Selective CRISPR enzyme kills cancer cells by shredding their DNA

Good news! Cancer is history (soon)!

"A peculiar type of CRISPR enzyme that slashes DNA into pieces has been used to control cancer in mice.

The enzyme, called Cas12a2, can be engineered to recognize a particular RNA sequence. When it finds its target, the enzyme goes wild, indiscriminately cutting up DNA.

Two separate research teams have used the CRISPR enzyme to target RNA made by cancer cells with mutated TP53 or KRAS genes. The treatment killed a proportion of lab-grown cancer cells and shrank tumours in mice. 

“There’s obviously still room to improve it,” ..."

"... A therapy that uses this approach to target head and neck cancers caused by human papillomavirus (HPV) is already in early development at Akribion Therapeutics, a biotechnology company in Zwingenberg, Germany. ..."

"... The research team engineered a CRISPR system called CRISPR-Cas12a2 to look for the specific RNA transcript produced only by cells with the mutated cancer gene.

In bacteria, this CRISPR acts as a suicide pill, intentionally killing a cell that has been infected by a virus to prevent its spread.

In the newly engineered version, once the system detects a cancer signature within a cell, the Cas12a2 enzyme activates and initiates “chromatin shredding,” slicing up all the genetic material inside that specific cell. This widespread genetic demolition triggers cell death, destroying mutated cells while leaving healthy cells completely untouched. ..."

From the abstract (1):

"Genetic mutations that drive cancer often occur in tumour-suppressor proteins such as the p53 transcription factor, which is altered in 40–50% of cases.

However, current therapies often fail to target these mutations because the mutant proteins typically lack defined drug-binding pockets and restoring their endogenous function has proven challenging.

Here we program Cas12a2, an RNA-guided CRISPR nuclease with trans-nucleolytic cleavage activity, to kill cancer cells selectively by targeting cancer-specific transcripts.

This approach limited cell growth by inducing trans shredding of chromatin and triggering DNA-damage responses and cell death.

In contrast to existing methods, RNA-guided Cas12a2 senses cellular RNA signatures, enabling precise targeting of undruggable mutations.

Transcript-activated chromatin shredding provides an innovative approach to precision disease treatments for undruggable targets."

From the abstract (2):

"Selectively eradicating target cells on the basis of their genetic or transcriptional identity remains important in basic research, medicine, biotechnology and agriculture.

For applications involving bacteria, CRISPR nucleases offer promising options due to their ability to enact RNA-guided counterselection; however, using these same nucleases for counterselection in eukaryotes has proven much more restrictive.

Here we show that Cas12a2, a recently discovered type V CRISPR nuclease, exhibits RNA-triggered DNA shredding, and enables programmable and sequence-specific elimination of yeast and human cells expressing a target transcript.

Triggering Cas12a2 elicits rampant double-stranded DNA breaks in trans, leading to cell death.

Cell killing can be activated by a wide range of target transcripts, with no observed off-target activation.

Leveraging this approach, we selectively eliminate cells that harbour human papillomavirus, cells that failed to undergo gene editing, or cells that encode a prevalent oncogenic point mutation in KRAS.

These findings expand the CRISPR toolbox to allow the selective elimination of eukaryotic cells on the basis of their transcriptional profile."

Nature Briefing: Cancer 

Bizarre CRISPR enzyme kills cancer cells by shredding their DNA "Early tests suggest the protein can be aimed at targets with tumour-causing mutations."

New CRISPR Technique Selectively Shreds Cancer Cells, Including “Undruggable” Cancers (original news release) "Innovative chromatin shredding technique shown to selectively destroy cancer cells carrying a mutation found in nearly half of all cancer cases"

Targeting cancer-specific mutations with RNA-triggered chromatin shredding (1; no public access, senior author is Jennifer A. Doudna, Nobel Prize winner in chemistry for CRISPR-Cas)

Selective Elimination of TP53 Mutant Cells by Transcript-Activated Chromatin Shredding (preprint of 1, open access, but preprint was apparently not updated to match the journal article)

RNA-triggered cell killing with CRISPR–Cas12a2 (2; open access)

Fig 2 Cas12a2 induces acute DNA damage and cell cycle arrest upon RNA targeting in mammalian cells.

Fig. 3: Cas12a2 has limited off-target activation in human cells.

An investigation into a hidden gene-editing therapy death of a 6-year old girl in China

Food for thought! Is the Communist Party of the superpower China pushing the envelope too fast in an attempt to stay ahead or just negligent and careless about human lives like other communist parties before? Or was this only a singular, isolated  incident?

Why the rush for this therapy? Perhaps, there would have been even a better therapy available in the near future.

Caveat: I did not read the two articles in their entirety. One of the articles is very long.

"On 24 March 2025, physicians and scientists in China tried to help a 6-year-old girl whose brain wasn’t developing properly because of a genetic mutation. They infused into her spinal fluid trillions of viruses that carried instructions to assemble a so-called base editor customized to fix the faulty gene in her brain cells.

Her condition was not likely to be fatal, but it could bring lifelong disability and serious medical issues. So her parents turned to an academic team, who developed the base editor and showed them evidence that it worked in the brains of mice and monkeys. The parents were told the risks were modest and decided treating their daughter was worth the gamble.

The trial went tragically wrong, however, according to a Science and Retraction Watch investigation written by Brendan Borell. The child, whose parents asked that she be called Mei (Chinese for “beautiful”), developed a fever and then signs of kidney damage. She died within a week after being treated. ..."

