Augmenting cGAS/STING-mediated innate immunity by copper-based nanoplatform for synergistic cancer immunotherapy via inducing irreversible DNA damage.

Innate Immune Dysfunction in Trauma Patients

Camptothecin (CPT) is a topoisomerase I inhibitor, derivatives of which are being used for cancer chemotherapy. CPT-induced DNA double-strand breaks (DSBs) are considered a major cause of its tumoricidal activity, and it has been shown that CPT induces DNA damage signaling through the phosphatidylinositol 3-kinase-related kinases, including ATM (ataxia telangiectasia mutated), ATR (ATM and Rad3-related), and DNA-PK (DNA-dependent protein kinase). In addition, CPT causes DNA strand breaks mediated by transcription, although the downstream signaling events are less well characterized. In this study, we show that CPT-induced activation of ATM requires transcription. Mechanistically, transcription inhibition suppressed CPT-dependent activation of ATM and blocked recruitment of the DNA damage mediator p53-binding protein 1 (53BP1) to DNA damage sites, whereas ATM inhibition abrogated CPT-induced G(1)/S and S phase checkpoints. Functional inactivation of ATM resulted in DNA replication-dependent hyperactivation of DNA-PK in CPT-treated cells and dramatic CPT hypersensitivity. On the other hand, simultaneous inhibition of ATM and DNA-PK partially restored CPT resistance, suggesting that activation of DNA-PK is proapoptotic in the absence of ATM. Correspondingly, comet assay and cell cycle synchronization experiments suggested that transcription collapse occurring as the result of CPT treatment are converted to frank double-strand breaks when ATM-deficient cells bypass the G(1)/S checkpoint. Thus, ATM suppresses DNA-PK-dependent cell death in response to topoisomerase poisons, a finding with potential clinical implications.

More than ten years ago, the “danger theory” challenged conservative immunology. At that time, the consensus was that the immune system is activated by antigens recognized as nonself. However, the self-nonself theory gave no explanation why, for example, a fetus with obvious foreign antigens does not lead to maternal immune activation whereas transplanted organs do. In an attempt to resolve these apparent paradoxes, the danger theory postulated that the immune system is triggered by “danger signals” released upon tissue injury and stress alerting the immune system that there is risk to the host [1]. The “danger theory” is supported by the growing number of endogenous ligands that can activate innate immune receptors such as toll-like receptors, RIG-I-like receptors, NOD-like receptors, and the inflammasome. However, it poses the challenge of identifying such signals and the mechanisms of their generation rigorously. Danger signals or danger associated molecular patterns (DAMPs) identified so far include factors like high-mobility group protein B1, mitochondrial DNA, heat shock protein (HSP), interleukin-1α, adenosine triphosphate, reactive oxygen intermediates, and uric acid [2]. A common feature of cardiovascular diseases, like myocardial infarction, heart failure, atherosclerosis, and so forth, is a robust inflammatory response. The reason for an immunologic reaction in mostly nonimmune diseases is not very well defined. However, the danger theory offers a good explanation: tissue damage, for example, in myocardial infarction, could lead to the release of danger signals and thereby cause an immune response. Indeed, in the current issue, several aspects of this process are highlighted: after a general introduction into DAMPs in the cardiovascular system [3], M. Ashri et al. discuss the theory of cardiotrophin-1 as a secondary DAMP in obesity “Update on the pathophysiological activities of the cardiac molecule cardiotrophin-1 in obesity,” whereas A. Schiopu et al. review S100A8 and S100A9, members of the calgranulin family, as potential DAMP in cardiovascular disease “S100A8 and S100A9: DAMPs at the crossroads between innate immunity, traditional risk factors, and cardiovascular disease.” F. van den Akker et al. highlight that danger signals might influence the phenotype of mesenchymal stem cells and secondarily outcome after myocardial infarction “Mesenchymal stem cell therapy for cardiac inflammation: immunomodulatory properties and the influence of toll-like receptors.” A few original articles deal with the role of oxidative stress as DAMP “Berberine protects against palmitate-induced endothelial dysfunction: involvements of upregulation of AMPK and eNOS and downregulation of NOX4” and “Natural antioxidant-isoliquiritigenin ameliorates contractile dysfunction of hypoxic cardiomyocytes via AMPK signaling pathway” and with actin or chitinase 3-like 1 as a trigger of immune activation in patients with advanced atherosclerotic plaques “Actin is a target of T-cell reactivity in patients with advanced carotid atherosclerotic plaques” and “Increased expression of chitinase 3-like 1 in aorta of patients with atherosclerosis and suppression of atherosclerosis in apolipoprotein E-knockout mice by chitinase 3-like 1 gene silencing” or complement factor C3 as marker of danger signal activation in patients with heart failure “Complement c3c as a biomarker in heart failure.” All manuscripts underline the importance of danger signals in cardiovascular disease in basic as well as clinical science. Clinical Implications. DAMPs may have great diagnostic, prognostic, and therapeutic potential. In theory, DAMPs may indicate active tissue injury. Since DAMP levels are related to the extent of injury, they may have prognostic implications. When DAMPs are the most important trigger for immune activation, pharmaceutical interference should allow tailoring an immune response. However, it has to be beard in mind that the activation of the immune system in the context of tissue injury makes evolutionary sense and is not necessarily negative. For example, after myocardial infarction depletion of macrophages causes the scar not to be cleared of cell debris and left ventricular thrombi to develop leading to adverse outcome in animals and potentially also in humans (Monocytes/macrophages prevent healing defects and left ventricular thrombus formation after myocardial infarction). Thus, an initial immune activation is necessary for a coordinated pathophysiologic and beneficial response to injury. However, a chronic immune activation might be detrimental, as has been shown by several groups. Therefore, timing will be crucial when interfering with DAMPs. In conclusion, a better understanding of DAMPs in cardiovascular disease might give us dual benefit: it will help us to identify and treat patients at the very core of the pathophysiological process. However, markers and potential drug targets warrant further research. Stefan Frantz Claudia Monaco Fatih Arslan

