Synthesis of a [18F]F Estradiol Derivative via Click Chemistry Using an Automated Synthesis Module: In Vitro Evaluation as Potential Radiopharmaceutical for Breast Cancer Imaging.

"Click reactions" are a very useful tool for the selective conjugation of different molecular subunits to produce complex structures in a simple way. In this paper, we present the application of Cu(I)-catalyzed biorthogonal reactions between alkynes and azides to the indirect radiofluorination of an estradiol derivative with potential applications in estrogen receptor imaging. The procedure was fully developed on an automated synthesis platform, and conditions were optimized to achieve the desired product with a reasonable yield without precipitation. Although the biological results were not adequate for a potential radiopharmaceutical, the outcome of this work is valuable since the use of automated platforms is required for the reliable and reproducible preparation of PET radiopharmaceuticals in GMP conditions while limiting the radiation dose rates to the personnel.

Mitofusin 1 silencing decreases the senescent associated secretory phenotype, promotes immune cell recruitment and delays melanoma tumor growth after chemotherapy.

Cellular senescence is a therapy endpoint in melanoma, and the senescence-associated secretory phenotype (SASP) can affect tumor growth and microenvironment, influencing treatment outcomes. Metabolic interventions can modulate the SASP, and mitochondrial energy metabolism supports resistance to therapy in melanoma. In a previous report we showed that senescence, induced by the DNA methylating agent temozolomide, increased the level of fusion proteins mitofusin 1 and 2 in melanoma, and silencing Mfn1 or Mfn2 expression reduced interleukin-6 secretion by senescent cells. Here we expanded these observations evaluating the secretome of senescent melanoma cells using shotgun proteomics, and explored the impact of silencing Mfn1 on the SASP. A significant increase in proteins reported to reduce the immune response towards the tumor was found in the media of senescent cells. The secretion of several of these immunomodulatory proteins was affected by Mfn1 silencing, among them was galectin-9. In agreement, tumors lacking mitofusin 1 responded better to treatment with the methylating agent dacarbazine, tumor size was reduced and a higher immune cell infiltration was detected in the tumor. Our results highlight mitochondrial dynamic proteins as potential pharmacological targets to modulate the SASP in the context of melanoma treatment.

Enhanced Tumor Targeting of Radiolabeled Mouse/Human Chimeric Anti-Tn Antibody in Losartan-Treated Mice Bearing Tn-Expressing Lung Tumors.

Aim: ChiTn, a mouse/human chimeric anti-Tn monoclonal antibody, was radiolabeled with iodine-131 (131I) and technetium-99m (99mTc) to assess its biodistribution and internalization in Tn-expressing (Tn+) and wild-type (Tn-) LL/2 lung cancer cells. Results: Selective accumulation and gradual internalization of ChiTn were observed in Tn+ cells. Biodistribution in mice with both Tn+ or Tn- lung tumors indicated that the uptake of radiolabeled ChiTn within tumors increased over time. Dual-labeling experiments with 99mTc and 131I showed different biodistribution patterns, with 99mTc exhibiting higher values in the liver, spleen, and kidneys, while 131I showed higher uptake in the thyroid and stomach. However, tumor uptake did not significantly differ between Tn+ and Tn- tumors. To improve tumor targeting, Losartan, an antihypertensive drug known to enhance tumor perfusion and drug delivery, was investigated. Biodistribution studies in Losartan-treated mice revealed significantly higher radiolabeled ChiTn uptake in Tn+ tumors. No significant changes were observed in the uptake of the control molecule IgG-HYNIC-99mTc. Conclusions: These findings demonstrate the enhanced tumor targeting of radiolabeled ChiTn in Losartan-treated mice with Tn-expressing lung tumors. They highlight the potential of ChiTn as a theranostic agent for cancer treatment and emphasize the importance of Losartan as an adjunctive treatment to improve tumor perfusion and drug delivery.

Oxidative Stress, Atherogenic Dyslipidemia, and Cardiovascular Risk.

