bims-cagime Biomed News
on Cancer, aging and metabolism
Issue of 2024–11–17
57 papers selected by
Kıvanç Görgülü, Technical University of Munich
  1. Genetics and biology of pancreatic ductal adenocarcinoma.
  2. Role of myofiber-specific FoxP1 in pancreatic cancer-induced muscle wasting.
  3. Cancer-associated fibroblasts maintain critical pancreatic cancer cell lipid homeostasis in the tumor microenvironment.
  4. Lfng-expressing centroacinar cell is a unique cell-of-origin for p53 deficient pancreatic cancer.
  5. Measuring the impact of therapy-induced senescence on NK cell phenotypes in cancer.
  6. Clinical functional proteomics of intercellular signalling in pancreatic cancer.
  7. Metabolic landscape of disseminated cancer dormancy.
  8. Senescence as a therapeutic target in cancer and age-related diseases.
  9. Mouse models for pancreatic ductal adenocarcinoma are affected by the cre-driver used to promote KRASG12D activation.
  10. Functionally overlapping intra- and extralysosomal pathways promote bis(monoacylglycero)phosphate synthesis in mammalian cells.
  11. Autophagy-dependent splicing control directs translation toward inflammation during senescence.
  12. A β-hydroxybutyrate shunt pathway generates anti-obesity ketone metabolites.
  13. PRDX6 dictates ferroptosis sensitivity by directing cellular selenium utilization.
  14. Global metabolomic profiling of tumor tissue and paired serum samples to identify biomarkers for response to neoadjuvant FOLFIRINOX treatment of human pancreatic cancer.
  15. Iron-Induced Lipid Oxidation Alters Membrane Mechanics Favoring Permeabilization.
  16. Calcium signaling from damaged lysosomes induces cytoprotective stress granules.
  17. A metabolic switch to the pentose-phosphate pathway induces radiation resistance in pancreatic cancer.
  18. CAFomics: convergence to translation for precision stroma approaches.
  19. Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
  20. Clearance of p21 highly expressing senescent cells accelerates cutaneous wound healing.
  21. Cell response to extracellular matrix viscous energy dissipation outweighs high-rigidity sensing.
  22. Pathogenic proteotoxicity of cryptic splicing is alleviated by ubiquitination and ER-phagy.
  23. Mechanisms of cancer cachexia and targeted therapeutic strategies.
  24. Dietary omega-3 (ω-3) fatty acids mitigate intestinal barrier integrity alterations in mice fed a high-fat diet: Implications for pancreatic carcinogenesis.
  25. Mapping membrane biophysical nano-environments.
  26. The pro-oncogenic noncanonical activity of a RAS•GTP:RanGAP1 complex facilitates nuclear protein export.
  27. Exploring the interplay between yeast cell membrane lipid adaptation and physiological response to acetic acid stress.
  28. GlutaR: A High-Performance Fluorescent Protein-Based Sensor for Spatiotemporal Monitoring of Glutamine Dynamics In Vivo.
  29. From precursor to cancer: decoding the intrinsic and extrinsic pathways of pancreatic intraepithelial neoplasia progression.
  30. A tuneable minimal cell membrane reveals that two lipid species suffice for life.
  31. The Warburg Effect is the result of faster ATP production by glycolysis than respiration.
  32. Daily GDF15 treatment has sex-specific effects on body weight and food intake and does not enhance the effects of voluntary physical activity in mice.
  33. Pancreatic ductal adenocarcinoma, β-blockers and antihistamines: A clinical trial is needed.
  34. Leader cells promote immunosuppression to drive ovarian cancer progression in vivo.
  35. Could senescent cells be the prescription for therapeutic cancer vaccines?
  36. Organoid-based precision medicine in pancreatic cancer.
  37. Tissue-colonizing disseminated tumor cells secrete prostaglandin E2 to promote NK cell dysfunction and evade anti-metastatic immunity.
  38. Opposing roles for AMPK in regulating distinct mitophagy pathways.
  39. Tumor heterogeneity and cooperating cancer hallmarks driven by divergent EMT programs.
  40. Early detection of pancreatic cancer: Study design and analytical considerations in biomarker discovery and early phase validation studies.
  41. Lactate reprograms glioblastoma immunity through CBX3-regulated histone lactylation.
  42. DNA methylation classifier to diagnose pancreatic ductal adenocarcinoma metastases from different anatomical sites.
  43. A Phase 2 study of nivolumab in combination with modified FOLFIRINOX for metastatic pancreatic cancer.
  44. Nonequilibrium Molecular Dynamics Method to Generate Poiseuille-Like Flow between Lipid Bilayers.
  45. Coherent Raman microscopy visualizes ongoing cellular senescence through amide I peak shifts originating from β sheets in disordered nucleolar proteins.
  46. Phase I Study of Adjuvant Allogeneic GM-CSF-Transduced Pancreatic Tumor Cell Vaccine, Low Dose Cyclophosphamide, and SBRT followed by FFX in High-Risk Resected Pancreatic Ductal Adenocarcinoma.
  47. Chronic social stress induces p16-mediated senescent cell accumulation in mice.
  48. Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer.
  49. EasyFlow: An open-source, user-friendly cytometry analyzer with graphic user interface (GUI).
  50. VEGF-C propagates 'onward' colorectal cancer metastasis from liver to lung.
  51. Spatiotemporal modeling of molecular holograms.
  52. Prospective observational study and mechanistic evidence showing lipolysis of circulating triglycerides worsens hypertriglyceridemic acute pancreatitis.
  53. EvoAI enables extreme compression and reconstruction of the protein sequence space.
  54. Understanding Mitochondrial and Lysosomal Dynamics by Fluorescent Microscopy.
  55. The cellular dogma.
  56. Biophysical and Morphological Cell Features Retrieved by Digital Holographic Microscopy Correlate with Drug-Induced Changes in Cell Migration Behavior.
  57. Charge-Stabilized Nanodiscs as a New Class of Lipid Nanoparticles.
  1. Genes Dev. 2024 Nov 07.
    Genetics and biology of pancreatic ductal adenocarcinoma.
    Haoqiang Ying, Alec C Kimmelman, Nabeel Bardeesy, Raghu Kalluri, Anirban Maitra, Ronald A DePinho.
      Pancreatic ductal adenocarcinoma (PDAC) poses a grim prognosis for patients. Recent multidisciplinary research efforts have provided critical insights into its genetics and tumor biology, creating the foundation for rational development of targeted and immune therapies. Here, we review the PDAC genomic landscape and the role of specific oncogenic events in tumor initiation and progression, as well as their contributions to shaping its tumor biology. We further summarize and synthesize breakthroughs in single-cell and metabolic profiling technologies that have illuminated the complex cellular composition and heterotypic interactions of the PDAC tumor microenvironment, with an emphasis on metabolic cross-talk across cancer and stromal cells that sustains anabolic growth and suppresses tumor immunity. These conceptual advances have generated novel immunotherapy regimens, particularly cancer vaccines, which are now in clinical testing. We also highlight the advent of KRAS targeted therapy, a milestone advance that has transformed treatment paradigms and offers a platform for combined immunotherapy and targeted strategies. This review provides a perspective summarizing current scientific and therapeutic challenges as well as practice-changing opportunities for the PDAC field at this major inflection point.
    Keywords:  cancer-associated fibroblast; immune regulation; kras; pancreatic cancer; targeted therapy; tumor heterogeneity; tumor microenvironment
    DOI:  https://doi.org/10.1101/gad.351863.124
  2. Am J Physiol Cell Physiol. 2024 Nov 15.
    Role of myofiber-specific FoxP1 in pancreatic cancer-induced muscle wasting.
    Martin M Schonk, Jeremy B Ducharme, Daria Neyroud, Rachel L Nosacka, Haley O Tucker, Sarah M Judge, Andrew R Judge.
      Cancer cachexia affects up to 80% of cancer patients and results in reduced quality of life and survival. We previously demonstrated that the transcriptional repressor Forkhead box P1 (FoxP1) is upregulated in skeletal muscle of cachectic mice and people with cancer, and when overexpressed in skeletal muscle is sufficient to induce pathological features characteristic of cachexia. However, the role of myofiber-derived FoxP1 in both normal muscle physiology and cancer-induced muscle wasting remains largely unexplored. To address this gap, we generated a conditional mouse line with myofiber-specific ablation of FoxP1 (FoxP1SkmKO) and found that in cancer-free mice, deletion of FoxP1 in skeletal myofibers resulted in increased myofiber size in both males and females, with a significant increase in muscle mass in males. In response to murine KPC pancreatic tumor burden, we found that myofiber-derived FoxP1 is required for cancer-induced muscle wasting and diaphragm muscle weakness in male mice. In summary, our findings identify myofiber-specific FoxP1 as a negative regulator of skeletal muscle with sex-specific differences in the context of cancer.
    Keywords:  Biological sex; Cancer cachexia; Forkhead box P1; Muscle wasting; Pancreatic cancer
    DOI:  https://doi.org/10.1152/ajpcell.00701.2024
  3. Cell Rep. 2024 Nov 12. pii: S2211-1247(24)01323-8. [Epub ahead of print]43(11): 114972
    Cancer-associated fibroblasts maintain critical pancreatic cancer cell lipid homeostasis in the tumor microenvironment.
    Xu Han, Michelle Burrows, Laura C Kim, Jimmy P Xu, Will Vostrejs, Tran Ngoc Van Le, Carson Poltorack, Yanqing Jiang, Edna Cukierman, Ben Z Stanger, Kim A Reiss, Sydney M Shaffer, Clementina Mesaros, Brian Keith, M Celeste Simon.
      Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with abundant cancer-associated fibroblasts (CAFs) creating hallmark desmoplasia that limits oxygen and nutrient delivery. This study explores the importance of lipid homeostasis under stress. Exogenous unsaturated lipids, rather than de novo synthesis, sustain PDAC cell viability by relieving endoplasmic reticulum (ER) stress under nutrient scarcity. Furthermore, CAFs are less hypoxic than adjacent malignant cells in vivo, nominating them as a potential source of unsaturated lipids. CAF-conditioned medium promotes PDAC cell survival upon nutrient and oxygen deprivation, an effect reversed by delipidation. Lysophosphatidylcholines (LPCs) are particularly enriched in CAF-conditioned medium and preferentially taken up by PDAC cells, where they are converted to phosphatidylcholine (PC) to sustain membrane integrity. Blocking LPC-to-PC conversion inhibits PDAC cell survival and increases ER stress. These findings show a critical lipid "cross-feeding" mechanism that promotes PDAC cell survival, offering a potential metabolic target for treatment.
    Keywords:  CP: Cancer; CP: Metabolism; fibroblasts; hypoxia; lipids; pancreatic cancer; tumor microenvironment; unsaturated fatty acids
    DOI:  https://doi.org/10.1016/j.celrep.2024.114972
  4. Oncogene. 2024 Nov 15.
    Lfng-expressing centroacinar cell is a unique cell-of-origin for p53 deficient pancreatic cancer.
    Wen-Cheng Chung, Shubing Zhang, Azeddine Atfi, Keli Xu.
      Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies with limited understanding of etiology. Studies in mice showed that both acinar and ductal cells of the pancreas can be targeted by combination of oncogenic Kras and p53 mutations to form PDAC. How the transforming capacities of pancreatic cells are constrained, and whether a subset of cells could serve as a prime target for oncogenic transformation, remain obscure. Here we report that expression of a Notch modulator, Lunatic Fringe (Lfng), is restricted to a limited number of cells with centroacinar location and morphology in the adult pancreas. Lfng-expressing cells are preferentially targeted by oncogenic Kras along with p53 deletion to form PDAC, and deletion of Lfng blocks tumor initiation from these cells. Notch3 is a functional Notch receptor for PDAC initiation and progression in this context. Lfng is upregulated in acinar- and ductal-derived PDAC and its deletion suppresses these tumors. Finally, high LFNG expression is associated with high grade and poor survival in human patients. Taken together, Lfng marks a centroacinar subpopulation that is uniquely susceptible to oncogenic transformation when p53 is lost, and Lfng functions as an oncogene in all three lineages of the exocrine pancreas.
    DOI:  https://doi.org/10.1038/s41388-024-03226-7
  5. Methods Cell Biol. 2024 ;pii: S0091-679X(24)00182-1. [Epub ahead of print]190 171-201
    Measuring the impact of therapy-induced senescence on NK cell phenotypes in cancer.
    Shreya R Chowdhury, Katherine C Murphy, Chaitanya N Parikh, Kelly D DeMarco, Lin Zhou, Marcus Ruscetti.
      Cellular senescence is a damage-induced condition characterized by enduring cell cycle arrest and a heightened secretory profile known as the senescence-associated secretory phenotype (SASP). The SASP consists not only of release of inflammatory cytokines and chemokines that attract and activate a diverse repertoire of innate and adaptive immune cells, but also the upregulation of immunomodulatory cell surface molecules that promote immune clearance of senescent cells. Natural Killer (NK) cells are particularly adept at sensing and eliminating senescent cells. In the setting of cancer, commonly administered cytotoxic and cytostatic therapies can elicit senescence and in turn reactivate NK cell immune surveillance against tumors. Here, we detail a series of in vivo, ex vivo, and in vitro assays to assess the impact of therapy-induced senescence on NK cell phenotypes, including their activation, exhaustion, migration, and killing capacity in the context of pancreatic cancer. Importantly, this methodology can be adapted to investigate NK cell biology across various disease states and treatment modalities and help inform NK cell-based immunotherapies for cancer.
