bims-mibica 2024-06-30 papers

bims-mibica

Biomed News

on Mitochondrial bioenergetics in cancer

Issue of 2024‒06‒30
twenty-one papers selected by
Kelsey Fisher-Wellman, East Carolina University

Implications of mitochondrial membrane potential gradients on signaling and ATP production analyzed by correlative multi-parameter microscopy.
Mitochondrial respiratory complex I can be inhibited via bypassing the ubiquinone-accessing tunnel.
Human mitochondrial carriers of the SLC25 family function as monomers exchanging substrates with a ping-pong kinetic mechanism.
Active DNA Demethylase, TET1, Increases Oxidative Phosphorylation and Sensitizes Ovarian Cancer Stem Cells to Mitochondrial Complex I Inhibitor.
CHCHD4 regulates the expression of mitochondrial genes that are essential for tumour cell growth.
Canonical and Noncanonical Functions of the BH3 Domain Protein Bid in Apoptosis, Oncogenesis, Cancer Therapeutics, and Aging.
Fluorescence Lifetime Super-Resolution Imaging Unveil the Dynamic Relationship between Mitochondrial Membrane Potential and Cristae Structure Using the Förster Resonance Energy Transfer Strategy.
Triphenylphosphonium-Conjugated Palmitic Acid for Mitochondrial Targeting of Pancreatic Cancer Cells: Proteomic and Molecular Evidence.
Raman micro-spectroscopy reveals the spatial distribution of fumarate in cells and tissues.
Multi-State Gene Cluster Switches Determine the Adaptive Mitochondrial And Metabolic Landscape of Breast Cancer.
Synthesis and anti-proliferative effect of novel 4-Aryl-1, 3-Thiazole-TPP conjugates via mitochondrial uncoupling process.
Targeted inhibition of DHODH is synergistic with BCL2 blockade in HGBCL with concurrent MYC and BCL2 rearrangement.
ACAT1 suppresses clear cell renal cell carcinoma progression by AMPK mediated fatty acid metabolism.
Disarib, a Specific BCL2 Inhibitor, Induces Apoptosis in Triple-Negative Breast Cancer Cells and Impedes Tumour Progression in Xenografts by Altering Mitochondria-Associated Processes.
SIRT4 Has an Anti-Cancer Role in Cervical Cancer by Inhibiting Glutamine Metabolism via the MEK/ERK/C-Myc Axis.
Branched-Chain Amino Acid Degradation Pathway was Inactivated in Colorectal Cancer: Results from a Proteomics Study.
Synthesis and biological characterization of an orally bioavailable lactate dehydrogenase-A inhibitor against pancreatic cancer.
The catalytic activity of methyltransferase METTL15 is dispensable for its role in mitochondrial ribosome biogenesis.
Intra-organ cell specific mitochondrial quantitative interactomics.
Stable Isotope Tracing Analysis in Cancer Research: Advancements and Challenges in Identifying Dysregulated Cancer Metabolism and Treatment Strategies.
Knockout Mouse Studies Show That Mitochondrial CLPP Peptidase and CLPX Unfoldase Act in Matrix Condensates near IMM, as Fast Stress Response in Protein Assemblies for Transcript Processing, Translation, and Heme Production.

Sci Rep. 2024 06 26. 14(1): 14784

Implications of mitochondrial membrane potential gradients on signaling and ATP production analyzed by correlative multi-parameter microscopy.

Benjamin Gottschalk, Zhanat Koshenov, Roland Malli, Wolfgang F Graier.

  The complex architecture and biochemistry of the inner mitochondrial membrane generate ultra-structures with different phospholipid and protein compositions, shapes, characteristics, and functions. The crista junction (CJ) serves as an important barrier separating the cristae (CM) and inner boundary membranes (IBM). Thereby CJ regulates the movement of ions and ensures distinct electrical potentials across the cristae (ΔΨC) and inner boundary (ΔΨIBM) membranes. We have developed a robust and flexible approach to visualize the CJ permeability with super-resolution microscopy as a readout of local mitochondrial membrane potential (ΔΨmito) fluctuations. This method involves analyzing the distribution of TMRM fluorescence intensity in a model that is restricted to the mitochondrial geometry. We show that mitochondrial Ca2+ elevation hyperpolarizes the CM most likely caused by Ca2+ sensitive increase of mitochondrial tricarboxylic acid cycle (TCA) and subsequent oxidative phosphorylation (OXPHOS) activity in the cristae. Dynamic multi-parameter correlation measurements of spatial mitochondrial membrane potential gradients, ATP levels, and mitochondrial morphometrics revealed a CJ-based membrane potential overflow valve mechanism protecting the mitochondrial integrity during excessive cristae hyperpolarization.

Keywords:  Correlative microscopy; Cristae junctions; Membrane potential gradient; Mitochondria; Mitochondrial membranes

DOI:  https://doi.org/10.1038/s41598-024-65595-z
FEBS Lett. 2024 Jun 25.

Mitochondrial respiratory complex I can be inhibited via bypassing the ubiquinone-accessing tunnel.

Ryohei Otani, Takahiro Masuya, Hideto Miyoshi, Masatoshi Murai.

