Impact of Early Antiviral Therapy on SARS-CoV-2 Clearance Time in High-Risk COVID-19 subjects: A Propensity Score Matching Study.

BACKGROUND: Effective treatments for COVID-19 are needed to mitigate disease progression and reduce the burden on healthcare systems. This study investigates the impact of early treatments on SARS-CoV-2 viral shedding duration among high-risk individuals with mild symptoms. METHODS: A single-centre, retrospective observational study was conducted at Luigi Sacco Hospital in Milan from December 2021 to March 2023. Hospitalized and non-hospitalized adults with a confirmed SARS-CoV-2 infection and at high-risk of disease progression were enrolled. Unadjusted and adjusted negative binomial regression models and a Random Forest regression model were performed before and after matching subjects based on their propensity of being treated or not. RESULTS: Results from 518 subjects (428 treated and 90 untreated) revealed a significant reduction in SARS-CoV-2 viral shedding duration among those who received early treatment compared to untreated individuals. Propensity score matching and multivariable regression analyses confirmed this finding. Early treatment significantly reduced the risk of COVID-19-related hospitalization and pneumonia development. Subgroup analysis identified COPD as a potential factor influencing effectiveness of early treatments. CONCLUSIONS: Early treatments play a crucial role in reducing SARS-CoV-2 viral shedding and preventing disease progression among high-risk individuals. Shorter viral shedding duration also contributes to improved healthcare resource utilization and infection control measures.

COVID-19 Vaccine: A Potential Risk Factor for Accelerating the Onset of Bullous Pemphigoid.

Bullous pemphigoid (BP) is the most common autoimmune bullous disease, whose main autoantigens are hemidesmosomal components BP180 and BP230. Although recent studies found no association between COVID-19 vaccines and BP, since mass vaccinations started, more than 90 vaccine-associated BP cases have been reported. To find an agreement among real-life clinical observations and recent epidemiologic data, we further investigated this topic. A total of 64 patients with BP onset in 2021 were demographically, clinically, and serologically characterized: 14 (21.9%) vaccine-associated patients (VA) developed BP within 5 weeks from the first/second vaccine dose. VA and vaccine-non-associated (VNA) patients had similar demographics and clinical and immunological characteristics. Noteworthy, the monthly distribution of BP onset during mass vaccinations paralleled vaccine administration to the elderly in the same catchment area. Additionally, in 2021, BP onsets in April-May and June-July significantly increased (p = 0.004) and declined (p = 0.027), respectively, compared to the three years before vaccination campaigns (2018-2020). Interestingly, VA and VNA patients showed statistically significant differences in the use of inhalers and diuretics. Our findings suggest that the COVID-19 vaccine may constitute an accelerating factor that, together with other triggering factors, could act in genetically predisposed individuals with possible sub-clinical autoreactivity against BP antigens, slightly accelerating BP onset.

Epidemiology and Clinical Insights of Catheter-Related Candidemia in Non-ICU Patients with Vascular Access Devices.

INTRODUCTION: Vascular access devices (VADs), namely peripheral VADs (PVADs) and central venous VADs (CVADs), are crucial in both intensive care unit (ICU) and non-ICU settings. However, VAD placement carries risks, notably catheter-related bloodstream infections (CRBSIs). Candida spp. is a common pathogen in CRBSIs, yet its clinical and microbiological characteristics, especially in non-ICU settings, are underexplored. METHODS: We conducted a monocentric, retrospective observational study at Luigi Sacco Hospital from 1 May 2021 to 1 September 2023. We reviewed medical records of non-ICU adult patients with CVADs and PVADs. Data on demographics, clinical and laboratory results, VAD placement, and CRBSI occurrences were collected. Statistical analysis compared Candida spp. CRBSI and bacterial CRBSI groups. RESULTS: Out of 1802 VAD placements in 1518 patients, 54 cases of CRBSI were identified, and Candida spp. was isolated in 30.9% of episodes. The prevalence of CRBSI was 3.05%, with Candida spp. accounting for 0.94%. Incidence rates were 2.35 per 1000 catheter days for CRBSI, with Candida albicans and Candida non-albicans at 0.47 and 0.26 per 1000 catheter days, respectively-patients with Candida spp. CRBSI had more frequent SARS-CoV-2 infection, COVID-19 pneumonia, and hypoalbuminemia. CONCLUSIONS: During the COVID-19 pandemic, Candida spp. was a notable cause of CRBSIs in our center, underscoring the importance of considering Candida spp. in suspected CRBSI cases, including those in non-ICU settings and in those with PVADs.

