Extremely Differentiated T Cell Subsets Contribute to Tissue Deterioration During Aging.

There is a dramatic remodeling of the T cell compartment during aging. The most notorious changes are the reduction of the naive T cell pool and the accumulation of memory-like T cells. Memory-like T cells in older people acquire a phenotype of terminally differentiated cells, lose the expression of costimulatory molecules, and acquire properties of senescent cells. In this review, we focus on the different subsets of age-associated T cells that accumulate during aging. These subsets include extremely cytotoxic T cells with natural killer properties, exhausted T cells with altered cytokine production, and regulatory T cells that gain proinflammatory features. Importantly, all of these subsets lose their lymph node homing capacity and migrate preferentially to nonlymphoid tissues, where they contribute to tissue deterioration and inflammaging.

Glycome dynamics in T and B cell development: basic immunological mechanisms and clinical applications.

Glycans cover the surfaces of all mammalian cells through a process called glycosylation. Nearly all proteins and receptors that integrate the intricate series of co-stimulatory/inhibitory pathways of the immune system are glycosylated. Growing evidence indicates that the development of the immune system at the origins of T and B cell development is tightly regulated by glycosylation. In this opinion, we hypothesize that the glycome composition of developing T and B cells is developmentally regulated. We discuss how glycans play fundamental roles in lymphocyte development and how glycans early define T and B cell functionality in multiple aspects of adaptive immunity. These advances can provide opportunities for the discovery of novel disease factors and more effective candidate treatments for various conditions.

Population Dynamics is a Cancer Driver.

Most tissues are continuously renovated through the division of stem cells and the death of old or damaged cells, which is known as cell turnover rate (CTOR). Despite being in steady state, tissues have different population dynamics and leading to diverse clonality levels. Here, we propose and test that cell population dynamics can be a cancer driver. We employed the evolutionary software esiCancer to show that CTOR, within a range comparable to what is observed in human tissues, can amplify the risk of a mutation due to ancestral selection (ANSEL). In a high CTOR tissue, a mutated ancestral cell is likely to be selected and persist over generations, which leads to a scenario of elevated ANSEL profile, characterized by few niches of large clones, which does not occur in low CTOR. We found that CTOR is significantly associated with the risk of developing cancer, even when correcting for mutation load, indicating that population dynamics per se is a cancer driver. This concept is central to understanding cancer risk and for the design of new therapeutic interventions that minimize the contribution of ANSEL in cancer growth.

Rapid Peptide Cyclization Inspired by the Modular Logic of Nonribosomal Peptide Synthetases.

Nonribosomal cyclic peptides (NRcPs) are structurally complex natural products and a vital pool of therapeutics, particularly antibiotics. Their structural diversity arises from the ability of the multidomain enzyme assembly lines, nonribosomal peptide synthetases (NRPSs), to utilize bespoke nonproteinogenic amino acids, modify the linear peptide during elongation, and catalyze an array of cyclization modes, e.g., head to tail, side chain to tail. The study and drug development of NRcPs are often limited by a lack of easy synthetic access to NRcPs and their analogues, with selective macrolactamization being a major bottleneck. Herein, we report a generally applicable chemical macrocyclization method of unprecedented speed and selectivity. Inspired by biosynthetic cyclization, it combines the deprotected linear biosynthetic precursor peptide sequence with a highly reactive C-terminus to produce NRcPs and analogues in minutes. The method was applied to several NRcPs of varying sequences, ring sizes, and cyclization modes including rufomycin, colistin, and gramicidin S with comparable success. We thus demonstrate that the linear order of modules in NRPS enzymes that determines peptide sequence encodes the key structural information to produce peptides conformationally biased toward macrocyclization. To fully exploit this conformational bias synthetically, a highly reactive C-terminal acyl azide is also required, alongside carefully balanced pH and solvent conditions. This allows for consistent, facile cyclization of exceptional speed, selectivity, and atom efficiency. This exciting macrolactamization method represents a new enabling technology for the biosynthetic study of NRcPs and their development as therapeutics.

Kinetic Resolution of Epimeric Proteins Enables Stereoselective Chemical Mutagenesis.

