Pbmc Immunophenotyping and Plasma Inflammatory Profile of Children with Long Covid

Background: Long COVID can be developed by individuals after an infection with SARS-CoV-2 as described by the WHO. Although this condition is more commonly described in adults, it can occur in children and adolescents with a wide range of estimated prevalence of 1-25%. Little is known about the role of the immune system in long COVID. However, one of the main hypotheses about the underlying mechanism in long COVID is that there is an immune and inflammatory dysregulation that persists after the acute infection. The objective of this study is to compare immune cells populations, and inflammatory biomarkers in paediatric populations with and without long COVID. Method(s): We analyzed 55 blood samples from the pediaCOVID cohort (Hospital Germans Trias i Pujol), which includes more than 130 children diagnosed with long COVID and 23 controls. We measured different immune cell populations using spectral cytometry with a panel of 37 cellular markers, and 42 inflammatory markers using Luminex or ELISA. EdgeR was used for statistical analysis of the spectral data;p-values of inflammatory markers were calculated using the likelihood ratio test and they were corrected for multiple comparisons. Result(s): The study cohort had a median age of 14.3 (IQR, 12.5-15.2) and 69.1% female. Patients had at least 3 symptoms associated with long COVID (median [IQR];10 [7-16]). The most common symptom was asthenia/fatigue (98.2%). Compared to the control cohort, children with long COVID had increased numbers of CD4+CD8+ T cells, IgA+CD21+CD27+ memory B cells, and IgA+CD21-CD27- memory B cells, while CD4+ TEMRA cells (CD45RA+, CCR7-), intermediate monocytes (CD14+, CD16+) and classical monocytes (CD14+, CD16-) were decreased (all p< 0.05;q=n.s.). None of the 42 inflammatory biomarkers showed significant differences between children with and without long COVID. Conclusion(s): The results of this study suggest that specific populations of peripheral blood immune cells might be involved in the mechanisms underlying prolonged COVID in children and adolescents. The increase in both IgA+CD21-CD27- and IgA+CD21+CD27+ memory B cells could be associated with the persistence of viral antigen in the gut and/or gut dysbiosis. Moreover, the decrease in CD4+ TEMRA cells could be related to autoantibodies against G-protein coupled receptors (GPCRs), since this cell population can express GPR56, and autoantibodies against GPCRs were previously reported to be elevated in adults with long COVID.

SARS-CoV-2 Kerala Isolate Spike Protein Induces Cancer Proliferating Markers for Lung and Breast Cancer: An In Silico Approach

Background: Coronavirus disease (COVID 19) has been emerging as a major threat to humans all over the world. Severe Acute Respiratory Syndrome CoronaVirus 2 (nSARS-CoV-2) is the causative agent for the disease resulting in severe acute respiratory illness. Earlier, it took several years to come up with a vaccine or other sorts of treatments for viral diseases. But now with the advent of biotechnology and development of bio-informatic tools, the process has been accelerated. The WHO reports 39,806,488 affected cases and 1,112,208 deaths till today all over the world (17 Oct 2020). nSARS-CoV-2 has a greater influence on people with comorbidities mainly cancer. Objective(s): The study herein attempts to understand the binding affinity of the spike protein of the novel coronavirus with the lung and breast cancer marker proteins by docking and ClusPro analysis. Method(s): The analysis was conducted in reference to hACE2 (human Angiotensin Converting Enzyme 2), the receptor of nSARS-CoV-2. Total 22 different marker proteins were analyzed using ClusPro. Result(s): BRCA1 (Breast Cancer type 1 susceptibility protein) and CXCR4 (a chemokine receptor belong-ing to the G protein coupled receptor family) were found to exhibit higher binding affinities.-73.82 kcal/mol and-66.45 kcal/mol were the global energies they showed upon binding to S protein respective-ly. Conclusion(s): Therefore, novel SARS-CoV-2 has a higher chance of inducing cancer in non-cancerous individuals and aids in cancer acceleration in cancer patients. This poses a threat to cancer patients and immunocompromised individuals. The study can be exploited to identify the optimal drug delivery system for novel SARS CoV2.Copyright © 2022 Bentham Science Publishers.