Exploring early COVID-19 therapies, variants, and viral clearance dynamics: Insights from a high-risk outpatients study.

This retrospective observational study investigates the impact of early COVID-19 therapies, including antivirals and monoclonal antibodies (mAbs), on time to achieve negative swab results in high-risk outpatients infected with specific Omicron sublineages. The study enrolled 104 patients from Luigi Sacco Hospital in Milan between December 2021 and March 2023, categorizing them based on the Omicron sublineage they were infected with (BA.1, BA.2, BA.4/BA.5) and the early treatment they received (antivirals or mAbs). Key data collected included demographic and clinical characteristics, initial and follow-up cycle threshold (Ct) values from qPCR tests, and the interval between swabs. The median age of the participants was 63 years (Interquartile Range [IQR] 54.0-76.5), and 55.8% were male. Among the patients, 15 received mAbs (14.4%), and 99 received antiviral treatments (95.2%) - specifically, Paxlovid (51.9%), Molnupiravir (21.1%), and Remdesivir (12.5%). No patients required hospitalization or experienced mortality during the one-month follow-up period. Regarding Omicron sublineages, 23 patients (22.1%) were infected with BA.1, 53 (51%) with BA.2, and 28 (26.9%) with BA.4/BA.5. The median interval between the initial and follow-up swabs was 6 days (IQR 6.0-7.0). Initial Ct values had a median of 18.5 (IQR 16.5-22.1), which increased to a median of 30.5 (IQR 27.1-33.0) at follow-up, indicating a reduction in viral load. A non-significant trend suggested that patients infected with BA.2 and BA.4/BA.5 sublineages might experience a faster increase in Ct values-indicating quicker viral load reduction - compared to those infected with BA.1, regardless of treatment type. However, this trend did not achieve statistical significance (p=0.609), likely due to the limited sample size and the absence of a clear trend curve. In summary, the study did not find a significant association between specific early therapies and the time to achieve swab negativization. These findings underscore the complex dynamics of viral clearance and highlight the need for further research with larger patient cohorts to refine treatment protocols for high-risk COVID-19 patients.

Understanding the burden of antibiotic resistance: a decade of carbapenem-resistant Gram-negative bacterial infections in Italian intensive care units.

Introduction: In patients admitted to intensive care units (ICUs), Gram-negative bacteria (GNB) infections pose significant challenges due to their contribution to morbidity, mortality, and healthcare costs. During the SARS-CoV-2 pandemic, Italy witnessed a rise in healthcare-associated infections (HAIs), with GNBs involved in a substantial proportion of cases. Concerningly, carbapenem-resistant GNBs (CR-GNBs) have increased worldwide, posing therapeutic challenges. Methods: Retrospective multicentre study analysing data from over 299,000 patients admitted to Italian ICUs from 2013 to 2022. Results: The study revealed an average of 1.5 infections per patient, with HAIs peaking during the pandemic years. Ventilator associated pneumonia (VAP) emerged as the most common HAI, with Klebsiella spp. and Pseudomonas aeruginosa predominating. Alarmingly, CR-GNBs accounted for a significant proportion of infections, particularly in VAP, bloodstream infections, and intra-abdominal infections. Discussion: Our findings underscore the pressing need for enhanced infection control measures, particularly in the ICU setting, to mitigate the rising prevalence of CR-GNBs and their impact on patient outcomes. The study provides valuable insights into the epidemiology of HAIs in Italian ICUs and highlights the challenges posed by CR-GNBs, especially in the context of the SARS-CoV-2 pandemic, which exacerbated the issue and may serve as a crucial example for the management of future viral pandemics.

Histone deacetylase inhibition mitigates fibrosis-driven disease progression in recessive dystrophic epidermolysis bullosa.