"... Doctors hoped to repair that mistake while her brain was still building itself. It would be a clinical trial of one, funded in part by $860,000 the parents had scraped together from their own savings and from relatives. ...

The parents felt they were in good hands. Xinhua Hospital, which is affiliated with the Shanghai Jiao Tong University School of Medicine, was acclaimed for its pediatrics department. ...

Although news that base editing saved “Baby KJ” would soon rocket around the world—Science named the feat one of the runners-up for its 2025 Breakthrough of the Year—the story of what happened at Xinhua Hospital has remained hidden. An entry for the study posted to ClinicalTrials.gov has not been updated for more than a year. And when Qiu and his colleagues published proof-of-concept animal studies related to the trial in Nature early this year, they stripped the paper of references to the family and its financial contributions ...

That vague language glossed over tragedy: Seven days after the girl’s medical team infused trillions of viruses carrying the recipe for the base editor into her spinal fluid, she died of a severe immune reaction linked to the therapy, Science and Retraction Watch can now reveal. ..."

Four takeaways from our investigation into a hidden gene-editing death | Science | AAAS "Girl’s death in China raises questions about the safety and cost of gene therapy, China’s biotech push, and scientific publishing practices"

A fatal reaction A cutting-edge gene-editing trial in China went disastrously wrong. A family wants accountability

TAPIR, a new CRISPR-based technology

Good news!

"Which comes first: cellular dysregulation or ribosomal RNA? This classic chicken-and-egg problem has long flummoxed scientists studying diseases associated with altered levels of ribosomes ... researchers have struggled to confirm whether unusual ribosomal readings in patients with certain cancers and congenital conditions were contributing to the diseases’ progressions or were side effects from other cellular issues.

To better understand this relationship, scientists created a new gene editing technology called Targeted Activation of Protein Translation—TAPIR, for short. ... the new technology allows them to tweak the quantities of the ribosomal RNA (rRNA) that makes up the bulk of the ribosomes themselves. By adjusting how much rRNA a cell generates and then observing how that affects a given medical condition, the team could finally establish a cause-and-effect relationship between ribosomes and disease.

The researchers first put TAPIR to the test by prompting ribosome formation in mice with a rare congenital disease that impedes rRNA creation. Once the rRNA was switched back on, the mice showed some improvement in their condition. Then, they activated rRNA in a different set of mice with pancreatic cancer, a disease associated with heightened ribosomal activity; in response, the cancer hastened its already rapid expansion.

Both results point to the same conclusion: Changes in rRNA levels are causes of disease symptoms, not symptoms themselves."

"... New Perspectives for Rare Diseases and Cancer

The results could be particularly relevant for diseases in which ribosome function is disturbed. These include ribosomopathies such as Treacher-Collins syndrome, a rare congenital disease that causes facial malformations. In a mouse model, the researchers successfully managed to partially compensate for disease-related alterations by stimulating rRNA production in a targeted way.

In addition, the research team observed that similar mechanisms also play a role in pancreatic cancer. Tumor cells seem to use increased rRNA production to maintain their rapid growth. In the mouse model for pancreatic cancer, TAPIR was able to increase rRNA production and promote the growth of the cancer cells. This shows that the increased rRNA production has a causal effect in contributing to tumor growth and is not just a side effect. ..."

From the abstract:

"Ribosomal RNA (rRNA) transcription rates vary during development, and their dysregulation is linked to diseases such as cancer and ribosomopathies. Owing to their high abundance and genomic redundancy, the functional significance of rRNA-levels remains unclear.

Here, we developed TAPIR (Targeted Activation of Protein Translation), a CRISPR-based approach to elevate rRNA-levels by inducing 47S rDNA transcription. TAPIR increased nucleolar size and enhanced protein synthesis, even in rapidly proliferating cells.

In neural stem cells, elevated translation promoted self-renewal and proliferation in vitro and in vivo.

Furthermore, TAPIR enabled the modeling and partial rescue of associated disease phenotypes.

Our findings revealed that rRNA-levels directly regulate translational output and that protein synthesis capacity can act as a key determinant of mammalian stem cell behavior."

ScienceAdviser

New CRISPR Method Makes It Possible to Control Protein Production in Cells (original news release) "A research team led by Prof. Stefan Stricker at Helmholtz Munich and Ludwig Maximilian University has developed TAPIR, a CRISPR-based technology that enables precise control of protein production in cells. The method provides new insights into the regulation of stem cells, cancer, and rare diseases, while opening up new opportunities for biomedical research."

Manipulation of protein translation and stem cell self-renewal by CRISPR activation of rRNA transcription (no public access)

First use of precision editing to alter a single gene to study human embryo development reveals role of master gene

Amazing stuff!

Notice we are now going way beyond the famous CRISPR/Cas9 gene editing!

In my opinion, the first gene to be removed from humans is the sneezing gene!

"Research ... has shown that a genome editing technique can be used to alter a single gene in human embryonic cells, enabling the study of very early human development in unparalleled detail.

The technique, called base editing, is a more precise version of the genome editing technique CRISPR/Cas9. It can change a single nucleotide base pair - the basic building block of DNA - within a human genome of approximately 3 billion base pairs. 

Using base editing, the researchers blocked a gene called NANOG in very early-stage human embryos, and found that the cells of the early embryo could not develop into more specialised pluripotent cells called the epiblast - which later form the body. ..."

