondria; noncanonical TCA cycle; others; reactive oxygen species

DOI:  https://doi.org/10.1111/cas.16347
Sci Adv. 2024 Nov;10(44): eadk8801

Pathogenic mitochondrial DNA mutations inhibit melanoma metastasis.

Spencer D Shelton, Sara House, Luiza Martins Nascentes Melo, Vijayashree Ramesh, Zhenkang Chen, Tao Wei, Xun Wang, Claire B Llamas, Siva Sai Krishna Venigalla, Cameron J Menezes, Gabriele Allies, Jonathan Krystkiewicz, Jonas Rösler, Sven W Meckelmann, Peihua Zhao, Florian Rambow, Dirk Schadendorf, Zhiyu Zhao, Jennifer G Gill, Ralph J DeBerardinis, Sean J Morrison, Alpaslan Tasdogan, Prashant Mishra.

Mitochondrial DNA (mtDNA) mutations are frequent in cancer, yet their precise role in cancer progression remains debated. To functionally evaluate the impact of mtDNA variants on tumor growth and metastasis, we developed an enhanced cytoplasmic hybrid (cybrid) generation protocol and established isogenic human melanoma cybrid lines with wild-type mtDNA or pathogenic mtDNA mutations with partial or complete loss of mitochondrial oxidative function. Cybrids with homoplasmic levels of pathogenic mtDNA reliably established tumors despite dysfunctional oxidative phosphorylation. However, these mtDNA variants disrupted spontaneous metastasis from primary tumors and reduced the abundance of circulating tumor cells. Migration and invasion of tumor cells were reduced, indicating that entry into circulation is a bottleneck for metastasis amid mtDNA dysfunction. Pathogenic mtDNA did not inhibit organ colonization following intravenous injection. In heteroplasmic cybrid tumors, single-cell analyses revealed selection against pathogenic mtDNA during melanoma growth. Collectively, these findings experimentally demonstrate that functional mtDNA is favored during melanoma growth and supports metastatic entry into the blood.

DOI: https://doi.org/10.1126/sciadv.adk8801
bioRxiv. 2024 Oct 14. pii: 2024.10.10.617261. [Epub ahead of print]

Glucose-dependent glycosphingolipid biosynthesis fuels CD8+ T cell function and tumor control.

Joseph Longo, Lisa M DeCamp, Brandon M Oswald, Robert Teis, Alfredo Reyes-Oliveras, Michael S Dahabieh, Abigail E Ellis, Michael P Vincent, Hannah Damico, Kristin L Gallik, Shelby E Compton, Colt D Capan, Kelsey S Williams, Corinne R Esquibel, Zachary B Madaj, Hyoungjoo Lee, Dominic G Roy, Connie M Krawczyk, Brian B Haab, Ryan D Sheldon, Russell G Jones.

  Glucose is essential for T cell proliferation and function, yet its specific metabolic roles in vivo remain poorly defined. Here, we identify glycosphingolipid (GSL) biosynthesis as a key pathway fueled by glucose that enables CD8+ T cell expansion and cytotoxic function in vivo. Using 13C-based stable isotope tracing, we demonstrate that CD8+ effector T cells use glucose to synthesize uridine diphosphate-glucose (UDP-Glc), a precursor for glycogen, glycan, and GSL biosynthesis. Inhibiting GSL production by targeting the enzymes UGP2 or UGCG impairs CD8+ T cell expansion and cytolytic activity without affecting glucose-dependent energy production. Mechanistically, we show that glucose-dependent GSL biosynthesis is required for plasma membrane lipid raft integrity and aggregation following TCR stimulation. Moreover, UGCG-deficient CD8+ T cells display reduced granzyme expression and tumor control in vivo. Together, our data establish GSL biosynthesis as a critical metabolic fate of glucose-independent of energy production-required for CD8+ T cell responses in vivo.

Keywords:  CD8+ T cells; UGCG; cytotoxic function; glucose; glycosphingolipids; immunometabolism; lipid rafts; lipidomics; metabolomics; nucleotide sugar metabolism

DOI:  https://doi.org/10.1101/2024.10.10.617261
Sci Adv. 2024 Nov;10(44): eadp3481

High mitochondrial DNA levels accelerate lung adenocarcinoma progression.

Mara Mennuni, Stephen E Wilkie, Pauline Michon, David Alsina, Roberta Filograna, Markus Lindberg, David E Sanin, Florian Rosenberger, Alina Schaaf, Erik Larsson, Erika L Pearce, Nils-Göran Larsson.

Lung adenocarcinoma is a common aggressive cancer and a leading cause of mortality worldwide. Here, we report an important in vivo role for mitochondrial DNA (mtDNA) copy number during lung adenocarcinoma progression in the mouse. We found that lung tumors induced by KRASG12D expression have increased mtDNA levels and enhanced mitochondrial respiration. To experimentally assess a possible causative role in tumor progression, we induced lung cancer in transgenic mice with a general increase in mtDNA copy number and found that they developed a larger tumor burden, whereas mtDNA depletion in tumor cells reduced tumor growth. Immune cell populations in the lung and cytokine levels in plasma were not affected by increased mtDNA levels. Analyses of large cancer databases indicate that mtDNA copy number is also important in human lung cancer. Our study thus reports experimental evidence for a tumor-intrinsic causative role for mtDNA in lung cancer progression, which could be exploited for development of future cancer therapies.