BACKGROUND: Recessive dystrophic epidermolysis bullosa (RDEB) is a blistering disease caused by mutations in the gene encoding type VII collagen (C7). RDEB is associated with fibrosis, which is responsible for severe complications. The phenotypic variability observed in siblings with RDEB suggests that epigenetic modifications contribute to disease severity. Identifying epigenetic changes may help to uncover molecular mechanisms underlying RDEB pathogenesis and new therapeutic targets. OBJECTIVES: To investigate histone acetylation in RDEB skin and to explore histone deacetylase inhibitors (HDACi) as therapeutic molecules capable of counteracting fibrosis and disease progression in RDEB mice. METHODS: Acetylated histone levels were detected in human skin by immunofluorescence and in RDEB fibroblasts by enzyme-linked immunosorbent assay (ELISA). The effects of givinostat and valproic acid (VPA) on RDEB fibroblast fibrotic behaviour were assessed by a collagen-gel contraction assay, Western blot and immunocytofluorescence for α-smooth muscle actin, and ELISA for released transforming growth factor (TGF)-ß1. RNA sequencing was performed in HDACi- and vehicle-treated RDEB fibroblasts. VPA was systemically administered to RDEB mice and effects on overt phenotype were monitored. Fibrosis was investigated in the skin using histological and immunofluorescence analyses. Eye and tongue defects were examined microscopically. Mass spectrometry proteomics was performed on skin protein extracts from VPA-treated RDEB and control mice. RESULTS: Histone acetylation decreases in RDEB skin and primary fibroblasts. RDEB fibroblasts treated with HDACi lowered fibrotic traits, including contractility, TGF-ß1 release and proliferation. VPA administration to RDEB mice mitigated severe manifestations affecting the eyes and paws. These effects were associated with fibrosis inhibition. Proteomic analysis of mouse skin revealed that VPA almost normalized protein sets involved in protein synthesis and immune response, processes linked to the increased susceptibility to cancer and bacterial infections seen in people with RDEB. CONCLUSIONS: Dysregulated histone acetylation contributes to RDEB pathogenesis by facilitating the progression of fibrosis. Repurposing of HDACi could be considered for disease-modifying treatments in RDEB.

Recessive dystrophic epidermolysis bullosa (or 'RDEB') is a rare skin disease that affects fewer than 5,000 people in the USA. A similar number of people in Europe are affected. RDEB is caused by mutations in the gene that controls the production of a protein called 'type VII collagen' (or 'C7'). A shortage of C7 causes fragile skin that blisters. In severe forms of RDEB, wounds heal slowly and can even affect a person's life expectancy. Differences in the disease are common in people (even identical twins) with RDEB who have similar levels of C7. This suggests that how severe the disease is could be affected by molecular processes that control other genes. Understanding these processes may help us to find treatments for RDEB. This study was done in Italy, in collaboration with centres in Germany and Switzerland. We wanted to see whether a chemical modification called 'histone acetylation' (which influences gene activity) is different in RDEB and whether it can be targeted by a specific treatment. We found that histone acetylation is reduced in RDEB skin and in skin cells grown in the lab called 'fibroblasts'. When we increased histone acetylation in fibroblasts with two drugs called givinostat and valproic acid, the amount of scar tissue produced decreased. This is important because scar tissue can lead to severe symptoms. We carried out more experiments to study the effects of givinostat and valproic acid in mice with RDEB. We found that valproic acid reduces the severity of RDEB by decreasing the disease's harmful effects and reducing the amount of scar tissue. Our findings suggest that abnormal histone acetylation contributes to the scar tissue seen in RDEB. Our study shows that valproic acid could be useful in treating the scarring seen in RDEB and in reducing the effects of the disease. As this drug is used to treat other diseases, there could be potential for rapid repurposing of it for RDEB.

Evaluating the Clinical Meaning of Dermatology Life Quality Index Scores Between Different Phenotypes of Atopic Dermatitis in Patients Before and After Biologic Therapy With Dupilumab.

Background and Objective: Atopic Dermatitis (AD) is the most prevalent inflammatory skin disorder resulting in an intense impact on patients quality of life. The aim of this study is to evaluate the clinical meaning of the DLQI scores documented between different phenotypes of AD patients under biologic therapy with Dupilumab. Method: We conducted a retrospective analysis of 209 patients with AD treated with Dupilumab for 2 years. These patients were categorized into different clinical phenotypes. Severity of the disease was assessed by using the Eczema Area and Severity Index (EASI), Numerical Scale Rating (NRS) for sleep (NRS sleep), pruritus (NRS pruritus) and Dermatology Life Quality Index (DLQI) at baseline and subsequently at 4,12 and 24 months. Results: Our results show that the higher DLQI scores (mean: 18.6, range:9-30) achieved at T0 are associated with a prurigo nodularis AD pattern, while after 24 months (T3) of therapy with Dupilumab, the worst quality of life index results were reported in Flexural and Head-Neck combined clinical phenotypes. Conclusions: Quality of life is probably what matters most as an overall endpoint in AD. Assessing the clinical meaning of DLQI scores across different AD phenotypes could be a further aid when considering decision making factors in patient management.

