In Silico Assisted Identification, Synthesis, and In Vitro Pharmacological Characterization of Potent and Selective Blockers of the Epilepsy-Associated KCNT1 Channel.

Gain-of-function (GoF) variants in KCNT1 channels cause severe, drug-resistant forms of epilepsy. Quinidine is a known KCNT1 blocker, but its clinical use is limited due to severe drawbacks. To identify novel KCNT1 blockers, a homology model of human KCNT1 was built and used to screen an in-house library of compounds. Among the 20 molecules selected, five (CPK4, 13, 16, 18, and 20) showed strong KCNT1-blocking ability in an in vitro fluorescence-based assay. Patch-clamp experiments confirmed a higher KCNT1-blocking potency of these compounds when compared to quinidine, and their selectivity for KCNT1 over hERG and Kv7.2 channels. Among identified molecules, CPK20 displayed the highest metabolic stability; this compound also blocked KCNT2 currents, although with a lower potency, and counteracted GoF effects prompted by 2 recurrent epilepsy-causing KCNT1 variants (G288S and A934T). The present results provide solid rational basis for future design of novel compounds to counteract KCNT1-related neurological disorders.

Case report: Marked electroclinical improvement by fluoxetine treatment in a patient with KCNT1-related drug-resistant focal epilepsy.

Variants in KCNT1 are associated with a wide spectrum of epileptic phenotypes, including epilepsy of infancy with migrating focal seizures (EIMFS), non-EIMFS developmental and epileptic encephalopathies, autosomal dominant or sporadic sleep-related hypermotor epilepsy, and focal epilepsy. Here, we describe a girl affected by drug-resistant focal seizures, developmental delay and behavior disorders, caused by a novel, de novo heterozygous missense KCNT1 variant (c.2809A > G, p.S937G). Functional characterization in transiently transfected Chinese Hamster Ovary (CHO) cells revealed a strong gain-of-function effect determined by the KCNT1 p.S937G variant compared to wild-type, consisting in an increased maximal current density and a hyperpolarizing shift in current activation threshold. Exposure to the antidepressant drug fluoxetine inhibited currents expressed by both wild-type and mutant KCNT1 channels. Treatment of the proband with fluoxetine led to a prolonged electroclinical amelioration, with disappearance of seizures and better EEG background organization, together with an improvement in behavior and mood. Altogether, these results suggest that, based on the proband's genetic and functional characteristics, the antidepressant drug fluoxetine may be repurposed for the treatment of focal epilepsy caused by gain-of-function variants in KCNT1. Further studies are needed to verify whether this approach could be also applied to other phenotypes of the KCNT1-related epilepsies spectrum.

De novo variants in KCNA3 cause developmental and epileptic encephalopathy.

OBJECTIVE: Variants in several potassium channel genes, including KCNA1 and KCNA2, cause Developmental and Epileptic Encephalopathies (DEEs). We investigated whether variants in KCNA3, another mammalian homologue of the Drosophila shaker family and encoding for Kv1.3 subunits, can cause DEE. METHODS: Genetic analysis of study individuals was performed by routine exome or genome sequencing, usually of parent-offspring trios. Phenotyping was performed via a standard clinical questionnaire. Currents from wild-type and/or mutant Kv1.3 subunits were investigated by whole-cell patch-clamp upon their heterologous expression. RESULTS: Fourteen individuals, each carrying a de novo heterozygous missense variant in KCNA3, were identified. Most (12/14; 86%) had DEE with marked speech delay with or without motor delay, intellectual disability, epilepsy, and autism spectrum disorder. Functional analysis of Kv1.3 channels carrying each variant revealed heterogeneous functional changes, ranging from "pure" loss-of-function (LoF) effects due to faster inactivation kinetics, depolarized voltage-dependence of activation, slower activation kinetics, increased current inactivation, reduced or absent currents with or without dominant-negative effects, to "mixed" loss- and gain-of-function (GoF) effects. Compared to controls, Kv1.3 currents in lymphoblasts from 1 of the proband displayed functional changes similar to those observed upon heterologous expression of channels carrying the same variant. The antidepressant drug fluoxetine inhibited with similar potency the currents from wild-type and 1 of the Kv1.3 GoF variant. INTERPRETATION: We describe a novel association of de novo missense variants in KCNA3 with a human DEE, and provide evidence that fluoxetine might represent a potential targeted treatment for individuals carrying variants with significant GoF effects. ANN NEUROL 2024;95:365-376.

