Changes In Cellular Metabolism Research Articles

Small Cell Lung Cancer-An Update on Chemotherapy Resistance.

Compared to other types of lung cancer, small cell lung cancer (SCLC) exhibits aggressive characteristics that promote drug resistance. Despite platinum-etoposide chemotherapy combined with immunotherapy being the current standard treatment, the rapid development of drug resistance has led to unsatisfactory clinical outcomes. This review focuses on the mechanisms contributing to the chemotherapy resistance phenotype in SCLC, such as increased intra-tumoral heterogeneity, alterations in the tumor microenvironment, changes in cellular metabolism, and dysregulation of apoptotic pathways. A comprehensive understanding of these drug resistance mechanisms in SCLC is imperative for ushering in a new era in cancer research, which will promise revolutionary advancements in cancer diagnosis and treatment methodologies.

Upregulation of peroxisome proliferator-activated receptor γ with resorcinol alleviates reactive oxygen species generation and lipid accumulation in neuropathic lysosomal storage diseases

Neuropathic lysosomal storage diseases (NLSDs), including ceroid lipofuscinosis neuronal 3 (CLN3) disease and Gaucher disease type 2 (GD2), are typically present in adolescents; however, there are no approved therapies. CLN3 disease is the most common of the 13 types of neuronal ceroid lipofuscinosis, and Gaucher disease is the most common type of lysosomal storage disease. These NLSDs share oxidative stress and lysosomal dysfunction with Parkinson’s disease. In this study, we used patient-derived cells (PDCs) and resorcinol to develop a therapeutic agent based on peroxisome proliferator-activated receptor γ (PPARγ) activation. PPARγ is a major regulator of autophagy and reactive oxygen species (ROS). Resorcinol, a polyphenolic compound, has been reported to exhibit PPARγ agonistic potential. Protein levels were analyzed by immunoblotting and immunofluorescence microscopy. Changes in cellular metabolism, including ROS levels, lipid droplet content, and lysosomal activity, were measured by flow cytometry. Resorcinol reduced ROS levels by suppressing hypoxia-inducible factor 1α levels in CLN3-PDCs. Resorcinol upregulated autophagy and reduced lipid accumulation in CLN3-PDCs; however, these effects were abolished by autophagy inhibitors. Resorcinol increased nuclear PPARγ levels in CLN3-PDCs, and PPARγ antagonists abolished the therapeutic effects of resorcinol. Moreover, Resorcinol upregulated nuclear PPARγ levels and lysosomal activity in GD2-PDCs, and reduced lipid accumulation and ROS levels. In summary, resorcinol alleviated the shared pathogenesis of CLN3 disease and GD2 through PPARγ upregulation. These findings suggest that resorcinol is a potential therapeutic candidate for alleviating NLSD progression.

Impact of the redox-active MnTnHex-2-PyP5+ and cisplatin on the metabolome of non-small cell lung cancer cells

Redox-based cancer therapeutic strategies aim to raise reactive oxygen species (ROS) levels in cancer cells, thus modifying their redox status, and eventually inducing cell death. Promising compounds, known as superoxide dismutase mimics (SODm), e.g. MnTnHex-2-Py5+ (MnTnHex), could increase intracellular H2O2 in cancer cells with deficient ROS removal systems and therefore enhance radio- and chemotherapy efficacy. We have previously shown that MnTnHex was cytotoxic either alone or combined with cisplatin to non-small cell lung cancer (NSCLC) cells. To gain a deeper understanding of the effects and safety of this compound, it is crucial to analyze the metabolic alterations that take place within the cell. Our goal was thus to study the intracellular metabolome (intracellular metabolites) of NSCLC cells (A549 and H1975) using nuclear magnetic resonance (NMR) spectroscopy-based metabolomics to evaluate the changes in cellular metabolism upon exposure to MnTnHex per se or in combination with cisplatin. 1H NMR metabolomics revealed a higher number of significantly altered metabolites in A549 cells exposed to MnTnHex alone or combined with cisplatin in comparison with non-treated cells (nine dysregulated metabolites), suggesting an impact on aminoacyl-tRNA biosynthesis, glycolysis/gluconeogenesis, taurine, hypotaurine, glycerophospholipid, pyruvate, arginine and proline metabolisms. Regarding H1975 cells, significant alterations in the levels of six metabolites were observed upon co-treatment with MnTnHex and cisplatin, suggesting dysregulations in aminoacyl-tRNA biosynthesis, arginine and proline metabolism, pyruvate metabolism, and glycolysis/gluconeogenesis. These findings help us to understand the impact of MnTnHex on NSCLC cells. Importantly, specific altered metabolites, such as taurine, may contribute to the chemosensitizing effects of MnTnHex.