Early administration of nirmatrelvir/ritonavir leads to faster negative SARS-CoV-2 nasal swabs than monoclonal antibodies in COVID 19 patients at high-risk for severe disease.

PURPOSE: Besides the well-established efficacy in preventing severe COVID-19, the impact of early treatments, namely antivirals and monoclonal antibodies (mAbs), on the time length to negativization of SARS-CoV-2 nasal swabs is still unclear. The aim of this study was to compare the efficacy of different early treatments in reducing the SARS-CoV-2 viral shedding, identifying a single drug that might potentially lead to a more rapid negativization of SARS-CoV-2 nasal swab. METHODS: This was a single-centre, retrospective, observational study conducted at Ospedale Luigi Sacco in Milan. Data of high-risk COVID-19 patients who received early treatments between 23 December 2021 and March 2023 were extracted. The comparison across treatments was conducted using the Kruskall-Wallis test for continuous variables. Dunn's test with Bonferroni adjustment was performed for post-hoc comparisons of days to negativization. Secondly, a negative binomial regression adjusted for age, sex, number of comorbidities, immunosuppression, and SARS-CoV-2 vaccination status was implemented. RESULTS: Data from 428 patients receiving early treatments were collected. The majority were treated with Nirmatrelvir/Ritonavir and were affected by SARS-CoV-2 Omicron infection with BA.2 sublineage. The median length time to SARS-CoV-2 nasal swab negativization was 9 days [IQR 7-13 days]. We found that Nirmatrelvir/Ritonavir determined a significant decrease of the length time to SARS-CoV-2 nasal swab negativization compared to mAbs (p = 0.003), but not compared to Remdesivir (p = 0.147) and Molnupiravir (p = 0.156). CONCLUSION: Our findings highlight the importance of promptly treating high-risk COVID-19 patients with Nirmatrelvir/Ritonavir, as it also contributes to achieving a faster time to negative SARS-CoV-2 nasal swabs.

Identification of immunological patterns characterizing immune-related psoriasis reactions in oncological patients in therapy with anti-PD-1 checkpoint inhibitors.

Introduction: Immunotherapy with biologics targeting programmed cell death protein-1 (PD-1) is highly effective in the treatment of various malignancies. Nevertheless, it is frequently responsible for unexpected cutaneous manifestations, including psoriasis-like dermatitis. The pathogenesis of anti-PD-1-induced psoriasis has yet to be clarified, even though it is plausible that some innate and adaptive immunity processes are in common with canonical psoriasis. The genetic predisposition to psoriasis of patients could also be a contributing factor. Here, we investigated the immunological and genetic profiles of two patients with metastatic melanoma and one patient affected by lung cancer, who developed severe psoriasis after receiving anti-PD-1 nivolumab therapy. Methods: The immune patterns of the three patients were compared with those detectable in classical, chronic plaque-type psoriasis or paradoxical psoriasis induced by anti-TNF-α therapy, mostly sustained by adaptive and innate immunity processes, respectively. Therefore, immunohistochemistry and mRNA analyses of innate and adaptive immunity molecules were conducted on skin biopsy of patients. Genetic analysis of polymorphisms predisposing to psoriasis was carried out by NGS technology. Results: We found that anti-PD-1-induced psoriasis showed immunological features similar to chronic psoriasis, characterized by the presence of cellular players of adaptive immunity, with abundant CD3+, CD8+ T cells and CD11c+ dendritic cells infiltrating skin lesions, and producing IL-23, IL-6, TNF-α, IFN-γ and IL-17. On the contrary, a lower number of innate immunity cells (BDCA2+ plasmacytoid dendritic cells, CD15+ neutrophils, CD117+ mast cells) and reduced IFN-α/ß, lymphotoxin (LT)-α/ß, were observed in anti-PD-1-induced psoriasis lesions, as compared with anti-TNF-α-induced paradoxical psoriasis. Importantly, the disintegrin and metalloprotease domain containing thrombospondin type 1 motif-like 5 (ADAMTSL5) psoriasis autoantigen was significantly upregulated in psoriasis lesions of anti-PD-1-treated patients, at levels comparable with chronic plaque-type psoriasis. Finally, NGS analysis revealed that all patients carried several allelic variants in psoriasis susceptibility genes, such as HLA-C, ERAP1 and other genes of the major psoriasis susceptibility PSORS1 locus. Discussion: Our study showed that adaptive immunity predominates over innate immunity in anti-PD-1-induced psoriasis lesions, consistently with the local ADAMTSL5 overexpression. The presence of numerous SNPs in psoriasis susceptibility genes of the three patients also suggested their strong predisposition to the disease.

