bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2024–04–07
forty-five papers selected by
Kıvanç Görgülü, Technical University of Munich



  1. BMC Cancer. 2024 Apr 04. 24(1): 414
       BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) presents with a high mortality rate. Two important features of PDAC contribute to this poor outcome. The first is metastasis which occurs in ~ 80% of PDAC patients. The second is cachexia, which compromises treatment tolerance for patients and reduces their quality of life. Although various mouse models of PDAC exist, recapitulating both metastatic and cachectic features have been challenging.
    METHODS: Here, we optimize an orthotopic mouse model of PDAC by altering several conditions, including the subcloning of parental murine PDAC cells, implantation site, number of transplanted cells, and age of recipient mice. We perform spatial profiling to compare primary and metastatic immune microenvironments and RNA sequencing to gain insight into the mechanisms of muscle wasting in PDAC-induced cachexia, comparing non-metastatic to metastatic conditions.
    RESULTS: These modifications extend the time course of the disease and concurrently increase the rate of metastasis to approximately 70%. Furthermore, reliable cachexia endpoints are achieved in both PDAC mice with and without metastases, which is reminiscent of patients. We also find that cachectic muscles from PDAC mice with metastasis exhibit a similar transcriptional profile to muscles derived from mice and patients without metastasis.
    CONCLUSION: Together, this model is likely to be advantageous in both advancing our understanding of the mechanism of PDAC cachexia, as well as in the evaluation of novel therapeutics.
    Keywords:  Cachexia; Metastasis; Pancreatic ductal adenocarcinoma; Skeletal muscle
    DOI:  https://doi.org/10.1186/s12885-024-12104-0
  2. bioRxiv. 2024 Mar 20. pii: 2024.03.18.585580. [Epub ahead of print]
      Pancreatic ductal adenocarcinoma (PDAC) subsists in a nutrient-deregulated microenvironment, making it particularly susceptible to treatments that interfere with cancer metabolism 1 2 . For example, PDAC utilizes and is dependent on high levels of autophagy and other lysosomal processes 3-5 . Although targeting these pathways has shown potential in preclinical studies, progress has been hampered by the challenge of identifying and characterizing favorable targets for drug development 6 . Here, we characterize PIKfyve, a lipid kinase integral to lysosomal functioning 7 , as a novel and targetable vulnerability in PDAC. In human patient and murine PDAC samples, we discovered that PIKFYVE is overexpressed in PDAC cells compared to adjacent normal cells. Employing a genetically engineered mouse model, we established the essential role of PIKfyve in PDAC progression. Further, through comprehensive metabolic analyses, we found that PIKfyve inhibition obligated PDAC to upregulate de novo lipid synthesis, a relationship previously undescribed. PIKfyve inhibition triggered a distinct lipogenic gene expression and metabolic program, creating a dependency on de novo lipid metabolism pathways, by upregulating genes such as FASN and ACACA . In PDAC, the KRAS-MAPK signaling pathway is a primary driver of de novo lipid synthesis, specifically enhancing FASN and ACACA levels. Accordingly, the simultaneous targeting of PIKfyve and KRAS-MAPK resulted in the elimination of tumor burden in a syngeneic orthotopic model and tumor regression in a xenograft model of PDAC. Taken together, these studies suggest that disrupting lipid metabolism through PIKfyve inhibition induces synthetic lethality in conjunction with KRAS-MAPK-directed therapies for PDAC.
    DOI:  https://doi.org/10.1101/2024.03.18.585580
  3. Nat Metab. 2024 Apr 02.
      White adipocytes function as major energy reservoirs in humans by storing substantial amounts of triglycerides, and their dysfunction is associated with metabolic disorders; however, the mechanisms underlying cellular specialization during adipogenesis remain unknown. Here, we generate a spatiotemporal proteomic atlas of human adipogenesis, which elucidates cellular remodelling as well as the spatial reorganization of metabolic pathways to optimize cells for lipid accumulation and highlights the coordinated regulation of protein localization and abundance during adipocyte formation. We identify compartment-specific regulation of protein levels and localization changes of metabolic enzymes to reprogramme branched-chain amino acids and one-carbon metabolism to provide building blocks and reduction equivalents. Additionally, we identify C19orf12 as a differentiation-induced adipocyte lipid droplet protein that interacts with the translocase of the outer membrane complex of lipid droplet-associated mitochondria and regulates adipocyte lipid storage by determining the capacity of mitochondria to metabolize fatty acids. Overall, our study provides a comprehensive resource for understanding human adipogenesis and for future discoveries in the field.
    DOI:  https://doi.org/10.1038/s42255-024-01025-8
  4. bioRxiv. 2024 Mar 18. pii: 2024.03.17.585402. [Epub ahead of print]
      Driver gene mutations can increase the metastatic potential of the primary tumor 1-3 , but their role in sustaining tumor growth at metastatic sites is poorly understood. A paradigm of such mutations is inactivation of SMAD4 - a transcriptional effector of TGFβ signaling - which is a hallmark of multiple gastrointestinal malignancies 4,5 . SMAD4 inactivation mediates TGFβ's remarkable anti-to pro-tumorigenic switch during cancer progression and can thus influence both tumor initiation and metastasis 6-14 . To determine whether metastatic tumors remain dependent on SMAD4 inactivation, we developed a mouse model of pancreatic ductal adenocarcinoma (PDAC) that enables Smad4 depletion in the pre-malignant pancreas and subsequent Smad4 reactivation in established metastases. As expected, Smad4 inactivation facilitated the formation of primary tumors that eventually colonized the liver and lungs. By contrast, Smad4 reactivation in metastatic disease had strikingly opposite effects depending on the tumor's organ of residence: suppression of liver metastases and promotion of lung metastases. Integrative multiomic analysis revealed organ-specific differences in the tumor cells' epigenomic state, whereby the liver and lungs harbored chromatin programs respectively dominated by the KLF and RUNX developmental transcription factors, with Klf4 depletion being sufficient to reverse Smad4 's tumor-suppressive activity in liver metastases. Our results show how epigenetic states favored by the organ of residence can influence the function of driver genes in metastatic tumors. This organ-specific gene-chromatin interplay invites consideration of anatomical site in the interpretation of tumor genetics, with implications for the therapeutic targeting of metastatic disease.
    DOI:  https://doi.org/10.1101/2024.03.17.585402
  5. Mol Metab. 2024 Apr 01. pii: S2212-8778(24)00061-9. [Epub ahead of print] 101930
       OBJECTIVE: Tumour progression drives profound alterations in host metabolism, such as adipose tissue depletion, an early event of cancer cachexia. As fatty acid consumption by cancer cells increases upon acidosis of the tumour microenvironment, we reasoned that fatty acids derived from distant adipose lipolysis may sustain tumour fatty acid craving, leading to the adipose tissue loss observed in cancer cachexia.
    METHODS: To evaluate the pro-lipolytic capacities of acid-exposed cancer cells, primary mouse adipocytes from subcutaneous and visceral adipose tissue were exposed to pH-matched conditioned medium from human and murine acid-exposed cancer cells (pH 6.5), compared to naive cancer cells (pH 7.4). To further address the role of tumoral acidosis on adipose tissue loss, a pH-low insertion peptide was injected into tumour-bearing mice, and tumoral acidosis was neutralised with a sodium bicarbonate buffer. Prolipolytic mediators were identified by transcriptomic approaches and validated on murine and human adipocytes.