From the abstract:

"Understanding how the first cell lineages in human development are specified and maintained has fundamental importance and clinical implications for regenerative medicine, infertility and pregnancy loss.

While mouse models have provided valuable insights into transcription factors regulating early development, translating these findings to human embryos has been limited by ethical, technical and biological constraints.

Functional studies of transcription factors in human embryos have been hindered by nuclease-based genome-editing approaches that induce genotoxicity. 

To overcome this, we applied adenine base editing (ABE8e) to precisely target an exon splice donor site, resulting in a splicing defect and functional knockout of NANOG, representing the first application of base editing to study a developmental regulator in human embryos.

This approach did not trigger genotoxicity and showed limited off-target editing.

Loss of NANOG disrupts pluripotent epiblast specification and instead cells differentiate toward a primitive endoderm (yolk sac) or trophectoderm (placental) transcriptional programme. Retention of primitive endoderm differentiation in NANOG-edited human embryos reveals a functional compensation distinct from mouse, underscoring the importance of directly investigating human development. Our findings demonstrate an essential role for NANOG in human pluripotency and epiblast specification, and highlight the utility of base editing for functional interrogation of human development."

First use of precision editing to study human embryo development reveals role of master gene | University of Cambridge "Scientists have, for the first time, used an extremely precise genome editing technique called base editing to study gene function in human embryos. They found that a gene called NANOG is essential for forming the future body from an embryo."

Base editing reveals an essential role for NANOG in human embryogenesis (no public access)

Day 6 human embryos showing the effect of NANOG presence versus absence. Normal embryo (left)

Könnten wir Stechmücken ausrotten?

Empfehlenswert! Wer braucht schon diese Blutsauger!

"... Keine dieser Methoden [der Vergangenheit} ist dazu geeignet, eine Mückenart komplett auszurotten. Aber es gibt eine Technik, die es schaffen könnte: der Gene-Drive. Eine einzige im Labor genetisch veränderte Mücke könnte ihre ganze Art ausrotten. ..."

NZZ Quantensprung

CRISPR takes a bold leap toward silencing Down syndrome's extra chromosome

Good news!

P.S. Google Scholar and Semantic Scholar (see screen print below) do not yet list this research paper published on 4/13/2026 as of writing this blog. This paper was published in the prestigious PNAS!

"Scientists have taken an important step toward a gene therapy that could one day turn off the extra genetic material that causes Down syndrome (DS). Down syndrome is a genetic condition caused by an extra chromosome 21 (and consequently hundreds of triplicate genes) that leads to developmental and neurological issues. ...

The team used a modified version of the gene-editing technique CRISPR/Cas9 ...  to insert the XIST gene into the extra chromosome 21 to silence it.

They tested their technique in the lab using human stem cells that contained an extra chromosome 21. After running several experiments, the team found that CRISPR was effective at pasting the XIST silencing gene exactly where it needed to go. ..."

From the highlights and abstract:

"Significance

Down syndrome (DS) results from trisomy 21 and remains without a molecularly targeted therapy. 

Prior work demonstrated that ectopic expression of the long noncoding RNA XIST could epigenetically silence the extra chromosome 21, but technical limitations, including low gene integration efficiency, have hindered translational progress.

Here, we present a CRISPR-based approach that markedly improves the efficiency and specificity of XIST integration into an extra copy of chromosome 21. By engineering Cas9-exonuclease fusion, designing SNP-specific sgRNAs, and enhancing donor–acceptor DNA pairing, we achieve a significant improvement in silencing efficiency. Our findings demonstrate partial transcriptional correction of trisomic gene dosage and offer a scalable, targeted platform for chromosomal therapy in DS and other aneuploidies.

Abstract

Down syndrome (DS) is one of the most common developmental human genetic disorders and is due to triplication of chromosome 21 (HSA21).

Although previous studies using epigenetic suppression of HSA21 by the long noncoding RNA XIST showed a potential for DS treatment, integration efficiency of XIST by conventional zinc finger nucleases is too low to allow for practical implementation.

Here, we report a modified CRISPR/Cas9 approach, which enhances the efficiency of XIST gene integration.

First, a codon-optimized λ-phage exonuclease (exo) was fused with Cas9 to create 5’- and 3’-end overhangs at cutting sites of donor DNA and acceptor chromosome DNA.

Second, four sgRNAs, two of which selectively targeted each the acceptor or donor DNA, were assembled tandemly into one Cas9 plasmid (PX459) to increase the Cas9-cutting efficiency and promote donor DNA integration.

Third, sgRNAs were designed by searching for unique single nucleotide polymorphism nucleotides distinct between the three HSA21 copies, as a protospacer adjacent motif site to specifically target one HSA21 copy.

Fourth, donor DNA plasmid containing XIST was modified to disable replication and inhibit transcription function and allow for inducible expression.

Our modified CRISPR method significantly enhanced the integration efficiency (20 to 40%) of long XIST gene (14 kb) into an extra chromosome 21 (HSA21), as was identified with PCR, cell cloning, immunostaining, and FISH.

RNA sequencing results showed that imbalance of gene transcription across extra HSA21 can be partially corrected by XIST gene integration. The modified CRISPR method with XIST paves a road for therapeutic treatment for DS."

CRISPR takes a bold leap toward silencing Down syndrome's extra chromosome

A modified CRISPR/Cas9 approach in silencing the triplication in Down syndrome: A treatment path XISTs (no public access)

Bacterial Retrons Revolutionize Gene Editing for Multiple complex Diseases

Amazing stuff!