Clinical characteristics and outcomes of vaccinated patients hospitalised with SARS-CoV-2 breakthrough infection: Multi-IPV, a multicentre study in Northern Italy.

BACKGROUND: Despite the well-known efficacy of anti-COVID-19 vaccines in preventing morbidity and mortality, several vaccinated individuals are diagnosed with SARS-CoV-2 breakthrough infection, which might require hospitalisation. This multicentre, observational, and retrospective study aimed to investigate the clinical characteristics and outcomes of vaccinated vs. non-vaccinated patients, both hospitalised with SARS-CoV-2 infection in 3 major hospitals in Northern Italy. METHODS: Data collection was retrospective, and paper and electronic medical records of adult patients with a diagnosed SARS-CoV-2 infection were pseudo-anonymised and analysed. Vaccinated and non-vaccinated individuals were manually paired, using a predetermined matching criterion (similar age, gender, and date of hospitalisation). Demographic, clinical, treatment, and outcome data were compared between groups differing by vaccination status using Pearson's Chi-square and Mann-Whitney tests. Moreover, multiple logistic regression analyses were performed to assess the impact of vaccination status on ICU admission or intra-hospital mortality. RESULTS: Data from 360 patients were collected. Vaccinated patients presented with a higher prevalence of relevant comorbidities, like kidney replacement therapy or haematological malignancy, despite a milder clinical presentation at the first evaluation. Non-vaccinated patients required intensive care more often than their vaccinated counterparts (8.8% vs. 1.7%, p = 0.002). Contrariwise, no difference in intra-hospital mortality was observed between the two groups (19% vs. 20%, p = 0.853). These results were confirmed by multivariable logistic regressions, which showed that vaccination was significantly associated with decreased risk of ICU admission (aOR=0.172, 95%CI: 0.039-0.542, p = 0.007), but not of intra-hospital mortality (aOR=0.996, 95%CI: 0.582-1.703, p = 0.987). CONCLUSIONS: This study provides real-world data on vaccinated patients hospitalised with COVID-19 in Northern Italy. Our results suggest that COVID-19 vaccination has a protective role in individuals with higher risk profiles, especially regarding the need for ICU admission. These findings contribute to our understanding of SARS-CoV-2 infection outcomes among vaccinated individuals and emphasise the importance of vaccination in preventing severe disease, particularly in those countries with lower first-booster uptake rates.

One-step NGS molecular analysis of the CFTR gene on newborn dried blood spots gives a higher diagnostic sensitivity in affected and carrier subjects: A pilot study.

BACKGROUND: Cystic fibrosis is the most common hereditary recessive disease with an incidence of about 1:2500/3000. It has long been known that the disease is caused by deleterious mutations in the CFTR gene. Conventionally, the disease is diagnosed in several phases. The analysis of all the possible disease-causing molecular alterations is time consuming and may not lead to a definitive diagnosis in several cases. Consequently, we propose, in this paper, a rapid sequencing method that, in a single procedural asset, reveals the presence of small mutations and also the copy number variants (CNVs) from the DNA extracted from the Guthrie Spot. MATERIALS AND METHODS: We first sequenced 30 blood spots, then we validated the method on 100 spots that underwent both traditional analyses and this complete NGS sequencing, and lastly, we tested the strategy on patients who normally do not reach the molecular sequencing step because of low level of Immune-Reactive Trypsinogen. RESULTS: Using this procedure, we identified 97 variants in the CFTR gene of our samples and 6 CNVs. Notably, the significant data were obtained in the group of patients with borderline or negative IRT who routinely would not undergo molecular testing. We also identified 6 carriers of "disease-causing" variants. CONCLUSION: This method is very robust. Indeed, there was a 100% concordance with Sanger sequencing validation, and 6 mutation carriers were identified who normally escaped molecular testing with actual conventional procedure. There were also 3 duplications of almost the entire gene in heterozygosity, which were not seen with traditional methods. Being quick and easy to perform, we suggest that complete sequencing of the CFTR gene, as in this study be considered for all newborns.