    Keywords:  Co-culture assays; Flow cytometry; Migration assays; Mouse models; Natural Killer cells; Pancreatic cancer; RAS targeted therapies; SASP; Senescence; Tumor immunology
    DOI:  https://doi.org/10.1016/bs.mcb.2024.07.010
  6. Nature. 2024 Nov 13.
    Clinical functional proteomics of intercellular signalling in pancreatic cancer.
    Peiwu Huang, Weina Gao, Changying Fu, Min Wang, Yunguang Li, Bizhu Chu, An He, Yuan Li, Xiaomei Deng, Yehan Zhang, Qian Kong, Jingxiong Yuan, Hebin Wang, Yu Shi, Dong Gao, Renyi Qin, Tony Hunter, Ruijun Tian.
      Pancreatic ductal adenocarcinoma (PDAC) has an atypical, highly stromal tumour microenvironment (TME) that profoundly contributes to its poor prognosis1. Here, to better understand the intercellular signalling between cancer and stromal cells directly in PDAC tumours, we developed a multidimensional proteomic strategy called TMEPro. We applied TMEPro to profile the glycosylated secreted and plasma membrane proteome of 100 human pancreatic tissue samples to a great depth, define cell type origins and identify potential paracrine cross-talk, especially that mediated through tyrosine phosphorylation. Temporal dynamics during pancreatic tumour progression were investigated in a genetically engineered PDAC mouse model. Functionally, we revealed reciprocal signalling between stromal cells and cancer cells mediated by the stromal PDGFR-PTPN11-FOS signalling axis. Furthermore, we examined the generic shedding mechanism of plasma membrane proteins in PDAC tumours and revealed that matrix-metalloprotease-mediated shedding of the AXL receptor tyrosine kinase ectodomain provides an additional dimension of intercellular signalling regulation in the PDAC TME. Importantly, the level of shed AXL has a potential correlation with lymph node metastasis, and inhibition of AXL shedding and its kinase activity showed a substantial synergistic effect in inhibiting cancer cell growth. In summary, we provide TMEPro, a generically applicable clinical functional proteomic strategy, and a comprehensive resource for better understanding the PDAC TME and facilitating the discovery of new diagnostic and therapeutic targets.
    DOI:  https://doi.org/10.1038/s41586-024-08225-y
  7. Trends Cancer. 2024 Nov 06. pii: S2405-8033(24)00224-3. [Epub ahead of print]
    Metabolic landscape of disseminated cancer dormancy.
    Stanislav Drapela, Bruna M Garcia, Ana P Gomes, Ana Luísa Correia.
      Cancer dormancy is a phenomenon defined by the entry of cancer cells into a reversible quiescent, nonproliferative state, and represents an essential part of the metastatic cascade responsible for cancer recurrence and mortality. Emerging evidence suggests that metabolic reprogramming plays a pivotal role in enabling entry, maintenance, and exit from dormancy in the face of the different environments of the metastatic cascade. Here, we review the current literature to understand the dynamics of metabolism during dormancy, highlighting its fine-tuning by the host micro- and macroenvironment, and put forward the importance of identifying metabolic vulnerabilities of the dormant state as therapeutic targets to eradicate recurrent disease.
    Keywords:  dormancy; metabolism; metastasis; metastatic microenvironment
    DOI:  https://doi.org/10.1016/j.trecan.2024.10.005
  8. Nat Rev Drug Discov. 2024 Nov 15.
    Senescence as a therapeutic target in cancer and age-related diseases.
    Domhnall McHugh, Imanol Durán, Jesús Gil.
      Cellular senescence is a stress response that restrains the growth of aged, damaged or abnormal cells. Thus, senescence has a crucial role in development, tissue maintenance and cancer prevention. However, lingering senescent cells fuel chronic inflammation through the acquisition of a senescence-associated secretory phenotype (SASP), which contributes to cancer and age-related tissue dysfunction. Recent progress in understanding senescence has spurred interest in the development of approaches to target senescent cells, known as senotherapies. In this Review, we evaluate the status of various types of senotherapies, including senolytics that eliminate senescent cells, senomorphics that suppress the SASP, interventions that mitigate senescence and strategies that harness the immune system to clear senescent cells. We also summarize how these approaches can be combined with cancer therapies, and we discuss the challenges and opportunities in moving senotherapies into clinical practice. Such therapies have the potential to address root causes of age-related diseases and thus open new avenues for preventive therapies and treating multimorbidities.
    DOI:  https://doi.org/10.1038/s41573-024-01074-4
  9. Cell Mol Gastroenterol Hepatol. 2024 Nov 13. pii: S2352-345X(24)00183-8. [Epub ahead of print] 101428
    Mouse models for pancreatic ductal adenocarcinoma are affected by the cre-driver used to promote KRASG12D activation.
    Fatemeh Mousavi, Joyce Thompson, Justine Lau, Nur Renollet, Mickenzie B Martin, Jake McGue, Oneeb Hassan, Timothy Frankel, Parisa Shooshtari, Christopher L Pin, Filip Bednar.
       BACKGROUND AND AIMS: The fundamental biology of pancreatic ductal adenocarcinoma has been greatly impacted by the characterization of genetically engineered mouse models that allow temporal and spatial activation of oncogenic KRAS (KRASG12D). One of the most commonly used models involves targeted insertion of a cre-recombinase into the Ptf1a gene. However, this approach disrupts the Ptf1a gene, resulting in haploinsufficiency that likely affects sensitivity to oncogenic KRAS (KRASG12D). This study aims to determine if Ptf1a haploinsufficiency affected the acinar cell response to KRASG12D before and after induction of pancreatic injury.
    METHODS: We performed morphological and molecular analysis of three genetically engineered mouse models that express a tamoxifen-inducible cre-recombinase to activate KrasG12D in acinar cells of the pancreas. The cre-recombinase was targeted to the acinar-specific transcription factor genes, Ptf1a or Mist1/Bhlha15, or expressed within a BAC-derived Elastase transgene. Histological and RNA-seq analyses were used to delineate differences between the models.
    RESULTS: Up to two months after tamoxifen induction of KRASG12D, morphological changes were negligible. However, induction of pancreatic injury by cerulein resulted in widespread PanIN lesions in Ptf1acreERT pancreata within seven days and maintained for at least five weeks post-injury, which was not seen in the models with two functional Ptf1a alleles. RNA-seq analysis prior to injury induction suggested Ptf1acreERT and Mist1creERT mice have unique profiles of gene expression that predict a differential response to injury. Multiplex analysis of pancreatic tissue confirmed different inflammatory responses between the models.
    CONCLUSIONS: These findings suggest Ptf1a haploinsufficiency in Ptf1acreERT mouse models promotes KRASG12D priming of genes for promotion of PDAC.
    Keywords:  Cre Recombinase; Elastase; Genetically Engineered Mouse Models; Mist1; Ptf1a; RNA-sequencing
    DOI:  https://doi.org/10.1016/j.jcmgh.2024.101428
  10. Nat Commun. 2024 Nov 16. 15(1): 9937
    Functionally overlapping intra- and extralysosomal pathways promote bis(monoacylglycero)phosphate synthesis in mammalian cells.
    Dominik Bulfon, Johannes Breithofer, Gernot F Grabner, Nermeen Fawzy, Anita Pirchheim, Heimo Wolinski, Dagmar Kolb, Lennart Hartig, Martin Tischitz, Clara Zitta, Greta Bramerdorfer, Achim Lass, Ulrike Taschler, Dagmar Kratky, Peter Greimel, Robert Zimmermann.
      Bis(monoacylglycero)phosphate (BMP) is a major phospholipid constituent of intralumenal membranes in late endosomes/lysosomes, where it regulates the degradation and sorting of lipid cargo. Recent observations suggest that the Batten disease-associated protein CLN5 functions as lysosomal BMP synthase. Here, we show that transacylation reactions catalyzed by cytosolic and secreted enzymes enhance BMP synthesis independently of CLN5. The transacylases identified in this study are capable of acylating the precursor lipid phosphatidylglycerol (PG), generating acyl-PG, which is subsequently hydrolyzed to BMP. Extracellularly, acyl-PG and BMP are generated by endothelial lipase in cooperation with other serum enzymes of the pancreatic lipase family. The intracellular acylation of PG is catalyzed by several members of the cytosolic phospholipase A2 group IV (PLA2G4) family. Overexpression of secreted or cytosolic transacylases was sufficient to correct BMP deficiency in HEK293 cells lacking CLN5. Collectively, our observations suggest that functionally overlapping pathways promote BMP synthesis in mammalian cells.
    DOI:  https://doi.org/10.1038/s41467-024-54213-1
  11. Dev Cell. 2024 Nov 01. pii: S1534-5807(24)00627-0. [Epub ahead of print]
    Autophagy-dependent splicing control directs translation toward inflammation during senescence.
    Jaejin Kim, Yeonghyeon Lee, Taerang Jeon, Seonmin Ju, Jong-Seo Kim, Mi-Sung Kim, Chanhee Kang.
      The cellular proteome determines the functional state of cells and is often skewed to direct pathological conditions. Autophagy shapes cellular proteomes primarily through lysosomal degradation of either damaged or unnecessary proteins. Here, we show that autophagy directs the senescence-specific translatome to fuel inflammation by coupling selective protein degradation with alternative splicing. RNA splicing is significantly altered during senescence, some of which surprisingly depend on autophagy, including exon 5 skipping of the translation regulator EIF4H. Systematic translatome profiling indicates that this event is key to the translational bias toward inflammation in senescence. Autophagy promotes these changes by selectively degrading the splicing regulator splicing factor proline and glutamine rich (SFPQ) via the autophagy receptor NBR1. These autophagy-centric inflammatory controls appear to be conserved during human tissue aging and cancer. Our work highlights the role of autophagy in the on-demand functional remodeling of cellular proteomes as well as the crosstalk between autophagy, alternative splicing, and inflammatory translation.
    Keywords:  RNA homeostasis; aging; alternative splicing; autophagy; cancer; cellular senescence; inflammation; protein translation; selective autophagy
    DOI:  https://doi.org/10.1016/j.devcel.2024.10.008
  12. Cell. 2024 Nov 07. pii: S0092-8674(24)01214-5. [Epub ahead of print]
    A β-hydroxybutyrate shunt pathway generates anti-obesity ketone metabolites.
    Maria Dolores Moya-Garzon, Mengjie Wang, Veronica L Li, Xuchao Lyu, Wei Wei, Alan Sheng-Hwa Tung, Steffen H Raun, Meng Zhao, Laetitia Coassolo, Hashim Islam, Barbara Oliveira, Yuqin Dai, Jan Spaas, Antonio Delgado-Gonzalez, Kenyi Donoso, Aurora Alvarez-Buylla, Francisco Franco-Montalban, Anudari Letian, Catherine P Ward, Lichao Liu, Katrin J Svensson, Emily L Goldberg, Christopher D Gardner, Jonathan P Little, Steven M Banik, Yong Xu, Jonathan Z Long.
      β-Hydroxybutyrate (BHB) is an abundant ketone body. To date, all known pathways of BHB metabolism involve the interconversion of BHB and primary energy intermediates. Here, we identify a previously undescribed BHB secondary metabolic pathway via CNDP2-dependent enzymatic conjugation of BHB and free amino acids. This BHB shunt pathway generates a family of anti-obesity ketone metabolites, the BHB-amino acids. Genetic ablation of CNDP2 in mice eliminates tissue amino acid BHB-ylation activity and reduces BHB-amino acid levels. The most abundant BHB-amino acid, BHB-Phe, is a ketosis-inducible congener of Lac-Phe that activates hypothalamic and brainstem neurons and suppresses feeding. Conversely, CNDP2-KO mice exhibit increased food intake and body weight following exogenous ketone ester supplementation or a ketogenic diet. CNDP2-dependent amino acid BHB-ylation and BHB-amino acid metabolites are also conserved in humans. Therefore, enzymatic amino acid BHB-ylation defines a ketone shunt pathway and bioactive ketone metabolites linked to energy balance.
    Keywords:  BHB; enzyme; ketone; metabolite; metabolomics; obesity
    DOI:  https://doi.org/10.1016/j.cell.2024.10.032
  13. Mol Cell. 2024 Nov 05. pii: S1097-2765(24)00868-2. [Epub ahead of print]
    PRDX6 dictates ferroptosis sensitivity by directing cellular selenium utilization.
    Junya Ito, Toshitaka Nakamura, Takashi Toyama, Deng Chen, Carsten Berndt, Gereon Poschmann, André Santos Dias Mourão, Sebastian Doll, Mirai Suzuki, Weijia Zhang, Jiashuo Zheng, Dietrich Trümbach, Naoya Yamada, Koya Ono, Masana Yazaki, Yasutaka Kawai, Mieko Arisawa, Yusuke Ohsaki, Hitoshi Shirakawa, Adam Wahida, Bettina Proneth, Yoshiro Saito, Kiyotaka Nakagawa, Eikan Mishima, Marcus Conrad.