Biophysical Determinants for the Viscosity of Concentrated Monoclonal Antibody Solutions.

Monoclonal antibodies (mAbs) are particularly relevant for therapeutics due to their high specificity and versatility, and mAb-based drugs are hence used to treat numerous diseases. The increased patient compliance of self-administration motivates the formulation of products for subcutaneous (SC) administration. The associated challenge is to formulate highly concentrated antibody solutions to achieve a significant therapeutic effect, while limiting their viscosity and preserving their physicochemical stability. Protein-protein interactions (PPIs) are in fact the root cause of several potential problems concerning the stability, manufacturability, and delivery of a drug product. The understanding of macroscopic viscosity requires an in-depth knowledge on protein diffusion, PPIs, and self-association/aggregation. Here, we study the self-diffusion of different mAbs of the IgG1 subtype in aqueous solution as a function of the concentration and temperature by quasi-elastic neutron scattering (QENS). QENS allows us to probe the short-time self-diffusion of the molecules and therefore to determine the hydrodynamic mAb cluster size and to gain information on the internal mAb dynamics. Small-angle neutron scattering (SANS) is jointly employed to probe structural details and to understand the nature and intensity of PPIs. Complementary information is provided by molecular dynamics (MD) simulations and viscometry, thus obtaining a comprehensive picture of mAb diffusion.

A five-year (2017-2021) time series evaluation of patient-reported informal healthcare payments in Romania.

Low wages of health professionals are widely recognized as one of the drivers of informal payments in Romania's healthcare system. In January 2018, the government increased wages by an average of 70% to 172% in the public healthcare sector. This study examined the trends in patient-reported informal healthcare payments, discussing the effect of a one-time wage increase in 2018 and the impact of the COVID-19 pandemic in 2020 and 2021. It draws on monthly survey data of patient-reported informal payments collected between January 2017 and December 2021. We analyzed three periods: before the wage rise ("low pay"), between the wage rise and the COVID-19 pandemic ("high pay"), and during the COVID-19 pandemic. We found that patient-reported informal payments decreased between the "low pay" and "high pay" period but with a sharper decline during the COVID-19 pandemic. The share of respondents willing to report informal payments increased during the "high pay" period, indicating a stronger willingness to voice dissatisfaction with health services and informal payments, but slowed down during the first lockdown in 2020. Informal payments were more frequently reported in larger hospitals and the poorest geographical areas. While the 2018 wage increase may have contributed to less prevalent informal payments, survey coverage and design must be improved to draw robust, system-level conclusions to inform tailored policy actions.

A novel KCNC1 gain-of-function variant causing developmental and epileptic encephalopathy: "Precision medicine" approach with fluoxetine.

Variable phenotypes, including developmental encephalopathy with (DEE) or without seizures and myoclonic epilepsy and ataxia due to potassium channel mutation, are caused by pathogenetic variants in KCNC1, encoding for Kv3.1 channel subunits. In vitro, channels carrying most KCNC1 pathogenic variants display loss-of-function features. Here, we describe a child affected by DEE with fever-triggered seizures, caused by a novel de novo heterozygous missense KCNC1 variant (c.1273G>A; V425M). Patch-clamp recordings in transiently transfected CHO cells revealed that, compared to wild-type, Kv3.1 V425M currents (1) were larger, with membrane potentials between -40 and +40 mV; (2) displayed a hyperpolarizing shift in activation gating; (3) failed to inactivate; and (4) had slower activation and deactivation kinetics, consistent with a mixed functional pattern with prevalent gain-of-function effects. Exposure to the antidepressant drug fluoxetine inhibited currents expressed by both wild-type and mutant Kv3.1 channels. Treatment of the proband with fluoxetine led to a rapid and prolonged clinical amelioration, with the disappearance of seizures and an improvement in balance, gross motor skills, and oculomotor coordination. These results suggest that drug repurposing based on the specific genetic defect may provide an effective personalized treatment for KCNC1-related DEEs.