  Mitochondrial NADH-ubiquinone oxidoreductase (complex I) couples electron transfer from NADH to ubiquinone with proton translocation in its membrane part. Structural studies have identified a long (~ 30 Å), narrow, tunnel-like cavity within the enzyme, through which ubiquinone may access a deep reaction site. Although various inhibitors are considered to block the ubiquinone reduction by occupying the tunnel's interior, this view is still debatable. We synthesized a phosphatidylcholine-quinazoline hybrid compound (PC-Qz1), in which a quinazoline-type toxophore was attached to the sn-2 acyl chain to prevent it from entering the tunnel. However, PC-Qz1 inhibited complex I and suppressed photoaffinity labeling by another quinazoline derivative, [125I]AzQ. This study provides further experimental evidence that is difficult to reconcile with the canonical ubiquinone-accessing tunnel model.

Keywords:  complex I; inhibitor; mitochondria; proteoliposomes; respiratory enzymes; ubiquinone

DOI:  https://doi.org/10.1002/1873-3468.14967
EMBO J. 2024 Jun 27.

Human mitochondrial carriers of the SLC25 family function as monomers exchanging substrates with a ping-pong kinetic mechanism.

Camila Cimadamore-Werthein, Martin S King, Denis Lacabanne, Eva Pyrihová, Stephany Jaiquel Baron, Edmund Rs Kunji.

  Members of the SLC25 mitochondrial carrier family link cytosolic and mitochondrial metabolism and support cellular maintenance and growth by transporting compounds across the mitochondrial inner membrane. Their monomeric or dimeric state and kinetic mechanism have been a matter of long-standing debate. It is believed by some that they exist as homodimers and transport substrates with a sequential kinetic mechanism, forming a ternary complex where both exchanged substrates are bound simultaneously. Some studies, in contrast, have provided evidence indicating that the mitochondrial ADP/ATP carrier (SLC25A4) functions as a monomer, has a single substrate binding site, and operates with a ping-pong kinetic mechanism, whereby ADP is imported before ATP is exported. Here we reanalyze the oligomeric state and kinetic properties of the human mitochondrial citrate carrier (SLC25A1), dicarboxylate carrier (SLC25A10), oxoglutarate carrier (SLC25A11), and aspartate/glutamate carrier (SLC25A13), all previously reported to be dimers with a sequential kinetic mechanism. We demonstrate that they are monomers, except for dimeric SLC25A13, and operate with a ping-pong kinetic mechanism in which the substrate import and export steps occur consecutively. These observations are consistent with a common transport mechanism, based on a functional monomer, in which a single central substrate-binding site is alternately accessible.

Keywords:  Bioenergetics; Kinetic Analysis; Mitochondria; SLC25 Mitochondrial Carrier Family; Transport

DOI:  https://doi.org/10.1038/s44318-024-00150-0
Antioxidants (Basel). 2024 Jun 17. pii: 735. [Epub ahead of print]13(6):

Active DNA Demethylase, TET1, Increases Oxidative Phosphorylation and Sensitizes Ovarian Cancer Stem Cells to Mitochondrial Complex I Inhibitor.

Lin-Yu Chen, Yao-An Shen, Ling-Hui Chu, Po-Hsuan Su, Hui-Chen Wang, Yu-Chun Weng, Shiou-Fu Lin, Kuo-Chang Wen, Phui-Ly Liew, Hung-Cheng Lai.

  Ten-eleven translocation 1 (TET1) is a methylcytosine dioxygenase involved in active DNA demethylation. In our previous study, we demonstrated that TET1 reprogrammed the ovarian cancer epigenome, increased stem properties, and activated various regulatory networks, including metabolic networks. However, the role of TET1 in cancer metabolism remains poorly understood. Herein, we uncovered a demethylated metabolic gene network, especially oxidative phosphorylation (OXPHOS). Contrary to the concept of the Warburg effect in cancer cells, TET1 increased energy production mainly using OXPHOS rather than using glycolysis. Notably, TET1 increased the mitochondrial mass and DNA copy number. TET1 also activated mitochondrial biogenesis genes and adenosine triphosphate production. However, the reactive oxygen species levels were surprisingly decreased. In addition, TET1 increased the basal and maximal respiratory capacities. In an analysis of tricarboxylic acid cycle metabolites, TET1 increased the levels of α-ketoglutarate, which is a coenzyme of TET1 dioxygenase and may provide a positive feedback loop to modify the epigenomic landscape. TET1 also increased the mitochondrial complex I activity. Moreover, the mitochondrial complex I inhibitor, which had synergistic effects with the casein kinase 2 inhibitor, affected ovarian cancer growth. Altogether, TET1-reprogrammed ovarian cancer stem cells shifted the energy source to OXPHOS, which suggested that metabolic intervention might be a novel strategy for ovarian cancer treatment.

Keywords:  DNA demethylation; casein kinase-2 subunit alpha (CK2α) inhibitor; mitochondria; mitochondrial complex I inhibitor; ovarian cancer stem cells; oxidative phosphorylation (OXPHOS); ten-eleven translocation 1 (TET1)

DOI:  https://doi.org/10.3390/antiox13060735
Biochim Biophys Acta Mol Basis Dis. 2024 Jun 21. pii: S0925-4439(24)00271-0. [Epub ahead of print] 167282

CHCHD4 regulates the expression of mitochondrial genes that are essential for tumour cell growth.

Luke W Thomas, Jenna M Stephen, Margaret Ashcroft.