Exploring the targetome of IsrR, an iron-regulated sRNA controlling the synthesis of iron-containing proteins in Staphylococcus aureus.

Staphylococcus aureus is a common colonizer of the skin and nares of healthy individuals, but also a major cause of severe human infections. During interaction with the host, pathogenic bacteria must adapt to a variety of adverse conditions including nutrient deprivation. In particular, they encounter severe iron limitation in the mammalian host through iron sequestration by haptoglobin and iron-binding proteins, a phenomenon called "nutritional immunity." In most bacteria, including S. aureus, the ferric uptake regulator (Fur) is the key regulator of iron homeostasis, which primarily acts as a transcriptional repressor of genes encoding iron acquisition systems. Moreover, Fur can control the expression of trans-acting small regulatory RNAs that play an important role in the cellular iron-sparing response involving major changes in cellular metabolism under iron-limiting conditions. In S. aureus, the sRNA IsrR is controlled by Fur, and most of its predicted targets are iron-containing proteins and other proteins related to iron metabolism and iron-dependent pathways. To characterize the IsrR targetome on a genome-wide scale, we combined proteomics-based identification of potential IsrR targets using S. aureus strains either lacking or constitutively expressing IsrR with an in silico target prediction approach, thereby suggesting 21 IsrR targets, of which 19 were negatively affected by IsrR based on the observed protein patterns. These included several Fe-S cluster- and heme-containing proteins, such as TCA cycle enzymes and catalase encoded by katA. IsrR affects multiple metabolic pathways connected to the TCA cycle as well as the oxidative stress response of S. aureus and links the iron limitation response to metabolic remodeling. In contrast to the majority of target mRNAs, the IsrR-katA mRNA interaction is predicted upstream of the ribosome binding site, and further experiments including mRNA half-life measurements demonstrated that IsrR, in addition to inhibiting translation initiation, can downregulate target protein levels by affecting mRNA stability.

Development of an organ-on-chip model for the detection of volatile organic compounds as potential biomarkers of tumour progression

Early detection of tumours remains a significant challenge due to their invasive nature and the limitations of current monitoring techniques. Liquid biopsies have emerged as a minimally invasive diagnostic approach, wherein volatile organic compounds (VOCs) show potential as compelling candidates. However, distinguishing tumour-specific VOCs is difficult due to the presence of gases from non-tumour tissues and environmental factors. Therefore, it is essential to develop preclinical models that accurately mimic the intricate tumour microenvironment to induce cellular metabolic changes and secretion of tumour-associated VOCs. In this study, a microfluidic device was used to recreate the ischaemic environment within solid tumours for the detection of tumour-derived VOCs. The system represents a significant advance in understanding the role of VOCs as biomarkers for early tumour detection and holds the potential to improve patient prognosis; particularly for inaccessible and rapidly progressing tumours such as glioblastoma.

Exposure to Various Degrees and Durations of Hypobaric Hypoxia Causes a Reduction in Body Weight of Female Adult Rats

Abstract Background: Hypobaric hypoxia refers to a condition where there is a decreased oxygen partial pressure in the air due to low atmospheric pressure. It is known to affect the metabolism, leading to increased basal metabolic rate, alterations in appetite, and changes in cellular metabolism and energy homeostasis. The effects of hypoxia on metabolism and weight loss are influenced by genetic factors, gender, and the duration and severity of exposure to hypoxia. Currently, there are no reports which elucidate the impact of hypobaric hypoxia on female laboratory rats. Objective: The aim of this study was to observe the effect of varying degrees and durations of hypobaric hypoxia on the body weight of female rats. Materials and Methods: In this study, the body weight of 36 laboratory rats divided into six groups was taken at day 0, and then, the rats were exposed to hypobaric hypoxia in a specially designed hypoxia chamber and their body weights were recorded after 5 days and 10 days of hypoxia exposure. The change in body weight at 5 days and 10 days was compared to that of their body weight before the exposure to hypoxia. Data analysis was performed using IBM SPSS version 20. Results: Body weight was reduced in all rats subjected to varying degrees and duration of hypoxia. The percentage change in body weight was higher in moderate and severe hypoxia than in the mild hypoxia group. No significant difference was observed in rats exposed to varying degrees of hypoxia for 5 days as compared to those exposed for 10 days. Conclusion: Hypoxia may cause a reduction in body weight of female rats proportionate to the increasing severity of hypoxia and this reduction remains independent of the duration of exposure to hypoxia.