    RESULTS: Here, we reveal that acid-exposed cancer cells promote lipolysis from subcutaneous and visceral adipocyte and that dampening acidosis in vivo inhibits adipose tissue depletion. We further found a set of well-known prolipolytic factors enhanced upon acidosis adaptation and unravelled a role for β-glucuronidase as a promising new actor in adipocyte lipolysis.
    CONCLUSIONS: Tumoral acidosis promotes the mobilization of fatty acids derived from adipocytes via the release of soluble factors by cancer cells. Our work paves the way for dual therapeutic approaches aimed at tackling cachexia by targeting the tumour acidic compartment.
    Keywords:  Adipocytes; Adipose tissue; Beta-glucuronidase; Cancer cachexia; Lipolysis; Tumor acidosis
    DOI:  https://doi.org/10.1016/j.molmet.2024.101930
  6. J Cachexia Sarcopenia Muscle. 2024 Apr 04.
       BACKGROUND: Cancer cachexia is a life-threatening, inflammation-driven wasting syndrome that remains untreatable. Adiponectin, the most abundant adipokine, plays an important role in several metabolic processes as well as in inflammation modulation. Our aim was to test whether administration of AdipoRon (AR), a synthetic agonist of the adiponectin receptors, prevents the development of cancer cachexia and its related muscle atrophy.
    METHODS: The effect of AR on cancer cachexia was investigated in two distinct murine models of colorectal cancer. First, 7-week-old CD2F1 male mice were subcutaneously injected with colon-26 carcinoma cells (C26) or vehicle (CT). Six days after injection, mice were treated for 5 days with AdipoRon (50 mg/kg/day; C26 + AR) or the corresponding vehicle (CT and C26). Additionally, a genetic model, the ApcMin/+ mouse, that develops spontaneously numerous intestinal polyps, was used. Eight-week-old male ApcMin/+ mice were treated with AdipoRon (50 mg/kg/day; Apc + AR) or the corresponding vehicle (Apc) over a period of 12 weeks, with C57BL/6J wild-type mice used as controls. In both models, several parameters were assessed in vivo: body weight, grip strength and serum parameters, as well as ex vivo: molecular changes in muscle, fat and liver.
    RESULTS: The protective effect of AR on cachexia development was observed in both cachectic C26 and ApcMin/+ mice. In these mice, AR administration led to a significant alleviation of body weight loss and muscle wasting, together with rescued muscle strength (P < 0.05 for all). In both models, AR had a strong anti-inflammatory effect, reflected by lower systemic interleukin-6 levels (-55% vs. C26, P < 0.001 and -80% vs. Apc mice, P < 0.05), reduced muscular inflammation as indicated by lower levels of Socs3, phospho-STAT3 and Serpina3n, an acute phase reactant (P < 0.05 for all). In addition, AR blunted circulating levels of corticosterone (-46% vs. C26 mice, P < 0.001 and -60% vs. Apc mice, P < 0.05), the predominant murine glucocorticoid known to induce muscle atrophy. Accordingly, key glucocorticoid-responsive factors implicated in atrophy programmes were-or tended to be-significantly blunted in skeletal muscle by AR. Finally, AR protected against lipid metabolism alterations observed in ApcMin/+ mice, as it mitigated the increase in circulating triglyceride levels (-38%, P < 0.05) by attenuating hepatic triglyceride synthesis and fatty acid uptake by the liver.
    CONCLUSIONS: Altogether, these results show that AdipoRon rescued the cachectic phenotype by alleviating body weight loss and muscle atrophy, along with restraining inflammation and hypercorticism in preclinical murine models. Therefore, AdipoRon could represent an innovative therapeutic strategy to counteract cancer cachexia.
    Keywords:  adiponectin; cachexia; cancer; inflammation; skeletal muscle
    DOI:  https://doi.org/10.1002/jcsm.13454
  7. Dev Cell. 2024 Apr 02. pii: S1534-5807(24)00183-7. [Epub ahead of print]
      UBE2F, a neddylation E2, neddylates CUL5 to activate cullin-RING ligase-5, upon coupling with neddylation E3 RBX2/SAG. Whether and how UBE2F controls pancreatic tumorigenesis is previously unknown. Here, we showed that UBE2F is essential for the growth of human pancreatic cancer cells with KRAS mutation. In the mouse KrasG12D pancreatic ductal adenocarcinoma (PDAC) model, Ube2f deletion suppresses cerulein-induced pancreatitis, and progression of acinar-to-ductal metaplasia (ADM) and pancreatic intraepithelial neoplasia. Mechanistically, Ube2f deletion inactivates the Mapk-c-Myc signals via blocking ubiquitylation of Diras2, a substrate of CRL5Asb11 E3 ligase. Biologically, DIRAS2 suppresses growth and survival of human pancreatic cancer cells harboring mutant KRAS, and Diras2 deletion largely rescues the phenotypes induced by Ube2f deletion. Collectively, Ube2f or Diras2 plays a tumor-promoting or tumor-suppressive role in the mouse KrasG12D PDAC model, respectively. The UBE2F-CRL5ASB11 axis could serve as a valid target for pancreatic cancer, whereas the levels of UBE2F or DIRAS2 may serve as prognostic biomarkers for PDAC patients.
    Keywords:  CRL5; Kras; MLN4924; Ube2F; neddylation; oncogene; pancreatic tumorigenesis; pancreatitis; tumor suppressor; ubiquitylation
    DOI:  https://doi.org/10.1016/j.devcel.2024.03.018
  8. Trends Cancer. 2024 Apr 04. pii: S2405-8033(24)00053-0. [Epub ahead of print]
      Metastasis has a major part in the severity of disease and lethality of cancer. Circulating tumour cells (CTCs) represent a reservoir of metastatic precursors in circulation, most of which cannot survive due to hostile conditions in the bloodstream. Surviving cells colonise a secondary site based on a combination of physical, metabolic, and oxidative stress protection states required for that environment. Recent advances in CTC isolation methods and high-resolution 'omics technologies are revealing specific metabolic pathways that support this selection of CTCs. In this review, we discuss recent advances in our understanding of CTC biology and discoveries of adaptations in metabolic pathways during their selection. Understanding these traits and delineating mechanisms by which they confer acquired resistance or vulnerability in CTCs is crucial for developing successful prognostic and therapeutic strategies in cancer.
    Keywords:  cancer; circulating tumour cells (CTCs); ferroptosis; metabolism; metastasis; oxidative stress
    DOI:  https://doi.org/10.1016/j.trecan.2024.03.004
  9. J Cell Biol. 2024 Jul 01. pii: e202309057. [Epub ahead of print]223(7):
      Autophagy serves as a stress response pathway by mediating the degradation of cellular material within lysosomes. In autophagy, this material is encapsulated in double-membrane vesicles termed autophagosomes, which form from precursors referred to as phagophores. Phagophores grow by lipid influx from the endoplasmic reticulum into Atg9-positive compartments and local lipid synthesis provides lipids for their expansion. How phagophore nucleation and expansion are coordinated with lipid synthesis is unclear. Here, we show that Faa1, an enzyme activating fatty acids, is recruited to Atg9 vesicles by directly binding to negatively charged membranes with a preference for phosphoinositides such as PI3P and PI4P. We define the membrane-binding surface of Faa1 and show that its direct interaction with the membrane is required for its recruitment to phagophores. Furthermore, the physiological localization of Faa1 is key for its efficient catalysis and promotes phagophore expansion. Our results suggest a positive feedback loop coupling phagophore nucleation and expansion to lipid synthesis.