  CHCHD4 (MIA40) is a central component of the mitochondrial disulfide relay system (DRS), is essential and evolutionarily conserved. Previously, we have shown CHCHD4 to be a critical regulator of tumour cell growth. Here, we use genome-wide CRISPR/Cas9 and SILAC proteomic analyses to delineate mechanisms of CHCHD4 essentiality in cancer. We identify a short-list of common essential genes/proteins associated with CHCHD4 essentiality in tumour cells, which includes subunits of complex I that are known DRS substrates, and genes/proteins involved in key metabolic pathways. Our study highlights a range of nuclear encoded mitochondrial genes essential for CHCHD4-regulated tumour cell growth.

Keywords:  CHCHD4; CRISPR/Cas9 genome-wide deletion screen; Gene essentiality; Mitochondria; Proteomics; SILAC analysis

DOI:  https://doi.org/10.1016/j.bbadis.2024.167282
Cancers (Basel). 2024 Jun 12. pii: 2199. [Epub ahead of print]16(12):

Canonical and Noncanonical Functions of the BH3 Domain Protein Bid in Apoptosis, Oncogenesis, Cancer Therapeutics, and Aging.

Yetunde Makinwa, Yibo Luo, Phillip R Musich, Yue Zou.

  Effective cancer therapy with limited adverse effects is a major challenge in the medical field. This is especially complicated by the development of acquired chemoresistance. Understanding the mechanisms that underlie these processes remains a major effort in cancer research. In this review, we focus on the dual role that Bid protein plays in apoptotic cell death via the mitochondrial pathway, in oncogenesis and in cancer therapeutics. The BH3 domain in Bid and the anti-apoptotic mitochondrial proteins (Bcl-2, Bcl-XL, mitochondrial ATR) it associates with at the outer mitochondrial membrane provides us with a viable target in cancer therapy. We will discuss the roles of Bid, mitochondrial ATR, and other anti-apoptotic proteins in intrinsic apoptosis, exploring how their interaction sustains cellular viability despite the initiation of upstream death signals. The unexpected upregulation of this Bid protein in cancer cells can also be instrumental in explaining the mechanisms behind acquired chemoresistance. The stable protein associations at the mitochondria between tBid and anti-apoptotic mitochondrial ATR play a crucial role in maintaining the viability of cancer cells, suggesting a novel mechanism to induce cancer cell apoptosis by freeing tBid from the ATR associations at mitochondria.

Keywords:  ATR; BH3 domain; Bid; LB-100; apoptosis; cancer; cancer therapy; chemoresistance; mitochondria; oncogenesis; senescence

DOI:  https://doi.org/10.3390/cancers16122199
Anal Chem. 2024 Jun 26.

Fluorescence Lifetime Super-Resolution Imaging Unveil the Dynamic Relationship between Mitochondrial Membrane Potential and Cristae Structure Using the Förster Resonance Energy Transfer Strategy.

Fei Peng, Xiangnan Ai, Jing Sun, Xichuan Ge, Meiqi Li, Peng Xi, Baoxiang Gao.

Mitochondrial cristae, invaginations of the inner mitochondrial membrane (IMM) into the matrix, are the main site for the generation of ATP via oxidative phosphorylation, and mitochondrial membrane potential (MMP). Synchronous study of the dynamic relationship between cristae and MMP is very important for further understanding of mitochondrial function. Due to the lack of suitable IMM probes and imaging techniques, the dynamic relationship between MMP and cristae structure alterations remains poorly understood. We designed a pair of FRET-based molecular probes, with the donor (OR-LA) being rhodamine modified with mitochondrial coenzyme lipoic acid and the acceptor (SiR-BA) being silicon-rhodamine modified with a butyl chain, for simultaneous dynamic monitoring of mitochondrial cristae structure and MMP. The FRET process of the molecular pair in mitochondria is regulated by MMP, enabling more precise visualization of MMP through fluorescence intensity ratio and fluorescence lifetime. By combining FRET with FLIM super-resolution imaging technology, we achieved simultaneous dynamic monitoring of mitochondrial cristae structure and MMP, revealing that during the decline of MMP, there is a progression involving cristae dilation, fragmentation, mitochondrial vacuolization, and eventual rupture. Significantly, we successfully observed that the rapid decrease in MMP at the site of mitochondrial membrane rupture may be a critical factor in mitochondrial fragmentation. These data collectively reveal the dynamic relationship between cristae structural alterations and MMP decline, laying a foundation for further investigation into cellular energy regulation mechanisms and therapeutic strategies for mitochondria-related diseases.

DOI: https://doi.org/10.1021/acs.analchem.4c01905
Int J Mol Sci. 2024 Jun 20. pii: 6790. [Epub ahead of print]25(12):

Triphenylphosphonium-Conjugated Palmitic Acid for Mitochondrial Targeting of Pancreatic Cancer Cells: Proteomic and Molecular Evidence.

Giuliana Siragusa, Jessica Brandi, Tristan Rawling, Michael Murray, Daniela Cecconi.

