Search Results (16)

Recent decades have witnessed a remarkable pace of innovation and performance improvements in integrated circuits (ICs), which have become indispensable in an array of critical applications ranging from military infrastructure to personal healthcare. Meanwhile, recent developments have brought physical security to the forefront of concern, particularly considering the valuable assets handled and stored within ICs. Among the various invasive attack vectors, micro-probing attacks have risen as a particularly menacing threat. These attacks leverage advanced focused ion beam (FIB) systems to enable post-silicon secret eavesdropping and circuit modifications with minimal traceability. As an evolved variant of micro-probing attacks, reroute attacks possess the ability to actively disable built-in shielding measures, granting access to the security-sensitive signals concealed beneath. To address and counter these emerging challenges, we introduce a layout-level framework known as Detour-RS. This framework is designed to automatically assess potential vulnerabilities, offering a systematic approach to identifying and mitigating exploitable weaknesses. Specifically, we employed a combination of linear and nonlinear programming-based approaches to identify the layout-aware attack costs in reroute attempts given specific target assets. The experimental results indicate that shielded designs outperform non-shielded structures against reroute attacks. Furthermore, among the two-layer shield configurations, the orthogonal layout exhibits better performance compared to the parallel arrangement. Furthermore, we explore both independent and dependent scenarios, where the latter accounts for potential interference among circuit edit locations. Notably, our results demonstrate a substantial near 50% increase in attack cost when employing the more realistic dependent estimation approach. In addition, we also propose time and gas consumption metrics to evaluate the resource consumption of the attackers, which provides a perspective for evaluating reroute attack efforts. We have collected the results for different categories of target assets and also the average resource consumption for each via, required during FIB reroute attack. Full article

This contribution explores the potential of PFIB for the post-production circuit editing of custom ASICs. The reworking of integrated circuits with ion beam is an effective tool for testing design modifications rapidly, and in small-volume productions, it proves to be a valuable substitute for the microfabrication of chips with a revised layout, thus reducing the cost and lead time. In the case study in this work, the PFIB intervention resulted in the recovery of ~90% of defective channels in a multichannel ASIC design by disconnecting some parts of the internal circuit. This contribution describes several implemented optimization strategies and their statistical effectiveness. Full article

Fully homomorphic encryption (FHE) cryptographic systems enable limitless computations over encrypted data, providing solutions to many of today’s data security problems. While effective FHE platforms can address modern data security concerns in unsecure environments, the extended execution time for these platforms hinders their broader application. This project aims to enhance FHE systems through an efficient parallel framework, specifically building upon the existing torus FHE (TFHE) system chillotti2016faster. The TFHE system was chosen for its superior bootstrapping computations and precise results for countless Boolean gate evaluations, such as AND and XOR. Our first approach was to expand upon the gate operations within the current system, shifting towards algebraic circuits, and using graphics processing units (GPUs) to manage cryptographic operations in parallel. Then, we implemented this GPU-parallel FHE framework into a needed genomic data operation, specifically string search. We utilized popular string distance metrics (hamming distance, edit distance, set maximal matches) to ascertain the disparities between multiple genomic sequences in a secure context with all data and operations occurring under encryption. Our experimental data revealed that our GPU implementation vastly outperforms the former method, providing a 20-fold speedup for any 32-bit Boolean operation and a 14.5-fold increase for multiplications.This paper introduces unique enhancements to existing FHE cryptographic systems using GPUs and additional algorithms to quicken fundamental computations. Looking ahead, the presented framework can be further developed to accommodate more complex, real-world applications. Full article

Eriodictyol is a hydroxylated flavonoid displaying multiple pharmaceutical activities, such as antitumoral, antiviral or neuroprotective. However, its industrial production is limited to extraction from plants due to its inherent limitations. Here, we present the generation of a Streptomyces albidoflavus bacterial factory edited at the genome level for an optimized de novo heterologous production of eriodictyol. For this purpose, an expansion of the Golden Standard toolkit (a Type IIS assembly method based on the Standard European Vector Architecture (SEVA)) has been created, encompassing a collection of synthetic biology modular vectors (adapted for their use in actinomycetes). These vectors have been designed for the assembly of transcriptional units and gene circuits in a plug-and-play manner, as well as for genome editing using CRISPR-Cas9-mediated genetic engineering. These vectors have been used for the optimization of the eriodictyol heterologous production levels in S. albidoflavus by enhancing the flavonoid-3′-hydroxylase (F3’H) activity (by means of a chimera design) and by replacing three native biosynthetic gene clusters in the bacterial chromosome with the plant genes matBC (involved in extracellular malonate uptake and its intracellular activation into malonyl-CoA), therefore allowing more malonyl-CoA to be devoted to the heterologous production of plant flavonoids in this bacterial factory. These experiments have allowed an increase in production of 1.8 times in the edited strain (where the three native biosynthetic gene clusters have been deleted) in comparison with the wild-type strain and a 13 times increase in eriodictyol overproduction in comparison with the non-chimaera version of the F3′H enzyme. Full article