"... Using bacterial retrons, this method can "reno" multiple dangerous mutations simultaneously, far more efficiently than previous gene-editing methods that might succeed against a single mutation, but are powerless against additional mutations not identical to the first. ..."

"Some genetic disorders—such as cystic fibrosis, hemophilia and Tay Sachs disease—involve many mutations in a person’s genome, often with enough variation that even two individuals who share the same disorder might have a different combination of mutations. Complexities like these make it challenging to develop broadly applicable gene therapies for these disorders.

Researchers ... now have developed an improved method of gene editing that is precise, more efficient than other similar methods and can correct many disease-causing mutations at once in mammalian cells. They also demonstrated its effectiveness in correcting scoliosis-causing mutations in zebrafish embryos. The new method uses genetic elements from bacteria called retrons that help protect the microbes from viral infection. This is the first time researchers have corrected a disease-causing mutation in vertebrates using retrons, raising hopes of new gene therapies for human disorders. ..."

From the abstract:

"Retrons can produce multicopy single-stranded DNA in cells through self-primed reverse transcription. However, their potential for inserting genetic cargos in eukaryotes remains largely unexplored.

Here we report the discovery and engineering of highly efficient retron-based gene editors for mammalian cells and vertebrates.

Through bioinformatic analysis of metagenomic data and functional screening, we identify retron reverse transcriptases that are highly active in mammalian cells. Rational design further improves the editing efficiency to levels comparable with conventional single-stranded oligodeoxynucleotide donors but from a genetically encoded cassette.

Retron editors exhibit robust activity with Cas12a nuclease and Cas9 nickase, expanding the genomic target scope and bypassing the need for a DNA double-stranded break.

Using a rationally engineered retron editor, we incorporate a split GFP epitope tag for live-cell imaging.

Lastly, we develop an all-RNA delivery strategy to enable DNA-free gene editing in cells and vertebrate embryos. This work establishes retron editors as a versatile and efficient tool for precise gene editing."

Bacterial Retrons Revolutionize Gene Editing for Multiple Diseases

New Gene-Editing Tech Holds Promise for Treating Complex Genetic Diseases (original news release) "Advanced biotechnology repurposes two bacterial immune systems to correct large stretches of DNA."

Discovery and engineering of retrons for precise genome editing (no public access)

Discovery and Engineering of Retrons for Precise Genome Editing (preprint from July 2024, open access)

Figure 1: A metagenomic survey reveals highly active RTs in mammalian cells.

Researchers unveil a powerful new gene-switch tool

Good news! Using a relatively nontoxic poison exon! 😊

"Investigators ... have developed a versatile and nontoxic technology for controlling the activity of any gene in a cell. Such “gene-switch” tools allow scientists to turn a target gene on or off to study how it works, model diseases and design new therapies.

The tool potentially could be adopted throughout biomedical research, including in the development of gene therapies. 

The new tool, called Cyclone (acyclovir-controlled poison exon) ... It was inspired by a natural feature of some genes, a DNA segment called a “poison exon,” which under certain conditions can block the gene from being translated into a protein.

To make Cyclone, the researchers engineered a poison exon that can be inserted into any target gene to suppress its activity. When the researchers are ready to turn up gene activity, a drug that binds to the poison exon is introduced and activity resumes. Unlike existing popular gene-switch tools, Cyclone uses a relatively nontoxic molecule, the antiviral drug acyclovir, to switch on the target gene. ..."

From the abstract:

"The ability to precisely control gene expression using small-molecule drugs is a valuable tool in research and has important therapeutic potential. However, existing systems are often limited by the toxicity of the drugs and the need to alter gene sequences or endogenous regulatory elements.

Here, we introduce Cyclone (acyclovir-controlled poison exon), an acyclovir-controlled poison exon cassette that can be used for small-molecule control of both transgene and endogenous gene expression.

Cyclone is a portable ‘intron–poison exon–intron’ element that can be inserted into nearly any gene and is completely removed upon acyclovir treatment, leaving the native transcript intact.

Cyclone offers tunable, reversible gene expression with nearly undetectable background and a ~295-fold activation.

We also present Pac-Cyclone, a cassette that simplifies the generation of cell lines with acyclovir-controlled endogenous gene expression.

Finally, we demonstrate the programmability of Cyclone, underscoring its potential for developing diverse genetic circuits controlled by various ligands."

Researchers unveil a powerful new gene-switch tool | Cornell Chronicle

Versatile gene-switch tool uses non-toxic molecule for safer research

A portable poison exon for small-molecule control of mammalian gene expression (no public access)

Endogenous gene regulation by Pac-Cyclone

Precise gene editing technique changes one DNA base to correct heart disease

Good news! Amazing stuff!

"Faulty versions of the LMNA gene can cause a wide range of health problems, including heart muscle disease (dilated cardiomyopathy) and muscle weakness (muscular dystrophies). Many of these diseases are caused by single-point mutations, which are changes to one DNA "letter" (base). Treatments include physical therapy and lifelong medication, but there are currently no cures. That could change following the work of a team of scientists who have developed and successfully tested a gene editing technique to correct the underlying genetic mutations.

Researchers led by Eric Olson at the University of Texas Southwestern Medical Center used a method called base editing, which has been used previously to correct other genetic heart diseases, but never before on these specific LMNA mutations. Unlike older editing tools, which create double-strand breaks in the DNA, base editing works like a "pencil" and "eraser" to change a single base in a patient's genome. ..."

