Out-of-Hospital Cardiac Arrest in the Paediatric Patient: An Observational Study in the Context of National Regulations.

Introduction: Cardiac arrest results in a high death rate if cardiopulmonary resuscitation and early defibrillation are not performed. Mortality is strongly linked to regulations, in terms of prevention and emergency-urgency system organization. In Italy, training of lay rescuers and the presence of defibrillators were recently made mandatory in schools. Our analysis aims to analyze Out-of-Hospital Cardiac Arrest (OHCA) events in pediatric patients (under 18 years old), to understand the epidemiology of this phenomenon and provide helpful evidence for policy-making. Methods: A retrospective observational analysis was conducted on the emergency databases of Lombardy Region, considering all pediatric OHCAs managed between 1 January 2016, and 31 December 2019. The demographics of the patients and the logistics of the events were statistically analyzed. Results: The incidence in pediatric subjects is 4.5 (95% CI 3.6-5.6) per 100,000 of the population. School buildings and sports facilities have relatively few events (1.9% and 4.4%, respectively), while 39.4% of OHCAs are preventable, being due to violent accidents or trauma, mainly occurring on the streets (23.2%). Conclusions: Limiting violent events is necessary to reduce OHCA mortality in children. Raising awareness and giving practical training to citizens is a priority in general but specifically in schools.

Is Mental Health Worse in Medical Students than in the General Population? A Cross-Sectional Study.

Background and Objectives: In recent years, there has been a notable increase in university students experiencing severe mental illness. The transition to university life can be demanding, leading to mental health disorders. Persistent stress and anxiety can cause demotivation, difficulties with concentration, cognitive impairment, and reduced academic performance. Mental health issues can also impact social relationships and overall well-being. This cross-sectional study aims to investigate the mental health of medical students and compare it with the mental health of the non-student population. Materials and Methods: The survey collected demographic information such as age and gender. Participants were questioned about their self-perceived mental distress, diagnosed mental disorders, and history of therapy for mental distress. Various validated assessment tools were utilized to assess mental health and quality of life. Results: Medical students exhibit a higher self-perception of mental symptoms that does not translate into a significantly higher prevalence of diagnosed mental disorders. Medical students experience higher levels of anxiety and subclinical depressive symptoms and lower quality of life. Female participants reported lower QoL scores and higher levels of anxiety symptoms compared with male participants. While the prevalence of mental disorders did not differ significantly between genders (except for clinical anxiety), females tended to perceive a higher burden of mental health challenges. Conclusions: By addressing mental health issues among medical students, especially females, institutions can create a more supportive and conducive learning environment. Encouraging open conversations about mental health and providing accessible mental health services can help in destigmatizing mental health challenges and promoting early intervention when needed.

Social Illness Before and After the COVID-19 Pandemic: A Regional Study.

INTRODUCTION: Coronavirus disease 2019 (COVID-19) has dramatically changed the epidemiology of several diseases. Much evidence on this has been published in the pandemic phase. In addition, many studies have shown that phenomena such as stress, substance abuse, and burnout increased in the general population during the lockdown. Unfortunately, few studies analyze the post-pandemic phase. STUDY OBJECTIVE: The study aimed to evaluate the trend of broad social problems, such as a diagnosis by the emergency department (ED), in the post-pandemic phase in the Lombardy (Italy) region. METHODS: The study is a retrospective observational cohort study. All admissions to emergency rooms in the Lombardy region registered in the Emergency Urgency OnLine (EUOL) portal made from January through June 2019 were analyzed, having as main causes: psychiatric disorders, self-harm, substance abuse, social disadvantage, and violence. All accesses in emergency rooms in the Lombardy region registered in the EUOL portal made from January 1, 2019 through June 30, 2019 were analyzed and compared with the same period in 2022. RESULTS: The study recorded an increase in the likelihood of events of self-harm (OR = 2.1; 95% CI, 1.8-2.6; P <.0001), substance abuse (OR = 1.2; 95% CI, 1.1-1.3; P <.0001), violence by others (OR = 1.3; 95% CI, 1.2-1.4; P <.0001), and social disadvantage (OR = 1.2; 95% CI, 1.1-1.4; P = .0045). The events are more concentrated in suburban areas (OR = 1.3; 95% CI, 1.2-1.4; P <.001). CONCLUSION: The increase in diagnoses of these social problems in the ED is only the culmination of a phenomenon that hides an underlying rise in social illness. In the post-COVID-19 phase, there is a need to invest in community care and social illness prevention policies.