Anosmia in COVID-19 and post-COVID syndrome

Anosmia is one of the most frequent complaints in COVID-19 and post-COVID syndrome. The exact mechanism through which the virus leads to this symptom is still unclear. A proposed mechanism is that in "immune-balanced” individuals, the coronavirus damages olfactory epithelium and induce acute local inflammation with edema and the development of anosmia, which is recovered after acute phase. However, in subjects with genetical predisposition for developing autoimmunity, the immune response to the virus could create a parallel autoimmune response creating autoantibodies targeting the olfactory receptors and similar peptides (G-protein-coupled receptors), permitting an inflammatory process in the central nervous system through the olfactory bulb, where induces inflammation with the development of neurological symptoms, including persistent anosmia, fatigue, brain fog, etc. © 2023 Elsevier Inc. All rights reserved.

Human T2R38 Bitter Taste Receptor Expression and COVID-19: From Immunity to Prognosis

Background: Bitter taste-sensing type 2 receptor (T2Rs or TAS2Rs) found on ciliated epithelial cells and solitary chemosensory cells have a role in respiratory tract immuni-ty. T2Rs have shown protection against SARS-CoV-2 by enhancing the innate immune response. The purpose of this review is to outline the current sphere of knowledge re-garding this association. Method(s): A narrative review of the literature was done by searching (T2R38 OR bitter taste receptor) AND (COVID-19 OR SARS-CoV-2) keywords in PubMed and google scholar. Result(s): T2R38, an isoform of T2Rs encoded by the TAS2R38 gene, may have a potential association between phenotypic expression of T2R38 and prognosis of COVID-19. Current studies suggest that due to different genotypes and widespread distributions of T2Rs within the respiratory tract and their role in innate immunity, treatment protocols for COVID-19 and other respiratory diseases may change accordingly. Based on the phenotypic expression of T2R38, it varies in innate immunity and host response to respiratory infection, systemic symptoms and hospitalization. Conclusion(s): This review reveals that patients' innate immune response to SARS-COV-2 could be influenced by T2R38 receptor allelic variations.Copyright © 2023, Avicenna Journal of Medical Biotechnology.

Polysaturated fatty acids and their derivatives regulate respiratory infections.

The regulation of inflammation is a complex pathophysiological process that depends on the production of oxygenated lipid derivatives essential polyunsaturated fatty acids, like omega-3 and omega-6, among which are the lipoxins resolvins and protectins, called specialized pro-resolving lipid mediators (SPM). Their activity is associated with the control of respiratory infection processes to modulate the production of proinflammatory cytokines, avoiding damage due to inflammation-associated necrosis, reducing microbial loads, and promoting tissue remodeling. Therefore, we review some of the biochemical, physiological and immunological aspects of SPM in the regulation of inflammation in respiratory infections.Copyright © 2022, Instituto Nacional de Enfermedades Respiratorias. All rights reserved.

ß-arrestins and G protein-coupled receptor kinases in viral entry: A graphical review.

Viruses rely on host-cell machinery in order to invade host cells and carry out a successful infection. G-protein coupled receptor (GPCR)-mediated signaling pathways are master regulators of cellular physiological processing and are an attractive target for viruses to rewire cells during infection. In particular, the GPCR-associated scaffolding proteins ß-arrestins and GPCR signaling effectors G-protein receptor kinases (GRKs) have been identified as key cellular factors that mediate viral entry and orchestrate signaling pathways that reprogram cells for viral replication. Interestingly, a broad range of viruses have been identified to activate and/or require GPCR-mediated pathways for infection, including polyomaviruses, flaviviruses, influenza virus, and SARS-CoV-2, demonstrating that these viruses may have conserved mechanisms of host-cell invasion. Thus, GPCR-mediated pathways highlight an attractive target for the development of broad antiviral therapies.

Clostridium butyricum-induced ω-3 fatty acid 18-HEPE elicits anti-influenza virus pneumonia effects through interferon-λ upregulation.