From the significance and abstract:

"Significance

Point mutations in the Lamin A (LMNA) gene cause devastating human diseases that preferentially affect skeletal muscles and the heart. There is currently no cure for these conditions, and existing therapies are designed to mitigate the symptoms. Base editors represent a powerful approach to correct disease-causing point mutations because they enable single-nucleotide changes without introducing double-strand DNA breaks.

In this study, we developed two distinct base editing approaches to edit point mutations in the LMNA gene associated with cardiac disease. We demonstrated their efficacy using patient-derived cardiomyocytes and humanized mouse models. Our work represents an important step toward the potential clinical correction of cardiomyopathies using gene editing tools.

Abstract

Mutations in the Lamin A (LMNA) gene, which encodes the Lamin A and C proteins, cause severe human diseases collectively known as laminopathies. These conditions are often devastating and lack effective therapies.

In this study, we developed precise base editing (BE) strategies targeting the human LMNA gene variants L35P and R249Q, which cause congenital muscular dystrophy (CMD) and dilated cardiomyopathy with conduction defects (DCM-CD), respectively.

Induced pluripotent stem cell–derived cardiomyocytes (iPSC-CMs) carrying the R249Q mutation displayed nuclear aberrations, DNA damage, and abnormal Ca2+ transients.

Similarly, L35P iPSC-CMs exhibited abnormal contraction, DNA damage, and reduced Lamin A/C protein expression.

We also generated “humanized” mouse models carrying these pathogenic human mutations. R249Q homozygous mice exhibited cardiac conduction abnormalities, cardiac arrhythmias, and premature death. Mice with the homozygous L35P mutation displayed severe muscle-wasting and reduced lifespan, while heterozygous L35P mice displayed DCM.

We developed an adenine base editing (ABE) approach for correcting the R249Q mutation and a cytosine base editing (CBE) strategy for the L35P variant.

Precise correction of these mutations in iPSC-CMs successfully rescued all of the in vitro abnormalities.

Furthermore, delivery of the BE components using adeno-associated virus prevented the pathological phenotypes and extended longevity of mice carrying the LMNA L35P and the R249Q mutations.

These results demonstrate the efficacy of ABE and CBE in correcting pathogenic LMNA mutations that cause cardiac disease, highlighting BE as a promising therapeutic approach for human laminopathies."

Precise gene editing technique changes one DNA base to correct heart disease

Precise gene editing of pathogenic Lamin A mutations corrects cardiac disease (no public access)

Apparently, there was some very similar work done very recently by researchers mainly from Spain:

CRISPR/Cas9-mediated elimination of the LMNA c.745C>G pathogenic mutation enhances survival and cardiac function in LMNA-associated congenital muscular dystrophy

Researchers used a technique called base editing to correct a mutation in the lamin A gene. At 8 months old, heart cross sections from treated mice resembled those of normal healthy mice.

Candy giant Mars partners with biotech firm Pairwise to gene-edit cocoa supply

Good news for chocolate lovers and chocoholics! Maybe one day we can grow cocoa in Canada? Just kidding!

"Key Points

• Candy maker Mars said Wednesday it has partnered with biotech company Pairwise to speed up the development of more resilient cocoa using CRISPR-based gene editing technology. ...
• The goal is to create cacao plants that can better withstand disease, heat and other climate-related stresses that can put global chocolate supply at risk.

..."

"... Mars, Incorporated has licensed Pairwise's Fulcrum® gene editing tools for cacao research and development. This licensing agreement grants Mars access to Pairwise's CRISPR tools, including the SHARC™ enzyme. The Pairwise Fulcrum™ platform encompasses gene editing tools, enzymes and trait libraries, enabling precise changes that unlock the plant’s inherent potential. This capability significantly accelerates the development of impactful crop traits compared to traditional breeding methods, allowing for the activation or deactivation of characteristics and fine-tuning traits, much like adjusting a dimmer switch to achieve optimal results.  ..."

Candy giant Mars partners with biotech firm to gene-edit cocoa supply

Mars and Pairwise Collaborate to Accelerate Cacao Research and Development (original news release) "Mars, Incorporated has licensed Pairwise’s Fulcrum® CRISPR platform"

Credits: The Flyover

Make them bigger, grow faster and grow anywhere

The durable cure for cystic fibrosis might start in the womb

Good news!

"... Using in utero gene editing, the researchers developed a method to deliver corrective genetic material to the fetus of mice with CF via tiny particles called nanoparticles. ...

Cystic fibrosis arises from mutations in the CF transmembrane conductance regulator, or CFTR. Because the disease is caused by a single mutation, researchers saw it as a great candidate for gene editing, specifically in utero gene editing.  ...

In the study, researchers used synthetic molecules similar to DNA — called peptide nucleic acids, or PNAs — to correct CFTR. PNA can be tweaked to bind to a particular gene containing a mutation in a strand of DNA, which can cause a lesion that the cell then removes itself. The gene is then corrected when a strand of DNA without the mutation takes its place. ...

The researchers delivered the PNA to the fetuses of mice using tiny synthetic particles called nanoparticles, which are the same size as viruses to correct the CFTR mutation before the mice were born. ..."

From the significance and abstract:

"Significance

Cystic fibrosis (CF), a monogenic disease resulting from mutations in the CF transmembrane conductance regulator (CFTR) gene, affects multiple organs, including the respiratory, gastrointestinal, and reproductive systems.