Chemical mutagenesis via dehydroalanine (Dha) is a powerful method to tailor protein structure and function, allowing the site-specific installation of post-translational modifications and non-natural functional groups. Despite the impressive versatility of this method, applications have been limited, as products are formed as epimeric mixtures, whereby the modified amino acid is present as both the desired l-configuration and a roughly equal amount of the undesired d-isomer. Here, we describe a simple remedy for this issue: removal of the d-isomer via proteolysis using a d-stereoselective peptidase, alkaline d-peptidase (AD-P). We demonstrate that AD-P can selectively cleave the d-isomer of epimeric residues within histone H3, GFP, Ddx4, and SGTA, allowing the installation of non-natural amino acids with stereochemical control. Given the breadth of modifications that can be introduced via Dha and the simplicity of our method, we believe that stereoselective chemoenzymatic mutagenesis will find broad utility in protein engineering and chemical biology applications.

Evolving patterns and clinical outcome of genetic studies performed at diagnosis in acute myeloid leukemia patients: Real life data from the PETHEMA Registry.

BACKGROUND: There are no studies assessing the evolution and patterns of genetic studies performed at diagnosis in acute myeloid leukemia (AML) patients. Such studies could help to identify potential gaps in our present diagnostic practices, especially in the context of increasingly complex procedures and classifications. METHODS: The REALMOL study (NCT05541224) evaluated the evolution, patterns, and clinical impact of performing main genetic and molecular studies performed at diagnosis in 7285 adult AML patients included in the PETHEMA AML registry (NCT02607059) between 2000 and 2021. RESULTS: Screening rates increased for all tests across different time periods (2000-2007, 2008-2016, and 2017-2021) and was the most influential factor for NPM1, FLT3-ITD, and next-generation sequencing (NGS) determinations: NPM1 testing increased from 28.9% to 72.8% and 95.2% (p < .001), whereas FLT3-ITD testing increased from 38.1% to 74.1% and 95.9% (p < .0001). NGS testing was not performed between 2000-2007 and only reached 3.5% in 2008-2016, but significantly increased to 72% in 2017-2021 (p < .001). Treatment decision was the most influential factor to perform karyotype (odds ratio [OR], 6.057; 95% confidence interval [CI], 4.702-7.802), and fluorescence in situ hybridation (OR, 2.273; 95% CI, 1.901-2.719) studies. Patients ≥70 years old or with an Eastern Cooperative Oncology Group ≥2 were less likely to undergo these diagnostic procedures. Performing genetic studies were associated with a favorable impact on overall survival, especially in patients who received intensive chemotherapy. CONCLUSIONS: This unique study provides relevant information about the evolving landscape of genetic and molecular diagnosis for adult AML patients in real-world setting, highlighting the increased complexity of genetic diagnosis over the past 2 decades.

FRET Sensor-Modified Synthetic Hydrogels for Real-Time Monitoring of Cell-Derived Matrix Metalloproteinase Activity using Fluorescence Lifetime Imaging.

Matrix remodeling plays central roles in a range of physiological and pathological processes and is driven predominantly by the activity of matrix metalloproteinases (MMPs), which degrade extracellular matrix (ECM) proteins. Our understanding of how MMPs regulate cell and tissue dynamics is often incomplete as in vivo approaches are lacking and many in vitro strategies cannot provide high-resolution, quantitative measures of enzyme activity in situ within tissue-like 3D microenvironments. Here, we incorporate a Förster resonance energy transfer (FRET) sensor of MMP activity into fully synthetic hydrogels that mimic many properties of the native ECM. We then use fluorescence lifetime imaging to provide a real-time, fluorophore concentration-independent quantification of MMP activity, establishing a highly accurate, readily adaptable platform for studying MMP dynamics in situ. MCF7 human breast cancer cells encapsulated within hydrogels highlight the detection of MMP activity both locally, at the sub-micron level, and within the bulk hydrogel. Our versatile platform may find use in a range of biological studies to explore questions in the dynamics of cancer metastasis, development, and tissue repair by providing high-resolution, quantitative and in situ readouts of local MMP activity within native tissue-like environments.

Outcomes after intensive chemotherapy for secondary and myeloid-related changes acute myeloid leukemia patients aged 60 to 75 years old: a retrospective analysis from the PETHEMA registry.

