Randomized phase II selection trial of FLASH and conventional radiotherapy for patients with localized cutaneous squamous cell carcinoma or basal cell carcinoma: A study protocol.

Background: Cutaneous basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) are the most prevalent skin cancers in western countries. Surgery is the standard of care for these cancers and conventional external radiotherapy (CONV-RT) with conventional dose rate (0.03-0.06 Gy/sec) represents a good alternative when the patients or tumors are not amenable to surgery but routinely generates skin side effects. Low energy electron FLASH radiotherapy (FLASH-RT) is a new form of radiotherapy exploiting the biological advantage of the FLASH effect, which consists in delivering radiation dose in milliseconds instead of minutes in CONV-RT. In pre-clinical studies, when compared to CONV-RT, FLASH-RT induced a robust, reproducible and remarkable sparing of the normal healthy tissues, while the efficacy on tumors was preserved. In this context, we aim to prospectively evaluate FLASH-RT versus CONV-RT with regards to toxicity and oncological outcome in localized cutaneous BCC and SCC. Methods: This is a randomized selection, non-comparative, phase II study of curative FLASH-RT versus CONV-RT in patients with T1-T2 N0 M0 cutaneous BCC and SCC. Patients will be randomly allocated to low energy electron FLASH-RT (dose rate: 220-270 Gy/s) or to CONV-RT arm. Small lesions (T1) will receive a single dose of 22 Gy and large lesions (T2) will receive 30 Gy in 5 fractions of 6 Gy over two weeks.The primary endpoint evaluates safety at 6 weeks after RT through grade ≥ 3 toxicity and efficacy through local control rate at 12 months. Approximately 60 patients in total will be randomized, considering on average 1-2 lesions and a maximum of 3 lesions per patients corresponding to the total of 96 lesions required. FLASH-RT will be performed using the Mobetron® (IntraOp, USA) with high dose rate functionality.LANCE (NCT05724875) is the first randomized trial evaluating FLASH-RT and CONV-RT in a curative setting.

ß2-Adrenoceptor Activation Favor Acquisition of Tumorigenic Properties in Non-Tumorigenic MCF-10A Breast Epithelial Cells.

Noradrenaline and adrenaline, and their cognate receptors, are currently accepted to participate in cancer progression. They may also participate in cancer initiation, although their role in this phase is much less explored. The aim of this work was to study the influence of adrenergic stimulation in several processes related to breast cancer carcinogenesis, using several adrenergic agonists in the MCF-10A non-tumorigenic breast cells. Activation of the ß-adrenoceptors promoted an epithelial phenotype in MCF-10A cells, revealed by an increased expression of the epithelial marker E-cadherin and a decrease in the mesenchymal markers, N-cadherin and vimentin. MCF-10A cell motility and migration were also impaired after the ß-adrenoceptors activation. Concomitant with this effect, ß-adrenoceptors decrease cell protrusions (lamellipodia and filopodia) while increasing cell adhesion. Activation of the ß-adrenoceptors also decreases MCF-10A cell proliferation. When the MCF-10A cells were cultured under low attachment conditions, activation the of ß- (likely ß2) or of α2-adrenoceptors had protective effects against cell death, suggesting a pro-survival role of these adrenoceptors. Overall, our results showed that, in breast cells, adrenoceptor activation (mainly through ß-adrenoceptors) may be a risk factor in breast cancer by inducing some cancer hallmarks, providing a mechanistic explanation for the increase in breast cancer incidences that may be associated with conditions that cause massive adrenergic stimulation, such as stress.

2,4,5-Triaminopyrimidines as blue fluorescent probes for cell viability monitoring: synthesis, photophysical properties, and microscopy applications.