The layout design of analog integrated circuits has been defying all automation attempts, and it is still primarily a handcrafting process carried by circuit designers on traditional layout editing frameworks. This paper presents a toolbox based on deep learning techniques and a sturdy graphical user interface to assist designers during that process. The underlying mechanism of this toolbox relies on a simple pairwise device interaction circuit description, i.e., the circuits’ topological constraints, to propose valid floorplan solutions for block-level structures, including topologies and deep nanometer technology nodes not used for its training, at push-button speed. Despite its automatic functionalities, the toolbox is focused on explainable artificial intelligence, involving the designer in the synthesis flow via filtering and editing options over the candidate floorplan solutions. This constant state of human-machine feedback environment turns the designer aware of the impact of each device’s position change and inherent tradeoffs while suggesting subsequent moves, ultimately increasing the designers’ productivity in this time-consuming and iterative task. Finally, the toolbox is shown to instantly generate floorplans with similar or better constraint fulfilment than human designed solutions for state-of-the-art analog circuit blocks. Full article

RNAs play many essential roles in gene expression and are involved in various human diseases. Although genome editing technologies have been established, the engineering of sequence-specific RNA-binding proteins that manipulate particular cellular RNA molecules is immature, in contrast to nucleotide-based RNA manipulation technology, such as siRNA- and RNA-targeting CRISPR/Cas. Here, we demonstrate a versatile RNA manipulation technology using pentatricopeptide-repeat (PPR)-motif-containing proteins. First, we developed a rapid construction and evaluation method for PPR-based designer sequence-specific RNA-binding proteins. This system has enabled the steady construction of dozens of functional designer PPR proteins targeting long 18 nt RNA, which targets a single specific RNA in the mammalian transcriptome. Furthermore, the cellular functionality of the designer PPR proteins was first demonstrated by the control of alternative splicing of either a reporter gene or an endogenous CHK1 mRNA. Our results present a versatile protein-based RNA manipulation technology using PPR proteins that facilitates the understanding of unknown RNA functions and the creation of gene circuits and has potential for use in future therapeutics. Full article

Glioblastoma multiforme (GBM) is a lethal brain tumor, characterized by enhanced proliferation and invasion, as well as increased vascularization and chemoresistance. The expression of the hyaluronan receptor CD44 has been shown to correlate with GBM progression and poor prognosis. Here, we sought to elucidate the molecular mechanisms by which CD44 promotes GBM progression by knocking out (KO) CD44, employing CRISPR/Cas9 gene editing in U251MG cells. CD44-depleted cells exhibited an impaired proliferation rate, as shown by the decreased cell numbers, decreased Ki67-positive cell nuclei, diminished phosphorylation of CREB, and increased levels of the cell cycle inhibitor p16 compared to control cells. Furthermore, the CD44 KO cells showed decreased stemness and increased senescence, which was manifested upon serum deprivation. In stem cell-like enriched spheres, RNA-sequencing analysis of U251MG cells revealed a CD44 dependence for gene signatures related to hypoxia, the glycolytic pathway, and G2 to M phase transition. Partially similar results were obtained when cells were treated with the γ-secretase inhibitor DAPT, which inhibits CD44 cleavage and therefore inhibits the release of the intracellular domain (ICD) of CD44, suggesting that certain transcriptional responses are dependent on CD44-ICD. Interestingly, the expression of molecules involved in hyaluronan synthesis, degradation, and interacting matrix proteins, as well as of platelet-derived growth factor (PDGF) isoforms and PDGF receptors, were also deregulated in CD44 KO cells. These results were confirmed by the knockdown of CD44 in another GBM cell line, U2990. Notably, downregulation of hyaluronan synthase 2 (HAS2) impaired the hypoxia-related genes and decreased the CD44 protein levels, suggesting a CD44/hyaluronan feedback circuit contributing to GBM progression. Full article