Oxidative stress is the consequence of an overproduction of reactive oxygen species (ROS) that exceeds the antioxidant defense mechanisms. Increased levels of ROS contribute to the development of cardiovascular disorders through oxidative damage to macromolecules, particularly by oxidation of plasma lipoproteins. One of the most prominent features of atherogenic dyslipidemia is plasma accumulation of small dense LDL (sdLDL) particles, characterized by an increased susceptibility to oxidation. Indeed, a considerable and diverse body of evidence from animal models and epidemiological studies was generated supporting oxidative modification of sdLDL particles as the earliest event in atherogenesis. Lipid peroxidation of LDL particles results in the formation of various bioactive species that contribute to the atherosclerotic process through different pathophysiological mechanisms, including foam cell formation, direct detrimental effects, and receptor-mediated activation of pro-inflammatory signaling pathways. In this paper, we will discuss recent data on the pathophysiological role of oxidative stress and atherogenic dyslipidemia and their interplay in the development of atherosclerosis. In addition, a special focus will be placed on the clinical applicability of novel, promising biomarkers of these processes.

Molecular Imaging of Monoamine Oxidase A Expression in Highly Aggressive Prostate Cancer: Synthesis and Preclinical Evaluation of Positron Emission Tomography Tracers.

The role of monoamine oxidase A (MAO-A) in the aggressiveness of prostate cancer (PCa) has been established in recent years. The molecular imaging of MAO-A expression could offer a noninvasive tool for the visualization and quantification of highly aggressive PCa. This study reports the synthesis and preclinical evaluation of 11C- and 18F-labeled MAO-A inhibitors as positron emission tomography (PET) tracers for proof-of-concept studies in animal models of PCa. Good manufacturing practice production and quality control of these radiotracers using an automated platform was achieved. PET imaging was performed in an LNCaP tumor model with high MAO-A expression. The tumor-to-muscle (T/M) uptake ratio of [11C]harmine (4.5 ± 0.5) was significantly higher than that for 2-[18F]fluoroethyl-harmol (2.3 ± 0.7) and [11C]clorgyline (2.0 ± 0.1). A comparable ex vivo biodistribution pattern in all radiotracers was observed. Furthermore, the tumor uptake of [11C]harmine showed a dramatic reduction (T/M = 1) in a PC3 tumor model with limited MAO-A expression, and radioactivity uptake in LNCaP tumors was blocked in the presence of nonradioactive harmine. Our findings suggest that [11C]harmine may serve as an attractive PET probe for the visualization of MAO-A expression in highly aggressive PCa. These radiotracers have the potential for clinical translation and may aid in the development of personalized therapeutic strategies for PCa patients.

Automated prostate multi-regional segmentation in magnetic resonance using fully convolutional neural networks.

OBJECTIVE: Automatic MR imaging segmentation of the prostate provides relevant clinical benefits for prostate cancer evaluation such as calculation of automated PSA density and other critical imaging biomarkers. Further, automated T2-weighted image segmentation of central-transition zone (CZ-TZ), peripheral zone (PZ), and seminal vesicle (SV) can help to evaluate clinically significant cancer following the PI-RADS v2.1 guidelines. Therefore, the main objective of this work was to develop a robust and reproducible CNN-based automatic prostate multi-regional segmentation model using an intercontinental cohort of prostate MRI. METHODS: A heterogeneous database of 243 T2-weighted prostate studies from 7 countries and 10 machines of 3 different vendors, with the CZ-TZ, PZ, and SV regions manually delineated by two experienced radiologists (ground truth), was used to train (n = 123) and test (n = 120) a U-Net-based model with deep supervision using a cyclical learning rate. The performance of the model was evaluated by means of dice similarity coefficient (DSC), among others. Segmentation results with a DSC above 0.7 were considered accurate. RESULTS: The proposed method obtained a DSC of 0.88 ± 0.01, 0.85 ± 0.02, 0.72 ± 0.02, and 0.72 ± 0.02 for the prostate gland, CZ-TZ, PZ, and SV respectively in the 120 studies of the test set when comparing the predicted segmentations with the ground truth. No statistically significant differences were found in the results obtained between manufacturers or continents. CONCLUSION: Prostate multi-regional T2-weighted MR images automatic segmentation can be accurately achieved by U-Net like CNN, generalizable in a highly variable clinical environment with different equipment, acquisition configurations, and population. KEY POINTS: • Deep learning techniques allows the accurate segmentation of the prostate in three different regions on MR T2w images. • Multi-centric database proved the generalization of the CNN model on different institutions across different continents. • CNN models can be used to aid on the diagnosis and follow-up of patients with prostate cancer.