PPARG in osteocytes controls cell bioenergetics and systemic energy metabolism independently of sclerostin levels in circulation

ObjectiveThe skeleton is one of the largest organs in the body, wherein metabolism is integrated with systemic energy metabolism. However, the bioenergetic programming of osteocytes, the most abundant bone cells coordinating bone metabolism, is not well defined. Here, using a mouse model with partial penetration of an osteocyte-specific PPARG deletion, we demonstrate that PPARG controls osteocyte bioenergetics and their contribution to systemic energy metabolism independently of circulating sclerostin levels, which were previously correlated with metabolic status of extramedullary fat depots. MethodsIn vivo and in vitro models of osteocyte-specific PPARG deletion, i.e. Dmp1CrePparγflfl male and female mice (γOTKO) and MLO-Y4 osteocyte-like cells with either siRNA-silenced or CRISPR/Cas9-edited Pparγ. As applicable, the models were analyzed for levels of energy metabolism, glucose metabolism, and metabolic profile of extramedullary adipose tissue, as well as the osteocyte transcriptome, mitochondrial function, bioenergetics, insulin signaling, and oxidative stress. ResultsCirculating sclerostin levels of γOTKO male and female mice were not different from control mice. Male γOTKO mice exhibited a high energy phenotype characterized by increased respiration, heat production, locomotion and food intake. This high energy phenotype in males did not correlate with “beiging” of peripheral adipose depots. However, both sexes showed a trend for reduced fat mass and apparent insulin resistance without changes in glucose tolerance, which correlated with decreased osteocytic responsiveness to insulin measured by AKT activation. The transcriptome of osteocytes isolated from γOTKO males suggested profound changes in cellular metabolism, fuel transport, mitochondria dysfunction, insulin signaling and increased oxidative stress. In MLO-Y4 osteocytes, PPARG deficiency correlated with highly active mitochondria, increased ATP production, and accumulation of reactive oxygen species (ROS). ConclusionsPPARG in male osteocytes acts as a molecular break on mitochondrial function, and protection against oxidative stress and ROS accumulation. It also regulates osteocyte insulin signaling and fuel usage to produce energy. These data provide insight into the connection between osteocyte bioenergetics and their sex-specific contribution to the balance of systemic energy metabolism. These findings support the concept that the skeleton controls systemic energy expenditure via osteocyte metabolism.

Untargeted metabolic profiling of high-dose methotrexate toxicity shows alteration in betaine metabolism

Cardiotoxicity is a well-established adverse effect of several drugs across multiple therapeutic indications. It is particularly prevalent following anticancer therapy. In order to evaluate the changes in cellular metabolism associated with methotrexate cardiotoxicity, we treated Wistar rats with a single high dose of methotrexate (HDMTX), and after five days, the animals were sacrificed. We then analyzed the cardiotoxicity parameters in serum like Cardiac enzymes(CK-MB, Troponin T, ALP), Inflammatory markers (TNF-α and IL-6), oxidative stress markers (NO, NOX-2), histopathology and cardiac tissue with the goal of identifying a metabolic signature of cardiotoxicity using discovery-based metabolomics. The biochemical parameters for cardiac enzymes, oxidative stress and inflammatory markers showed a significant increase in all three categories in rats treated with HDMTX. These findings were mirrored in the histopathological analysis confirming cardiotoxicity due to HDMTX. The results showed a total of 95 metabolites that were found to be significantly (p < 0.05) modulated: either up- or downregulated in the HDMTX-treated group when compared with the control group. Using integrated pathway analysis we found these metabolites were associated with many important cardiac tissue metabolic pathways, such as the malate aspartate shuttle, taurine and hypotaurine metabolism, betaine metabolism, spermidine biosynthesis, and homocysteine degradation. Among them, L-arginine, homocysteine, and betaine were significantly upregulated, suggesting their possible association with cardiac tissue injury. Overall, we provided evidence for using untargeted metabolomics to identify novel metabolites associated with HDMTX cardiac toxicity.

Cellular metabolism and hypoxia interfacing with allergic diseases.

