bims-cagime 2024-08-18 papers

bims-cagime

Biomed News

on Cancer, aging and metabolism

Issue of 2024–08–18
38 papers selected by
Kıvanç Görgülü, Technical University of Munich

Guidelines for minimal information on cellular senescence experimentation in vivo.
Mitochondrial complex I promotes kidney cancer metastasis.
Stromal KITL/SCF promotes pancreas tissue homeostasis and restrains tumor progression.
3D histology reveals that immune response to pancreatic precancers is heterogeneous and depends on global pancreas structure.
Capturing RAS oligomerization on a membrane.
Senescent fibroblasts in the tumor stroma rewire lung cancer metabolism and plasticity.
Targeting BCL2 with Venetoclax Enhances the Efficacy of the KRASG12D Inhibitor MRTX1133 in Pancreatic Cancer.
Conjugation of ATG8s to single membranes at a glance.
Metabolomics and cellular altered pathways in cancer biology: A review.
Protocol for monitoring fatty acid trafficking from lipid droplets to mitochondria in cultured cells.
Autophagy in Skeletal Muscle.
Mouse sarcopenia model reveals sex- and age-specific differences in phenotypic and molecular characteristics.
Circadian period is compensated for repressor protein turnover rates in single cells.
Enhanced cytokine signaling and ferroptosis defense interplay initiates obesity-associated pancreatic ductal adenocarcinoma.
A high-throughput microfabricated platform for rapid quantification of metastatic potential.
Remodelling of the translatome controls diet and its impact on tumorigenesis.
Dynamical modelling of lipid droplet formation suggests a key function of membrane phospholipids.
Transformers in single-cell omics: a review and new perspectives.
Prevalence of and survival with cachexia among patients with cancer: A systematic review and meta-analysis.
Macrophage-mediated myelin recycling fuels brain cancer malignancy.
Mitochondrial Ca 2+ controls pancreatic cancer growth and metastasis by regulating epithelial cell plasticity.
Aging human abdominal subcutaneous white adipose tissue at single cell resolution.
Rewiring cancer cell death: LPCAT1 shapes lipid composition and ferroptosis resistance.
Recycling the recyclers: lysophagy emerges as a new pharmacological target for retinal degeneration.
3Din vitromodeling of the exocrine pancreatic unit using tomographic volumetric bioprinting.
Clinical and molecular features of early onset pancreatic adenocarcinoma.
An Experimental Protocol for the Boyden Chamber Invasion Assay With Absorbance Readout.
Unraveling cysteine deficiency-associated rapid weight loss.
SOS1 inhibitor combinations overcome KRAS inhibitor resistance.
Proximity sequencing for the detection of mRNA, extracellular proteins and extracellular protein complexes in single cells.
Methods and Protocol for Single-Cell Motility Assays under Topological Constraints.
Ceas: An R package for Seahorse data analysis and visualization.
Analytical evaluation of the automated interleukin-6 assay on the Roche cobas e602 analyzer.
Cysteine depletion triggers adipose tissue thermogenesis and weight-loss.
Multiomics2Targets identifies targets from cancer cohorts profiled with transcriptomics, proteomics, and phosphoproteomics.
Identification of senescent cell subpopulations by CITE-seq analysis.
Profiling Dynamic Patterns of Single-Cell Motility.
Tracking live-cell single-molecule dynamics enables measurements of heterochromatin-associated protein-protein interactions.

Cell. 2024 Aug 08. pii: S0092-8674(24)00640-8. [Epub ahead of print]187(16): 4150-4175

Guidelines for minimal information on cellular senescence experimentation in vivo.

Mikolaj Ogrodnik, Juan Carlos Acosta, Peter D Adams, Fabrizio d'Adda di Fagagna, Darren J Baker, Cleo L Bishop, Tamir Chandra, Manuel Collado, Jesus Gil, Vassilis Gorgoulis, Florian Gruber, Eiji Hara, Pidder Jansen-Dürr, Diana Jurk, Sundeep Khosla, James L Kirkland, Valery Krizhanovsky, Tohru Minamino, Laura J Niedernhofer, João F Passos, Nadja A R Ring, Heinz Redl, Paul D Robbins, Francis Rodier, Karin Scharffetter-Kochanek, John M Sedivy, Ewa Sikora, Kenneth Witwer, Thomas von Zglinicki, Maximina H Yun, Johannes Grillari, Marco Demaria.

  Cellular senescence is a cell fate triggered in response to stress and is characterized by stable cell-cycle arrest and a hypersecretory state. It has diverse biological roles, ranging from tissue repair to chronic disease. The development of new tools to study senescence in vivo has paved the way for uncovering its physiological and pathological roles and testing senescent cells as a therapeutic target. However, the lack of specific and broadly applicable markers makes it difficult to identify and characterize senescent cells in tissues and living organisms. To address this, we provide practical guidelines called "minimum information for cellular senescence experimentation in vivo" (MICSE). It presents an overview of senescence markers in rodent tissues, transgenic models, non-mammalian systems, human tissues, and tumors and their use in the identification and specification of senescent cells. These guidelines provide a uniform, state-of-the-art, and accessible toolset to improve our understanding of cellular senescence in vivo.

Keywords:  aging; humans; in vivo; mouse; senescence; senotherapy

DOI:  https://doi.org/10.1016/j.cell.2024.05.059
Nature. 2024 Aug 14.

Mitochondrial complex I promotes kidney cancer metastasis.

Divya Bezwada, Luigi Perelli, Nicholas P Lesner, Ling Cai, Bailey Brooks, Zheng Wu, Hieu S Vu, Varun Sondhi, Daniel L Cassidy, Stacy Kasitinon, Sherwin Kelekar, Feng Cai, Arin B Aurora, McKenzie Patrick, Ashley Leach, Rashed Ghandour, Yuanyuan Zhang, Duyen Do, Phyllis McDaniel, Jessica Sudderth, Dennis Dumesnil, Sara House, Tracy Rosales, Alan M Poole, Yair Lotan, Solomon Woldu, Aditya Bagrodia, Xiaosong Meng, Jeffrey A Cadeddu, Prashant Mishra, Javier Garcia-Bermudez, Ivan Pedrosa, Payal Kapur, Kevin D Courtney, Craig R Malloy, Giannicola Genovese, Vitaly Margulis, Ralph J DeBerardinis.

Most kidney cancers are metabolically dysfunctional1-4, but how this dysfunction affects cancer progression in humans is unknown. We infused 13C-labelled nutrients in over 80 patients with kidney cancer during surgical tumour resection. Labelling from [U-13C]glucose varies across subtypes, indicating that the kidney environment alone cannot account for all tumour metabolic reprogramming. Compared with the adjacent kidney, clear cell renal cell carcinomas (ccRCCs) display suppressed labelling of tricarboxylic acid (TCA) cycle intermediates in vivo and in ex vivo organotypic cultures, indicating that suppressed labelling is tissue intrinsic. [1,2-13C]acetate and [U-13C]glutamine infusions in patients, coupled with measurements of respiration in isolated human kidney and tumour mitochondria, reveal lower electron transport chain activity in ccRCCs that contributes to decreased oxidative and enhanced reductive TCA cycle labelling. However, ccRCC metastases unexpectedly have enhanced TCA cycle labelling compared with that of primary ccRCCs, indicating a divergent metabolic program during metastasis in patients. In mice, stimulating respiration or NADH recycling in kidney cancer cells is sufficient to promote metastasis, whereas inhibiting electron transport chain complex I decreases metastasis. These findings in humans and mice indicate that metabolic properties and liabilities evolve during kidney cancer progression, and that mitochondrial function is limiting for metastasis but not growth at the original site.

DOI: https://doi.org/10.1038/s41586-024-07812-3
bioRxiv. 2024 Jul 30. pii: 2024.07.29.605485. [Epub ahead of print]

Stromal KITL/SCF promotes pancreas tissue homeostasis and restrains tumor progression.

M Kathrina Oñate, Chet Oon, Sohinee Bhattacharyya, Vivien Low, Canping Chen, Xiaofan Zhao, Ziqiao Yan, Yan Hang, Seung K Kim, Zheng Xia, Mara H Sherman.

