A Hybrid Machine Learning and Network Analysis Approach Reveals Two Parkinson’s Disease Subtypes from 115 RNA-Seq Post-Mortem Brain Samples

Precision medicine emphasizes fine-grained diagnostics, taking individual variability into account to enhance treatment effectiveness. Parkinson’s disease (PD) heterogeneity among individuals proves the existence of disease subtypes, so subgrouping patients is vital for better understanding disease mechanisms and designing precise treatment. The purpose of this study was to identify PD subtypes using RNA-Seq data in a combined pipeline including unsupervised machine learning, bioinformatics, and network analysis. Two hundred and ten post mortem brain RNA-Seq samples from PD (n = 115) and normal controls (NCs, n = 95) were obtained with systematic data retrieval following PRISMA statements and a fully data-driven clustering pipeline was performed to identify PD subtypes. Bioinformatics and network analyses were performed to characterize the disease mechanisms of the identified PD subtypes and to identify target genes for drug repurposing. Two PD clusters were identified and 42 DEGs were found (p adjusted ≤ 0.01). PD clusters had significantly different gene network structures (p < 0.0001) and phenotype-specific disease mechanisms, highlighting the differential involvement of the Wnt/β-catenin pathway regulating adult neurogenesis. NEUROD1 was identified as a key regulator of gene networks and ISX9 and PD98059 were identified as NEUROD1-interacting compounds with disease-modifying potential, reducing the effects of dopaminergic neurodegeneration. This hybrid data analysis approach could enable precision medicine applications by providing insights for the identification and characterization of pathological subtypes. This workflow has proven useful on PD brain RNA-Seq, but its application to other neurodegenerative diseases is encouraged.     

Immune Checkpoint Blockade via PD-L1 Potentiates More CD28-Based than 4-1BB-Based Anti-Carbonic Anhydrase IX Chimeric Antigen Receptor T Cells

The complete regression of clear cell renal cell carcinoma (ccRCC) obtained pre-clinically with anti-carbonic anhydrase IX (CAIX) G36 chimeric antigen receptor (CAR) T cells in doses equivalent to ≅10⁸ CAR T cells/kg renewed the potential of this target to treat ccRCC and other tumors in hypoxia. The immune checkpoint blockade (ICB) brought durable clinical responses in advanced ccRCC and other tumors. Here, we tested CD8α/4-1BB compared to CD28-based anti-CAIX CAR peripheral blood mononuclear cells (PBMCs) releasing anti-programmed cell death ligand-1 (PD-L1) IgG4 for human ccRCC treatment in vitro and in an orthotopic NSG mice model in vivo. Using a ≅10⁷ CAR PBMCs cells/kg dose, anti-CAIX CD28 CAR T cells releasing anti-PD-L1 IgG highly decrease both tumor volume and weight in vivo, avoiding the occurrence of metastasis. This antitumoral superiority of CD28-based CAR PBMCs cells compared to 4-1BB occurred under ICB via PD-L1. Furthermore, the T cell exhaustion status in peripheral CD4 T cells, additionally to CD8, was critical for CAR T cells efficiency. The lack of hepatotoxicity and nephrotoxicity upon the administration of a 10⁷ CAR PMBCs cells/kg dose is the basis for carrying out clinical trials using anti-CAIX CD28 CAR PBMCs cells releasing anti-PD-L1 antibodies or anti-CAIX 4-1BB CAR T cells, offering exciting new prospects for the treatment of refractory ccRCC and hypoxic tumors.     

Insights into Network of Hot Spots of Aggregation in Nucleophosmin 1

In a protein, point mutations associated with diseases can alter the native structure and provide loss or alteration of functional levels, and an internal structural network defines the connectivity among domains, as well as aggregate/soluble states’ equilibria. Nucleophosmin (NPM)1 is an abundant nucleolar protein, which becomes mutated in acute myeloid leukemia (AML) patients. NPM1-dependent leukemogenesis, which leads to its aggregation in the cytoplasm (NPMc+), is still obscure, but the investigations have outlined a direct link between AML mutations and amyloid aggregation. Protein aggregation can be due to the cooperation among several hot spots located within the aggregation-prone regions (APR), often predictable with bioinformatic tools. In the present study, we investigated potential APRs in the entire NPM1 not yet investigated. On the basis of bioinformatic predictions and experimental structures, we designed several protein fragments and analyzed them through typical aggrsegation experiments, such as Thioflavin T (ThT), fluorescence and scanning electron microscopy (SEM) experiments, carried out at different times; in addition, their biocompatibility in SHSY5 cells was also evaluated. The presented data clearly demonstrate the existence of hot spots of aggregation located in different regions, mostly in the N-terminal domain (NTD) of the entire NPM1 protein, and provide a more comprehensive view of the molecular details potentially at the basis of NPMc+-dependent AML.     

