bims-mibica Biomed News
on Mitochondrial bioenergetics in cancer
Issue of 2022‒12‒18
34 papers selected by
Kelsey Fisher-Wellman
East Carolina University

  1. Clin Exp Med. 2022 Dec 17.
      Altered mitochondrial function contributes greatly to pathogenesis and progression of colorectal cancer. In this study, we report a functional pool of Src homology 2 domain-containing F (SHF) in mitochondria controlling the response of colorectal cancer cells to radiation therapy. We found that elevated expression of SHF in cancer cells is essential for promoting mitochondrial function by increasing mitochondrial DNA copy number, thus reducing the sensitivity of colorectal cancer cells to radiation. Mechanistically, SHF binds to mitochondrial DNA and promotes POLG/SSBP1-mediated mitochondrial DNA synthesis. Importantly, SHF loss-mediated radiosensitization was phenocopied by depletion of mitochondrial DNA. Thus, our data demonstrate that mitochondrial SHF is an important regulator of radioresistance in colorectal cancer cells, identifying SHF as a promising therapeutic target to enhance radiotherapy efficacy in colorectal cancer.
    Keywords:  Colorectal cancer; Mitochondrial DNA copy number; Mitochondrial function; Radioresistance; Radiosensitivity
  2. Cell Rep. 2022 Dec 13. pii: S2211-1247(22)01710-7. [Epub ahead of print]41(11): 111818
      Oncogenic KRas activates mitochondrial fission through Erk-mediated phosphorylation of the mitochondrial fission GTPase Drp1. Drp1 deletion inhibits tumorigenesis of KRas-driven pancreatic cancer, but the role of mitochondrial dynamics in other Ras-driven malignancies is poorly defined. Here we show that in vitro and in vivo growth of KRas-driven lung adenocarcinoma is unaffected by deletion of Drp1 but is inhibited by deletion of Opa1, the GTPase that regulates inner membrane fusion and proper cristae morphology. Mechanistically, Opa1 knockout disrupts cristae morphology and inhibits electron transport chain (ETC) assembly and activity, which inhibits tumor cell proliferation through loss of NAD+ regeneration. Simultaneous inactivation of Drp1 and Opa1 restores cristae morphology, ETC activity, and cell proliferation indicating that mitochondrial fission activity drives ETC dysfunction induced by Opa1 knockout. Our results support a model in which mitochondrial fission events disrupt cristae structure, and tumor cells with hyperactive fission activity require Opa1 activity to maintain ETC function.
    Keywords:  CP: Cancer; Drp1; ETC; KRas; NAD; Opa1; cancer; cristae; fission; fusion; mitochondria
  3. Sci Adv. 2022 Dec 16. 8(50): eabp8293
      Targeting metabolic vulnerabilities has been proposed as a therapeutic strategy in renal cell carcinoma (RCC). Here, we analyzed the metabolism of patient-derived xenografts (tumorgrafts) from diverse subtypes of RCC. Tumorgrafts from VHL-mutant clear cell RCC (ccRCC) retained metabolic features of human ccRCC and engaged in oxidative and reductive glutamine metabolism. Genetic silencing of isocitrate dehydrogenase-1 or isocitrate dehydrogenase-2 impaired reductive labeling of tricarboxylic acid (TCA) cycle intermediates in vivo and suppressed growth of tumors generated from tumorgraft-derived cells. Glutaminase inhibition reduced the contribution of glutamine to the TCA cycle and resulted in modest suppression of tumorgraft growth. Infusions with [amide-15N]glutamine revealed persistent amidotransferase activity during glutaminase inhibition, and blocking these activities with the amidotransferase inhibitor JHU-083 also reduced tumor growth in both immunocompromised and immunocompetent mice. We conclude that ccRCC tumorgrafts catabolize glutamine via multiple pathways, perhaps explaining why it has been challenging to achieve therapeutic responses in patients by inhibiting glutaminase.
  4. Chem Sci. 2022 Nov 23. 13(45): 13489-13498
      The mitochondrial electron transport chain comprises a series of protein complexes embedded in the inner mitochondrial membrane that generate a proton motive force via oxidative phosphorylation, ultimately generating ATP. These protein complexes can oligomerize to form larger structures called supercomplexes. Cardiolipin (CL), a conical lipid, unique within eukaryotes to the inner mitochondrial membrane, has proven essential in maintaining the stability and function of supercomplexes. Monolysocardiolipin (MLCL) is a CL variant that accumulates in people with Barth syndrome (BTHS). BTHS is caused by defects in CL biosynthesis and characterised by abnormal mitochondrial bioenergetics and destabilised supercomplexes. However, the mechanisms by which MLCL causes pathogenesis remain unclear. Here, multiscale molecular dynamics characterise the interactions of CL and MLCL with yeast and mammalian mitochondrial supercomplexes containing complex III (CIII) and complex IV (CIV). Coarse-grained simulations reveal that both CL and MLCL bind to sites at the interface between CIII and CIV of the supercomplex. Free energy perturbation calculations show that MLCL interaction is weaker than that of CL and suggest that interaction with CIV drives this difference. Atomistic contact analyses show that, although interaction with CIII is similar for CL and MLCL, CIV makes more contacts with CL than MLCL, demonstrating that CL is a more successful "glue" between the two complexes. Simulations of the human CIII2CIV supercomplex show that this interface site is maintained between species. Our study suggests that MLCL accumulation in people with BTHS disrupts supercomplex stability by formation of relatively weak interactions at the interface lipid binding site.