Allergic diseases display significant heterogeneity in their pathogenesis. Understanding the influencing factors, pathogenesis, and advancing new treatments for allergic diseases is becoming more and more vital as currently, prevalence continues to rise, and mechanisms of allergic diseases are not fully understood. The upregulation of the hypoxia response is linked to an elevated infiltration of activated inflammatory cells, accompanied by elevated metabolic requirements. An enhanced hypoxia response may potentially contribute to inflammation, remodeling, and the onset of allergic diseases. It has become increasingly clear that the process underlying immune and stromal cell activation during allergic sensitization requires well-tuned and dynamic changes in cellular metabolism. The purpose of this review is to examine current perspectives regarding metabolic dysfunction in allergic diseases. In the past decade, new technological platforms such as "omic" techniques have been applied, allowing for the identification of different biomarkers in multiple models ranging from altered lipid species content, increased nutrient transporters, and altered serum amino acids in various allergic diseases. Better understanding, recognition, and integration of these alterations would increase our knowledge of pathogenesis and potentially actuate a novel repertoire of targeted treatment approaches that regulate immune metabolic pathways.

Targeting the Hippo pathway to prevent radioresistance brain metastases from the lung (Review).

The prognosis for patients with non‑small cell lung cancer (NSCLC), a cancer type which represents 85% of all lung cancers, is poor with a 5‑year survival rate of 19%, mainly because NSCLC is diagnosed at an advanced and metastatic stage. Despite recent therapeutic advancements, ~50% of patients with NSCLC will develop brain metastases (BMs). Either surgical BM treatment alone for symptomatic patients and patients with single cerebral metastases, or in combination with stereotactic radiotherapy (RT) for patients who are not suitable for surgery or presenting with fewer than four cerebral lesions with a diameter range of 5‑30 mm, or whole‑brain RT for numerous or large BMs can be administered. However, radioresistance (RR) invariably prevents the action of RT. Several mechanisms of RR have been described including hypoxia, cellular stress, presence of cancer stem cells, dysregulation of apoptosis and/or autophagy, dysregulation of the cell cycle, changes in cellular metabolism, epithelial‑to‑mesenchymal transition, overexpression of programmed cell death‑ligand 1 and activation several signaling pathways; however, the role of the Hippo signaling pathway in RR is unclear. Dysregulation of the Hippo pathway in NSCLC confers metastatic properties, and inhibitors targeting this pathway are currently in development. It is therefore essential to evaluate the effect of inhibiting the Hippo pathway, particularly the effector yes‑associated protein‑1, on cerebral metastases originating from lung cancer.

Nucleobase adducts bind MR1 and stimulate MR1-restricted T cells.

MR1T cells are a recently found class of T cells that recognize antigens presented by the major histocompatibility complex-I-related molecule MR1 in the absence of microbial infection. The nature of the self-antigens that stimulate MR1T cells remains unclear, hampering our understanding of their physiological role and therapeutic potential. By combining genetic, pharmacological, and biochemical approaches, we found that carbonyl stress and changes in nucleobase metabolism in target cells promote MR1T cell activation. Stimulatory compounds formed by carbonyl adducts of nucleobases were detected within MR1 molecules produced by tumor cells, and their abundance and antigenicity were enhanced by drugs that induce carbonyl accumulation. Our data reveal carbonyl-nucleobase adducts as MR1T cell antigens. Recognizing cells under carbonyl stress allows MR1T cells to monitor cellular metabolic changes with physiological and therapeutic implications.

Fluorinated hydroxyapatite conditions a favorable osteo-immune microenvironment via triggering metabolic shift from glycolysis to oxidative phosphorylation