    DOI:  https://doi.org/10.1083/jcb.202309057
  10. Elife. 2024 Apr 04. pii: RP92559. [Epub ahead of print]13
      Oncogenic mutations in KRAS are among the most common in cancer. Classical models suggest that loss of epithelial characteristics and the acquisition of mesenchymal traits are associated with cancer aggressiveness and therapy resistance. However, the mechanistic link between these phenotypes and mutant KRAS biology remains to be established. Here, we identify STAT3 as a genetic modifier of TGF-beta-induced epithelial to mesenchymal transition. Gene expression profiling of pancreatic cancer cells identifies more than 200 genes commonly regulated by STAT3 and oncogenic KRAS. Functional classification of the STAT3-responsive program reveals its major role in tumor maintenance and epithelial homeostasis. The signatures of STAT3-activated cell states can be projected onto human KRAS mutant tumors, suggesting that they faithfully reflect characteristics of human disease. These observations have implications for therapeutic intervention and tumor aggressiveness.
    Keywords:  KRAS; STAT3; cancer biology; cancer dependency; epithelial carcinogenesis; mouse
    DOI:  https://doi.org/10.7554/eLife.92559
  11. bioRxiv. 2024 Mar 21. pii: 2024.03.20.585922. [Epub ahead of print]
      Cancer cell fate has been widely ascribed to mutational changes within protein-coding genes associated with tumor suppressors and oncogenes. In contrast, the mechanisms through which the biophysical properties of membrane lipids influence cancer cell survival, dedifferentiation and metastasis have received little scrutiny. Here, we report that cancer cells endowed with a high metastatic ability and cancer stem cell-like traits employ ether lipids to maintain low membrane tension and high membrane fluidity. Using genetic approaches and lipid reconstitution assays, we show that these ether lipid-regulated biophysical properties permit non-clathrin-mediated iron endocytosis via CD44, leading directly to significant increases in intracellular redox-active iron and enhanced ferroptosis susceptibility. Using a combination of in vitro three-dimensional microvascular network systems and in vivo animal models, we show that loss of ether lipids also strongly attenuates extravasation, metastatic burden and cancer stemness. These findings illuminate a mechanism whereby ether lipids in carcinoma cells serve as key regulators of malignant progression while conferring a unique vulnerability that can be exploited for therapeutic intervention.
    DOI:  https://doi.org/10.1101/2024.03.20.585922
  12. Cancer Discov. 2024 Apr 04. 14(4): 669-673
       SUMMARY: The field of cancer neuroscience has begun to define the contributions of nerves to cancer initiation and progression; here, we highlight the future directions of basic and translational cancer neuroscience for malignancies arising outside of the central nervous system.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1495
  13. bioRxiv. 2024 Mar 11. pii: 2024.03.08.584164. [Epub ahead of print]
      The intrinsic pathways that control membrane organization in immune cells and the impact of such pathways on cellular function are not well defined. Here we report that the non-vesicular cholesterol transporter Aster-A links plasma membrane (PM) cholesterol availability in T cells to immune signaling and systemic metabolism. Aster-A is recruited to the PM during T-cell receptor (TCR) activation, where it facilitates the removal of newly generated "accessible" membrane cholesterol. Loss of Aster-A leads to excess PM cholesterol accumulation, resulting in enhanced TCR nano-clustering and signaling, and Th17 cytokine production. Finally, we show that the mucosal Th17 response is restrained by PM cholesterol remodeling. Ablation of Aster-A in T cells leads to enhanced IL-22 production, reduced intestinal fatty acid absorption, and resistance to diet-induced obesity. These findings delineate a multi-tiered regulatory scheme linking immune cell lipid flux to nutrient absorption and systemic physiology.
    DOI:  https://doi.org/10.1101/2024.03.08.584164
  14. Cell Rep Med. 2024 Apr 02. pii: S2666-3791(24)00166-6. [Epub ahead of print] 101498
      Progressive weakness and muscle loss are associated with multiple chronic conditions, including muscular dystrophy and cancer. Cancer-associated cachexia, characterized by dramatic weight loss and fatigue, leads to reduced quality of life and poor survival. Inflammatory cytokines have been implicated in muscle atrophy; however, available anticytokine therapies failed to prevent muscle wasting in cancer patients. Here, we show that oncostatin M (OSM) is a potent inducer of muscle atrophy. OSM triggers cellular atrophy in primary myotubes using the JAK/STAT3 pathway. Identification of OSM targets by RNA sequencing reveals the induction of various muscle atrophy-related genes, including Atrogin1. OSM overexpression in mice causes muscle wasting, whereas muscle-specific deletion of the OSM receptor (OSMR) and the neutralization of circulating OSM preserves muscle mass and function in tumor-bearing mice. Our results indicate that activated OSM/OSMR signaling drives muscle atrophy, and the therapeutic targeting of this pathway may be useful in preventing muscle wasting.
    Keywords:  JAK/STAT3 signaling; cancer cachexia; oncostatin M; skeletal muscle atrophy
    DOI:  https://doi.org/10.1016/j.xcrm.2024.101498
  15. bioRxiv. 2024 Mar 13. pii: 2024.03.12.584718. [Epub ahead of print]
      Autophagy is an intracellular degradation process that maintains homeostasis, responds to stress, and plays key roles in the prevention of aging and disease. Autophagosome biogenesis, vesicle rocketing, and autolysosome tubulation are controlled by multiple actin nucleation factors, but the impact of actin assembly on completion of the autophagic pathway is not well understood. Here we studied autophagosome and lysosome remodeling in fibroblasts harboring an inducible knockout (iKO) of the Arp2/3 complex, an essential actin nucleator. Arp2/3 complex ablation resulted in increased basal levels of autophagy receptors and lipidated membrane proteins from the LC3 and GABARAP families. Under both steady-state and starvation conditions, Arp2/3 iKO cells accumulated abnormally high numbers of autolysosomes, suggesting a defect in autophagic flux. The inability of Arp2/3 complex-deficient cells to complete autolysosome degradation and turnover is explained by the presence of damaged, leaky lysosomes. In cells treated with an acute lysosomal membrane-damaging agent, the Arp2/3-activating protein WHAMM is recruited to lysosomes, where Arp2/3 complex-dependent actin assembly is crucial for restoring intact lysosomal structure. These results establish the Arp2/3 complex as a central player late in the canonical autophagy pathway and reveal a new role for the actin nucleation machinery in maintaining lysosomal integrity.
    DOI:  https://doi.org/10.1101/2024.03.12.584718
  16. Cancer Cell. 2024 Mar 31. pii: S1535-6108(24)00090-4. [Epub ahead of print]
      The tumor microenvironment (TME) in pancreatic ductal adenocarcinoma (PDAC) involves a significant accumulation of cancer-associated fibroblasts (CAFs) as part of the host response to tumor cells. The origins and functions of transcriptionally diverse CAF populations in PDAC remain poorly understood. Tumor cell-intrinsic genetic mutations and epigenetic dysregulation may reshape the TME; however, their impacts on CAF heterogeneity remain elusive. SETD2, a histone H3K36 trimethyl-transferase, functions as a tumor suppressor. Through single-cell RNA sequencing, we identify a lipid-laden CAF subpopulation marked by ABCA8a in Setd2-deficient pancreatic tumors. Our findings reveal that tumor-intrinsic SETD2 loss unleashes BMP2 signaling via ectopic gain of H3K27Ac, leading to CAFs differentiation toward lipid-rich phenotype. Lipid-laden CAFs then enhance tumor progression by providing lipids for mitochondrial oxidative phosphorylation via ABCA8a transporter. Together, our study links CAF heterogeneity to epigenetic dysregulation in tumor cells, highlighting a previously unappreciated metabolic interaction between CAFs and pancreatic tumor cells.