Development of new N-Arylbenzamides as STAT3 Dimerization Inhibitors.

The O-tosylsalicylamide S3I-201 (10) was used as a starting point for design and synthesis of novel STAT-3 dimerization inhibitors with improved drug-like qualities. The phosphonic acid 12d and salicylic acids 13f, 13g with a shorter amide linker lacking the O-tosyl group had improved STAT-3 inhibitory activity. The equivalent potencies observed by the replacement of phosphonic acid moiety of 12d with 5-amino-2-hydroxybenzoic acid group as in 13f further validates 5-amino-2-hydroxybenzoic acid as a phosphotyrosine mimic. The salicylic acid 13f displayed improved whole cell activity. The focused library of salicylic acids 13 with benzamide linker indicated that hydrophobic heptyl and cyclohexyl are the best tolerated R groups and a biphenyl ether (as the Ar group) significantly contributes to STAT3 inhibitory activity. Our docking studies indicated that the acidic groups of 12d, 13f and 13g interact in the p-Tyr-705 binding site in a broadly similar manner, while the phenoxybenzoyl group and the cyclohexylbenzyl group occupying pY+1 and pY-X hydrophobic pockets respectively. The in vitro and cell based potency of 13f warrants further development of this scaffold as STAT3 inhibitors.

A novel inhibitor of STAT3 homodimerization selectively suppresses STAT3 activity and malignant transformation.

STAT3-STAT3 dimerization, which involves reciprocal binding of the STAT3-SH2 domain to phosphorylated tyrosine-705 (Y-705), is required for STAT3 nuclear translocation, DNA binding, and transcriptional regulation of downstream target genes. Here, we describe a small molecule S3I-1757 capable of disrupting STAT3-STAT3 dimerization, activation, and malignant transforming activity. Fluorescence polarization assay and molecular modeling suggest that S3I-1757 interacts with the phospho-Y-705-binding site in the SH2 domain and displaces fluorescein-labeled GpYLPQTV phosphotyrosine peptide from binding to STAT3. We generated hemagglutinin (HA)-tagged STAT3 and FLAG-tagged STAT3 and showed using coimmunoprecipitation and colocalization studies that S3I-1757 inhibits STAT3 dimerization and STAT3-EGF receptor (EGFR) binding in intact cells. Treatment of human cancer cells with S3I-1757 (but not a closely related analog, S3I-1756, which does not inhibit STAT3 dimerization), inhibits selectively the phosphorylation of STAT3 over AKT1 and ERK1/2 (MAPK3/1), nuclear accumulation of P-Y705-STAT3, STAT3-DNA binding, and transcriptional activation and suppresses the expression levels of STAT3 target genes, such as Bcl-xL (BCL2L1), survivin (BIRC5), cyclin D1 (CCND1), and matrix metalloproteinase (MMP)-9. Furthermore, S3I-1757, but not S3I-1756, inhibits anchorage-dependent and -independent growth, migration, and invasion of human cancer cells, which depend on STAT3. Finally, STAT3-C, a genetically engineered mutant of STAT3 that forms a constitutively dimerized STAT3, rescues cells from the effects of S3I-1757 inhibition. Thus, we have developed S3I-1757 as a STAT3-STAT3 dimerization inhibitor capable of blocking hyperactivated STAT3 and suppressing malignant transformation in human cancer cells that depend on STAT3.

RKI-1447 is a potent inhibitor of the Rho-associated ROCK kinases with anti-invasive and antitumor activities in breast cancer.