Monitoring cell viability is critical in cell biology, pathology, and drug discovery. Most cell viability assays are cell-destructive, time-consuming, expensive, and/or hazardous. Herein, we present a series of newly synthesized 2,4,5-triaminopyrimidine derivatives able to discriminate between live and dead cells. To our knowledge, these compounds are the first fluorescent nucleobase analogues (FNAs) with cell viability monitoring potential. These new fluorescent molecules are synthesized using highly efficient and cost-effective methods and feature unprecedented photophysical properties (longer absorption and emission wavelengths, environment-sensitive emission, and unprecedented brightness within FNAs). Using a live-dead Saccharomyces cerevisiae cell and theoretical assays, the fluorescent 2,4,5-triaminopyrimidine derivatives were found to specifically accumulate inside dead cells by interacting with dsDNA grooves, thus paving the way for the emergence of novel and safe fluorescent cell viability markers emitting in the blue region. As the majority of commercially available viability dyes emit in the green to red region of the visible spectrum, these novel markers might be useful to meet the needs of blue markers for co-staining combinations.

Synthesis of 6,8-diaminopurines via acid-induced cascade cyclization of 5-aminoimidazole precursors and preliminary anticancer evaluation.

Inspired by the pharmacological interest generated by 6-substituted purine roscovitine for cancer treatment, 5-aminoimidazole-4-carboxamidine precursors containing a cyanamide unit were prepared by condensation of 5-amino-N-cyanoimidazole-4-carbimidoyl cyanides with a wide range of primary amines. When these amidine precursors were combined with acids, a fast cascade cyclization occurred at room temperature, affording new 6,8-diaminopurines with the N-3 and N-6 substituents changed relatively to the original positions they occupied in the amidine and imidazole moieties of precursors. The efficacy and wide scope of this method was well demonstrated by an easy and affordable synthesis of 22 6,8-diaminopurines decorated with a wide diversity of substituents at the N-3 and N-6 positions of the purine ring. Preliminary in silico and in vitro assessments of these 22 compounds were carried out and the results showed that 13 of these tested compounds not only exhibited IC50 values between 1.4 and 7.5 µM against the colorectal cancer cell line HCT116 but also showed better binding energies than known inhibitors in docking studies with different cancer-related target proteins. In addition, good harmonization observed between in silico and in vitro results strengthens and validates this preliminary evaluation, suggesting that these novel entities are good candidates for further studies as new anticancer agents.

Machine discovery of partial differential equations from spatiotemporal data: A sparse Bayesian learning framework.

This study presents a general framework, namely, Sparse Spatiotemporal System Discovery (S3d), for discovering dynamical models given by Partial Differential Equations (PDEs) from spatiotemporal data. S3d is built on the recent development of sparse Bayesian learning, which enforces sparsity in the estimated PDEs. This approach enables a balance between model complexity and fitting error with theoretical guarantees. The proposed framework integrates Bayesian inference and a sparse priori distribution with the sparse regression method. It also introduces a principled iterative re-weighted algorithm to select dominant features in PDEs and solve for the sparse coefficients. We have demonstrated the discovery of the complex Ginzburg-Landau equation from a traveling-wave convection experiment, as well as several other PDEs, including the important cases of Navier-Stokes and sine-Gordon equations, from simulated data.

Challenges in Pharmacological Intervention in Perilipins (PLINs) to Modulate Lipid Droplet Dynamics in Obesity and Cancer.

Perilipins (PLINs) are the most abundant proteins in lipid droplets (LD). These LD-associated proteins are responsible for upgrading LD from inert lipid storage structures to fully functional organelles, fundamentally integrated in the lipid metabolism. There are five distinct perilipins (PLIN1-5), each with specific expression patterns and metabolic activation, but all capable of regulating the activity of lipases on LD. This plurality creates a complex orchestrated mechanism that is directly related to the healthy balance between lipogenesis and lipolysis. Given the essential role of PLINs in the modulation of the lipid metabolism, these proteins can become interesting targets for the treatment of lipid-associated diseases. Since reprogrammed lipid metabolism is a recognized cancer hallmark, and obesity is a known risk factor for cancer and other comorbidities, the modulation of PLINs could either improve existing treatments or create new opportunities for the treatment of these diseases. Even though PLINs have not been, so far, directly considered for pharmacological interventions, there are many established drugs that can modulate PLINs activity. Therefore, the aim of this study is to assess the involvement of PLINs in diseases related to lipid metabolism dysregulation and whether PLINs can be viewed as potential therapeutic targets for cancer and obesity.

Systematic analysis and optimization of early warning signals for critical transitions using distribution data.