    Keywords:  OXPHOS; Pancreatic cancer; SETD2; cell communication; epigenetic dysregulation; lipid-laden CAF
    DOI:  https://doi.org/10.1016/j.ccell.2024.03.005
  17. Mol Cancer. 2024 04 01. 23(1): 68
      Cancer progression is continuously controlled by the immune system which can identify and destroy nascent tumor cells or inhibit metastatic spreading. However, the immune system and its deregulated activity in the tumor microenvironment can also promote tumor progression favoring the outgrowth of cancers capable of escaping immune control, in a process termed cancer immunoediting. This process, which has been classified into three phases, i.e. "elimination", "equilibrium" and "escape", is influenced by several cancer- and microenvironment-dependent factors. Senescence is a cellular program primed by cells in response to different pathophysiological stimuli, which is based on long-lasting cell cycle arrest and the secretion of numerous bioactive and inflammatory molecules. Because of this, cellular senescence is a potent immunomodulatory factor promptly recruiting immune cells and actively promoting tissue remodeling. In the context of cancer, these functions can lead to both cancer immunosurveillance and immunosuppression. In this review, the authors will discuss the role of senescence in cancer immunoediting, highlighting its context- and timing-dependent effects on the different three phases, describing how senescent cells promote immune cell recruitment for cancer cell elimination or sustain tumor microenvironment inflammation for immune escape. A potential contribution of senescent cells in cancer dormancy, as a mechanism of therapy resistance and cancer relapse, will be discussed with the final objective to unravel the immunotherapeutic implications of senescence modulation in cancer.
    Keywords:  Cancer; Dormancy; Immunoediting; Immunosurveillance; SASP; Senescence; Senotherapy
    DOI:  https://doi.org/10.1186/s12943-024-01973-5
  18. Methods Mol Biol. 2024 ;2797 351-362
      KRAS mutations occur in approximately ~50% of colorectal cancers (CRCs) and are associated with poor prognosis and resistance to therapy. While these most common mutations found at amino acids G12, G13, Q61, and A146 have long been considered oncogenic drivers of CRC, emerging clinical data suggest that each mutation may possess different biological functions, resulting in varying consequences in oncogenesis. Currently, the mechanistic underpinnings associated with each allelic variation remain unclear. Elucidating the unique effectors of each KRAS mutant could both increase the understanding of KRAS biology and provide a basis for allele-specific therapeutic opportunities. Biotinylation identification (BioID) is a method to label and identify proteins located in proximity of a protein of interest. These proteins are captured through the strong interaction between the biotin label and streptavidin bead and subsequently identified by mass spectrometry. Here, we developed a protocol using CRISPR-mediated gene editing to generate endogenous BioID2-tagged KrasG12D and KrasG12V isogenic murine colon epithelial cell lines to identify unique protein proximity partners by BioID.
    Keywords:  BioID; BirA; Colorectal cancer; Gene editing; KRAS; Proximity labeling; Signal transduction; Small G proteins
    DOI:  https://doi.org/10.1007/978-1-0716-3822-4_25
  19. Proc Natl Acad Sci U S A. 2024 Apr 09. 121(15): e2400675121
      Atherosclerosis is fueled by a failure to resolve lipid-driven inflammation within the vasculature that drives plaque formation. Therapeutic approaches to reverse atherosclerotic inflammation are needed to address the rising global burden of cardiovascular disease (CVD). Recently, metabolites have gained attention for their immunomodulatory properties, including itaconate, which is generated from the tricarboxylic acid-intermediate cis-aconitate by the enzyme Immune Responsive Gene 1 (IRG1/ACOD1). Here, we tested the therapeutic potential of the IRG1-itaconate axis for human atherosclerosis. Using single-cell RNA sequencing (scRNA-seq), we found that IRG1 is up-regulated in human coronary atherosclerotic lesions compared to patient-matched healthy vasculature, and in mouse models of atherosclerosis, where it is primarily expressed by plaque monocytes, macrophages, and neutrophils. Global or hematopoietic Irg1-deficiency in mice increases atherosclerosis burden, plaque macrophage and lipid content, and expression of the proatherosclerotic cytokine interleukin (IL)-1β. Mechanistically, absence of Irg1 increased macrophage lipid accumulation, and accelerated inflammation via increased neutrophil extracellular trap (NET) formation and NET-priming of the NLRP3-inflammasome in macrophages, resulting in increased IL-1β release. Conversely, supplementation of the Irg1-itaconate axis using 4-octyl itaconate (4-OI) beneficially remodeled advanced plaques and reduced lesional IL-1β levels in mice. To investigate the effects of 4-OI in humans, we leveraged an ex vivo systems-immunology approach for CVD drug discovery. Using CyTOF and scRNA-seq of peripheral blood mononuclear cells treated with plasma from CVD patients, we showed that 4-OI attenuates proinflammatory phospho-signaling and mediates anti-inflammatory rewiring of macrophage populations. Our data highlight the relevance of pursuing IRG1-itaconate axis supplementation as a therapeutic approach for atherosclerosis in humans.
    Keywords:  atherosclerosis; immunometabolism; innate immunity; macrophage; neutrophil
    DOI:  https://doi.org/10.1073/pnas.2400675121
  20. J Phys Chem B. 2024 Apr 04.
      The lateral mobility of molecules within the cell membrane is ultimately governed by the local environment of the membrane. Confined regions induced by membrane structures, such as protein aggregates or the actin meshwork, occur over a wide range of length scales and can impede or steer the diffusion of membrane components. However, a detailed picture of the origins and nature of these confinement effects remains elusive. Here, we prepare model lipid systems on substrates patterned with confined domains of varying geometries constructed with different materials to explore the influences of physical boundary conditions and specific molecular interactions on diffusion. We demonstrate a platform that is capable of significantly altering and steering the long-range diffusion of lipids by using simple oxide deposition approaches, enabling us to systematically explore how confinement size and shape impact diffusion over multiple length scales. While we find that a "boundary condition" description of the system captures underlying trends in some cases, we are also able to directly compare our systems to analytical models, revealing the unexpected breakdown of several approximate solutions. Our results highlight the importance of considering the length scale dependence when discussing properties such as diffusion.
    DOI:  https://doi.org/10.1021/acs.jpcb.3c07388
  21. Methods Mol Biol. 2024 ;2797 1-12
      RAS research has entered the world of translational and clinical science. Progress has been based on our appreciation of the role of RAS mutations in different types of cancer and the effects of these mutations on the biochemical, structural, and biophysical properties of the RAS proteins themselves, particularly KRAS, on which most attention has been focused. This knowledge base, while still growing, has enabled creative chemical approaches to targeting KRAS directly. Our understanding of RAS signaling pathways in normal and cancer cells plays an important role for developing RAS inhibitors but also continues to reveal new approaches to targeting RAS through disruption of signaling complexes and downstream pathways.