  Components of normal tissue architecture serve as barriers to tumor progression. Inflammatory and wound-healing programs are requisite features of solid tumorigenesis, wherein alterations to immune and non-immune stromal elements enable loss of homeostasis during tumor evolution. The precise mechanisms by which normal stromal cell states limit tissue plasticity and tumorigenesis, and which are lost during tumor progression, remain largely unknown. Here we show that healthy pancreatic mesenchyme expresses the paracrine signaling molecule KITL, also known as stem cell factor, and identify loss of stromal KITL during tumorigenesis as tumor-promoting. Genetic inhibition of mesenchymal KITL in the contexts of homeostasis, injury, and cancer together indicate a role for KITL signaling in maintenance of pancreas tissue architecture, such that loss of the stromal KITL pool increased tumor growth and reduced survival of tumor-bearing mice. Together, these findings implicate loss of mesenchymal KITL as a mechanism for establishing a tumor-permissive microenvironment.

Keywords:  mesenchymal cells; pancreatic cancer; paracrine signaling; tissue plasticity

DOI:  https://doi.org/10.1101/2024.07.29.605485
bioRxiv. 2024 Aug 06. pii: 2024.08.03.606493. [Epub ahead of print]

3D histology reveals that immune response to pancreatic precancers is heterogeneous and depends on global pancreas structure.

Ashley L Kiemen, Cristina Almagro-Pérez, Valentina Matos, Andre Forjaz, Alicia M Braxton, Lucie Dequiedt, Jeeun Parksong, Courtney D Cannon, Xuan Yuan, Sarah M Shin, Jaanvi Mahesh Babu, Elizabeth D Thompson, Toby C Cornish, Won Jin Ho, Laura D Wood, Pei-Hsun Wu, Arrate Muñoz Barrutia, Ralph H Hruban, Denis Wirtz.

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal cancer for which few effective therapies exist. Immunotherapies specifically are ineffective in pancreatic cancer, in part due to its unique stromal and immune microenvironment. Pancreatic intraepithelial neoplasia, or PanIN, is the main precursor lesion to PDAC. Recently it was discovered that PanINs are remarkably abundant in the grossly normal pancreas, suggesting that the vast majority will never progress to cancer. Here, through construction of 48 samples of cm 3 -sized human pancreas tissue, we profiled the immune microenvironment of 1,476 PanINs in 3D and at single-cell resolution to better understand the early evolution of the pancreatic tumor microenvironment and to determine how inflammation may play a role in cancer progression. We found that bulk pancreatic inflammation strongly correlates to PanIN cell fraction. We found that the immune response around PanINs is highly heterogeneous, with distinct immune hotspots and cold spots that appear and disappear in a span of tens of microns. Immune hotspots generally mark locations of higher grade of dysplasia or locations near acinar atrophy. The immune composition at these hotspots is dominated by naïve, cytotoxic, and regulatory T cells, cancer associated fibroblasts, and tumor associated macrophages, with little similarity to the immune composition around less-inflamed PanINs. By mapping FOXP3+ cells in 3D, we found that regulatory T cells are present at higher density in larger PanIN lesions compared to smaller PanINs, suggesting that the early initiation of PanINs may not exhibit an immunosuppressive response. This analysis demonstrates that while PanINs are common in the pancreases of most individuals, inflammation may play a pivotal role, both at the bulk and the microscopic scale, in demarcating regions of significance in cancer progression.

DOI: https://doi.org/10.1101/2024.08.03.606493
Proc Natl Acad Sci U S A. 2024 Aug 20. 121(34): e2405986121

Capturing RAS oligomerization on a membrane.

Sangho D Yun, Elena Scott, Jing-Yuan Chang, Hanieh Bahramimoghaddam, Michael Lynn, Carter Lantz, David H Russell, Arthur Laganowsky.

  RAS GTPases associate with the biological membrane where they function as molecular switches to regulate cell growth. Recent studies indicate that RAS proteins oligomerize on membranes, and disrupting these assemblies represents an alternative therapeutic strategy. However, conflicting reports on RAS assemblies, ranging in size from dimers to nanoclusters, have brought to the fore key questions regarding the stoichiometry and parameters that influence oligomerization. Here, we probe three isoforms of RAS [Kirsten Rat Sarcoma viral oncogene (KRAS), Harvey Rat Sarcoma viral oncogene (HRAS), and Neuroblastoma oncogene (NRAS)] directly from membranes using mass spectrometry. We show that KRAS on membranes in the inactive state (GDP-bound) is monomeric but forms dimers in the active state (GTP-bound). We demonstrate that the small molecule BI2852 can induce dimerization of KRAS, whereas the binding of effector proteins disrupts dimerization. We also show that RAS dimerization is dependent on lipid composition and reveal that oligomerization of NRAS is regulated by palmitoylation. By monitoring the intrinsic GTPase activity of RAS, we capture the emergence of a dimer containing either mixed nucleotides or GDP on membranes. We find that the interaction of RAS with the catalytic domain of Son of Sevenless (SOScat) is influenced by membrane composition. We also capture the activation and monomer to dimer conversion of KRAS by SOScat. These results not only reveal the stoichiometry of RAS assemblies on membranes but also uncover the impact of critical factors on oligomerization, encompassing regulation by nucleotides, lipids, and palmitoylation.

Keywords:  RAS; cancer; mass spectrometry; peripheral membrane protein

DOI:  https://doi.org/10.1073/pnas.2405986121
bioRxiv. 2024 Jul 30. pii: 2024.07.29.605645. [Epub ahead of print]

Senescent fibroblasts in the tumor stroma rewire lung cancer metabolism and plasticity.

Jin Young Lee, Nabora Reyes, Sang-Ho Woo, Sakshi Goel, Fia Stratton, Chaoyuan Kuang, Aaron S Mansfield, Lindsay M LaFave, Tien Peng.

Senescence has been demonstrated to either inhibit or promote tumorigenesis. Resolving this paradox requires spatial mapping and functional characterization of senescent cells in the native tumor niche. Here, we identified senescent p16 Ink4a + cancer-associated fibroblasts with a secretory phenotype that promotes fatty acid uptake and utilization by aggressive lung adenocarcinoma driven by Kras and p53 mutations. Furthermore, rewiring of lung cancer metabolism by p16 Ink4a + cancer-associated fibroblasts also altered tumor cell identity to a highly plastic/dedifferentiated state associated with progression in murine and human LUAD. Our ex vivo senolytic screening platform identified XL888, a HSP90 inhibitor, that cleared p16 Ink4a + cancer-associated fibroblasts in vivo. XL888 administration after establishment of advanced lung adenocarcinoma significantly reduced tumor burden concurrent with the loss of plastic tumor cells. Our study identified a druggable component of the tumor stroma that fulfills the metabolic requirement of tumor cells to acquire a more aggressive phenotype.

DOI: https://doi.org/10.1101/2024.07.29.605645
Cancer Res. 2024 Aug 13.

Targeting BCL2 with Venetoclax Enhances the Efficacy of the KRASG12D Inhibitor MRTX1133 in Pancreatic Cancer.

Jeffrey H Becker, Anastasia E Metropulos, Christina Spaulding, Alejandra M Marinelarena, Mario A Shields, Daniel R Principe, Thao D Pham, Hidayatullah G Munshi.

MRTX1133 is currently being evaluated in patients with pancreatic ductal adenocarcinoma (PDAC) tumors harboring a KRASG12D mutation. Combination strategies have the potential to enhance the efficacy of MRTX1133 to further promote cell death and tumor regression. In this study, we demonstrated that MRTX1133 increased the levels of the pro-apoptotic protein BIM in PDAC cells and conferred sensitivity to the FDA-approved BCL2 inhibitor venetoclax. Combined treatment with MRTX1133 and venetoclax resulted in cell death and growth suppression in 3D cultures. BIM was required for apoptosis induced by the combination treatment. Consistently, BIM was induced in tumors treated with MRTX1133, and venetoclax enhanced the efficacy of MRTX1133 in vivo. Venetoclax could also re-sensitize MRTX1133-resistant PDAC cells to MRTX1133 in 3D cultures, and tumors established from resistant cells responded to the combination of MRTX1133 and venetoclax. These results provide a rationale for the clinical testing of MRTX1133 and venetoclax in PDAC patients.