DOI: https://doi.org/10.1126/sciadv.adp3481
Nat Commun. 2024 Oct 29. 15(1): 9340

Structure of the turnover-ready state of an ancestral respiratory complex I.

Bozhidar S Ivanov, Hannah R Bridges, Owen D Jarman, Judy Hirst.

Respiratory complex I is pivotal for cellular energy conversion, harnessing energy from NADH:ubiquinone oxidoreduction to drive protons across energy-transducing membranes for ATP synthesis. Despite detailed structural information on complex I, its mechanism of catalysis remains elusive due to lack of accompanying functional data for comprehensive structure-function analyses. Here, we present the 2.3-Å resolution structure of complex I from the α-proteobacterium Paracoccus denitrificans, a close relative of the mitochondrial progenitor, in phospholipid-bilayer nanodiscs. Three eukaryotic-type supernumerary subunits (NDUFS4, NDUFS6 and NDUFA12) plus a novel L-isoaspartyl-O-methyltransferase are bound to the core complex. Importantly, the enzyme is in a single, homogeneous resting state that matches the closed, turnover-ready (active) state of mammalian complex I. Our structure reveals the elements that stabilise the closed state and completes P. denitrificans complex I as a unified platform for combining structure, function and genetics in mechanistic studies.

DOI: https://doi.org/10.1038/s41467-024-53679-3
Methods Enzymol. 2024 ;pii: S0076-6879(24)00385-9. [Epub ahead of print]706 519-532

Assays to monitor mitochondrial translation.

Sung-Jun Jung, Martin Ott.

  The complexes of the oxidative phosphorylation (OXPHOS) system found in the mitochondrial inner membrane comprises nuclear and mitochondrial-encoded proteins. The mitochondrial-encoded subunits of the OXPHOS complexes play vital catalytic roles for OXPHOS. These subunits are inserted co-translationally into the inner membrane, where they are matured and assembled with nuclear encoded subunits, requiring a set of OXPHOS assembly and quality control factors. Hence, monitoring the fate of newly synthesized mitochondrial-encoded polypeptides is a basic and essential approach for exploring OXPHOS biogenesis and the related protein quality control processes. Here, we describe a detailed protocol for labeling mitochondrial encoded proteins with 35S-methionine for pulse and pulse/chase experiments, both in vivo and in organello, using the yeast Saccharomyces cerevisiae as the model. These methods enable analyses of the early steps during the biogenesis and turnover of mitochondrial-encoded proteins.

Keywords:  35S-methionine; Mitochondrial translation; isolated mitochondria; protein stability; protein synthesis; yeast

DOI:  https://doi.org/10.1016/bs.mie.2024.07.045
Sci Adv. 2024 Nov;10(44): eadp7725

Membrane potential stimulates ADP import and ATP export by the mitochondrial ADP/ATP carrier due to its positively charged binding site.

Vasiliki Mavridou, Martin S King, Andre Bazzone, Roger Springett, Edmund R S Kunji.

The mitochondrial adenosine 5'-diphosphate (ADP)/adenosine 5'-triphosphate (ATP) carrier imports ADP into the mitochondrion and exports ATP to the cell. Here, we demonstrate that 3.3 positive charges are translocated with the negatively charged substrate in each transport step. They can be assigned to three positively charged residues of the central substrate-binding site and two asparagine/arginine pairs. In this way, the membrane potential stimulates not only the ATP4- export step, as a net -0.7 charge is transported, but also the ADP3- import step, as a net +0.3 charge is transported with the electric field. These positive charge movements also inhibit the import of ATP and export of ADP in the presence of a membrane potential, allowing these nucleotides to be maintained at high concentrations in the cytosol and mitochondrial matrix to drive the hydrolysis and synthesis of ATP, respectively. Thus, this is the mechanism by which the membrane potential drives adenine nucleotide exchange with high directional fluxes to fuel the cellular processes.

DOI: https://doi.org/10.1126/sciadv.adp7725
bioRxiv. 2024 Oct 25. pii: 2024.10.24.616714. [Epub ahead of print]

Site of breast cancer metastasis is independent of single nutrient levels.

Keene L Abbott, Sonu Subudhi, Raphael Ferreira, Yetiş Gültekin, Sophie C Steinbuch, Muhammad Bin Munim, Sophie E Honeder, Ashwin S Kumar, Diya L Ramesh, Michelle Wu, Jacob A Hansen, Sharanya Sivanand, Lisa M Riedmayr, Mark Duquette, Ahmed Ali, Nicole Henning, Anna Shevzov-Zebrun, Florian Gourgue, Anna M Barbeau, Millenia Waite, Tenzin Kunchok, Gino B Ferraro, Brian T Do, Virginia Spanoudaki, Francisco J Sánchez-Rivera, Xin Jin, George M Church, Rakesh K Jain, Matthew G Vander Heiden.

