Riisedamm3891

The CRISPR-based genome editing holds immense potential to fix disease-causing mutations, however, must also handle substantial natural genetic variations between individuals. Previous studies have shown that mismatches between the single guide RNA (sgRNA) and genomic DNA may negatively impact sgRNA efficiencies and lead to imprecise specificity prediction. find more Hence, the genetic variations bring about a great challenge for designing platinum sgRNAs in large human populations. However, they also provide a promising entry for designing allele-specific sgRNAs for the treatment of each individual. The CRISPR system is rather specific, with the potential ability to discriminate between similar alleles, even based on a single nucleotide difference. Genetic variants contribute to the discrimination capabilities, once they generate a novel protospacer adjacent motif (PAM) site or locate in the seed region near an available PAM. Therefore, it can be leveraged to establish allele-specific targeting in numerous dominant human disorders, by selectively ablating the deleterious alleles. link2 So far, allele-specific CRISPR has been increasingly implemented not only in treating dominantly inherited diseases, but also in research areas such as genome imprinting, haploinsufficiency, spatiotemporal loci imaging and immunocompatible manipulations. In this review, we will describe the working principles of allele-specific genome manipulations by virtue of expanding engineering tools of CRISPR. And then we will review new advances in the versatile applications of allele-specific CRISPR targeting in treating human genetic diseases, as well as in a series of other interesting research areas. Lastly, we will discuss their potential therapeutic utilities and considerations in the era of precision medicine. © The author(s).Recent advances in the field of immunotherapy have profoundly opened up the potential for improved cancer therapy and reduced side effects. However, the tumor microenvironment (TME) is highly immunosuppressive, therefore, clinical outcomes of currently available cancer immunotherapy are still poor. Recently, nanomaterial-based strategies have been developed to modulate the TME for robust immunotherapeutic responses. In this review, the immunoregulatory cell types (cells relating to the regulation of immune responses) inside the TME in terms of stimulatory and suppressive roles are described, and the technologies used to identify and quantify these cells are provided. In addition, recent examples of nanomaterial-based cancer immunotherapy are discussed, with particular emphasis on those designed to overcome barriers caused by the complexity and diversity of TME. © The author(s).Over a decade of research has confirmed the critical role of cancer stem-like cells (CSCs) in tumor initiation, chemoresistance, and metastasis. Increasingly, CSC hierarchies have begun to be defined with some recurring themes. This includes evidence that these hierarchies are 'flexible,' with both cell state transitions and dedifferentiation events possible. These findings pose therapeutic hurdles and opportunities. Here, we review cancer stem cell hierarchies and their interactions with the tumor microenvironment. We also discuss the current therapeutic approaches designed to target CSC hierarchies and initial clinical trial results for CSC targeting agents. While cancer stem cell targeted therapies are still in their infancy, we are beginning to see encouraging results that suggest a positive outlook for CSC-targeting approaches. © The author(s).A successful matching of a PEG group size with the EPR effect for an off-to-on responsive NIR-fluorophore conjugate has been accomplished which allows two distinct in vivo tumor imaging periods, the first being the switch on during the initial tumor uptake via enhanced permeability into the ROI (as background is suppressed) and a second, later, due to enhanced retention within the tumor. Methods Software simulation (https//mihaitodor.github.io/particle_simulation/index.html), synthetic chemistry, with in vitro and in vivo imaging have been synergistically employed to identify an optimal PEG conjugate of a bio-responsive NIR-AZA fluorophore for in vivo tumor imaging. Results A bio-responsive NIR-AZA fluorophore conjugated to a 10 kDa PEG group has shown excellent in vivo imaging performance with sustained high tumor to background ratios and peak tumor emission within 24 h. Analysis of fluorescence profiles over 7 days has provided evidence for the EPR effect playing a positive role. link3 Conclusion Preclinical results show that exploiting the EPR effect by utilizing an optimized PEG substituent on a bio-responsive fluorophore may offer a means for intraoperative tumor margin delineation. The off-to-on responsive nature of the fluorophore makes tumor imaging achievable without waiting for clearance from normal tissue. © The author(s).Simultaneously targeted treatment of tumor cells and their surrounding growth-supporting immune cells is a promising strategy to reshape immunosuppressive tumor microenvironment (TME) and potentiate host innate and adaptive antitumor immune responses. Methods We designed a series of melittin-(RADA)n hybrid peptide sequences with varying self-assembling motifs of RADA and screened out a melittin-(RADA)6 peptide that has an optimal gel-formation ability and in vitro antitumor activity. Results The formed melittin-(RADA)6 (MR52) hydrogel scaffold could be loaded with a specific Ca2+/calmodulin-dependent protein kinase II (CAMKII) inhibitor, KN93, originally found to have both direct tumoricidal activity and macrophages-reprogramming ability, for potent immunotherapy against melanoma and hepatoma ascites in mice models. Our MR52 hydrogel has an interweaving nanofiber-like structure, possesses direct antitumor and controlled drug release properties, and promotes the enhanced intracellular