CMN Weekly (24 January 2025) - Your Weekly CRISPR Medicine News

Top picks

• Researchers in the UK have developed CRISPR MiRAGE (miRNA-activated genome editing) that enables tissue-specific gene editing by using miRNA-sensing single-guide RNAs to activate CRISPR activity in response to endogenous miRNA signatures. The approach achieved muscle-specific editing in in vivo mouse models of Duchenne muscular dystrophy, demonstrating its potential to enhance cell specificity and minimise off-target effects. MiRAGE offers a promising strategy for advancing RNA-controlled, tissue-targeted CRISPR therapies for various diseases.
• Intellia Therapeutics has dosed the first patient in a Phase 3 trial of NTLA-2002, an in vivo CRISPR-Cas therapy for hereditary angioedema (HAE). NTLA-2002 inactivates the KLKB1 gene to prevent HAE attacks. The single-dose treatment showed promising durability in earlier trials. If successful, it could become the first one-time therapy for HAE, with a U.S. launch anticipated in 2027.

Research

• Researchers have developed CRISPR-CAR-NK cells by integrating CAR sequences into the GAPDH 3'UTR locus of NK-92MI cells, achieving precise gene insertion and endogenous regulation of CAR expression. These cells showed superior anti-tumour activity, enhanced receptor expression, and reduced metabolic reliance compared to lentiviral CAR-NKs. Their distinct PI3K-AKT independence offers potential synergy with AKT inhibitors, presenting a promising strategy to optimise CAR-NK therapies for solid tumours.
• CRISPR-dCas9 methylation editing has been used to study DNA methylation dynamics in breast cancer progression. The study identified six methylation-related differentially expressed genes (CAVIN2, ARL4D, DUSP1, TENT5B, P3H2, MMP28) where hypermethylation correlated with reduced gene expression. Notably, silencing TENT5B promoted cell proliferation.
• German researchers have developed a novel micelleplex, C14-PEI, to co-deliver Cas9 mRNA and sgRNA for editing KRAS mutations in lung cancer cells. With 60% indel efficiency and 48.5% gene editing efficiency, C14-PEI specifically targets the KRAS G12S mutation, reducing tumour cell migration and increasing apoptosis.
• Highly branched poly(β-amino ester)s (HPAEs) with phenyl guanidine (PG) groups (HPAE-PG) have been developed as non-viral vectors for CRISPR-Cas9 RNP delivery, enabling efficient large DNA fragment deletion. HPAE-PG used in combination with dual sgRNAs allowed for the successful deletion of exon 80 (56 base pairs) of the COL7A1 gene in HEK cells and keratinocytes from dystrophic epidermolysis bullosa patients.
• Researchers have used a two-guide CRISPR knock-in technique to replace the entire coding sequence (CDS) of the synaptic LRRTM2 protein (Leucine-Rich Repeat Transmembrane neuronal protein 2) with a freely editable custom donor sequence. The approach overcomes the limitations of traditional editing, which often disrupts neuronal function, and it allows for the detailed study of the critical adhesion molecule's trafficking and function.
• A study by researchers in Saudi Arabia reveals that CRISPR-Cas9 editing frequently induces unintended large insertions (LgIns), including retrotransposable elements and host genomic fragments, via DNA repair mechanisms. The use of phosphorylated dsDNA donors significantly reduces LgIns and large deletions while maintaining HDR efficiency.
• A DNA nanostructure (TUBE) modified with aptamers was developed to deliver CRISPR-Cas9, enabling precise and dynamic genomic imaging in living cells. This method tracks loci like Chr3q29 during mitosis, revealing distinct spatial behaviours compared to telomeres. It supports high-resolution, multicolour labelling and multiplexed tracking, offering a versatile tool for studying genome regulation and cellular processes in real-time.

Clinical

• YolTech Therapeutics has launched a clinical trial for YOLT-204, an in vivo CRISPR-Cas therapy targeting transfusion-dependent beta-thalassemia (TDT). Delivered via lipid nanoparticles, YOLT-204 edits the haemoglobin regulatory region to induce fetal haemoglobin expression, potentially restoring red blood cell production. Preclinical studies showed sustained efficacy.

Industry

• IntelliGenome's CRISPR-TB Blood Test has received FDA Breakthrough Device Designation. Combining CRISPR with real-time PCR, it detects Mycobacterium tuberculosis cfDNA in blood with high sensitivity, enabling rapid and sputum-free diagnosis of pulmonary and extra-pulmonary tuberculosis (TB). Designed for CLIA-certified labs, this cost-effective test improves diagnostic access for vulnerable populations, including children and critically ill patients, advancing early TB detection and public health efforts.