Abrupt shifts between alternative regimes occur in complex systems, from cell regulation to brain functions to ecosystems. Several model-free early warning signals (EWS) have been proposed to detect impending transitions, but failure or poor performance in some systems have called for better investigation of their generic applicability. Notably, there are still ongoing debates whether such signals can be successfully extracted from data in particular from biological experiments. In this work, we systematically investigate properties and performance of dynamical EWS in different deteriorating conditions, and we propose an optimized combination to trigger warnings as early as possible, eventually verified on experimental data from microbiological populations. Our results explain discrepancies observed in the literature between warning signs extracted from simulated models and from real data, provide guidance for EWS selection based on desired systems and suggest an optimized composite indicator to alert for impending critical transitions using distribution data.

Involvement of P2Y1, P2Y6, A1 and A2A Receptors in the Purinergic Inhibition of NMDA-Evoked Noradrenaline Release in the Rat Brain Cortex.

In the cerebral cortex, glutamate activates NMDA receptors (NMDARs), localized in noradrenergic neurons, inducing noradrenaline release that may have a permissive effect on glutamatergic transmission, and therefore, on the modulation of long-term plasticity. ATP is co-released with noradrenaline, and with its metabolites (ADP and adenosine) is involved in the purinergic modulation of electrically-evoked noradrenaline release. However, it is not known if noradrenaline release evoked by activation of NMDARs is also under purinergic modulation. The present study aimed to investigate and to characterize the purinergic modulation of noradrenaline release evoked by NMDARs. Stimulation of rat cortical slices with 30 µM NMDA increased noradrenaline release, which was inhibited by ATP upon metabolization into ADP and adenosine and by the selective agonists of A1 and A2A receptors, CPA and CGS2680, respectively. It was also inhibited by UTP and UDP, which are mainly released under pathophysiological situations. Characterization of the effects mediated by these compounds indicated the involvement of P2Y1, P2Y6, A1 and A2A receptors. It is concluded that, in the rat brain cortex, NMDA-evoked noradrenaline release is modulated by several purinergic receptors that may represent a relevant mechanism to regulate the permissive effect of noradrenaline on NMDA-induced neuroplasticity.

Independent Reproduction of the FLASH Effect on the Gastrointestinal Tract: A Multi-Institutional Comparative Study.

FLASH radiation therapy (RT) is a promising new paradigm in radiation oncology. However, a major question that remains is the robustness and reproducibility of the FLASH effect when different irradiators are used on animals or patients with different genetic backgrounds, diets, and microbiomes, all of which can influence the effects of radiation on normal tissues. To address questions of rigor and reproducibility across different centers, we analyzed independent data sets from The University of Texas MD Anderson Cancer Center and from Lausanne University (CHUV). Both centers investigated acute effects after total abdominal irradiation to C57BL/6 animals delivered by the FLASH Mobetron system. The two centers used similar beam parameters but otherwise conducted the studies independently. The FLASH-enabled animal survival and intestinal crypt regeneration after irradiation were comparable between the two centers. These findings, together with previously published data using a converted linear accelerator, show that a robust and reproducible FLASH effect can be induced as long as the same set of irradiation parameters are used.

Potential use of renin-angiotensin-aldosterone system inhibitors to reduce COVID-19 severity.