    Keywords:  GTP hydrolysis; Mutation frequency in cancer; Oncogenic alleles; RAS; Targeting RAS
    DOI:  https://doi.org/10.1007/978-1-0716-3822-4_1
  22. Cancer Res. 2024 Apr 01. 84(7): 956-957
      I was recently surprised to hear a medical doctor on a TV show refute the role of stress in cancer, assuming that "the whole population would have cancer if this was the case." This statement illustrates a long and winding road since Hippocrates suggested the potential relationship between cancer and psychologic disturbances. The 20th and 21st centuries have finally witnessed the evidence of how physical or psychosocial stress situations contribute to the development and progression of cancer, and it is now assumed that psychologic stress does affect multiple aspects of cancer such as angiogenesis, immunologic escape, invasion, and metastasis. The 2010 publication by Sloan and colleagues in Cancer Research achieved a mechanistic step toward the understanding of how physical distress enhances metastasis through perturbation of the tumor immune system and paves the way for future cancer research in psychoneuroimmunology. This Landmark commentary places this publication in the historical context of science, discusses major advances in the field, and asks questions to be answered while drawing perspectives on the key role of the peripheral and central nervous systems in cancer. See related article by Sloan and colleagues, Cancer Res 2010;70:7042-52.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-24-0628
  23. Cancer Discov. 2024 Apr 04. 14(4): 663-668
       SUMMARY: We are building the world's first Virtual Child-a computer model of normal and cancerous human development at the level of each individual cell. The Virtual Child will "develop cancer" that we will subject to unlimited virtual clinical trials that pinpoint, predict, and prioritize potential new treatments, bringing forward the day when no child dies of cancer, giving each one the opportunity to lead a full and healthy life.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1500
  24. Cancer Discov. 2024 Apr 04. 14(4): 630-634
       SUMMARY: Beyond lipid membrane compartments, cells including cancer cells utilize various membraneless compartments, often termed biomolecular condensates, to regulate or organize key cellular processes underlying physiologic or pathologic phenotypes. In this commentary, the emergence of biomolecular condensation in cancer biology is highlighted, with a focus on key unanswered questions and with implications for improving the understanding of cancer pathogenesis and developing innovative cancer management strategies.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1551
  25. Cell Death Discov. 2024 Apr 05. 10(1): 166
      Although increased aerobic glycolysis is common in various cancers, pancreatic ductal adenocarcinoma (PDAC) cells can survive a state of glycolysis suppression. We aimed to identify potential therapeutic targets in glycolysis-suppressed PDAC cells. By screening anticancer metabolic compounds, we identified SP-2509, an inhibitor of lysine-specific histone demethylase 1A (LSD1), which dramatically decreased the growth of PDAC PANC-1 cells and showed an anti-tumoral effect in tumor-bearing mice. The growth of glycolysis-suppressed PANC-1 cells was also inhibited by another LSD1 inhibitor, OG-L002. Similarly, the other two PDAC cells (PK-1 and KLM-1) with suppressed glycolysis exhibited anticancer effects against SP-2509. However, the anticancer effects on PDAC cells were unrelated to LSD1. To investigate how PDAC cells survive in a glycolysis-suppressed condition, we conducted proteomic analyses. These results combined with our previous findings suggested that glucose-starvation causes PDAC cells to enhance mitochondrial oxidative phosphorylation. In particular, mitochondrial fatty acid metabolism was identified as a key factor contributing to the survival of PDAC cells under glycolysis suppression. We further demonstrated that SP-2509 and OG-L002 disturbed fatty acid metabolism and induced lipid droplet accumulation through the impairment of lipophagy, but not bulk autophagy. These findings indicate a significant potential association of lipophagy and anticancer effects in glycolysis-suppressed PDAC cells, offering ideas for new therapeutic strategies for PDAC by dual inhibition of glycolysis and fatty acids metabolism.
    DOI:  https://doi.org/10.1038/s41420-024-01933-4
  26. J Biochem. 2024 Apr 02. pii: mvae032. [Epub ahead of print]
      Senescent cells accumulate in various organs with aging, and its accumulation induces chronic inflammation and age-related physiological dysfunctions. Several remodeling of intracellular environments has been identified in senescent cells, including enlargement of cell / nuclear size and intracellular acidification. Although these alterations of intracellular environments were reported to be involved in unique characteristics of senescent cells, the contribution of intracellular acidification to senescence-associated cellular phenotypes is poorly understood. Here, we identified that upregulation of TXNIP and its paralog ARRDC4 as a hallmark of intracellular acidification in addition to KGA-type GLS1. These genes were also upregulated in response to senescence-associated intracellular acidification. Neutralization of the intracellular acidic environment ameliorated not only senescence-related upregulation of TXNIP, ARRDC4, and KGA, but also inflammation-related genes, possibly through suppression of PDK-dependent anaerobic glycolysis. Furthermore, we found that expression of the intracellular acidification-induced genes, TXNIP and ARRDC4, correlated with inflammatory gene expression in heterogeneous senescent cell population in vitro and even in vivo, implying that the contribution of intracellular pH to senescence-associated cellular features, such as SASP.
    Keywords:  SASP; glycolysis; intracellular acidification; senescence
    DOI:  https://doi.org/10.1093/jb/mvae032
  27. Histopathology. 2024 Apr 04.
       AIMS: Following the increased use of neoadjuvant therapy for pancreatic cancer, grading of tumour regression (TR) has become part of routine diagnostics. However, it suffers from marked interobserver variation, which is mainly ascribed to the subjectivity of the defining criteria of the categories in TR grading systems. We hypothesized that a further cause for the interobserver variation is the use of divergent and nonspecific morphological criteria to identify tumour regression.
    METHODS AND RESULTS: Twenty treatment-naïve pancreatic cancers and 20 pancreatic cancers treated with neoadjuvant chemotherapy were reviewed by three experienced pancreatic pathologists who, blinded for treatment status, categorized each tumour as treatment-naïve or neoadjuvantly treated, and annotated all tissue areas they considered showing tumour regression. Only 50%-65% of the cases were categorized correctly, and the annotated tissue areas were highly discrepant (only 3%-41% overlap). When the prevalence of various morphological features deemed to indicate TR was compared between treatment-naïve and neoadjuvantly treated tumours, only one pattern, characterized by reduced cancer cell density and prominent stroma affecting a large area of the tumour bed, occurred significantly more frequently, but not exclusively, in the neoadjuvantly treated group. Finally, stromal features, both morphological and biological, were investigated as possible markers for tumour regression, but failed to distinguish TR from native tumour stroma.
    CONCLUSION: There is considerable divergence in opinion between pathologists when it comes to the identification of tumour regression. Reliable identification of TR is only possible if it is extensive, while lesser degrees of treatment effect cannot be recognized with certainty.
    Keywords:  neoadjuvant chemotherapy; observer variation; pancreatic cancer; pathology; therapeutic effect
    DOI:  https://doi.org/10.1111/his.15190
  28. EMBO Mol Med. 2024 Apr 03.
      Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with an overall 5-year survival rate of <12% due to the lack of effective treatments. Novel treatment strategies are urgently needed. Here, PKMYT1 is identified through genome-wide CRISPR screens as a non-mutant, genetic vulnerability of PDAC. Higher PKMYT1 expression levels indicate poor prognosis in PDAC patients. PKMYT1 ablation inhibits tumor growth and proliferation in vitro and in vivo by regulating cell cycle progression and inducing apoptosis. Moreover, pharmacological inhibition of PKMYT1 shows efficacy in multiple PDAC cell models and effectively induces tumor regression without overt toxicity in PDAC cell line-derived xenograft and in more clinically relevant patient-derived xenograft models. Mechanistically, in addition to its canonical function of phosphorylating CDK1, PKMYT1 functions as an oncogene to promote PDAC tumorigenesis by regulating PLK1 expression and phosphorylation. Finally, TP53 function and PRKDC activation are shown to modulate the sensitivity to PKMYT1 inhibition. These results define PKMYT1 dependency in PDAC and identify potential therapeutic strategies for clinical translation.