DOI: https://doi.org/10.1158/0008-5472.CAN-23-3574
J Cell Sci. 2024 Aug 01. pii: jcs261031. [Epub ahead of print]137(15):

Conjugation of ATG8s to single membranes at a glance.

Carmen Figueras-Novoa, Lewis Timimi, Elena Marcassa, Rachel Ulferts, Rupert Beale.

  Autophagy refers to a set of degradative mechanisms whereby cytoplasmic contents are targeted to the lysosome. This is best described for macroautophagy, where a double-membrane compartment (autophagosome) is generated to engulf cytoplasmic contents. Autophagosomes are decorated with ubiquitin-like ATG8 molecules (ATG8s), which are recruited through covalent lipidation, catalysed by the E3-ligase-like ATG16L1 complex. LC3 proteins are ATG8 family members that are often used as a marker for autophagosomes. In contrast to canonical macroautophagy, conjugation of ATG8s to single membranes (CASM) describes a group of non-canonical autophagy processes in which ATG8s are targeted to pre-existing single-membrane compartments. CASM occurs in response to disrupted intracellular pH gradients, when the V-ATPase proton pump recruits ATG16L1 in a process called V-ATPase-ATG16L1-induced LC3 lipidation (VAIL). Recent work has demonstrated a parallel, alternative axis for CASM induction, triggered when the membrane recruitment factor TECPR1 recognises sphingomyelin exposed on the cytosolic face of a membrane and forms an alternative E3-ligase-like complex. This sphingomyelin-TECPR1-induced LC3 lipidation (STIL) is independent of the V-ATPase and ATG16L1. In light of these discoveries, this Cell Science at a Glance article summarises these two mechanisms of CASM to highlight how they differ from canonical macroautophagy, and from each other.

Keywords:  ATG8ylation; Autophagy; CASM; STIL; TECPR1; V-ATPase; VAIL

DOI:  https://doi.org/10.1242/jcs.261031
J Biochem Mol Toxicol. 2024 Sep;38(9): e23807

Metabolomics and cellular altered pathways in cancer biology: A review.

Jitender Kumar Bhardwaj, Anshu Siwach, Som Nath Sachdeva.

  Cancer is a deadly disease that affects a cell's metabolism and surrounding tissues. Understanding the fundamental mechanisms of metabolic alterations in cancer cells would assist in developing cancer treatment targets and approaches. From this perspective, metabolomics is a great analytical tool to clarify the mechanisms of cancer therapy as well as a useful tool to investigate cancer from a distinct viewpoint. It is a powerful emerging technology that detects up to thousands of molecules in tissues and biofluids. Like other "-omics" technologies, metabolomics involves the comprehensive investigation of micromolecule metabolites and can reveal important details about the cancer state that is otherwise not apparent. Recent developments in metabolomics technologies have made it possible to investigate cancer metabolism in greater depth and comprehend how cancer cells utilize metabolic pathways to make the amino acids, nucleotides, and lipids required for tumorigenesis. These new technologies have made it possible to learn more about cancer metabolism. Here, we review the cellular and systemic effects of cancer and cancer treatments on metabolism. The current study provides an overview of metabolomics, emphasizing the current technologies and their use in clinical and translational research settings.

Keywords:  bioinformatics; biomarker; cancer; metabolic reprogramming; metabolomics

DOI:  https://doi.org/10.1002/jbt.23807
STAR Protoc. 2024 Aug 13. pii: S2666-1667(24)00401-5. [Epub ahead of print]5(3): 103236

Protocol for monitoring fatty acid trafficking from lipid droplets to mitochondria in cultured cells.

Gregory E Miner, Sarah Cohen.

  Intracellular trafficking of fatty acids (FAs) between organelles is critical for cells to adjust their metabolism in response to stimuli such as exercise, fasting, and cold exposure. Here, we describe a protocol to monitor trafficking of FAs from lipid droplets to mitochondria. We describe the labeling of organelles in cultured C2C12 myoblasts with transfection and dyes. We detail a pulse-chase labeling paradigm using a fluorescent FA analog, live-cell imaging to visualize trafficking of FAs, and steps to quantify FA trafficking. For complete details on the use and execution of this protocol, please refer to Miner et al.1.

Keywords:  Cell Biology; Metabolism; Microscopy

DOI:  https://doi.org/10.1016/j.xpro.2024.103236
Cold Spring Harb Perspect Biol. 2024 Aug 12. pii: a041565. [Epub ahead of print]

Autophagy in Skeletal Muscle.

Anais Franco-Romero, Marco Sandri, Stefano Schiaffino.

Skeletal muscle fibers possess, like all cells of our body, an evolutionary conserved autophagy machinery, which allows them to segregate unfolded proteins and damaged organelles within autophagosomes, and to induce fusion of autophagosomes with lysosomes, leading to degradation of those altered cell constituents. This process may be selective for specific cell components, as in the case of glycogen (glycophagy) or organelles, as with mitochondria (mitophagy). The autophagic flux is activated by fasting, and contributes with the proteasome to provide the organism with amino acids required for survival. Autophagy is also essential for the normal turnover of muscle proteins and organelles, as shown by the degenerative changes induced by genetic block of the autophagic mechanism, and in several myopathies. Autophagy is enhanced in muscle by exercise and impaired during aging, suggesting that aging-dependent muscle dysfunction could be delayed by boosting autophagy.

DOI: https://doi.org/10.1101/cshperspect.a041565
J Clin Invest. 2024 Jun 11. pii: e172890. [Epub ahead of print]134(16):

Mouse sarcopenia model reveals sex- and age-specific differences in phenotypic and molecular characteristics.

Haiming L Kerr, Kora Krumm, Barbara Anderson, Anthony Christiani, Lena Strait, Theresa Li, Brynn Irwin, Siyi Jiang, Artur Rybachok, Amanda Chen, Elizabeth Dacek, Lucas Caeiro, Gennifer E Merrihew, James W MacDonald, Theo K Bammler, Michael J MacCoss, Jose M Garcia.

  Our study was to characterize sarcopenia in C57BL/6J mice using a clinically relevant definition to investigate the underlying molecular mechanisms. Aged male (23-32 months old) and female (27-28 months old) C57BL/6J mice were classified as non-, probable-, or sarcopenic based on assessments of grip strength, muscle mass, and treadmill running time, using 2 SDs below the mean of their young counterparts as cutoff points. A 9%-22% prevalence of sarcopenia was identified in 23-26 month-old male mice, with more severe age-related declines in muscle function than mass. Females aged 27-28 months showed fewer sarcopenic but more probable cases compared with the males. As sarcopenia progressed, a decrease in muscle contractility and a trend toward lower type IIB fiber size were observed in males. Mitochondrial biogenesis, oxidative capacity, and AMPK-autophagy signaling decreased as sarcopenia progressed in males, with pathways linked to mitochondrial metabolism positively correlated with muscle mass. No age- or sarcopenia-related changes were observed in mitochondrial biogenesis, OXPHOS complexes, AMPK signaling, mitophagy, or atrogenes in females. Our results highlight the different trajectories of age-related declines in muscle mass and function, providing insights into sex-dependent molecular changes associated with sarcopenia progression, which may inform the future development of novel therapeutic interventions.

Keywords:  Aging; Mitochondria; Mouse models; Muscle biology; Skeletal muscle

DOI:  https://doi.org/10.1172/JCI172890
Proc Natl Acad Sci U S A. 2024 Aug 20. 121(34): e2404738121

Circadian period is compensated for repressor protein turnover rates in single cells.

Christian H Gabriel, Marta Del Olmo, Arunya Rizki Widini, Rashin Roshanbin, Jonas Woyde, Ebrahim Hamza, Nica-Nicoleta Gutu, Amin Zehtabian, Helge Ewers, Adrian Granada, Hanspeter Herzel, Achim Kramer.

  Most mammalian cells have molecular circadian clocks that generate widespread rhythms in transcript and protein abundance. While circadian clocks are robust to fluctuations in the cellular environment, little is known about the mechanisms by which the circadian period compensates for fluctuating metabolic states. Here, we exploit the heterogeneity of single cells both in circadian period and a metabolic parameter-protein stability-to study their interdependence without the need for genetic manipulation. We generated cells expressing key circadian proteins (CRYPTOCHROME1/2 (CRY1/2) and PERIOD1/2 (PER1/2)) as endogenous fusions with fluorescent proteins and simultaneously monitored circadian rhythms and degradation in thousands of single cells. We found that the circadian period compensates for fluctuations in the turnover rates of circadian repressor proteins and uncovered possible mechanisms using a mathematical model. In addition, the stabilities of the repressor proteins are circadian phase dependent and correlate with the circadian period in a phase-dependent manner, in contrast to the prevailing model.