  Pancreatic ductal adenocarcinoma (PDAC)'s resistance to therapies is mainly attributed to pancreatic cancer stem cells (PCSCs). Mitochondria-impairing agents can be used to hamper PCSC propagation and reduce PDAC progression. Therefore, to develop an efficient vector for delivering drugs to the mitochondria, we synthesized tris(3,5-dimethylphenyl)phosphonium-conjugated palmitic acid. Triphenylphosphonium (TPP) is a lipophilic cationic moiety that promotes the accumulation of conjugated agents in the mitochondrion. Palmitic acid (PA), the most common saturated fatty acid, has pro-apoptotic activity in different types of cancer cells. TPP-PA was prepared by the reaction of 16-bromopalmitic acid with TPP, and its structure was characterized by 1H and 13C NMR and HRMS. We compared the proteomes of TPP-PA-treated and untreated PDAC cells and PCSCs, identifying dysregulated proteins and pathways. Furthermore, assessments of mitochondrial membrane potential, intracellular ROS, cardiolipin content and lipid peroxidation, ER stress, and autophagy markers provided information on the mechanism of action of TPP-PA. The findings showed that TPP-PA reduces PDAC cell proliferation through mitochondrial disruption that leads to increased ROS, activation of ER stress, and autophagy. Hence, TPP-PA might offer a new approach for eliminating both the primary population of cancer cells and PCSCs, which highlights the promise of TPP-derived compounds as anticancer agents for PDAC.

Keywords:  mitochondria; palmitic acid; pancreatic cancer; reactive oxygen species; triphenylphosphonium

DOI:  https://doi.org/10.3390/ijms25126790
Nat Commun. 2024 Jun 25. 15(1): 5386

Raman micro-spectroscopy reveals the spatial distribution of fumarate in cells and tissues.

Marlous Kamp, Jakub Surmacki, Marc Segarra Mondejar, Tim Young, Karolina Chrabaszcz, Fadwa Joud, Vincent Zecchini, Alyson Speed, Christian Frezza, Sarah E Bohndiek.

Aberrantly accumulated metabolites elicit intra- and inter-cellular pro-oncogenic cascades, yet current measurement methods require sample perturbation/disruption and lack spatio-temporal resolution, limiting our ability to fully characterize their function and distribution. Here, we show that Raman spectroscopy (RS) can directly detect fumarate in living cells in vivo and animal tissues ex vivo, and that RS can distinguish between Fumarate hydratase (Fh1)-deficient and Fh1-proficient cells based on fumarate concentration. Moreover, RS reveals the spatial compartmentalization of fumarate within cellular organelles in Fh1-deficient cells: consistent with disruptive methods, we observe the highest fumarate concentration (37 ± 19 mM) in mitochondria, where the TCA cycle operates, followed by the cytoplasm (24 ± 13 mM) and then the nucleus (9 ± 6 mM). Finally, we apply RS to tissues from an inducible mouse model of FH loss in the kidney, demonstrating RS can classify FH status. These results suggest RS could be adopted as a valuable tool for small molecule metabolic imaging, enabling in situ non-destructive evaluation of fumarate compartmentalization.

DOI: https://doi.org/10.1038/s41467-024-49403-w
Cancer Res. 2024 Jun 26.

Multi-State Gene Cluster Switches Determine the Adaptive Mitochondrial And Metabolic Landscape of Breast Cancer.

Michela Menegollo, Robert B Bentham, Tiago Henriques, Seow Qi Ng, Ziyu Ren, Clarinde Esculier, Sia Agarwal, Emily T Y Tong, Clement Lo, Sanjana Ilangovan, Zorka Szabadkai, Matteo Suman, Neill Patani, Avinash Ghanate, Kevin Bryson, Robert C Stein, Mariia Yuneva, Gyorgy Szabadkai.

Adaptive metabolic switches are proposed to underlie conversions between cellular states during normal development as well as in cancer evolution. Metabolic adaptations represent important therapeutic targets in tumors, highlighting the need to characterize the full spectrum, characteristics, and regulation of the metabolic switches. To investigate the hypothesis that metabolic switches associated with specific metabolic states can be recognized by locating large alternating gene expression patterns, we developed a method to identify interspersed gene sets by massive correlated biclustering (MCbiclust) and to predict their metabolic wiring. Testing the method on breast cancer transcriptome datasets revealed a series of gene sets with switch-like behavior that could be used to predict mitochondrial content, metabolic activity, and central carbon flux in tumors. The predictions were experimentally validated by bioenergetic profiling and metabolic flux analysis of 13C-labelled substrates. The metabolic switch positions also distinguished between cellular states, correlating with tumor pathology, prognosis, and chemosensitivity. The method is applicable to any large and heterogeneous transcriptome dataset to discover metabolic and associated pathophysiological states.

DOI: https://doi.org/10.1158/0008-5472.CAN-23-3172
Bioorg Chem. 2024 Jun 23. pii: S0045-2068(24)00493-0. [Epub ahead of print]150 107588

Synthesis and anti-proliferative effect of novel 4-Aryl-1, 3-Thiazole-TPP conjugates via mitochondrial uncoupling process.

Yixin Hu, Yang Zhang, Jie Guo, Shihao Chen, Jie Jin, Pengyu Li, Yuchen Pan, Shuwen Lei, Jiaqi Li, Suheng Wu, Buzhou Bu, Lei Fu.

  With the advent of mitochondrial targeting moiety such as triphenlyphosphonium cation (TPP+), targeting mitochondria in cancer cells has become a promising strategy for combating tumors. Herein, a series of novel 4-aryl-1,3-thiazole derivatives linked to TPP+ moiety were designed and synthesized. The cytotoxicity against a panel of four cancer cell lines was evaluated by CCK-8 assay. Most of these compounds exhibited moderate to good inhibitory activity over HeLa, PC-3 and HCT-15 cells while MCF-7 cells were less sensitive to most compounds. Among them, compound 12a exhibited a significant anti-proliferative activity against HeLa cells, and prompted for further investigation. Specifically, 12a decreased mitochondrial membrane potential and enhanced levels of reactive oxygen species (ROS). The flow cytometry analysis revealed that compound 12a could induce apoptosis and cell cycle arrest at G0/G1 phase in HeLa cells. In addition, mitochondrial bioenergetics assay revealed that 12a displayed mild mitochondrial uncoupling effect. Taken together, these findings suggest the therapeutic potential of compound 12a as an antitumor agent targeting mitochondria.