Treatment options for patients with secondary acute myeloid leukemia (sAML) and AML with myeloid-related changes (AMLMRC) aged 60 to 75 years are scarce and unsuitable. A pivotal trial showed that CPX-351 improved complete remission with/without incomplete recovery (CR/CRi) and overall survival (OS) as compared with standard "3+7" regimens. We retrospectively analyze outcomes of 765 patients with sAML and AML-MRC aged 60 to 75 years treated with intensive chemotherapy, reported to the PETHEMA registry before CPX-351 became available. The CR/CRi rate was 48%, median OS was 7.6 months (95% confidence interval [CI]: 6.7-8.5) and event-free survival (EFS) 2.7 months (95% CI: 2-3.3), without differences between intensive chemotherapy regimens and AML type. Multivariate analyses identified age ≥70 years, Eastern Cooperative Oncology Group performance status ≥1 as independent adverse prognostic factors for CR/CRi and OS, while favorable/intermediate cytogenetic risk and NPM1 were favorable prognostic factors. Patients receiving allogeneic stem cell transplant (HSCT), autologous HSCT, and those who completed more consolidation cycles showed improved OS. This large study suggests that classical intensive chemotherapy could lead to similar CR/CRi rates with slightly shorter median OS than CPX-351.

Electronic effects of conjugated aryl groups on the properties and reactivities of di(arylethynyl)tetracenes.

The photochemical oxidations of acenes can cause challenges with their optoelectronic applications, such as singlet fission and organic transistors. At the same time, these reactions form the basis for many luminescent sensing schemes for 1O2. While diethynyl substitution is arguably the most widely adopted of the various substitution strategies to control oxidation and also improve solubility and processability of long acenes, the extent to which differences between the alkyne groups can influence key properties of long acenes remains largely unknown. This report therefore describes the effects of various arenes and heteroarenes on the electronic structures, optical properites, and reactivity with singlet oxygen for eight 5,12-di(arylethynyl)tetracenes. The fluorescence spectra of these tetracenes span approximately 100 nm, while their observed rate constants for reaction with singlet oxygen correlates strongly with the HOMO level, spanning one order of magnitude. They are also amenable to fluorescent materials that respond ratiometrically to singlet oxygen. Therefore, electronic effects of groups directly conjugated to ethynylacenes offer a useful chemical space for rational acene design.

Simulation of the THF hydrate-water interfacial free energy from computer simulation.

In this work, the tetrahydrofuran (THF) hydrate-water interfacial free energy is determined at 500 bar, at one point of the univariant two-phase coexistence line of the THF hydrate, by molecular dynamics simulation. The mold integration-host methodology, an extension of the original mold integration technique to deal with hydrate-fluid interfaces, is used to calculate the interfacial energy. Water is described using the well-known TIP4P/Ice model, and THF is described using a rigid version of the TraPPE model. We have recently used the combination of these two models to accurately describe the univariant two-phase dissociation line of the THF hydrate in a wide range of pressures from computer simulation [Algaba et al., J. Chem. Phys. 160, 164718 (2024)]. The THF hydrate-water interfacial free energy predicted in this work is compared with the only experimental data available in the literature. The value obtained, 27(2) mJ/m2, is in excellent agreement with the experimental data taken from the literature, 24(8) mJ/m2. To the best of our knowledge, this is the first time that the THF hydrate-water interfacial free energy is predicted from computer simulation. This work confirms that the mold integration technique can be used with confidence to predict the solid-fluid interfaces of complex structures, including hydrates that exhibit sI and sII crystallographic structures.

Modeling oceanic sedimentary methane hydrate growth through molecular dynamics simulation.

The crystallization process of methane hydrates in a confined geometry resembling seabed porous silica sedimentary conditions has been studied using molecular dynamics simulations. With this objective in mind, a fully atomistic quartz silica slit pore has been designed, and the temperature stability of a methane hydrate crystalline seed in the presence of water and guest molecule methane has been analyzed. NaCl ion pairs have been added in different concentrations, simulating salinity conditions up to values higher than average oceanic conditions. The structure obtained when the hydrate crystallizes inside the pore is discussed, paying special attention to the presence of ionic doping inside the hydrate and the subsequent induced structural distortion. The shift in the hydrate stability conditions due to the increasing water salinity is discussed and compared with the case of unconfined hydrate, concluding that the influence of the confinement geometry and pore hydrophilicity produces a larger deviation in the confined hydrate phase equilibria.

Molecular Simulation of SO2 Separation and Storage Using a Cryptophane-Based Porous Liquid.

A theoretical molecular simulation study of the encapsulation of gaseous SO2 at different temperature conditions in a type II porous liquid is presented here. The system is composed of cage cryptophane-111 molecules that are dispersed in dichloromethane, and it is described using an atomistic modelling of molecular dynamics. Gaseous SO2 tended to almost fully occupy cryptophane-111 cavities throughout the simulation. Calculations were performed at 300 K and 283 K, and some insights into the different adsorption found in each case were obtained. Simulations with different system sizes were also studied. An experimental-like approach was also employed by inserting a SO2 bubble in the simulation box. Finally, an evaluation of the radial distribution function of cryptophane-111 and gaseous SO2 was also performed. From the results obtained, the feasibility of a renewable separation and storage method for SO2 using porous liquids is mentioned.