KCNT2-Related Disorders: Phenotypes, Functional, and Pharmacological Properties.

OBJECTIVE: Pathogenic variants in KCNT2 are rare causes of developmental epileptic encephalopathy (DEE). We herein describe the phenotypic and genetic features of patients with KCNT2-related DEE, and the in vitro functional and pharmacological properties of KCNT2 channels carrying 14 novel or previously untested variants. METHODS: Twenty-five patients harboring KCNT2 variants were investigated: 12 were identified through an international collaborative network, 13 were retrieved from the literature. Clinical data were collected and included in a standardized phenotyping sheet. Novel variants were detected using exome sequencing and classified using ACMG criteria. Functional and pharmacological studies were performed by whole-cell electrophysiology in HEK-293 and SH-SY5Y cells. RESULTS: The phenotypic spectrum encompassed: (a) intellectual disability/developmental delay (21/22 individuals with available information), ranging from mild to severe/profound; (b) epilepsy (15/25); (c) neurological impairment, with altered muscle tone (14/22); (d) dysmorphisms (13/20). Nineteen pathogenic KCNT2 variants were found (9 new, 10 reported previously): 16 missense, 1 in-frame deletion of a single amino acid, 1 nonsense, and 1 frameshift. Among tested variants, 8 showed gain-of-function (GoF), and 6 loss-of-function (LoF) features when expressed heterologously in vitro. Quinidine and fluoxetine blocked all GoF variants, whereas loxapine and riluzole activated some LoF variants while blocking others. INTERPRETATION: We expanded the phenotypic and genotypic spectrum of KCNT2-related disorders, highlighting novel genotype-phenotype associations. Pathogenic KCNT2 variants cause GoF or LoF in vitro phenotypes, and each shows a unique pharmacological profile, suggesting the need for in vitro functional and pharmacological investigation to enable targeted therapies based on the molecular phenotype. ANN NEUROL 2023;94:332-349.

KCNQ2 R144 variants cause neurodevelopmental disability with language impairment and autistic features without neonatal seizures through a gain-of-function mechanism.

BACKGROUND: Prior studies have revealed remarkable phenotypic heterogeneity in KCNQ2-related disorders, correlated with effects on biophysical features of heterologously expressed channels. Here, we assessed phenotypes and functional properties associated with KCNQ2 missense variants R144W, R144Q, and R144G. We also explored in vitro blockade of channels carrying R144Q mutant subunits by amitriptyline. METHODS: Patients were identified using the RIKEE database and through clinical collaborators. Phenotypes were collected by a standardized questionnaire. Functional and pharmacological properties of variant subunits were analyzed by whole-cell patch-clamp recordings. FINDINGS: Detailed clinical information on fifteen patients (14 novel and 1 previously published) was analyzed. All patients had developmental delay with prominent language impairment. R144Q patients were more severely affected than R144W patients. Infantile to childhood onset epilepsy occurred in 40%, while 67% of sleep-EEGs showed sleep-activated epileptiform activity. Ten patients (67%) showed autistic features. Activation gating of homomeric Kv7.2 R144W/Q/G channels was left-shifted, suggesting gain-of-function effects. Amitriptyline blocked channels containing Kv7.2 and Kv7.2 R144Q subunits. INTERPRETATION: Patients carrying KCNQ2 R144 gain-of-function variants have developmental delay with prominent language impairment, autistic features, often accompanied by infantile- to childhood-onset epilepsy and EEG sleep-activated epileptiform activity. The absence of neonatal seizures is a robust and important clinical differentiator between KCNQ2 gain-of-function and loss-of-function variants. The Kv7.2/7.3 channel blocker amitriptyline might represent a targeted treatment. FUNDING: Supported by FWO, GSKE, KCNQ2-Cure, Jack Pribaz Foundation, European Joint Programme on Rare Disease 2020, the Italian Ministry for University and Research, the Italian Ministry of Health, the European Commission, the University of Antwerp, NINDS, and Chalk Family Foundation.