BackgroundBiological-derived hydroxyapatite is widely used as a bone substitute for addressing bone defects, but its limited osteoconductive properties necessitate further improvement. The osteo-immunomodulatory properties hold crucial promise in maintaining bone homeostasis, and precise modulation of macrophage polarization is essential in this process. Metabolism serves as a guiding force for immunity, and fluoride modification represents a promising strategy for modulating the osteoimmunological environment by regulating immunometabolism. In this context, we synthesized fluorinated porcine hydroxyapatite (FPHA), and has demonstrated its enhanced biological properties and osteogenic capacity. However, it remains unknown whether and how FPHA affects the immune microenvironment of the bone defects.MethodsFPHA was synthesized and its composition and structural properties were confirmed. Macrophages were cultured with FPHA extract to investigate the effects of FPHA on their polarization and the related osteo-immune microenvironment. Furthermore, total RNA of these macrophages was extracted, and RNA-seq analysis was performed to explore the underlying mechanisms associated with the observed changes in macrophages. The metabolic states were evaluated with a Seahorse analyzer. Additionally, immunohistochemical staining was performed to evaluate the macrophages response after implantation of the novel bone substitutes in critical size calvarial defects in SD rats.ResultsThe incorporation of fluoride ions in FPHA was validated. FPHA promoted macrophage proliferation and enhanced the expression of M2 markers while suppressing the expression of M1 markers. Additionally, FPHA inhibited the expression of inflammatory factors and upregulated the expression of osteogenic factors, thereby enhancing the osteogenic differentiation capacity of the rBMSCs. RNA-seq analysis suggested that the polarization-regulating function of FPHA may be related to changes in cellular metabolism. Further experiments confirmed that FPHA enhanced mitochondrial function and promoted the metabolic shift of macrophages from glycolysis to oxidative phosphorylation. Moreover, in vivo experiments validated the above results in the calvarial defect model in SD rats.ConclusionIn summary, our study reveals that FPHA induces a metabolic shift in macrophages from glycolysis to oxidative phosphorylation. This shift leads to an increased tendency toward M2 polarization in macrophages, consequently creating a favorable osteo-immune microenvironment. These findings provide valuable insights into the impact of incorporating an appropriate concentration of fluoride on immunometabolism and macrophage mitochondrial function, which have important implications for the development of fluoride-modified immunometabolism-based bone regenerative biomaterials and the clinical application of FPHA or other fluoride-containing materials.Graphical . FPHA was successfully prepared through the chemical and thermal process. The immunomodulatory effects of FPHA were investigated through in vitro and in vivo studies, revealing its ability to induce a metabolic shift in macrophages from glycolysis to mitochondrial oxidative phosphorylation (OxPhos). This metabolic remodeling resulted in a notable suppression of M1 macrophage polarization and promotion of M2 macrophage polarization. Furthermore, FPHA was found to enhance osteogenic differentiation and facilitate bone repair. These findings underscore the promising potential of FPHA as a biomaterial for bone regenerative applications, providing valuable insights for the development of bioactive materials with metabolic-immunoregulatory propertiesGraphical

Gestational Long Term High-Altitude Hypoxia and Metabolomic Reprogramming in Uterine Arteries of Adult Sheep

Long term hypoxic stress during pregnancy causes a reduction in uterine blood flow, compromising oxygen and nutrient delivery to the fetus. There are multiple epigenetic factors involved in the structural and functional impairments that underlie the impairments in uterine arterial function. Evidence indicates that oxidative stress plays an important role to the decrements in blood flow following high altitude pregnancy. The purpose of the current study was to test the hypothesis that changes in oxylipin and endocannabinoid signaling are important to the increases in oxidative stress along with changes in glucose and fat metabolism. To further understand the mechanisms associated with disruption of uterine artery blood flow due to high altitude pregnancy, we examined a high altitude pregnant sheep model. Pregnant and non-pregnant sheep were housed either at low (700m) or high altitude (3801 meters) for 110+ days of pregnancy. Uterine arteries were excised from pregnant and non-pregnant sheep and targeted as well as untargeted metabolomic analyses were performed to test the hypothesis that hypoxia increases oxidative stress and reduces oxidative metabolism. Enrichment analysis of the metabolites show that hypoxia impacted several key biochemical pathways. At high altitude there were decreases in Alpha Linolenic Acid and Linoleic Acid Metabolism along with reductions in Pyruvate Metabolism and antioxidants. Further to this, there was an increase in the metabolism of Arachidonic Acid, Glycerolipids, and Methionine, which are linked to inflammation, cell signaling, and cell growth. Fully understanding how increased oxidative stress and inflammatory pathways along with decreased antioxidant capacity and changes in cellular metabolism is tied to malformations in uterine arterial structure and function following high altitude gestation is an active area of investigation. This work was supported by the National Institutes of Health Grants NIH R01HL155295, R01HL149608, R03HD098477, P01HD083132, and U24DK097154. Additional support was provided by the Loma Linda University School of Medicine and West Coast Metabolomics Center. This is the full abstract presented at the American Physiology Summit 2024 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.

