CMN Weekly (1 September 2023) - Your Weekly CRISPR Medicine News

Top picks

• Researchers in Ireland have successfully used adenine base editing (ABE) and homology-independent targeted integration strategies to correct the cystic fibrosis-causing variant, W1282X. In primary human nasal epithelial cells homozygous for W1282X, ABE corrected 27% of alleles but with a notably lower level of bystander editing. Moreover, CFTR channel function was restored to 16% of wild-type levels.
• Whole genomic analysis reveals that CRISPR-Cas9-mediated genome editing induces atypical non-homologous off-target large structural variants, according to researchers in Taiwan. The observed large structural variants caused by CRISPR-Cas9 editing in dividing cells may result in pathogenic consequences and thus limit the usefulness of the CRISPR-Cas9 editing system for disease modelling and gene therapy.

Research

• Italian researchers have developed lipid nanoparticles (LNPs) that enable efficient and harmless ex vivo gene editing of human hematopoietic stem/progenitor cells (HSPCs). Notably, LNP-based HSPC editing dampened p53 pathway induction and supported higher clonogenic activity and similar or higher reconstitution by long-term repopulating HSPCs compared with electroporation, reaching comparable editing efficiencies.
• David Liu and colleagues have used protein evolution and engineering to generate 516–810 base pairs smaller prime editors with up to 22-fold better efficiencies than the current-generation editor PEmax. The engineering was based on observations that different reverse transcriptases specialise in various edits.
• American researchers have shown that epitope base editing of the pan-leukocyte marker CD45 in hematopoietic cells enables universal blood cancer immune therapy. Epitope-edited hematopoietic stem cells (HSCs) were protected from CAR T cells and, unlike CD45 knockout cells, could engraft, persist, and differentiate in vivo.
• A new CRISPR-based treatment for sickle cell disease - OTQ923, sponsored by Novartis Pharmaceuticals - has shown stable induction of fetal haemoglobin, reducing disease manifestations in three patients in a multicenter, phase 1-2 clinical trial. OTQ923 is an autologous CRISPR-edited hematopoietic stem cell product that targets the HBG1 and HBG2 gene promoters and increases fetal haemoglobin levels in red cell progeny.
• A potential new CRISPR-based approach to treating glioblastoma brain tumours targets the DNA methylation status of the O6-methylguanine DNA methyltransferase (MGMT) promoter - a known biomarker affecting treatment efficacy. By modifying this region, German researchers successfully downregulated MGMT expression in temozolomide (TMZ)-resistant glioblastoma cell lines, reversing their resistance to the drug.
• Researchers in Japan have established a pancreatic beta cell-specific gene knockout system based on CRISPR-Cas9 technology with AAV8-mediated gRNA delivery. The system utilises transgenic mice expressing Cas9 in pancreatic beta cells.
• A new system for cascade dynamic assembly/disassembly of DNA nanoframework (NF) enables the controlled delivery of Cas9 ribonucleoprotein (RNP). In vitro and in vivo investigations demonstrated the high gene editing efficiency in cancer cells, the hypotoxicity in normal cells, and notable antitumor efficacy in a breast cancer mouse model.

Detection

• Researchers in the US have developed a new ultrasensitive CRISPR-Cas12a detection platform that exploits the nuclease's unidirectional trans-cleaving behaviour. The authors designed a new hybrid DNA reporter as a nonoptical substrate for the CRISPR-Cas12a detection platform, which sensitively detected ssDNA targets at the picomolar level.
• Researchers in Thailand have developed a rapid diagnostic test for the early detection of leptospirosis using a combination of RPA-CRISPR/Cas12a and Leptospira IgM. The combination of the two assays exhibited significant sensitivity for detecting leptospires on various days after the onset of fever, reducing the likelihood of misdiagnosis.
• Chinese researchers report a DNA tile and invading stacking primer-assisted CRISPR–Cas12a multiple amplification strategy to construct an entropy-controlled electrochemical biosensor for detecting miRNA with tunable sensitivity and dynamic range. The strategy has shown a tunable detection limit for miRNA from 0.31 fM to 0.56 pM and a dynamic range from ∼2200-fold to ∼270,000-fold.
• American researchers report a 3D-printed composable microfluidic plate (cPlate) device that utilises miniaturised wells and microfluidic loading for a multiplexed CRISPR-Cas12a assay. The device combines loop-mediated isothermal amplification (LAMP) and CRISPR-Cas12a readout in a simple, high-throughput workflow with low reagent consumption.

Reviews

• Biotechnological advances in gene therapy of hematopoietic stem cells: Systematic review and meta-analysis. This review summarises research evaluating the most effective gene editing method in HSC for translational medicine. It concludes that although promising, recent technologies like CRISPR-Cas have yet to be ready for efficient long-term immune system restoration.
• Novel epigenetic molecular therapies for imprinting disorders. This review highlights molecular approaches of therapeutic candidates in preclinical and clinical studies for individual imprinting disorders. These include significant progress in discovering and testing small molecules, antisense oligonucleotides, and CRISPR-mediated genome editing approaches as new therapeutic strategies.
• CRISPR/Cas systems for in situ imaging of intracellular nucleic acids: Concepts and applications. This review presents various CRISPR methods for visualising intracellular genomic sequences and RNA based on their detection principles and application scenarios. Furthermore, it discusses the advantages and drawbacks of the existing CRISPR imaging methods and future research directions.
• Clustered Regularly Interspaced Short Palindromic Repeats and CRISPR-associated protein 9 System: Factors Affecting Precision Gene Editing Efficiency and Optimisation Strategies. This review provided novel insights for improving the repair efficiency of the CRISPR-Cas9 gene editing system, which may enable the development and improvement of gene editing tools.

Conferences and meetings

• Precision BioSciences will host its in vivo gene editing R&D Day on Tuesday, September 12, 2023, from 9:00-11:30 AM ET. The R&D Day event will reflect Precision's go-forward singular focus on in vivo gene editing through ARCUS, its proprietary, wholly-owned genome editing platform. The agenda will include an overview of the broad potential and differentiation of ARCUS, new preclinical data, and timelines for leading in vivo gene editing programs.