    Keywords:  CRISPR Screens; PKMYT1; Pancreatic Ductal Adenocarcinoma
    DOI:  https://doi.org/10.1038/s44321-024-00060-y
  29. Cancer Discov. 2024 Apr 04. 14(4): 610-614
       SUMMARY: Cancer is traditionally perceived through a genetic lens, with therapeutic strategies targeting oncogenic driver mutations. We advocate an overarching framework recognizing tumors as comprising driver, passenger, and trailer cell states: Tailoring therapies to simultaneously target driver genetics and cell states may enhance effectiveness and durability.
    SIGNIFICANCE: We redefine cancer progression by introducing a model that categorizes tumor cells into "driver," "passenger," and "trailer" phenotypes, expanding the focus on genetic aberrations to cellular behavior. This approach offers a roadmap to guide refining therapeutic strategies for more precise and durable cancer treatments that address tumor heterogeneity and plasticity.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1510
  30. Lipids Health Dis. 2024 Apr 02. 23(1): 94
       BACKGROUND: Insulin-stimulated glucose uptake into skeletal muscle occurs via translocation of GLUT4 from intracellular storage vesicles to the plasma membrane. Elevated free fatty acid (FFA) availability via a lipid infusion reduces glucose disposal, but this occurs in the absence of impaired proximal insulin signalling. Whether GLUT4 localisation to the plasma membrane is subsequently affected by elevated FFA availability is not known.
    METHODS: Trained (n = 11) and sedentary (n = 10) individuals, matched for age, sex and body mass index, received either a 6 h lipid or glycerol infusion in the setting of a concurrent hyperinsulinaemic-euglycaemic clamp. Sequential muscle biopsies (0, 2 and 6 h) were analysed for GLUT4 membrane localisation and microvesicle size and distribution using immunofluorescence microscopy.
    RESULTS: At baseline, trained individuals had more small GLUT4 spots at the plasma membrane, whereas sedentary individuals had larger GLUT4 spots. GLUT4 localisation with the plasma membrane increased at 2 h (P = 0.04) of the hyperinsulinemic-euglycemic clamp, and remained elevated until 6 h, with no differences between groups or infusion type. The number of GLUT4 spots was unchanged at 2 h of infusion. However, from 2 to 6 h there was a decrease in the number of small GLUT4 spots at the plasma membrane (P = 0.047), with no differences between groups or infusion type.
    CONCLUSION: GLUT4 localisation with the plasma membrane increases during a hyperinsulinemic-euglycemic clamp, but this is not altered by elevated FFA availability. GLUT4 appears to disperse from small GLUT4 clusters located at the plasma membrane to support glucose uptake during a hyperinsulinaemic-euglycaemic clamp.
    Keywords:  GLUT4 translocation; Lipid infusion; Skeletal muscle
    DOI:  https://doi.org/10.1186/s12944-024-02079-z
  31. Dev Cell. 2024 Mar 29. pii: S1534-5807(24)00184-9. [Epub ahead of print]
      The function of many organs, including skeletal muscle, depends on their three-dimensional structure. Muscle regeneration therefore requires not only reestablishment of myofibers but also restoration of tissue architecture. Resident muscle stem cells (SCs) are essential for regeneration, but how SCs regenerate muscle architecture is largely unknown. We address this problem using genetic labeling of mouse SCs and whole-mount imaging to reconstruct, in three dimensions, muscle regeneration. Unexpectedly, we found that myofibers form via two distinct phases of fusion and the residual basement membrane of necrotic myofibers is critical for promoting fusion and orienting regenerated myofibers. Furthermore, the centralized myonuclei characteristic of regenerated myofibers are associated with myofibrillogenesis and endure months post injury. Finally, we elucidate two cellular mechanisms for the formation of branched myofibers, a pathology characteristic of diseased muscle. We provide a synthesis of the cellular events of regeneration and show that these differ from those used during development.
    Keywords:  basement membrane; fusion; macrophage; muscle; muscle stem cells; regeneration; satellite cells
    DOI:  https://doi.org/10.1016/j.devcel.2024.03.017
  32. Dev Cell. 2024 Mar 28. pii: S1534-5807(24)00188-6. [Epub ahead of print]
      Telomere dynamics are linked to aging hallmarks, and age-associated telomere loss fuels the development of epithelial cancers. In Apc-mutant mice, the onset of DNA damage associated with telomere dysfunction has been shown to accelerate adenoma initiation via unknown mechanisms. Here, we observed that Apc-mutant mice engineered to experience telomere dysfunction show accelerated adenoma formation resulting from augmented cell competition and clonal expansion. Mechanistically, telomere dysfunction induces the repression of EZH2, resulting in the derepression of Wnt antagonists, which causes the differentiation of adjacent stem cells and a relative growth advantage to Apc-deficient telomere dysfunctional cells. Correspondingly, in this mouse model, GSK3β inhibition countered the actions of Wnt antagonists on intestinal stem cells, resulting in impaired adenoma formation of telomere dysfunctional Apc-mutant cells. Thus, telomere dysfunction contributes to cancer initiation through altered stem cell dynamics, identifying an interception strategy for human APC-mutant cancers with shortened telomeres.
    Keywords:  EZH2; Wnt signaling; colorectal cancer; genomic instability; initiation; microenvironment; telomeres
    DOI:  https://doi.org/10.1016/j.devcel.2024.03.020
  33. Cell Metab. 2024 Mar 28. pii: S1550-4131(24)00085-8. [Epub ahead of print]
      Bone secretory proteins, termed osteokines, regulate bone metabolism and whole-body homeostasis. However, fundamental questions as to what the bona fide osteokines and their cellular sources are and how they are regulated remain unclear. In this study, we analyzed bone and extraskeletal tissues, osteoblast (OB) conditioned media, bone marrow supernatant (BMS), and serum, for basal osteokines and those responsive to aging and mechanical loading/unloading. We identified 375 candidate osteokines and their changes in response to aging and mechanical dynamics by integrating data from RNA-seq, scRNA-seq, and proteomic approaches. Furthermore, we analyzed their cellular sources in the bone and inter-organ communication facilitated by them (bone-brain, liver, and aorta). Notably, we discovered that senescent OBs secrete fatty-acid-binding protein 3 to propagate senescence toward vascular smooth muscle cells (VSMCs). Taken together, we identified previously unknown candidate osteokines and established a dynamic regulatory network among them, thus providing valuable resources to further investigate their systemic roles.
    Keywords:  FABP3; aging; global regulation; mechanical loading; multi-omics; osteokine
    DOI:  https://doi.org/10.1016/j.cmet.2024.03.006
  34. Ann Hepatobiliary Pancreat Surg. 2024 Apr 01.
    Barts Pancreas Tissue Bank
       Backgrounds/Aims: This study aimed to investigate patterns and factors affecting recurrence after curative resection for pancreatic ductal adenocarcinoma (PDAC).