Radiosynthesis and validation of [18 F]fluoroestradiol in a Synthra plus research platform for use in routine clinical practice.

In this practitioner protocol, the optimization of the radiochemical synthesis of [18 F]fluoroestradiol (FES) on the Synthra RNplus research automated platform is described in detail and a quality control (QC) summary of three validation productions is presented. In comparison with published synthesis methods developed on other platforms, the yield was considerably improved (40%-45% ndc). The other important improvement is the reduction of the required concentration of H2 SO4 avoiding the production of high concentrations of acidic vapors that can deteriorate the module. Purification was achieved by solid phase extraction, and the required adaptation of an external heating plate to the module to evaporate the ethanol is also described. The product was obtained with high radiochemical purity and fulfilled all the requirements of current Good Manufacturing Practice (cGMP). The final product is formulated as a sterile, pyrogen-free solution suitable for human injection. To the best of our knowledge, this is the first report of FES production using this type of module.

Oxidative Stress and Inflammation: From Mechanisms to Therapeutic Approaches.

Oxidative stress and inflammation are two phenomena that are directly involved in practically all pathologies and especially in aging [...].

Transcriptomic profile of epileptic children treated with ketogenic therapies.

Background: Ketogenic dietary therapies (KDT) are used as a treatment in childhood epilepsy. However, their mechanism has not yet been established. The main objective of this study was to determine the changes in the transcriptomic profile induced by KDT in children with epilepsy in order to shed light on its possible mechanisms. Methods: Eight children with refractory epilepsy were enrolled in the study. Peripheral blood mononuclear cells were obtained before and after the children were treated with KDT for a minimum of 6 months. RNA was extracted and mRNA and miRNA profiling were performed and analyzed. Results: Our intervention with KDT significantly reduced the seizure number in seven of the eight paediatric patients treated and caused important changes in their gene expression profile. Our study reveals modifications in the transcription of 4630 genes and 230 miRNAs. We found that the genes involved in the protection against epileptic crises were among those mainly changed. These genes collectively encode for ion channels, neurotransmitter receptors, and synapse structural proteins. Conclusions: Together our results explain the possible mechanisms of KDT and reinforce its clinical importance in the treatment of epilepsy.

Lifelong soya consumption in males does not increase lifespan but increases health span under a metabolic stress such as type 2 diabetes mellitus.

Soya consumption can decrease oxidative stress in animal models. Moreover, phytoestrogens such as genistein, present in soya, can mimic some of the beneficial effects of estrogens and are devoid of significant side effects, such as cancer. In this study, we have performed a controlled lifelong study with male OF1 mice that consumed either a soya-free diet or a soya-rich diet. We show that, although we found an increase in the expression and activity of antioxidant enzymes in soya-consuming mice, it did not increase lifespan. We reasoned that the soya diet could not increase lifespan in a very healthy population, but perhaps it could extend health span in stressed animals such as type 2 diabetic Goto Kakizaki (GK) rats. Indeed, this was the case: we found that male GK rats consuming a soya-rich diet developed the disease at a lower rate and, therefore, lived longer than soya-free diet-consuming rats.

Anti-Inflammatory Properties of Diet: Role in Healthy Aging.

