CMN Weekly (7 June 2024) - Your Weekly CRISPR Medicine News

Top picks

• Dengue lacks approved antivirals, but systemic delivery of mRNA-encoded Cas13a and guide RNAs in lipid nanoparticles shows promise. Guides against DENV2 and DENV3 were effective in vitro. A single dose of this formulation, given one-day post-infection, increased survival and reduced viral titres in mice. RNA sequencing revealed no off-target effects, highlighting mRNA-encoded Cas13's potential as a pan-DENV treatment.
• OliTag-seq, an in-cellulo assay, improves CRISPR-Cas9 off-target detection with stable oligonucleotide tagging and triple-priming amplification. It surpasses traditional methods in coverage and accuracy, especially in iPSCs. Enhanced by prolonged Cas9 expression and HDAC inhibitor treatments, OliTag-seq offers high sensitivity and practical application, making it essential for precise genome editing assessment in research and clinical environments.

Research

• Using a CRISPR-Cas9-based knockout system, researchers identified genes crucial for human mast cell differentiation. Electroporation and ribonucleoprotein techniques showed that the transcription factor MITF is essential, with its loss suppressing mast cell formation. Conversely, CITED2 was found to be dispensable despite its upregulation. This framework enables efficient gene disruption in primary human hematopoietic progenitors, uncovering key regulators of mast cell differentiation.
• A conditional AID-dead Cas9 (dCas9) system has been optimised for base editing, showing more significant edit heterogeneity than the BE4 cytosine base editor. Using this system with a custom sgRNA library, researchers identified EGFR and BRAF variants that confer resistance to EGFR inhibitors. This scalable platform aids in drug-modifying variant discovery and insights into protein structure-function relationships.
• An American study has used Random Forest classifiers to distinguish Cas9 and Cas12 proteins from non-Cas proteins based on 13,494 protein features. The models achieved 92-95% accuracy. Key features include quasi-sequence-order descriptors for Cas12 and amino acid composition for Cas9. These findings offer insights for designing Cas proteins with improved gene-editing properties, enhancing their application in correcting genetic defects.
• TALE base editors (TALEBs) convert cytidines to uracils in DNA using TALE domains and split-DddA deaminase. A high-throughput screen showed that linker nature and spacer length between TALE regions affect editing precision and bystander activity, enabling fine-tuning of editing windows and more accurate or permissive editing profiles based on surrounding bases.
• Cytosine base editors face sequence context and off-target limitations. Using AlphaFold2, researchers identified highly efficient cytidine deaminases with improved on-target ratios. These deaminases converted C-to-T at diverse sites and introduced stop codons without double-strand breaks.
• Using a genome-wide in vivo CRISPR screen, a team at McGill University Health Centre in Canada identified transforming growth factor beta-3 (TGFβ3) as an important determinant of palbociclib sensitivity in triple-negative breast cancer (TNBC). The study suggests the synergistic interaction between CDK4/6 and TGFβ3 as a potentially new combination treatment approach to TNBC.
• A new redundancy-based modular CRISPR-Cas12a synergistic activation platform (MCSAP) addresses molecular biology and engineering, extending to diagnostics, biochemistry circuits, and information storage. It works by finely regulating Cas12a activity through split DNA activators and integrates with molecular networks, performing nucleic acid detection, enzyme detection, and logic operations.

Industry

• Precision Biosciences has presented preclinical safety data for its PBGENE-HBV gene editing program that supports advancement to clinical trials as a potentially curative treatment for chronic hepatitis B. The therapy employs the company's novel proprietary ARCUS platform for in vivo gene editing.

CRISPR screens

• A phenotypic CRISPR-Cas9 scan of 17,065 genes in human cells revealed a 'proximity bias', where knockouts unexpectedly affect unrelated genes on the same chromosome arm. This bias, consistent across various conditions, is likely due to telomeric truncations influenced by cell cycle and apoptosis pathways. A correction method mitigates this bias, impacting functional genomic studies, drug-target identification, and genetic therapies.

Reviews

• The genetic engineering Swiss army knife. This brief review discusses how CRISPR-Cas9, a multifunctional molecular tool of genetic engineering, is revolutionising multiple fields.
• Nucleic acid-responsive smart systems for controlled cargo delivery. This review discusses several types of nucleic acid-responsive controlled delivery vehicles based on locks and keys, including DNA/RNA-responsive, aptamer-responsive, and CRISPR-responsive. It summarises their advantages and limitations.

Webinars

• CRISPR Medicine News and GeneHumdi present a webinar in which Dr Annarita Miccio from the Imagine Institute in Paris will discuss CRISPR Clinical Trials: current progress and future perspectives in ex vivo approaches. The webinar airs for free on Tuesday, June 11, 2024, from 15:00 to 16:00 CEST.

News from CRISPR Medicine News

• On Monday, we published a report about a major study that shed new light on mechanisms used by CRISPR-associated transposons, with commentary from study leader Guillermo Montoya (University of Copenhagen) about the impact of the study's findings on therapeutic applications.
• On Wednesday, we brought a brief update about the latest news from three gene-editing clinical trials sponsored by Cellectis, Intellia Therapeutics and Caribou Biosciences. The gene-editing therapies covered in that update are being developed to treat acute lymphoblastic leukaemia, hereditary angioedema, and second-line large B cell lymphoma, respectively.