Cancer metastasis is a major contributor to patient morbidity and mortality 1 , yet the factors that determine the organs where cancers can metastasize are incompletely understood. In this study, we quantify the absolute levels of over 100 nutrients available across multiple tissues in mice and investigate how this relates to the ability of breast cancer cells to grow in different organs. We engineered breast cancer cells with broad metastatic potential to be auxotrophic for specific nutrients and assessed their ability to colonize different organs. We then asked how tumor growth in different tissues relates to nutrient availability and tumor biosynthetic activity. We find that single nutrients alone do not define the sites where breast cancer cells can grow as metastases. Additionally, we identify purine synthesis as a requirement for tumor growth and metastasis across many tissues and find that this phenotype is independent of tissue nucleotide availability or tumor de novo nucleotide synthesis activity. These data suggest that a complex interplay of multiple nutrients within the microenvironment dictates potential sites of metastatic cancer growth, and highlights the interdependence between extrinsic environmental factors and intrinsic cellular properties in influencing where breast cancer cells can grow as metastases.

DOI: https://doi.org/10.1101/2024.10.24.616714
bioRxiv. 2024 Oct 17. pii: 2024.10.15.618543. [Epub ahead of print]

Engineering mtDNA Deletions by Reconstituting End-Joining in Human Mitochondria.

Yi Fu, Max Land, Ruobing Cui, Tamar Kavlashvili, Minsoo Kim, Toby Lieber, Keun Woo Ryu, Emily DeBitetto, Ignas Masilionis, Rahul Saha, Meril Takizawa, Daphne Baker, Marco Tigano, Ed Reznik, Roshan Sharma, Ronan Chaligne, Craig B Thompson, Dana Pe'er, Agnel Sfeir.

Recent breakthroughs in the genetic manipulation of mitochondrial DNA (mtDNA) have enabled the precise introduction of base substitutions and the effective removal of genomes carrying harmful mutations. However, the reconstitution of mtDNA deletions responsible for severe mitochondrial myopathies and age-related diseases has not yet been achieved in human cells. Here, we developed a method to engineer specific mtDNA deletions in human cells by co-expressing end-joining (EJ) machinery and targeted endonucleases. As a proof-of-concept, we used mito-EJ and mito-ScaI to generate a panel of clonal cell lines harboring a ∼3.5 kb mtDNA deletion with the full spectrum of heteroplasmy. Investigating these isogenic cells revealed a critical threshold of ∼75% deleted genomes, beyond which cells exhibited depletion of OXPHOS proteins, severe metabolic disruption, and impaired growth in galactose-containing media. Single-cell multiomic analysis revealed two distinct patterns of nuclear gene deregulation in response to mtDNA deletion accumulation; one triggered at the deletion threshold and another progressively responding to increasing heteroplasmy. In summary, the co-expression of mito-EJ and programable nucleases provides a powerful tool to model disease-associated mtDNA deletions in different cell types. Establishing a panel of cell lines with a large-scale deletion at varying levels of heteroplasmy is a valuable resource for understanding the impact of mtDNA deletions on diseases and guiding the development of potential therapeutic strategies.
Highlights: Combining prokaryotic end-joining with targeted endonucleases generates specific mtDNA deletions in human cellsEngineering a panel of cell lines with a large-scale deletion that spans the full spectrum of heteroplasmy75% heteroplasmy is the threshold that triggers mitochondrial and cellular dysfunctionTwo distinct nuclear transcriptional programs in response to mtDNA deletions: threshold-triggered and heteroplasmy-sensing.

DOI: https://doi.org/10.1101/2024.10.15.618543
Oncogene. 2024 Oct 29.

Methylmalonic acid induces metabolic abnormalities and exhaustion in CD8+ T cells to suppress anti-tumor immunity.

Joanne D Tejero, Rebecca S Hesterberg, Stanislav Drapela, Didem Ilter, Devesh Raizada, Felicia Lazure, Hossein Kashfi, Min Liu, Leonardo Silvane, Dorina Avram, Juan Fernández-García, John M Asara, Sarah-Maria Fendt, John L Cleveland, Ana P Gomes.

Systemic levels of methylmalonic acid (MMA), a byproduct of propionate metabolism, increase with age and MMA promotes tumor progression via its direct effects in tumor cells. However, the role of MMA in modulating the tumor ecosystem remains to be investigated. The proliferation and function of CD8+ T cells, key anti-tumor immune cells, declines with age and in conditions of vitamin B12 deficiency, which are the two most well-established conditions that lead to increased systemic levels of MMA. Thus, we hypothesized that increased circulatory levels of MMA would lead to a suppression of CD8+ T cell immunity. Treatment of primary CD8+ T cells with MMA induced a dysfunctional phenotype characterized by robust immunosuppressive transcriptional reprogramming and marked increases in the expression of the exhaustion regulator, TOX. Accordingly, MMA treatment upregulated exhaustion markers in CD8+ T cells and decreased their effector functions, which drove the suppression of anti-tumor immunity in vitro and in vivo. Mechanistically, MMA-induced CD8+ T cell exhaustion was associated with a suppression of NADH-regenerating reactions in the TCA cycle and concomitant defects in mitochondrial function. Thus, MMA has immunomodulatory roles, thereby highlighting MMA as an important link between aging, immune dysfunction, and cancer.

DOI: https://doi.org/10.1038/s41388-024-03191-1
Cancer Metab. 2024 Oct 28. 12(1): 33

CYP19A1 regulates chemoresistance in colorectal cancer through modulation of estrogen biosynthesis and mitochondrial function.