Keywords:  Anticancer; Mitochondrial bioenergetics; Mitochondrial targeting; Mitochondrial uncoupler; Oxidative phosphorylation

DOI:  https://doi.org/10.1016/j.bioorg.2024.107588
BMC Cancer. 2024 Jun 25. 24(1): 761

Targeted inhibition of DHODH is synergistic with BCL2 blockade in HGBCL with concurrent MYC and BCL2 rearrangement.

Lin Liu, Wenbin Mo, Miao Chen, Yi Qu, Pingping Wang, Ying Liang, Xiaojing Yan.

  High-grade B-cell lymphoma (HGBCL), the subtype of non-Hodgkin lymphoma, to be relapsed or refractory in patients after initial therapy or salvage chemotherapy. Dual dysregulation of MYC and BCL2 is one of the important pathogenic mechanisms. Thus, combined targeting of MYC and BCL2 appears to be a promising strategy. Dihydroorotate dehydrogenase (DHODH) is the fourth rate-limiting enzyme for the de novo biosynthesis of pyrimidine. It has been shown to be a potential therapeutic target for multiple diseases. In this study, the DHODH inhibitor brequinar exhibited growth inhibition, cell cycle blockade, and apoptosis promotion in HGBCL cell lines with MYC and BCL2 rearrangements. The combination of brequinar and BCL2 inhibitors venetoclax had a synergistic inhibitory effect on the survival of DHL cells through different pathways. Venetoclax could upregulate MCL-1 and MYC expression, which has been reported as a resistance mechanism of BCL2 inhibitors. Brequinar downregulated MCL-1 and MYC, which could potentially overcome drug resistance to venetoclax in HGBCL cells. Furthermore, brequinar could downregulate a broad range of genes, including ribosome biosynthesis genes, which might contribute to its anti-tumor effects. In vivo studies demonstrated synergetic tumor growth inhibition in xenograft models with brequinar and venetoclax combination treatment. These results provide preliminary evidence for the rational combination of DHODH and BCL2 blockade in HGBCL with abnormal MYC and BCL2.

Keywords:  BCL2; DHODH inhibitor; High-grade B-cell lymphoma; MYC; Venetoclax

DOI:  https://doi.org/10.1186/s12885-024-12534-w
Transl Oncol. 2024 Jun 22. pii: S1936-5233(24)00170-0. [Epub ahead of print]47 102043

ACAT1 suppresses clear cell renal cell carcinoma progression by AMPK mediated fatty acid metabolism.

Ming Zheng, Shenghu Zhang, Jiajie Zhou, Ming Lin, Yixiang Liao.

  Renal cell carcinoma (RCC) stands as a prevalent malignancy within urological pathology, exhibiting a noteworthy escalation in its incidence. Despite being a mitochondrial enzyme, the precise role of Acetyl-CoA Acetyltransferase 1 (ACAT1) in RCC remains elusive. In this investigation, we employed bioinformatics methodologies to assess the expression patterns and prognostic significance across various RCC subtypes, encompassing clear cell renal cell carcinoma (ccRCC), papillary cell carcinoma, and chromophobe cell carcinoma. Our findings unveil a close correlation between ACAT1 expression and the prognostic implications specifically within ccRCC. Through both in vitro and in vivo overexpression studies, we delineated the functional and mechanistic facets of ACAT1 in impeding the progression of ccRCC. Our results unequivocally demonstrated that ACAT1 overexpression markedly curtailed proliferation, invasion, and metastasis of ccRCC cells in both in vivo models and cell cultures. Mechanistically, ACAT1's inhibitory effect on the AMPK signaling pathway orchestrated a regulatory role in modulating fatty acid metabolism, thereby effectively restraining the advancement of ccRCC. Collectively, our findings underscore ACAT1 as a pivotal tumor suppressor, instrumental in curtailing the proliferation, migration, and invasion of ccRCC by governing fatty acid metabolism through the AMPK signaling pathway. These insights posit ACAT1 as a potential predictive biomarker and therapeutic target warranting further exploration in RCC management.

Keywords:  Acetyl-CoA Acetyltransferase 1; Clear cell renal cell carcinoma; Fatty acid metabolism; Progression

DOI:  https://doi.org/10.1016/j.tranon.2024.102043
Int J Mol Sci. 2024 Jun 12. pii: 6485. [Epub ahead of print]25(12):

Disarib, a Specific BCL2 Inhibitor, Induces Apoptosis in Triple-Negative Breast Cancer Cells and Impedes Tumour Progression in Xenografts by Altering Mitochondria-Associated Processes.

Meghana Manjunath, Febina Ravindran, Shivangi Sharma, Humaira Siddiqua, Sathees C Raghavan, Bibha Choudhary.