Keywords:  circadian rhythms; fluorescence microscopy; metabolic compensation; protein degradation; single-cell imaging

DOI:  https://doi.org/10.1073/pnas.2404738121
Cancer Lett. 2024 Aug 09. pii: S0304-3835(24)00557-3. [Epub ahead of print]601 217162

Enhanced cytokine signaling and ferroptosis defense interplay initiates obesity-associated pancreatic ductal adenocarcinoma.

Rexiati Ruze, Yuan Chen, Jianlu Song, Ruiyuan Xu, Xinpeng Yin, Qiang Xu, Chengcheng Wang, Yupei Zhao.

  Obesity is a significant risk factor for various cancers, including pancreatic cancer (PC), but the underlying mechanisms are still unclear. In our study, pancreatic ductal epithelial cells were cultured using serum from human subjects with diverse metabolic statuses, revealing that serum from patients with obesity alters inflammatory cytokine signaling and ferroptosis, where a mutual enhancement between interleukin 34 (IL-34) expression and ferroptosis defense was observed in these cells. Notably, oncogenic KRASG12D amplified their interaction and this leads to the initiation of pancreatic ductal adenocarcinoma (PDAC) in diet-induced obese mice via macrophage-mediated immunosuppression. Single-cell RNA sequencing (scRNA-seq) of human samples showed that cytokine signaling, ferroptosis defense, and immunosuppression are correlated with the patients' body mass index (BMI) during PDAC progression. Our findings provide a mechanistic link between obesity, inflammation, ferroptosis defense, and pancreatic cancer, suggesting novel therapeutic targets for the prevention and treatment of obesity-associated PDAC.

Keywords:  Carcinogenesis; Ferroptosis defense; IL-34; Obesity; Pancreatic cancer

DOI:  https://doi.org/10.1016/j.canlet.2024.217162
Sci Adv. 2024 Aug 16. 10(33): eadk0015

A high-throughput microfabricated platform for rapid quantification of metastatic potential.

Smiti Bhattacharya, Abora Ettela, Jonathan Haydak, Chad M Hobson, Alan Stern, Miran Yoo, Teng-Leong Chew, G Luca Gusella, Emily J Gallagher, James C Hone, Evren U Azeloglu.

Assays that measure morphology, proliferation, motility, deformability, and migration are used to study the invasiveness of cancer cells. However, native invasive potential of cells may be hidden from these contextual metrics because they depend on culture conditions. We created a micropatterned chip that mimics the native environmental conditions, quantifies the invasive potential of tumor cells, and improves our understanding of the malignancy signatures. Unlike conventional assays, which rely on indirect measurements of metastatic potential, our method uses three-dimensional microchannels to measure the basal native invasiveness without chemoattractants or microfluidics. No change in cell death or proliferation is observed on our chips. Using six cancer cell lines, we show that our system is more sensitive than other motility-based assays, measures of nuclear deformability, or cell morphometrics. In addition to quantifying metastatic potential, our platform can distinguish between motility and invasiveness, help study molecular mechanisms of invasion, and screen for targeted therapeutics.

DOI: https://doi.org/10.1126/sciadv.adk0015
Nature. 2024 Aug 14.

Remodelling of the translatome controls diet and its impact on tumorigenesis.

Haojun Yang, Vincenzo Andrea Zingaro, James Lincoff, Harrison Tom, Satoshi Oikawa, Juan A Oses-Prieto, Quinn Edmondson, Ian Seiple, Hardik Shah, Shingo Kajimura, Alma L Burlingame, Michael Grabe, Davide Ruggero.

Fasting is associated with a range of health benefits1-6. How fasting signals elicit changes in the proteome to establish metabolic programmes remains poorly understood. Here we show that hepatocytes selectively remodel the translatome while global translation is paradoxically downregulated during fasting7,8. We discover that phosphorylation of eukaryotic translation initiation factor 4E (P-eIF4E) is induced during fasting. We show that P-eIF4E is responsible for controlling the translation of genes involved in lipid catabolism and the production of ketone bodies. Inhibiting P-eIF4E impairs ketogenesis in response to fasting and a ketogenic diet. P-eIF4E regulates those messenger RNAs through a specific translation regulatory element within their 5' untranslated regions (5' UTRs). Our findings reveal a new signalling property of fatty acids, which are elevated during fasting. We found that fatty acids bind and induce AMP-activated protein kinase (AMPK) kinase activity that in turn enhances the phosphorylation of MAP kinase-interacting protein kinase (MNK), the kinase that phosphorylates eIF4E. The AMPK-MNK-eIF4E axis controls ketogenesis, revealing a new lipid-mediated kinase signalling pathway that links ketogenesis to translation control. Certain types of cancer use ketone bodies as an energy source9,10 that may rely on P-eIF4E. Our findings reveal that on a ketogenic diet, treatment with eFT508 (also known as tomivosertib; a P-eIF4E inhibitor) restrains pancreatic tumour growth. Thus, our findings unveil a new fatty acid-induced signalling pathway that activates selective translation, which underlies ketogenesis and provides a tailored diet intervention therapy for cancer.

DOI: https://doi.org/10.1038/s41586-024-07781-7
FEBS J. 2024 Aug 12.

Dynamical modelling of lipid droplet formation suggests a key function of membrane phospholipids.

Hermann-Georg Holzhütter.

  Cells store triacylglycerol (TAG) within lipid droplets (LDs). A dynamic model describing complete LD formation at the endoplasmic reticulum (ER) membrane does not yet exist. A biochemical-biophysical model of LD synthesis is proposed. It describes the time-dependent accumulation of TAG in the ER membrane as the formation of a potential LD (pLD) bounded by spherical caps of the inner and outer monolayers of the membrane. The expansion rate of the pLD depends on the TAG supply, the elastic properties of the ER membrane, and the recruitment of phospholipids (PLs) to the cap-covering monolayers. Model simulations provided the following insights: (a) Marginal differences in the surface tension of the cap monolayers are sufficient to fully drive the expansion of the pLD towards the cytosol or lumen. (b) Selective reduction of PL supply to the luminal monolayer ensures stable formation of cytosolic LDs, irrespective of variations in the elasto-mechanical properties of the ER membrane. (c) The rate of TAG supply to the cytosolic monolayer has a major effect on the size and maturation time of LDs but has no significant effect on the TAG export per individual LD. The recruitment of additional PLs to the cap monolayers of pLDs critically controls the budding direction, size, and maturation time of LDs. The ability of cells to acquire additional LD initiation sites appears to be key to coping with acutely high levels of potentially toxic free fatty acids.

Keywords:  computer simulations; dynamic model; lipid droplet; triacylglycerol storage

DOI:  https://doi.org/10.1111/febs.17238
Nat Methods. 2024 Aug;21(8): 1430-1443

Transformers in single-cell omics: a review and new perspectives.

Artur Szałata, Karin Hrovatin, Sören Becker, Alejandro Tejada-Lapuerta, Haotian Cui, Bo Wang, Fabian J Theis.

Recent efforts to construct reference maps of cellular phenotypes have expanded the volume and diversity of single-cell omics data, providing an unprecedented resource for studying cell properties. Despite the availability of rich datasets and their continued growth, current single-cell models are unable to fully capitalize on the information they contain. Transformers have become the architecture of choice for foundation models in other domains owing to their ability to generalize to heterogeneous, large-scale datasets. Thus, the question arises of whether transformers could set off a similar shift in the field of single-cell modeling. Here we first describe the transformer architecture and its single-cell adaptations and then present a comprehensive review of the existing applications of transformers in single-cell analysis and critically discuss their future potential for single-cell biology. By studying limitations and technical challenges, we aim to provide a structured outlook for future research directions at the intersection of machine learning and single-cell biology.

