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The RNA binding protein Human Antigen R (HuR) has been identified as a main regulator of the innate immune response and its inhibition can lead to beneficial anti-inflammatory effects. To this aim, we previously synthesized a novel class of small molecules named Tanshinone Mimics (TMs) able to interfere with HuR-RNA binding, and that dampen the LPS-induced immune response. Herein, we present a novel series of TMs, encompassing thiophene 3/TM9 and 4/TM10, furan 5/TM11 and 6/TM12, pyrrole 7b/TM13, and pyrazole 8. The furan-containing 5(TM11) showed the greatest inhibitory effect of the series on HuR-RNA complex formation, as suggested by RNA Electromobility Shift Assay and Time-Resolved FRET. Molecular Dynamics Calculation of HuR - 5/TM11 interaction, quantum mechanics approaches and Surface Plasmon Resonance data, all indicates that, within the novel heteroaryl substituents, the furan ring better recapitulates the chemical features of the RNA bound to HuR. Compound 5/TM11 also showed improved aqueous solubility compared to previously reported TMs. Real-time monitoring of cell growth and flow cytometry analyses showed that 5/TM11 preferentially reduced cell proliferation rather than apoptosis in murine macrophages at immunomodulatory doses. We observed its effects on the innate immune response triggered by lipopolysaccharide (LPS) in macrophages, showing that 5/TM11 significantly reduced the expression of proinflammatory cytokines as Cxcl10 and Il1b., (© 2024. The Author(s).)

The maturation of Arabica coffee fruits is influenced by both endogenous and external factors. The stage of fruit maturation affects the chemical composition of the beans, which in turn impacts the quality of the coffee beverage. During maturation, the fruit peel changes colour from green to red (cherry), signalling the optimal harvest time and suggesting high fruit quality. However, the degree of redness can vary, indicating different levels of maturity. This study aimed to explore the variation in soluble sugar accumulation in relation to the redness of coffee fruit tissues. We classified ripe fruits into six ripeness categories based on the intensity of the red colour of the epicarp, measured using a colourimeter. We analysed total soluble sugar, sucrose, and starch in three parts: coat (exocarp + mesocarp), coat juice (obtained by squeezing the coat), and beans. Our findings reveal that the variation in sugar in the endosperm does not correspond to changes in the coat, suggesting separate regulation of sugar accumulation, particularly sucrose, which is crucial for coffee quality. Our data indicate that there is no transfer of sucrose and reducing sugars from the red coat to the bean.

Today it is widely recognized that the PD-1/PD-L1 axis plays a fundamental role in escaping the immune system in cancers, so that anti-PD-1/PD-L1 antibodies have been evaluated for their antitumor properties in more than 1000 clinical trials. As a result, some of them have entered the market revolutionizing the treatment landscape of specific cancer types. Nonetheless, a new era based on the development of small molecules as anti PD-L1 drugs has begun. There are, however, some limitations to advancing these compounds into clinical stages including the possible difficulty in counteracting the PD-1/PD-L1 interaction in vivo, the discrepancy between the in vitro IC 50 (HTFR assay) and cellular EC 50 (immune checkpoint blockade co-culture assay), and the differences in ligands' affinity between human and murine PD-L1, which can affect their preclinical evaluation. Here, an extensive theoretical study, assisted by MicroScale Thermophoresis binding assays and NMR experiments, was performed to provide an atomistic picture of the binding event of three representative biphenyl-based compounds in both human and murine PD-L1. Structural determinants of the species' specificity were unraveled, providing unprecedented details useful for the design of next generation anti-PD-L1 molecules., Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (© 2023 Published by Elsevier B.V. on behalf of Research Network of Computational and Structural Biotechnology.)

Lipopolysaccharide (LPS) exposure to macrophages induces an inflammatory response, which is regulated at the transcriptional and post-transcriptional levels. HuR (ELAVL1) is an RNA-binding protein that regulates cytokines and chemokines transcripts containing AU/U-rich elements (AREs) and mediates the LPS-induced response. Here, we show that small-molecule tanshinone mimics (TMs) inhibiting HuR-RNA interaction counteract LPS stimulus in macrophages. TMs exist in solution in keto-enolic tautomerism, and molecular dynamic calculations showed the ortho-quinone form inhibiting binding of HuR to mRNA targets. TM activity was lost in vitro by blocking the diphenolic reduced form as a diacetate, but resulted in prodrug-like activity in vivo. RNA and ribonucleoprotein immunoprecipitation sequencing revealed that LPS induces a strong coupling between differentially expressed genes and HuR-bound genes, and TMs reduced such interactions. TMs decreased the association of HuR with genes involved in chemotaxis and immune response, including Cxcl10, Il1b and Cd40, reducing their expression and protein secretion in primary murine bone marrow-derived macrophages and in an LPS-induced peritonitis model. Overall, TMs show anti-inflammatory properties in vivo and suggest HuR as a potential therapeutic target for inflammation-related diseases., Competing Interests: Competing interests The authors declare no competing or financial interests., (© 2023. Published by The Company of Biologists Ltd.)