SARS-CoV-2 infection and its clinical manifestations (COVID-19) quickly evolved to a pandemic and a global public health emergency. The limited effectivity of available treatments aimed at reducing virus replication and the lessons learned from other coronavirus infections (SARS-CoV-1 or NL63) that share the internalization process of SARS-CoV-2, led us to revisit the COVID-19 pathogenesis and potential treatments. Virus protein S binds to the angiotensin-converting enzyme 2 (ACE2) initiating the internalization process. Endosome formation removes ACE2 from the cellular membrane preventing its counter-regulative effect mediated by the metabolism of angiotensin II to angiotensin (1-7). Internalized virus-ACE2 complexes have been identified for these coronaviruses. SARS-CoV-2 presents the highest affinity for ACE2 and produces the most severe symptoms. Assuming ACE2 internalization is the trigger for COVID-19 pathogenesis, accumulation of angiotensin II can be viewed as the potential cause of symptoms. Angiotensin II is a strong vasoconstrictor, but has also important roles in hypertrophy, inflammation, remodeling, and apoptosis. Higher levels of ACE2 in the lungs explain the acute respiratory distress syndrome as primary symptoms. Most of the described findings and clinical manifestations of COVID-19, including increased interleukin levels, endothelial inflammation, hypercoagulability, myocarditis, dysgeusia, inflammatory neuropathies, epileptic seizures and memory disorders can be explained by excessive angiotensin II levels. Several meta-analyses have demonstrated that previous use of angiotensin-converting enzyme inhibitors or angiotensin receptor blockers were associated with better prognosis for COVID-19. Therefore, pragmatic trials to assess the potential therapeutic benefits of renin-angiotensin-aldosterone system inhibitors should be urgently promoted by health authorities to widen the therapeutic options for COVID-19.

Long-term 1,2-dimethylhydrazine triggers pathological remodeling of colon mucosa through repression of sestrin2, nuclear factor (erythroid-derived 2)-like 2, and sirtuin4 stimulating mitochondrial stress and metabolic reprogramming.

1,2-Dimethylhydrazine (DMH) is a plant toxicant that enters the food web through the diet. It is biotransformed into azoxymethane, a colon carcinogen, during the first hepatic passage. In mice, this study assessed the role of glutamate dehydrogenase (GDH), a key glutaminolysis enzyme in DMH-induced colorectal cancer (CRC). Colon samples were taken from mice given 6 or 15 weekly doses of 20 mg/kg DMH and serially sacrificed. Repeated DMH doses induced early aberrant crypt foci that evolved into irreversible adenocarcinomas over 24 weeks, along with an increase in GDH and lactate dehydrogenase activities (+ 122%, + 238%, P < 0.001), indicating a switch to aerobic glycolysis and glutaminolysis. Transcriptional downregulation of the endogenous GDH inhibitor, sirtuin4, and two redox regulators, mitochondrial sestrin2 and nuclear factor (erythroid derivative 2)-like 2 (- 26% and - 22%, P < 0, 05; and - 30%, P < 0.01), exacerbated mitochondrial stress by boosting mitochondrial superoxide dismutase activity (+ 240% (P < 0.001) while depressing catalase activity and GSH levels (- 57% and - 60%, P < 0.001). In vitro, allosteric GDH inhibition by 50 µM epigallocatechin gallate decreased human carcinoma (HCT-116) cells' viability, clonogenicity, and migration (- 43% and - 57%, P < 0.001, 41%, P < 0.05), while stimulating ROS release (+ 57%, P < 0.001). Dimethylfumarate (DMF), a linear electrophile and mitochondrial fumarate analog, rebalanced ROS levels (- 34%, P < 0.05) and improved GDH activity, cell viability, and tumorogenic capacity (+ 20%, 20%, P < 0.001; and 33%, P < 0.05). Thus, the pathological remodeling of colon mucosa is supported by metabolic reprogramming bypassing uncoupled mitochondria. DMF highlights the critical role of electrophile response elements in modulating redox mithormesis and redox homeostasis during CRC.

Activation of ß-Adrenoceptors Promotes Lipid Droplet Accumulation in MCF-7 Breast Cancer Cells via cAMP/PKA/EPAC Pathways.