Metabolomics analysis of CEF cells infected with avian leukosis virus subgroup J based on UHPLC-QE-MS

Avian leukosis virus subgroup J (ALV-J) is a retrovirus that can cause immunosuppression and tumors in chicken. However, relative pathogenesis is still not clear. At present, metabolomics has shown great potential in the screening of tumor metabolic markers, prognostic evaluation, and drug target design. In this study, we utilize an untargeted metabolomics approach based on ultrahigh-performance liquid chromatography-quadrupole time-of-flight tandem mass spectrometry (UHPLC-QTOF-MS) to analyze the metabolic changes in chicken embryo fibroblast (CEF) cells infected by ALV-J. We found that ALV-J infection significantly altered a wealth of metabolites compared with control group. Additionally, most of the differentially expressed metabolites belonged to lipid metabolism, purine nucleotide metabolism and amino acid metabolism. Among them, the proportion of lipid metabolites account for the highest proportion (around 31%). Results suggest that these changes may be conductive to the formation of virion, thereby promoting the replication of ALV-J. These data provided metabolic evidence and potential biomarkers for the cellular metabolic changes induced by ALV-J, and provided important insight for further understanding the replication needs and pathogenesis of ALV-J.

Abstract 392: MYB regulates a specific subset of hypoxia-induced genes and exhibits altered genomic occupancy under hypoxia

Abstract Hypoxia, an unavoidable microenvironmental stress, greatly influences the tumor cell behavior and therapeutic outcomes. Under low oxygen condition, tumor cells undergo transcriptional reprogramming, largely mediated through hypoxia-inducible factors (HIFs), triggering adaptive response pathways. We recently demonstrated a significant role of MYB in hypoxic survival of pancreatic cancer cells by facilitating adaptive changes in cellular metabolism in collaboration with HIF1α. Here, we employed genome-wide deep sequencing approaches to study the impact of hypoxia on transcriptional reprogramming of pancreatic cancer cells and investigate the role of MYB in the hypoxia-dependent processes. A large number of differentially expressed genes (DEGs) were identified in pancreatic cancer cells subjected to hypoxia (1% O2) and associated with multiple adaptive response pathways. Specifically, we found that pathways related to autophagy, cell migration, epithelial-to-mesenchymal transition, endocytosis, cytoskeleton reorganization were induced, while those associated with RNA processing, ribosome biogenesis, DNA replication and cell cycle were suppressed. To analyze the role of MYB in these adaptive responses, we compared the relative abundance of transcripts in MYB-overexpressing control and MYB knockout (KO) pancreatic cancer cells when cultured under normoxic or hypoxic conditions. We observed that the transcriptional output of MYB was drastically changed under hypoxia with a vast majority of genes being distinctly altered in response to the loss of MYB expression under normoxia and hypoxia. To examine if it resulted from altered genomic occupancy of MYB under normoxia and hypoxia, we compared MYB ChIP-seq peaks under these conditions. We observed a decrease in overall MYB-bound genomic regions under hypoxia despite the fact that we did not observe a decrease in nuclear MYB expression. More interestingly, the genomic occupancy of MYB shifted substantially from regions proximal to transcription start site to distal intergenic or intronic regions under hypoxia. Studies are currently underway to examine the mechanisms underlying the altered MYB genomic occupancy and its impact on adaptive physiological response of pancreatic cancer cells under hypoxia. Together, our findings bring unprecedented insight into the mechanistic role of MYB in hypoxia adaptation and pancreatic cancer pathobiology. Citation Format: Shashi Anand, Mohammad Aslam Khan, Kunwar Somesh Vikramdeo, Seema Singh, Ajay Pratap Singh. MYB regulates a specific subset of hypoxia-induced genes and exhibits altered genomic occupancy under hypoxia [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 392.

Abstract 1274: Investigating PLOD2 as a therapeutic target to overcome metastasis in radiorecurrent prostate cancer