  Targeted cancer therapy aims to disrupt the functions of proteins that regulate cancer progression, mainly by using small molecule inhibitors (SMIs). SMIs exert their effect by modulating signalling pathways, organelle integrity, chromatin components, and several biosynthetic processes essential for cell division and survival. Antiapoptotic protein BCL2 is highly upregulated in many cancers compared with normal cells, making it an ideal target for cancer therapy. Around 75% of primary breast cancers overexpress BCL2, providing an opportunity to explore BCL2 inhibitors as a therapeutic option. Disarib is an SMI that has been developed as a selective BCL2 inhibitor. Disarib works by disrupting BCL2-BAK interaction and activating intrinsic apoptotic pathways in leukemic cells while sparing normal cells. We investigated the effects of Disarib, a BCL2 specific inhibitor, on breast cancer cells and xenografts. Cytotoxicity and fluorometric assays revealed that Disarib induced cell death by increasing reactive oxygen species and activating intrinsic apoptotic pathways in Triple-Negative Breast Cancer cells (MDA-MB-231 and MDA-MB-468). Disarib also affected the colony-forming properties of these cells. MDA-MB-231- and MDA-MB-468-derived xenografts showed a significant reduction in tumours upon Disarib treatment. Through the transcriptomics approach, we also explored the influence of BCL2 inhibitors on energy metabolism, mitochondrial dynamics, and epithelial-to-mesenchymal transition (EMT). Mitochondrial dynamics and glucose metabolism mainly regulate energy metabolism. The change in energetics regulates tumour growth through epithelial-mesenchymal transition, and angiogenesis. RNA sequencing (RNAseq) analysis revealed that BCL2 inhibitors ABT-199 and Disarib maintain Oxphos levels in MDA-MB-231. However, key glycolytic genes were significantly downregulated. Mitochondrial fission genes were seen to be downregulated both in RNAseq data and semi quantitative real time polymerase chain reaction (qRTPCR) in Disarib-treated TNBC cells and xenografts. Lastly, Disarib inhibited wound healing and epithelial-to-mesenchymal transition. This study showed that Disarib disrupts mitochondrial function, activates the intrinsic apoptotic pathway in breast cancer, and inhibits epithelial-to-mesenchymal transition both in vitro and in vivo. These findings highlight Disarib's potential as a multifaceted therapeutic strategy for patients with Triple-Negative Breast Cancer.

Keywords:  BCL2 inhibitors; energy metabolism; gene expression; triple-negative breast cancer

DOI:  https://doi.org/10.3390/ijms25126485
Anticancer Res. 2024 Jul;44(7): 2861-2870

SIRT4 Has an Anti-Cancer Role in Cervical Cancer by Inhibiting Glutamine Metabolism via the MEK/ERK/C-Myc Axis.

Yina Zhang, Xiaocen Niu, Yuanhe Wang, Shuting Bao, Peiyue Jiang, Xiaorui Dong.

  BACKGROUND/AIM: Glutamine metabolism is crucial in cell proliferation, aging, and apoptosis across various cancer types. Existing research indicates that Sirtuin 4 (SIRT4), primarily located in mitochondria, modulates this process. This study aimed to clarify the regulatory relationship between SIRT4 and glutamine metabolism in cervical cancer.MATERIALS AND METHODS: SIRT4 mRNA levels and their clinical correlation to cervical cancer were analyzed using the UALCAN database. Immunohistochemistry (IHC) was performed to assess SIRT4 protein expression in tissue samples from cervical cancer patients. Transient transfection was employed to create Hela and Siha cell lines with overexpressed SIRT4, mitogen-activated extracellular signal-regulated kinase (MEK), and glutaminase 1 (GLS1). The impact on cellular functions was studied using MTT, soft agar, transwell, and western blotting assays. Glutamate and ATP levels were also measured to evaluate metabolic changes.
RESULTS: Low levels of SIRT4 mRNA in cervical cancer tissues correlated with tumor metastasis and poor survival rates. Overexpression of SIRT4 led to suppressed cell proliferation, colony growth, and motility, along with significant down-regulation of GLS expression, a key contributor to glutamine metabolism. Additionally, SIRT4 overexpression resulted in the inactivation of the MEK/ERK/c-myc signaling pathway, while overexpression of MEK reversed these effects. Notably, the inhibitory effects of SIRT4 on cell proliferation, colony formation, migration, and invasion in Hela and Siha cells were significantly attenuated following GLS1 overexpression.
CONCLUSION: SIRT4 acts as an anti-cancer agent in cervical cancer by inhibiting glutamine metabolism through the MEK/ERK/c-myc signaling pathway, providing a novel sight for cervical cancer therapy.

Keywords:  MEK/ERK/c-myc signal; SIRT4; cervical cancer; glutamine metabolism

DOI:  https://doi.org/10.21873/anticanres.17098
J Cancer. 2024 ;15(12): 3724-3737

Branched-Chain Amino Acid Degradation Pathway was Inactivated in Colorectal Cancer: Results from a Proteomics Study.

Shixian Lian, Siyuan Liu, Ao Wu, Lin Yin, Lei Li, Liyan Zeng, Mingkun Zhao, Lijun Zhang.