The precise mechanism by which butyrate-producing bacteria in the gut contribute to resistance to respiratory viral infections remains to be elucidated. Here, we describe a gut-lung axis mechanism and report that orally administered Clostridium butyricum (CB) enhances influenza virus infection resistance through upregulation of interferon (IFN)-λ in lung epithelial cells. Gut microbiome-induced ω-3 fatty acid 18-hydroxy eicosapentaenoic acid (18-HEPE) promotes IFN-λ production through the G protein-coupled receptor (GPR)120 and IFN regulatory factor (IRF)-1/-7 activations. CB promotes 18-HEPE production in the gut and enhances ω-3 fatty acid sensitivity in the lungs by promoting GPR120 expression. This study finds a gut-lung axis mechanism and provides insights into the treatments and prophylaxis for viral respiratory infections.

Dysregulated autoantibodies targeting vaso- and immunoregulatory receptors in Post COVID Syndrome correlate with symptom severity.

Most patients with Post COVID Syndrome (PCS) present with a plethora of symptoms without clear evidence of organ dysfunction. A subset of them fulfills diagnostic criteria of myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS). Symptom severity of ME/CFS correlates with natural regulatory autoantibody (AAB) levels targeting several G-protein coupled receptors (GPCR). In this exploratory study, we analyzed serum AAB levels against vaso- and immunoregulatory receptors, mostly GPCRs, in 80 PCS patients following mild-to-moderate COVID-19, with 40 of them fulfilling diagnostic criteria of ME/CFS. Healthy seronegative (n=38) and asymptomatic post COVID-19 controls (n=40) were also included in the study as control groups. We found lower levels for various AABs in PCS compared to at least one control group, accompanied by alterations in the correlations among AABs. Classification using random forest indicated AABs targeting ADRB2, STAB1, and ADRA2A as the strongest classifiers (AABs stratifying patients according to disease outcomes) of post COVID-19 outcomes. Several AABs correlated with symptom severity in PCS groups. Remarkably, severity of fatigue and vasomotor symptoms were associated with ADRB2 AAB levels in PCS/ME/CFS patients. Our study identified dysregulation of AAB against various receptors involved in the autonomous nervous system (ANS), vaso-, and immunoregulation and their correlation with symptom severity, pointing to their role in the pathogenesis of PCS.

The GNB3 c.825C>T (rs5443) polymorphism and protection against fatal outcome of corona virus disease 2019 (COVID-19).

Background and aims: Albeit several factors which influence the outcome of corona virus disease (COVID-19) are already known, genetic markers which may predict the outcome of the disease in hospitalized patients are still very sparse. Thus, in this study, we aimed to analyze whether the single-nucleotide polymorphism (SNP) rs5443 in the gene GNB3, which was associated with higher T cell responses in previous studies, might be a suitable biomarker to predict T cell responses and the outcome of COVID-19 in a comprehensive German cohort. Methods: We analyzed the influence of demographics, pre-existing disorders, laboratory parameters at the time of hospitalization, and GNB3 rs5443 genotype in a comprehensive cohort (N = 1570) on the outcome of COVID-19. In a sub cohort, we analyzed SARS-CoV-2-specific T cell responses and associated GNB3 rs5443 genotypes. We investigated the influence of all factors on COVID-19 fatality in multivariable analysis. Results: We found a younger patient age, normotension or absence of diabetes mellitus or cardiovascular diseases, normal blood cell counts, and low inflammatory markers at hospital admission were protective factors against fatal course of disease. In addition, the rs5443 TT genotype was significantly associated with protection against COVID-19 fatality (OR: 0.60, 95% CI: 0.40-0.92, p = 0.02). We also observed significantly increased SARS-CoV-2-specific T cell responses in rs5443 TT genotype carriers (p = 0.01). Although we observed a significant association of the factors described previously in univariate analysis, only a younger age of the patients, normal blood cell counts, and the GNB3 rs5443 TT genotype remained independent predictors against COVID-19 fatality in multivariable analysis. Conclusion: Immutable predictors for COVID-19 fatality are relatively rare. In this study we could show that the TT genotype of the SNP rs5443 in the gene GNB3 is associated with protection against COVID-19 fatality. It was as well correlated to higher SARS-CoV-2-specific T cell responses, which could result in a milder course of disease in those patients. Based on those observations we hereby provide a further prognostic biomarker, which might be used in routine diagnostics as a predictive factor for COVID-19 mortality already upon hospitalization.