Irreversible pathology in these tissues is often already present at birth, suggesting that a true curative therapy would require intervention during fetal development.

Here, we demonstrate CFTR gene correction in multiple tissues affected by CF after a single in utero administration of nanoparticles containing gene editing cargo. We found that gene correction was durable into adulthood in mice, which resulted in functional CFTR activity in both the respiratory and gastrointestinal systems. Our work establishes the possibility that CF could be treated, or possibly cured, by a single in utero gene editing treatment.

Abstract

In utero gene editing has the potential to modify disease-causing genes in multiple developing tissues before birth, possibly allowing for normal organ development, disease improvement, and conceivably, cure. 

In cystic fibrosis (CF), a disease that arises from mutations in the CF transmembrane conductance regulator (CFTR) gene, there are signs of multiorgan disease affecting the function of the respiratory, gastrointestinal, and reproductive systems already present at birth. 

Thus, treating CF patients early is crucial for preventing or delaying irreversible organ damage. Here, we demonstrate proof-of-concept of multiorgan mutation correction in CF using peptide nucleic acids encapsulated in polymeric nanoparticles and delivered systemically in utero.

In utero editing was associated with sustained postnatal CFTR activity, at a level similar to that of wild-type mice, in both respiratory and gastrointestinal tissues, without detection of off-target mutations in partially homologous loci. This work suggests that systemic in utero gene editing represents a viable strategy for treating monogenic diseases before birth that impact multiple tissue types."

The cure for cystic fibrosis might start in the womb | Yale News "In a new study, Yale researchers develop a method that has the potential to cure cystic fibrosis prior to birth."

Systemic in utero gene editing as a treatment for cystic fibrosis (no public access)

Marie Egan, a senior author of the study

UK approves regulations for gene-edited crops under Precision Breeding Act while EU struggles with outdated legislation

Good news! More good reasons in support of the Brexit in 2020!

The EU has become a Leviathan!

"The UK parliament has signed into law rules for its Precision Breeding Act (2023), bringing the sale of gene-edited products to consumers closer. However, the new law applies to England only.

Precision breeding refers to altering an animal’s or plant’s DNA using specialised enzymes to change its genome at specific points; the modification must be possible using traditional breeding to qualify. They do not apply in cases where genes are transferred from unrelated species – deemed genetically modified organisms (GMO). ...

The new rules make it easier for researchers to develop and commercialise genetically-edited plants. They should clear a path to crops with enhanced resistance to diseases and pests or greater resilience to drought or waterlogging. ..."

UK approves regulations for gene-edited crops under Precision Breeding Act while EU struggles with outdated legislation | Chemistry World

Mosquitoes could be driven to extinction with gene editing. We finally may be able to rid the world of mosquitoes. But should we?

What a rhetorical and silly question!

Of course, we should get rid of all blood suckers! Don't hesitate, eradicate! Caveat: Mosquitoes may have important functions like pollination.

Hematophagy is an aberration of evolution (or a dead end in the tree of life) in my dim, narrow view! What are they good for other than spreading diseases? Do they help train our immune system this way?

Mosquitoes could be driven to extinction with gene editing - The Washington Post "Gene editing holds the potential of suppressing mosquito species that carry deadly diseases — and raises ethical questions."

Mutation agnostic evolved gene editor inserts entire genes in human cells of various cell types

Amazing stuff! It is getting crisper! 😊

"Researchers ... have developed a way to insert entire ... genes into human cells efficiently enough for potential therapeutic applications. It lays a foundation for gene-editing therapies for patients with different mutations that cause a genetic disease. ..."

"... The gene editor—called evoCAST—goes a long way toward solving a problem that has confounded the development of gene therapies from the field’s beginnings: How to add long stretches of DNA to defined locations in the human genome without creating unwanted modifications. ..."

From the editor's summary and abstract:

"Editor’s summary

The ability to install large DNA sequences into specified locations in the human genome has far-reaching implications, including paving the way for single-drug treatments of mutationally diverse loss-of-function genetic diseases. CRISPR-associated transposases (CASTs) are naturally occurring systems that support RNA-programmable insertion of gene-sized DNA but have shown minimal activity in human cells.

Witte et al. developed a continuous evolution platform to improve CAST activity, yielding an evolved CAST with more than 200-fold increased activity in human cells.

This enzyme enables efficient gene integration across a variety of therapeutically relevant genomic sites in multiple human cell types, representing a versatile new platform for mammalian cell genome editing. ...

Structured Abstract

INTRODUCTION

The efficient insertion of gene-sized DNA sequences at user-specified genomic sites is a long-standing goal in genome editing.

Although current editing methods can correct most disease-causing mutations, the genetic diversity underlying many disorders will require the design and regulatory approval of many mutation-specific strategies—substantially limiting the number of patients who can benefit from therapeutic genome editing. Programmed genomic integration of a healthy gene copy could offer a mutation-agnostic treatment for loss-of-function genetic diseases. Additionally, targeted gene integration enables other applications, including cancer immunotherapies, transgenic cell and animal models for basic research, and metabolic engineering.

RATIONALE

CRISPR-associated transposases (CASTs) are naturally occurring bacterial systems that exploit nuclease-deficient CRISPR machinery to integrate DNA at genomic locations specified by guide RNAs.

CASTs offer many attractive qualities as a genome editing tool, including facile programmability, compatibility with multi-kilobase-scale DNA cargo, and avoidance of genomic double-strand DNA breaks.