    Methods: Consecutive patients who underwent curative resection for PDAC (2011-21) and consented to data and tissue collection (Barts Pancreas Tissue Bank) were followed up until May 2023. Clinico-pathological variables were analysed using Cox proportional hazards model.
    Results: Of 91 people (42 males [46%]; median age, 71 years [range, 43-86 years]) with a median follow-up of 51 months (95% confidence intervals [CIs], 40-61 months), the recurrence rate was 72.5% (n = 66; 12 loco-regional alone, 11 liver alone, 5 lung alone, 3 peritoneal alone, 29 simultaneous loco-regional and distant metastases, and 6 multi-focal distant metastases at first recurrence diagnosis). The median time to recurrence was 8.5 months (95% CI, 6.6-10.5 months). Median survival after recurrence was 5.8 months (95% CI, 4.2-7.3 months). Stratification by recurrence location revealed significant differences in time to recurrence between loco-regional only recurrence (median, 13.6 months; 95% CI, 11.7-15.5 months) and simultaneous loco-regional with distant recurrence (median, 7.5 months; 95% CI, 4.6-10.4 months; p = 0.02, pairwise log-rank test). Significant predictors for recurrence were systemic inflammation index (SII) ≥ 500 (hazard ratio [HR], 4.5; 95% CI, 1.4-14.3), lymph node ratio ≥ 0.33 (HR, 2.8; 95% CI, 1.4-5.8), and adjuvant chemotherapy (HR, 0.4; 95% CI, 0.2-0.7).
    Conclusions: Timing to loco-regional only recurrence was significantly longer than simultaneous loco-regional with distant recurrence. Significant predictors for recurrence were SII, lymph node ration, and adjuvant chemotherapy.
    Keywords:  Disease free survival; Lymph node ratio; Multivariate analysis; Overall survival; Systemic inflammation index
    DOI:  https://doi.org/10.14701/ahbps.23-149
  35. bioRxiv. 2024 Mar 22. pii: 2024.03.21.586194. [Epub ahead of print]
      Taurine is a conditionally essential micronutrient and one of the most abundant amino acids in humans 1-3 . In endogenous taurine metabolism, dedicated enzymes are involved in biosynthesis of taurine from cysteine as well as the downstream derivatization of taurine into secondary taurine metabolites 4,5 . One such taurine metabolite is N-acetyltaurine 6 . Levels of N-acetyltaurine are dynamically regulated by diverse physiologic perturbations that alter taurine and/or acetate flux, including endurance exercise 7 , nutritional taurine supplementation 8 , and alcohol consumption 6,9 . While taurine N-acetyltransferase activity has been previously detected in mammalian cells 6,7 , the molecular identity of this enzyme, and the physiologic relevance of N-acetyltaurine, have remained unknown. Here we show that the orphan body mass index-associated enzyme PTER (phosphotriesterase-related) 10 is the principal mammalian taurine N-acetyltransferase/hydrolase. In vitro, recombinant PTER catalyzes bidirectional taurine N-acetylation with free acetate as well as the reverse N-acetyltaurine hydrolysis reaction. Genetic ablation of PTER in mice results in complete loss of tissue taurine N-acetyltransferase/hydrolysis activities and systemic elevation of N-acetyltaurine levels. Upon stimuli that increase taurine levels, PTER-KO mice exhibit lower body weight, reduced adiposity, and improved glucose homeostasis. These phenotypes are recapitulated by administration of N-acetyltaurine to wild-type mice. Lastly, the anorexigenic and anti-obesity effects of N-acetyltaurine require functional GFRAL receptors. Together, these data uncover enzymatic control of a previously enigmatic pathway of secondary taurine metabolism linked to energy balance.
    DOI:  https://doi.org/10.1101/2024.03.21.586194
  36. Cell Rep. 2024 Apr 02. pii: S2211-1247(24)00372-3. [Epub ahead of print]43(4): 114044
      We identify a senescence restriction point (SeRP) as a critical event for cells to commit to senescence. The SeRP integrates the intensity and duration of oncogenic stress, keeps a memory of previous stresses, and combines oncogenic signals acting on different pathways by modulating chromatin accessibility. Chromatin regions opened upon commitment to senescence are enriched in nucleolar-associated domains, which are gene-poor regions enriched in repeated sequences. Once committed to senescence, cells no longer depend on the initial stress signal and exhibit a characteristic transcriptome regulated by a transcription factor network that includes ETV4, RUNX1, OCT1, and MAFB. Consistent with a tumor suppressor role for this network, the levels of ETV4 and RUNX1 are very high in benign lesions of the pancreas but decrease dramatically in pancreatic ductal adenocarcinomas. The discovery of senescence commitment and its chromatin-linked regulation suggests potential strategies for reinstating tumor suppression in human cancers.
    Keywords:  CP: Cancer; ERK; ETV4; RUNX1; chromatin; commitment; oncogenic memory; pancreatic cancer; restriction point; senescence
    DOI:  https://doi.org/10.1016/j.celrep.2024.114044
  37. bioRxiv. 2024 Mar 14. pii: 2024.03.13.583573. [Epub ahead of print]
      Circulating Tumor Cells (CTCs), interrogated by sampling blood from patients with cancer, contain multiple analytes, including intact RNA, high molecular weight DNA, proteins, and metabolic markers. However, the clinical utility of tumor cell-based liquid biopsy has been limited since CTCs are very rare, and current technologies cannot process the blood volumes required to isolate a sufficient number of tumor cells for in-depth assays. We previously described a high-throughput microfluidic prototype utilizing high-flow channels and amplification of cell sorting forces through magnetic lenses. Here, we apply this technology to analyze patient-derived leukapheresis products, interrogating a mean blood volume of 5.83 liters from patients with metastatic cancer, with a median of 2,799 CTCs purified per patient. Isolation of many CTCs from individual patients enables characterization of their morphological and molecular heterogeneity, including cell and nuclear size and RNA expression. It also allows robust detection of gene copy number variation, a definitive cancer marker with potential diagnostic applications. High-volume microfluidic enrichment of CTCs constitutes a new dimension in liquid biopsies.
    DOI:  https://doi.org/10.1101/2024.03.13.583573
  38. Elife. 2024 Apr 05. pii: RP89303. [Epub ahead of print]12
      Poly(ADP-ribose)ylation or PARylation by PAR polymerase 1 (PARP1) and dePARylation by poly(ADP-ribose) glycohydrolase (PARG) are equally important for the dynamic regulation of DNA damage response. PARG, the most active dePARylation enzyme, is recruited to sites of DNA damage via pADPr-dependent and PCNA-dependent mechanisms. Targeting dePARylation is considered an alternative strategy to overcome PARP inhibitor resistance. However, precisely how dePARylation functions in normal unperturbed cells remains elusive. To address this challenge, we conducted multiple CRISPR screens and revealed that dePARylation of S phase pADPr by PARG is essential for cell viability. Loss of dePARylation activity initially induced S-phase-specific pADPr signaling, which resulted from unligated Okazaki fragments and eventually led to uncontrolled pADPr accumulation and PARP1/2-dependent cytotoxicity. Moreover, we demonstrated that proteins involved in Okazaki fragment ligation and/or base excision repair regulate pADPr signaling and cell death induced by PARG inhibition. In addition, we determined that PARG expression is critical for cellular sensitivity to PARG inhibition. Additionally, we revealed that PARG is essential for cell survival by suppressing pADPr. Collectively, our data not only identify an essential role for PARG in normal proliferating cells but also provide a potential biomarker for the further development of PARG inhibitors in cancer therapy.