Possible Impact of functional active GPCRautoantibodies on retinal microcirculation in Long-COVID

Purpose : Long-COVID is assumed to have an autoimmune component. As a seropositivity of functional active autoantibodies against G-protein coupled receptors (GPCR-AAb) and an impaired retinal microcirculation, measured by OCT-angiography (OCT-A), was observed after COVID-19 infection, it was the aim of the present study to investigate the impact of GPCR-AAbs on retinal microcirculation in patients with LongCOVID. Methods : Seventy-six patients with Long-COVID (COVID-19 infection was confirmed by real time rt-PCR) were recruited. A seropositivity for GPCR-AAbs (Nociceptin-AAb, ß2-AAb, AT1-AAb, α1-AAb, MAS-AAb, M2-AAb, ETA-AAb) was analyzed by a specific rat cardiomyocyte bioassay in vitro. Macular (superficial vascular plexus (SVP), intermediate capillary plexus (ICP) and deep capillary plexus (DCP)) and peripapillary vessel density (VD) were measured by OCT-A (Spectralis II, Heidelberg, Germany). VD analysis was done by the Erlangen-Angio tool, including an APSifyed and Bruch's membrane opening (BMO) based analysis. Data were analyzed by mixed model (SAS version 9.4). Gender and age were set as covariates. The study was approved by the local ethics committee and was done in accordance with the tenets of the Declaration of Helsinki. Informed consent was achieved. Results : All patients with Long-COVID showed a seropositivity for GPCR-AAbs (100%). Female patients with Long-COVID showed lower macular and peripapillary VD compared to men. A significant effect of a seropositivity of AT1-AAb on age-corrected least squares means (LS-means) overall VD of DCP ([CI: 0,25;6.07], p=0.03). A seropositivity of α1-AAb showed a significant effect on age-corrected LS-mean overall VD of SVP ([CI: 0.07;2.69], p=0.04), of DCP ([CI: 0.36;3.25], p=0.01), and of PH ([CI: 1.17;5.59], p=0.01), respectively. A seropositivity of MAS-AAb yielded a significant effect on age-corrected LS-mean overall VD of DH (CI: 1.48;6.07, p=0.001). A seropositivity of ß2-AAb has a significant effect on agecorrected LS-mean overall VD of MH (CI: 0,02;1,94, p=0.04). For those effects, the covariate age was significant in the type III tests (p<0.05), thus analysis was done considering this age-effect. Conclusions : As autoimmune mechanisms were reported to be involved in the pathogenesis of Long-COVID, we postulate that functional active GPCR-AAb may have an impact on retinal microcirculation, being a propable correlate to systemic disease.

The REEP family of proteins: Molecular targets and role in pathophysiology.

Receptor expression-enhancing proteins (REEPs) are an evolutionarily conserved protein family that is pivotal to the structure and function of the endoplasmic reticulum (ER). The REEP family can be classified into two major subfamilies in higher species, the REEP1-4 and REEP5-6 subfamilies. Within the REEP1-4 subfamily, REEP1 and REEP2 are closely related, and REEP3 and REEP4 are similarly related. The REEP family is widely distributed in various tissues. Recent studies indicate that the REEP family is involved in many pathological and physiological processes, such as ER morphogenesis and remodeling, microtubule cytoskeleton regulation, and the trafficking and expression of G protein-coupled receptors (GPCRs). Moreover, the REEP family plays crucial roles in the occurrence and development of many diseases, including neurological diseases, diabetes, retinal diseases, cardiac diseases, infertility, obesity, oligoarticular juvenile idiopathic arthritis (OJIA), COVID-19, and cancer. In the present review, we describe the distribution and structure of the REEP family. Furthermore, we summarize the functions and the associated diseases of this family. Based on the pleiotropic actions of the REEP family, the study of its family members is crucial to understanding the relevant pathophysiological processes and developing strategies to modulate and control these related diseases.