Yang Wang, Qiang Ji, Ning Cao, Guijie Ge, Xiaomin Li, Xiangdong Liu, Yanqi Mi.

  Chemoresistance remains a major challenge in the effective treatment of colorectal cancer (CRC), contributing to poor patient outcomes. While the molecular mechanisms underlying chemoresistance are complex and multifaceted, emerging evidence suggests that altered mitochondrial function and hormone signaling play crucial roles. In this study, we investigated the role of CYP19A1, a key enzyme in estrogen biosynthesis, in regulating chemoresistance in CRC. Using a combination of in vitro functional assays, transcriptomic analysis, and clinical data mining, we demonstrate that CYP19A1 expression is significantly upregulated in CRC cells and patient-derived samples compared to normal controls. Mechanistically, we found that CYP19A1 regulates chemoresistance through modulation of mitochondrial function and complex I activity, which is mediated by CYP19A1-dependent estrogen biosynthesis. Notably, targeted inhibition of CYP19A1 and complex I using specific inhibitors effectively reversed the chemoresistance of CRC cells to chemotherapeutic drugs. Moreover, analysis of the TCGA CRC dataset revealed that high CYP19A1 expression correlates with poor overall survival in chemotherapy-treated patients. Taken together, our findings uncover a novel role for CYP19A1 in regulating chemoresistance in CRC through modulation of mitochondrial function and estrogen signaling, and highlight the potential of targeting the CYP19A1/estrogen/complex I axis as a therapeutic strategy to overcome chemoresistance and improve patient outcomes.

Keywords:  CYP19A1; Chemotherapy resistance; Colorectal cancer; Mitochondria; Mitochondrial respiratory chain complexes

DOI:  https://doi.org/10.1186/s40170-024-00360-4
bioRxiv. 2024 Oct 17. pii: 2024.10.16.617214. [Epub ahead of print]

The Integrated Stress Response Suppresses PINK1-dependent Mitophagy by Preserving Mitochondrial Import Efficiency.

Mingchong Yang, Zengshuo Mo, Kelly Walsh, Wen Liu, Xiaoyan Guo.

Mitophagy is crucial for maintaining mitochondrial health, but how its levels adjust to different stress conditions remains unclear. In this study, we investigated the role of the DELE1-HRI axis of integrated stress response (ISR) in regulating mitophagy, a key mitochondrial stress pathway. Our findings show that the ISR suppresses mitophagy under non-depolarizing mitochondrial stress by positively regulating mitochondrial protein import, independent of ATF4 activation. Mitochondrial protein import is regulated by the rate of protein synthesis under both depolarizing and non-depolarizing stress. Without ISR, increased protein synthesis overwhelms the mitochondrial import machinery, reducing its efficiency. Under depolarizing stress, mitochondrial import is heavily impaired even with active ISR, leading to significant PINK1 accumulation. In contrast, non-depolarizing stress allows more efficient protein import in the presence of ISR, resulting in lower mitophagy. Without ISR, mitochondrial protein import becomes severely compromised, causing PINK1 accumulation to reach the threshold necessary to trigger mitophagy. These findings reveal a novel link between ISR-regulated protein synthesis, mitochondrial import, and mitophagy, offering potential therapeutic targets for diseases associated with mitochondrial dysfunction.

DOI: https://doi.org/10.1101/2024.10.16.617214
Cancer Discov. 2024 Oct 31.

Copper drives remodeling of metabolic state and progression of clear cell renal cell carcinoma.

Megan E Bischoff, Behrouz Shamsaei, Juechen Yang, Dina Secic, Bhargav Vemuri, Julie A Reisz, Angelo D'Alessandro, Caterina Bartolacci, Rafal Adamczak, Lucas Schmidt, Jiang Wang, Amelia Martines, Jahnavi Venkat, Vanina Toffessi Tcheuyap, Jacek Biesiada, Catherine A Behrmann, Katherine E Vest, James Brugarolas, Pier Paolo Scaglioni, David R Plas, Krushna C Patra, Shuchi Gulati, Julio A Landero Figueroa, Jarek Meller, John T Cunningham, Maria F Czyzyk-Krzeska.

Copper (Cu) is a cofactor of cytochrome c oxidase (CuCOX), indispensable for aerobic mitochondrial respiration. This study reveals that advanced clear cell renal cell carcinomas (ccRCCs) accumulate Cu, allocating it to CuCOX. Using a range of orthogonal approaches, including metabolomics, lipidomics, isotope-labeled glucose and glutamine flux analysis, and transcriptomics across tumor samples, cell lines, xenografts, and PDX models, combined with genetic and pharmacological interventions, we explored Cu's role in ccRCC. Elevated Cu levels stimulate CuCOX biogenesis, providing bioenergetic and biosynthetic benefits that promote tumor growth. This effect is complemented by glucose-dependent glutathione production, which facilitates detoxification and mitigates Cu-H2O2 toxicity. Single-cell RNA sequencing (scRNA-seq) and spatial transcriptomics reveal increased oxidative metabolism, altered glutathione and Cu metabolism, and diminished HIF activity during ccRCC progression. Thus, Cu drives an integrated oncogenic remodeling of bioenergetics, biosynthesis, and redox homeostasis, fueling ccRCC growth, which can be targeted for new therapeutic approaches.