The Rho-associated kinases ROCK1 and ROCK2 are critical for cancer cell migration and invasion, suggesting they may be useful therapeutic targets. In this study, we describe the discovery and development of RKI-1447, a potent small molecule inhibitor of ROCK1 and ROCK2. Crystal structures of the RKI-1447/ROCK1 complex revealed that RKI-1447 is a Type I kinase inhibitor that binds the ATP binding site through interactions with the hinge region and the DFG motif. RKI-1447 suppressed phosphorylation of the ROCK substrates MLC-2 and MYPT-1 in human cancer cells, but had no effect on the phosphorylation levels of the AKT, MEK, and S6 kinase at concentrations as high as 10 µmol/L. RKI-1447 was also highly selective at inhibiting ROCK-mediated cytoskeleton re-organization (actin stress fiber formation) following LPA stimulation, but does not affect PAK-meditated lamellipodia and filopodia formation following PDGF and Bradykinin stimulation, respectively. RKI-1447 inhibited migration, invasion and anchorage-independent tumor growth of breast cancer cells. In contrast, RKI-1313, a much weaker analog in vitro, had little effect on the phosphorylation levels of ROCK substrates, migration, invasion or anchorage-independent growth. Finally, RKI-1447 was highly effective at inhibiting the outgrowth of mammary tumors in a transgenic mouse model. In summary, our findings establish RKI-1447 as a potent and selective ROCK inhibitor with significant anti-invasive and antitumor activities and offer a preclinical proof-of-concept that justify further examination of RKI-1447 suitability as a potential clinical candidate.

Development of o-chlorophenyl substituted pyrimidines as exceptionally potent aurora kinase inhibitors.

The o-carboxylic acid substituted bisanilinopyrimidine 1 was identified as a potent hit (Aurora A IC(50) = 6.1 ± 1.0 nM) from in-house screening. Detailed structure-activity relationship (SAR) studies indicated that polar substituents at the para position of the B-ring are critical for potent activity. X-ray crystallography studies revealed that compound 1 is a type I inhibitor that binds the Aurora kinase active site in a DFG-in conformation. Structure-activity guided replacement of the A-ring carboxylic acid with halogens and incorporation of fluorine at the pyrimidine 5-position led to highly potent inhibitors of Aurora A that bind in a DFG-out conformation. B-Ring modifications were undertaken to improve the solubility and cell permeability. Compounds such as 9m with water-solubilizing moieties at the para position of the B-ring inhibited the autophosphorylation of Aurora A in MDA-MB-468 breast cancer cells.

Fragment-based and structure-guided discovery and optimization of Rho kinase inhibitors.

Using high concentration biochemical assays and fragment-based screening assisted by structure-guided design, we discovered a novel class of Rho-kinase inhibitors. Compound 18 was equipotent for ROCK1 (IC(50) = 650 nM) and ROCK2 (IC(50) = 670 nM), whereas compound 24 was more selective for ROCK2 (IC(50) = 100 nM) over ROCK1 (IC(50) = 1690 nM). The crystal structure of the compound 18-ROCK1 complex revealed that 18 is a type 1 inhibitor that binds the hinge region in the ATP binding site. Compounds 18 and 24 inhibited potently the phosphorylation of the ROCK substrate MLC2 in intact human breast cancer cells.

A novel mechanism by which small molecule inhibitors induce the DFG flip in Aurora A.

Most protein kinases share a DFG (Asp-Phe-Gly) motif in the ATP site that can assume two distinct conformations, the active DFG-in and the inactive DFG-out states. Small molecule inhibitors able to induce the DFG-out state have received considerable attention in kinase drug discovery. Using a typical DFG-in inhibitor scaffold of Aurora A, a kinase involved in the regulation of cell division, we found that halogen and nitrile substituents directed at the N-terminally flanking residue Ala273 induced global conformational changes in the enzyme, leading to DFG-out inhibitors that are among the most potent Aurora A inhibitors reported to date. The data suggest an unprecedented mechanism of action, in which induced-dipole forces along the Ala273 side chain alter the charge distribution of the DFG backbone, allowing the DFG to unwind. As the ADFG sequence and three-dimensional structure is highly conserved, DFG-out inhibitors of other kinases may be designed by specifically targeting the flanking alanine residue with electric dipoles.

Pyridylthiazole-based ureas as inhibitors of Rho associated protein kinases (ROCK1 and 2).

Potent ROCK inhibitors of a new class of 1-benzyl-3-(4-pyridylthiazol-2-yl)ureas have been identified. Remarkable differences in activity were observed for ureas bearing a benzylic stereogenic center. Derivatives with hydroxy, methoxy and amino groups at the meta position of the phenyl ring give rise to the most potent inhibitors (low nM). Substitutions at the para position result in substantial loss of potency. Changes at the benzylic position are tolerated resulting in significant potency in the case of methyl and methylenehydroxy groups. X-Ray crystallography was used to establish the binding mode of this class of inhibitors and provides an explanation for the observed differences of the enantiomer series. Potent inhibition of ROCK in human lung cancer cells was shown by suppression of the levels of phosphorylation of the ROCK substrate MYPT-1.