Homologous Recombination Deficiency (HRD) Scoring, by Means of Two Different Shallow Whole-Genome Sequencing Pipelines (sWGS), in Ovarian Cancer Patients: A Comparison with Myriad MyChoice Assay.

High-grade serous ovarian cancer (HGSOC) patients carrying the BRCA1/2 mutation or deficient in the homologous recombination repair system (HRD) generally benefit from treatment with PARP inhibitors. Some international recommendations suggest that BRCA1/2 genetic testing should be offered for all newly diagnosed epithelial ovarian cancer, along with HRD assessment. Academic tests (ATs) are continuously under development, in order to break down the barriers patients encounter in accessing HRD testing. Two different methods for shallow whole-genome sequencing (sWGS) were compared to the reference assay, Myriad. All these three assays were performed on 20 retrospective HGSOC samples. Moreover, HRD results were correlated with the progression-free survival rate (PFS). Both sWGS chemistries showed good correlation with each other and a complete agreement, even when compared to the Myriad score. Our academic HRD assay categorized patients as HRD-Deficient, HRM-Mild and HRN-Negative. These three groups were matched with PFS, providing interesting findings in terms of HRD scoring and months of survival. Both our sWGS assays and the Myriad test correlated with the patient's response to treatments. Finally, our AT confirms its capability of determining HRD status, with the advantage of being faster, cheaper, and easier to carry out. Our results showed a prognostic value for the HRD score.

The performance of multi-gene panels for breast/ovarian cancer predisposition.

BRCA1 and BRCA2 are the most mutated genes in breast cancer. We analyzed 48 breast cancer subjects using two methods that differ in terms of number of genes investigated and strategy used (primers: Panel A - 12 genes - vs probes: Panel B - 48 genes). Both the panels and procedures identified "pathogenic" or "likely pathogenic" variants in TP53, ATM, CHEK2 and BARD1 besides BRCA1 and BRCA2. Panel B identified two other putatively pathogenic variants in RNASEL and in RAD50. Identification of variants other than the BRCA genes can be useful in patient management. A total of 121 variants were distributed within the 12 genes and were correctly detected by both panels. However, the number of calls without divergence, namely ± 0.10 difference of allelic frequency, was 78.3%, while calls with a divergence below 0.10 was 16.7%, thus indicating that only 5% (n = 275) of 5,412 calls had a divergence above 0.10. Although these panels differ from each other, both are useful in different situations, particularly when patients should be tested for genes other than BRCA1/2 (as occurs in patients affected by a so called hereditary syndrome) or for therapeutic purposes.

Nasal Microbiome in COVID-19: A Potential Role of Corynebacterium in Anosmia.

The evolution and the development of the symptoms of Coronavirus disease 19 (COVID-19) are due to different factors, where the microbiome plays a relevant role. The possible relationships between the gut, lung, nasopharyngeal, and oral microbiome with COVID-19 have been investigated. We analyzed the nasal microbiome of both positive and negative SARS-CoV-2 individuals, showing differences in terms of bacterial composition in this niche of respiratory tract. The microbiota solution A (Arrow Diagnostics) was used to cover the hypervariable V1-V3 regions of the bacterial 16S rRNA gene. MicrobAT Suite and MicrobiomeAnalyst program were used to identify the operational taxonomic units (OTUs) and to perform the statistical analysis, respectively. The main taxa identified in nasal microbiome of COVID-19 patients and in Healthy Control subjects belonged to three distinct phyla: Proteobacteria (HC = 14%, Cov19 = 35.8%), Firmicutes (HC = 28.8%, Cov19 = 30.6%), and Actinobacteria (HC = 56.7%, Cov19 = 14.4%) with a relative abundance > 1% in all groups. A significant reduction of Actinobacteria in Cov19 group compared to controls (P < 0.001, FDR = 0.01) was found. The significant reduction of Actinobacteria was identified in all taxonomic levels down to the genus (P < 0.01) using the ANOVA test. Indeed, a significantly reduced relative abundance of Corynebacterium was found in the patients compared to healthy controls (P = 0.001). Reduced abundance of Corynebacterium has been widely associated with anosmia, a common symptom of COVID-19 as suffered from our patients. Contrastingly, the Corynebacterium genus was highly represented in the nasal mucosa of healthy subjects. Further investigations on larger cohorts are necessary to establish functional relationships between nasal microbiota content and clinical features of COVID-19.