LITERATURE Watch: Implications for transplantation

Cancer immunotherapy has transformed oncological treatment paradigms, yet tumor resistance and immune evasion continue to limit therapeutic efficacy. Mitochondria-targeting organic sensitizers (MTOSs) represent an emerging class of therapeutic agents that exploit mitochondrial dysfunction as a convergent node for tumor elimination and immune activation. As central regulators of cellular metabolism, apoptotic signaling, and immune cell function, mitochondria serve as critical determinants of tumor progression and the immunological landscape within the tumor microenvironment (TME). This comprehensive review synthesizes the latest advances (2023-2025) in MTOS-mediated cancer immunotherapy, systematically examining the capacity of MTOSs to induce diverse forms of regulated cell death and orchestrate antitumor immune responses. MTOSs demonstrate remarkable versatility in triggering mitochondria-dependent apoptosis, immunogenic cell death (ICD), necroptosis, pyroptosis, ferroptosis, and autophagic cell death through strategic disruption of mitochondrial homeostasis. These sensitizers modulate key mitochondrial functions including membrane potential dynamics, reactive oxygen species (ROS) generation, electron transport chain integrity, and calcium homeostasis, thereby releasing damage-associated molecular patterns (DAMPs) that potently activate both innate and adaptive immunity. Current MTOS platforms encompass small-molecule sensitizers, polymeric nanocarriers, metal-organic complexes, and biomimetic systems, each offering distinct advantages in mitochondrial targeting and therapeutic efficacy. Clinical translation faces significant challenges including variable mitochondrial targeting efficiency due to transmembrane transport limitations and TME pH fluctuations, systemic toxicity risks from nonspecific metal ion release in metal-organic complexes, insufficient long-term biocompatibility evaluation, and the predominant reliance on simplified tumor models that inadequately reflect clinical heterogeneity and complex spatiotemporal dynamics of mitochondrial damage-immune remodeling interactions. Future research directions emphasize the multidisciplinary integration of synthetic biology, nanotechnology, and computational approaches to engineer next-generation intelligent sensitizer platforms with enhanced TME-adaptive capabilities, enabling precise mitochondrial intervention and immune modulation for improved cancer immunotherapy outcomes.