      Selenium-dependent glutathione peroxidase 4 (GPX4) is the guardian of ferroptosis, preventing unrestrained (phospho)lipid peroxidation by reducing phospholipid hydroperoxides (PLOOH). However, the contribution of other phospholipid peroxidases in ferroptosis protection remains unclear. We show that cells lacking GPX4 still exhibit substantial PLOOH-reducing capacity, suggesting a contribution of alternative PLOOH peroxidases. By scrutinizing potential candidates, we found that although overexpression of peroxiredoxin 6 (PRDX6), a thiol-specific antioxidant enzyme with reported PLOOH-reducing activity, failed to prevent ferroptosis, its genetic loss sensitizes cancer cells to ferroptosis. Mechanistically, we uncover that PRDX6, beyond its known peroxidase activity, acts as a selenium-acceptor protein, facilitating intracellular selenium utilization and efficient selenium incorporation into selenoproteins, including GPX4. Its physiological significance was demonstrated by reduced GPX4 expression in Prdx6-deficient mouse brains and increased sensitivity to ferroptosis in PRDX6-deficient tumor xenografts in mice. Our study highlights PRDX6 as a critical player in directing cellular selenium utilization and dictating ferroptosis sensitivity.
    Keywords:  GPX4; LC-MS/MS; brain; cell death; cysteine; lipid peroxidation; selenite; selenocysteine; selenoproteins; tumor
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.028
  14. Mol Oncol. 2024 Nov 15.
    Global metabolomic profiling of tumor tissue and paired serum samples to identify biomarkers for response to neoadjuvant FOLFIRINOX treatment of human pancreatic cancer.
    Manoj Amrutkar, Sander Johannes Thorbjørnsen Guttorm, Anette Vefferstad Finstadsveen, Knut Jørgen Labori, Lars Eide, Helge Rootwelt, Katja Benedikte Prestø Elgstøen, Ivar P Gladhaug, Caroline S Verbeke.
      Neoadjuvant chemotherapy (NAT) is increasingly used for the treatment of non-metastatic pancreatic ductal adenocarcinoma (PDAC) and is established as a standard of care for borderline resectable and locally advanced PDAC. However, full exploitation of its clinical benefits is limited by the lack of biomarkers that assess treatment response. To address this unmet need, global metabolomic profiling was performed on tumor tissue and paired serum samples from patients with treatment-naïve (TN; n = 18) and neoadjuvant leucovorin calcium (folinic acid), fluorouracil, irinotecan hydrochloride and oxaliplatin (FOLFIRINOX)-treated (NAT; n = 17) PDAC using liquid chromatography mass spectrometry. Differentially abundant metabolites (DAMs) in TN versus NAT groups were identified and their correlation with various clinical parameters was assessed. Metabolomics profiling identified 40 tissue and five serum DAMs in TN versus NAT PDAC. In general, DAMs associated with amino acid and nucleotide metabolism were lower in NAT compared to TN. Four DAMs-3-hydroxybutyric acid (BHB), 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF), glycochenodeoxycholate and citrulline-were common to both tissue and serum and showed a similar pattern of differential abundance in both groups. A strong positive correlation was observed between serum carbohydrate 19-9 antigen (CA 19-9) and tissue carnitines (C12, C18, C18:2) and N8-acetylspermidine. The reduction in CA 19-9 following NAT correlated negatively with serum deoxycholate levels, and the latter correlated positively with survival. This study revealed neoadjuvant-chemotherapy-induced changes in metabolic pathways in PDAC, mainly amino acid and nucleotide metabolism, and these correlated with reduced CA 19-9 following neoadjuvant FOLFIRINOX treatment.
    Keywords:  FOLFIRINOX; LC–MS; neoadjuvant chemotherapy; pancreatic cancer; untargeted metabolomics
    DOI:  https://doi.org/10.1002/1878-0261.13759
  15. Langmuir. 2024 Nov 12.
    Iron-Induced Lipid Oxidation Alters Membrane Mechanics Favoring Permeabilization.
    Sara Lotfipour Nasudivar, Lohans Pedrera, Ana J García-Sáez.
      Ferroptosis is a form of regulated necrosis characterized by the iron-dependent accumulation of lipid peroxides in cell membranes. However, how lipid oxidation via iron-mediated Fenton reactions affects the biophysical properties of cellular membranes and how these changes contribute to the opening of plasma membrane pores are major questions in the field. Here, we characterized the dynamics of membrane alterations during lipid oxidation induced onsite by Fenton reactions in chemically defined in vitro model membrane systems. We find that lipid vesicle permeabilization kinetically correlates with the appearance of malondialdehyde (MDA), a product of lipid oxidation. Iron-induced lipid oxidation also alters the lateral organization of supported lipid bilayers (SLBs) with lipid phase coexistence in a time-dependent manner, reducing the lipid phase mismatch and the circularity of liquid ordered domains, which indicates a decrease in line tension at the phase boundaries. Further analysis of oxidized SLBs by force spectroscopy reveals a significant decrease in the average membrane breakthrough force upon oxidation, resulting from changes in lipid bilayer organization that make it more susceptible to permeabilization. Our findings suggest that lipid oxidation via iron-mediated Fenton-like reactions induces strong changes in membrane lipid interactions and mechanical properties leading to reduced line tension in the permeabilized state of the bilayer, which promotes membrane pore formation.
    DOI:  https://doi.org/10.1021/acs.langmuir.4c03294
  16. EMBO J. 2024 Nov 12.
    Calcium signaling from damaged lysosomes induces cytoprotective stress granules.
    Jacob Duran, Jay E Salinas, Rui Ping Wheaton, Suttinee Poolsup, Lee Allers, Monica Rosas-Lemus, Li Chen, Qiuying Cheng, Jing Pu, Michelle Salemi, Brett Phinney, Pavel Ivanov, Alf Håkon Lystad, Kiran Bhaskar, Jaya Rajaiya, Douglas J Perkins, Jingyue Jia.
      Lysosomal damage induces stress granule (SG) formation. However, the importance of SGs in determining cell fate and the precise mechanisms that mediate SG formation in response to lysosomal damage remain unclear. Here, we describe a novel calcium-dependent pathway controlling SG formation, which promotes cell survival during lysosomal damage. Mechanistically, the calcium-activated protein ALIX transduces lysosomal damage signals to SG formation by controlling eIF2α phosphorylation after sensing calcium leakage. ALIX enhances eIF2α phosphorylation by promoting the association between PKR and its activator PACT, with galectin-3 inhibiting this interaction; these regulatory events occur on damaged lysosomes. We further find that SG formation plays a crucial role in promoting cell survival upon lysosomal damage caused by factors such as SARS-CoV-2ORF3a, adenovirus, malarial pigment, proteopathic tau, or environmental hazards. Collectively, these data provide insights into the mechanism of SG formation upon lysosomal damage and implicate it in diseases associated with damaged lysosomes and SGs.
    Keywords:  ALG2-ALIX; Calcium-dependent Pathway; Lysosomal Damage; PACT-PKR-eIF2α; Stress Granules
    DOI:  https://doi.org/10.1038/s44318-024-00292-1
  17. Radiother Oncol. 2024 Nov 07. pii: S0167-8140(24)04268-3. [Epub ahead of print] 110606
    A metabolic switch to the pentose-phosphate pathway induces radiation resistance in pancreatic cancer.
    Ariel Shimoni-Sebag, Ifat Abramovich, Bella Agranovich, Rami Massri, Chani Stossel, Dikla Atias, Maria Raites-Gurevich, Keren Yizhak, Talia Golan, Eyal Gottlieb, Yaacov Richard Lawrence.
       PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is remarkably resistant to standard modalities, including radiotherapy. We hypothesized that metabolic reprogramming may underlie PDAC radioresistance, and moreover, that it would be possible to exploit these metabolic changes for therapeutic intent.
    METHODS AND MATERIALS: We established two matched models of radioresistant PDAC cells by exposing the AsPC-1 and MIAPaCa-2 human pancreatic cancer cells to incremental doses of radiation. The metabolic profile of parental and radioresistant cells was investigated using Nanostring technology, labeled-glucose tracing by liquid chromatography-mass spectrometry, Seahorse analysis and exposure to metabolic inhibitors. The synergistic effect of radiation combined with a pentose-phosphate pathway inhibitor, 6-aminonicotinamide (6-AN) was evaluated in a xenograft model established by subcutaneous injection of radioresistant-AsPC-1 cells into nude mice.
    RESULTS: The radioresistant cells overexpressed pyruvate dehydrogenase kinase (PDK) and consistently, displayed increased glycolysis and downregulated the tricarboxylic acid (TCA) cycle and oxidative phosphorylation. Metabolic flux through the pentose-phosphate pathway (PPP) was increased, as were levels of reduced glutathione; pharmacological inhibition of the PPP dramatically potentiated radiation-induced cell death. Furthermore, the combined treatment of radiation with the PPP inhibitor 6-AN synergistically inhibited tumor growth in-vivo.
    CONCLUSIONS: We provide a mechanistic understanding of the metabolic changes that underlie radioresistance in PDAC. Furthermore, we demonstrate that pancreatic cancer cells can be re-sensitized to radiation via metabolic manipulation, in particular, inhibition of the PPP. Exploitation of the metabolic vulnerabilities of radioresistant pancreatic cancer cells constitutes a new approach to pancreatic cancer, with a potential to improve clinical outcomes.
    Keywords:  DNA damage; Metabolic reprogramming; Pancreatic cancer; Pentose-phosphate pathway; Radiation resistance; Radiation therapy
    DOI:  https://doi.org/10.1016/j.radonc.2024.110606
  18. Carcinogenesis. 2024 Nov 08. pii: bgae063. [Epub ahead of print]
    CAFomics: convergence to translation for precision stroma approaches.
    Ian C McCabe, Xianlu L Peng, Joseph F Kearney, Jen Jen Yeh.
      A noticeable characteristic of pancreatic ductal adenocarcinoma (PDAC) tumors is a dense tumor microenvironment with abundant and dense, desmoplastic stroma woven tightly with both cellular and matrix components. The high stromal density is associated with higher intratumor pressures which, until the last decade, was largely assumed to be tumor protective, confirmed by early studies demonstrating that altering the stroma was effective in genetically engineered models of PDAC. However, clinical trials using these approaches have been disappointing. There is increasing recognition that stroma heterogeneity is much greater than initially thought with an explosion of investigation into cancer-associated fibroblast (CAF) subpopulations led by experimental and single-cell transcriptomic studies. This review summarizes and attempts to harmonize the current transcriptomic data of CAF subpopulations. Understanding the heterogeneity of CAFs, the matrix, and other tumor microenvironment features will be critical to developing effective therapeutic approaches. Identifying model systems that best recapitulate the clinical behavior and treatment response of human PDAC will be important. Examining subpopulations as defined by clinical outcome will remain a critical step in defining clinically impactful CAF subtypes in larger clinical cohorts. The future of precision oncology in PDAC will depend on the integration of precision tumor epithelial and precision stroma approaches.
    Keywords:  PDAC; cancer-associated fibroblast; tumor microenvironment
    DOI:  https://doi.org/10.1093/carcin/bgae063
  19. Proc Natl Acad Sci U S A. 2024 Nov 19. 121(47): e2413837121
    Parallel phosphoproteomics and metabolomics map the global metabolic tyrosine phosphoproteome.
    Alissandra L Hillis, Tigist Tamir, Grace E Perry, John M Asara, Jared L Johnson, Tomer M Yaron, Lewis C Cantley, Forest M White, Alex Toker.
      Tyrosine phosphorylation of metabolic enzymes is an evolutionarily conserved posttranslational modification that facilitates rapid and reversible modulation of enzyme activity, localization, or function. Despite the high abundance of tyrosine phosphorylation events detected on metabolic enzymes in high-throughput mass spectrometry-based studies, functional characterization of tyrosine phosphorylation sites has been limited to a subset of enzymes. Since tyrosine phosphorylation is dysregulated across human diseases, including cancer, understanding the consequences of metabolic enzyme tyrosine phosphorylation events is critical for informing disease biology and therapeutic interventions. To globally identify metabolic enzyme tyrosine phosphorylation events and simultaneously assign functional significance to these sites, we performed parallel phosphoproteomics and polar metabolomics in nontumorigenic mammary epithelial cells (MCF10A) stimulated with epidermal growth factor (EGF) in the absence or presence of the EGF receptor inhibitor erlotinib. We performed an integrated analysis of the phosphoproteomic and metabolomic datasets to identify tyrosine phosphorylation sites on metabolic enzymes with functional consequences. We identified two previously characterized (pyruvate kinase muscle isozyme, phosphoglycerate mutase 1) and two uncharacterized (glutathione S-transferase Pi 1, glutamate dehydrogenase 1) tyrosine phosphorylation sites on metabolic enzymes with purported functions based on metabolomic analyses. We validated these hits using a doxycycline-inducible CRISPR interference system in MCF10A cells, in which target metabolic enzymes were depleted with simultaneous reexpression of wild-type, phosphomutant, or phosphomimetic isoforms. Together, these data provide a framework for identification, prioritization, and characterization of tyrosine phosphorylation sites on metabolic enzymes with functional significance.
    Keywords:  EGFR; cancer metabolism; metabolomics; phosphotyrosine; proteomics
    DOI:  https://doi.org/10.1073/pnas.2413837121
  20. Nat Aging. 2024 Nov 13.
    Clearance of p21 highly expressing senescent cells accelerates cutaneous wound healing.