DOI: https://doi.org/10.1158/2159-8290.CD-24-0187
Cell Metab. 2024 Oct 25. pii: S1550-4131(24)00409-1. [Epub ahead of print]

A hierarchical hepatic de novo lipogenesis substrate supply network utilizing pyruvate, acetate, and ketones.

Adam J Rauckhorst, Ryan D Sheldon, Daniel J Pape, Adnan Ahmed, Kelly C Falls-Hubert, Ronald A Merrill, Reid F Brown, Kshitij Deshmukh, Thomas A Vallim, Stanislaw Deja, Shawn C Burgess, Eric B Taylor.

  Hepatic de novo lipogenesis (DNL) is a fundamental physiologic process that is often pathogenically elevated in metabolic disease. Treatment is limited by incomplete understanding of the metabolic pathways supplying cytosolic acetyl-CoA, the obligate precursor to DNL, including their interactions and proportional contributions. Here, we combined extensive 13C tracing with liver-specific knockout of key mitochondrial and cytosolic proteins mediating cytosolic acetyl-CoA production. We show that the mitochondrial pyruvate carrier (MPC) and ATP-citrate lyase (ACLY) gate the major hepatic lipogenic acetyl-CoA production pathway, operating in parallel with acetyl-CoA synthetase 2 (ACSS2). Given persistent DNL after mitochondrial citrate carrier (CiC) and ACSS2 double knockout, we tested the contribution of exogenous and leucine-derived acetoacetate to acetoacetyl-CoA synthetase (AACS)-dependent DNL. CiC knockout increased acetoacetate-supplied hepatic acetyl-CoA production and DNL, indicating that ketones function as mitochondrial-citrate reciprocal DNL precursors. By delineating a mitochondrial-cytosolic DNL substrate supply network, these findings may inform strategies to therapeutically modulate DNL.

Keywords:  AACS; ACLY; ACSS2; ATP-citrate lyase; CiC; DNL; MPC; acetoacetyl-CoA synthetase; acetyl-CoA synthetase 2; de novo lipogenesis; liver; metabolomics; mitochondrial citrate carrier; mitochondrial pyruvate carrier; stable isotope tracers

DOI:  https://doi.org/10.1016/j.cmet.2024.10.013
Blood. 2024 Oct 29. pii: blood.2024024341. [Epub ahead of print]

Venetoclax Dose Escalation Rapidly Activates a BAFF/BCL-2 Survival Axis in Chronic Lymphocytic Leukemia.

Meng-Xiao Luo, Tania Tan, Marie Trussart, Annika Poch, Minh-Hanh Thi Nguyen, Terence P Speed, Damien G Hicks, Esther Bandala-Sanchez, Hongke Peng, Stephane Chappaz, Charlotte Slade, Daniel T Utzschneider, Rachel M Koldej, David Ritchie, Andreas Strasser, Rachel Thijssen, Matthew E Ritchie, Constantine Si Lun Tam, Geoffrey J Lindeman, David Ching Siang Huang, Thomas E Lew, Mary Ann Anderson, Andrew W Roberts, Charis E Teh, Daniel H D Gray.

Venetoclax, a first-in-class BH3 mimetic drug targeting BCL-2, has improved outcomes for patients with chronic lymphocytic leukemia (CLL). Early measurements of the depth of the venetoclax treatment response, assessed by minimal residual disease, are strong predictors of long-term clinical outcomes. Yet, there are limited data concerning the early changes induced by venetoclax treatment that might inform strategies to improve responses. To address this gap, we conducted longitudinal mass cytometric profiling of blood cells from patients with CLL during the first five weeks of venetoclax monotherapy. At baseline, we resolved CLL heterogeneity at the single-cell level to define multiple subpopulations in all patients distinguished by proliferative, metabolic and cell survival proteins. Venetoclax induced significant reduction in all CLL subpopulations coincident with rapid upregulation of pro-survival BCL-2, BCL-XL and MCL-1 proteins in surviving cells, which had reduced sensitivity to the drug. Mouse models recapitulated the venetoclax-induced elevation of survival proteins in B cells and CLL-like cells that persisted in vivo, with genetic models demonstrating that extensive apoptosis and access to the B cell cytokine, BAFF, were essential. Accordingly, analysis of patients with CLL that were treated with venetoclax or the anti-CD20 antibody obinutuzumab exhibited marked elevation of BAFF and increased pro-survival proteins in leukemic cells that persisted. Overall, these data highlight the rapid adaptation of CLL cells to targeted therapies via homeostatic factors and support co-targeting of cytokine signals to achieve deeper and more durable long-term responses.

DOI: https://doi.org/10.1182/blood.2024024341
Int J Mol Sci. 2024 Oct 15. pii: 11093. [Epub ahead of print]25(20):

Targeting Oxidative Phosphorylation with a Novel Thiophene Carboxamide Increases the Efficacy of Imatinib against Leukemic Stem Cells in Chronic Myeloid Leukemia.

Kana Kusaba, Tatsuro Watanabe, Keisuke Kidoguchi, Yuta Yamamoto, Ayaka Tomoda, Toshimi Hoshiko, Naoto Kojima, Susumu Nakata, Shinya Kimura.