DOI: https://doi.org/10.1038/s41592-024-02353-z
Adv Nutr. 2024 Aug 08. pii: S2161-8313(24)00116-9. [Epub ahead of print] 100282

Prevalence of and survival with cachexia among patients with cancer: A systematic review and meta-analysis.

Tomoya Takaoka, Akinori Yaegashi, Daiki Watanabe.

   BACKGROUND: Cachexia is associated with lower overall survival in patients with cancer; however, the relationship between the two is reported to differ according to the definitive criteria for diagnosing cachexia.
OBJECTIVE: We aimed to investigate the (1) difference in the prevalence of cachexia in patients with cancer and (2) association between cachexia and overall survival, depending on the definitive criteria for diagnosing cachexia in patients with cancer.
METHODS: We searched PubMed and Web of Science from their inception until July 31, 2023 to identify eligible studies. We conducted a systematic review of the prevalence of cachexia in patients with cancer and performed a meta-analysis to investigate its relationship with overall survival.
RESULTS: A total of 125 articles comprising 137,960 patients were included in the systematic review, and 26 articles consisting of 11,118 patients underwent meta-analysis. The overall prevalence of cachexia in patients with cancer was 33.0% (95% confidence interval [CI], 32.8-33.3); however, it varied according to the definitive criteria for diagnosing cachexia (13.9-56.5%). According to the Fearon 2011 criteria, the prevalence of cachexia was associated with a high hazard ratio (HR) for overall survival compared to that of non-cachexia (HR: 1.58 [95% CI, 1.45-1.73]); according to the other criteria, the HR was 2.78 (95% CI, 1.88-4.11), indicating significant subgroup differences (p = 0.006). The dose-response curve indicated that the HR for overall survival plateaued at a cachexia prevalence range of 40-50% (L-shaped relationship).
CONCLUSIONS: The prevalence of cachexia in patients with cancer may vary depending on the definitive criteria used to diagnose cachexia. The HR for overall survival was higher for low cachexia prevalence. The definitive criteria should be carefully considered when assessing cachexia in patients with cancer. This study was registered with PROSPERO as CRD42023435474.
STATEMENT OF SIGNIFICANCE: This study provides crucial insights into the prognostic impact of the differential diagnostic criteria for cancer cachexia in clinical practice. We provide quantitative and qualitative evidence of a significant difference in the overall survival and prevalence of cachexia, depending on the definitive criteria for diagnosing cachexia in patients with cancer.

Keywords:  Cancer cachexia; definitive criteria; diagnostic accuracy; heterogeneity; prognosis

DOI:  https://doi.org/10.1016/j.advnut.2024.100282
Cell. 2024 Aug 07. pii: S0092-8674(24)00824-9. [Epub ahead of print]

Macrophage-mediated myelin recycling fuels brain cancer malignancy.

Daan J Kloosterman, Johanna Erbani, Menno Boon, Martina Farber, Shanna M Handgraaf, Masami Ando-Kuri, Elena Sánchez-López, Bauke Fontein, Marjolijn Mertz, Marja Nieuwland, Ning Qing Liu, Gabriel Forn-Cuni, Nicole N van der Wel, Anita E Grootemaat, Luuk Reinalda, Sander I van Kasteren, Elzo de Wit, Brian Ruffell, Ewa Snaar-Jagalska, Kevin Petrecca, Dieta Brandsma, Alexander Kros, Martin Giera, Leila Akkari.

  Tumors growing in metabolically challenged environments, such as glioblastoma in the brain, are particularly reliant on crosstalk with their tumor microenvironment (TME) to satisfy their high energetic needs. To study the intricacies of this metabolic interplay, we interrogated the heterogeneity of the glioblastoma TME using single-cell and multi-omics analyses and identified metabolically rewired tumor-associated macrophage (TAM) subpopulations with pro-tumorigenic properties. These TAM subsets, termed lipid-laden macrophages (LLMs) to reflect their cholesterol accumulation, are epigenetically rewired, display immunosuppressive features, and are enriched in the aggressive mesenchymal glioblastoma subtype. Engulfment of cholesterol-rich myelin debris endows subsets of TAMs to acquire an LLM phenotype. Subsequently, LLMs directly transfer myelin-derived lipids to cancer cells in an LXR/Abca1-dependent manner, thereby fueling the heightened metabolic demands of mesenchymal glioblastoma. Our work provides an in-depth understanding of the immune-metabolic interplay during glioblastoma progression, thereby laying a framework to unveil targetable metabolic vulnerabilities in glioblastoma.

Keywords:  cancer immunity; cholesterol; glioblastoma; lipid metabolism; macrophages; myelin recycling; tumor microenvironment

DOI:  https://doi.org/10.1016/j.cell.2024.07.030
bioRxiv. 2024 Aug 09. pii: 2024.08.08.607195. [Epub ahead of print]

Mitochondrial Ca 2+ controls pancreatic cancer growth and metastasis by regulating epithelial cell plasticity.

Jillian S Weissenrieder, Jessica Peura, Usha Paudel, Nikita Bhalerao, Natalie Weinmann, Calvin Johnson, Maximilian Wengyn, Rebecca Drager, Emma Elizabeth Furth, Karl Simin, Marcus Ruscetti, Ben Z Stanger, Anil K Rustgi, Jason R Pitarresi, J Kevin Foskett.

Endoplasmic reticulum to mitochondria Ca 2+ transfer is important for cancer cell survival, but the role of mitochondrial Ca 2+ uptake through the mitochondrial Ca 2+ uniporter (MCU) in pancreatic adenocarcinoma (PDAC) is poorly understood. Here, we show that increased MCU expression is associated with malignancy and poorer outcomes in PDAC patients. In isogenic murine PDAC models, Mcu deletion ( Mcu KO ) ablated mitochondrial Ca 2+ uptake, which reduced proliferation and inhibited self-renewal. Orthotopic implantation of MCU-null tumor cells reduced primary tumor growth and metastasis. Mcu deletion reduced the cellular plasticity of tumor cells by inhibiting epithelial-to-mesenchymal transition (EMT), which contributes to metastatic competency in PDAC. Mechanistically, the loss of mitochondrial Ca 2+ uptake reduced expression of the key EMT transcription factor Snail and secretion of the EMT-inducing ligand TGFβ. Snail re-expression and TGFβ treatment rescued deficits in Mcu KO cells and restored their metastatic ability. Thus, MCU may present a therapeutic target in PDAC to limit cancer-cell-induced EMT and metastasis.

DOI: https://doi.org/10.1101/2024.08.08.607195
Aging Cell. 2024 Aug 14. e14287

Aging human abdominal subcutaneous white adipose tissue at single cell resolution.

K L Whytock, A Divoux, Y Sun, M F Pino, G Yu, C A Jin, J J Robino, A Plekhanov, O Varlamov, S R Smith, M J Walsh, L M Sparks.

  White adipose tissue (WAT) is a robust energy storage and endocrine organ critical for maintaining metabolic health as we age. Our aim was to identify cell-specific transcriptional aberrations that occur in WAT with aging. We leveraged full-length snRNA-Seq and histology to characterize the cellular landscape of human abdominal subcutaneous WAT in a prospective cohort of 10 younger (≤30 years) and 10 older individuals (≥65 years) balanced for sex and body mass index (BMI). The older group had greater cholesterol, very-low-density lipoprotein, triglycerides, thyroid stimulating hormone, and aspartate transaminase compared to the younger group (p < 0.05). We highlight that aging WAT is associated with adipocyte hypertrophy, increased proportions of lipid-associated macrophages and mast cells, an upregulation of immune responses linked to fibrosis in pre-adipocyte, adipocyte, and vascular populations, and highlight CXCL14 as a biomarker of these processes. We show that older WAT has elevated levels of senescence marker p16 in adipocytes and identify the adipocyte subpopulation driving this senescence profile. We confirm that these transcriptional and phenotypical changes occur without overt fibrosis and in older individuals that have comparable WAT insulin sensitivity to the younger individuals.

Keywords:  White adipose tissue; aging; senescence; single nuclei RNA‐Seq

DOI:  https://doi.org/10.1111/acel.14287
Cell Death Differ. 2024 Aug 13.

Rewiring cancer cell death: LPCAT1 shapes lipid composition and ferroptosis resistance.

Hyemin Lee, Li Zhuang, Boyi Gan.

