CMN Weekly (3 September2021)

Top picks

• A new treatment approach for rheumatoid arthritis uses implanted cells that release drugs in response to inflammation. American researchers from the NIH used CRISPR-Cas9 genome editing of iPSCs to generate a synthetic gene circuit that senses changing levels of endogenous inflammatory cytokines to trigger a proportional therapeutic response. Bioengineered implants in a mouse model system significantly mitigated disease severity as measured by joint pain, structural damage, and systemic and local inflammation.
• Though RNA-targeting CRISPR–Cas13 systems have significant therapeutic potential, their applications have been hampered because Cas13-based RNA editors exceed the packaging capacity of adeno-associated virus (AAV). To solve this problem, Feng Shang and co-workers have identified and characterised an ultrasmall family of Cas13b proteins, Cas13bt. The researchers fused inactive Cas13bt to the catalytic domain of RNA adenosine or cytosine deaminases. The full constructs were then easily packed into AAV vectors. When the loaded AAVs were transfected to HEK293FT cells, they efficiently edited the RNA target sites.

Research

• The miniaturisation of Cas-proteins has also come to Cas12 in other versions than the previously reported Cas12e (CasX) and Cas12j (CasΦ). In a paper in Nature Chemical Biology, Chinese researchers report on discovering a novel RNA-guided miniature endonuclease of only 422 amino acids from Acidibacillus sulfuroxidans, AsCas12f. AsCas12f1 was shown to be an effective genome-editing tool in both bacteria and human cells using various delivery methods, including AAV.
• In another paper in Nature Biotechnology, researchers from South Korea report on an optimised version of the hypercompact archaeal Un1Cas12f1 that in its original form has very low activity in eukaryotic cells. By redesigning the natural guide RNA, the researchers increased the average indel frequency by 867-fold.
• Homologous recombination is essential for the accurate repair of double-stranded DNA breaks (DSBs) and is a crucial component of CRISPR-Cas gene-editing. In a Nature paper, Swedish researchers use CRISPR to induce DSB in E. coli with labelled RecA molecules. This approach allowed for imaging of the full homologous recombination process with fluorescence microscopy. The researchers found that a thin, highly dynamic RecA filament stretches throughout the cell to search for the sister loci and that the search takes less than nine minutes.
• A paper on the preprint server bioRxiv suggests an urgent need for the CRISPR community to agree upon a benchmark dataset for CRISPR-Cas off-target scoring. The authors curated a "TrueOT" dataset and compared the performance of existing models on it. They demonstrated that model complexity could only improve performance on TrueOT if transfer learning techniques are employed.

Industry

• Poseida Therapeutics announces that FDA has cleared its Investigational New Drug (IND) application for P-BCMA-ALLO1 - a fully allogeneic CAR-T product candidate for patients with relapsed/refractory multiple myeloma. P-BCMA-ALLO1 is produced using the proprietary Cas-CLOVER site-specific gene-editing system that combines CRISPR/Cas9 and TALEN or ZFN.
• In another announcement, Poseida Therapeutics presents preliminary results from its Phase 1 clinical trial of P-PSMA-101, the Company's - solid tumour autologous CAR-T product candidate to treat patients with metastatic castrate-resistant prostate cancer (mCRPC). The data from nine patients showed that one patient had complete tumour elimination and remains in a durable response of greater than five months at the time of this presentation. Three other patients showed significant improvement.

COVID-19

• Researchers in South Korea have developed a new assay for rapid point-of-care testing for various infectious viruses, including SARS-CoV-2, SARS-CoV and MERS-CoV. The assay is based on isothermal reverse transcription recombinase polymerase amplification (RT-RPA) for target amplification. CRISPR-Cas12a then generates a fluorescent signal that can be measured or detected by the naked eye on a lateral flow paper strip.

Reviews

• A review looks into the possibilities of nanoparticle-based CRISPR-Cas9 and RNAi systems for treatments that can address the root cause of type 2 diabetes. The study points out the safety issues with target-specific delivery using viral carriers and suggests that nanoparticles will be a better choice for delivery to the target site.
• Another review looks at CRISPR/Cas9 and other innovative multiplexed engineering approaches to use natural killer cells (NK) for cellular immunotherapy in cancer. The review focuses on how to overcome CAR-NK functional exhaustion and reprogram immune cell metabolism for enhanced potency.
• A review by Chinese researchers focuses on the investigations and applications that employ the CRISPR/Cas9 system to combat several human infectious viruses. It also discusses challenges and future perspectives of using this new platform in the preclinical and clinical settings as an antiviral strategy.

Huh, Heh, Wow

• Chinese researchers have developed a new tool for the rapid authentification of halal food. The assay uses CRISPR-Cas12 and a gRNA targeting the pork cytochrome b (Cyt b) gene to cleave single-stranded DNA probes and turn-on fluorescence. The method allowed specific discrimination of pork meat from beef, mutton, and chicken and yielded a detection limit of 2.7 ng/μL of total DNA from pork meat.