Medical science technology has improved tremendously over the decades with the invention of robotic surgery, gene editing, immune therapy, etc. However, scientists are now recognizing the significance of ‘biological circuits’ i.e., bodily innate electrical systems for the healthy functioning of the body or for any disease conditions. Therefore, the current trend in the medical field is to understand the role of these biological circuits and exploit their advantages for therapeutic purposes. Bioelectronics, devised with these aims, work by resetting, stimulating, or blocking the electrical pathways. Bioelectronics are also used to monitor the biological cues to assess the homeostasis of the body. In a way, they bridge the gap between drug-based interventions and medical devices. With this in mind, scientists are now working towards developing flexible and stretchable miniaturized bioelectronics that can easily conform to the tissue topology, are non-toxic, elicit no immune reaction, and address the issues that drugs are unable to solve. Since the bioelectronic devices that come in contact with the body or body organs need to establish an unobstructed interface with the respective site, it is crucial that those bioelectronics are not only flexible but also stretchable for constant monitoring of the biological signals. Understanding the challenges of fabricating soft stretchable devices, we review several flexible and stretchable materials used as substrate, stretchable electrical conduits and encapsulation, design modifications for stretchability, fabrication techniques, methods of signal transmission and monitoring, and the power sources for these stretchable bioelectronics. Ultimately, these bioelectronic devices can be used for wide range of applications from skin bioelectronics and biosensing devices, to neural implants for diagnostic or therapeutic purposes. Full article

Nowadays, cancers still represent a significant health burden, accounting for around 10 million deaths per year, due to ageing populations and inefficient treatments for some refractory cancers. Immunotherapy strategies that modulate the patient’s immune system have emerged as good treatment options. Among them, the adoptive transfer of B cells selected ex vivo showed promising results, with a reduction in tumor growth in several cancer mouse models, often associated with antitumoral immune responses. Aside from the benefits of their intrinsic properties, including antigen presentation, antibody secretion, homing and long-term persistence, B cells can be modified prior to reinfusion to increase their therapeutic role. For instance, B cells have been modified mainly to boost their immuno-stimulatory activation potential by forcing the expression of costimulatory ligands using defined culture conditions or gene insertion. Moreover, tumor-specific antigen presentation by infused B cells has been increased by ex vivo antigen loading (peptides, RNA, DNA, virus) or by the sorting/ engineering of B cells with a B cell receptor specific to tumor antigens. Editing of the BCR also rewires B cell specificity toward tumor antigens, and may trigger, upon antigen recognition, the secretion of antitumor antibodies by differentiated plasma cells that can then be recognized by other immune components or cells involved in tumor clearance by antibody-dependent cell cytotoxicity or complement-dependent cytotoxicity for example. With the expansion of gene editing methodologies, new strategies to reprogram immune cells with whole synthetic circuits are being explored: modified B cells can sense disease-specific biomarkers and, in response, trigger the expression of therapeutic molecules, such as molecules that counteract the tumoral immunosuppressive microenvironment. Such strategies remain in their infancy for implementation in B cells, but are likely to expand in the coming years. Full article

Chromodomain helicase domain 8 (CHD8) is one of the most frequently mutated and most penetrant genes in the autism spectrum disorder (ASD). Individuals with CHD8 mutations show leading symptoms of autism, macrocephaly, and facial dysmorphisms. The molecular and cellular mechanisms underpinning the early onset and development of these symptoms are still poorly understood and prevent timely and more efficient therapies of patients. Progress in this area will require an understanding of “when, why and how cells deviate from their normal trajectories”. High-throughput single-cell RNA sequencing (sc-RNAseq) directly quantifies information-bearing RNA molecules that enact each cell’s biological identity. Here, we discuss recent insights from sc-RNAseq of CRISPR/Cas9-editing of Chd8/CHD8 during mouse neocorticogenesis and human cerebral organoids. Given that the deregulation of the balance between excitation and inhibition (E/I balance) in cortical and subcortical circuits is thought to represent a major etiopathogenetic mechanism in ASD, we focus on the question of whether, and to what degree, results from current sc-RNAseq studies support this hypothesis. Beyond that, we discuss the pros and cons of these approaches and further steps to be taken to harvest the full potential of these transformative techniques. Full article