Abstract Prostate cancer (PCa) is the second most common cancer in males, with 1 in 8 men developing the disease in their lifetime. For PCa, a common treatment strategy is external beam radiation therapy to the prostate. However, when cancer recurs (radiorecurrent PCa), it often behaves aggressively by invading into surrounding organs or spreading distantly. Radiorecurrent PCa metastasis is therefore a significant cause of morbidity and mortality that must be overcome to improve survival of advanced PCa patients. Proteomic analysis revealed the procollagen enzyme Lysyl Hydroxylase 2 (PLOD2) to be upregulated in our radiorecurrent, and highly aggressive, conventionally fractionated DU145 (DU145-CF) PCa cell line. Given its established function as a mediator of invasion in various other cancers, we sought to characterize the role of PLOD2 in the aggressive phenotype of our radiorecurrent PCa cells. Bioinformatic analysis of clinical data revealed PLOD2 genomic amplification to be significantly associated with biochemical recurrence in PCa patients. Upon further examination in vitro, it was revealed that PLOD2 knockdown significantly reduces matrigel invasion and migration in our radiorecurrent DU145-CF cell line, in addition to numerous other PCa cell lines, including primary cells we derived directly from PCa patients. Using the in vivo chick Chorioallantoic Membrane (CAM) model, we confirmed that PLOD2 knockdown significantly reduces the ability of DU145-CF cells to extravasate from the CAM vasculature into the surrounding stroma, a critical step of the metastatic cascade. To explore mechanisms of metastatic cellular reprogramming downstream of PLOD2, RNA sequencing of DU145-CF cells was conducted; a total of 681 genes were discovered to be dysregulated by PLOD2 knockdown, with their functional analysis suggesting changes in cellular metabolism and respiration. Finally, since PLOD2 is known to be regulated by hypoxia-induced protein HIF1α, we explored whether PLOD2 expression could be inhibited by the HIF1α inhibitor, PX-478. Treatment with PX-478 reduced both HIF1α and PLOD2 protein expression, and significantly reduced invasion, migration, and in vivo extravasation in DU145-CF cells, thereby indicating its potential as a pharmacological inhibitor of HIF1α-associated PLOD2 in radiorecurrent PCa. Together, our results demonstrate for the first time the role of PLOD2 in radiorecurrent PCa invasiveness, and point towards its potential as a therapeutic target to reduce metastasis and improve survival outcomes in PCa patients. Citation Format: Gavin Frame, Hon Leong, Roni Haas, Xiaoyong Huang, Jessica Wright, Paul C. Boutros, Thomas Kislinger, Stanley K. Liu. Investigating PLOD2 as a therapeutic target to overcome metastasis in radiorecurrent prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 1274.

Metaplastic regeneration in the mouse stomach requires a reactive oxygen species pathway

In pyloric metaplasia, mature gastric chief cells reprogram via an evolutionarily conserved process termed paligenosis to re-enter the cell cycle and become spasmolytic polypeptide-expressing metaplasia (SPEM) cells. Here, we use single-cell RNA sequencing (scRNA-seq) following injury to the murine stomach to analyze mechanisms governing paligenosis at high resolution. Injury causes induced reactive oxygen species (ROS) with coordinated changes in mitochondrial activity and cellular metabolism, requiring the transcriptional mitochondrial regulator Ppargc1a (Pgc1α) and ROS regulator Nf2el2 (Nrf2). Loss of the ROS and mitochondrial control in Ppargc1a-/- mice causes the death of paligenotic cells through ferroptosis. Blocking the cystine transporter SLC7A11(xCT), which is critical in lipid radical detoxification through glutathione peroxidase 4 (GPX4), also increases ferroptosis. Finally, we show that PGC1α-mediated ROS and mitochondrial changes also underlie the paligenosis of pancreatic acinar cells. Altogether, the results detail how metabolic and mitochondrial changes are necessary for injury response, regeneration, and metaplasia in the stomach.

Screening by Q Exactive liquid chromatography/tandem mass spectrometry identified Choline, 25-hydroxyvitamin D2, and SM(d18:0/16:1(9Z) (OH)) as biomarkers for high-grade serous ovarian cancer

High-grade serous ovarian cancer (HGSOC) has a high death rate and poor prognosis. The main causes of poor prognosis are asymptomatic early disease, no effective screening method at present, and advanced disease. Changes in cellular metabolism are characteristic of cancer, and plasma metabolome analysis can be used to identify biomarkers. In this study, we used Q Exactive liquid chromatography tandem mass spectrometry (LC–MS/MS, QE) to compare the differentiation between plasma samples (22 HGSOC samples and 22 normal samples). In total, we detected 124 metabolites, and an orthogonal partial least-squares-discriminant analysis (OPLS-DA) model was useful to distinguish HGSOC patients from healthy controls. Choline, 25-hydroxyvitamin D2, and sphingomyelin (d18:0/16:1(9Z) (OH))/SM(d18:0/16:1(9Z) (OH)) showed significantly differential plasma levels in HGSOC patients under the conditions of variable importance in projection (VIP) > 1, p < 0.05 using Student's t-test, and fold change (FC) ≥ 1.5 or ≤ 0.667. Metabolic pathway analysis can provide valuable information to enhance the understanding of the underlying pathophysiology of HGSOC. In conclusion, the Q Exactive LC/MS/MS method validation-based plasma metabolomics approach may have potential as a convenient screening method for HGSOC and may be a method to monitor tumor recurrence in patients with HGSOC after surgery SignificanceHigh-grade serous ovarian cancer (HGSOC) has a high death rate and poor prognosis. The main causes of poor prognosis are asymptomatic early disease, no effective screening method at present, and advanced disease. Changes in cellular metabolism are characteristic of cancer, and plasma metabolome analysis can be used to identify biomarkers. In this study, we used Q Exactive liquid chromatography tandem mass spectrometry (LC–MS/MS, QE) to compare the differentiation between plasma samples (20 HGSOC samples and 20 normal samples). In total, we detected 124 metabolites, and an orthogonal partial least-squares-discriminant analysis (OPLS-DA) model was useful to distinguish HGSOC patients from healthy controls. Choline, 25-hydroxyvitamin D2, and sphingomyelin (d18:0/16:1(9Z) (OH))/SM(d18:0/16:1(9Z) (OH)) showed significantly differential plasma levels in HGSOC patients under the conditions of variable importance in projection (VIP) > 1, p < 0.05 using Student's t-test, and fold change (FC) ≥ 1.5 or ≤ 0.667. Metabolic pathway analysis can provide valuable information to enhance the understanding of the underlying pathophysiology of HGSOC. In conclusion, the Q Exactive LC/MS/MS method validation-based plasma metabolomics approach may have potential as a convenient screening method for HGSOC and may be a method to monitor tumor recurrence in patients with HGSOC after surgery.