Screens

• Using an in vivo CRISPR-Cas9 screen in Huntington's disease knock-in mice, researchers have identified modifiers of CAG repeat expansion in the huntingtin (HTT) gene, a key driver of disease onset. The study revealed DNA repair genes, particularly mismatch repair pathway components, as critical regulators. These findings provide insights into the mechanisms of somatic CAG expansion and highlight potential therapeutic targets for delaying Huntington's disease progression.
• Researchers used a CRISPR-based genome-wide knockout screen in HEK 293 cells to identify genes influencing recombinant AAV (rAAV) vector production. Knockout of heparan sulfate biosynthesis genes reduced rAAV yield while targeting specific vesicular trafficking proteins increased it. These findings suggest host proteins linked to viral infection are also involved in assembly, offering a strategy to optimise producer cells for higher rAAV production efficiency genetically.

Detection

• A new CRISPR-based electrochemical strategy allows for the ultrasensitive detection of breast cancer extracellular vesicles (BC-EVs). Using fluidity-enhanced magnetic nanoprobes and a DNA logic gate-based CRISPR-Cas12a amplification system, the method achieves precise identification of dual protein markers on BC-EVs with a high signal-to-noise ratio in clinical serum samples.
• A new method, CESBA, combines CRISPR-Cas13a with NESBA amplification to achieve ultrasensitive genomic RNA detection. Using SARS-CoV-2 as a model, CESBA detected as few as 0.05 copies/μL with high specificity via fluorescence and lateral flow assays. The new method was validated on clinical samples with 100% sensitivity and specificity, demonstrating it as a powerful tool for molecular diagnostics with exceptional sensitivity and versatility.
• A CRISPR-Cas12a-based diagnostic method targeting the Orientia tsutsugamushi 16S rRNA was developed for rapid scrub typhus detection, a neglected tropical disease. The one-pot isothermal assay detects as few as 100 copies within 1 hour and achieves 98% clinical sensitivity and high specificity in patient blood samples.

Reviews

• Transthyretin Cardiac Amyloidosis: Current and Emerging Therapies. This review describes current and newer transthyretin (TTR) stabilisers, TTR silencers, which include small interfering RNA agents (siRNA), antisense oligonucleotides (ASO) and CRISPR-Cas9 gene editing, and TTR depleters, which investigates the use of monoclonal antibodies to remove amyloid fibril deposits for patients with advanced disease.
• Advanced Delivery Systems for Gene Editing: A Comprehensive Review from the GenE-HumDi COST Action Working Group. This review examines the diverse systems developed for the delivery of CRISPR genome-editing tools. The study delves into both viral and non-viral delivery methods and highlights their respective advantages, limitations, and potential clinical applications.
• Treating genetic blood disorders in the era of CRISPR-mediated genome editing. Using β-hemoglobinopathies and haemophilia as exemplars, this review examines recent experimental breakthroughs utilising CRISPR-derived genome editing technology that has translated to significant improvements in the management of inherited hematologic disorders.
• New frontiers in CRISPR: Addressing antimicrobial resistance with Cas9, Cas12, Cas13, and Cas14. This review aims to explore the applications of CRISPR-Cas technologies in combating AMR, evaluating their effectiveness, challenges, and potential for integration into current antimicrobial strategies.
• Integrating malaria vaccine and CRISPR/Cas9 gene drive: a comprehensive strategy for accelerated malaria eradication. This review highlights the potential of integrating CRISPR-Cas gene drive technology with malaria vaccination efforts to improve vector control and reduce transmission.
• Unlocking the potential of CRISPR-Cas9 for cystic fibrosis: A detailed review. This review explores the application of CRISPR-Cas9 for cystic fibrosis (CF), particularly targeting mutations in the CFTR gene, and it provides a foundation for advancing CRISPR-Cas9 technologies for the treatment of CF and related disorders.
• CRISPR-Cas9 in Cardiovascular Medicine: Unlocking New Potential for Treatment. This review explores the potential of CRISPR-Cas9 to revolutionise cardiovascular disease treatment by targeting genetic causes while addressing technical, ethical, and translational challenges.

Patents

• The CVC group (University of California, University of Vienna, and Emmanuelle Charpentier) has secured the intention to grant for divisional patent EP4289948A, restoring broad CRISPR-Cas9 claim scope in Europe after prior patent revocations. The new claims cover single guide RNAs (sgRNAs), modified DNA-targeting RNAs, and combinations with naturally occurring Cas9, reinforcing CVC's IP position. This development coincides with ERS Genomics' new "Express License" platform for small enterprises, streamlining access to CRISPR-Cas9 research licenses. However, ongoing oppositions and claims in related patents suggest the European CRISPR-Cas9 IP landscape will remain contentious.