  5. Cell Metab. 2022 Dec 08. pii: S1550-4131(22)00502-2. [Epub ahead of print]
      The mammalian succinate dehydrogenase (SDH) complex has recently been shown as capable of operating bidirectionally. Here, we develop a method (Q-Flux) capable of measuring absolute rates of both forward (VSDH(F)) and reverse (VSDH(R)) flux through SDH in vivo while also deconvoluting the amount of glucose derived from four discreet carbon sources in the liver. In validation studies, a mitochondrial uncoupler increased net SDH flux by >100% in awake rodents but also increased SDH cycling. During hyperglucagonemia, attenuated pyruvate cycling enhances phosphoenolpyruvate carboxykinase efficiency to drive increased gluconeogenesis, which is complemented by increased glutaminase (GLS) flux, methylmalonyl-CoA mutase (MUT) flux, and glycerol conversion to glucose. During hyperinsulinemic-euglycemic clamp, both pyruvate carboxylase and GLS are suppressed, while VSDH(R) is increased. Unstimulated MUT is a minor anaplerotic reaction but is readily induced by small amounts of propionate, which elicits glucagon-like metabolic rewiring. Taken together, Q-Flux yields a comprehensive picture of hepatic mitochondrial metabolism and should be broadly useful to researchers.
    Keywords:  anaplerosis; glucagon; glutaminase; insulin; mass spectrometry; metabolic flux analysis; methylmalonyl-CoA mutase; mitochondrial metabolism; propionate; succinate dehydrogenase
  6. Adv Exp Med Biol. 2022 ;1395 367-372
      In intact mitochondria, the transport of electrons, respiration and generation of proton gradients across the inner membrane (proton motive force) are mutually coupled, according to Peter Mitchell's hypothesis on oxidative phosphorylation. Thus, the inhibition of electron transport at either respiratory complex III or IV in the electron transport chain leads to failure in producing proton motive force along with the abolition of respiration. Here, we determined the mitochondrial membrane potential (MMP), as a measure of proton motive force, and cellular respiration in various cultured cells and demonstrated that inhibition of complex IV by KCN abolished mitochondrial respiration while MMP was sustained. These results are unexpected and appear incompatible with Mitchell's chemiosmotic hypothesis.
    Keywords:  Electron transport; Mitchell’s chemiosmotic hypothesis; Mitochondria; Proton motive force
  7. Biochim Biophys Acta Bioenerg. 2022 Dec 09. pii: S0005-2728(22)00421-2. [Epub ahead of print]1864(2): 148951
      Respiratory complex I in mitochondria and bacteria catalyzes the transfer of electrons from NADH to quinone (Q). The free energy available from the reaction is used to pump protons and to establish a membrane proton electrochemical gradient, which drives ATP synthesis. Even though several high-resolution structures of complex I have been resolved, how Q reduction is linked with proton pumping, remains unknown. Here, microsecond long molecular dynamics (MD) simulations were performed on Yarrowia lipolytica complex I structures where Q molecules have been resolved in the ~30 Å long Q tunnel. MD simulations of several different redox/protonation states of Q reveal the coupling between the Q dynamics and the restructuring of conserved loops and ion pairs. Oxidized quinone stabilizes towards the N2 FeS cluster, a binding mode not previously described in Yarrowia lipolytica complex I structures. On the other hand, reduced (and protonated) species tend to diffuse towards the Q binding sites closer to the tunnel entrance. Mechanistic and physiological relevance of these results are discussed.
    Keywords:  Bioenergetics; Electron transfer; Molecular dynamics simulations; Proton pumping; Semiquinone
  8. Acta Histochem. 2022 Dec 09. pii: S0065-1281(22)00145-3. [Epub ahead of print]125(1): 151986
      Lon protease 1(Lonp1) is an ATP-dependent protease located in the mitochondrial matrix and plays a crucial role in preserving normal mitochondrial function. Lonp1 overexpression is associated with tumorigenesis in various cancer types, including cervical cancer. In the present study, we show that the Lonp1 content is elevated in cervical cancer tissues compared to cervical paracancerous tissues. Conversely, Lonp1 knockdown suppresses cervical cancer cell proliferation, migration and invasion but promotes apoptosis. Mechanistically, Lonp1 knockdown decreases area of mitochondrial networks and induces mitochondrial depolarization. Furthermore, Lonp1 inhibition reduces the level of LC3-II/I, PINK1 and Parkin, but promotes the level of p62. Collectively, our study suggests that the anti-cancer effect caused by Lonp1 downregulation likely contributes to mitochondrial remodeling and suppression of autophagy and mitophagy.
    Keywords:  Autophagy; Lonp1; Mitochondria
  9. Trends Cell Biol. 2022 Dec 12. pii: S0962-8924(22)00259-8. [Epub ahead of print]
      The mitochondrial calcium uniporter (MCU) controls mitochondrial bioenergetics, and its activity varies greatly between tissues. Here, we highlight a recently identified MCU-EMRE-UCP1 complex, named thermoporter, in the adaptive thermogenesis of brown adipose tissue (BAT). The thermoporter enhances MCU activity to promote thermogenic metabolism, demonstrating a BAT-specific regulation for MCU activity.