Physiologically, ß-adrenoceptors are major regulators of lipid metabolism, which may be reflected in alterations in lipid droplet dynamics. ß-adrenoceptors have also been shown to participate in breast cancer carcinogenesis. Since lipid droplets may be seen as a hallmark of cancer, the present study aimed to investigate the role of ß-adrenoceptors in the regulation of lipid droplet dynamics in MCF-7 breast cancer cells. Cells were treated for up to 72 h with adrenaline (an endogenous adrenoceptor agonist), isoprenaline (a non-selective ß-adrenoceptor agonist) and salbutamol (a selective ß2-selective agonist), and their effects on lipid droplets were evaluated using Nile Red staining. Adrenaline or isoprenaline, but not salbutamol, caused a lipid-accumulating phenotype in the MCF-7 cells. These effects were significantly reduced by selective ß1- and ß3-antagonists (10 nM atenolol and 100 nM L-748,337, respectively), indicating a dependence on both ß1- and ß3-adrenoceptors. These effects were dependent on the cAMP signalling pathway, involving both protein kinase A (PKA) and cAMP-dependent guanine-nucleotide-exchange (EPAC) proteins: treatment with cAMP-elevating agents (forskolin or 8-Br-cAMP) induced lipid droplet accumulation, whereas either 1 µM H-89 or 1 µM ESI-09 (PKA or EPAC inhibitors, respectively) abrogated this effect. Taken together, the present results demonstrate the existence of a ß-adrenoceptor-mediated regulation of lipid droplet dynamics in breast cancer cells, likely involving ß1- and ß3-adrenoceptors, revealing a new mechanism by which adrenergic stimulation may influence cancer cell metabolism.

DBSegment: Fast and robust segmentation of deep brain structures considering domain generalization.

Segmenting deep brain structures from magnetic resonance images is important for patient diagnosis, surgical planning, and research. Most current state-of-the-art solutions follow a segmentation-by-registration approach, where subject magnetic resonance imaging (MRIs) are mapped to a template with well-defined segmentations. However, registration-based pipelines are time-consuming, thus, limiting their clinical use. This paper uses deep learning to provide a one-step, robust, and efficient deep brain segmentation solution directly in the native space. The method consists of a preprocessing step to conform all MRI images to the same orientation, followed by a convolutional neural network using the nnU-Net framework. We use a total of 14 datasets from both research and clinical collections. Of these, seven were used for training and validation and seven were retained for testing. We trained the network to segment 30 deep brain structures, as well as a brain mask, using labels generated from a registration-based approach. We evaluated the generalizability of the network by performing a leave-one-dataset-out cross-validation, and independent testing on unseen datasets. Furthermore, we assessed cross-domain transportability by evaluating the results separately on different domains. We achieved an average dice score similarity of 0.89 ± 0.04 on the test datasets when compared to the registration-based gold standard. On our test system, the computation time decreased from 43 min for a reference registration-based pipeline to 1.3 min. Our proposed method is fast, robust, and generalizes with high reliability. It can be extended to the segmentation of other brain structures. It is publicly available on GitHub, and as a pip package for convenient usage.

Skin temperature response to thermal stimulus in patients with hyperhidrosis: A comparative study.

Primary hyperhidrosis (HH), the excessive sweating exceeding physiological demand, has been associated to a complex dysfunction of the autonomic nervous system which may explain the disfunction in sweating but may also cause unrevealed alterations in skin blood flow regulation. In fact, HH patients present a sympathetic over-function with less reflex bradycardia in response to the Valsalva maneuver and higher sympathetic skin responses. We aimed to identify response patterns to room thermal stimulus in HH patients compared to a control group in order to investigate putative differences in blood flow assuming that skin temperature in glabrous (non-hairy) areas reflect the sympathetic tone in arteriovenous anastomoses (AVAs). Infrared thermography images were obtained from a cohort of patients diagnosed with HH, followed at a hospital pediatric surgical department and to a sex- and age-matched control group of patients admitted for other surgical procedures. With the participants in Fowler's position, a set of 3 images were captured simultaneously and 44 regions of interest were analyzed, distributed on the palms of the hands, soles of the feet, axilla, and inner canthus. After an acclimatization period at 20 °C, the room temperature was increased to 24, 28 and 32 °C to obtain similar sets of thermograms. A total of 37 patients with HH and 16 participants in the control group were included in the study. At baseline (20 °C), body core temperature (measured in the inner canthus) was significantly higher in the HH patients compared to the controls (p = 0.019 and p = 0.003 in right and left inner canthi, respectively), without any significant differences in the other thermograms. When room temperature was increased, differences in core temperature disappeared, while differences appeared in axilla and palms of the hands with HH patients presenting significantly lower temperature at the three thermal stimulus stages. Patients with HH presented a lower thermoregulatory response when submitted to room temperature increase, which may reflect a vasomotor sympathetic over-function in AVAs.

Buffering variability in cell regulation motifs close to criticality.