Background: In the first few months of 2020, Lombardy was the hardest-hit region in Italy for COVID-19 cases. Our Rehabilitation Institute offered a telemedicine service to COVID-19 patients discharged after hospitalization. Methods: Patients transferred from Emergency, Intensive Care, and Pulmonology departments of the principal regional hospital hubs had an average stay in our hospital of 2-3 weeks. On discharge, at home, they underwent a telecare nursing and specialist teleconsultation program for 3 months, including monitoring of vital signs and symptoms. Patients completed the SF-12 questionnaire at the start and end of the program and rated their satisfaction with it. Results: The program involved 130 patients (51%). During the period, there were 14 ± 2 (1,800 in total) telenursing support phone calls per patient made, and 12.5 ± 3.4 oxygen saturation readings per patient (1,631 in total). Persisting symptoms, frequently in combination, were present at the start of the program in 124 (94%) patients. There was a significant reduction of symptoms ( p < 0.0000) after the telecare program. The physical component of SF-12 significantly improved at the end [Δ( t 1- t 0) = 6.7 ± 9.3, p < 0.0001]. On the contrary, the mental component of SF-12 remained unchanged or decreased slightly in patients ≤70 years of age [Δ( t 1- t 0) = -2.7 ± 12.3, ns], while it decreased significantly [Δ( t 1- t 0) = -5.4 ± 12.4, p = 0.0367] in older patients (although remaining mild). Patient satisfaction with the program was very high in all 130 patients. Co nclusions: Our Telehealth and Telecare Service offers an example of rapid scaling and adaptation of an existing program to meet the needs of COVID-19 patients. Our findings indicate that telemedicine can be an integral part of clinical practice if supported by the institution with training and IT support provided to patients, nurses, and clinicians.

The disaccharide trehalose is a well-established autophagy inducer, but its therapeutic application is severely hampered by its low potency and poor pharmacokinetic profile. Thus, we targeted the rational design and synthesis of trehalose-based small molecules and nano objects to overcome such issues. Among several rationally designed trehalose-centered putative autophagy inducers, we coupled trehalose via suitable spacers with known self-assembly inducer squalene to yield two nanolipid-trehalose conjugates. Squalene is known for its propensity, once linked to a bioactive compound, to assemble in aqueous media in controlled conditions, internalizing its payload and forming nanoassemblies with better pharmacokinetics. We assembled squalene conjugates to produce the corresponding nanoassemblies, characterized by a hydrodynamic diameter of 188 and 184 nm and a high stability in aqueous media as demonstrated by the measured Z-potential. Moreover, the nanoassemblies were characterized for their toxicity and capability to induce autophagy in vitro.

The Human antigen R (HuR) protein is an RNA-binding protein, ubiquitously expressed in human tissues, that orchestrates target RNA maturation and processing both in the nucleus and in the cytoplasm. A survey of known modulators of the RNA-HuR interactions is followed by a description of its structure and molecular mechanism of action - RRM domains, interactions with RNA, dimerization, binding modes with naturally occurring and synthetic HuR inhibitors. Then, the review focuses on HuR as a validated molecular target in oncology and briefly describes its role in inflammation. Namely, we show ample evidence for the involvement of HuR in the hallmarks and enabling characteristics of cancer, reporting findings from in vitro and in vivo studies; and we provide abundant experimental proofs of a beneficial role for the inhibition of HuR-mRNA interactions through silencing (CRISPR, siRNA) or pharmacological inhibition (small molecule HuR inhibitors)., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2021 Elsevier B.V. All rights reserved.)