  Patients with chronic myeloid leukemia (CML) respond to tyrosine kinase inhibitors (TKIs); however, CML leukemic stem cells (LSCs) exhibit BCR::ABL kinase-independent growth and are insensitive to TKIs, leading to disease relapse. To prevent this, new therapies targeting CML-LSCs are needed. Rates of mitochondria-mediated oxidative phosphorylation (OXPHOS) in CD34+CML cells within the primitive CML cell population are higher than those in normal undifferentiated hematopoietic cells; therefore, the inhibition of OXPHOS in CML-LSCs may be a potential cure for CML. NK-128 (C33H61NO5S) is a structurally simplified analog of JCI-20679, the design of which was based on annonaceous acetogenins. NK-128 exhibits antitumor activity against glioblastoma and human colon cancer cells by inhibiting OXPHOS and activating AMP-activated protein kinase (AMPK). Here, we demonstrate that NK-128 effectively suppresses the growth of CML cell lines and that the combination of imatinib and NK-128 is more potent than either alone in a CML xenograft mouse model. We also found that NK-128 inhibits colony formation by CD34+ CML cells isolated from the bone marrow of untreated CML patients. Taken together, these findings suggest that targeting OXPHOS is a beneficial approach to eliminating CML-LSCs, and may improve the treatment of CML.

Keywords:  chronic myeloid leukemia; oxidative phosphorylation; stem cell; thiophene carboxamide; tyrosine kinase

DOI:  https://doi.org/10.3390/ijms252011093
Clin Transl Radiat Oncol. 2024 Nov;49 100875

Inhibition of OXPHOS induces metabolic rewiring and reduces hypoxia in murine tumor models.

Daan F Boreel, Anne P M Beerkens, Sandra Heskamp, Milou Boswinkel, Johannes P W Peters, Gosse J Adema, Paul N Span, Johan Bussink.

   Introduction: Tumor hypoxia is a feature of many solid malignancies and is known to cause radio resistance. In recent years it has become clear that hypoxic tumor regions also foster an immunosuppressive phenotype and are involved in immunotherapy resistance. It has been proposed that reducing the tumors' oxygen consumption will result in an increased oxygen concentration in the tissue and improve radio- and immunotherapy efficacy. The aim of this study is to investigate the metabolic rewiring of cancer cells by pharmacological attenuation of oxidative phosphorylation (OXPHOS) and subsequently reduce tumor hypoxia.
Material and methods: The metabolic effects of three OXPHOS inhibitors IACS-010759, atovaquone and metformin were explored by measuring oxygen consumption rate, extra cellular acidification rate, and [18F]FDG uptake in 2D and 3D cell culture. Tumor cell growth in 2D cell culture and hypoxia in 3D cell culture were analyzed by live cell imaging. Tumor hypoxia and [18F]FDG uptake in vivo following treatment with IACS-010759 was determined by immunohistochemistry and ex vivo biodistribution respectively.
Results: In vitro experiments show that tumor cell metabolism is heterogeneous between different models. Upon OXPHOS inhibition, metabolism shifts from oxygen consumption through OXPHOS towards glycolysis, indicated by increased acidification and glucose uptake. Inhibition of OXPHOS by IACS-010759 treatment reduced diffusion limited tumor hypoxia in both 3D cell culture and in vivo. Although immune cell presence was lower in hypoxic areas compared with normoxic areas, it is not altered following short term OXPHOS inhibition.
Discussion: These results show that inhibition of OXPHOS causes a metabolic shift from OXPHOS towards increased glycolysis in 2D and 3D cell culture. Moreover, inhibition of OXPHOS reduces diffusion limited hypoxia in 3D cell culture and murine tumor models. Reduced hypoxia by OXPHOS inhibition might enhance therapy efficacy in future studies. However, caution is warranted as systemic metabolic rewiring can cause adverse effects.

Keywords:  Atovaquone; Hypoxia; IACS-010759; Metabolism; Metformin; OXPHOS

DOI:  https://doi.org/10.1016/j.ctro.2024.100875
bioRxiv. 2024 Oct 24. pii: 2024.10.24.620005. [Epub ahead of print]

TFAM as a sensor of UVC-induced mitochondrial DNA damage.

Dillon E King, Emily E Beard, Matthew J Satusky, Ian Ryde, Alex George, Caitlin Johnson, Emma L Dolan, Yuning Zhang, Wei Zhu, Hunter Wilkins, Evan Corden, Susan K Murphy, Dorothy Erie, Raluca Gordân, Joel N Meyer.

Mitochondria lack nucleotide excision DNA repair; however, mitochondrial DNA (mtDNA) is resistant to mutation accumulation following DNA damage. These observations suggest additional damage sensing or protection mechanisms. Transcription Factor A, Mitochondrial (TFAM) compacts mtDNA into nucleoids. As such, TFAM has emerged as a candidate for protecting DNA or sensing damage. To examine these possibilities, we used live-cell imaging, cell-based assays, atomic force microscopy, and high-throughput protein-DNA binding assays to characterize the binding properties of TFAM to UVC-irradiated DNA and cellular consequences of UVC irradiation. Our data indicate an increase in mtDNA degradation and turnover, without a loss in mitochondrial membrane potential that might trigger mitophagy. We identified a reduction in sequence specificity of TFAM associated with UVC irradiation and a redistribution of TFAM binding throughout the mitochondrial genome. Our AFM data show increased compaction of DNA by TFAM in the presence of damage. Despite the TFAM-mediated compaction of mtDNA, we do not observe any protective effect on DNA damage accumulation in cells or in vitro . Taken together, these studies indicate that UVC-induced DNA damage promotes compaction by TFAM, suggesting that TFAM may act as a damage sensor, sequestering damaged genomes to prevent mutagenesis by direct removal or suppression of replication.

