Expanding the Molecular Spectrum of ANKRD11 Gene Defects in 33 Patients with a Clinical Presentation of KBG Syndrome

KBG syndrome (KBGS) is a neurodevelopmental disorder caused by the Ankyrin Repeat Domain 11 (ANKRD11) haploinsufficiency. Here, we report the molecular investigations performed on a cohort of 33 individuals with KBGS clinical suspicion. By using a multi-testing genomic approach, including gene sequencing, Chromosome Microarray Analysis (CMA), and RT-qPCR gene expression assay, we searched for pathogenic alterations in ANKRD11. A molecular diagnosis was obtained in 22 out of 33 patients (67%). ANKRD11 sequencing disclosed pathogenic or likely pathogenic variants in 18 out of 33 patients. CMA identified one full and one terminal ANKRD11 pathogenic deletions, and one partial duplication and one intronic microdeletion, with both possibly being pathogenic. The pathogenic effect was established by RT-qPCR, which confirmed ANKRD11 haploinsufficiency only for the three deletions. Moreover, RT-qPCR applied to six molecularly unsolved KBGS patients identified gene downregulation in a clinically typical patient with previous negative tests, and further molecular investigations revealed a cryptic deletion involving the gene promoter. In conclusion, ANKRD11 pathogenic variants could also involve the regulatory regions of the gene. Moreover, the application of a multi-test approach along with the innovative use of RT-qPCR improved the diagnostic yield in KBGS suspected patients.     

Fatty-Acid-Based Membrane Lipidome Profile of Peanut Allergy Patients: An Exploratory Study of a Lifelong Health Condition

Peanut allergy is a lifelong, increasingly prevalent, and potentially life-threatening disease burdening families and communities. Dietary, particularly polyunsaturated fatty acids (PUFAs), intakes can exert positive effects on immune and inflammatory responses, and the red blood cell (RBC) membrane lipidome contains stabilized metabolic and nutritional information connected with such responses. The fatty-acid-based membrane lipidome profile has been exploratorily evaluated in a small cohort of patients (eight males and one female, age range 4.1–21.7 years old, body mass index BMI < 25) with angioedema and/or anaphylaxis after peanut ingestion. This analysis was performed according to an ISO 17025 certified robotic protocol, isolating mature RBCs, extracting membrane lipids, and transforming them to fatty acid methyl esters for gas chromatography recognition and quantification. Comparison with a group of age- and BMI-matched healthy individuals and with benchmark interval values of a healthy population evidenced significant differences, such as higher levels of ω-6 (arachidonic acid), lower values of ω-3 eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), together with an increased ω-6/ω-3 ratio in allergic patients. A significant inverse correlation was also found between specific immunoglobulin E (IgE) levels and ω-6 di-homo-gamma-linolenic acid (DGLA) and total PUFAs. Results of this preliminary study encourage screenings in larger cohorts, also in view of precision nutrition and nutraceuticals strategies, and stimulate interest to expand basic and applied research for unveiling molecular mechanisms that are still missing and individuating treatments in chronic allergic disorders.     

Analysis of Dormancy-Associated Transcriptional Networks Reveals a Shared Quiescence Signature in Lung and Colorectal Cancer

Quiescent cancer cells (QCCs) are a common feature of solid tumors, representing a major obstacle to the long-term success of cancer therapies. We isolated QCCs ex vivo from non-small cell lung cancer (NSCLC) and colorectal cancer (CRC) xenografts with a label-retaining strategy and compared QCCs gene expression profiles to identify a shared “quiescence signature”. Principal Component Analysis (PCA) revealed a specific component neatly discriminating quiescent and replicative phenotypes in NSCLC and CRC. The discriminating component showed significant overlapping, with 688 genes in common including ZEB2, a master regulator of stem cell plasticity and epithelial-to-mesenchymal transition (EMT). Gene set enrichment analysis showed that QCCs of both NSCLC and CRC had an increased expression of factors related to stemness/self renewal, EMT, TGF-β, morphogenesis, cell adhesion and chemotaxis, whereas proliferating cells overexpressed Myc targets and factors involved in RNA metabolism. Eventually, we analyzed in depth by means of a complex network approach, both the ‘morphogenesis module’ and the subset of differentially expressed genes shared by NCSLC and CRC. This allowed us to recognize different gene regulation network wiring for quiescent and proliferating cells and to underpin few genes central for network integration that may represent new therapeutic vulnerabilities. Altogether, our results highlight common regulatory pathways in QCCs of lung and colorectal tumors that may be the target of future therapeutic interventions.     