    Nathan S Gasek, Pengyi Yan, Junyu Zhu, K-Raman Purushothaman, Taewan Kim, Lichao Wang, Binsheng Wang, William F Flynn, Mingda Sun, Chun Guo, Billy Huggins, Roshanak Sharafieh, Yueying Zhou, Vojtech Parizek, Tamar Tchkonia, James L Kirkland, Saranya P Wyles, Ming Xu.
      While senescent cells have detrimental roles in several contexts, they are highly heterogeneous. p16 highly expressing senescent cells have been reported to exert beneficial functions in wound healing. Here we use Xenium spatial transcriptomics to identify a distinct p21 highly expressing senescent population induced on wounding, with a pro-inflammatory profile. We find that clearing p21 highly expressing cells expedites wound closure and is partially mediated by NF-κB inhibition, thus enhancing our understanding of the multifaceted functions of senescence in tissue remodeling.
    DOI:  https://doi.org/10.1038/s43587-024-00755-4
  21. Sci Adv. 2024 Nov 15. 10(46): eadf9758
    Cell response to extracellular matrix viscous energy dissipation outweighs high-rigidity sensing.
    Carla Huerta-López, Alejandro Clemente-Manteca, Diana Velázquez-Carreras, Francisco M Espinosa, Juan G Sanchez, Álvaro Martínez-Del-Pozo, María García-García, Sara Martín-Colomo, Andrea Rodríguez-Blanco, Ricardo Esteban-González, Francisco M Martín-Zamora, Luis I Gutierrez-Rus, Ricardo Garcia, Pere Roca-Cusachs, Alberto Elosegui-Artola, Miguel A Del Pozo, Elías Herrero-Galán, Pablo Sáez, Gustavo R Plaza, Jorge Alegre-Cebollada.
      The mechanics of the extracellular matrix (ECM) determine cell activity and fate through mechanoresponsive proteins including Yes-associated protein 1 (YAP). Rigidity and viscous relaxation have emerged as the main mechanical properties of the ECM steering cell behavior. However, how cells integrate coexisting ECM rigidity and viscosity cues remains poorly understood, particularly in the high-stiffness regime. Here, we have exploited engineered stiff viscoelastic protein hydrogels to show that, contrary to current models of cell-ECM interaction, substrate viscous energy dissipation attenuates mechanosensing even when cells are exposed to higher effective rigidity. This unexpected behavior is however readily captured by a pull-and-hold model of molecular clutch-based cell mechanosensing, which also recapitulates opposite cellular response at low rigidities. Consistent with predictions of the pull-and-hold model, we find that myosin inhibition can boost mechanosensing on cells cultured on dissipative matrices. Together, our work provides general mechanistic understanding on how cells respond to the viscoelastic properties of the ECM.
    DOI:  https://doi.org/10.1126/sciadv.adf9758
  22. Science. 2024 Nov 15. 386(6723): 768-776
    Pathogenic proteotoxicity of cryptic splicing is alleviated by ubiquitination and ER-phagy.
    Cristian Prieto-Garcia, Vigor Matkovic, Thorsten Mosler, Congxin Li, Jie Liang, James A Oo, Felix Haidle, Igor Mačinković, Alfredo Cabrera-Orefice, Rayene Berkane, Giulio Giuliani, Fenfen Xu, Anne-Claire Jacomin, Ines Tomaskovic, Marion Basoglu, Marina E Hoffmann, Rajeshwari Rathore, Ronay Cetin, Doha Boutguetait, Süleyman Bozkurt, María Clara Hernández Cañás, Mario Keller, Jonas Busam, Varun Jayeshkumar Shah, Ilka Wittig, Manuel Kaulich, Petra Beli, Wojciech P Galej, Ingo Ebersberger, Likun Wang, Christian Münch, Alexandra Stolz, Ralf P Brandes, William Ka Fai Tse, Stefan Eimer, Didier Y R Stainier, Stefan Legewie, Kathi Zarnack, Michaela Müller-McNicoll, Ivan Dikic.
      RNA splicing enables the functional adaptation of cells to changing contexts. Impaired splicing has been associated with diseases, including retinitis pigmentosa, but the underlying molecular mechanisms and cellular responses remain poorly understood. In this work, we report that deficiency of ubiquitin-specific protease 39 (USP39) in human cell lines, zebrafish larvae, and mice led to impaired spliceosome assembly and a cytotoxic splicing profile characterized by the use of cryptic 5' splice sites. Disruptive cryptic variants evaded messenger RNA (mRNA) surveillance pathways and were translated into misfolded proteins, which caused proteotoxic aggregates, endoplasmic reticulum (ER) stress, and, ultimately, cell death. The detrimental consequence of splicing-induced proteotoxicity could be mitigated by up-regulating the ubiquitin-proteasome system and selective autophagy. Our findings provide insight into the molecular pathogenesis of spliceosome-associated diseases.
    DOI:  https://doi.org/10.1126/science.adi5295
  23. Biochim Biophys Acta Rev Cancer. 2024 Nov 12. pii: S0304-419X(24)00139-2. [Epub ahead of print] 189208
    Mechanisms of cancer cachexia and targeted therapeutic strategies.
    Long Li, Zhi-Qiang Ling.
      Tumor cachexia is a multifactorial syndrome characterized by systemic dysfunction, including anorexia and severe weight loss that is resistant to standard nutritional interventions. It is estimated that approximately 20 % of cancer patients succumb to cachexia in the later stages of their disease. Thus, understanding its pathogenesis is vital for improving therapeutic outcomes. Recent research has focused on the imbalance between energy intake and expenditure in cachexia. Clinically, cachexia presents with anorexia, adipose tissue atrophy, and skeletal muscle wasting, each driven by distinct mechanisms. Anorexia arises primarily from tumor-secreted factors and cancer-induced hormonal disruptions that impair hypothalamic regulation of appetite. Adipose tissue atrophy is largely attributed to enhanced lipolysis, driven by increased activity of enzymes such as adipose triglyceride lipase and hormone-sensitive lipase, coupled with decreased lipoprotein lipase activity. The browning of white adipose tissue, facilitated by uncoupling protein 1, further accelerates fat breakdown by increasing energy expenditure. Skeletal muscle atrophy, a hallmark of cachexia, results from dysregulated protein turnover via the ubiquitin-proteasome and autophagy-lysosomal pathways, as well as mitochondrial dysfunction. Additionally, chemotherapy can exacerbate cachexia. This review examines the molecular mechanisms underlying cancer cachexia and discusses current therapeutic strategies, aiming to inform future research and improve treatment approaches.
    Keywords:  Anorexia; Lipoatrophy; Skeletal muscle atrophy; Treatment; cancer cachexia
    DOI:  https://doi.org/10.1016/j.bbcan.2024.189208
  24. J Nutr. 2024 Nov 05. pii: S0022-3166(24)01132-5. [Epub ahead of print]
    Dietary omega-3 (ω-3) fatty acids mitigate intestinal barrier integrity alterations in mice fed a high-fat diet: Implications for pancreatic carcinogenesis.
    Jazmin Machuca, Joanna Wirkus, Aya S Ead, Payam Vahmani, Karen E Matsukuma, Gerardo G Mackenzie, Patricia I Oteiza.
       BACKGROUND: Although body fatness is a recognized risk factor for pancreatic ductal adenocarcinoma (PDAC), the underlying mechanisms of how fat composition affects pancreatic carcinogenesis are poorly understood. High fat diets (HFD) can disrupt intestinal barrier function, potentially accelerating carcinogenesis. Omega-3 (ω-3) polyunsaturated fatty acids (FAs) have anti-inflammatory properties and help preserve intestinal integrity.
    OBJECTIVE: to evaluate how ω-3 FAs affect the colonic barrier in the context of HFD-induced changes, in a mouse model of PDAC [p48-Cre; LSL-KrasG12D (KC)].
    METHODS: Male and female KC mice were randomized into one of four groups: i) a control diet containing approximately 11% total calories from fat with an ω-6:ω-3 FA ratio of 10:1 (C); ii) the control diet with high levels of ω-3 FA with an ω-6:ω-3 FA ratio of 1:1 (Cω3); iii) a HFD containing 60% total calories from fat with an ω-6:ω-3 FA ratio of approximately 10:1 (HF); iv) a HFD with high levels of ω-3 FA with an ω-6:ω-3 FA ratio of 1:1 (HFω3).
    RESULTS: Consumption of a HFD for 8 weeks caused: i) disruption of tight junction structure and function; ii) decreased Goblet cell number, iii) higher colonic TLR4 and NOX1 expression; iv) activation of TLR4-triggered pathways, i.e. NF-κB, JNK1/2; v) elevated plasma LPS levels; v) higher pancreatic TLR4 expression, and vi) accelerated acinar-to-ductal metaplasia. All of these events were mitigated in mice fed the HFω3.
    CONCLUSIONS: Our findings support the concept that, in the context of obesity, ω-3 FA have protective effects during early-stage pancreatic carcinogenesis through the regulation of intestinal permeability and endotoxemia.
    Keywords:  high fat diet; intestinal permeability; obesity; omega-3 (ω-3) polyunsaturated fatty acids; pancreatic cancer; pancreatic carcinogenesis
    DOI:  https://doi.org/10.1016/j.tjnut.2024.10.054
  25. Nat Commun. 2024 Nov 07. 15(1): 9641
    Mapping membrane biophysical nano-environments.
    Luca Panconi, Jonas Euchner, Stanimir A Tashev, Maria Makarova, Dirk-Peter Herten, Dylan M Owen, Daniel J Nieves.
      The mammalian plasma membrane is known to contain domains with varying lipid composition and biophysical properties. However, studying these membrane lipid domains presents challenges due to their predicted morphological similarity to the bulk membrane and their scale being below the classical resolution limit of optical microscopy. To address this, we combine the solvatochromic probe di-4-ANEPPDHQ, which reports on its biophysical environment through changes in its fluorescence emission, with spectrally resolved single-molecule localisation microscopy. The resulting data comprises nanometre-precision localisation coordinates and a generalised polarisation value related to the probe's environment - a marked point pattern. We introduce quantification algorithms based on topological data analysis (PLASMA) to detect and map nano-domains in this marked data, demonstrating their effectiveness in both artificial membranes and live cells. By leveraging environmentally sensitive fluorophores, multi-modal single molecule localisation microscopy, and advanced analysis methods, we achieve nanometre scale mapping of membrane properties and assess changes in response to external perturbation with methyl-β-cyclodextrin. This integrated methodology represents an integrated toolset for investigating marked point pattern data at nanometre spatial scales.
    DOI:  https://doi.org/10.1038/s41467-024-53883-1
  26. Nat Cancer. 2024 Nov 11.
    The pro-oncogenic noncanonical activity of a RAS•GTP:RanGAP1 complex facilitates nuclear protein export.
    Brajendra K Tripathi, Nicole H Hirsh, Xiaolan Qian, Marian E Durkin, Dunrui Wang, Alex G Papageorge, Ross Lake, Yvonne A Evrard, Adam I Marcus, Suresh S Ramalingam, Mary Dasso, Karen H Vousden, James H Doroshow, Kylie J Walters, Douglas R Lowy.
      Canonical RAS signaling, including PI3K/AKT- and RAF/MEK-dependent activities, results mainly from RAS•GTP interaction with its effectors at the plasma membrane. Here, we identified a fundamental, oncogenic, noncanonical RAS•GTP activity that increases XPO1-dependent export of nuclear protein cargo into the cytoplasm and is independent of PI3K/AKT and RAF/MEK signaling. This RAS-dependent step acts downstream from XPO1 binding to nuclear protein cargo and is mediated by a perinuclear protein complex between RAS•GTP and RanGAP1 that facilitates hydrolysis of Ran•GTP to Ran•GDP, which promotes release of nuclear protein cargo into the cytoplasm. The export of nuclear EZH2, which promotes cytoplasmic degradation of the DLC1 tumor suppressor protein, is a biologically important component of this pro-oncogenic activity. Conversely, preventing nuclear protein export contributes to the antitumor activity of KRAS inhibition, which can be further augmented by reactivating the tumor suppressor activity of DLC1 or potentially combining RAS inhibitors with other cancer treatments.
    DOI:  https://doi.org/10.1038/s43018-024-00847-5
  27. Appl Environ Microbiol. 2024 Nov 13. e0121224
    Exploring the interplay between yeast cell membrane lipid adaptation and physiological response to acetic acid stress.
    Fei Wu, Maurizio Bettiga, Lisbeth Olsson.
      Acetic acid is a byproduct of lignocellulose pretreatment and a potent inhibitor of yeast-based fermentation processes. A thicker yeast plasma membrane (PM) is expected to retard the passive diffusion of undissociated acetic acid into the cell. Molecular dynamic simulations suggest that membrane thickness can be increased by elongating glycerophospholipids (GPL) fatty acyl chains. Previously, we successfully engineered Saccharomyces cerevisiae to increase GPL fatty acyl chain length but failed to lower acetic acid net uptake. Here, we tested whether altering the relative abundance of diacylglycerol (DAG) might affect PM permeability to acetic acid in cells with longer GPL acyl chains (DAGEN). To this end, we expressed diacylglycerol kinase α (DGKα) in DAGEN. The resulting DAGEN_Dgkα strain exhibited restored DAG levels, grew in medium containing 13 g/L acetic acid, and accumulated less acetic acid. Acetic acid stress and energy burden were accompanied by increased glucose uptake in DAGEN_Dgkα cells. Compared to DAGEN, the relative abundance of several membrane lipids changed in DAGEN_Dgkα in response to acetic acid stress. We propose that the ability to increase the energy supply and alter membrane lipid composition could compensate for the negative effect of high net acetic acid uptake in DAGEN_Dgkα under stressful conditions.