Inflammation is a physiological process involved in the defenses of the body and the repair of tissues. It is acutely activated by infections, trauma, toxins, or allergic reactions. However, if it becomes chronic, inflammation can end up stimulating the development of diseases such as cardiovascular disease, autoimmune disease, neurological disease, or cancer. Additionally, during aging, inflammation becomes increasingly more chronic. Furthermore, we found that certain foods, such as saturated fats, have pro-inflammatory activity. Taking this into account, in this review we have discussed different diets with possible anti-inflammatory activity, the commonly ingested components of each diet and their active compounds. In addition, we have proposed some dietary guidelines, as well as a list of compounds present in foods with anti-inflammatory activity, outlining how to combine them to achieve optimal anti-inflammatory effects. Therefore, we can conclude that the compounds in our diet with anti-inflammatory activity could help alleviate the inflammatory processes derived from diseases and unhealthy diets, and thereby promote healthy aging.

Pharmacological Properties of Polyphenols: Bioavailability, Mechanisms of Action, and Biological Effects in In Vitro Studies, Animal Models, and Humans.

Drugs are bioactive compounds originally discovered from chemical structures present in both the plant and animal kingdoms. These have the ability to interact with molecules found in our body, blocking them, activating them, or increasing or decreasing their levels. Their actions have allowed us to cure diseases and improve our state of health, which has led us to increase the longevity of our species. Among the molecules with pharmacological activity produced by plants are the polyphenols. These, due to their molecular structure, as drugs, also have the ability to interact with molecules in our body, presenting various pharmacological properties. In addition, these compounds are found in multiple foods in our diet. In this review, we focused on discussing the bioavailability of these compounds when we ingested them through diet and the specific mechanisms of action of polyphenols, focusing on studies carried out in vitro, in animals and in humans over the last five years. Knowing which foods have these pharmacological activities could allow us to prevent and aid as concomitant treatment against various pathologies.

Moderate Red Wine Consumption Increases the Expression of Longevity-Associated Genes in Controlled Human Populations and Extends Lifespan in Drosophila melanogaster.

The beneficial effects of moderate red wine consumption on cardiovascular health are well known. The composition of red wine includes several compounds, such as the phytoestrogen resveratrol, that exert these beneficial effects, although not all the mechanisms by which they act are known. Our aim was to study the effect of red wine consumption on longevity-related genes in controlled human populations, such as cloistered nuns. We found that the expression of catalase, manganese-superoxide dismutase, Sirt1, and p53 was increased in peripheral blood mononuclear cells after 14 days of moderate red wine consumption. This increase was accompanied by an enhanced metabolic wellness: fatty acids, cholesterol, branched chain amino acids (isoleucine and leucine), ketone bodies (acetoacetate), bacterial co-metabolites (trimethylamine), and cellular antioxidants (taurine) contributed to a change in metabolic profile after moderate red wine consumption by the nuns. No serious unwanted side effects were observed. Finally, we tested the effect of moderate red wine consumption on longevity in a controlled animal population, such as D. melanogaster, and found that it increased average life span by 7%. In conclusion, moderate red wine consumption increases the expression of key longevity-related genes and improves metabolic health in humans and increases longevity in flies.

99mTechnetium- or Cy7-Labeled Fab(Tocilizumab) as Potential Multiple Myeloma Imaging Agents.