Effect of linguistic framing and information provision on attitudes towards induced seismicity and seismicity regulation.

Shale gas is an expanding energy source worldwide, yet 'fracking' remains controversial. Amongst public concerns is induced seismicity (tremors). The UK had the most stringent induced seismicity regulations in the world, prior to instating a moratorium on shale gas development. The Government cited induced seismicity as the key rationale for its November 2019 English moratorium. Yet, little is known about how the public perceives induced seismicity, whether they support regulatory change, or how framing and information provision affect perceptions. Across three waves of a longitudinal experimental UK survey (N = 2777; 1858; 1439), we tested whether framing of induced seismicity influences support for changing regulations. The surveys compared (1) quantitative versus qualitative framings, (2) information provision about regulatory limits in other countries and (3) seismicity from other industries, and (4) framing a seismic event as an 'earthquake' or something else. We find low support for changing current policy, and that framing and information provision made little difference to this. The one strong influence on perceptions of seismic events came from the type of activity causing the event; shale gas extraction clearly led to the most negative reactions. We discuss implications for future UK policy on shale gas and geothermal energy in an evolving energy landscape.

Functional Characterization of Two Variants at the Intron 6-Exon 7 Boundary of the KCNQ2 Potassium Channel Gene Causing Distinct Epileptic Phenotypes.

Pathogenic variants in KCNQ2 encoding for Kv7.2 potassium channel subunits have been found in patients affected by widely diverging epileptic phenotypes, ranging from Self-Limiting Familial Neonatal Epilepsy (SLFNE) to severe Developmental and Epileptic Encephalopathy (DEE). Thus, understanding the pathogenic molecular mechanisms of KCNQ2 variants and their correlation with clinical phenotypes has a relevant impact on the clinical management of these patients. In the present study, the genetic, biochemical, and functional effects prompted by two variants, each found in a non-familial SLNE or a DEE patient but both affecting nucleotides at the KCNQ2 intron 6-exon 7 boundary, have been investigated to test whether and how they affected the splicing process and to clarify whether such mechanism might play a pathogenetic role in these patients. Analysis of KCNQ2 mRNA splicing in patient-derived lymphoblasts revealed that the SLNE-causing intronic variant (c.928-1G > C) impeded the use of the natural splice site, but lead to a 10-aa Kv7.2 in frame deletion (Kv7.2 p.G310Δ10); by contrast, the DEE-causing exonic variant (c.928G > A) only had subtle effects on the splicing process at this site, thus leading to the synthesis of a full-length subunit carrying the G310S missense variant (Kv7.2 p.G310S). Patch-clamp recordings in transiently-transfected CHO cells and primary neurons revealed that both variants fully impeded Kv7.2 channel function, and exerted strong dominant-negative effects when co-expressed with Kv7.2 and/or Kv7.3 subunits. Notably, Kv7.2 p.G310S, but not Kv7.2 p.G310Δ10, currents were recovered upon overexpression of the PIP2-synthesizing enzyme PIP5K, and/or CaM; moreover, currents from heteromeric Kv7.2/Kv7.3 channels incorporating either Kv7.2 mutant subunits were differentially regulated by changes in PIP2 availability, with Kv7.2/Kv7.2 G310S/Kv7.3 currents showing a greater sensitivity to PIP2 depletion when compared to those from Kv7.2/Kv7.2 G310Δ10/Kv7.3 channels. Altogether, these results suggest that the two variants investigated differentially affected the splicing process at the intron 6-exon 7 boundary, and led to the synthesis of Kv7.2 subunits showing a differential sensitivity to PIP2 and CaM regulation; more studies are needed to clarify how such different functional properties contribute to the widely-divergent clinical phenotypes.

Gain of function due to increased opening probability by two KCNQ5 pore variants causing developmental and epileptic encephalopathy.