  Background: Colorectal cancer (CRC) ranks third in terms of cancer incidence and fourth in terms of cancer-related deaths worldwide. Identifying potential biomarkers of CRC is crucial for treatment and drug development. Methods: In this study, we established a C57B/6N mouse model of colon carcinogenesis using azoxymethane-dextran sodium sulfate (AOM-DSS) treatment for 14 weeks to identify proteins associated with colon cancer. An isobaric tag for relative and absolute quantitation (iTRAQ)-based proteomic analysis was conducted on the cell membrane components enriched in the colonic mucosa. Additionally, tumor tissues and adjacent normal colon tissues were collected from patients with colon cancer for comparative protein and metabolite analyses. Results: In total, 74 differentially expressed proteins were identified in the tumor tissue samples from AOM/DSS-treated mice compared to both the adjacent tissue samples from AOM/DSS-treated mice and tissue samples from saline-treated control mice. Bioinformatics analysis revealed eight downregulated proteins enriched in the branched-chain amino acids pathway (valine, leucine, and isoleucine degradation). Moreover, these proteins are already known to be associated with the survival rate of patients with cancer. Targeted metabolomics showed increased levels of valine, leucine, and isoleucine in tumor tissues compared to those in adjacent normal tissues in patients with colon cancer. Furthermore, a real-time PCR experiment demonstrated that Aldehyde dehydrogenase, mitochondrial (short protein name ALDH2, gene name Aldh2) and Hydroxyacyl-coenzyme A dehydrogenase, mitochondrial (short protein name HCDH, gene name Hadh) (two genes) in the pathway of branched-chain amino acids) were downregulated in patients with colon cancer (colon tumor tissues vs. their adjacent colon tissues). ALDH2 expression was further validated by western blotting in AOM/DSS-treated mouse model and in clinical samples. Conclusion: This study highlighted the inactivation of the branched-chain amino acid degradation pathway in colon cancer and identified ALDH2 and HCDH as potential biomarkers for diagnosing colon cancer and developing new therapeutic strategies.

Keywords:  ALDH2; AOM/DSS; HCDH; branched chain amino acids.; colon cancer; proteomics

DOI:  https://doi.org/10.7150/jca.95454
Eur J Med Chem. 2024 Jun 17. pii: S0223-5234(24)00478-1. [Epub ahead of print]275 116598

Synthesis and biological characterization of an orally bioavailable lactate dehydrogenase-A inhibitor against pancreatic cancer.

Horrick Sharma, Somrita Mondal, Uzziah Urquiza, Colter Esparza, Seth Bartlett, Landon Santa-Pinter, Hanna Hill, Madalyn White, Pragya Sharma, Lerin Luckett-Chastain, Anne Cooper, Mohammad Rasel, Philip Gao, Kevin P Battaile, Surendra K Shukla, Scott Lovell, Michael A Ihnat.

  Lactate dehydrogenase-A (LDHA) is the major isoform of lactate dehydrogenases (LDH) that is overexpressed and linked to poor survival in pancreatic ductal adenocarcinoma (PDAC). Despite some progress, current LDH inhibitors have poor structural and physicochemical properties or exhibit unfavorable pharmacokinetics that have hampered their development. The present study reports the synthesis and biological evaluation of a novel class of LDHA inhibitors comprising a succinic acid monoamide motif. Compounds 6 and 21 are structurally related analogs that demonstrated potent inhibition of LDHA with IC50s of 46 nM and 72 nM, respectively. We solved cocrystal structures of compound 21-bound to LDHA that showed that the compound binds to a distinct allosteric site between the two subunits of the LDHA tetramer. Inhibition of LDHA correlated with reduced lactate production and reduction of glycolysis in MIA PaCa-2 pancreatic cancer cells. The lead compounds inhibit the proliferation of human pancreatic cancer cell lines and patient-derived 3D organoids and exhibit a synergistic cytotoxic effect with the OXPHOS inhibitor phenformin. Unlike current LDHA inhibitors, 6 and 21 have appropriate pharmacokinetics and ligand efficiency metrics, exhibit up to 73% oral bioavailability, and a cumulative half-life greater than 4 h in mice.

Keywords:  Lactate; Lactate Dehydrogenase-A; OXPHOS; Pancreatic cancer; Pharmacokinetics; Small molecule inhibitor

DOI:  https://doi.org/10.1016/j.ejmech.2024.116598
RNA Biol. 2024 Jan;21(1): 23-30

The catalytic activity of methyltransferase METTL15 is dispensable for its role in mitochondrial ribosome biogenesis.

Christian D Mutti, Lindsey Van Haute, Michal Minczuk.

  Ribosomes are large macromolecular complexes composed of both proteins and RNA, that require a plethora of factors and post-transcriptional modifications for their biogenesis. In human mitochondria, the ribosomal RNA is post-transcriptionally modified at ten sites. The N4-methylcytidine (m4C) methyltransferase, METTL15, modifies the 12S rRNA of the small subunit at position C1486. The enzyme is essential for mitochondrial protein synthesis and assembly of the mitoribosome small subunit, as shown here and by previous studies. Here, we demonstrate that the m4C modification is not required for small subunit biogenesis, indicating that the chaperone-like activity of the METTL15 protein itself is an essential component for mitoribosome biogenesis.

Keywords:  Mitochondrial ribosome; chaperone; epitranscriptomics; methyltransferase; mitochondria; ribosomal RNA

DOI:  https://doi.org/10.1080/15476286.2024.2369374
bioRxiv. 2024 Jun 12. pii: 2024.06.10.598354. [Epub ahead of print]

Intra-organ cell specific mitochondrial quantitative interactomics.

A A Bakhtina, M D Campbell, B D Sibley, M Sanchez-Contreras, M T Sweetwyne, J E Bruce.