DOI: https://doi.org/10.1038/s41418-024-01364-9
Autophagy. 2024 Aug 12.

Recycling the recyclers: lysophagy emerges as a new pharmacological target for retinal degeneration.

Juan Ignacio Jiménez-Loygorri, Patricia Boya.

  Dysregulated macroautophagy/autophagy is one of the hallmarks of aging and has also been linked to higher incidence of several age-associated diseases such as age-related macular degeneration (AMD). The main cell type affected in AMD is the retinal pigment epithelium (RPE), and this disease can lead to central vision loss. Despite affecting around 8.7% of the population between 45-85 years, its etiopathogenesis remains unknown. In our recent manuscript using the pharmacological sodium iodate (SI) model of AMD we identified severe lysosomal membrane permeabilization (LMP) in the RPE, that leads to autophagy flux blockage and proteostasis defects. Treatment with the natural compound urolithin A (UA) reduces RPE cell death and alleviates vision loss, concurrent with full autophagy restoration. While UA was initially described as a specific mitophagy inducer, we now show that it is also able to promote SQSTM1/p62-dependent lysophagy in the context of lysosomal damage and LMP. Genetic downregulation of SQSTM1/p62 fully abolishes the effect of UA on lysophagy while mitophagy stimulation remains unaffected. In summary, these findings highlight the wide range of pathways modulated by UA and its potential implementation in the management of AMD and other diseases involving lysosomal damage.

Keywords:  AMD; RPE; autophagy; lysosomal membrane permeabilization; lysosome

DOI:  https://doi.org/10.1080/15548627.2024.2391726
Biofabrication. 2024 Aug 09.

3Din vitromodeling of the exocrine pancreatic unit using tomographic volumetric bioprinting.

Viola Sgarminato, Jorge Madrid-Wolff, Antoine Boniface, Gianluca Ciardelli, Chiara Tonda-Turo, Christophe Moser.

  Pancreatic ductal adenocarcinoma (PDAC) is the most common type of pancreatic cancer, a leading cause of cancer-related deaths globally. Initial lesions of PDAC develop within the exocrine pancreas' functional units, with tumor progression driven by interactions between PDAC and stromal cells. Effective therapies require anatomically and functionally relevant in vitro human models of the pancreatic cancer microenvironment. We employed tomographic volumetric bioprinting, a novel biofabrication method, to create human fibroblast-laden constructs mimicking the tubuloacinar structures of the exocrine pancreas. Human pancreatic ductal epithelial (HPDE) cells overexpressing the KRAS oncogene (HPDE-KRAS) were seeded in the multiacinar cavity to replicate pathological tissue. HPDE cell growth and organization within the structure were assessed, demonstrating the formation of a thin epithelium covering the acini inner surfaces. Immunofluorescence assays showed significantly higher alpha smooth muscle actin (α-SMA) vs. F-actin expression in fibroblasts co-cultured with cancerous versus wild-type HPDE cells. Additionally, α-SMA expression increased over time and was higher in fibroblasts closer to HPDE cells. Elevated interleukin (IL)-6 levels were quantified in supernatants from co-cultures of stromal and HPDE-KRAS cells. These findings align with inflamed tumor-associated myofibroblast behavior, serving as relevant biomarkers to monitor early disease progression and target drug efficacy. To our knowledge, this is the first demonstration of a 3D bioprinted model of exocrine pancreas that recapitulates its true 3-dimensional microanatomy and shows tumor triggered inflammation.&#xD.

Keywords:  3D in vitro models; additive manufacturing; glandular structures; pancreatic cancer; tissue engineering; tomographic volumetric bioprinting

DOI:  https://doi.org/10.1088/1758-5090/ad6d8d
Int J Cancer. 2024 Aug 15.

Clinical and molecular features of early onset pancreatic adenocarcinoma.

Maxime Rémond, Cristina Smolenschi, Anthony Tarabay, Maximiliano Gelli, Elena Fernandez-de-Sevilla, Ali Mouawia, Simona Cosconea, Lambros Tselikas, Remy Barbe, Alina Fuerea, Mohamed A Bani, Marc Deloger, Benjamin Besse, Thomas Pudlarz, Marine Valéry, Valérie Boige, Antoine Hollebecque, Michel Ducreux, Alice Boilève.

  Pancreatic adenocarcinoma (PDAC) is a major health burden and may become the second cause of death by cancer in developed countries. The incidence of early-onset pancreatic cancer (EOPC, defined by an age at diagnosis <50 years old) is increasing. Here, we conducted a study of all PDAC patients followed at our institution. Patients were classified as EOPC or non-early onset (nEOPC, >50). Eight hundred and seventy eight patients were included, of which 113 EOPC, exhibiting a comparable performance status. EOPC were more often diagnosed at the metastatic stage (70.0% vs 58.3%) and liver metastases were more prevalent at diagnosis (60.2% vs. 43.9%). The median overall survival (OS) from diagnosis was 18.1 months, similar between EOPC and nEOPC. Among patients who underwent surgery, recurrence-free survival was similar between age groups. Among metastatic patients, first line progression free survival was similar but EOPC received more treatment lines (72.3% vs. 58.1% received ≥2 lines). Regarding molecular alterations, the mean tumor mutational burden (TMB) was lower in EOPC (1.42 vs. 2.95 mut/Mb). The prevalence of KRAS and BRCA1/2 mutations was similar, but EOPC displayed fewer alterations in CNKN2A/B. Fifty eight patients (18.6%) had actionable alterations (ESCAT I-III) and 31 of them received molecularly matched treatments. On the transcriptomic level, despite its clinical aggressiveness, EOPC was less likely to display a basal-like phenotype. To conclude, EOPC were diagnosed more frequently at the metastatic stage. OS and 1st line PFS were similar to nEOPC. EOPC displayed specific molecular features, such as a lower TMB and fewer alterations in CDKN2A/B.

Keywords:  early‐onset; pancreatic adenocarcinoma; pancreatic cancer; precision medicine; transcriptomic

DOI:  https://doi.org/10.1002/ijc.35135
Bio Protoc. 2024 Aug 05. 14(15): e5040

An Experimental Protocol for the Boyden Chamber Invasion Assay With Absorbance Readout.

Kathleen C Brown, Amanda M Sugrue, Kushal J Modi, Reagan S Light, Kaitlyn B Conley, Ashley J Cox, Christopher R Bender, Sarah L Miles, Monica A Valentovic, Piyali Dasgupta.

  The phenomenon of cell invasion is an essential step in angiogenesis, embryonic development, immune responses, and cancer metastasis. In the course of cancer progression, the ability of neoplastic cells to degrade the basement membrane and penetrate neighboring tissue (or blood vessels and lymph nodes) is an early event of the metastatic cascade. The Boyden chamber assay is one of the most prevalent methods implemented to measure the pro- or anti-invasive effects of drugs, investigate signaling pathways that modulate cell invasion, and characterize the role of extracellular matrix proteins in metastasis. However, the traditional protocol of the Boyden chamber assay has some technical challenges and limitations. One such challenge is that the endpoint of the assay involves photographing and counting stained cells (in multiple fields) on porous filters. This process is very arduous, requires multiple observers, and is very time-consuming. Our improved protocol for the Boyden chamber assay involves lysis of the dye-stained cells and reading the absorbance using an ELISA reader to mitigate this challenge. We believe that our improved Boyden chamber methodology offers a standardized, high-throughput format to evaluate the efficacy of various drugs and test compounds in influencing cellular invasion in normal and diseased states. We believe that our protocol will be useful for researchers working in the fields of immunology, vascular biology, drug discovery, cancer biology, and developmental biology. Key features • Measurement of tumor invasion using human cancer cells. • Ability to measure the pro-invasive/anti-invasive activity of small molecules and biological modifiers. • Measurement of chemotaxis, chemokines, trafficking of immune cells, and proteolytic activity of matrix metalloproteinases, lysosomal hydrolysates, collagenases, and plasminogen activators in physiological and pathological conditions. • Investigation of the role of extracellular matrix proteins in the crosstalk between endothelial, epithelial, muscle, or neuronal cells and their adjacent stroma.