Developmental and epileptic encephalopathies (DEEs) are neurodevelopmental diseases characterized by refractory epilepsy, distinct electroencephalographic and neuroradiological features, and various degrees of developmental delay. Mutations in KCNQ2, KCNQ3, and, more rarely, KCNQ5 genes encoding voltage-gated potassium channel subunits variably contributing to excitability control of specific neuronal populations at distinct developmental stages have been associated to DEEs. In the present work, the clinical features of two DEE patients carrying de novo KCNQ5 variants affecting the same residue in the pore region of the Kv7.5 subunit (G347S/A) are described. The in vitro functional properties of channels incorporating these variants were investigated with electrophysiological and biochemical techniques to highlight pathophysiological disease mechanisms. Currents carried by Kv7.5 G347 S/A channels displayed: 1) large (>10 times) increases in maximal current density, 2) the occurrence of a voltage-independent component, 3) slower deactivation kinetics, and 4) hyperpolarization shift in activation. All these functional features are consistent with a gain-of-function (GoF) pathogenetic mechanism. Similar functional changes were also observed when the same variants were introduced at the corresponding position in Kv7.2 subunits. Nonstationary noise analysis revealed that GoF effects observed for both Kv7.2 and Kv7.5 variants were mainly attributable to an increase in single-channel open probability, without changes in membrane abundance or single-channel conductance. The mutation-induced increase in channel opening probability was insensitive to manipulation of membrane levels of the critical Kv7 channel regulator PIP2. These results reveal a pathophysiological mechanism for KCNQ5-related DEEs, which might be exploited to implement personalized treatments.

Kv7.4 channels regulate potassium permeability in neuronal mitochondria.

Mitochondrial K+ permeability regulates neuronal apoptosis, energy metabolism, autophagy, and protection against ischemia-reperfusion injury. Kv7.4 channels have been recently shown to regulate K+ permeability in cardiac mitochondria and exert cardioprotective effects. Here, the possible expression and functional role of Kv7.4 channels in regulating membrane potential, radical oxygen species (ROS) production, and Ca2+ uptake in neuronal mitochondria was investigated in both clonal (F11 cells) and native brain neurons. In coupled mitochondria isolated from F11 cells, K+-dependent changes of mitochondrial membrane potential (ΔΨ) were unaffected by the selective mitoBKCa channel blocker iberiotoxin and only partially inhibited by the mitoKATP blockers glyburide or ATP. Interestingly, K+-dependent ΔΨ decrease was significantly reduced by the Kv7 blocker XE991 and enhanced by the Kv7 activator retigabine. Among Kv7s, western blot experiments showed the expression of only Kv7.4 subunits in F11 mitochondrial fractions; immunocytochemistry experiments showed a strong overlap between the Kv7.4 fluorescent signal and that of the mitochondrial marker Mitotracker. Silencing of Kv7.4 expression significantly suppressed retigabine-dependent decrease in ΔΨ in intact F11 cells. Expression of Kv7.4 subunits was also detected by western blot in isolated mitochondria from total mouse brain and by immunofluorescence in mouse primary cortical neurons. Pharmacological experiments revealed a relevant functional role for Kv7.4 channels in regulating membrane potential and Ca2+ uptake in isolated neuronal mitochondria, as well as ΔΨ and ROS production in intact cortical neurons. In conclusion, these findings provide the first experimental evidence for the expression of Kv7.4 channels and their contribution in regulating K+ permeability of neuronal mitochondria.

Își pot permite oamenii să plătească pentru servicii de sănătate? Noi dovezi privind protecția financiară în România