Almost every organ consists of many cell types, each with its unique functions. Proteomes of these cell types are thus unique too. But it is reasonable to assume that interactome (inter and intra molecular interactions of proteins) are also distinct since protein interactions are what ultimately carry out the function. Podocytes and tubules are two cell types within kidney with vastly different functions: podocytes envelop the blood vessels in the glomerulus and act as filters while tubules are located downstream of the glomerulus and are responsible for reabsorption of important nutrients. It has been long known that for tubules mitochondria plays an important role as they require a lot of energy to carry out their functions. In podocytes, however, it has been assumed that mitochondria might not matter as much in both normal physiology and pathology 1 . Here we have applied quantitative cross-linking mass spectrometry to compare mitochondrial interactomes of tubules and podocytes using a transgenic mitochondrial tagging strategy to immunoprecipitate cell-specific mitochondria directly from whole kidney. We have uncovered that mitochondrial proteomes of these cell types are quite similar, although still showing unique features that correspond to known functions, such as high energy production through TCA cycle in tubules and requirements for detoxification in podocytes which are on the frontline of filtration where they encounter toxic compounds and therefore, as a non-renewing cell type they must have ways to protect themselves from cellular toxicity. But we gained much deeper insight with the interactomics data. We were able to find pathways differentially regulated in podocytes and tubules based on changing cross-link levels and not just protein levels. Among these pathways are betaine metabolism, lysine degradation, and many others. We have also demonstrated how quantitative interactomics could be used to detect different activity levels of an enzyme even when protein abundances of it are the same between cell types. We have validated this finding with an orthogonal activity assay. Overall, this work presents a new view of mitochondrial biology for two important, but functionally distinct, cell types within the mouse kidney showing both similarities and unique features. This data can continue to be explored to find new aspects of mitochondrial biology, especially in podocytes, where mitochondria has been understudied. In the future this methodology can also be applied to other organs to uncover differences in the function of cell types.

DOI: https://doi.org/10.1101/2024.06.10.598354
Metabolites. 2024 May 31. pii: 318. [Epub ahead of print]14(6):

Stable Isotope Tracing Analysis in Cancer Research: Advancements and Challenges in Identifying Dysregulated Cancer Metabolism and Treatment Strategies.

Dalton Hilovsky, Joshua Hartsell, Jamey D Young, Xiaojing Liu.

  Metabolic reprogramming is a hallmark of cancer, driving the development of therapies targeting cancer metabolism. Stable isotope tracing has emerged as a widely adopted tool for monitoring cancer metabolism both in vitro and in vivo. Advances in instrumentation and the development of new tracers, metabolite databases, and data analysis tools have expanded the scope of cancer metabolism studies across these scales. In this review, we explore the latest advancements in metabolic analysis, spanning from experimental design in stable isotope-labeling metabolomics to sophisticated data analysis techniques. We highlight successful applications in cancer research, particularly focusing on ongoing clinical trials utilizing stable isotope tracing to characterize disease progression, treatment responses, and potential mechanisms of resistance to anticancer therapies. Furthermore, we outline key challenges and discuss potential strategies to address them, aiming to enhance our understanding of the biochemical basis of cancer metabolism.

Keywords:  cancer metabolism; high resolution mass spectrometry; stable isotope tracing

DOI:  https://doi.org/10.3390/metabo14060318
Genes (Basel). 2024 May 27. pii: 694. [Epub ahead of print]15(6):

Knockout Mouse Studies Show That Mitochondrial CLPP Peptidase and CLPX Unfoldase Act in Matrix Condensates near IMM, as Fast Stress Response in Protein Assemblies for Transcript Processing, Translation, and Heme Production.

Jana Key, Suzana Gispert, Georg Auburger.

  LONP1 is the principal AAA+ unfoldase and bulk protease in the mitochondrial matrix, so its deletion causes embryonic lethality. The AAA+ unfoldase CLPX and the peptidase CLPP also act in the matrix, especially during stress periods, but their substrates are poorly defined. Mammalian CLPP deletion triggers infertility, deafness, growth retardation, and cGAS-STING-activated cytosolic innate immunity. CLPX mutations impair heme biosynthesis and heavy metal homeostasis. CLPP and CLPX are conserved from bacteria to humans, despite their secondary role in proteolysis. Based on recent proteomic-metabolomic evidence from knockout mice and patient cells, we propose that CLPP acts on phase-separated ribonucleoprotein granules and CLPX on multi-enzyme condensates as first-aid systems near the inner mitochondrial membrane. Trimming within assemblies, CLPP rescues stalled processes in mitoribosomes, mitochondrial RNA granules and nucleoids, and the D-foci-mediated degradation of toxic double-stranded mtRNA/mtDNA. Unfolding multi-enzyme condensates, CLPX maximizes PLP-dependent delta-transamination and rescues malformed nascent peptides. Overall, their actions occur in granules with multivalent or hydrophobic interactions, separated from the aqueous phase. Thus, the role of CLPXP in the matrix is compartment-selective, as other mitochondrial peptidases: MPPs at precursor import pores, m-AAA and i-AAA at either IMM face, PARL within the IMM, and OMA1/HTRA2 in the intermembrane space.

Keywords:  ALAS; GFM1; ISC; OAT; PNPT1; Perrault syndrome type 3 (PRLTS3); RNA-G4; VWA8; iron toxicity; pyridoxal-5′-phosphate

DOI:  https://doi.org/10.3390/genes15060694