DNA is the source of genetic information, and preserving its integrity is essential in order to sustain life. The genome is continuously threatened by different types of DNA lesions, such as abasic sites, mismatches, interstrand crosslinks, or single-stranded and double-stranded breaks. As a consequence, cells have evolved specialized DNA damage response (DDR) mechanisms to sustain genome integrity. By orchestrating multilayer signaling cascades specific for the type of lesion that occurred, the DDR ensures that genetic information is preserved overtime. In the last decades, DNA repair mechanisms have been thoroughly investigated to untangle these complex networks of pathways and processes. As a result, key factors have been identified that control and coordinate DDR circuits in time and space. In the first part of this review, we describe the critical processes encompassing DNA damage sensing and resolution. In the second part, we illustrate the consequences of partial or complete failure of the DNA repair machinery. Lastly, we will report examples in which this knowledge has been instrumental to develop novel therapies based on genome editing technologies, such as CRISPR-Cas. Full article

Neurodegenerative diseases are disabling and fatal neurological disorders that currently lack effective treatment. Neural stem cell (NSC) transplantation has been studied as a potential therapeutic approach and appears to exert a beneficial effect against neurodegeneration via different mechanisms, such as the production of neurotrophic factors, decreased neuroinflammation, enhanced neuronal plasticity and cell replacement. Thus, NSC transplantation may represent an effective therapeutic strategy. To exploit NSCs’ potential, some of their essential biological characteristics must be thoroughly investigated, including the specific markers for NSC subpopulations, to allow profiling and selection. Another key feature is their secretome, which is responsible for the regulation of intercellular communication, neuroprotection, and immunomodulation. In addition, NSCs must properly migrate into the central nervous system (CNS) and integrate into host neuronal circuits, enhancing neuroplasticity. Understanding and modulating these aspects can allow us to further exploit the therapeutic potential of NSCs. Recent progress in gene editing and cellular engineering techniques has opened up the possibility of modifying NSCs to express select candidate molecules to further enhance their therapeutic effects. This review summarizes current knowledge regarding these aspects, promoting the development of stem cell therapies that could be applied safely and effectively in clinical settings. Full article

In this contribution, we compare the performance of Focused Electron Beam-induced Deposition (FEBID) and Focused Ion Beam-induced Deposition (FIBID) at room temperature and under cryogenic conditions (the prefix “Cryo” is used here for cryogenic). Under cryogenic conditions, the precursor material condensates on the substrate, forming a layer that is several nm thick. Its subsequent exposure to a focused electron or ion beam and posterior heating to 50 °C reveals the deposit. Due to the extremely low charge dose required, Cryo-FEBID and Cryo-FIBID are found to excel in terms of growth rate, which is typically a few hundred/thousand times higher than room-temperature deposition. Cryo-FIBID using the W(CO)₆ precursor has demonstrated the growth of metallic deposits, with resistivity not far from the corresponding deposits grown at room temperature. This paves the way for its application in circuit edit and the fast and direct growth of micro/nano-electrical contacts with decreased ion damage. The last part of the contribution is dedicated to the comparison of these techniques with other charge-based lithography techniques in terms of the charge dose required and process complexity. The comparison indicates that Cryo-FIBID is very competitive and shows great potential for future lithography developments. Full article

Spatiotemporal patterns of gene expression depend on enhancer elements and other factors during individual development and disease progression. The rapid progress of high-throughput techniques has led to well-defined enhancer chromatin properties. Various genome-wide methods have revealed a large number of enhancers and the discovery of three-dimensional (3D) genome architecture showing the distant interacting mechanisms of enhancers that loop to target gene promoters. Whole genome sequencing projects directed at cancer have led to the discovery of substantial enhancer dysfunction in misregulating gene expression and in tumor initiation and progression. Results from genome-wide association studies (GWAS) combined with functional genomics analyses have elucidated the functional impacts of many cancer risk-associated variants that are enriched within the enhancer regions of chromatin. Risk variants dysregulate the expression of enhancer variant-associated genes via 3D genomic interactions. Moreover, these enhancer variants often alter the chromatin binding affinity for cancer-relevant transcription factors, which in turn leads to aberrant expression of the genes associated with cancer susceptibility. In this review, we investigate the extent to which these genetic regulatory circuits affect cancer predisposition and how the recent development of genome-editing methods have enabled the determination of the impacts of genomic variation and alteration on cancer phenotype, which will eventually lead to better management plans and treatment responses to human cancer in the clinic. Full article