Această analiză face parte dintr-o serie de studii naționale menite să obțină dovezi noi privind protecția financiară în Europa. Protecția financiară este esențială pentru acoperirea universală cu servicii de sănătate și este un aspect esențial al performanței sistemelor sanitare. În România există o incidență mai mare a cheltuielilor pentru sănătate catastrofale decât în alte țări din Uniunea Europeană, și este mult mai mare decât ar fi fost de așteptat, ținând cont de faptul că România nu se bazează foarte mult pe plăți directe pentru a finanța sistemul de sănătate. Acestea sunt concentrate îndeosebi în rândul gospodăriilor mai sărace și al persoanelor în vârstă, demonstrând astfel lacunele la nivelul celor trei dimensiuni ale acoperirii medicale (drepturile populației, acoperirea cu servicii și contribuțiile suportate de beneficiari), punctele slabe ale politicii de cumpărare de servicii și nivelurile scăzute ale cheltuielilor publice pentru sănătate. Dificultățile financiare sunt determinate în principal de plățile directe pentru medicamentele din ambulatoriu și au crescut pe parcursul timpului, mai ales în rândul gospodăriilor mai sărace. Pentru a putea reduce nevoile nesatisfăcute și dificultățile financiare, Guvernul ar trebui să se axeze pe a face mai accesibile ca preț medicamentele din ambulatoriu (inclusiv cele eliberate fără prescripție medicală) și serviciile stomatologice. Aceasta se poate realiza prin identificarea de soluții pentru: asigurarea întregii populații în sistemul Casei Naționale de Asigurări de Sănătate (CNAS); introducerea unor excepții de la coplată pentru gospodăriile cu venituri mici și pentru persoanele cu afecțiuni cronice; introducerea unui plafon anual pentru toate coplățile, corelat cu venitul gospodăriei; îmbunătățirea politicii CNAS de cumpărare a serviciilor de sănătate; limitarea contribuțiilor personale și a plăților pentru servicii suplimentare; și luarea de măsuri în privința plăților informale și a calității serviciilor. Aceste eforturi vor necesita creșteri substanțiale și constante ale cheltuielilor publice pentru sănătate. Toate fondurile suplimentare noi ar trebui alocate cu atenție, astfel încât să îndeplinească obiectivele de echitate și eficiență.

Can people afford to pay for health care? New evidence on financial protection in Romania

This review is part of a series of country-based studies generating new evidence on financial protection in Europe. Financial protection is central to universal health coverage and a core dimension of health system performance. The incidence of catastrophic health spending is high in Romania compared to many countries in the European Union and higher than expected given Romania’s relatively low reliance on out-of-pocket payments to finance the health system. It is heavily concentrated among poorer households and older people, reflecting gaps in all three dimensions of health coverage (population entitlement, service coverage and user charges), weaknesses in purchasing policy and low levels of public spending on health. Financial hardship is mainly driven by out-of-pocket payments for outpatient medicines and has increased over time, particularly among poorer households. To reduce unmet need and financial hardship, the Government should focus on improving the affordability of outpatient medicines (including over-the-counter treatment) and dental care. This can be done by finding ways to: ensure the National Health Insurance House (NHIH) covers the whole population; introduce exemptions from co-payments for low-income households and people with chronic conditions; introduce an annual cap on all co-payments and link the cap to household income; improve NHIH purchasing policy; limit balance billing and extra billing; and pay attention to informal payments and service quality. These efforts will require sustained increases in public spending on health. Any new additional funds should be carefully allocated to meet equity and efficiency goals.

Дали луѓето можат да си дозволат да платат за здравствена заштита? Нови докази за финансиската заштита во Северна Македонија: резиме

Со овој преглед се оценува до кој степен луѓето во СевернаМакедонија доживуваат финансиски тешкотии кога користатздравствени услуги, вклучувајќи и лекови. Анализата го опфаќапериодот од 2006 година до денес, врз основа на податоци одгодишните анкети за потрошувачката на домаќинствата спроведенимеѓу 2006 и 2018 г. од Државниот завод за статистика во СевернаМакедонија, податоци за незадоволената потреба од здравственазаштита до 2019 година и информации за политиката за покриеностдо 2021 година. Се фокусира на два показатела за финансисказаштита: катастрофални трошоци за здравствена заштита и трошоциза здравствена заштита, што доведуваат до сиромаштија. Исто така,се разгледува присуството на бариери во пристапот до здравственатазаштита, што доведува до незадоволена потреба од здравствени истоматолошки услуги.