DOI: https://doi.org/10.1101/2024.10.24.620005
Nat Metab. 2024 Oct 25.

Quantitation of metabolic activity from isotope tracing data using automated methodology.

Shiyu Liu, Xiaojing Liu, Jason W Locasale.

DOI: https://doi.org/10.1038/s42255-024-01144-2
bioRxiv. 2024 Oct 23. pii: 2024.10.20.619301. [Epub ahead of print]

Enhancing Radiation-induced Reactive Oxygen Species Generation Through Mitochondrial Transplantation in Human Glioblastoma.

Kent L Marshall, Murugesan Velayutham, Valery V Khramtsov, Alan Mizener, Christopher P Cifarelli.

Glioblastoma (GBM) is the most aggressive primary brain malignancy in adults, with high recurrence rates and resistance to standard therapies. This study explores mitochondrial transplantation as a novel method to enhance the radiobiological effect (RBE) of ionizing radiation (IR) by increasing mitochondrial density in GBM cells, potentially boosting reactive oxygen species (ROS) production and promoting radiation-induced cell death. Using cell-penetrating peptides (CPPs), mitochondria were transplanted into GBM cell lines U3020 and U3035. Transplanted mitochondria were successfully incorporated into recipient cells, increasing mitochondrial density significantly. Mitochondrial chimeric cells demonstrated enhanced ROS generation post-irradiation, as evidenced by increased electron paramagnetic resonance (EPR) signal intensity and fluorescent ROS assays. The transplanted mitochondria retained functionality and viability for up to 14 days, with mitochondrial DNA (mtDNA) sequencing confirming high transfection and retention rates. Notably, mitochondrial transplantation was feasible in radiation-resistant GBM cells, suggesting potential clinical applicability. These findings support mitochondrial transplantation as a promising strategy to overcome therapeutic resistance in GBM by amplifying ROS-mediated cytotoxicity, warranting further investigation into its efficacy and mechanisms in vivo .

DOI: https://doi.org/10.1101/2024.10.20.619301
Cell Metab. 2024 Oct 19. pii: S1550-4131(24)00396-6. [Epub ahead of print]

Metabolic regulation of the glioblastoma stem cell epitranscriptome by malate dehydrogenase 2.

Deguan Lv, Deobrat Dixit, Andrea F Cruz, Leo J Y Kim, Likun Duan, Xin Xu, Qiulian Wu, Cuiqing Zhong, Chenfei Lu, Zachary C Gersey, Ryan C Gimple, Qi Xie, Kailin Yang, Xiaojing Liu, Xiaoguang Fang, Xujia Wu, Reilly L Kidwell, Xiuxing Wang, Shideng Bao, Housheng H He, Jason W Locasale, Sameer Agnihotri, Jeremy N Rich.

  Tumors reprogram their metabolism to generate complex neoplastic ecosystems. Here, we demonstrate that glioblastoma (GBM) stem cells (GSCs) display elevated activity of the malate-aspartate shuttle (MAS) and expression of malate dehydrogenase 2 (MDH2). Genetic and pharmacologic targeting of MDH2 attenuated GSC proliferation, self-renewal, and in vivo tumor growth, partially rescued by aspartate. Targeting MDH2 induced accumulation of alpha-ketoglutarate (αKG), a critical co-factor for dioxygenases, including the N6-methyladenosine (m6A) RNA demethylase AlkB homolog 5, RNA demethylase (ALKBH5). Forced expression of MDH2 increased m6A levels and inhibited ALKBH5 activity, both rescued by αKG supplementation. Reciprocally, targeting MDH2 reduced global m6A levels with platelet-derived growth factor receptor-β (PDGFRβ) as a regulated transcript. Pharmacological inhibition of MDH2 in GSCs augmented efficacy of dasatinib, an orally bioavailable multi-kinase inhibitor, including PDGFRβ. Collectively, stem-like tumor cells reprogram their metabolism to induce changes in their epitranscriptomes and reveal possible therapeutic paradigms.

Keywords:  ALKBH5; MDH2; PDGFRβ; alpha-ketoglutarate; cancer stem cell; epitranscriptomics; glioblastoma; m6A; malate-aspartate shuttle; metabolism

DOI:  https://doi.org/10.1016/j.cmet.2024.09.014
Nature. 2024 Oct 30.

RAS-mutant leukaemia stem cells drive clinical resistance to venetoclax.