BACKGROUND: Multiple Myeloma (MM) is a malignant hematologic disorder and the second most common blood cancer. Interleukin-6 (IL-6) has been identified as a crucial factor for the proliferation and survival of MM cells and the overexpression of IL-6 receptor is being studied as a molecular target for therapeutic and diagnostic use in myelomas and other comorbidities. Tocilizumab is a humanized monoclonal antibody that binds IL-6R. OBJECTIVE: We aim to label and evaluate Fab(Tocilizumab) with 99mTechnetium or Cy7 as potential MM imaging agents. METHODS: IL-6R distribution was analyzed by Laser Confocal Microscopy (LCM) in MM cell lines. Fab(Tocilizumab) was produced by the digestion of Tocilizumab with papain for 24h at 37°C, derivatized with NHS-HYNIC-Tfa and radiolabeled with 99mTc. Radiochemical stability and in vitro cell assays were evaluated. Biodistribution and SPECT/CT were performed. Also, Fab(Tocilizumab) was labeled with Cy7 for in vivo fluorescence imaging up to 72h. RESULTS: LCM analysis demonstrates IL-6R distribution on MM cell lines. Incubation with papain resulted in complete digestion of Tocilizumab and exhibited a good purity and homogeneity. Radiolabeling with 99mTc via NHS-HYNIC-Tfa was found to be fast, easy, reproducible and stable, revealing high radiochemical purity and without interfering with IL-6R recognition. Biodistribution and SPECT/CT studies showed a quick blood clearance and significant kidney and MM engrafted tumor uptake. Cy7-Fab(Tocilizumab) fluorescent imaging allowed MM1S tumor identification up to 72h p.i. CONCLUSION: These new molecular imaging agents could potentially be used in the clinical setting for staging and follow-up of MM through radioactive whole-body IL-6R expression visualization in vivo. The fluorescent version could be used for tissue sample evaluation and to guide surgical excision, if necessary.

Relationship between Diet, Microbiota, and Healthy Aging.

Due to medical advances and lifestyle changes, population life expectancy has increased. For this reason, it is important to achieve healthy aging by reducing the risk factors causing damage and pathologies associated with age. Through nutrition, one of the pillars of health, we are able to modify these factors through modulation of the intestinal microbiota. The Mediterranean and Oriental diets are proof of this, as well as the components present in them, such as fiber and polyphenols. These generate beneficial effects on the body thanks, in part, to their interaction with intestinal bacteria. Likewise, the low consumption of products with high fat content favors the state of the microbiota, contributing to the maintenance of good health.

The Relationship between Diet and Frailty in Aging.

The increase in lifespan in the 20th century entails an increase in the elderly population. This brings a new challenge for society, causing people to have physical and mental limitations caused by age-related diseases, such as frailty. Frailty is clinically characterized by multisystem pathophysiological processes, such as chronic inflammation, immune activation, dysregulation of the musculoskeletal and endocrine systems, oxidative stress, energy imbalances, mitochondrial dysfunction, and sarcopenia. The elderly should consume energy in amounts close to those in what is currently accepted as a balanced diet. However, an increase in protein intake may be recommended for elderly people as long as there is no kidney damage. This increase could help fight the loss of muscle mass associated with age. Additionally, vitamin and mineral intakes are often insufficient in their diets. Therefore, the diet should be adapted not only to their age, but also to the pathologies associated with aging. Through these measures, we can reduce the prevalence of comorbidity and thereby increase health span. Therefore, both physical and nutritional interventions, including functional foods and nutraceuticals, should be taken into account.

BCL-xL, a Mitochondrial Protein Involved in Successful Aging: From C. elegans to Human Centenarians.

B-Cell Lymphoma-extra-large (BCL-xL) is involved in longevity and successful aging, which indicates a role for BCL-xL in cell survival pathway regulation. Beyond its well described role as an inhibitor of apoptosis by preventing cytochrome c release, BCL-xL has also been related, indirectly, to autophagy and senescence pathways. Although in these latter cases, BCL-xL has dual roles, either activating or inhibiting, depending on the cell type and the specific conditions. Taken together, all these findings suggest a precise mechanism of action for BCL-xL, able to regulate the crosstalk between apoptosis, autophagy, and senescence, thus promoting cell survival or cell death. All three pathways can be both beneficial or detrimental depending on the circumstances. Thus, targeting BCL-xL would in turn be a "double-edge sword" and therefore, additional studies are needed to better comprehend this dual and apparently contradictory role of BCL-XL in longevity.

Extracellular vesicles and redox modulation in aging.

