Biology subjects

Crooks, D. R.

Publications and source records attributed to Crooks, D. R..

6 recordsLinked to original sources

bioRxiv2025

Low Field Magnetic Resonance Imaging to Detect Acute Kidney Injury

Renal oxygenation is essential for maintaining kidney function. Disruptions in oxygen delivery can lead to renal hypoxia, which can exacerbate kidney injury through multiple pathways, including inflammation, oxidative stress, and ischemia-reperfusion injury. Despite the recognized importance of oxygenation in renal pathology, non-invasive and reliable methods for assessing kidney oxygen levels are limited. Current techniques either lack sensitivity or involve invasive procedures, restricting their use in routine monitoring. Therefore, there is a pressing need for innovative approaches to assess renal oxygenation, particularly in kidney injury. This study evaluated Electron Paramagnetic Resonance (EPR)-based oxygen imaging using the paramagnetic tracer Ox071 to assess kidney oxygen levels in mice with cyclophosphamide-induced kidney injury. Urine pO2 was also assessed as a potential surrogate marker. EPR oximetry accurately measured kidney oxygen distribution, revealing a temporary increase in pO2 post-injury. Urine oximetry, however, did not reliably reflect changes in kidney oxygenation. Furthermore, EPR oximetry provided high-resolution spatial mapping of oxygen levels within the kidney, allowing for a detailed understanding of the impact of hypoxia on renal tissue. EPR oximetry is a promising, non-invasive tool for monitoring renal oxygenation, offering high-resolution mapping and longitudinal assessment. Its ability to provide detailed information about oxygen distribution within the kidney makes it a valuable tool for studying the pathophysiology of renal diseases and for developing novel therapeutic strategies. Translational Statement: Quantitative spatially resolved measurement of renal oxygenation has the potential to guide clinical decision making in renal disorders such as acute kidney injury. In this study we demonstrate the utility of electron paramagnetic resonance imaging to provide non-invasive and quantitative high-resolution mapping of kidney oxygen concentrations.

physiology↗

bioRxiv2025

Impaired oxidative phosphorylation drives primary tumor escape and metastasis

Metastasis causes most cancer deaths and reflects transitions from primary tumor escape to seeding and growth at metastatic sites. Epithelial-to-mesenchymal transition (EMT) is important early in metastasis to enable cancer cells to detach from neighboring cells, become migratory, and escape the primary tumor. While different phases of metastasis expose cells to variable nutrient environments and demands, the metabolic requirements and plasticity of each step are uncertain. Here we show that EMT and primary tumor escape are stimulated by disrupted oxidative metabolism. Using Renal Cell Carcinoma (RCC) patient samples, we identified the mitochondrial electron transport inhibitor NDUFA4L2 as upregulated in cells undergoing EMT. Deletion of NDUFA4L2 enhanced oxidative metabolism and prevented EMT and metastasis while NDUFA4L2 overexpression enhanced these processes. Mechanistically, NDUFA4L2 suppressed oxidative phosphorylation and caused citric acid cycle intermediates to accumulate, which modified chromatin accessibility of EMT-related loci to drive primary tumor escape. The effect of impaired mitochondrial metabolism to drive EMT appeared general, as renal cell carcinoma patient tumors driven by fumarate hydratase mutations with disrupted oxidative phosphorylation were highly metastatic and also had robust EMT. These findings highlight the importance of dynamic shifts in metabolism for cell migration and metastasis, with mitochondrial impairment driving early phases of this process. Understanding mitochondrial dynamics may have important implications in both basic and translational efforts to prevent cancer deaths.

cancer biology↗

bioRxiv2024

Pharmacological Ascorbate Induces Transient Hypoxia Sensitizing Pancreatic Ductal Adenocarcinoma to a Hypoxia Activated Prodrug

Hypoxic tumor microenvironments pose a significant challenge in cancer treatment. Hypoxia-activated prodrugs like evofosfamide aim to specifically target and eliminate these resistant cells. However, their effectiveness is often limited by reoxygenation after cell death. We hypothesized that ascorbates pro-oxidant properties could be harnessed to induce transient hypoxia, enhancing the efficacy of evofosfamide by overcoming reoxygenation. To test this hypothesis, we investigated the sensitivity of MIA Paca-2 and A549 cancer cells to ascorbate in vitro and in vivo. Ascorbate induced a cytotoxic effect at 5 mM that could be alleviated by endogenous administration of catalase, suggesting a role for hydrogen peroxide in its cytotoxic mechanism. In vitro, Seahorse experiments indicated generation of hydrogen peroxide consumes oxygen, which is offset at later time points by a reduction in oxygen consumption due to hydrogen peroxides cytotoxic effect. In vivo, photoacoustic imaging showed ascorbate treatment at sublethal levels triggered a complex, multi-phasic response in tumor oxygenation across both cell lines. Initially, ascorbate generated transient hypoxia within minutes through hydrogen peroxide production, via reactions that consume oxygen. This initial hypoxic phase peaked at around 150 seconds and then gradually subsided. However, at longer time scales (approximately 300 seconds) a vasodilation effect triggered by ascorbate resulted in increased blood flow and subsequent reoxygenation. Combining sublethal levels of ascorbate with evofosfamide significantly prolonged tumor doubling time in MIA Paca-2 and A549b xenografts compared to either treatment alone. This improvement, however, was only observed in a subpopulation of tumors, highlighting the complexity of the oxygenation response.