uptake of loaded cargo. Cntitumor applications. © The author(s).Background Esophageal squamous cell carcinoma (ESCC), one of the leading causes of cancer mortality worldwide, is a heterogeneous cancer with diverse clinical manifestations. However, little is known about the epigenetic heterogeneity and its clinical relevance for this prevalent cancer. Methods We generated 7.56 Tb single-base resolution whole-genome bisulfite sequencing data for 84 ESCC and paired paraneoplastic tissues. The analysis identified inter- and intratumor DNA methylation (DNAm) heterogeneity, epigenome-wide DNAm alterations together with the functional regulators involved in the hyper- or hypomethylated regions, and their association with clinical features. We then validated the correlation between the methylation level of specific regions and clinical outcomes of 96 ESCC patients in an independent cohort. Results ESCC manifested substantial inter- and intratumor DNAm heterogeneity. The high intratumor DNAm heterogeneity was associated with lymph node metastasis and worse overall survival. Interestingly, hypermethylated regions in ESCC were enriched in promoters of numerous transcription factors, and demethylated noncoding regions related to RXR transcription factor binding appeared to contribute to the development of ESCC. Furthermore, we identified numerous DNAm alterations associated with carcinogenesis and lymph node metastasis of ESCC. We also validated three novel prognostic markers for ESCC, including one each in the promoter of CLK1, the 3' untranslated region of ZEB2, and the intergenic locus surrounded by several lncRNAs. Conclusions This study presents the first population-level resource for dissecting base-resolution DNAm variation in ESCC and provides novel insights into the ESCC pathogenesis and progression, which might facilitate diagnosis and prognosis for this prevalent malignancy. © The author(s).Background and Purpose The humoral immune response in cancer patients can be used for early detection of the disease. Autoantibodies raised against tumor-associated antigens (TAAs) are promising clinical biomarkers for reliable cancer diagnosis, prognosis, and therapy monitoring. In this study, an electrochemical disposable multiplexed immunosensing platform able to integrate difficult- and easy-to-express colorectal cancer (CRC) TAAs is reported for the sensitive determination of eight CRC-specific autoantibodies. Methods The electrochemical immunosensing approach involves the use of magnetic microcarriers (MBs) as solid supports modified with covalently immobilized HaloTag fusion proteins for the selective capture of specific autoantibodies. After magnetic capture of the modified MBs onto screen-printed carbon working electrodes, the amperometric responses measured using the hydroquinone (HQ)/H2O2 system were related to the levels of autoantibodies in plasma. Results The biosensing platform was applied to the analysis of autoantibodies against 8 TAAs described for the first time in this work in plasma samples from healthy asymptomatic individuals (n=3), and patients with high-risk of developing CRC (n=3), and from patients already diagnosed with colorectal (n=3), lung (n=2) or breast (n=2) cancer. The developed bioplatform demonstrated an improved discrimination between CRC patients and controls (asymptomatic healthy individuals and breast and lung cancer patients) compared to an ELISA-like luminescence test. Conclusions The proposed methodology uses a just-in-time produced protein in a simpler protocol, with low sample volume, and involves cost-effective instrumentation, which could be used in a high-throughput manner for reliable population screening to facilitate the detection of early CRC patients at affordable cost. © The author(s).Rationale Contactin-associated protein-like 4 (CNTNAP4) belongs to the neurexin superfamily and has critical functions in neurological development and synaptic function. Loss of CNTNAP4 in interneurons has been linked to autism, schizophrenia, and epilepsy. CNTNAP4 is also highly enriched in dopaminergic (DA) neurons in the substantia nigra (SN), however, few studies have investigated the role of CNTNAP4 in DA neurons, and whether CNTNAP4 deficiency in DA neurons contributes to Parkinson's disease (PD) remains unclear. Methods Effects of CNTNAP4 knockdown or overexpression on the DA MN9D cell line were assessed via Western blotting, immunocytochemistry, and RNA sequencing. An in vivo animal model, including CNTNAP4 knockout mice and stereotaxic injections of adeno-associated viral short-hairpin RNA with the tyrosine-hydroxylase promotor to silence CNTNAP4 in the SN, as well as the resulting physiological/behavioral effects, were evaluated via behavioral tests, Western blotting, immunohistochemistry, and transich we elucidated some of the underlying mechanisms. © The author(s).Müller glia are specialized retinal cells with stem cell properties in fish and frogs but not in mammals. Current efforts to develop gene therapies to activate mammalian Müller glia for retinal repair will require safe and effective delivery strategies for recombinant adeno-associated viruses (AAVs), vectors of choice for clinical translation. Intravitreal and subretinal injections are currently used for AAV gene delivery in the eye, but less invasive methods efficiently targeting Müller glia have yet to be developed. Methods As gene delivery strategies have been more extensively studied in the brain, to validate our vectors, we initially compared the glial tropism of AAV-PHP.eB, an AAV9 that crosses the blood-brain and blood-retinal barriers, for its ability to drive fluorescent protein expression in glial cells in both the brain and retina. We then tested the glial transduction of AAV2/8-GFAP-mCherry, a virus that does not cross blood-brain and blood-retinal barriers, for its effectiveness in transducing Müller glia in murine retinal explants ex vivo.