    IMPORTANCE: In the present study, we successfully engineered a yeast strain that could grow under high acetic acid stress by regulating its diacylglycerol metabolism. We compared how the plasma membrane and total cell membranes responded to acetic acid by adjusting their lipid content. By combining physiological and lipidomics analyses in cells cultivated in the absence or presence of acetic acid, we found that the capacity of the membrane to adapt lipid composition together with sufficient energy supply influenced membrane properties in response to stress. We suggest that potentiating the intracellular energy system or enhancing lipid transport to destination membranes should be taken into account when designing membrane engineering strategies. The findings highlight new directions for future yeast cell factory engineering.
    Keywords:  Dgkα; diacylglycerol; energy burden; glucose uptake rate; lipidomics; membrane permeability
    DOI:  https://doi.org/10.1128/aem.01212-24
  28. Angew Chem Int Ed Engl. 2024 Nov 13. e202416608
    GlutaR: A High-Performance Fluorescent Protein-Based Sensor for Spatiotemporal Monitoring of Glutamine Dynamics In Vivo.
    Bingjie Liu, Zijie Zhao, Pengcheng Wang, Kamiran Aihemaiti, Lixin Zhu, Qingpeng Wei, Wenzhe Li, Xia Yuan, Jing Wu, Changtao Jiang, Min Hao, Jing Wang.
      Glutamine is the most abundant amino acid in human blood and muscle, and is integral to a wide variety of functions in cancer cells. However, the inability to monitor the subcellular distribution of glutamine in real-time has obscured understanding of glutamine metabolism under physiological and pathological conditions. Here, we report the development of a genetically encoded fluorescent sensor and demonstrate how this GlnBP-cpYFP fusion "GlutaR sensor" undergoes glutamine-induced conformational changes reflected in detectable fluorescence responses. Obtained after iterative screening of approximately 1,600 variants, GlutaR exhibits a ratiometric readout, fast response kinetics, and high responsivity, and we demonstrate its selectivity for monitoring glutamine fluctuations in multiple cell types. Additionally, using digitonin permeabilization of GlutaR HeLa cells, we generated a calibration curve and performed in situ titration to quantify free glutamine concentrations in subcellular compartments (cytosol, nucleus, mitochondria). Subsequently, we applied GlutaR to investigate how chemical and genetic inhibition of GS and GLS differentially alter glutamine levels in subcellular compartments. Finally, we demonstrate GlutaR's ability to monitor dynamic glutamine levels in muscle and liver tissues of diabetic mice in vivo. These findings collectively demonstrate GlutaR as a versatile tool for the spatiotemporal characterization of glutamine metabolism in living cells and tissues.
    Keywords:  glutamine* biosensor* fluorescence imaging * metabolism* live mice
    DOI:  https://doi.org/10.1002/anie.202416608
  29. Carcinogenesis. 2024 Nov 08. pii: bgae064. [Epub ahead of print]
    From precursor to cancer: decoding the intrinsic and extrinsic pathways of pancreatic intraepithelial neoplasia progression.
    Sarah Graham, Mariia Dmitrieva, Debora Barbosa Vendramini-Costa, Ralph Francescone, Maria A Trujillo, Edna Cukierman, Laura D Wood.
      This review explores the progression of pancreatic intraepithelial neoplasia (PanIN) to pancreatic ductal adenocarcinoma through a dual lens of intrinsic molecular alterations and extrinsic microenvironmental influences. PanIN development begins with Kirsten rat sarcoma viral oncogene (KRAS) mutations driving PanIN initiation. Key additional mutations in cyclin-dependent kinase inhibitor 2A (CDKN2A), tumor protein p53 (TP53), and mothers against decapentaplegic homolog 4 (SMAD4) disrupt cell cycle control and genomic stability, crucial for PanIN progression from low-grade to high-grade dysplasia. Additional molecular alterations in neoplastic cells, including epigenetic modifications and chromosomal alterations, can further contribute to neoplastic progression. In parallel with these alterations in neoplastic cells, the microenvironment, including fibroblast activation, extracellular matrix remodeling, and immune modulation, plays a pivotal role in PanIN initiation and progression. Crosstalk between neoplastic and stromal cells influences nutrient support and immune evasion, contributing to tumor development, growth, and survival. This review underscores the intricate interplay between cell-intrinsic molecular drivers and cell-extrinsic microenvironmental factors, shaping PanIN predisposition, initiation, and progression. Future research aims to unravel these interactions to develop targeted therapeutic strategies and early detection techniques, aiming to alleviate the severe impact of pancreatic cancer by addressing both genetic predispositions and environmental influences.
    Keywords:  cancer-associated fibroblasts; cell extrinsic; cell intrinsic; driver mutations; pancreatic ductal adenocarcinoma; pancreatic intraepithelial neoplasia; tumor microenvironment
    DOI:  https://doi.org/10.1093/carcin/bgae064
  30. Nat Commun. 2024 Nov 08. 15(1): 9679
    A tuneable minimal cell membrane reveals that two lipid species suffice for life.
    Isaac Justice, Petra Kiesel, Nataliya Safronova, Alexander von Appen, James P Saenz.
      All cells are encapsulated by a lipid membrane that facilitates their interactions with the environment. How cells manage diverse mixtures of lipids, which dictate membrane property and function, is experimentally challenging to address. Here, we present an approach to tune and minimize membrane lipid composition in the bacterium Mycoplasma mycoides and its derived 'minimal cell' (JCVI-Syn3A), revealing that a two-component lipidome can support life. Systematic reintroduction of phospholipids with different features demonstrates that acyl chain diversity is more important for growth than head group diversity. By tuning lipid chirality, we explore the lipid divide between Archaea and the rest of life, showing that ancestral lipidomes could have been heterochiral. However, in these simple organisms, heterochirality leads to impaired cellular fitness. Thus, our approach offers a tunable minimal membrane system to explore the fundamental lipidomic requirements for life, thereby extending the concept of minimal life from the genome to the lipidome.
    DOI:  https://doi.org/10.1038/s41467-024-53975-y
  31. Proc Natl Acad Sci U S A. 2024 Nov 12. 121(46): e2409509121
    The Warburg Effect is the result of faster ATP production by glycolysis than respiration.
    Matthew A Kukurugya, Saharon Rosset, Denis V Titov.
      Many prokaryotic and eukaryotic cells metabolize glucose to organism-specific by-products instead of fully oxidizing it to carbon dioxide and water-a phenomenon referred to as the Warburg Effect. The benefit to a cell is not fully understood, given that partial metabolism of glucose yields an order of magnitude less adenosine triphosphate (ATP) per molecule of glucose than complete oxidation. Here, we test a previously formulated hypothesis that the benefit of the Warburg Effect is to increase ATP production rate by switching from high-yielding respiration to faster glycolysis when excess glucose is available and respiration rate becomes limited by proteome occupancy. We show that glycolysis produces ATP faster per gram of pathway protein than respiration in Escherichia coli, Saccharomyces cerevisiae, and mammalian cells. We then develop a simple mathematical model of energy metabolism that uses five experimentally estimated parameters and show that this model can accurately predict absolute rates of glycolysis and respiration in all three organisms under diverse conditions, providing strong support for the validity of the ATP production rate maximization hypothesis. In addition, our measurements show that mammalian respiration produces ATP up to 10-fold slower than respiration in E. coli or S. cerevisiae, suggesting that the ATP production rate per gram of pathway protein is a highly evolvable trait that is heavily optimized in microbes. We also find that E. coli respiration is faster than fermentation, explaining the observation that E. coli, unlike S. cerevisiae or mammalian cells, never switch to pure fermentation in the presence of oxygen.
    Keywords:  Warburg Effect; cancer metabolism; energy metabolism; modeling; systems biology
    DOI:  https://doi.org/10.1073/pnas.2409509121
  32. J Physiol. 2024 Nov 09.
    Daily GDF15 treatment has sex-specific effects on body weight and food intake and does not enhance the effects of voluntary physical activity in mice.
    Stewart Jeromson, Michael Akcan, Bradley Baranowski, Meagan Arbeau, Annalaura Bellucci, David C Wright.
      Growth differentiation factor 15 (GDF15) is a stress-induced cytokine that suppresses food intake and causes weight loss. GDF15 also reduces voluntary physical activity and, thus, it is not clear whether combining GDF15 with exercise will be beneficial or if reductions in food intake would be offset by decreases in physical activity. We investigated how GDF15 treatment combined with voluntary wheel running (VWR) would impact weight gain, food intake, adiposity and indices of metabolic health in mice. High-fat fed male and female mice underwent daily GDF15 treatments and were given access to voluntary running wheels, or not, for 11 days. In both sexes, VWR prevented weight gain. In males, GDF15 reduced food intake, as well as attenuated weight gain and the accumulation of adipose tissue, with no additional effect of VWR. In female mice, GDF15 did not impact body weight gain or body composition. GDF15 acutely reduced food intake in female mice but this was followed by a period of rebound hyperphagia and consequently GDF15 did not reduce total food intake in female mice. GDF15 treatment reduced wheel running distance in both sexes. There were main effects of VWR to improve glucose tolerance in female but not male mice. These findings show that GDF15 has sex-specific effects on food intake and consequently weight gain and adiposity. There is no added benefit of combining GDF15 and voluntary physical activity for weight loss. Adaptive responses to acute caloric restriction induced by GDF15 might limit its effectiveness as a weight loss tool in females. KEY POINTS: GDF15 is a stress-induced signalling factor that reduces food intake and voluntary physical activity. It is not known whether combining GDF15 treatment with voluntary wheel running would impart beneficial combined effects in attenuating weight gain and the accumulation of adipose tissue. In the present study, we demonstrate that GDF15 reduces food intake and prevents weight gain in male but not female mice consuming a high-fat diet and also that combining GDF15 with voluntary wheel running (VWR) does not lead to a greater dampening of weight gain. In female mice, GDF15 acutely reduced food intake, but this was followed by a period of rebound hyperphagia resulting in no differences in total food intake. In both sexes, VWR was equivalent, or superior to GDF15 in preventing weight gain.
    Keywords:  GDF15; food intake; mice; obesity; physical activity
    DOI:  https://doi.org/10.1113/JP287256
  33. Function (Oxf). 2024 Nov 15. pii: zqae050. [Epub ahead of print]
    Pancreatic ductal adenocarcinoma, β-blockers and antihistamines: A clinical trial is needed.
    Jillian G Baker, Erica K Sloan, Kevin Pfleger, Peter J McCormick, Cristina Salmerón, Paul A Insel.
      Survival in pancreatic cancer (pancreatic ductal adenocarcinoma, PDAC) at just 5 months is the worst of all cancers. It is predicted to become the second highest cause of cancer deaths worldwide this decade and unlike most cancers, there has been little progress in improving survival in PDAC. Numerous studies including molecular and mechanistic studies, cancer biology studies and retrospective human epidemiological studies suggest that two well-known, approved drug classes - β-blockers and H1-antihistamines - may be beneficial and thus may potentially prolong life in patients with PDAC. In our opinion, the body of evidence has reached a point where the potential gains outweigh the very low risks involved in a clinical study in PDAC. We thus believe that it is now time for a clinical trial involving these two agents in PDAC patients. As a repurposing of generic drugs, this is not likely to be appealing to pharmaceutical companies and therefore is likely to require governmental, philanthropic and /or charitable organisational input. In this article, we opine and propose that an urgent clinical trial is needed to determine if repurposing these two orally administered, inexpensive, largely safe drug classes, either alone or in combination, could prolong survival in PDAC and thus improve the outcome for the 10,000 people worldwide who die from PDAC each week.
    Keywords:  antihistamine; cancer; gastrointestinal; histamine receptor; pancreas; pancreatic cancer; repurposing; β-adrenoceptor; β-antagonist; β-blocker
    DOI:  https://doi.org/10.1093/function/zqae050
  34. Cell Rep. 2024 Nov 13. pii: S2211-1247(24)01330-5. [Epub ahead of print]43(11): 114979
    Leader cells promote immunosuppression to drive ovarian cancer progression in vivo.
    Amy L Wilson, Laura R Moffitt, Brittany R Doran, Bashira Basri, Jennie Do, Thomas W Jobling, Magdalena Plebanski, Andrew N Stephens, Maree Bilandzic.