Properties and Potential Antiproliferative Activity of Thrombin-Binding Aptamer (TBA) Derivatives with One or Two Additional G-Tetrads

In this paper, we study the biological properties of two TBA analogs containing one and two extra G-tetrads, namely TBAG3 and TBAG4, respectively, and two further derivatives in which one of the small loops at the bottom (TBAG41S) or the large loop at the top (TBAG4GS) of the TBAG4 structure has been completely modified by replacing all loop residues with abasic site mimics. The therapeutical development of the TBA was hindered by its low thermodynamic and nuclease stability, while its potential as an anticancer/antiproliferative molecule is also affected by the anticoagulant activity, being a side effect in this case. In order to obtain suitable TBA analogs and to explore the involvement of specific aptamer regions in biological activity, the antiproliferative capability against DU 145 and MDAMB 231 cancer cell lines (MTT), the anticoagulant properties (PT), the biological degradability (nuclease stability assay) and nucleolin (NCL) binding ability (SPR) of the above described TBA derivatives have been tested. Interestingly, none of the TBA analogs exhibits an anticoagulant activity, while all of them show antiproliferative properties to the same extent. Furthermore, TBAG4 displays extraordinary nuclease stability and promising antiproliferative properties against breast cancer cells binding NCL efficiently. These results expand the range of G4-structures targeting NCL and the possibility of developing novel anticancer and antiviral drugs.     

Characterization of a Novel Splicing Variant in Acylglycerol Kinase (AGK) Associated with Fatal Sengers Syndrome

Mitochondrial functional integrity depends on protein and lipid homeostasis in the mitochondrial membranes and disturbances in their accumulation can cause disease. AGK, a mitochondrial acylglycerol kinase, is not only involved in lipid signaling but is also a component of the TIM22 complex in the inner mitochondrial membrane, which mediates the import of a subset of membrane proteins. AGK mutations can alter both phospholipid metabolism and mitochondrial protein biogenesis, contributing to the pathogenesis of Sengers syndrome. We describe the case of an infant carrying a novel homozygous AGK variant, c.518+1G>A, who was born with congenital cataracts, pielic ectasia, critical congenital dilated myocardiopathy, and hyperlactacidemia and died 20 h after birth. Using the patient’s DNA, we performed targeted sequencing of 314 nuclear genes encoding respiratory chain complex subunits and proteins implicated in mitochondrial oxidative phosphorylation (OXPHOS). A decrease of 96-bp in the length of the AGK cDNA sequence was detected. Decreases in the oxygen consumption rate (OCR) and the OCR:ECAR (extracellular acidification rate) ratio in the patient’s fibroblasts indicated reduced electron flow through the respiratory chain, and spectrophotometry revealed decreased activity of OXPHOS complexes I and V. We demonstrate a clear defect in mitochondrial function in the patient’s fibroblasts and describe the possible molecular mechanism underlying the pathogenicity of this novel AGK variant. Experimental validation using in vitro analysis allowed an accurate characterization of the disease-causing variant.     

Statin-Associated Myopathy: Emphasis on Mechanisms and Targeted Therapy

Hyperlipidemia is a major risk factor for cardiovascular morbidity and mortality. Statins are the first-choice therapy for dyslipidemias and are considered the cornerstone of atherosclerotic cardiovascular disease (ASCVD) in both primary and secondary prevention. Despite the statin-therapy-mediated positive effects on cardiovascular events, patient compliance is often poor. Statin-associated muscle symptoms (SAMS) are the most common side effect associated with treatment discontinuation. SAMS, which range from mild-to-moderate muscle pain, weakness, or fatigue to potentially life-threatening rhabdomyolysis, are reported by 10% to 25% of patients receiving statin therapy. There are many risk factors associated with patient features and hypolipidemic agents that seem to increase the risk of developing SAMS. Due to the lack of a “gold standard”, the diagnostic test for SAMS is based on a clinical criteria score, which is independent of creatine kinase (CK) elevation. Mechanisms that underlie the pathogenesis of SAMS remain almost unclear, though a high number of risk factors may increase the probability of myotoxicity induced by statin therapy. Some of these, related to pharmacokinetic properties of statins and to concomitant therapies or patient characteristics, may affect statin bioavailability and increase vulnerability to high-dose statins.     