  10. Cancer Cell Int. 2022 Dec 12. 22(1): 402
      BACKGROUND: Metabolic reprogramming is an important issue in tumor biology. A recently-identified actor in this regard is the molecular chaperone TRAP1, that is considered an oncogene in several cancers for its high expression but an oncosuppressor in others with predominant oxidative metabolism. TRAP1 is mainly localized in mitochondria, where it interacts with respiratory complexes, although alternative localizations have been described, particularly on the endoplasmic reticulum, where it interacts with the translational machinery with relevant roles in protein synthesis regulation.RESULTS: Herein we show that, inside mitochondria, TRAP1 binds the complex III core component UQCRC2 and regulates complex III activity. This decreases respiration rate during basal conditions but allows sustained oxidative phosphorylation when glucose is limiting, a condition in which the direct TRAP1-UQCRC2 binding is disrupted, but not TRAP1-complex III binding. Interestingly, several complex III components and assembly factors show an inverse correlation with survival and response to platinum-based therapy in high grade serous ovarian cancers, where TRAP1 inversely correlates with stage and grade and directly correlates with survival. Accordingly, drug-resistant ovarian cancer cells show high levels of complex III components and high sensitivity to complex III inhibitory drug antimycin A.
    CONCLUSIONS: These results shed new light on the molecular mechanisms involved in TRAP1-dependent regulation of cancer cell metabolism and point out a potential novel target for metabolic therapy in ovarian cancer.
    Keywords:  Ovarian cancer; Platinum resistance; Respiratory complex III; TRAP1
  11. Biochim Biophys Acta Bioenerg. 2022 Dec 09. pii: S0005-2728(22)00420-0. [Epub ahead of print]1864(2): 148950
      The F1FO-ATP synthase uses the energy stored in the electrochemical proton gradient to synthesize ATP. This complex is found in the inner mitochondrial membrane as a monomer and dimer. The dimer shows higher ATPase activity than the monomer and is essential for cristae folding. The monomer-monomer interface is constituted by subunits a, i/j, e, g, and k. The role of the subunit g in a strict respiratory organism is unknown. A gene knockout was generated in Ustilago maydis to study the role of subunit g on mitochondrial metabolism and cristae architecture. Deletion of the ATP20 gene, encoding the g subunit, did not affect cell growth or glucose consumption, but biomass production was lower in the mutant strain (gΔ strain). Ultrastructure observations showed that mitochondrial size and cristae shape were similar in wild-type and gΔ strains. The mitochondrial membrane potential in both strains had a similar magnitude, but oxygen consumption was higher in the WT strain. ATP synthesis was 20 % lower in the gΔ strain. Additionally, the mutant strain expressed the alternative oxidase in the early stages of growth (exponential phase), probably as a response to ROS stress. Dimer from mutant strain was unstable to digitonin solubilization, avoiding its isolation and kinetic characterization. The isolated monomeric state activated by n-dodecyl-β-D-maltopyranoside showed similar kinetic constants to the monomer from the WT strain. A decrease in mitochondrial ATP synthesis and the presence of the AOX during the exponential growth phase suggests that deletion of the g gene induces ROS stress.
    Keywords:  ATPase activity; Dimer of complex V; Oxidative phosphorylation; ROS production; g subunit
  12. J Mol Cell Biol. 2022 Dec 16. pii: mjac070. [Epub ahead of print]
      Mitochondria in many fungi are inherited uniparentally during meiosis. It has remained unclear whether parental mitochondria in the fission yeast Schizosaccharomyces pombe are inherited uniparentally or biparentally. Here, we assessed the mixing of parental mitochondria carefully by live-cell microscopy and developed an algorithm to determine the degree of mitochondrial mixing in a quantitative manner. We found that parental mitochondria in fission yeast cells were mixed progressively as meiosis progressed. Moreover, we established that mitochondrial fission and the size of the conjugation neck are the limiting factors in restricting the mixing of parental mitochondria. We further employed a combination of quantitative polymerase chain reaction, fluorescent live-cell microscopy, and transmission electron microscopy approaches to examine the mitochondrial inheritance of progeny cells derived from a cross between wild-type and Rho0 (mitochondrial DNA absent) cells. The results show that all progeny cells of the cross carry mitochondrial DNA. Hence, our data support the model in which parental mitochondria in the fission yeast Schizosaccharomyces pombe are inherited biparentally during meiosis.
    Keywords:  fission yeast; mitochondria; mitochondrial fission; mitochondrial inheritance
  13. Trends Endocrinol Metab. 2022 Dec 15. pii: S1043-2760(22)00218-1. [Epub ahead of print]
      A de novo heterozygous variant in the catalytic subunit of mitochondrial F1FO-ATPase has been recently discovered by Ganetzky et al. to be the main cause of an autosomal dominant syndrome of hypermetabolism associated with defective ATP production. We describe how the 'rotor free-wheeling' causes this F1FO-ATPase dysfunction in primary congenital hypothyroidism.
    Keywords:  ATP production; F(1)F(O)-ATPase; congenital hypermetabolism; mitochondrial bioenergetics; mutation