Deficiency in one or more subsystems of the DNA damage response and repair system hierarchy is a hallmark of cancer and makes DNA repair an attractive target for physical or chemical therapies. An accurate, complete, yet parsimonious model of the systems and pathways comprising the hierarchy of DNA repair mechanisms can be expected to further our understanding of cancer genesis and progression and to facilitate the development of improved cancer therapies. Previous attempts in building a hierarchical model of DNA repair include methods based on literature curation (Gene Ontology [1]) and expert consensus (2). However, as these models rely on manual curation, rapid integration of novel omics-scale experimental data sets is not practicable. Moreover, Gene Ontology is by its definition only concerned with the modeling of pathways in the healthy cell and does not aim to reflect cancer-specific aspects. To bridge this gap, we here describe the generation of an entirely data-driven model of DNA repair, inferred from a large meta-compendium describing the physical interactions between proteins based on an integration of over 110 experimental data sets from different omics levels (genome, transcriptome, proteome). Augmenting a technique that we developed earlier, Active Interaction Mapping (AIM) (3), we define a hierarchical model of DNA repair consisting of 293 genes organized in 40 systems. Briefly, we integrate the interaction evidence using a random forest regressor to generate a weighted, integrated network; we then transform the network into a consensus matrix and feed this into an algorithm based on previous work (4) to identify the hierarchical structure embedded in the network. The hierarchical model takes the form of a directed acyclic graph (DAG), which allows us to model pleiotropic aspects of molecular systems, as some proteins and systems play functional roles in different ancestor systems. Using a network biology approach, we suggest a list of 293 genes playing central roles in DNA repair and find evidence that genome replication and RNA splicing processes are more intimately connected to DNA repair than previously appreciated. We find novel roles for about one third of the genes in our model, identify novel subsystems of already known systems in DNA repair, and can describe novel interactions between already known DNA repair systems. Finally, we perform perturbations of the DNA repair hierarchy in different cell lines, measure the impact on the protein-protein and genetic interaction networks, and use the identified interactions to search for impacted connections within and between systems of the DNA repair model.

2017 American Transplant Congress Focuses on Hot Issues.

The gene mutated in Ataxia telangiectasia termed Ataxia Telangiectasia Mutated (ATM) encodes for a Serine/Threonine protein kinase which functions at the core of signalling network responsible for DNA damage recognition and repair. Recent evidence has supported the role of ATM beyond DNA repair and has been shown to function in metabolic regulation and cellular homeostasis. Here, we show that ATM regulates mitochondrial biogenesis in MCF-7 and A549 cancer cell lines. Inhibition of ATM kinase activity with its specific inhibitor KU55933 (KU) both with and without DNA damage by Doxorubicin or Bleomycin resulted in evoked cytosolic Ca2+ ions leading to rapid, but sustained induction of mitochondrial biogenesis in these cell lines. This induction was independent of AMPK, a kinase previously shown to be involved in mitochondrial biogenesis as KU treatment resulted in downregulation of phospho AMPK. Moreover siRNA mediated knockdown of AMPK could not prevent KU dependent induction of mitochondrial biogenesis. Interestingly, KU treatment resulted in induction of Extracellular signal-regulated kinase(ERK) and CREB phosphorylation. Inhibition of ERK signalling by using MEK inhibitor U0126 disrupted KU dependent induction of mitochondrial biogenesis demonstrating the involvement of ERK signalling cascade in mitochondrial biogenesis upon ATM or AMPK inhibition. These results demonstrate that ATM maintains cellular homeostasis by regulating AMPK mediated mitochondrial biogenesis and that perturbation in this pathway by KU treatment causes a switch in mitochondrial biogenesis in ERK dependent manner. This study indicates cancer cell's fail-safe mechanism in maintaining its homeostasis by up-regulating mitochondrial biogenesis upon inhibition of ATM. Citation Format: Hilal S. Khalil, Hemanth Tummala, Laura DeCaris, Nikolai Zhelev. Pharmacological inhibition of ATM results in mitochondrial biogenesis in AMPK independent manner. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1697. doi:10.1158/1538-7445.AM2013-1697

Ethics Approval This study was approved by C.E.I.M. reference 20/1615 (E.C.P.S.) (Spain); CUHK-NTEC CREC 2021.294-T (Hong Kong); CMO Arnhem-Nijmegen Region NL-number: NL76406.091.21 (Netherlands). Consent The patient was identified as an...