Despite this promise, wild-type CASTs reported to date support minimal integration in human cells (often ≤0.1% of treated cells). We reasoned that this low efficiency may stem from naturally evolved, suboptimal transposition catalysis that mitigates mobilization-induced fitness cost to the host. To enable efficient CAST integration in human cells, we developed a phage-assisted continuous evolution (PACE) system that rapidly evolves CAST variants capable of fast targeted transposition and applied CAST-PACE to a prototypical Type I-F CAST system from Pseudoalteromonas.

RESULTS

We linked on-target DNA integration in Escherichia coli to the propagation of continuously mutating phage genomes encoding evolving CAST components. After hundreds of generations of continuous selection, replication, and mutation in which the resulting phage survived an overall 10322-fold dilution, we generated an evolved variant of the CAST transposase protein TnsB that mediated >200-fold improved integration activity in human cells. The evolved TnsB contains 10 activity-enhancing mutations located throughout the protein, which likely modulate several distinct interactions with other CAST components. Notably, the evolved TnsB mediated efficient integration activity in human cells without requiring codelivery of the bacterial CAST accessory protein, ClpX, which is cytotoxic.

We combined this evolved TnsB with other PACE-evolved and rationally engineered CAST components to yield evoCAST, a system optimized for human-cell integration activity. EvoCAST achieved 10 to 30% integration efficiencies across 14 genomic targets in human cells, representing a 420-fold average improvement over wild-type CAST.

EvoCAST supported large DNA cargoes >10 kb and mediated the integration of several therapeutic payloads at disease-relevant genomic sites, including safe harbor loci, sites for cancer immunotherapy engineering, and genes implicated in loss-of-function genetic diseases.

EvoCAST also performed targeted integration in multiple human cell types, including primary human fibroblasts, and exhibited high product purity, with no detected insertions and deletions (indels), predominantly unidirectional cargo insertion, single–base pair precision of integration, and low levels of off-target integration.

CONCLUSION

This work establishes CAST as a powerful platform technology for efficient, RNA-guided gene integration in human cells. The advantages of evoCAST—including its simple programmability, single-step integration mechanism, and avoidance of genomic double-strand breaks—make it well-suited for many applications in the life sciences and therapeutics, including the capability to address genetically diverse patient populations through a single editing agent.

The CAST PACE system developed in this work also provides a strategy for improving the properties of other naturally occurring CASTs toward their use for efficient human-cell genome editing."

Evolved gene editor inserts entire genes in human cells | Broad Institute "The new system is the first to use a DNA-mobilizing enzyme called a CRISPR-associated transposase to make targeted gene-sized edits at therapeutically useful levels in human cells."

New Gene Editor Enables Greater Precision

Programmable gene insertion in human cells with a laboratory-evolved CRISPR-associated transposase (no public access)

PACE-evolved CASTs mediate efficient, programmable gene integration in human cells.

An ancient RNA-guided system could simplify delivery of gene editing therapies

Amazing stuff!

"A vast search of natural diversity has led scientists ... to uncover ancient systems with the potential to expand the genome editing toolbox. These systems, which the researchers call TIGR (Tandem Interspaced Guide RNA) systems, use RNA to guide them to specific sites on DNA. TIGR systems can be reprogrammed to target any DNA sequence of interest, and they have distinct functional modules that can act on the targeted DNA. In addition to its modularity, TIGR is very compact compared to other RNA-guided systems, like CRISPR, which is a major advantage for delivering it in a therapeutic context.  ...

The TIGR-associated (Tas) proteins that Zhang’s team found share a characteristic RNA-binding component that interacts with an RNA guide that directs it to a specific site in the genome.

They note that CRISPR systems can only be directed to segments of DNA that are flanked by short motifs known as PAMs (protospacer adjacent motifs). TIGR Tas proteins, in contrast, have no such requirement. “This means theoretically that any site in the genome should be targetable,” ... The team’s experiments also show that TIGR systems have ...  a “dual-guide system,” interacting with both strands of the DNA double helix to home in on their target sequences, which should ensure they act only where they are directed by their RNA guide. What’s more, Tas proteins are compact—a quarter of the size Cas9 on average—making them easier to deliver, which could overcome a major obstacle to therapeutic deployment of gene editing tools.  "

From the abstract:

"RNA-guided systems provide remarkable versatility, enabling diverse biological functions. Through iterative structural and sequence homology-based mining starting with a guide RNA-interaction domain of Cas9, we identified a family of RNA-guided DNA-targeting proteins in phage and parasitic bacteria.

Each system consists of a Tandem Interspaced Guide RNA (TIGR) array and a TIGR-associated (Tas) protein containing a Nop domain, sometimes fused to HNH (TasH) or RuvC (TasR) nuclease domains.

We show that TIGR arrays are processed into 36-nt RNAs (tigRNAs) that direct sequence-specific DNA binding through a tandem-spacer targeting mechanism.

TasR can be reprogrammed for precise DNA cleavage, including in human cells. The structure of TasR reveals striking similarities to box C/D snoRNPs and IS110 RNA-guided transposases, providing insights into the evolution of diverse RNA-guided systems."

An ancient RNA-guided system could simplify delivery of gene editing therapies | Broad Institute "The programmable proteins are compact, modular, and can be directed to modify DNA in human cells."

An ancient RNA-guided system could simplify delivery of gene editing therapies "The programmable proteins are compact, modular, and can be directed to modify DNA in human cells."