  14. Chem Biol Drug Des. 2022 Dec 16.
      Two natural products, bongkrekic acid and carboxyatractyloside, are known to specifically inhibit the mitochondrial ADP/ATP carrier from its matrix side and cytosolic side, respectively, in concentration ranges of 10-6 M. In the present study, we investigated the manner of action of a synthetic bongkrekic acid derivative, KH-17, lacking three methyl groups, one methoxy group and five internal double bonds, on the mitochondrial ADP/ATP carrier. At slightly acidic pH, KH-17 inhibited mitochondrial [3 H]ADP uptake, but its inhibitory action was about 10 times weaker than that of its parental compound, bongkrekic acid. The main site of action of KH-17 was confirmed as the matrix side of the ADP/ATP carrier by experiments using submitochondrial particles, which have an inside-out orientation of the inner mitochondrial membrane. However, when we added KH-17 to mitochondria at neutral pH, it had a weak inhibitory effect on [3 H]ADP uptake, and its inhibitory strength was similar to that of bongkrekic acid. These results indicated that KH-17 weakly inhibits the ADP/ATP carrier not only from the matrix side, but also from the cytosolic side. To ascertain whether this interpretation was correct, we examined the effects of KH-17 and carboxyatractyloside on mitochondrial [3 H]ADP uptake at two [3 H]ADP concentrations. We found that both KH-17 and carboxyatractyloside showed a stronger inhibitory effect at the lower [3 H]ADP concentration. Therefore, we concluded that the bongkrekic acid derivative, KH-17, weakly inhibits the mitochondrial ADP/ATP carrier from both sides of the inner mitochondrial membrane. These results suggested that the elimination of three methyl groups, one methoxy group and five internal double bonds present in bongkrekic acid altered its manner of action towards the mitochondrial ADP/ATP carrier. Our data will help to improve our understanding of the interaction between bongkrekic acid and the mitochondrial ADP/ATP carrier.
    Keywords:  ADP/ATP carrier; bongkrekic acid; inhibitor; mitochondria
  15. Cancers (Basel). 2022 Nov 24. pii: 5779. [Epub ahead of print]14(23):
      The human isocitrate dehydrogenase (IDH) gene encodes for the isoenzymes IDH1, 2, and 3, which catalyze the conversion of isocitrate and α-ketoglutarate (α-KG) and are required for normal mammalian metabolism. Isocitrate dehydrogenase 1 and 2 catalyze the reversible conversion of isocitrate to α-KG. Isocitrate dehydrogenase 3 is the key enzyme that mediates the production of α-KG from isocitrate in the tricarboxylic acid (TCA) cycle. In the TCA cycle, the decarboxylation reaction catalyzed by isocitrate dehydrogenase mediates the conversion of isocitrate to α-KG accompanied by dehydrogenation, a process commonly known as oxidative decarboxylation. The formation of 6-C isocitrate from α-KG and CO2 catalyzed by IDH is termed reductive carboxylation. This IDH-mediated reversible reaction is of great importance in tumor cells. We outline the role of the various isocitrate dehydrogenase isoforms in cancer, discuss the metabolic implications of interference with IDH, summarize therapeutic interventions targeting changes in IDH expression, and highlight areas for future research.
    Keywords:  cancer metabolism; isocitrate dehydrogenase (IDH); nicotinamide adenine dinucleotide phosphate (NADPH); oxidative decarboxylation; reductive carboxylation; α-ketoglutarate (α-KG)
  16. J Cell Sci. 2022 Dec 01. pii: jcs259748. [Epub ahead of print]135(23):
      Mitochondria are crucial organelles that play a central role in various cell signaling and metabolic pathways. A healthy mitochondrial population is maintained through a series of quality control pathways and requires a fine-tuned balance between mitochondrial biogenesis and degradation. Defective targeting of dysfunctional mitochondria to lysosomes through mitophagy has been linked to several diseases, but the underlying mechanisms and the relative importance of distinct mitophagy pathways in vivo are largely unknown. In this Cell Science at a Glance and the accompanying poster, we describe our current understanding of how parts of, or whole, mitochondria are recognized by the autophagic machinery and targeted to lysosomes for degradation. We also discuss how this might be regulated under different physiological conditions to maintain mitochondrial and cellular health.
    Keywords:  BNIP3; HIF1; Mitochondria; Mitophagy; NIX; PINK1; Parkin; SLR; Selective autophagy
  17. EMBO J. 2022 Dec 14. e111372
      Mitophagy, the elimination of mitochondria via the autophagy-lysosome pathway, is essential for the maintenance of cellular homeostasis. The best characterised mitophagy pathway is mediated by stabilisation of the protein kinase PINK1 and recruitment of the ubiquitin ligase Parkin to damaged mitochondria. Ubiquitinated mitochondrial surface proteins are recognised by autophagy receptors including NDP52 which initiate the formation of an autophagic vesicle around the mitochondria. Damaged mitochondria also generate reactive oxygen species (ROS) which have been proposed to act as a signal for mitophagy, however the mechanism of ROS sensing is unknown. Here we found that oxidation of NDP52 is essential for the efficient PINK1/Parkin-dependent mitophagy. We identified redox-sensitive cysteine residues involved in disulphide bond formation and oligomerisation of NDP52 on damaged mitochondria. Oligomerisation of NDP52 facilitates the recruitment of autophagy machinery for rapid mitochondrial degradation. We propose that redox sensing by NDP52 allows mitophagy to function as a mechanism of oxidative stress response.
    Keywords:  NDP52; autophagy; mitophagy; p62; redox
  18. Commun Biol. 2022 Dec 13. 5(1): 1366
      Cellular metabolism influences immune cell function, with mitochondrial fatty acid β-oxidation and oxidative phosphorylation required for multiple immune cell phenotypes. Carnitine palmitoyltransferase 1a (Cpt1a) is considered the rate-limiting enzyme for mitochondrial metabolism of long-chain fatty acids, and Cpt1a deficiency is associated with infant mortality and infection risk. This study was undertaken to test the hypothesis that impairment in Cpt1a-dependent fatty acid oxidation results in increased susceptibility to infection. Screening the Cpt1a gene for common variants predicted to affect protein function revealed allele rs2229738_T, which was associated with pneumonia risk in a targeted human phenome association study. Pharmacologic inhibition of Cpt1a increases mortality and impairs control of the infection in a murine model of bacterial pneumonia. Susceptibility to pneumonia is associated with blunted neutrophilic responses in mice and humans that result from impaired neutrophil trafficking to the site of infection. Chemotaxis responsible for neutrophil trafficking requires Cpt1a-dependent mitochondrial fatty acid oxidation for amplification of chemoattractant signals. These findings identify Cpt1a as a potential host determinant of infection susceptibility and demonstrate a requirement for mitochondrial fatty acid oxidation in neutrophil biology.