DAG-Lactone Radiosensitization of Human Prostate Cancer Cells Is Mediated by ATM Down-regulation But Not Due to Abnormal DNA Repair

Until recently, research on transplantation rejection and tolerance has been directed toward deciphering the mechanisms of the adaptive immune system. However, the emergence that the innate immune system, the body's first-line defense against pathogens, has a strong influence on adaptive immunity has galvanized interest in elucidating the interplay between these two arms of the immune system. The discovery of Toll-like receptors and the characterization of the cellular mediators involved in innate immunity have provided growing evidence that innate immunity affects the adaptive immune response. Emerging evidence has also shown that early "danger signals"' associated with ischemia-reperfusion injury or brain death contribute to innate immune activation, promoting rejection, and inhibiting tolerance induction. In addition, nonspecific stimuli such as increased donor age or patient disease may also serve to exert a synergistic influence on innate immune activation. Ultimately, controlling the events in innate immune activation may help drive tolerance induction and reduce the rate of rejection.

Pathogenesis of Liver Injury in Acute Liver Failure

During the last few years, mitochondrial DNA has attained much attention as a modulator of immune responses. Due to common evolutionary origin, mitochondrial DNA shares various characteristic features with DNA of bacteria, as it consists of a remarkable number of unmethylated DNA as 2′-deoxyribose cytidine-phosphate-guanosine (CpG) islands. Due to this particular feature, mitochondrial DNA seems to be recognized as a pathogen-associated molecular pattern by the innate immune system. Under the normal physiological situation, mitochondrial DNA is enclosed in the double membrane structure of mitochondria. However, upon pathological conditions, it is usually released into the cytoplasm. Growing evidence suggests that this cytosolic mitochondrial DNA induces various innate immune signaling pathways involving NLRP3, toll-like receptor 9, and stimulator of interferon genes (STING) signaling, which participate in triggering downstream cascade and stimulating to produce effector molecules. Mitochondrial DNA is responsible for inflammatory diseases after stress and cellular damage. In addition, it is also involved in the anti-viral and anti-bacterial innate immunity. Thus, instead of entire mitochondrial importance in cellular metabolism and energy production, mitochondrial DNA seems to be essential in triggering innate anti-microbial immunity. Here, we describe existing knowledge on the involvement of mitochondrial DNA in the anti-microbial immunity by modulating the various immune signaling pathways.

ATM Signaling Facilitates Repair of DNA Double-Strand Breaks Associated with Heterochromatin

Leishmania parasites are the causative agents of a group of neglected tropical diseases known as leishmaniasis. The molecular mechanisms employed by these parasites to adapt to the adverse conditions found in their hosts are not yet completely understood. DNA repair pathways can be used by Leishmania to enable survival in the interior of macrophages, where the parasite is constantly exposed to oxygen reactive species. In higher eukaryotes, DNA repair pathways are coordinated by the central protein kinases ataxia telangiectasia mutated (ATM) and ataxia telangiectasia and Rad3 related (ATR). The enzyme Exonuclease-1 (EXO1) plays important roles in DNA replication, repair, and recombination, and it can be regulated by ATM- and ATR-mediated signaling pathways. In this study, the DNA damage response pathways in promastigote forms of L. major were investigated using bioinformatics tools, exposure of lineages to oxidizing agents and radiation damage, treatment of cells with ATM and ATR inhibitors, and flow cytometry analysis. We demonstrated high structural and important residue conservation for the catalytic activity of the putative LmjEXO1. The overexpression of putative LmjEXO1 made L. major cells more susceptible to genotoxic damage, most likely due to the nuclease activity of this enzyme and the occurrence of hyper-resection of DNA strands. These cells could be rescued by the addition of caffeine or a selective ATM inhibitor. In contrast, ATR-specific inhibition made the control cells more susceptible to oxidative damage in an LmjEXO1 overexpression-like manner. We demonstrated that ATR-specific inhibition results in the formation of extended single-stranded DNA, most likely due to EXO1 nucleasic activity. Antagonistically, ATM inhibition prevented single-strand DNA formation, which could explain the survival phenotype of lineages overexpressing LmjEXO1. These results suggest that an ATM homolog in Leishmania could act to promote end resection by putative LmjEXO1, and an ATR homologue could prevent hyper-resection, ensuring adequate repair of the parasite DNA.