Allelic Variants of HLA-C Upstream Region, PSORS1C3, MICA, TNFA and Genes Involved in Epidermal Homeostasis and Barrier Function Influence the Clinical Response to Anti-IL-12/IL-23 Treatment of Patients with Psoriasis.

Several biologic therapies have been developed to treat moderate-to-severe psoriasis, with patients exhibiting different clinical benefits, possibly due to the heterogeneity of pathogenic processes underlying their conditions. Ustekinumab targets the IL-12/IL-23-p40 subunit and inhibits type-1 and type-17 T-cell responses. Although ustekinumab is effective as both short- and long-term treatment, therapeutic response varies considerably among patients. Ustekinumab biosimilars will be commercialized in the very next future, likely broadening the use of this drug in the treatment of psoriasis patients. Our pharmacogenomic study evaluated the influence of 417 single-nucleotide polymorphisms (SNPs) in psoriasis-risk alleles on the clinical response to ustekinumab in a cohort of 152 patients affected by moderate-to-severe plaque-type psoriasis. Differences in SNP pattern characterizing HLA-Cw6+ or HLA-Cw6- patients, showing high or low responses to ustekinumab, were also analysed. We identified twelve SNPs in HLA-C upstream region (rs12189871, rs4406273, rs9348862 and rs9368670), PSORS1C3 (rs1265181), MICA (rs2523497), LCE3A-B intergenic region (rs12030223, rs6701730), CDSN (rs1042127, rs4713436), CCHCR1 (rs2073719) and in TNFA (rs1800610) genes associated with excellent response to ustekinumab. We also found that HLA-Cw6+ and HLA-Cw6- patients carried out distinct patterns of SNPs associated with different clinical responses. The assessment of HLA-C alleles, together with other genetic variants, could be helpful for defining patients who better benefit from anti-IL-12/IL-23 therapy.

TMBleR: a bioinformatic tool to optimize TMB estimation and predictive power.

MOTIVATION: Tumor mutational burden (TMB) has been proposed as a predictive biomarker for immunotherapy response in cancer patients, as it is thought to enrich for tumors with high neoantigen load. TMB assessed by whole-exome sequencing is considered the gold standard but remains confined to research settings. In the clinical setting, targeted gene panels sampling various genomic sizes along with diverse strategies to estimate TMB were proposed and no real standard has emerged yet. RESULTS: We provide the community with TMBleR, a tool to measure the clinical impact of various strategies of panel-based TMB measurement. AVAILABILITY AND IMPLEMENTATION: R package and docker container (GPL-3 Open Source license): https://acc-bioinfo.github.io/TMBleR/. Graphical-user interface website: https://bioserver.ieo.it/shiny/app/tmbler. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

PI3Kδ Sustains Keratinocyte Hyperproliferation and Epithelial Inflammation: Implications for a Topically Druggable Target in Psoriasis.

The phosphatidylinositol 3-kinase (PI3K)-dependent signaling pathway is aberrantly activated in psoriatic lesions and contributes to disease pathogenesis. Among PI3Ks enzymes, PI3Kα, ß, and δ isoforms are known to bind the p85 regulatory subunit and mediate activation of AKT and other downstream effectors. In this study, we deepened our understanding of the expression and function of PI3Kδ in skin lesions of patients affected by psoriasis. For the first time, we found that PI3Kδ is overexpressed in psoriatic plaques, and its expression is not only confined to infiltrating immune cells but also accumulates in proliferating keratinocytes of the epidermal basal layer. We investigated the function of PI3Kδ in psoriatic skin by evaluating the impact of seletalisib, a newly developed selective PI3Kδ inhibitor, in both in vitro and in vivo experimental models of psoriasis. Of note, we found that PI3Kδ sustains keratinocyte hyperproliferation and impaired terminal differentiation induced by IL-22, as well as induces epithelial inflammation and resistance to apoptosis mediated by TNF-α in human keratinocytes. Mechanistically, PI3Kδ promotes PDK1 phosphorylation and signals through AKT-dependent or -independent pathways. It is worth mentioning that PI3Kδ inhibition by seletalisib attenuates the severity of psoriasiform phenotype induced in the Imiquimod-induced mouse model of psoriasis by restoring the physiological proliferation and differentiation programs in epidermal keratinocytes and contrasting the cutaneous inflammatory responses. Therefore, we suggest PI3Kδ as a potential topically druggable target in psoriasis and skin diseases characterized by epidermal hyperproliferation and skin inflammation.