  19. Cancers (Basel). 2022 Nov 29. pii: 5900. [Epub ahead of print]14(23):
      The commonly used therapeutic management of PC involves androgen deprivation therapy (ADT) followed by treatment with AR signaling inhibitors (ARSI). However, nearly all patients develop drug-resistant disease, with a median progression-free survival of less than 2 years in chemotherapy-naïve men. Acetyl-coenzyme A (acetyl-CoA) is a central metabolic signaling molecule with key roles in biosynthetic processes and cancer signaling. In signaling, acetyl-CoA serves as the acetyl donor for acetylation, a critical post-translational modification. Acetylation affects the androgen receptor (AR) both directly and indirectly increasing expression of AR dependent genes. Our studies reveal that PC cells respond to the treatment with ARSI by increasing expression of ATP-citrate lyase (ACLY), a major enzyme responsible for cytosolic acetyl-CoA synthesis, and up-regulation of acetyl-CoA intracellular levels. Inhibition of ACLY results in a significant suppression of ligand-dependent and -independent routes of AR activation. Accordingly, the addition of exogenous acetyl-CoA, or its precursor acetate, augments AR transcriptional activity and diminishes the anti-AR activity of ARSI. Taken together, our findings suggest that PC cells respond to antiandrogens by increasing activity of the acetyl-coA pathway in order to reinstate AR signaling.
    Keywords:  abiraterone; acetyl-coenzyme A; androgen receptor; enzalutamide; prostate cancer
  20. Biomed Pharmacother. 2022 Dec 12. pii: S0753-3322(22)01504-9. [Epub ahead of print]158 114115
      Phosphoglycerate kinase 1(PGK1) is an important enzyme in the metabolic glycolysis pathway. Nowadays, PGK1 is an appealing therapeutic target for multiple cancers. However, no effective inhibitor of PGK1 has been reported. In this study, we demonstrate that Ilicicolin H a 5-(4-hydroxyphenyl)-pyridone with a decalin ring system and a non-ATP-competitive inhibitor of PGK1, inhibits the proliferation and promotes apoptosis of Hepatocellular carcinoma (HCC). Many cancer cells display enhanced glycolysis which is critical for tumor development. Here we identified that Ilicicolin H can target PGK1 in vitro to inhibit the lactate production and glucose uptake of HCC cells. These findings suggest that the PGK1 inhibitor- Ilicicolin H is a promising anticancer agent and may provide a better therapeutic strategy for HCC treatment in the future.
    Keywords:  Anti-cancer; Hepatocellular carcinoma; Inhibitor; PGK1
  21. FEBS J. 2022 Dec 14.
      Advances in cancer biology over the past decades have revealed that metabolic adaptation of cancer cells is an essential aspect of tumorigenesis. However, recent insights into tumor metabolism in vivo have revealed dissimilarities with results obtained in vitro. This is partly due to the reductionism of in vitro cancer models that struggle to reproduce the complexity of tumor tissues. This review describes some of the discrepancies in cancer cell metabolism between in vitro and in vivo conditions, and presents current methodological approaches and tools used to bridge the gap with the clinically relevant microenvironment. As such, these approaches should generate new knowledge that could be more effectively translated into therapeutic opportunities.
    Keywords:  3D models; metabolic heterogeneity; metabolic sensors; tissue culture media; tumor cell metabolism
  22. PeerJ. 2022 ;10 e14285
      Colorectal carcinoma (CRC) is the second most frequent cancer worldwide. MiR-491-3p, a tumor-suppressive microRNA (miRNA, miR), has been revealed to be abnormally expressed in CRC tissues. Meanwhile, up-regulated ubiquitous mitochondrial creatine kinase (uMtCK) contributes to CRC cell proliferation. Here we aim to explore whether aberrant miR-491-3p expression promotes CRC progression through regulating uMtCK. To this end, miR-491-3p and uMtCK levels were assessed in CRC tissues using quantitative real-time PCR (qRT-PCR). The biological roles of miR-491-3p and uMtCK in regulating CRC growth were evaluated using colony formation assay and mouse Xenograft tumour model. We found that miR-491-3p expression was decreased in CRC tissues compared with matched para-cancerous tissues, whereas uMtCK expression was increased. Functionally, miR-491-3p overexpression repressed SW480 cell growth, whereas miR-491-3p depletion accelerated SW620 cell proliferation and growth. Inversely, uMtCK positively regulated CRC cell proliferation. Mechanistically, miR-491-3p post-transcriptionally downregulated uMtCK expression by binding to 3'-UTR of uMtCK. Consequently, restoring uMtCK expression markedly eliminated the role of miR-491-3p in suppressing CRC growth. Collectively, miR-491-3p functions as a tumour suppressor gene by repressing uMtCK, and may be a potential target for CRC treatment.