cancer biology↗

bioRxiv2023

Metabolic and imaging phenotypes associated with RB1 loss in castrate resistant prostate cancer

Advanced prostate cancer is treated with androgen receptor (AR) signaling inhibitors, which are initially effective, but most patients eventually develop resistance and progress to castrate-resistant prostate cancer (CRPC). Loss of RB1 in CRPC tumors is correlated with rapid progression and poor patient survival and, in combination with TP53 loss, predisposes patients to the development of transitional neuroendocrine prostate cancer (NEPC). Although progressive CRPC is clinically associated with higher 18FDG-PET SUVmax values, it is unknown whether inactivation of RB1 and/or TP53 is a driver of increased glucose import. Using a cohort of patient-derived xenograft (PDX)-derived CRPC organoids, we found that NEPC could not be conclusively distinguished from adenocarcinoma by 18FDG uptake alone, and PSMA protein levels did not correlate with cancer phenotype or 18FDG uptake. Castration-resistant models showed higher 18FDG uptake, but lower pyruvate-to-lactate conversion compared to their castration-sensitive counterparts. In parallel studies using castration-sensitive prostate cancer models, RB1/TP53 knockdown did not affect 18FDG uptake, but increased basal respiration and glycolytic activity, with combined depletion leading to glucose diversion into glycogenesis. These metabolic changes were reflected in increased lactate dehydrogenase flux detected by 13C-hyperpolarized magnetic resonance spectroscopy upon RB1 loss, but not in 18FDG uptake. The metabolic heterogeneity revealed here suggests that a multimodal molecular imaging approach can improve tumor characterization, potentially leading to a better prognosis in cancer treatment.

cancer biology↗

bioRxiv2023

Rewiring of RNA methylation by the oncometabolite fumarate in renal cell carcinoma

Metabolic reprogramming is a hallmark of cancer that facilitates changes in many adaptive biological processes. Mutations in the tricarboxylic acid (TCA) cycle enzyme fumarate hydratase (FH) lead to fumarate accumulation and cause hereditary leiomyomatosis and renal cell cancer (HLRCC). HLRCC is a rare, inherited disease characterized by the development of non-cancerous smooth muscle tumors of the uterus and skin, and an increased risk of a highly metastatic and aggressive form of kidney cancer. Fumarate has been shown to inhibit 2-oxyglutarate-dependent dioxygenases (2OGDDs) involved in the hydroxylation of HIF1, as well as in DNA and histone demethylation. However, the link between fumarate accumulation and changes in RNA post-transcriptional modifications has not been defined. Here, we determine the consequences of fumarate accumulation on the activity of different members of the 2OGDD family targeting RNA modifications. By evaluating multiple RNA modifications in patient-derived HLRCC cell lines, we show that mutation of FH selectively alters the activity of demethylases acting upon N6-methyladenosine (m6A), while the demethylase acting upon N1-methyladenosine (m1A) and 5-formylcytosine (f5C) in mitochondrial RNA are unaffected. The observation that metabolites modulate specific subsets of RNA-modifying enzymes offers new insights into the intersection between metabolism and the epitranscriptome.

molecular biology↗

bioRxiv2020

Heterogeneous adaptation of cysteine reactivity to a covalent oncometabolite

ABSTRACTMetabolism and signaling intersect in the genetic cancer syndrome hereditary leiomyomatosis and renal cell carcinoma (HLRCC), a disease in which mutation of the TCA cycle enzyme fumarate hydratase (FH) causes hyperaccumulation of fumarate. This electrophilic oncometabolite can alter gene activity at the level of transcription, via reversible inhibition of epigenetic dioxygenases, as well as posttranslationally, via covalent modification of cysteine residues. To better understand how metabolites function as covalent signals, here we report a chemoproteomic analysis of a kidney-derived HLRCC cell line. Building on previous studies, we applied a general reactivity probe to compile a dataset of cysteine residues sensitive to rescue of cellular FH activity. This revealed a broad upregulation of cysteine reactivity upon FH rescue, caused by an approximately equal proportion of transcriptional and posttranslational regulation in the rescue cell line. Gene ontology analysis highlights new targets and pathways potentially modulated by FH mutation. Comparison of the new dataset to literature studies highlights considerable heterogeneity in the adaptive response of cysteine-containing proteins in different models of HLRCC. Our analysis provides a resource for understanding the proteomic adaptation to fumarate accumulation, and a foundation for future efforts to exploit this knowledge for cancer therapy.Competing Interest StatementThe authors have declared no competing interest.View Full Text

cancer biology↗