      Over 75% of patients with ovarian cancer present with late-stage disease, often accompanied by extensive metastasis. The metastatic cascade is driven by a sub-population of transcriptionally plastic cells known as "leader cells" (LCs), which play a critical role in collective invasion yet remain poorly understood. LCs are marked by the expression of keratin-14 (KRT14), which determines their migratory and invasive capacity in ovarian cancer. This study demonstrates that KRT14+ LCs promote tumor progression through immunosuppression and immune privilege in vivo. In the ID8 syngeneic epithelial ovarian cancer mouse model, tumor-specific loss of KRT14+ LCs impairs tumor progression and metastatic spread without affecting cellular proliferation. Immune profiling shows reduced immunosuppressive regulatory T cells (Tregs) and M2 macrophages and improved CD8+ T cell/Treg ratios in LC knockout (LCKO) mice. Conversely, forced LC overexpression accelerates metastasis and increases the secretion of immunosuppressive chemokines, such as CCL22 and CCL5, highlighting the role of KRT14+ LCs in immune suppression and metastatic progression.
    Keywords:  CP: Cancer; T regulatory cells; cancer metastasis; chemokines; immunosuppression; keratin-14; leader cells; ovarian cancer
    DOI:  https://doi.org/10.1016/j.celrep.2024.114979
  35. Immunotherapy. 2024 Nov 15. 1-3
    Could senescent cells be the prescription for therapeutic cancer vaccines?
    Yue Liu, Stephen J Kron.
      
    Keywords:  cancer vaccine; immune checkpoint blockade; immunotherapy; senescence; senescence-associated secretory phenotype (SASP)
    DOI:  https://doi.org/10.1080/1750743X.2024.2422813
  36. United European Gastroenterol J. 2024 Nov 14.
    Organoid-based precision medicine in pancreatic cancer.
    Alica K Beutel, Menar Ekizce, Thomas J Ettrich, Thomas Seufferlein, Jessica Lindenmayer, Johann Gout, Alexander Kleger.
      Pancreatic ductal adenocarcinoma (PDAC) ranks among the leading causes of cancer-related deaths worldwide. Despite advances in precision oncology in other malignancies, treatment of PDAC still largely relies on conventional chemotherapy. Given the dismal prognosis and heterogeneity in PDAC, there is an urgent need for personalized therapeutic strategies to improve treatment response. Organoids, generated from patients' tumor tissue, have emerged as a powerful tool in cancer research. These three-dimensional models faithfully recapitulate the morphological and genetic features of the parental tumor and retain patient-specific heterogeneity. This review summarizes existing precision oncology approaches in PDAC, explores current applications and limitations of organoid cultures in personalized medicine, details preclinical studies correlating in vitro organoid prediction and patient treatment response, and provides an overview of ongoing organoid-based clinical trials.
    Keywords:  PDAC; drug response prediction; oncology; pancreatic ductal adenocarcinoma; patient‐derived organoids; personalized therapy; pharmacotyping; precision oncology; treatment response
    DOI:  https://doi.org/10.1002/ueg2.12701
  37. Cell Rep. 2024 Nov 13. pii: S2211-1247(24)01206-3. [Epub ahead of print]43(11): 114855
    Tissue-colonizing disseminated tumor cells secrete prostaglandin E2 to promote NK cell dysfunction and evade anti-metastatic immunity.
    Anna-Marie Pedde, Hyunu Kim, Sainitin Donakonda, Tobias Baumann, Felix Bayerl, Philippa Meiser, Anna Hirschberger, Christine Klement, Simon Grassmann, Rupert Öllinger, Norbert Hüser, Daniel Hartmann, Melanie Laschinger, Joseph A Trapani, Alfred Zippelius, Tobias Bald, Gabriela M Wiedemann, Roland Rad, Joseph C Sun, Bastian Höchst, Jan P Böttcher.
      Natural killer (NK) cells are critical for anti-metastatic immunity and can eliminate metastasizing tumor cells within circulation and sites of metastatic seeding. Here, we show that disseminated tumor cells (DTCs) colonizing the mouse lung secrete prostaglandin E2 (PGE2) to locally induce NK cell dysfunction, allowing outgrowing metastases to escape immune control and establish metastatic disease. Mechanistically, PGE2 signaling through its receptors EP2 and EP4 mediates NK cell dysfunction, which leads to reprogramming of NK cell gene expression and results in impaired production of anti-metastatic cytokines. In human cancer patients, the PGE2-EP2/EP4 axis is associated with NK cell dysfunction within distant organ metastases. Disabling EP2/EP4 signaling in NK cells prevents their dysfunction in DTC-colonized lungs and achieves effective NK cell-mediated control of metastatic disease. Our findings reveal a suppressive signaling axis exploited by metastasizing tumor cells to escape immune control in distant organs that could be targeted for metastatic cancer therapy.
    Keywords:  CP: Cancer; CP: Immunology
    DOI:  https://doi.org/10.1016/j.celrep.2024.114855
  38. Mol Cell. 2024 Nov 05. pii: S1097-2765(24)00865-7. [Epub ahead of print]
    Opposing roles for AMPK in regulating distinct mitophagy pathways.
    Marianna Longo, Aniketh Bishnu, Pierpaolo Risiglione, Lambert Montava-Garriga, Joyceline Cuenco, Kei Sakamoto, Carol MacKintosh, Ian G Ganley.
      Mitophagy degrades damaged mitochondria, but we show here that it can also target functional mitochondria. This latter scenario occurs during programmed mitophagy and involves the mitophagy receptors NIX and BNIP3. Although AMP-activated protein kinase (AMPK), the energy-sensing protein kinase, can influence damaged-induced mitophagy, its role in programmed mitophagy is unclear. We found that AMPK directly inhibits NIX-dependent mitophagy by triggering 14-3-3-mediated sequestration of ULK1, via ULK1 phosphorylation at two sites: Ser556 and an additional identified site, Ser694. By contrast, AMPK activation increases Parkin phosphorylation and enhances the rate of depolarization-induced mitophagy, independently of ULK1. We show that this happens both in cultured cells and tissues in vivo, using the mito-QC mouse model. Our work unveils a mechanism whereby AMPK activation downregulates mitophagy of functional mitochondria but enhances that of dysfunctional/damaged ones.
    Keywords:  14-3-3; AMPK; NIX; Parkin; ULK1; autophagy; liver; mito-QC; mitophagy; skeletal muscle
    DOI:  https://doi.org/10.1016/j.molcel.2024.10.025
  39. Cancer Res. 2024 Nov 15.
    Tumor heterogeneity and cooperating cancer hallmarks driven by divergent EMT programs.
    Phoebe Carter, Yibin Kang.
      Epithelial-to-mesenchymal transition (EMT) is known to play roles in orchestrating cellular plasticity across many physiological and pathological contexts. Partial EMT, wherein cells maintain both epithelial and mesenchymal features, is gaining recognition for its functional importance in cancer in recent years. There are many factors regulating both partial and full EMT, and the precise mechanisms underlying these processes vary depending on the biological context. Furthermore, how different EMT states cooperate to create a heterogeneous tumor population and promote different pro-malignant features remains largely undefined. In a recent study published in Nature Cancer, Youssef and colleagues described how two disparate EMT programs, active in either organ fibrosis or embryonic development, are utilized within different cells within the same murine mammary tumor model. This work provides mechanistic insight into the development of intratumoral heterogeneity, providing evidence for the cooperation between the two EMT trajectories.
    DOI:  https://doi.org/10.1158/0008-5472.CAN-24-4309
  40. Pancreatology. 2024 Oct 29. pii: S1424-3903(24)00784-1. [Epub ahead of print]
    Early detection of pancreatic cancer: Study design and analytical considerations in biomarker discovery and early phase validation studies.
    Lynette M Smith, Douglas W Mahoney, William R Bamlet, Fang Yu, Suyu Liu, Michael G Goggins, Sourat Darabi, Shounak Majumder, Qiao-Li Wang, Gregory A Coté, Michael J Demeure, Zhen Zhang, Sudhir Srivastava, Akhil Chawla, Grant Izmirlian, Janet E Olson, Brian M Wolpin, Jeanine M Genkinger, Kenneth S Zaret, Randall Brand, Eugene J Koay, Ann L Oberg.
       OBJECTIVES: Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease that is challenging to detect at an early stage. Biomarkers are needed that can detect PDAC early in the course of disease when interventions lead to the best outcomes. We highlight study design and statistical considerations that inform pancreatic cancer early detection biomarker evaluation.
    METHODS: We describe experimental design strategies in this setting useful for streamlining biomarker evaluation at each Early Detection Research Network (EDRN) phase of biomarker development. We break the early EDRN phases into sub-phases, proposing objectives, study design strategies, and biomarker performance benchmarks.
    RESULTS: The goal of early detection in populations at high-risk of PDAC is described. Evaluating biomarker behavior in patients under surveillance without disease can winnow candidate biomarkers. Potential resources for biomarker validation studies are described.
    CONCLUSIONS: Multisite and multidisciplinary collaboration can facilitate study design strategies in this lethal but low incidence disease and streamline the path from biomarker discovery to clinical use. Improvements in analytical and experimental design methods could help accelerate biomarker evaluation through the phases of biomarker development.
    Keywords:  Biomarker development; Early detection; Experimental design; Pancreatic cancer; Validation
    DOI:  https://doi.org/10.1016/j.pan.2024.10.012
  41. J Clin Invest. 2024 Nov 15. pii: e176851. [Epub ahead of print]134(22):
    Lactate reprograms glioblastoma immunity through CBX3-regulated histone lactylation.
    Shuai Wang, Tengfei Huang, Qiulian Wu, Huairui Yuan, Xujia Wu, Fanen Yuan, Tingting Duan, Suchet Taori, Yingming Zhao, Nathaniel W Snyder, Dimitris G Placantonakis, Jeremy N Rich.
      Glioblastoma (GBM), an aggressive brain malignancy with a cellular hierarchy dominated by GBM stem cells (GSCs), evades antitumor immunity through mechanisms that remain incompletely understood. Like most cancers, GBMs undergo metabolic reprogramming toward glycolysis to generate lactate. Here, we show that lactate production by patient-derived GSCs and microglia/macrophages induces tumor cell epigenetic reprogramming through histone lactylation, an activating modification that leads to immunosuppressive transcriptional programs and suppression of phagocytosis via transcriptional upregulation of CD47, a "don't eat me" signal, in GBM cells. Leveraging these findings, pharmacologic targeting of lactate production augments efficacy of anti-CD47 therapy. Mechanistically, lactylated histone interacts with the heterochromatin component chromobox protein homolog 3 (CBX3). Although CBX3 does not possess direct lactyltransferase activity, CBX3 binds histone acetyltransferase (HAT) EP300 to induce increased EP300 substrate specificity toward lactyl-CoA and a transcriptional shift toward an immunosuppressive cytokine profile. Targeting CBX3 inhibits tumor growth by both tumor cell-intrinsic mechanisms and increased tumor cell phagocytosis. Collectively, these results suggest that lactate mediates metabolism-induced epigenetic reprogramming in GBM that contributes to CD47-dependent immune evasion, which can be leveraged to augment efficacy of immuno-oncology therapies.
    Keywords:  Adult stem cells; Brain cancer; Epigenetics; Metabolism; Oncology
    DOI:  https://doi.org/10.1172/JCI176851
  42. Clin Epigenetics. 2024 Nov 10. 16(1): 156
    DNA methylation classifier to diagnose pancreatic ductal adenocarcinoma metastases from different anatomical sites.
    Teodor G Calina, Eilís Perez, Elena Grafenhorst, Jamal Benhamida, Simon Schallenberg, Adrian Popescu, Ines Koch, Tobias Janik, BaoQing Chen, Jana Ihlow, Stephanie Roessler, Benjamin Goeppert, Bruno Sinn, Marcus Bahra, George A Calin, Eliane T Taube, Uwe Pelzer, Christopher C M Neumann, David Horst, Erik Knutsen, David Capper, Mihnea P Dragomir.
       BACKGROUND: We have recently constructed a DNA methylation classifier that can discriminate between pancreatic ductal adenocarcinoma (PAAD) liver metastasis and intrahepatic cholangiocarcinoma (iCCA) with high accuracy (PAAD-iCCA-Classifier). PAAD is one of the leading causes of cancer of unknown primary and diagnosis is based on exclusion of other malignancies. Therefore, our focus was to investigate whether the PAAD-iCCA-Classifier can be used to diagnose PAAD metastases from other sites.
    METHODS: For this scope, the anomaly detection filter of the initial classifier was expanded by 8 additional mimicker carcinomas, amounting to a total of 10 carcinomas in the negative class. We validated the updated version of the classifier on a validation set, which consisted of a biological cohort (n = 3579) and a technical one (n = 15). We then assessed the performance of the classifier on a test set, which included a positive control cohort of 16 PAAD metastases from various sites and a cohort of 124 negative control samples consisting of 96 breast cancer metastases from 18 anatomical sites and 28 carcinoma metastases to the brain.
    RESULTS: The updated PAAD-iCCA-Classifier achieved 98.21% accuracy on the biological validation samples, and on the technical validation ones it reached 100%. The classifier also correctly identified 15/16 (93.75%) metastases of the positive control as PAAD, and on the negative control, it correctly classified 122/124 samples (98.39%) for a 97.85% overall accuracy on the test set. We used this DNA methylation dataset to explore the organotropism of PAAD metastases and observed that PAAD liver metastases are distinct from PAAD peritoneal carcinomatosis and primary PAAD, and are characterized by specific copy number alterations and hypomethylation of enhancers involved in epithelial-mesenchymal-transition.