    Keywords:  Cell proliferation; Colorectal cancer; miR-491-3p; microRNA; uMtCK
  23. Am J Cancer Res. 2022 ;12(11): 4977-4987
      Pancreatic cancer (PC) is the fourth-most-deadly cancer in the United States with a 5-year survival rate of only 8%. The majority of patients with locally advanced pancreatic cancer undergo chemotherapy and/or radiation therapy (RT). However, current treatments are inadequate and novel strategies are desperately required. 3-Bromopyruvate (3-BP) is a promising anticancer drug against pancreatic cancer. It exerts potent anticancer effects by inhibiting hexokinase II enzyme (HK2) of the glycolytic pathway in cancer cells while not affecting the normal cells. 3-BP killed 95% of Panc-2 cells at 15 μM concentration and severely inhibited ATP production by disrupting the interaction between HK2 and mitochondrial Voltage Dependent Anion Channel-1 (VDAC1) protein. Electron microscopy data revealed that 3-BP severely damaged mitochondrial membrane in cancer cells. We further examined therapeutic effect of 3-BP in syngeneic mouse pancreatic cancer model by treating animals with 10, 15 and 20 mg/kg dose. 3-BP at 15 & 20 mg/kg dose level significantly reduced tumor growth by approximately 75-80% in C57BL/6 female mice. Immunohistochemistry data showed complete inhibition of hexokinase II (HK2) and TGFβ, in animals treated with 3-BP drug. We also observed enhanced expression of active caspase-3 in tumor tissues exhibited apoptotic death. Flow Cytometry analysis showed significant inhibition in MDSC (CD11b) population in treated tumor which may have allowed infiltration of CD8+ T cells and inhibited tumor growth. Notably, metabolomic data also revealed severe inhibition in glycolysis, NADP, ATP and lactic acid production in cancer cells treated with 40 μM 3-BP. Importantly, we also observed inhibition in lactic acid production responsible for tumor aggression. These results provide new evidence that 3-BP severely inhibit glucose metabolism in cancer cells by blocking hexokinase II, and disrupting mitochondria by suppressing BCL2L1 in pancreatic cancer.
    Keywords:  3-Bromopyruvate; ATP production; Pancreatic cancer; apoptosis; glycolysis; hexokinase II; mitochondria; voltage dependent anion channel 1 (VDAC1)
  24. Blood. 2022 Dec 12. pii: blood.2021011094. [Epub ahead of print]
      Myeloid neoplasms with erythroid or megakaryocytic differentiation include pure erythroid leukemia (PEL), myelodysplastic syndrome (MDS) with erythroid features, and acute megakaryoblastic leukemia (FAB M7) and are characterized by poor prognosis and limited treatment options. Here, we investigate the drug sensitivity landscape of these rare malignancies. We show that acute myeloid leukemia (AML) cells with erythroid or megakaryocytic differentiation depend on the anti-apoptotic protein BCL-XL, rather than BCL-2, using combined ex vivo drug sensitivity testing, genetic perturbation, and transcriptomic profiling. High-throughput screening of > 500 compounds identified the BCL-XL-selective inhibitor A-1331852 and navitoclax as highly effective against erythroid/megakaryoblastic leukemia cell lines. In contrast, these AML subtypes were resistant to the BCL-2 inhibitor venetoclax used clinically in the treatment of AML. Consistently, genome-scale CRISPR-Cas9 and RNAi screening data demonstrated striking essentiality of BCL2L1 encoding BCL-XL, but not BCL2 or MCL1, for the survival of erythroid/megakaryoblastic leukemia cell lines. Single-cell and bulk transcriptomics of patient samples with erythroid and megakaryoblastic leukemias identified high BCL2L1 expression compared to other subtypes of AML and other hematological malignancies, where BCL2 and MCL1 were more prominent. BCL-XL inhibition effectively killed blasts in AML patient samples with erythroid or megakaryocytic differentiation ex vivo and reduced tumor burden in a mouse erythroleukemia xenograft model. Combining BCL-XL inhibitor with the JAK inhibitor ruxolitinib showed synergistic and durable responses in cell lines. Our results suggest targeting BCL-XL as a potential therapy option in erythroid/megakaryoblastic leukemias and highlight an AML subgroup with potentially reduced sensitivity to venetoclax-based treatments.
  25. J Exp Med. 2023 Mar 06. pii: e20221316. [Epub ahead of print]220(3):
      Fatty acid uptake is essential for cell physiological function, but detailed mechanisms remain unclear. Here, we generated an acetyl-CoA carboxylases (ACC1/2) double-knockout cell line, which lacked fatty acid biosynthesis and survived on serum fatty acids and was used to screen for fatty acid uptake inhibitors. We identified a Food and Drug Administration-approved tricyclic antidepressant, nortriptyline, that potently blocked fatty acid uptake both in vitro and in vivo. We also characterized underlying mechanisms whereby nortriptyline provoked lysosomes to release protons and induce cell acidification to suppress macropinocytosis, which accounted for fatty acid endocytosis. Furthermore, nortriptyline alone or in combination with ND-646, a selective ACC1/2 inhibitor, significantly repressed tumor growth, lipogenesis, and hepatic steatosis in mice. Therefore, we show that cells actively take up fatty acids through macropinocytosis, and we provide a potential strategy suppressing tumor growth, lipogenesis, and hepatic steatosis through controlling the cellular level of fatty acids.