INCREASED PROTEASE-ACTIVATED RECEPTOR 1 AUTOANTIBODIES ARE ASSOCIATED with SEVERE COVID-19

Background: In acute COVID-19 infection, growing evidence hints towards a broad activation of plasma cells and the presence of pathologic autoantibodies (abs). A systematic screening for abs confrmed induction of diverse functional abs by SARS-CoV-2 infection (1, 2). Immune-mediated thrombosis, involving platelet activation, has been identifed as one of the key pathogenic mechanisms in COVID-19 and is linked to morbidity and mortality (3). As natural abs against G protein-coupled receptors, functional abs against the thrombin receptor type-1 (PAR-1) might predispose for increased activation of the coagulation system present in COVID-19 infection. Objectives: The aim of this study is to identify the diagnostic value of anti-PAR1 antibodies and their capacity to predict the outcome of COVID-19 infection. Methods: 82 serum samples from 55 individuals with COVID-19 derived from three different hospitals in Schleswig-Holstein, Germany, and 88 single time point samples from healthy controls were subjected to ELISA-based quantifcation of anti-PAR-1 abs (CellTrend GmbH Luckenwalde, Germany). The levels of anti-AT1R abs were compared with clinical and laboratory parameters. Results: COVID-19 patients revealed markedly increased levels of circulating anti-PAR1 abs in hospitalized patients particularly in those required intensive care treatment in comparison to controls (p < 0.0001, Figure 1a). Anti-PAR1 ab levels were highest in patients with fatal outcome (p = 0.006, Figure 1a). Receiver operating characteristic (ROC) analysis of PAR1 abs levels in COVID-19 patients revealed a sensitivity of 84.00% and a specifcity 79.25% for patients requiring intensive care unit (ICU) treatment and a sensitivity of 87.50 % and a specifcity 84.51 % to distinguish fatal vs. non-fatal disease outcome (Figure 1b). We found correlation of circulating anti-PAR1 abs with D dimers. Conclusion: The increased anti-PAR1 abs, their prediction to identify patients requiring ICU and fatal outcome, and the correlation with markers for blood clotting suggest a role for antibodies against PAR1 in the disease development of blood clotting in COVID-19.

21stRSC/SCI Medicinal Chemistry Symposium Cambridge/Virtual - September 13-15, 2021

The Royal Chemistry Society/Society of Chemistry Industry (RSC/SCI) Medicinal Chemistry Symposium is a key symposium in the field of medicinal chemistry that takes place every 2 years at the University of Cambridge, U.K., under the auspices of the RSC and the SCI. This year, in its 21st edition, it was run as a hybrid event, simultaneously attended by both in-person and virtual attendees. Its theme was 'Improving Success', and 25 orals and 30 posters were presented. The scientific program covered recent medicinal chemistry achievements in major therapeutic areas, with a few first-time structure disclosure presentations and particular emphasis on emerging early-stage drug discovery and lead optimization strategies, including reports of successful case studies illustrating fragment-based screening and phenotypic target engagement approaches. Applications of artificial intelligence (AI), mass spectrometry and photochemistry in drug design and discovery were also highlighted. This report will cover some of the medicinal chemistry presentations delivered at the meeting.

Role of adenosine receptors in cardiovascular disorders and novel corona virus infection

Objective: To assess the importance of adenosine signaling in cardiovascular disorders (thrombosis, ischemia) and novel corona virus infection. Methodology: A specified web search was done to gather the relevant information using different scientific research forums and databases like WHO database, Pubmed and Google Scholar etc. Results: Adenosine receptors are P1 type of purinergic receptors and belong to G protein-coupled receptors (GPCRs), which is the largest family of integral membrane bound proteins receptors. Adenosine receptors are further classified into four subclasses known as A1, A2A, A2B, and A3. All four subclasses are being mediated by extracellular adenosine and perform a key role in a wide range of physiological functions such as immune system modulation, angiogenesis and sleep regulation. Adenosine receptors are thought to play a significant role in many pathophysiological conditions including cardiovascular disorders such as ischemia and thrombosis and novel corona virus infection making it a key target against these disorders. Conclusion: We suggest that modulation of adenosine receptor activity could increase the regenerative phase in these disorders by increasing the proliferation and differentiation rates of damaged tissue.