Thermally annealed large-scale gold nanostructure platform for long-term and label-free electrochemical monitoring of cellular metabolism

In recent developments, an electrochemical method has emerged as a promising tool that enables real-time and non-destructive monitoring of cell viability and functionality in vitro. However, the restricted cell cultivation area and poor adhesion properties of the gold nanostructure film impede its potential as a new analytical tool for cellular research. In this study, we present an advanced platform designed for prolonged cell growth, differentiation, and drug screening, while providing electrochemical detection capability. The thermal annealing process, implemented after gold nanostructure deposition, was found to induce structural reformation on the indium tin oxide (ITO) film through Ostwald ripening and coalescence, leading to increased surface coverage and reduced height. The resulting optimized platform, referred to as the thermally annealed large-scale gold nanostructure (TLGN), offers an expanded cell culture area (about 227 mm2) with sustained resistance to gold film detachment, a critical challenge observed in previously reported electrodes, lasting up to 4 weeks under exposure to cell culture medium. Using TLGN, redox signals were successfully detected from a substantial number of human cervical cancer cells (470,000 cells) at days in vitro 5, a cell count that is 10.35 times higher compared to our previous gold nanostructure electrode. The cells were continuously monitored for up to 4 days under treatment with the anticancer drug (CB-839). Furthermore, the TLGN platform demonstrated its effectiveness in electrochemically monitoring the osteogenesis of human mesenchymal stem cells over a differentiation period of 28 days, based on observed changes in cellular metabolism. In summary, the TLGN platform proves invaluable not only for assessing drug accumulation and potential cytotoxicity at the cellular level but also for real-time monitoring of mitochondrial dynamics during cellular differentiation.

Exploring the Role of Surface and Mitochondrial ATP-Sensitive Potassium Channels in Cancer: From Cellular Functions to Therapeutic Potentials.

ATP-sensitive potassium (KATP) channels are found in plasma membranes and mitochondria. These channels are a type of ion channel that is regulated by the intracellular concentration of adenosine triphosphate (ATP) and other nucleotides. In cell membranes, they play a crucial role in linking metabolic activity to electrical activity, especially in tissues like the heart and pancreas. In mitochondria, KATP channels are involved in protecting cells against ischemic damage and regulating mitochondrial function. This review delves into the role of KATP channels in cancer biology, underscoring their critical function. Notably responsive to changes in cellular metabolism, KATP channels link metabolic states to electrical activity, a feature that becomes particularly significant in cancer cells. These cells, characterized by uncontrolled growth, necessitate unique metabolic and signaling pathways, differing fundamentally from normal cells. Our review explores the intricate roles of KATP channels in influencing the metabolic and ionic balance within cancerous cells, detailing their structural and operational mechanisms. We highlight the channels' impact on cancer cell survival, proliferation, and the potential of KATP channels as therapeutic targets in oncology. This includes the challenges in targeting these channels due to their widespread presence in various tissues and the need for personalized treatment strategies. By integrating molecular biology, physiology, and pharmacology perspectives, the review aims to enhance the understanding of cancer as a complex metabolic disease and to open new research and treatment avenues by focusing on KATP channels. This comprehensive approach provides valuable insights into the potential of KATP channels in developing innovative cancer treatments.