  26. Cells. 2022 Dec 06. pii: 3940. [Epub ahead of print]11(23):
      Lonp1 is a mitochondrial protease that degrades oxidized and damaged proteins, assists protein folding, and contributes to the maintenance of mitochondrial DNA. A higher expression of LonP1 has been associated with higher tumour aggressiveness. Besides the full-length isoform (ISO1), we identified two other isoforms of Lonp1 in humans, resulting from alternative splicing: Isoform-2 (ISO2) lacking aa 42-105 and isoform-3 (ISO3) lacking aa 1-196. An inspection of the public database TSVdb showed that ISO1 was upregulated in lung, bladder, prostate, and breast cancer, ISO2 in all the cancers analysed (including rectum, colon, cervical, bladder, prostate, breast, head, and neck), ISO3 did not show significant changes between cancer and normal tissue. We overexpressed ISO1, ISO2, and ISO3 in SW620 cells and found that the ISO1 isoform was exclusively mitochondrial, ISO2 was present in the organelle and in the cytoplasm, and ISO3 was exclusively cytoplasmatic. The overexpression of ISO1 and, at a letter extent, of ISO2 enhanced basal, ATP-linked, and maximal respiration without altering the mitochondria number or network, mtDNA amount. or mitochondrial dynamics. A higher extracellular acidification rate was observed in ISO1 and ISO2, overexpressing cells, suggesting an increase in glycolysis. Cells overexpressing the different isoforms did not show a difference in the proliferation rate but showed a great increase in anchorage-independent growth. ISO1 and ISO2, but not ISO3, determined an upregulation of EMT-related proteins, which appeared unrelated to higher mitochondrial ROS production, nor due to the activation of the MEK ERK pathway, but rather to global metabolic reprogramming of cells.
    Keywords:  Lon protease; SW620; mitochondria; mitochondrial DNA
  27. Blood Adv. 2022 Dec 15. pii: bloodadvances.2022008221. [Epub ahead of print]
      Venetoclax is an effective treatment for certain blood cancers, such as chronic lymphocytic leukemia (CLL) and acute myeloid leukemia (AML). However, most patients relapse while on venetoclax and further treatment options become limited. Combining venetoclax with immunotherapies is an attractive approach; however, a detailed understanding of how venetoclax treatment impacts normal immune cells in patients is lacking. In this study, we performed deep profiling of peripheral blood cells from CLL and AML patients before and after short-term treatment with venetoclax using mass cytometry (CyTOF) and found no impact on the concentrations of key T cell subsets nor their expression of checkpoint molecules. We also analyzed peripheral blood from breast cancer patients receiving venetoclax long-term using a single-cell multi-omics approach (CITE-seq) and functional assays. We found significant depletion of B cell populations with low expression of MCL-1 relative to other immune cells, attended by extensive transcriptomic changes. By contrast, there was less impact on circulating T cells and natural killer (NK) cells, with no changes in their subset composition, transcriptome or function following venetoclax treatment. Our data indicate that venetoclax has minimal impact on circulating T or NK cells, supporting the rationale of combining this BH3 mimetic drug with cancer immunotherapies for more durable anti-tumour responses.
  28. Cancer Res. 2022 Dec 13. pii: CAN-22-2145. [Epub ahead of print]
      Metabolic reprogramming can contribute to colorectal cancer (CRC) progression and therapy resistance. Identification of key regulators of CRC metabolism could provide new approaches to improve treatment and reduce recurrence. Here, we demonstrate a critical role for the COP9 signalosome subunit CSN6 in rewiring nucleotide metabolism in CRC. Transcriptomic analysis of CRC patient samples revealed a correlation between CSN6 expression and purine and pyrimidine metabolism. A colitis-associated colorectal cancer model established that Csn6 intestinal conditional deletion decreased tumor development and altered nucleotide metabolism. CSN6 knockdown increased the chemosensitivity of CRC cells in vitro and in vivo, which could be partially reversed with nucleoside supplementation. Isotope metabolite tracing showed that CSN6 loss reduced de novo nucleotide synthesis. Mechanistically, CSN6 upregulated purine and pyrimidine biosynthesis by increasing expression of PHGDH, a key enzyme in the serine synthesis pathway. CSN6 inhibited β-Trcp-mediated DDX5 polyubiquitination and degradation, which in turn promoted DDX5-mediated PHGDH mRNA stabilization, leading to metabolic reprogramming and CRC progression. Butyrate treatment decreased CSN6 expression and improved chemotherapy efficacy. These findings unravel the oncogenic role of CSN6 in regulating nucleotide metabolism and chemosensitivity in CRC.
  29. Br J Cancer. 2022 Dec 16.
      CONTEXT: Many cancer patients who initially respond to chemotherapy eventually develop chemoresistance, and to address this, we previously conducted a RNAi screen to identify genes contributing to resistance. One of the hits from the screen was branched-chain α-keto acid dehydrogenase kinase (BCKDK). BCKDK controls the metabolism of branched-chain amino acids (BCAAs) through phosphorylation and inactivation of the branched-chain α-keto acid dehydrogenase complex (BCKDH), thereby inhibiting catabolism of BCAAs.METHODS: We measured the impact on paclitaxel sensitivity of inhibiting BCKDK in ovarian and breast cancer cell lines.
    RESULTS: Inhibition of BCKDK using siRNA or two chemical inhibitors (BCKDKi) was synergistic with paclitaxel in both breast and ovarian cancer cells. BCKDKi reduced levels of BCAA and the addition of exogenous BCAA suppressed this synergy. BCKDKi inactivated the mTORC1-Aurora pathway, allowing cells to overcame M-phase arrest induced by paclitaxel. In some cases, cells almost completed cytokinesis, then reverted to a single cell, resulting in multinucleate cells.
    CONCLUSION: BCKDK is an attractive target to augment the sensitivity of cancer cells to paclitaxel.