Keywords:  Boyden; Chemoattractant; Inserts; Invasion; Matrigel; Metastasis

DOI:  https://doi.org/10.21769/BioProtoc.5040
bioRxiv. 2024 Jul 31. pii: 2024.07.30.605703. [Epub ahead of print]

Unraveling cysteine deficiency-associated rapid weight loss.

Alan Varghese, Ivan Gusarov, Begoña Gamallo-Lana, Daria Dolgonos, Yatin Mankan, Ilya Shamovsky, Mydia Phan, Rebecca Jones, Maria Gomez-Jenkins, Eileen White, Rui Wang, Drew Jones, Thales Papagiannakopoulos, Michael E Pacold, Adam C Mar, Dan R Littman, Evgeny Nudler.

Forty percent of the US population and 1 in 6 individuals worldwide are obese, and the incidence of this disease is surging globally1,2. Various dietary interventions, including carbohydrate and fat restriction, and more recently amino acid restriction, have been explored to combat this epidemic3-6. We sought to investigate the impact of removing individual amino acids on the weight profiles of mice. Compared to essential amino acid restriction, induction of conditional cysteine restriction resulted in the most dramatic weight loss, amounting to 20% within 3 days and 30% within one week, which was readily reversed. This weight loss occurred despite the presence of substantial cysteine reserves stored in glutathione (GSH) across various tissues7. Further analysis demonstrated that the weight reduction primarily stemmed from an increase in the utilization of fat mass, while locomotion, circadian rhythm and histological appearance of multiple other tissues remained largely unaffected. Cysteine deficiency activated the integrated stress response (ISR) and NRF2-mediated oxidative stress response (OSR), which amplify each other, leading to the induction of GDF15 and FGF21, hormones associated with increased lipolysis, energy homeostasis and food aversion8-10. We additionally observed rapid tissue coenzyme A (CoA) depletion, resulting in energetically inefficient anaerobic glycolysis and TCA cycle, with sustained urinary excretion of pyruvate, orotate, citrate, α-ketoglutarate, nitrogen rich compounds and amino acids. In summary, our investigation highlights that cysteine restriction, by depleting GSH and CoA, exerts a maximal impact on weight loss, metabolism, and stress signaling compared to other amino acid restrictions. These findings may pave the way for innovative strategies for addressing a range of metabolic diseases and the growing obesity crisis.

DOI: https://doi.org/10.1101/2024.07.30.605703
Nat Cancer. 2024 Aug 12.

SOS1 inhibitor combinations overcome KRAS inhibitor resistance.

DOI: https://doi.org/10.1038/s43018-024-00801-5
Nat Protoc. 2024 Aug 15.

Proximity sequencing for the detection of mRNA, extracellular proteins and extracellular protein complexes in single cells.

Luke Vistain, Bijentimala Keisham, Junjie Xia, Hoang Van Phan, Savaş Tay.

Complex cellular functions occur via the coordinated formation and dissociation of protein complexes. Functions such as the response to a signaling ligand can incorporate dozens of proteins and hundreds of complexes. Until recently, it has been difficult to measure multiple protein complexes at the single-cell level. Here, we present a step-by-step procedure for proximity sequencing, which enables the simultaneous measurement of proteins, mRNA and hundreds of protein complexes located on the outer membrane of cells. We guide the user through probe creation, sample preparation, staining, sequencing and computational quantification of protein complexes. This protocol empowers researchers to study, for example, the interplay between transcriptional states and cellular functions by coupling measurements of transcription to measurements of linked effector molecules, yet could be generalizable to other paired events. The protocol requires roughly 16 h spread over several days to complete by users with expertise in basic molecular biology and single-cell sequencing.

DOI: https://doi.org/10.1038/s41596-024-01030-x
Methods Mol Biol. 2024 ;2828 11-21

Methods and Protocol for Single-Cell Motility Assays under Topological Constraints.

Carlotta Ficorella, Federico Sala, Rebeca Martínez Vázquez, Roberto Osellame, Josef A Käs.

  The extracellular environment plays a crucial role in many physiological and pathological processes involving cell motility, such as metastatic invasion in cancer development, by heavily impacting the migration strategies adopted by the cells. The study of how mechanical constraints affect the dynamics of cell migration may be relevant to gain more insight into such processes, and it may prove to be a powerful tool in the hands of biologists. In this chapter, we describe the methods used to investigate the ability of neoplastic cells to migrate through narrowing, rigid microstructures upon chemoattractant stimulation.

Keywords:  Cancer cell migration; Immunostaining; Live fluorescent staining; Microconstriction

DOI:  https://doi.org/10.1007/978-1-0716-4023-4_2
Bioinformatics. 2024 Aug 12. pii: btae503. [Epub ahead of print]

Ceas: An R package for Seahorse data analysis and visualization.

Rachel Rae J House, James P Eapen, Hui Shen, Carrie R Graveel, Matthew R Steensma.

   SUMMARY: Measuring cellular energetics is essential to understanding a matrix's (e.g. cell, tissue or biofluid) metabolic state. The Agilent Seahorse machine is a common method to measure real-time cellular energetics, but existing analysis tools are highly manual or lack functionality. The Cellular Energetics Analysis Software (ceas) R package fills this analytical gap by providing modular and automated Seahorse data analysis and visualization.
AVAILABILITY AND IMPLEMENTATION: Ceas is available on CRAN (https://cran.r-project.org/package=ceas). Source code and installable tarballs are freely available for download at https://github.com/jamespeapen/ceas/releases/ under the MIT license. Package documentation may be found at https://jamespeapen.github.io/ceas/. ceas is implemented in R and is supported on macOS, Windows and Linux.
SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Keywords:  ATP; Seahorse; bioenergetics; metabolism

DOI:  https://doi.org/10.1093/bioinformatics/btae503
Lab Med. 2024 Aug 10. pii: lmae064. [Epub ahead of print]

Analytical evaluation of the automated interleukin-6 assay on the Roche cobas e602 analyzer.

Anastasia Gant Kanegusuku, Timothy Carll, Kiang-Teck J Yeo.

   BACKGROUND: Interleukin-6 (IL-6) is a proinflammatory cytokine that is associated with many inflammatory diseases. This validation study evaluates the automated Roche Elecsys IL-6 electrochemiluminescent immunoassay that has been granted emergency use authorization by the US Food and Drug Administration.
METHODS: The Elecsys IL-6 assay was evaluated for precision, linearity, interference (by hemoglobin, bilirubin, triglycerides, and biotin) and clinical performance was compared to the V-PLEX Human IL-6 immunoassay (Meso Scale Discovery), performed by a reference laboratory.
RESULTS: The Elecsys IL-6 assay is precise (intra-assay <3% coefficient of variation [CV], interassay <5% CV), exhibits an analytical measurable range of 1.5-4790 pg/mL, and is tolerant of significant interferences (H < 2522, I <62, L<2101, biotin <50 ng/mL). Comparison with the V-PLEX assay revealed a 2.95 slope bias in patient samples evaluated for IL-6 concentration (n = 43, range = 1.5-1891 pg/mL, y = 2.95x - 32.7, r2 = 0.84). Bland-Altman analysis revealed an absolute mean bias of 152 pg/mL (SD = 254 pg/mL), or a mean percentage difference of 73%.
CONCLUSION: The Roche IL-6 assay showed good analytical performance. The large systematic bias compared with another reference method precludes using multiple methods to monitor IL-6 response. The random-access nature of an automated IL-6 assay on the Roche platform makes the test available on demand.

Keywords:  IL-6; automated; chemistry; clinical chemistry; immunoassay; laboratory developed test

DOI:  https://doi.org/10.1093/labmed/lmae064
bioRxiv. 2024 Aug 08. pii: 2024.08.06.606880. [Epub ahead of print]

Cysteine depletion triggers adipose tissue thermogenesis and weight-loss.

Aileen H Lee, Lucie Orliaguet, Yun-Hee Youm, Rae Maeda, Tamara Dlugos, Yuanjiu Lei, Daniel Coman, Irina Shchukina, Sairam Andhey, Steven R Smith, Eric Ravussin, Krisztian Stadler, Fahmeed Hyder, Maxim N Artyomov, Yuki Sugiura, Vishwa Deep Dixit.