Enhanced NAMPT-Mediated NAD Salvage Pathway Contributes to Psoriasis Pathogenesis by Amplifying Epithelial Auto-Inflammatory Circuits.

Dysregulated cross-talk between immune cells and epithelial compartments is responsible for the onset and amplification of pathogenic auto-inflammatory circuits occurring in psoriasis. NAMPT-mediated NAD salvage pathway has been recently described as an immunometabolic route having inflammatory function in several disorders, including arthritis and inflammatory bowel diseases. To date, the role of NAD salvage pathway has not been explored in the skin of patients affected by psoriasis. Here, we show that NAD content is enhanced in lesional skin of psoriatic patients and is associated to high NAMPT transcriptional levels. The latter are drastically reduced in psoriatic skin following treatment with the anti-IL-17A biologics secukinumab. We provide evidence that NAMPT-mediated NAD+ metabolism fuels the immune responses executed by resident skin cells in psoriatic skin. In particular, intracellular NAMPT, strongly induced by Th1/Th17-cytokines, acts on keratinocytes by inducing hyper-proliferation and impairing their terminal differentiation. Furthermore, NAMPT-mediated NAD+ boosting synergizes with psoriasis-related cytokines in the upregulation of inflammatory chemokines important for neutrophil and Th1/Th17 cell recruitment. In addition, extracellular NAMPT, abundantly released by keratinocytes and dermal fibroblasts, acts in a paracrine manner on endothelial cells by inducing their proliferation and migration, as well as the expression of ICAM-1 membrane molecule and chemokines important for leukocyte recruitment into inflamed skin. In conclusion, our results showed that NAMPT-mediated NAD salvage pathway contributes to psoriasis pathogenic processes by amplifying epithelial auto-inflammatory responses in psoriasis.

Nasopharyngeal Microbiome Signature in COVID-19 Positive Patients: Can We Definitively Get a Role to Fusobacterium periodonticum?

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) caused the pandemic Coronavirus Disease 2019 (COVID-19). This virus is highly transmissible among individuals through both droplets and aerosol leading to determine severe pneumonia. Among the various factors that can influence both the onset of disease and the severity of its complications, the microbiome composition has also been investigated. Recent evidence showed the possible relationship between gut, lung, nasopharyngeal, or oral microbiome and COVID-19, but very little is known about it. Therefore, we aimed to verify the relationships between nasopharyngeal microbiome and the development of either COVID-19 or the severity of symptoms. To this purpose, we analyzed, by next generation sequencing, the hypervariable V1-V2-V3 regions of the bacterial 16S rRNA in nasopharyngeal swabs from SARS-CoV-2 infected patients (n=18) and control (CO) individuals (n=12) using Microbiota solution A (Arrow Diagnostics). We found a significant lower abundance of Proteobacteria and Fusobacteria in COVID-19 patients in respect to CO (p=0.003 and p<0.0001, respectively) from the phylum up to the genus (p<0.001). The Fusobacterium periodonticum (FP) resulted as the most significantly reduced species in COVID-19 patients respect to CO. FP is reported as being able to perform the surface sialylation. Noteworthy, some sialic acids residues on the cell surface could work as additional S protein of SARS-CoV-2 receptors. Consequently, SARS-CoV-2 could use sialic acids as receptors to bind to the epithelium of the respiratory tract, promoting its clustering and the disease development. We can therefore speculate that the significant reduction of FP in COVID-19 patients could be directly or indirectly linked to the modulation of sialic acid metabolism. Finally, viral or environmental factors capable of interfering with sialic metabolism could determine a fall in the individual protection from SARS-CoV-2. Further studies are necessary to clarify the precise role of FP in COVID-19.

Case Report: Detection of a Novel Germline PALB2 Deletion in a Young Woman With Hereditary Breast Cancer: When the Patient's Phenotype History Doesn't Lie.

The partner and localizer of BRCA2 (PALB2) is a major BRCA2 binding partner that participates in homologous recombination repair in response to DNA double-strand breaks. Germline alterations of the PALB2 gene have recently been associated with a high risk of developing breast cancer. We investigated a 37-year-old Caucasian woman with breast cancer and family history of breast cancer using targeted next generation sequencing. A novel heterozygous deletion involving exons 5 and 6 was found in the PALB2 gene, and resulted in the production of a truncated PALB2 protein. These findings expand the mutational spectra of PALB2-associated breast cancer, and may improve the mutation-based screening and genetic diagnosis of breast cancer.