  30. Cell Rep. 2022 Dec 13. pii: S2211-1247(22)01697-7. [Epub ahead of print]41(11): 111809
      The gut microbiota influences acetylation on host histones by fermenting dietary fiber into butyrate. Although butyrate could promote histone acetylation by inhibiting histone deacetylases, it may also undergo oxidation to acetyl-coenzyme A (CoA), a necessary cofactor for histone acetyltransferases. Here, we find that epithelial cells from germ-free mice harbor a loss of histone H4 acetylation across the genome except at promoter regions. Using stable isotope tracing in vivo with 13C-labeled fiber, we demonstrate that the microbiota supplies carbon for histone acetylation. Subsequent metabolomic profiling revealed hundreds of labeled molecules and supported a microbial contribution to host fatty acid metabolism, which declined in response to colitis and correlated with reduced expression of genes involved in fatty acid oxidation. These results illuminate the flow of carbon from the diet to the host via the microbiota, disruptions to which may affect energy homeostasis in the distal gut and contribute to the development of colitis.
    Keywords:  CP: Microbiology; colitis; epigenetics; fatty acid metabolism; histone acetylation; host-microbiota interactions
  31. J Gerontol A Biol Sci Med Sci. 2022 Dec 13. pii: glac247. [Epub ahead of print]
      Aging is the largest risk factor for the development of cancer. A a growing body of literature indicates that aging and cancer often play a somewhat reciprocal relationship at various times. On the one hand, aging is a "driver" of cancer, and on the other, cancer is a "disease driver" of aging. Here, we synthesize our reflections on the current literature linking cancer and aging, with an eye on fundamental aging processes, such as cellular senescence. Additionally, we consider how interventions that target fundamental aging processes can potentially transform cancer care, from preventing cancer development and progression to reducing the burden of accelerated aging in cancer survivors. Finally, we conclude with a reflection highlighting our vision for future directions to advance the science of cancer and aging and its applicability to improve the care of older adults with cancer.
    Keywords:  aging; aging biology; cancer; geroscience; hallmarks
  32. Cells. 2022 Nov 24. pii: 3754. [Epub ahead of print]11(23):
      The ability of animal orthologs of human mitochondrial transcription factor A (hTFAM) to support the replication of human mitochondrial DNA (hmtDNA) does not follow a simple pattern of phylogenetic closeness or sequence similarity. In particular, TFAM from chickens (Gallus gallus, chTFAM), unlike TFAM from the "living fossil" fish coelacanth (Latimeria chalumnae), cannot support hmtDNA replication. Here, we implemented the recently developed GeneSwap approach for reverse genetic analysis of chTFAM to obtain insights into this apparent contradiction. By implementing limited "humanization" of chTFAM focused either on amino acid residues that make DNA contacts, or the ones with significant variances in side chains, we isolated two variants, Ch13 and Ch22. The former has a low mtDNA copy number (mtCN) but robust respiration. The converse is true of Ch22. Ch13 and Ch22 complement each other's deficiencies. Opposite directionalities of changes in mtCN and respiration were also observed in cells expressing frog TFAM. This led us to conclude that TFAM's contributions to mtDNA replication and respiratory chain biogenesis are genetically separable. We also present evidence that TFAM residues that make DNA contacts play the leading role in mtDNA replication. Finally, we present evidence for a novel mode of regulation of the respiratory chain biogenesis by regulating the supply of rRNA subunits.
    Keywords:  GeneSwap approach; TFAM; mtDNA instability; mtDNA replication; mtDNA transcription
  33. Nat Struct Mol Biol. 2022 Dec 15.
      Glycerol-3-phosphate acyltransferase (GPAT)1 is a mitochondrial outer membrane protein that catalyzes the first step of de novo glycerolipid biosynthesis. Hepatic expression of GPAT1 is linked to liver fat accumulation and the severity of nonalcoholic fatty liver diseases. Here we present the cryo-EM structures of human GPAT1 in substrate analog-bound and product-bound states. The structures reveal an N-terminal acyltransferase domain that harbors important catalytic motifs and a tightly associated C-terminal domain that is critical for proper protein folding. Unexpectedly, GPAT1 has no transmembrane regions as previously proposed but instead associates with the membrane via an amphipathic surface patch and an N-terminal loop-helix region that contains a mitochondrial-targeting signal. Combined structural, computational and functional studies uncover a hydrophobic pathway within GPAT1 for lipid trafficking. The results presented herein lay a framework for rational inhibitor development for GPAT1.
  34. Haematologica. 2022 Dec 15.
      The BCL2 inhibitor venetoclax has revolutionized the treatment of acute myeloid leukemia (AML) patients not benefitting from intensive chemotherapy. Nevertheless, treatment failure remains a challenge, and predictive markers are needed, particularly for relapsed or refractory (R/R) AML. Ex vivo drug sensitivity testing may correlate with outcomes, but its prospective predictive value remains unexplored. Here we report the results of the first stage of the prospective Phase 2 VenEx trial evaluating the utility and predictiveness of venetoclax sensitivity testing using different cell culture conditions and cell viability assays in patients receiving venetoclax-azacitidine (NCT04267081). Participants with de novo AML ineligible for intensive chemotherapy, R/R AML, or secondary AML were included. The primary endpoint was the treatment response in ex vivo sensitive participants and the key secondary endpoints were the correlation of sensitivity with responses and survival. Venetoclax sensitivity testing was successful in 38/39 participants. Experimental conditions significantly influenced predictive accuracy. Blast-specific venetoclax sensitivity measured in conditioned medium most accurately correlated with treatment outcomes; 88% of sensitive participants achieved treatment response. Median survival was significantly longer for ex vivo sensitive participants (14. 6 months for s ensitive, 3. 5 for insensitive, p < 0 . 001). T his analysis illustrates the feasibility of integrating drug-response profiling into clinical practice and demonstrates excellent predictivity.