Image perception and reception in wordless picture books: Eye movements of children with intellectual disabilities.

Wordless picture books enhance comprehension and vocabulary growth and motivate children with intellectual disabilities (ID) to participate in literary activities. However, the reception of picture books can be challenging because deliberate selective attention processes and recognition of the image's meaning are often delayed. Examining eye movements may help explore these cognitive processes. Therefore, we examined eye movements in 29 children with mild and moderate ID as they explored a wordless picture book, presented on a screen and compared them to 14 typically developing children using a Tobii Pro X3-120 eye tracker. The findings showed that children with moderate ID had shorter fixation duration, fixated less often, and revisited regions of interest less frequently. Our results suggest that children with moderate ID have greater difficulties in selectively directing their attention toward regions of visual input with a high level of informativeness and expend less cognitive effort to understand their meaning.

Enzymatic Fluoromethylation as a Tool for ATP-Independent Ligation.

S-adenosylmethionine-dependent methyltransferases are involved in countless biological processes, including signal transduction, epigenetics, natural product biosynthesis, and detoxification. Only a handful of carboxylate methyltransferases have evolved to participate in amide bond formation. In this report we show that enzyme-catalyzed F-methylation of carboxylate substrates produces F-methyl esters that readily react with N- or S-nucleophiles under physiological conditions. We demonstrate the applicability of this approach to the synthesis of small amides, hydroxamates, and thioesters, as well as to site-specific protein modification and native chemical ligation.

Glycans as shapers of tumour microenvironment: A sweet driver of T-cell-mediated anti-tumour immune response.

Essentially all cells are covered with a dense coat of different glycan structures/sugar chains, giving rise to the so-called glycocalyx. Changes in cellular glycosylation are a hallmark of cancer, affecting most of the pathophysiological processes associated with malignant transformation, including tumour immune responses. Glycans are chief macromolecules that define T-cell development, differentiation, fate, activation and signalling. Thus, the diversity of glycans expressed at the surface of T cells constitutes a fundamental molecular interface with the microenvironment by regulating the bilateral interactions between T-cells and cancer cells, fine-tuning the anti-tumour immune response. In this review, we will introduce the power of glycans as orchestrators of T-cell-mediated immune response in physiological conditions and in cancer. We discuss how glycans modulate the glyco-metabolic landscape in the tumour microenvironment, and whether glycans can synergize with immunotherapy as a way of rewiring T-cell effector functions against cancer cells.

Altered IgG glycosylation at COVID-19 diagnosis predicts disease severity.

The nature of the immune responses associated with COVID-19 pathogenesis and disease severity, as well as the breadth of vaccine coverage and duration of immunity, is still unclear. Given the unpredictability for developing a severe/complicated disease, there is an urgent need in the field for predictive biomarkers of COVID-19. We have analyzed IgG Fc N-glycan traits of 82 SARS-CoV-2+ unvaccinated patients, at diagnosis, by nano-LC-ESI-MS. We determined the impact of IgG Fc glyco-variations in the induction of NK cells activation, further evaluating the association between IgG Fc N-glycans and disease severity/prognosis. We found that SARS-CoV-2+ individuals display, at diagnosis, variations in the glycans composition of circulating IgGs. Importantly, levels of galactose and sialic acid structures on IgGs are able to predict the development of a poor COVID-19 disease. Mechanistically, we demonstrated that a deficiency on galactose structures on IgG Fc in COVID-19 patients appears to induce NK cells activation associated with increased release of IFN-γ and TNF-α, which indicates the presence of pro-inflammatory immunoglobulins and higher immune activation, associated with a poor disease course. This study brings to light a novel blood biomarker based on IgG Fc glycome composition with capacity to stratify patients at diagnosis.

Cysteine-Selective Modification of Peptides and Proteins via Desulfurative C-C Bond Formation.

The site-selective modification of peptides and proteins facilitates the preparation of targeted therapeutic agents and tools to interrogate biochemical pathways. Among the numerous bioconjugation techniques developed to install groups of interest, those that generate C(sp3 )-C(sp3 ) bonds are significantly underrepresented despite affording proteolytically stable, biogenic linkages. Herein, a visible-light-mediated reaction is described that enables the site-selective modification of peptides and proteins via desulfurative C(sp3 )-C(sp3 ) bond formation. The reaction is rapid and high yielding in peptide systems, with comparable translation to proteins. Using this chemistry, a range of moieties is installed into model systems and an effective PTM-mimic is successfully integrated into a recombinantly expressed histone.