Dietary interventions such as caloric restriction (CR) 1 and methionine restriction 2 that prolong lifespan induce the 'browning' of white adipose tissue (WAT), an adaptive metabolic response that increases heat production to maintain health 3,4 . However, how diet influences adipose browning and metabolic health is unclear. Here, we identified that weight-loss induced by CR in humans 5 reduces cysteine concentration in WAT suggesting depletion of this amino-acid may be involved in metabolic benefits of CR. To investigate the role of cysteine on organismal metabolism, we created a cysteine-deficiency mouse model in which dietary cysteine was eliminated and cystathionine γ-lyase (CTH) 6 , the enzyme that synthesizes cysteine was conditionally deleted. Using this animal model, we found that systemic cysteine-depletion causes drastic weight-loss with increased fat utilization and browning of adipose tissue. The restoration of dietary cysteine in cysteine-deficient mice rescued weight loss together with reversal of adipose browning and increased food-intake in an on-demand fashion. Mechanistically, cysteine deficiency induced browning and weight loss is dependent on sympathetic nervous system derived noradrenaline signaling via β3-adrenergic-receptors and does not require UCP1. Therapeutically, in high-fat diet fed obese mice, one week of cysteine-deficiency caused 30% weight-loss and reversed inflammation. These findings thus establish that cysteine is essential for organismal metabolism as removal of cysteine in the host triggers adipose browning and rapid weight loss.

DOI: https://doi.org/10.1101/2024.08.06.606880
Cell Rep Methods. 2024 Aug 04. pii: S2667-2375(24)00212-1. [Epub ahead of print] 100839

Multiomics2Targets identifies targets from cancer cohorts profiled with transcriptomics, proteomics, and phosphoproteomics.

Eden Z Deng, Giacomo B Marino, Daniel J B Clarke, Ido Diamant, Adam C Resnick, Weiping Ma, Pei Wang, Avi Ma'ayan.

  The availability of data from profiling of cancer patients with multiomics is rapidly increasing. However, integrative analysis of such data for personalized target identification is not trivial. Multiomics2Targets is a platform that enables users to upload transcriptomics, proteomics, and phosphoproteomics data matrices collected from the same cohort of cancer patients. After uploading the data, Multiomics2Targets produces a report that resembles a research publication. The uploaded matrices are processed, analyzed, and visualized using the tools Enrichr, KEA3, ChEA3, Expression2Kinases, and TargetRanger to identify and prioritize proteins, genes, and transcripts as potential targets. Figures and tables, as well as descriptions of the methods and results, are automatically generated. Reports include an abstract, introduction, methods, results, discussion, conclusions, and references and are exportable as citable PDFs and Jupyter Notebooks. Multiomics2Targets is applied to analyze version 3 of the Clinical Proteomic Tumor Analysis Consortium (CPTAC3) pan-cancer cohort, identifying potential targets for each CPTAC3 cancer subtype. Multiomics2Targets is available from https://multiomics2targets.maayanlab.cloud/.

Keywords:  CP: Cancer biology; CP: Systems biology; ChEA3; Expression2Kinases; GPT; KEA3; LLM; X2K; multiomics; transcriptomics

DOI:  https://doi.org/10.1016/j.crmeth.2024.100839
Aging Cell. 2024 Aug 14. e14297

Identification of senescent cell subpopulations by CITE-seq analysis.

Kotb Abdelmohsen, Krystyna Mazan-Mamczarz, Rachel Munk, Dimitrios Tsitsipatis, Qiong Meng, Martina Rossi, Apala Pal, Chang Hoon Shin, Jennifer L Martindale, Yulan Piao, Jinshui Fan, Hagai Yanai, Supriyo De, Isabel Beerman, Myriam Gorospe.

  Cellular senescence, a state of persistent growth arrest, is closely associated with aging and age-related diseases. Deciphering the heterogeneity within senescent cell populations and identifying therapeutic targets are paramount for mitigating senescence-associated pathologies. In this study, proteins on the surface of cells rendered senescent by replicative exhaustion and by exposure to ionizing radiation (IR) were identified using mass spectrometry analysis, and a subset of them was further studied using single-cell CITE-seq (Cellular Indexing of Transcriptomes and Epitopes by Sequencing) analysis. Based on the presence of proteins on the cell surface, we identified two distinct IR-induced senescent cell populations: one characterized by high levels of CD109 and CD112 (cluster 3), the other characterized by high levels of CD112, CD26, CD73, HLA-ABC, CD54, CD49A, and CD44 (cluster 0). We further found that cluster 0 represented proliferating and senescent cells in the G1 phase of the division cycle, and CITE-seq detection of cell surface proteins selectively discerned those in the senescence group. Our study highlights the heterogeneity of senescent cells and underscores the value of cell surface proteins as tools for distinguishing senescent cell programs and subclasses, paving the way for targeted therapeutic strategies in disorders exacerbated by senescence.

Keywords:  CITE‐seq; cell cycle; proteome; senescence; single‐cell transcriptome; surface proteins; surfaceome

DOI:  https://doi.org/10.1111/acel.14297
Adv Sci (Weinh). 2024 Aug 13. e2400918

Profiling Dynamic Patterns of Single-Cell Motility.

Debonil Maity, Nikita Sivakumar, Pratik Kamat, Nahuel Zamponi, Chanhong Min, Wenxuan Du, Hasini Jayatilaka, Adrian Johnston, Bartholomew Starich, Anshika Agrawal, Deanna Riley, Leandro Venturutti, Ari Melnick, Leandro Cerchietti, Jeremy Walston, Jude M Phillip.

  Cell motility plays an essential role in many biological processes as cells move and interact within their local microenvironments. Current methods for quantifying cell motility typically involve tracking individual cells over time, but the results are often presented as averaged values across cell populations. While informative, these ensemble approaches have limitations in assessing cellular heterogeneity and identifying generalizable patterns of single-cell behaviors, at baseline and in response to perturbations. In this study, CaMI is introduced, a computational framework designed to leverage the single-cell nature of motility data. CaMI identifies and classifies distinct spatio-temporal behaviors of individual cells, enabling robust classification of single-cell motility patterns in a large dataset (n = 74 253 cells). This framework allows quantification of spatial and temporal heterogeneities, determination of single-cell motility behaviors across various biological conditions and provides a visualization scheme for direct interpretation of dynamic cell behaviors. Importantly, CaMI reveals insights that conventional cell motility analyses may overlook, showcasing its utility in uncovering robust biological insights. Together, a multivariate framework is presented to classify emergent patterns of single-cell motility, emphasizing the critical role of cellular heterogeneity in shaping cell behaviors across populations.

Keywords:  cell motility; high‐throughput cell phenotyping; single‐cell behaviors

DOI:  https://doi.org/10.1002/advs.202400918
Nucleic Acids Res. 2024 Aug 15. pii: gkae692. [Epub ahead of print]

Tracking live-cell single-molecule dynamics enables measurements of heterochromatin-associated protein-protein interactions.

Ziyuan Chen, Melissa Seman, Yekaterina Fyodorova, Ali Farhat, Amanda Ames, Alexander Levashkevich, Saikat Biswas, Fengting Huang, Lydia Freddolino, Julie S Biteen, Kaushik Ragunathan.

Visualizing and measuring molecular-scale interactions in living cells represents a major challenge, but recent advances in single-molecule super-resolution microscopy are bringing us closer to achieving this goal. Single-molecule super-resolution microscopy enables high-resolution and sensitive imaging of the positions and movement of molecules in living cells. HP1 proteins are important regulators of gene expression because they selectively bind and recognize H3K9 methylated (H3K9me) histones to form heterochromatin-associated protein complexes that silence gene expression, but several important mechanistic details of this process remain unexplored. Here, we extended live-cell single-molecule tracking studies in fission yeast to determine how HP1 proteins interact with their binding partners in the nucleus. We measured how genetic perturbations that affect H3K9me alter the diffusive properties of HP1 proteins and their binding partners, and we inferred their most likely interaction sites. Our results demonstrate that H3K9 methylation spatially restricts HP1 proteins and their interactors, thereby promoting ternary complex formation on chromatin while simultaneously suppressing off-chromatin binding. As opposed to being an inert platform to direct HP1 binding, our studies propose a novel function for H3K9me in promoting ternary complex formation by enhancing the specificity and stimulating the assembly of HP1-protein complexes in living cells.

DOI: https://doi.org/10.1093/nar/gkae692