Rb-Raf-1 interaction disruptor RRD-251 induces apoptosis in metastatic melanoma cells and synergizes with dacarbazine.

Metastatic melanoma is an aggressive cancer with very low response rate against conventional chemotherapeutic agents such as dacarbazine (DTIC). Inhibitor of Rb-Raf-1 interaction RRD-251 was tested against the melanoma cell lines SK-MEL-28, SK-MEL-5, and SK-MEL-2. RRD-251 was found to be a potent inhibitor of melanoma cell proliferation, irrespective of V600E B-Raf mutation status of the cell lines. In a SK-MEL-28 xenograft experiment, RRD-251 exerted a significant suppression of tumor growth compared with vehicle (P = 0.003). Similar to in vitro effects, tumors from RRD-251-treated animals showed decreased Rb-Raf-1 interaction in vivo. Growth suppressive effects of RRD-251 were associated with induction of apoptosis as well as a G(1) arrest, with an accompanying decrease in S-phase cells. RRD-251 inhibited Rb phosphorylation and downregulated E2F1 protein levels in these cells. Real-time PCR analysis showed that RRD-251 caused downregulation of cell-cycle regulatory genes thymidylate synthase (TS) and cdc6 as well as the antiapoptotic gene Mcl-1. Combinatorial treatment of RRD-251 and DTIC resulted in a significantly higher apoptosis in DTIC resistant cell lines SK-MEL-28 and SK-MEL-5, as revealed by increased caspase-3 activity and PARP cleavage. Because aberrant Rb/E2F pathway is associated with melanoma progression and resistance to apoptosis, these results suggest that the Rb-Raf-1 inhibitor could be an effective agent for melanoma treatment, either alone or in combination with DTIC.

A critical role for phosphatase haplodeficiency in the selective suppression of deletion 5q MDS by lenalidomide.

Lenalidomide is the first karyotype-selective therapeutic approved for the treatment of myelodysplastic syndromes (MDS) owing to high rates of erythroid and cytogenetic response in patients with chromosome 5q deletion [del(5q)]. Although haploinsufficiency for the RPS14 gene and others encoded within the common deleted region (CDR) have been implicated in the pathogenesis of the del(5q) phenotype, the molecular basis of the karyotype specificity of lenalidomide remains unexplained. We focused our analysis on possible haplodeficient enzymatic targets encoded within the CDR that play key roles in cell-cycle regulation. We show that the dual specificity phosphatases, Cdc25C and PP2Acalpha, which are coregulators of the G(2)-M checkpoint, are inhibited by lenalidomide. Gene expression was lower in MDS and acute myeloid leukemia (AML) specimens with del(5q) compared with those with alternate karyotypes. Lenalidomide inhibited phosphatase activity either directly (Cdc25C) or indirectly (PP2A) with corresponding retention of inhibitory phospho-tyrosine residues. Treatment of del(5q) AML cells with lenalidomide induced G(2) arrest and apoptosis, whereas there was no effect in nondel(5q) AML cells. Small interfering RNA (shRNA) suppression of Cdc25C and PP2Acalpha gene expression recapitulated del(5q) susceptibility to lenalidomide with induction of G(2) arrest and apoptosis in both U937 and primary nondel(5q) MDS cells. These data establish a role for allelic haplodeficiency of the lenalidomide inhibitable Cdc25C and PP2Acalpha phosphatases in the selective drug sensitivity of del(5q) MDS.

Inhibitors of Src homology-2 domain containing protein tyrosine phosphatase-2 (Shp2) based on oxindole scaffolds.

Screening of the NCI diversity set of compounds has led to the identification of 5 (NSC-117199), which inhibits the protein tyrosine phosphatase (PTP) Shp2 with an IC50 of 47 microM. A focused library incorporating an isatin scaffold was designed and evaluated for inhibition of Shp2 and Shp1 PTP activities. Several compounds were identified that selectively inhibit Shp2 over Shp1 and PTP1B with low to submicromolar activity. A model for the binding of the active compounds is proposed.