Insight Into Factors Governing Formation, Synthesis and Stereochemical Configuration of DNA Adducts Formed by Mitomycins.

Mitomycin C, (MC), an antitumor drug used in the clinics, is a DNA alkylating agent. Inert in its native form, MC is reduced to reactive mitosenes in cellulo which undergo nucleophilic attack by DNA bases to form monoadducts as well as interstrand crosslinks (ICLs). These properties constitute the molecular basis for the cytotoxic effects of the drug. The mechanism of DNA alkylation by mitomycins has been studied for the past 30 years and, until recently, the consensus was that drugs of the mitomycins family mainly target CpG sequences in DNA. However, that paradigm was recently challenged. Here, we relate the latest research on both MC and dicarbamoylmitomycin C (DMC), a synthetic derivative of MC which has been used to investigate the regioselectivity of mitomycins DNA alkylation as well as the relationship between mitomycins reductive activation pathways and DNA adducts stereochemical configuration. We also review the different synthetic routes to access mitomycins nucleoside adducts and oligonucleotides containing MC/DMC DNA adducts located at a single position. Finally, we briefly describe the DNA structural modifications induced by MC and DMC adducts and how site specifically modified oligonucleotides have been used to elucidate the role each adduct plays in the drugs cytotoxicity.

Visualization of fibroblast activation using 68Ga-FAPI PET/CT after pulmonary vein isolation with pulsed field compared with cryoballoon ablation.

BACKGROUND: Pulsed-field ablation (PFA) is a novel ablation modality for atrial fibrillation (AF) ablating myocardium by electroporation without tissue-heating. With its different mechanism of tissue ablation, it is assumed that lesion creation is divergent to thermal energy sources. 68Ga-fibroblast-activation protein inhibitor (FAPI) PET/CT targets FAP-alpha expressed by activated fibroblasts. We aimed to assess 68Ga-FAPI uptake in pulmonary veins as surrogate for ablation damage after PFA and cryoballoon ablation (CBA). METHODS: 26 patients (15 PFA, 11 CBA) underwent 68Ga-FAPI-PET/CT after ablation. Standardized uptake values (SUV) and fibroblast-activation volumes of localized tracer uptake were assessed. RESULTS: Patient characteristics were comparable between groups. In PFA, focal FAPI uptake was only observed in 3/15 (20%) patients, whereas in the CBA cohort, 10/11 (90.9%) patients showed atrial visual uptake. We observed lower values of SUVmax (2.85 ± 0.56 vs 4.71 ± 2.06, P = 0.025) and FAV (1.13 ± 0.84 cm3 vs 3.91 ± 2.74 cm3, P = 0.014) along with a trend towards lower SUVpeak and SUVmean in PFA vs CBA patients, respectively. CONCLUSION: Tissue response with respect to fibroblast activation seems to be less pronounced in PFA compared to established thermal ablation systems. This functional assessment might contribute to a better understanding of lesion formation in thermal and PFA ablation potentially contributing to better safety outcomes.

Parasite assemblages in volatile host stocks: inter- and intra-cohort variability restrict their value as biological tags for squid stock assessment.

The Argentine shortfin squid, Illex argentinus, inhabits in the southwest Atlantic; it is a semelparous species which grows rapidly along its 1 year lifespan. The identification of its stocks is critical for sustainable fishery exploitation. Parasites have been used as biological indicators in a lower number of studies dealing with squids, therefore a validation of this methodology is necessary. The intra- and inter-cohort variability of parasite assemblages in the summer-spawning stock of I. argentinus was analysed to assess their value as indicators of stock structure. Four squid samples from the continental shelf of central Patagonia, corresponding to 3 consecutive cohorts, were examined for metazoan parasites. Results evidenced heterogeneity in terms of parasite assemblage composition and structure, dominated by short-lived gastrointestinal parasites, with a strong influence of host size, but no effect of squid sex. These changes are related to their recent habitats and diets, which change with ontogeny and migrations, clouding any interpretation of patterns when samples spatially or temporally separated are compared. Many squid species share these characteristics; therefore, it is recommended that the use of parasites as biological tags should be restricted to simultaneous sampling, while size or age must be considered for deriving proper conclusions.