TALQUETAMAB, A G PROTEIN-COUPLED RECEPTOR FAMILY C GROUP 5 MEMBER D X CD3 BISPECIFIC ANTIBODY, IN PATIENTS WITH RELAPSED/REFRACTORY MULTIPLE MYELOMA (RRMM): UPDATED PHASE 1 RESULTS FROM MONUMENTAL-1

G protein-coupled receptor family C group 5 member D (GPRC5D) has limited expression in healthy human tissue but is highly expressed in malignant plasma cells, making it a promising target for immunotherapy approaches for MM. Talquetamab (JNJ-64407564) is a first-in-class bispecific antibody that binds to both GPRC5D and CD3 receptors to redirect T cells to kill MM cells. Updated and new results of talquetamab at the recommended phase 2 doses (RP2Ds) are reported (NCT03399799). Eligible patients had RRMM or were intolerant to standard therapies. Patients who were previously treated with B-cell maturation antigen (BCMA)-directed therapies were eligible. This analysis focuses on patients who received talquetamab subcutaneously (SC;range: 5.0-800 μg/kg) weekly (QW) or biweekly (Q2W) with step-up dosing. The primary objectives were to identify the RP2D (part 1) and assess talquetamab safety and tolerability at the RP2Ds (part 2). Adverse events (AEs) were graded by CTCAE v4.03;cytokine release syndrome (CRS) was graded per Lee et al 2014 criteria. Responses were investigator-assessed per IMWG criteria. As of July 19, 2021, 95 patients had received SC talquetamab. The original RP2D was 405 μg/kg SC talquetamab QW with step-up doses, and a second RP2D of 800 μg/kg SC talquetamab Q2W with step-up doses was also identified. 30 patients received 405 μg/kg QW (median 61.5 years [range 46-80];63% male;100% triple-class exposed;80% penta-drug exposed;77% triple-class refractory, 20% penta-drug refractory;30% prior BCMA-directed therapy;median follow-up [mF/U]: 7.5 mo [range 0.9-15.2]). 23 patients received 800 μg/kg Q2W (median 60.0 years [range 47-84];48% male;96% triple-class exposed;70% penta-drug exposed;65% triple-class refractory, 22% penta-drug refractory;17% prior BCMA-directed therapy;mF/U: 3.7 mo [range 0.0-12.0]). No treatment discontinuations due to AEs were reported at either RP2Ds. Most common AEs at the 405 μg/kg QW were CRS (73%;1 grade 3 CRS), neutropenia (67%;grade 3/4: 60%), and dysgeusia (60%;grade 2: 29%). Skin-related AEs occurred in 77% of patients and were all grade 1/2 (nail disorders: 30%). Infections occurred in 37% of patients (1 grade 3 COVID-19 pneumonia). Most common AEs at 800 μg/kg Q2W were CRS (78%;all grade 1/2), dry mouth (44%;all grade 1/2), and neutropenia (44%;grade 3/4: 35%). Skin-related AEs occurred in 65% of patients with grade 3 events in 13% (nail disorders: 17%). Infections occurred in 13% of patients (1 grade 3 pneumococcal sepsis). In 30 response-evaluable patients treated at 405 μg/kg QW, the overall response rate (ORR) was 70% (very good partial response or better [≥VGPR]: 57%). In 17 response-evaluable patients treated at 800 μg/ kg Q2W, the ORR was 71% (≥VGPR: 53%). Responses were durable and deepened over time with both RP2Ds (Figure). Median duration of response (DOR) was not reached at either RP2D;6-month DOR rate was 67% (95% CI: 41-84) at 405 μg/kg QW. Serum trough levels of talquetamab were comparable at both RP2Ds. Pharmacodynamic data at both RP2Ds showed peripheral T cell activation and induction of cytokines. SC talquetamab is well tolerated and highly effective at both RP2Ds. Preliminary data suggest that less frequent, higher doses of SC talquetamab do not negatively impact the safety profile. Further evaluation of talquetamab as monotherapy (phase 2;NCT04634552) and in combination with other therapies in patients with RRMM is underway. (Figure Presented) .