    CONCLUSIONS: The updated PAAD-iCCA-Classifier (available at https://classifier.tgc-research.de/ ) can accurately classify PAAD samples from various metastatic sites and it can serve as a diagnostic aid.
    Keywords:  Cancer of unknown primary; DNA methylation; Epigenetics; Molecular diagnosis; Pancreatic ductal adenocarcinoma
    DOI:  https://doi.org/10.1186/s13148-024-01768-x
  43. BJC Rep. 2024 Jan 23. 2(1): 3
    A Phase 2 study of nivolumab in combination with modified FOLFIRINOX for metastatic pancreatic cancer.
    Chigusa Morizane, Makoto Ueno, Masafumi Ikeda, Kentaro Sudo, Yoshinori Hirashima, Masataka Kuroda, Shinji Ueno, Takuji Okusaka, Junji Furuse.
       BACKGROUND: Nivolumab with modified FOLFIRINOX (mFOLFIRINOX) may have additive antitumour effects while minimising chemotherapy cytotoxicity. We assessed the efficacy and safety of nivolumab+mFOLFIRINOX in metastatic pancreatic cancer.
    METHODS: Thirty-one treatment-naïve patients aged ≥20 years with metastatic unresectable/recurrent pancreatic cancer (≥1 measurable lesion per Response Evaluation Criteria in Solid Tumours version 1.1) and Eastern Cooperative Oncology Group 0/1 score and life expectancy ≥90 days received nivolumab (480 mg, every 4 weeks) plus mFOLFIRINOX. The primary endpoint was objective response rate (ORR). Secondary endpoints included overall survival (OS), progression-free survival (PFS) and safety.
    RESULTS: At the median follow-up of 13.4 months, the ORR was 32.3% (complete response 0%; partial response 32.3%) and the median duration of response was 7.4 (range: 3.5-21.9) months; the primary endpoint was not met. Median OS and PFS were 13.4 (95% confidence interval [CI]: 10.6-16.6) months and 7.4 (95% CI: 3.9-9.2) months, respectively. The 1-year survival rate was 54.8% (95% CI: 36.0%-70.3%). Drug-related serious adverse events were reported in 29.0% of the patients; 3.2% drug-related adverse events led to discontinuation, and none led to death within 30-day safety window.
    CONCLUSION: Nivolumab+mFOLFIRINOX was tolerable in patients with metastatic pancreatic cancer. ORR and survival were comparable to previously reported data. (JapicCTI-184230).
    DOI:  https://doi.org/10.1038/s44276-023-00028-4
  44. J Chem Theory Comput. 2024 Nov 11.
    Nonequilibrium Molecular Dynamics Method to Generate Poiseuille-Like Flow between Lipid Bilayers.
    Masaki Otawa, Satoru G Itoh, Hisashi Okumura.
      There are various flows inside and outside cells in vivo. Nonequilibrium molecular dynamics (NEMD) simulation is a useful tool for understanding the effects of these flows on the dynamics of biomolecules. We propose an NEMD method to generate a Poiseuille-like flow between lipid bilayers. We extended the conventional equilibrium MD method to produce a flow by adding constant external force terms to the water molecules. Using the Lagrange multiplier method, the center of mass of the lipid bilayer is constrained so that the flow does not sweep away the lipid bilayer, but the individual lipid molecules fluctuate. The temperature of the system is controlled properly in the solution and membrane by using the Nosé-Hoover thermostat. We found that the flow velocity increases linearly as the applied external force term increases. It is possible to estimate the appropriate value of acceleration to generate a flow with an arbitrary velocity using this proportional relation once a single short MD simulation is performed. We also found that the flow between two lipid bilayers is slower than the analytical solution of the Navier-Stokes equations between rigid parallel plates due to the interactions between water molecules and the membrane. This method can be applied not only to a flow on lipid membranes but also to a flow on soft surfaces generally.
    DOI:  https://doi.org/10.1021/acs.jctc.4c00750
  45. Sci Rep. 2024 11 11. 14(1): 27584
    Coherent Raman microscopy visualizes ongoing cellular senescence through amide I peak shifts originating from β sheets in disordered nucleolar proteins.
    Shigeo Ishibashi, Akihito Inoko, Yuki Oka, Philippe Leproux, Hideaki Kano.
      Cellular senescence occurs through the accumulation of many kinds of stresses. Senescent cells in tissues also cause various age-related disorders. Therefore, detecting them without labeling is beneficial for medical research and developing diagnostic methods. However, existing biomarkers have limitations of requiring fixation and labeling, or their molecular backgrounds are uncertain. Coherent anti-Stokes Raman scattering (CARS) spectroscopic imaging is a novel option because it can assess and visualize molecular structures based on their molecular fingerprint. Here, we present a new label-free method to visualize cellular senescence using CARS imaging in nucleoli. We found the peak of the nucleolar amide I band shifted to a higher wavenumber in binuclear senescent cells, which reflects changes in the protein secondary structure from predominant α-helices to β-sheets originating from amyloid-like aggregates. Following this, we developed a procedure that can visualize the senescent cells by providing the ratios and subtractions of these two components. We also confirmed that the procedure can visualize nucleolar aggregates due to unfolded/misfolded proteins produced by proteasome inhibition. Finally, we found that this method can help visualize the nucleolar defects in naïve cells even before binucleation. Thus, our method is beneficial to evaluate ongoing cellular senescence through label-free imaging of nucleolar defects.
    Keywords:  Amide I; Cellular senescence; Coherent anti-stokes Raman scattering (CARS) microscopy; LLPS (liquid-liquid phase separation); Nucleolus; Raman
    DOI:  https://doi.org/10.1038/s41598-024-78899-x
  46. Int J Radiat Oncol Biol Phys. 2024 Nov 13. pii: S0360-3016(24)03561-2. [Epub ahead of print]
    Phase I Study of Adjuvant Allogeneic GM-CSF-Transduced Pancreatic Tumor Cell Vaccine, Low Dose Cyclophosphamide, and SBRT followed by FFX in High-Risk Resected Pancreatic Ductal Adenocarcinoma.
    Colin S Hill, Rose Parkinson, Elizabeth M Jaffee, Elizabeth Sugar, Lei Zheng, Beth Onners, Matthew J Weiss, Christopher L Wolfgang, John L Cameron, Timothy M Pawlik, Lauren Rosati, Dung T Le, Amy Hacker-Prietz, Eric R Lutz, Richard Schulick, Amol K Narang, Daniel A Laheru, Joseph M Herman.
       PURPOSE: Local and distant progression remain common following resection of resectable pancreatic ductal adenocarcinoma (PDAC) despite adjuvant multiagent chemotherapy. We report a prospective institutional phase I trial incorporating adjuvant GVAX vaccine, low-dose cyclophosphamide (Cy) and SBRT followed by FOLFIRINOX (FFX) among patients who underwent resection of high-risk PDAC.
    PATIENTS AND METHODS: The study design was a modified 3+3. Cohort 1 received 5-fraction SBRT to 33 Gy to the tumor bed and 25 Gy to elective nodes followed by 6 cycles of full dose FFX. After toxicity review, cohort 2 had SBRT and were switched to modified FFX (mFFX). Cohort 3 had 1 cycle of Cy/GVAX followed by SBRT, mFFX, and 4 cycles of maintenance Cy/GVAX with 6-month Cy/GVAX boosts until progression.
    RESULTS: 19 patients were enrolled with a median follow-up of 36.2 months. To be eligible, patients were required to have close/positive margins (within ≤1 mm) (71%) and/or lymph node metastasis (79%). Overall, 63% of patients had both. In cohort 1, 67% of patients received 6 cycles of FFX; in cohort 2, 75% received 6 cycles of modified FFX. In cohort 3, 12 patients received the first dose of Cy/GVAX and SBRT with 10 individuals (83%) receiving 6 cycles of mFFX. Cohort 3 had acceptable levels of grade ≥3 thrombocytopenia, neutropenia, and diarrhea after two cycles of mFFX. Median OS/DFS for the overall cohort and cohort 3 was 36.2/18.2 months and 61.3/24.1 months, respectively. 1-year and 2-year OS for cohort 3 was 83%/75%, respectively. At last follow-up (median= x), 5 patients were alive (42%) in cohort 3.
    CONCLUSION: This is the first prospective trial to evaluate adjuvant GVAX, Cy, SBRT, and mFFX in resected PDAC patients with high-risk features. This combination regimen was well tolerated with limited toxicity and promising survival outcomes, warranting future studies to validate this regimen in the adjuvant setting.
    Keywords:  Adjuvant; FOLFINRINOX; Radiation; SBRT; Vaccine
    DOI:  https://doi.org/10.1016/j.ijrobp.2024.10.039
  47. Nat Aging. 2024 Nov 11.
    Chronic social stress induces p16-mediated senescent cell accumulation in mice.
    Carey E Lyons, Jean Pierre Pallais, Seth McGonigle, Rachel P Mansk, Charles W Collinge, Matthew J Yousefzadeh, Darren J Baker, Patricia R Schrank, Jesse W Williams, Laura J Niedernhofer, Jan M van Deursen, Maria Razzoli, Alessandro Bartolomucci.
      Life stress can shorten lifespan and increase risk for aging-related diseases, but the biology underlying this phenomenon remains unclear. Here we assessed the effect of chronic stress on cellular senescence-a hallmark of aging. Exposure to restraint stress, a psychological non-social stress model, increased p21Cip1 exclusively in the brains of male, but not female mice, and in a p16Ink4a-independent manner. Conversely, exposure to chronic subordination stress (only males were tested) increased key senescent cell markers in peripheral blood mononuclear cells, adipose tissue and brain, in a p16Ink4a-dependent manner. p16Ink4a-positive cells in the brain of chronic subordination stress-exposed mice were primarily hippocampal and cortical neurons with evidence of DNA damage that could be reduced by p16Ink4a cell clearance. Clearance of p16Ink4a-positive cells was not sufficient to ameliorate the adverse effects of social stress on measured metrics of healthspan. Overall, our findings indicate that social stress induces an organ-specific and p16Ink4a-dependent accumulation of senescent cells, illuminating a fundamental way by which the social environment can contribute to aging.
    DOI:  https://doi.org/10.1038/s43587-024-00743-8
  48. Nat Genet. 2024 Nov 11.
    Brca1 haploinsufficiency promotes early tumor onset and epigenetic alterations in a mouse model of hereditary breast cancer.
    Carman Man-Chung Li, Alyssa Cordes, Michael U J Oliphant, S Aidan Quinn, Mayura Thomas, Laura M Selfors, Francesca Silvestri, Nomeda Girnius, Gianmarco Rinaldi, Jason J Zoeller, Hana Shapiro, Christina Tsiobikas, Kushali P Gupta, Shailja Pathania, Aviv Regev, S Cigall Kadoch, Senthil K Muthuswamy, Joan S Brugge.
      Germline BRCA1 mutation carriers face a high breast cancer risk; however, the underlying mechanisms for this risk are not completely understood. Using a new genetically engineered mouse model of germline Brca1 heterozygosity, we demonstrate that early tumor onset in a Brca1 heterozygous background cannot be fully explained by the conventional 'two-hit' hypothesis, suggesting the existence of inherent tumor-promoting alterations in the Brca1 heterozygous state. Single-cell RNA sequencing and assay for transposase-accessible chromatin with sequencing analyses uncover a unique set of differentially accessible chromatin regions in ostensibly normal Brca1 heterozygous mammary epithelial cells, distinct from wild-type cells and partially mimicking the chromatin and RNA-level changes in tumor cells. Transcription factor analyses identify loss of ELF5 and gain of AP-1 sites in these epigenetically primed regions; in vivo experiments further implicate AP-1 and Wnt10a as strong promoters of Brca1-related breast cancer. These findings reveal a previously unappreciated epigenetic effect of Brca1 haploinsufficiency in accelerating tumorigenesis, advancing our mechanistic understanding and informing potential therapeutic strategies.
    DOI:  https://doi.org/10.1038/s41588-024-01958-6
  49. PLoS One. 2024 ;19(11): e0308873
    EasyFlow: An open-source, user-friendly cytometry analyzer with graphic user interface (GUI).
    Yitong Ma, Inbal Eizenberg-Magar, Yaron Antebi.
      Flow cytometry enables quantitative measurements of fluorescence in single cells. The technique was widely used for immunology to identify populations with different surface protein markers. More recently, the usage of flow cytometry has been extended to additional readouts, including intracellular proteins and fluorescent protein transgenes, and is widely utilized to study developmental biology, systems biology, microbiology, and many other fields. A common file format (FCS format, defined by the International Society for Advancement of Cytometry (ISAC)) has been universally adopted, facilitating data exchange between different machines. A diverse spectrum of software packages has been developed for the analysis of flow cytometry data. However, those are either 1) costly proprietary softwares, 2) open source packages with prerequisite installation of R or Python and sometimes require users to have experience in coding, or 3) online tools that are limiting for analysis of large data sets. Here, we present EasyFlow, an open-source flow cytometry analysis graphic user interface (GUI) based on Matlab or Python, that can be installed and run locally across platforms (Windows, MacOS, and Linux) without requiring previous coding knowledge. The Python version (EasyFlowQ) is also developed on a popular plotting framework (Matplotlib) and modern user interface toolkit (Qt), allowing more advanced users to customize and keep contributing to the software, as well as its tutorials. Overall, EasyFlow serves as a simple-to-use tool for inexperienced users with little coding experience to use locally, as well as a platform for advanced users to further customize for their own needs.