Bistable biological regulatory systems need to cope with stochastic noise to fine tune their function close to bifurcation points. Here, we study stability properties of this regime in generic systems to demonstrate that cooperative interactions buffer system variability, hampering noise-induced regime shifts. Our analysis also shows that, in the considered cooperativity range, impending regime shifts can be generically detected by statistical early warning signals from distributional data. Our generic framework, based on minimal models, can be used to extract robustness and variability properties of more complex models and empirical data close to criticality.

Metagenomics-resolved genomics provides novel insights into chitin turnover, metabolic specialization, and niche partitioning in the octocoral microbiome.

BACKGROUND: The role of bacterial symbionts that populate octocorals (Cnidaria, Octocorallia) is still poorly understood. To shed light on their metabolic capacities, we examined 66 high-quality metagenome-assembled genomes (MAGs) spanning 30 prokaryotic species, retrieved from microbial metagenomes of three octocoral species and seawater. RESULTS: Symbionts of healthy octocorals were affiliated with the taxa Endozoicomonadaceae, Candidatus Thioglobaceae, Metamycoplasmataceae, unclassified Pseudomonadales, Rhodobacteraceae, unclassified Alphaproteobacteria and Ca. Rhabdochlamydiaceae. Phylogenomics inference revealed that the Endozoicomonadaceae symbionts uncovered here represent two species of a novel genus unique to temperate octocorals, here denoted Ca. Gorgonimonas eunicellae and Ca. Gorgonimonas leptogorgiae. Their genomes revealed metabolic capacities to thrive under suboxic conditions and high gene copy numbers of serine-threonine protein kinases, type 3-secretion system, type-4 pili, and ankyrin-repeat proteins, suggesting excellent capabilities to colonize, aggregate, and persist inside their host. Contrarily, MAGs obtained from seawater frequently lacked symbiosis-related genes. All Endozoicomonadaceae symbionts harbored endo-chitinase and chitin-binging protein-encoding genes, indicating that they can hydrolyze the most abundant polysaccharide in the oceans. Other symbionts, including Metamycoplasmataceae and Ca. Thioglobaceae, may assimilate the smaller chitin oligosaccharides resulting from chitin breakdown and engage in chitin deacetylation, respectively, suggesting possibilities for substrate cross-feeding and a role for the coral microbiome in overall chitin turnover. We also observed sharp differences in secondary metabolite production potential between symbiotic lineages. Specific Proteobacteria taxa may specialize in chemical defense and guard other symbionts, including Endozoicomonadaceae, which lack such capacity. CONCLUSION: This is the first study to recover MAGs from dominant symbionts of octocorals, including those of so-far unculturable Endozoicomonadaceae, Ca. Thioglobaceae and Metamycoplasmataceae symbionts. We identify a thus-far unanticipated, global role for Endozoicomonadaceae symbionts of corals in the processing of chitin, the most abundant natural polysaccharide in the oceans and major component of the natural zoo- and phytoplankton feed of octocorals. We conclude that niche partitioning, metabolic specialization, and adaptation to low oxygen conditions among prokaryotic symbionts likely contribute to the plasticity and adaptability of the octocoral holobiont in changing marine environments. These findings bear implications not only for our understanding of symbiotic relationships in the marine realm but also for the functioning of benthic ecosystems at large. Video Abstract.

Rényi Entropy in Statistical Mechanics.

Rényi entropy was originally introduced in the field of information theory as a parametric relaxation of Shannon (in physics, Boltzmann-Gibbs) entropy. This has also fuelled different attempts to generalise statistical mechanics, although mostly skipping the physical arguments behind this entropy and instead tending to introduce it artificially. However, as we will show, modifications to the theory of statistical mechanics are needless to see how Rényi entropy automatically arises as the average rate of change of free energy over an ensemble at different temperatures. Moreover, this notion is extended by considering distributions for isospectral, non-isothermal processes, resulting in relative versions of free energy, in which the Kullback-Leibler divergence or the relative version of Rényi entropy appear within the structure of the corrections to free energy. These generalisations of free energy recover the ordinary thermodynamic potential whenever isothermal processes are considered.