Extracellular vesicles (EVs) are nowadays known to be mediators of cell-to-cell communication involved in physiological and pathological processes. The current expectation is their use as specific biomarkers and therapeutic tools due to their inner characteristics. However, several investigations still need to be done before we can use them in the clinic. First, their categorization is still under debate, although an accurate classification of EVs subtypes should be based on physical characteristics, biochemical composition or condition description of the cell of origin. Second, EVs carry lipids, proteins and nucleic acids that can induce epigenetic modifications on target cells. These cargos, as well as EVs biogenesis, shedding and uptake is both ageing and redox sensitive. More specifically, senescence and oxidative stress increase EVs release, and their altered content can trigger antioxidant but also prooxidant responses in target cells thereby modulating the redox status. Further analysis would help to asses EVs role in the development and progression of oxidative stress-related pathologies. In this review we aimed to summarize the current knowledge on EVs and their involvement in redox modulation on age-related pathologies. We also discuss future directions and prospective that could be performed to improve EVs usage as biomarkers or therapeutic tools.

Quantitative comparison between single-photon emission computed tomography and positron emission tomography imaging of lung ventilation with 99mTc-technegas and 68Ga-gallgas in patients with chronic obstructive pulmonary disease: A pilot study.

The aim of this study was quantitative comparison between 68Ga-Gallgas positron emission tomography (PET) and 99mTc-Technegas single photon emission computed tomography (SPECT) for lung ventilation function assessment in patients with moderate-to-severe obstructive pulmonary disease and to identify image-derived texture features correlating to the physiologic parameters. Five patients with moderate-to-severe chronic obstructive pulmonary disease with PET and SPECT lung ventilation scans were selected for this study. Threshold-based segmentations were used to compare ventilated regions between both imaging techniques. Histograms of both scans were compared to reveal main differences in distributions of radiotracers. Volumes of segmentation as well as 50 textural features measured in the pulmonary region were correlated to the forced expiratory volume in 1 s (FEV1) as the relevant physiological variable. A better peripheral distribution of the radiotracer was observed in PET scans for three out of five patients. A segmentation threshold of 27% and 31% for normalized scans, for PET and SPECT respectively, was found optimal for volume correlation with FEV1. A high correlation (Pearson correlation coefficient >0.9) was found between 16 texture features measured from SPECT and 7 features measured from PET and FEV1. Quantitative measurements revealed different tracer distribution in both techniques. These results suggest that tracer distribution patterns may depend on the cause of the pulmonary obstruction. We found several texture features measured from SPECT to correlate to FEV1.

Sex Differences in Age-Associated Type 2 Diabetes in Rats-Role of Estrogens and Oxidative Stress.

Females live longer than males, and the estrogens are one of the reasons for this difference. We reported some years ago that estrogens are able to protect rats against oxidative stress, by inducing antioxidant genes. Type 2 diabetes is an age-associated disease in which oxidative stress is involved, and moreover, some studies show that the prevalence is higher in men than in women, and therefore there are sex-associated differences. Thus, the aim of this study was to evaluate the role of estrogens in protecting against oxidative stress in type 2 diabetic males and females. For this purpose, we used Goto-Kakizaki rats, which develop type 2 diabetes with age. We found that female diabetic rats showed lower glycaemia levels with age than did diabetic males and that estrogens enhanced insulin sensitivity in diabetic females. Moreover, glucose uptake, measured by positron emission tomography, was higher in the female brain, cerebellum, and heart than in those from male diabetic rats. There were also sex-associated differences in the plasma metabolic profile as determined by metabolomics. The metabolic profile was similar between estrogen-replaced and control diabetic rats and different from ovariectomized diabetic rats. Oxidative stress is involved in these differences. We showed that hepatic mitochondria from females produced less hydrogen peroxide levels and exhibited lower xanthine oxidase activity. We also found that hepatic mitochondrial glutathione oxidation and lipid oxidation levels were lower in diabetic females when compared with diabetic males. Ovariectomy induced oxidative stress, and estrogen replacement therapy prevented it. These findings provide evidence for estrogen beneficial effects in type 2 diabetes and should be considered when prescribing estrogen replacement therapy to menopausal women.