    DOI:  https://doi.org/10.1371/journal.pone.0308873
  50. Br J Cancer. 2024 Nov 09.
    VEGF-C propagates 'onward' colorectal cancer metastasis from liver to lung.
    Susanna Poghosyan, Nicola Frenkel, Lotte van den Bent, Danielle Raats, Tessa Spaapen, Jamila Laoukili, Inne Borel Rinkes, Onno Kranenburg, Jeroen Hagendoorn.
       BACKGROUND: The formation of lung metastasis as part of the progression of colon cancer is a poorly understood process. Theoretically, liver metastases could seed lung metastases.
    METHODS: To assess the contribution of the liver lymphatic vasculature to metastatic spread to the lungs, we generated murine liver-metastasis-derived organoids overexpressing vascular endothelial growth factor (VEGF)-C. The organoids were reimplanted into the mouse liver for tumour generation and onward metastasis.
    RESULTS: Liver metastases from patients with concomitant lung metastases showed higher expression of VEGF-C, lymphatic vessel hyperplasia, and tumour cell invasion into lymphatic vessels when compared to those without lung metastases. Reimplantation of VEGF-C overexpressing organoids into the mouse liver showed that VEGF-C caused peritumoral lymphatic vessel hyperplasia, lymphatic tumour cell invasion, and lung metastasis formation. This change in metastatic organotropism was accompanied by reduced expression of WNT-driven adult stem cell markers, and increased expression of fetal stem cell markers and NOTCH pathway genes. Further NOTCH pathway inhibition with γ-secretase inhibitor (DAPT) in vivo results in a slight reduction in lung metastases and a decrease in lymphatic hyperplasia and invasion in VEGF-C-overexpressing tumours.
    CONCLUSION: Collectively, these data indicate that VEGF-C can drive onward metastasis from the liver to the lung and suggest that targeting VEGF-C/NOTCH pathways may impair the progression of colorectal cancer.
    DOI:  https://doi.org/10.1038/s41416-024-02892-4
  51. Cell. 2024 Nov 11. pii: S0092-8674(24)01159-0. [Epub ahead of print]
    Spatiotemporal modeling of molecular holograms.
    Xiaojie Qiu, Daniel Y Zhu, Yifan Lu, Jiajun Yao, Zehua Jing, Kyung Hoi Min, Mengnan Cheng, Hailin Pan, Lulu Zuo, Samuel King, Qi Fang, Huiwen Zheng, Mingyue Wang, Shuai Wang, Qingquan Zhang, Sichao Yu, Sha Liao, Chao Liu, Xinchao Wu, Yiwei Lai, Shijie Hao, Zhewei Zhang, Liang Wu, Yong Zhang, Mei Li, Zhencheng Tu, Jinpei Lin, Zhuoxuan Yang, Yuxiang Li, Ying Gu, David Ellison, Ao Chen, Longqi Liu, Jonathan S Weissman, Jiayi Ma, Xun Xu, Shiping Liu, Yinqi Bai.
      Quantifying spatiotemporal dynamics during embryogenesis is crucial for understanding congenital diseases. We developed Spateo (https://github.com/aristoteo/spateo-release), a 3D spatiotemporal modeling framework, and applied it to a 3D mouse embryogenesis atlas at E9.5 and E11.5, capturing eight million cells. Spateo enables scalable, partial, non-rigid alignment, multi-slice refinement, and mesh correction to create molecular holograms of whole embryos. It introduces digitization methods to uncover multi-level biology from subcellular to whole organ, identifying expression gradients along orthogonal axes of emergent 3D structures, e.g., secondary organizers such as midbrain-hindbrain boundary (MHB). Spateo further jointly models intercellular and intracellular interaction to dissect signaling landscapes in 3D structures, including the zona limitans intrathalamica (ZLI). Lastly, Spateo introduces "morphometric vector fields" of cell migration and integrates spatial differential geometry to unveil molecular programs underlying asymmetrical murine heart organogenesis and others, bridging macroscopic changes with molecular dynamics. Thus, Spateo enables the study of organ ecology at a molecular level in 3D space over time.
    Keywords:  3D reconstruction; Spateo; Stereo-seq; intercellular and intracellular interactions; ligand-receptor interactions, cell-cell interactions; morphometric vector field; spatial differential geometry analyses; spatial domain digitization; whole-embryo 3D spatial transcriptomics
    DOI:  https://doi.org/10.1016/j.cell.2024.10.011
  52. J Clin Invest. 2024 Nov 07. pii: e184785. [Epub ahead of print]
    Prospective observational study and mechanistic evidence showing lipolysis of circulating triglycerides worsens hypertriglyceridemic acute pancreatitis.
    Prasad Rajalingamgari, Biswajit Khatua, Megan J Summers, Sergiy Kostenko, Yu-Hui H Chang, Mohamed Elmallahy, Arti Anand, Anoop Narayana Pillai, Mahmoud Morsy, Shubham Trivedi, Bryce McFayden, Sarah Jahangir, Christine Lh Snozek, Vijay P Singh.
       BACKGROUND: While most hypertriglyceridemia is asymptomatic, hypertriglyceridemia-associated acute pancreatitis (HTG-AP) can be more severe than other AP etiologies. The reasons underlying this are unclear. We thus studied whether lipolytic generation of non-esterified fatty acids (NEFA) from circulating triglycerides (TGs) could worsen clinical outcomes.
    METHODS: Admission serum TGs, NEFA compositions and concentrations were analyzed prospectively in 269 patients with AP. These and demographics, clinical outcomes were compared between HTGAP (TGs >500mg/dL; American Heart Association 2018 guidelines) and other AP etiologies. Serum NEFAs were correlated with the serum triglyceride fatty acids (TGFAs) alone, and with the product of TGFA x serum lipase (NEFA-TGFA x lipase). Studies in mice, rats were done to understand the role of HTG lipolysis in organ failure and to interpret the NEFA-TGFA correlations.
    RESULTS: HTG-AP patients had higher serum NEFAs and TGs and more severe AP (19% vs. 7% p<0.03) than other etiologies. Correlations of long-chain unsaturated NEFA with corresponding TGFAs increased with TG concentrations up to 500mg/dL and declined thereafter. However, NEFA-TGFA x lipase correlations got stronger with TGs >500mg/dL. AP, and intravenous lipase infusion in rodents caused lipolysis of circulating TGs to NEFA. This led to multi-system organ failure, which was prevented by pancreatic triglyceride lipase deletion, or lipase inhibition.
    CONCLUSIONS: HTG-AP is made severe by the NEFAs generated form intravascular lipolysis of circulating TGs. Strategies that prevent TG lipolysis may be effective in improving clinical outcomes of HTG-AP.
    TRIAL REGISTRATION: Not applicable.
    FUNDING: This project was supported by Grant numbers RO1DK092460, R01DK119646 from the NIDDK, PR191945 under W81XWH-20-1-0400 from the DOD (VPS), and R01AA031257 from the NIAAA (VPS).
    Keywords:  Gastroenterology; Lipoproteins
    DOI:  https://doi.org/10.1172/JCI184785
  53. Nat Methods. 2024 Nov 11.
    EvoAI enables extreme compression and reconstruction of the protein sequence space.
    Ziyuan Ma, Wenjie Li, Yunhao Shen, Yunxin Xu, Gengjiang Liu, Jiamin Chang, Zeju Li, Hong Qin, Boxue Tian, Haipeng Gong, David R Liu, B W Thuronyi, Christopher A Voigt, Shuyi Zhang.
      Designing proteins with improved functions requires a deep understanding of how sequence and function are related, a vast space that is hard to explore. The ability to efficiently compress this space by identifying functionally important features is extremely valuable. Here we establish a method called EvoScan to comprehensively segment and scan the high-fitness sequence space to obtain anchor points that capture its essential features, especially in high dimensions. Our approach is compatible with any biomolecular function that can be coupled to a transcriptional output. We then develop deep learning and large language models to accurately reconstruct the space from these anchors, allowing computational prediction of novel, highly fit sequences without prior homology-derived or structural information. We apply this hybrid experimental-computational method, which we call EvoAI, to a repressor protein and find that only 82 anchors are sufficient to compress the high-fitness sequence space with a compression ratio of 1048. The extreme compressibility of the space informs both applied biomolecular design and understanding of natural evolution.
    DOI:  https://doi.org/10.1038/s41592-024-02504-2
  54. Methods Mol Biol. 2025 ;2878 211-221
    Understanding Mitochondrial and Lysosomal Dynamics by Fluorescent Microscopy.
    Pedro A Dionísio, Vilma A Sardão, Nuno Raimundo.
      Live cell imaging is a robust method to visualize dynamic cellular structures, especially organelles with network-like structures such as mitochondria. In this regard, mitochondrial dynamics, namely mitochondrial fission and fusion, are highly dynamic processes that regulate mitochondrial size and morphology depending on a plethora of cellular cues. Likewise, lysosome size and distribution may hint at their function and state.Here, we describe how to perform live cell confocal imaging using commercially available organelle dyes (MitoTracker, LysoTracker), followed by either 2D or 3D analyses of mitochondrial morphology/network connectivity and lysosomal morphology using the freely available Mitochondria Analyzer plugin for ImageJ/Fiji.
    Keywords:  Cell imaging; Fluorescent probes; LysoTracker; Lysosomes; Microscopy; MitoTracker; Mitochondria; Mitochondrial dynamics
    DOI:  https://doi.org/10.1007/978-1-0716-4264-1_11
  55. Cell. 2024 Nov 14. pii: S0092-8674(24)01211-X. [Epub ahead of print]187(23): 6421-6423
    The cellular dogma.
    Stephen R Quake.
      In this essay, I will put forth what I see as a major conceptual challenge for biology in the next decade, one that is inspired by Crick's Central Dogma: understanding information flow in the cell in the most general sense.
    DOI:  https://doi.org/10.1016/j.cell.2024.10.029
  56. Anal Chem. 2024 Nov 08.
    Biophysical and Morphological Cell Features Retrieved by Digital Holographic Microscopy Correlate with Drug-Induced Changes in Cell Migration Behavior.
    Xiaoxi Ren, Feiyang Zhang, Ge Gao, Dan Gao, Ping Su.
      Quantitative analysis of cancer cell migration is critical for developing effective therapies to curb cancer metastasis. However, traditional methods are time-consuming and labor-intensive and lack quantitative capabilities. Cell volume change, a key physiological indicator of cell migration, is directly linked to phase change. In this work, we have developed a model that connects phase features from digital holographic microscopy (DHM) with cell healrate values from the wound healing assay. This approach aims to provide a rapid and quantitative evaluation of the breast cancer cell migration capability. Using DHM, six phase features of 231 cells treated with varying drug concentrations were extracted. It was observed that the rate of change of these phase features, termed characteristic parameters, showed a high linear correlation with cell healrate values from wound healing assays. Based on these linear correlations, a composite coefficient was derived by linearly combining the characteristic parameters of the six phase features. This composite coefficient was then linearly correlated with the cell healrate values to create a correlation model. This model establishes a strong connection between DHM-extracted morphological/biophysical features and cell migration metrics from a complementary assay. It provides a new, rapid, and quantitative method for assessing cancer cell migration in vitro and delivering valuable insights for cancer research.
    DOI:  https://doi.org/10.1021/acs.analchem.4c04872
  57. Adv Mater. 2024 Nov 14. e2408307
    Charge-Stabilized Nanodiscs as a New Class of Lipid Nanoparticles.
    Ivan S Pires, Alexander Hostetler, Gil Covarrubias, Isabella S Carlo, Jack R Suggs, B J Kim, Andrew J Pickering, Ezra Gordon, Darrell J Irvine, Paula T Hammond.
      Nanoparticles have the potential to improve disease treatment and diagnosis due to their ability to incorporate drugs, alter pharmacokinetics, and enable tissue targeting. While considerable effort is placed on developing spherical lipid-based nanocarriers, recent evidence suggests that high aspect ratio lipid nanocarriers can exhibit enhanced disease site targeting and altered cellular interactions. However, the assembly of lipid-based nanoparticles into non-spherical morphologies has typically required incorporating additional agents such as synthetic polymers, proteins, lipid-polymer conjugates, or detergents. Here, charged lipid headgroups are used to generate stable discoidal lipid nanoparticles from mixed micelles, which are termed charge-stabilized nanodiscs (CNDs). The ability to generate CNDs in buffers with physiological ionic strength is restricted to lipids with more than one anionic group, whereas monovalent lipids only generate small nanoliposomal assemblies. In mice, the smaller size and anisotropic shape of CNDs promote higher accumulation in subcutaneous tumors than spherical liposomes. Further, the surface chemistry of CNDs can be modified via layer-by-layer (LbL) assembly to improve their tumor-targeting properties over state-of-the-art LbL-liposomes when tested using a metastatic model of ovarian cancer. The application of charge-mediated anisotropy in lipid-based assemblies can aid in the future design of biomaterials and cell-membrane mimetic structures.
    Keywords:  anisotropy; layer‐by‐layer; lipid nanoparticles; nanodiscs; self‐assembly; tumor targeting
    DOI:  https://doi.org/10.1002/adma.202408307
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