Integration of TETRA with Satellite Networks: A Contribution to the IMT-A Vision

The paper addresses the integration architecture (I-concept) between a terrestrial technology—TETRA (TErrestrial Trunked Radio)—and satellite systems. This approach, that enhances and harmonises the features of both technologies, could provide an interesting contribution to the effectiveness of the International Mobile Telecommunications-Advanced (IMT-A) and, hence, to the 4G vision. TETRA can represent an interesting building block of an integrated network devoted to both civil and military scenarios; it meets the “suitable technological capability” requirement for integration, because it represents a consolidated terrestrial technology that can be trusted, hence focusing the integration effort on the definition, design and implementation of proper interfaces. System architectures are here proposed referring to short, medium term and long term scenarios.

Casein phosphopeptides modulate calcium uptake and apoptosis in Caco2 cells through their interaction with the TRPV6 calcium channel

Dietary calcium intake is associated with colon cancer incidence due to its ability to modulate proliferation and apoptosis, cellular function directly linked to normal or tumour cell phenotype. In milk calcium is associated with casein whose hydrolysis produces the casein phosphopeptides (CPPs). CPPs induce calcium uptake in differentiated HT-29 and Caco2 cells. The aim of the present study was to explore: (i) the interaction between CPPs and the TRPV6 calcium channel in HT-29 and Caco2 cells; (ii) CPP effects on Caco2 cell functions. By reducing the expression of TRPV6 through small interfering RNA (siRNA), a decrease in cell response to CPPs was monitored in Caco2 cells (about 56%) but not in HT-29 cells. CPPs increased apoptosis both in undifferentiated and in transfected Caco2 cells. Based on the reported involvement of TRPV6 in cancer development, the results presented for CPPs may be helpful for their consideration as functional food ingredients.     

Sourdough on quinoa and buckwheat gluten-free breads: Evaluation of autochthonous starter fermentation on bread nutritional and technological properties

The aim of this work was to evaluate the effect of two autochthonous strains of lactic acid bacteria as sourdough (SD) starters on gluten-free bread technological quality and in vitro starch hydrolysis and antioxidant activity. Two strains of Limosilactobacillus fermentum isolated from buckwheat (BW) and quinoa flours were used. SDs were prepared from wholegrain quinoa or BW flours. Bread technological properties, total polyphenol content and in vitro antioxidant activity and starch hydrolysis and dialysability were assessed. Specific bread volumes were between 2.61 and 2.76 cm³ g⁻¹. Breads with quinoa-based SD had overall higher technological quality than breads made with BW SDs, shown by a softer, well-aerated crumb, with an opener crumb structure and larger air area. Crumb firming during storage was reduced up to 29% when quinoa SD was used, and up to 42% when BW SD was added. Quinoa breads showed higher polyphenols and FRAP values, whereas BW breads had significantly higher ABTS values. Total starch hydrolysis and dialysability were significantly reduced when SD was used. Starch hydrolysis was reduced up to 42% and 25% when quinoa and BW SD were used, respectively. SD application produced breads with higher technological quality and modified in vitro starch hydrolysis and antioxidant capacity. Even though these effects seemed to be influenced by pH, a biological effect was also observed.     

A Novel Concept for Highly Oriented Carbon Nanotube Composite Tapes or Fibres with High Strength and Electrical Conductivity

A new concept is described for the creation of multifunctional polymer nanocomposite tapes (or fibres) that combines high stiffness and strength with good electrical properties and a low percolation threshold of carbon nanotubes (CNTs). The concept is based on a bicomponent tape (or fibre) construction consisting of a highly oriented polymer core and a conductive polymer composite (CPC) skin based on a polymer with a lower melting temperature than the core, enabling thermal annealing of these skins to improve conductivity through a dynamic percolation process while retaining the properties of the core and hence those of the tape (or fibre). The percolation threshold in the CPC skins of the highly drawn conductive bicomponent tapes could be decreased from 5.3 to 1.1 wt.‐% after annealing.