Gut Microbiota Functional Traits,Blood pH,and Anti-GAD Antibodies Concur in the Clinical Characterization of T1D at Onset

Alterations of gut microbiota have been identified before clinical manifestation of type 1 diabetes (T1D). To identify the associations amongst gut microbiome profile, metabolism and disease markers, the 16S rRNA-based microbiota profiling and ¹H-NMR metabolomic analysis were performed on stool samples of 52 T1D patients at onset, 17 T1D siblings and 57 healthy subjects (CTRL). Univariate, multivariate analyses and classification models were applied to clinical and -omic integrated datasets. In T1D patients and their siblings, Clostridiales and Dorea were increased and Dialister and Akkermansia were decreased compared to CTRL, while in T1D, Lachnospiraceae were higher and Collinsella was lower, compared to siblings and CTRL. Higher levels of isobutyrate, malonate, Clostridium, Enterobacteriaceae, Clostridiales, Bacteroidales, were associated to T1D compared to CTRL. Patients with higher anti-GAD levels showed low abundances of Roseburia, Faecalibacterium and Alistipes and those with normal blood pH and low serum HbA_1c levels showed high levels of purine and pyrimidine intermediates. We detected specific gut microbiota profiles linked to both T1D at the onset and to diabetes familiarity. The presence of specific microbial and metabolic profiles in gut linked to anti-GAD levels and to blood acidosis can be considered as predictive biomarker associated progression and severity of T1D.     

Dissecting Immunotherapy Strategies for Small Cell Lung Cancer: Antibodies,Ionizing Radiation and CAR-T

Small cell lung cancer (SCLC) is a highly aggressive malignancy that accounts for about 14% of all lung cancers. Platinum-based chemotherapy has been the only available treatment for a long time, until the introduction of immune checkpoint inhibitors (ICIs) recently changed first-line standard of care and shed light on the pivotal role of the immune system. Despite improved survival in a subset of patients, a lot of them still do not benefit from first-line chemo-immunotherapy, and several studies are investigating whether different combination strategies (with both systemic and local treatments, such as radiotherapy) may improve patient outcomes. Moreover, research of biomarkers that may be used to predict patients’ outcomes is ongoing. In addition to ICIs, immunotherapy offers other different strategies, including naked monoclonal antibodies targeting tumor associated antigens, conjugated antibody, bispecific antibodies and cellular therapies. In this review, we summarize the main evidence available about the use of immunotherapy in SCLC, the rationale behind combination strategies and the studies that are currently ongoing in this setting, in order to give the reader a clear and complete view of this rapidly expanding topic.     

Saliva and Plasma Neutralizing Activity Induced by the Administration of a Third bnt162b2 Vaccine Dose

The BNT162b2 vaccine induces neutralizing activity (NA) in serum, but no data are available on whether a third-dose activates specific-immunity within the oral mucosa, representing the primary route of viral-entry. To carefully address this issue, we investigated if such immunity is boosted by SARS-CoV-2-infection; how long it is maintained over-time; and if it protects against the SARS-CoV-2 lineage B.1 (EU) and the emerging Delta and Omicron variants. NA was measured in plasma and saliva samples from: uninfected SARS-CoV-2-Vaccinated (SV), subjects infected prior to vaccination (SIV), and subjects who were infected after the second (SIV2) or the third (SIV3) vaccine dose. Samples were collected immediately before (T0), 15 days (T1), and 90 days (T2) post third-dose administration (SV and SIV), or 15 days post-infection (SIV2 and SIV3). In all the enrolled groups, NA in plasma and saliva: (i) was higher against EU compared to the other variants at all time-points (SV: T0 and T1, EU vs. both Delta and Omicron p < 0.001; T2 p < 0.01) (SIV: T0, EU vs. Delta p < 0.05; EU vs. Omi p < 0.01; T1 and T2 EU vs. Delta p < 0.01; EU vs. Omi p < 0.001); (ii) was boosted by the administration of the third dose; iii) declined over-time, albeit being detectable in almost all subjects at T2. The monitoring of NA over time will be important in clarifying if different NA levels may influence either acquisition or course of infection to properly plan the timing of a fourth vaccine dose administration.     