Junya Sango, Saul Carcamo, Maria Sirenko, Abhishek Maiti, Hager Mansour, Gulay Ulukaya, Lewis E Tomalin, Nataly Cruz-Rodriguez, Tiansu Wang, Malgorzata Olszewska, Emmanuel Olivier, Manon Jaud, Bettina Nadorp, Benjamin Kroger, Feng Hu, Lewis Silverman, Stephen S Chung, Elvin Wagenblast, Ronan Chaligne, Ann-Kathrin Eisfeld, Deniz Demircioglu, Dan A Landau, Piro Lito, Elli Papaemmanuil, Courtney D DiNardo, Dan Hasson, Marina Konopleva, Eirini P Papapetrou.

Cancer driver mutations often show distinct temporal acquisition patterns, but the biological basis for this, if any, remains unknown. RAS mutations occur invariably late in the course of acute myeloid leukaemia, upon progression or relapsed/refractory disease1-6. Here, by using human leukaemogenesis models, we first show that RAS mutations are obligatory late events that need to succeed earlier cooperating mutations. We provide the mechanistic explanation for this in a requirement for mutant RAS to specifically transform committed progenitors of the myelomonocytic lineage (granulocyte-monocyte progenitors) harbouring previously acquired driver mutations, showing that advanced leukaemic clones can originate from a different cell type in the haematopoietic hierarchy than ancestral clones. Furthermore, we demonstrate that RAS-mutant leukaemia stem cells (LSCs) give rise to monocytic disease, as observed frequently in patients with poor responses to treatment with the BCL2 inhibitor venetoclax. We show that this is because RAS-mutant LSCs, in contrast to RAS-wild-type LSCs, have altered BCL2 family gene expression and are resistant to venetoclax, driving clinical resistance and relapse with monocytic features. Our findings demonstrate that a specific genetic driver shapes the non-genetic cellular hierarchy of acute myeloid leukaemia by imposing a specific LSC target cell restriction and critically affects therapeutic outcomes in patients.

DOI: https://doi.org/10.1038/s41586-024-08137-x
Sci Rep. 2024 10 29. 14(1): 25970

Oxidative phosphorylation related gene COA6 is a novel indicator for the prognosis and immune response in lung adenocarcinoma.

Wenting Liu, Yantao Jiang, Guoli Li, Dingzhi Huang, Tingting Qin.

  Although the initial research focused on glycolysis, mitochondrial oxidative phosphorylation has become a major target of cancer cells. Cytochrome C oxidase assembly factor 6 (COA6) is a conserved assembly factor necessary for complex IV biogenesis. Nevertheless, the clinical predictive value of COA6, especially its correlation with immune cell infiltration in lung adenocarcinoma (LUAD), has not yet been elucidated. COA6 exhibited higher expression levels in LUAD cells and tumor tissues compared to normal tissues. Additionally, heightened COA6 expression was associated with reduced overall survival (OS) and advanced tumor stage. Apart from its role in mitochondrial respiratory processes, COA6 may be involved in the process of antigen binding, immunoglobulin receptor binding. Interestingly, we observed a positive correlation between COA6 expression and tumor mutational burden (TMB), as well as a significant association with decreased immune cell infiltration. COA6 was linked to resistance against gemcitabine and etoposide. We verified that COA6 was highly expressed in LUAD experimentally and cell proliferation was inhibited after COA6 knockdown. Thus, we conclude that the expression of COA6 was correlated with reduced immune cell infiltration. Additionally, COA6 functioned as a biomarker for drug sensitivity and the prognosis of lung adenocarcinoma.

Keywords:  Biomarker; COA6; Immune infiltration; Lung adenocarcinoma; Prognosis

DOI:  https://doi.org/10.1038/s41598-024-77775-y
EMBO Mol Med. 2024 Oct 30.

Aberrant fragmentomic features of circulating cell-free mitochondrial DNA as novel biomarkers for multi-cancer detection.

Yang Liu, Fan Peng, Siyuan Wang, Huanmin Jiao, Miao Dang, Kaixiang Zhou, Wenjie Guo, Shanshan Guo, Huanqin Zhang, Wenjie Song, Jinliang Xing.

  Fragmentomic features of circulating cell free mitochondrial DNA (ccf-mtDNA) including fragmentation profile, 5' end base preference and motif diversity are poorly understood. Here, we generated ccf-mtDNA sequencing data of 1607 plasma samples using capture-based next generation sequencing. We firstly found that fragmentomic features of ccf-mtDNA were remarkably different from those of circulating cell free nuclear DNA. Furthermore, region-specific fragmentomic features of ccf-mtDNA were observed, which was associated with protein binding, base composition and special structure of mitochondrial DNA. When comparing to non-cancer controls, six types of cancer patients exhibited aberrant fragmentomic features. Then, cancer detection models were built based on the fragmentomic features. Both internal and external validation cohorts demonstrated the excellent capacity of our model in distinguishing cancer patients from non-cancer control, with all area under curve higher than 0.9322. The overall accuracy of tissue-of-origin was 89.24% and 87.92% for six cancer types in two validation cohort, respectively. Altogether, our study comprehensively describes cancer-specific fragmentomic features of ccf-mtDNA and provides a proof-of-principle for the ccf-mtDNA fragmentomics-based multi-cancer detection and tissue-of-origin classification.

Keywords:  Biomarker; Circulating Cell-Free Mitochondrial DNA; Fragmentomics; Multi-Cancer Detection; Tissue-of-Origin Classification

DOI:  https://doi.org/10.1038/s44321-024-00163-6