The inhibition of the PD-1/PD-L1 axis by monoclonal antibodies has achieved remarkable success in treating a growing number of cancers. However, a novel class of small organic molecules, with BMS-202 ( 1 ) as the lead, is emerging as direct PD-L1 inhibitors. Herein, we report a series of 2,4,6-tri- and 2,4-disubstituted 1,3,5-triazines, which were synthesized and assayed for their PD-L1 binding by NMR and homogeneous time-resolved fluorescence. Among them, compound 10 demonstrated to strongly bind with the PD-L1 protein and challenged it in a co-culture of PD-L1 expressing cancer cells (PC9 and HCC827 cells) and peripheral blood mononuclear cells enhanced antitumor immune activity of the latter. Compound 10 significantly increased interferon γ release and apoptotic induction of cancer cells, with low cytotoxicity in healthy cells when compared to 1 , thus paving the way for subsequent preclinical optimization and medical applications.

A small set of trehalose-centered putative autophagy inducers was rationally designed and synthesized, with the aim to identify more potent and bioavailable autophagy inducers than free trehalose, and to acquire information about their molecular mechanism of action. Several robust, high yield routes to key trehalose intermediates and small molecule prodrugs (2-5), putative probes (6-10) and inorganic nanovectors (12a - thiol-PEG-triazole-trehalose constructs 11) were successfully executed, and compounds were tested for their autophagy-inducing properties. While small molecules 2-11 showed no pro-autophagic behavior at sub-millimolar concentrations, trehalose-bearing PEG-AuNPs 12a caused measurable autophagy induction at an estimated 40 μM trehalose concentration without any significant toxicity at the same concentration., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Objective: Respicard is a clinical scoring system used during tele-assistance of patients with chronic obstructive pulmonary disease (COPD) in order to assess variations during nurse phone contacts. Strategies to prevent occurrence of unreported COPD exacerbations are needed. We evaluated (1) comparison between administration of Respicard by the nurse during phone calls with self-administration by the patient and (2) patient compliance with Respicard use., Subjects and Methods: From January to June 2011, every week for 6 months, patients with COPD enrolled in a tele-assistance program in Italy were invited to fill in Respicard at home according to their symptoms. Concurrently, on a weekly basis, a nurse telephoned the patients and filled in a similar card. The scoring of both Respicards was compared. At the end of the study, the patient's subjective utility and ease in implementing or discomfort in recording Respicard was assessed., Results: In the 39 patients studied, the mean value of recorded nurse cards was 3.7±2.9, and the mean time spent by nurses to measure the first 52 cards was 7.7±2.7 min. Patient compliance was 78.2±33.4%. There was a significant positive correlation between nurse and patient card scores (R=0.98; p<0.0001). A similar correlation was observed when data from patients with or without exacerbations and with high or poor adherence to the program were considered (p<0.0001). Patients' acceptance of the self-recording was good., Conclusions: In patients with COPD with sufficient self-autonomy and followed up with tele-assistance, Respicard is a reliable system for recording symptoms of COPD and could be a good cost-cutting modality for continuing management of patients.

Purpose: To describe i) nurse's utilisation and associated costs during 4-year tele-assistance in amyotrophic lateral sclerosis (ALS) ii) optimal time for initiating tele-assistance., Method: 73 ALS patients after hospital discharge were followed up on voice by phone calls conducted by a nurse through a clinical card which was tested for feasibility, efficacy and utility. Number of patients simultaneously assisted/month, calls/patient, fixed and variable costs were calculated., Results: Time between disease's diagnosis and the beginning of tele-assistance was 720 ± 971 days. ALS patients' percentage managed by nurses through tele-assistance increased over time by 628%. Tele-assistance integrated care (TAIC) followed up patients with a total of 5073 telephone calls. Steady state activity was reached at the 3rd year employing 5 ± 3 calls/pt/month (time/call 64 ± 44 min). Implementing the card was feasible, useful and effective by operator. Survival from diagnosis of ALS was 1224 ± 1150 days. Survival was related to tracheotomy., Conclusion: TAIC is a feasible tool to manage up to 25 ALS patients/month/nurse and costs about €105.00 per patient per month. Tele-assistance is proposed at 2/3 of time course of the disease.