Sex Differences in Neuropathy: The Paradigmatic Case of MetFormin

As a widely prescribed anti-diabetic drug, metformin has been receiving novel attention for its analgesic potential. In the study of the complex etiology of neuropathic pain (NeP), male and female individuals exhibit quite different responses characterized by higher pain sensitivity and greater NeP incidence in women. This “gender gap” in our knowledge of sex differences in pain processing strongly limits the sex-oriented treatment of patients suffering from NeP. Besides, the current investigation of the analgesic potential of metformin has not addressed the “gender gap” problem. Hence, this study focuses on metformin and sex-dependent analgesia in a murine model of NeP induced by chronic constriction injury of the sciatic nerve. We investigated sexual dimorphism in signaling pathways involved by 7 days of metformin administration, such as changes in AMP-activated protein kinase and the positive regulation of autophagy machinery, discovering that metformin affected in a sexually dimorphic manner the immunological and inflammatory response to nerve lesion. These effects were complemented by morphological and adaptive changes occurring after peripheral nerve injury. Altogether these data can contribute to explaining a number of potential mechanisms responsible for the complete recovery from NeP found in male mice, as opposed to the failure of long-lasting recovery in female animals.     

Transthyretin Stabilization: An Emerging Strategy for the Treatment of Alzheimer’s Disease?

Transthyretin (TTR), previously named prealbumin is a plasma protein secreted mainly by the liver and choroid plexus (CP) that is a carrier for thyroid hormones (THs) and retinol (vitamin A). The structure of TTR, with four monomers rich in β-chains in a globular tetrameric protein, accounts for the predisposition of the protein to aggregate in fibrils, leading to a rare and severe disease, namely transthyretin amyloidosis (ATTR). Much effort has been made and still is required to find new therapeutic compounds that can stabilize TTR (“kinetic stabilization”) and prevent the amyloid genetic process. Moreover, TTR is an interesting therapeutic target for neurodegenerative diseases due to its recognized neuroprotective properties in the cognitive impairment context and interestingly in Alzheimer’s disease (AD). Much evidence has been collected regarding the neuroprotective effects in AD, including through in vitro and in vivo studies as well as a wide range of clinical series. Despite this supported hypothesis of neuroprotection for TTR, the mechanisms are still not completely clear. The aim of this review is to highlight the most relevant findings on the neuroprotective role of TTR, and to summarize the recent progress on the development of TTR tetramer stabilizers.     

Seminal Plasma Proteomic Biomarkers of Oxidative Stress

The prevalence of idiopathic male infertility is high, up to 75% of patients with abnormal sperm parameters. Hence, the research of its causes is mandatory. Oxidative stress (OS) can be responsible for male infertility in 30–80% of cases. In recent years, seminal plasma (SP) proteomics has developed as a useful tool to provide biomarkers of specific diseases. This systematic review aims to collect the available evidence on the changes of SP proteome in patients exposed to OS to provide possible SP biomarkers of sperm OS. To accomplish this, the following keyterms “seminal fluid proteome”, “seminal plasma proteome”, “oxidative stress”, and “sperm oxidative stress” were used and 137 records were found. Among these, 17 were finally included. Nine proteins involved with OS were found overexpressed in patients with OS. Twenty-three proteins were found differentially expressed in patients with clinical conditions associated with OS, such as varicocele, male accessory gland infection/inflammation, cigarette smoke, and obesity. These proteins do not seem to overlap among the clinical conditions taken into account. We speculate that specific SP proteins may mediate OS in different clinical conditions. Altogether, these results suggest that proteomics could help to better understand some of the molecular mechanisms involved in the pathogenesis of infertility. However, further studies are needed to identify potential biomarkers of male infertility with valuable clinical significance.