    Keywords:  PARG; PARG inhibitor; PARP1; PARylation; biochemistry; cancer biology; chemical biology; nad; parg; parg inhibitor; parp inhibitor; parylation; ribosylation
    DOI:  https://doi.org/10.7554/eLife.89303
  39. Cell Rep. 2024 Apr 04. pii: S2211-1247(24)00394-2. [Epub ahead of print]43(4): 114066
      In human cells and yeast, an intact "hydrophobic patch" substrate docking site is needed for mitotic cyclin centrosomal localization. A hydrophobic patch mutant (HPM) of the fission yeast mitotic cyclin Cdc13 cannot enter mitosis, but whether this is due to defective centrosomal localization or defective cyclin-substrate docking more widely is unknown. Here, we show that artificially restoring Cdc13-HPM centrosomal localization promotes mitotic entry and increases CDK (cyclin-dependent kinase) substrate phosphorylation at the centrosome and in the cytoplasm. We also show that the S-phase B-cyclin hydrophobic patch is required for centrosomal localization but not for S phase. We propose that the hydrophobic patch is essential for mitosis due to its requirement for the local concentration of cyclin-CDK with CDK substrates and regulators at the centrosome. Our findings emphasize the central importance of the centrosome as a hub coordinating cell-cycle control and explain why the cyclin hydrophobic patch is essential for mitosis.
    Keywords:  CDK; CP: Cell biology; cell cycle; centrosome; cyclin B; hydrophobic patch; mitosis
    DOI:  https://doi.org/10.1016/j.celrep.2024.114066
  40. Biochim Biophys Acta Mol Cell Biol Lipids. 2024 Mar 31. pii: S1388-1981(24)00041-6. [Epub ahead of print] 159491
      Inborn errors of metabolism (IEM) represent a heterogeneous group of more than 1800 rare disorders, many of which are causing significant childhood morbidity and mortality. More than 100 IEM are linked to dyslipidaemia, but yet our knowledge in connecting genetic information with lipidomic data is limited. Stable isotope tracing studies of the lipid metabolism (STL) provide insights on the dynamic of cellular lipid processes and could thereby facilitate the delineation of underlying metabolic (patho)mechanisms. This mini-review focuses on principles as well as technical limitations of STL and describes potential clinical applications by discussing recently published STL focusing on IEM.
    Keywords:  Flux studies; Lipids; Mass spectrometry; Metabolism; Rare diseases
    DOI:  https://doi.org/10.1016/j.bbalip.2024.159491
  41. Cancer Discov. 2024 Apr 03.
      Activating point mutations in the MET tyrosine kinase domain (TKD) are oncogenic in a subset of papillary renal cell carcinomas (PRCC). Here, using comprehensive genomic profiling among >600,000 patients, we identify activating MET TKD point mutations as putative oncogenic driver across diverse cancers, with a frequency of ~0.5%. The most common mutations in the MET TKD defined as oncogenic or likely oncogenic according to OncoKB resulted in amino acid substitutions at positions H1094, L1195, F1200, D1228, Y1230, M1250, and others. Preclinical modeling of these alterations confirmed their oncogenic potential, and also demonstrated differential patterns of sensitivity to type I and type II MET inhibitors. Two patients with metastatic lung adenocarcinoma harboring MET TKD mutations (H1094Y, F1200I) and no other known oncogenic drivers achieved confirmed partial responses to a type I MET inhibitor. Activating MET TKD mutations occur in multiple malignancies and may confer clinical sensitivity to currently available MET inhibitors.
    DOI:  https://doi.org/10.1158/2159-8290.CD-23-1217
  42. Nat Cell Biol. 2024 Apr 01.
      Metabolic crosstalk of the major nutrients glucose, amino acids and fatty acids (FAs) ensures systemic metabolic homeostasis. The coordination between the supply of glucose and FAs to meet various physiological demands is especially important as improper nutrient levels lead to metabolic disorders, such as diabetes and metabolic dysfunction-associated steatohepatitis (MASH). In response to the oscillations in blood glucose levels, lipolysis is thought to be mainly regulated hormonally to control FA liberation from lipid droplets by insulin, catecholamine and glucagon. However, whether general cell-intrinsic mechanisms exist to directly modulate lipolysis via glucose sensing remains largely unknown. Here we report the identification of such an intrinsic mechanism, which involves Golgi PtdIns4P-mediated regulation of adipose triglyceride lipase (ATGL)-driven lipolysis via intracellular glucose sensing. Mechanistically, depletion of intracellular glucose results in lower Golgi PtdIns4P levels, and thus reduced assembly of the E3 ligase complex CUL7FBXW8 in the Golgi apparatus. Decreased levels of the E3 ligase complex lead to reduced polyubiquitylation of ATGL in the Golgi and enhancement of ATGL-driven lipolysis. This cell-intrinsic mechanism regulates both the pool of intracellular FAs and their extracellular release to meet physiological demands during fasting and glucose deprivation. Moreover, genetic and pharmacological manipulation of the Golgi PtdIns4P-CUL7FBXW8-ATGL axis in mouse models of simple hepatic steatosis and MASH, as well as during ex vivo perfusion of a human steatotic liver graft leads to the amelioration of steatosis, suggesting that this pathway might be a promising target for metabolic dysfunction-associated steatotic liver disease and possibly MASH.
    DOI:  https://doi.org/10.1038/s41556-024-01386-y
  43. J Lipid Res. 2024 Apr 01. pii: S0022-2275(24)00045-2. [Epub ahead of print] 100540
      Intestinal epithelial cells convert excess fatty acids into triglyceride (TAG) for storage in cytoplasmic lipid droplets and secretion in chylomicrons. Nuclear lipid droplets (nLDs) are present in intestinal cells but their origin and relationship to cytoplasmic TAG synthesis and secretion is unknown. nLDs and related lipid-associated promyelocytic leukemia (PML) structures (LAPS) were abundant in oleate-treated Caco2 but less frequent in other human colorectal cancer cell lines and mouse intestinal organoids. nLDs and LAPS in undifferentiated oleate-treated Caco2 cells harboured the phosphatidate phosphatase Lipin1, its product diacylglycerol (DAG), and CTP:phosphocholine cytidylyltransferase (CCT)α. CCTα knockout Caco2 cells had fewer but larger nLDs indicating a reliance on de novo PC synthesis for assembly. Differentiation of Caco2 cells caused large nLDs and LAPS to form regardless of oleate treatment or CCTα expression. nLDs and LAPS in Caco2 cells did not associate with apoCIII and apoAI, and formed dependently of microsomal triglyceride transfer protein (MTP) expression and activity, indicating they are not derived from endoplasmic reticulum luminal LDs precursors. Instead, undifferentiated Caco2 cells harboured a constitutive pool of nLDs and LAPS in proximity to the nuclear envelope (NE) that expanded in size and number with oleate treatment. Inhibition of TAG synthesis did affect the number of nascent nLDs and LAPS but prevented their association with PML, Lipin1α and DAG, which instead accumulated on the nuclear membranes. Thus, nLD and LAPS biogenesis in Caco2 cells is not linked to lipoprotein secretion but involves biogenesis and/or expansion of nascent nLDs by de novo lipid synthesis.
    Keywords:  CCTα; MTP; Nuclear lipid droplets; phosphatidylcholine; promyelocytic leukemia protein
    DOI:  https://doi.org/10.1016/j.jlr.2024.100540