Objective: To establish a cost-effective telephone-accessed consultation and mechanical in-exsufflation (MI-E) and manually assisted coughing, oximetry feedback program for 39 patients with amyotrophic lateral sclerosis., Design: Rapid access to healthcare consultation and to MI-E was provided to treat episodes of distress as a result of secretion encumbrance not reversed by suctioning and associated with a decrease in oxyhemoglobin saturation (SpO(2)) baseline. Avoided hospitalizations, defined by relief of respiratory distress and return of SpO(2) baseline to >or=95% by continuous ventilator use and assisted coughing, were recorded. Patient satisfaction was queried at 6 mos, and a cost analysis was performed of continuous vs. on demand MI-E use., Results: Thirty-nine patients made a total of 1661 calls in 7.46 +/- 5.8 mos of follow-up. Twenty-seven patients had 66 home care visits by a respiratory therapist for a total time commitment of 89.7 +/- 99.3 min/patient/mo. Twelve patients, all ventilator users, were also brought mechanical in-exsufflators for mechanically assisted coughing for 47 respiratory episodes. Thirty hospitalizations were avoided. Seventy-five percent of the patients were extremely satisfied. Mean monthly cost per patient for on-demand telephone consultation, professional home healthcare visits, and MI-E as deemed necessary was euro403 +/- euro420 or 59% less than for continuous MI-E rental. Hospitalization costs were also spared., Conclusions: An on-demand consult and MI-E access program can avoid hospitalizations for patients with amyotrophic lateral sclerosis with significant cost savings.

Background: The issue of how to optimize the routine use of tele-assistance (TA) in the clinical care remains to be addressed. Skills and costs of human resources represent the major component to care for chronic patients with TA., Methods: We investigated (1) the change in patient population, mortality, and staff utilization/cost during the first 5-year activity of a TA program (24-h availability of a call center, pulse oxygen device, and second medical opinion) dedicated to chronic respiratory failure patients and (2) the staff time dedicated to each new patient admitted to the service., Results: Three-hundred and ninety-six patients (age 64 +/- 17 years; 296 men) were reviewed across 5 years of activity. Patients followed/year increased over time, particularly for amyotrophic lateral sclerosis subjects. Calls/month dramatically increased from 60 to 290, with a 5-year number of calls equal to 12.952. The doctor's time dedicated to TA decreased over time, whereas the nurse's time increased allowing a cost saving of 39% when compared with budgeted salary costs. The number of home ventilated patients did not change, remaining over 78%. The mortality rate increased over time (from 6% to 11%). Both chronic obstructive pulmonary disease (COPD) and No-COPD patients used the TA service more frequently during the winter, and COPD used it also in the summer., Conclusions: A stable TA service dedicated to chronic respiratory failure may be reached after 4 years. Across years, (1) number of patients increased, with COPD and ventilated subjects being the most representative; (2) calls varied during seasons; (3) doctor's workload decreased, saving salary costs; and (4) each new enrolled patient may require 73 and 27 min/month of nurse and doctor, respectively.

Patients with amyotrophic lateral sclerosis (ALS) need a care programme as the disease progresses. We used telemedicine-assisted integrated care (TAIC) in 40 patients with ALS, for a mean duration of 8.6 months (range 1-12). A nurse-tutor played the key role, supported by respiratory physicians, neurologists and psychologists. Each patient used a portable pulse oximeter during the daily telephone contacts to assess clinical/oxygen variations. Patients also completed a satisfaction questionnaire. During the study period, each patient used TAIC at least five times per month. There were 1907 scheduled telephone calls (86% of the total) and 317 unscheduled calls. Of the unscheduled calls, 84% were managed by the nurse-tutor and only 16% of them required specialist intervention. The most common item was the ALS clinical interview (58%), followed by the description of acute symptoms, cough ability and oxygenation. TAIC staff recommended 4 out of 12 emergency hospital admissions (33%) and 77% of the other hospitalizations. Patients and caregivers were extremely satisfied (79%) with the nurse assistance provided and the patients' confidence in handling their disease improved in 71% of the cases. TAIC provides a nurse-centred, home-monitoring programme that can be a useful way of following up ALS patients.

We describe the use of telemedicine in support of weaning from invasive mechanical ventilation on a 63-year-old woman of at home by means of a telepneumology program (TPP). Under telephone assistance of a pulmonologist and a TPP nurse tutor, the pulsed arterial saturimetric (pSaT), heart rate (HR), breathing pattern tracing monitoring transmitted via a home telephone line and the aid of the caregiver, the patient was able to maintain diurnal spontaneous breathing after 24 spontaneous breathing trial (SBT) steps twice daily. The duration of each SBT period progressively increased starting from 30 minutes up to 8 hours. This case report shows that many patients at home on ventilators could possibly be weaned through the use of remote monitoring and call center response, with only family/caregivers on-site.