Dysfunctional Heteroreceptor Complexes as Novel Targets for the Treatment of Major Depressive and Anxiety Disorders.

Among mental diseases, major depressive disorder (MDD) and anxiety deserve a special place due to their high prevalence and their negative impact both on society and patients suffering from these disorders. Consequently, the development of novel strategies designed to treat them quickly and efficiently, without or at least having limited side effects, is considered a highly important goal. Growing evidence indicates that emerging properties are developed on recognition, trafficking, and signaling of G-protein coupled receptors (GPCRs) upon their heteromerization with other types of GPCRs, receptor tyrosine kinases, and ionotropic receptors such as N-methyl-D-aspartate (NMDA) receptors. Therefore, to develop new treatments for MDD and anxiety, it will be important to identify the most vulnerable heteroreceptor complexes involved in MDD and anxiety. This review focuses on how GPCRs, especially serotonin, dopamine, galanin, and opioid heteroreceptor complexes, modulate synaptic and volume transmission in the limbic networks of the brain. We attempt to provide information showing how these emerging concepts can contribute to finding new ways to treat both MDD and anxiety disorders.

High-throughput drug screening allowed identification of entry inhibitors specifically targeting different routes of SARS-CoV-2 Delta and Omicron/BA.1.

The Severe Acute Respiratory Syndrome Coronavirus type 2 (SARS-CoV-2) has continuously evolved, resulting in the emergence of several variants of concern (VOCs). To study mechanisms of viral entry and potentially identify specific inhibitors, we pseudotyped lentiviral vectors with different SARS-CoV-2 VOC spike variants (D614G, Alpha, Beta, Delta, Omicron/BA.1), responsible for receptor binding and membrane fusion. These SARS-CoV-2 lentiviral pseudoviruses were applied to screen 774 FDA-approved drugs. For the assay we decided to use CaCo2 cells, since they equally allow cell entry through both the direct membrane fusion pathway mediated by TMPRSS2 and the endocytosis pathway mediated by cathepsin-L. The active molecules which showed stronger differences in their potency to inhibit certain SARS-CoV-2 VOCs included antagonists of G-protein coupled receptors, like phenothiazine-derived antipsychotic compounds such as Chlorpromazine, with highest activity against the Omicron pseudovirus. In general, our data showed that the various VOCs differ in their preferences for cell entry, and we were able to identify synergistic combinations of inhibitors. Notably, Omicron singled out by relying primarily on the endocytosis pathway while Delta preferred cell entry via membrane fusion. In conclusion, our data provide new insights into different entry preferences of SARS-CoV-2 VOCs, which might help to identify new drug targets.

Pre-miRNA Hsa-Let-7a-2: a Novel Intracellular Partner of Angiotensin II Type 2 Receptor Negatively Regulating its Signals

G protein-coupled receptors (GPCRs) are the largest family of druggable targets, and their biological functions depend on different ligands and intracellular interactomes. Some microRNAs (miRNAs) bind as ligands to RNA-sensitive toll-like receptor 7 to regulate the inflammatory response, thereby contributing to the pathogenesis of cancer or neurodegeneration. It is unknown whether miRNAs bind to angiotensin II (Ang II) type 2 receptor (AGTR2), a critical protective GPCR in cardiovascular diseases, as ligands or intracellular interactomes. Here, screening for miRNAs that bind to AGTR2, we identified and confirmed that the pre-miRNA hsa-let-7a-2 non-competitively binds to the intracellular third loop of AGTR2. Functionally, intracellular hsa-let-7a-2 overexpression suppressed the Ang II-induced AGTR2 effects such as cAMP lowering, RhoA inhibition, and activation of Src homology 2 domain-containing protein-tyrosine phosphatase 1, whereas hsa-let-7a-2 knockdown enhanced these effects. Consistently, overexpressed hsa-let-7a-2 restrained the AGTR2-induced antiproliferation, antimigration, and proapoptosis of cells, and vasodilation of mesenteric arteries. Our findings demonstrated that hsa-let-7a-2 is a novel intracellular partner of AGTR2 that negatively regulates AGTR2-activated signals.