Дали луѓето можат да си дозволат да платат за здравствена заштита? Нови докази за финансиската заштита во Северна Македонија

Овој преглед е дел од серијата националнистудии за собирање нови докази за финансискатазаштита во здравствените системи во Европа.Финансиската заштита има клучно значење зауниверзалната здравствена заштита и претставуваосновен елемент на функционирањето наздравствениот систем.Иако пристапот и финансиската заштита воСеверна Македонија, во последните годинисе подобрени, катастрофалните трошоци заздравствена заштита и натаму претставуваатпроблем, посебно за посиромашнитедомаќинства, а тие во голема мера се поврзани соприватните плаќања за лекови во аптеките.За да се подобри пристапот и финансискатазаштита во здравствениот систем треба: да сеотстрани врската меѓу остварувањето на праватаи плаќањето придонеси за целото население,така што пристапот до здравствена заштитаповеќе нема да зависи од статусот на здравственоосигурување; да се поедностави сложениотсистем на учество на корисниците и да сезголеми заштитата при плаќање партиципацијаза лекови на рецепт; да се подобри достапностана лековите на рецепт преку подобрување населекцијата и стратешкото купување лекови иредовно ажурирање на позитивната листа; да сеадресираат неформалните плаќања, почнувајќи сововедување подобар мониторинг и да се зголематјавните вложувања во здравствениот систем,преку давање поголем приоритет на здравствотопри распределба на јавните средства.

Can people afford to pay for health care? New evidence on financial protection in North Macedonia

This review is part of a series of country-based studies generating new evidence on financial protection in health systems in Europe. Financial protection is central to universal health coverage and a core dimensionof health system performance. Although access and financial protection have improved in North Macedonia in recent years, catastrophic health spending remains a problem, particularly for poorer households, and is largely driven by out-of-pocket payments for outpatient medicines. To improve access and financial protection, the health system should: de-link entitlement from payment of contributions for the whole population, so that access to health care no longer depends on health insurance status; simplify the complex design of user charges and strengthen protection against co-payments for outpatient medicines; improve the affordability of outpatient prescribed medicines by enhancing the selection and purchasing of medicines and regularly updating the positive list; address informal payments, starting with better monitoring; and increase public investment in the health system through sustained rises in the priority given to health in allocating government spending.

Can people afford to pay for health care? New evidence on financial protection in North Macedonia: summary

This review assesses the extent to which people in North Macedonia experience financial hardship when they use health services, including medicines. The analysis covers the period from 2006 to the present day, drawing on data from household budget surveys carried out annually between 2006 and 2018 by the State Statistical Office in North Macedonia, data on unmet need for health care up to 2019 and information on coverage policy up to 2021. It focuses on two indicators of financial protection: catastrophic health spending and impoverishing health spending. It also considers the presence of access barriers leading to unmet need for health and dental care.

Calcium cytotoxicity sensitizes prostate cancer cells to standard-of-care treatments for locally advanced tumors.

Therapy resistance is a major roadblock in oncology. Exacerbation of molecular dysfunctions typical of cancer cells have proven effective in twisting oncogenic mechanisms to lethal conditions, thus offering new therapeutic avenues for cancer treatment. Here, we demonstrate that selective agonists of Transient Receptor Potential cation channel subfamily M member 8 (TRPM8), a cation channel characteristic of the prostate epithelium frequently overexpressed in advanced stage III/IV prostate cancers (PCa), sensitize therapy refractory models of PCa to radio, chemo or hormonal treatment. Overall, our study demonstrates that pharmacological-induced Ca2+ cytotoxicity is an actionable strategy to sensitize cancer cells to standard therapies.

Gabapentin treatment in a patient with KCNQ2 developmental epileptic encephalopathy.

De novo variants in KCNQ2 encoding for Kv7.2 voltage-dependent neuronal potassium (K+) channel subunits are associated with developmental epileptic encephalopathy (DEE). We herein describe the clinical and electroencephalographic (EEG) features of a child with early-onset DEE caused by the novel KCNQ2 p.G310S variant. In vitro experiments demonstrated that the mutation induces loss-of-function effects on the currents produced by channels incorporating mutant subunits; these effects were counteracted by the selective Kv7 opener retigabine and by gabapentin, a recently described Kv7 activator. Given these data, the patient started treatment with gabapentin, showing a rapid and sustained clinical and EEG improvement over the following months. Overall, these results suggest that gabapentin can be regarded as a precision therapy for DEEs due to KCNQ2 loss-of-function mutations.