TIGR-Tas: A family of modular RNA-guided DNA-targeting systems in prokaryotes and their viruses (no public access)

The Tas protein uses an RNA guide to recognize a specific target DNA sequence.

Serious side effect of using CRISPR-Cas gene scissors uncovered: AZD7648 molecule can destroy parts of genome

CRISPR can be a blessing or a curse! Caution is advised! Double checking of innovations is needed! No rush please!

"... To get the cell to use homology-directed repair, the researchers recently began using a molecule called AZD7648, which blocks fast repair and forces the cell to use homology-directed repair. This approach is expected to accelerate the development of more efficient gene therapies. Initial studies with these new therapies have been good. Too good to be true, as it turned out.

A research group ... discovered that the use of AZD7648 has serious side effects. The study has been published in the journal Nature Biotechnology. ...

Extent of damage is large

The extent of the negative effects surprised the researchers.  ...

The ETH researchers then analyzed the sequence of DNA building blocks not only around the edited site but also in the wider environment. They discovered these unwanted and catastrophic side effects caused by using AZD7648. ..."

From the abstract:

''The DNA-PKcs inhibitor AZD7648 enhances CRISPR–Cas9-directed homology-directed repair efficiencies, with potential for clinical utility, but its possible on-target consequences are unknown. We found that genome editing with AZD7648 causes frequent kilobase-scale and megabase-scale deletions, chromosome arm loss and translocations. These large-scale chromosomal alterations evade detection through typical genome editing assays, prompting caution in deploying AZD7648 and reinforcing the need to investigate multiple types of potential editing outcomes."

Serious side effect of using CRISPR-Cas gene scissors uncovered: AZD7648 molecule can destroy parts of genome

Crispr-Cas: die Balance finden zwischen Effizienz und Sicherheit (original news release in German) "ETH-Forschende decken eine gravierende Nebenwirkung bei einer Anwendung der Genschere Crispr-Cas auf. Ein Molekül, das den Vorgang effizienter machen soll, zerstört Bereiche des Genoms."

Genome editing with the HDR-enhancing DNA-PKcs inhibitor AZD7648 causes large-scale genomic alterations

Fig. 2: AZD7648 increases the frequency of megabase-scale deletions, chromosome arm loss and translocations.

New CRISPR System Pauses/silences Genes Rather than Turning Them Off and without cutting DNA

Amazing stuff!

"“Researchers at Vilnius University in Lithuania introduced a new, more versatile genetic toolkit called the type IV-A CRISPR system. Described in a study published Oct. 29 in the journal Nature Communications, the system deactivate genes in an impermanent manner, giving researchers more control over gene activity…

by unraveling DNA’s double helix, the type IV-A system can stably but reversibly suppress a gene’s activity without having to cut its DNA.”"

"Scientists ... have discovered a unique way for cells to silence specific genes without cutting DNA. This groundbreaking research ... reveals a new way to silence genes akin to pressing a "pause" button on specific genetic instructions within cells.

The research team  ... uncovered how cells use a specific system to locate and silence unwanted DNA. This system, which could eventually enable safer gene modifications, shows promise for repairing faulty genes that cause diseases.

“Unlike the well-known CRISPR gene-editing system, often described as molecular 'scissors,' the newly studied type IV-A CRISPR system does not cut genes. Instead, it uses an RNA-guided ‘effector’ complex to recruit an enzyme called DinG, which moves along DNA and silences targeted genes more subtly,” ..."

From the abstract:

"CRISPR-Cas mediated DNA-interference typically relies on sequence-specific binding and nucleolytic degradation of foreign genetic material. Type IV-A CRISPR-Cas systems diverge from this general mechanism, using a nuclease-independent interference pathway to suppress gene expression for gene regulation and plasmid competition. To understand how the type IV-A system associated effector complex achieves this interference, we determine cryo-EM structures of two evolutionarily distinct type IV-A complexes (types IV-A1 and IV-A3) bound to cognate DNA-targets in the presence and absence of the type IV-A signature DinG effector helicase. The structures reveal how the effector complexes recognize the protospacer adjacent motif and target-strand DNA to form an R-loop structure. Additionally, we reveal differences between types IV-A1 and IV-A3 in DNA interactions and structural motifs that allow for in trans recruitment of DinG. Our study provides a detailed view of type IV-A mediated DNA-interference and presents a structural foundation for engineering type IV-A-based genome editing tools."

New CRISPR System Pauses Genes Rather than Turning Them Off - Human Progress

New CRISPR system pauses genes, rather than turning them off permanently "Researchers in Lithuania present the molecular structure of a new, more-versatile CRISPR system for gene editing."

Vilnius University Researchers Uncover New Gene Silencing Mechanism (original news release)

Structural variation of types IV-A1- and IV-A3-mediated CRISPR interference (open access)

Fig. 1: Structures of types IV-A1 and IV-A3.

Fig. 9: Model of type IV-A–mediated interference.

South Africa's latest research ethics guidelines include heritable gene editing

Good news? Will humans overtake nature when it comes to evolution? What could possibly go wrong in a world with plenty of dictators and megalomaniacs?

Maybe the old theocrats in Iran want to correct young women to obey hijab laws. Just an unlikely scenario?

From the "Cradle of Humankind" comes progress in gene editing?

"South Africa has amended its health-research ethics guidelines to include a new section on heritable (or germline) human genome editing. The move could put the nation one step closer to being the first to explicitly allow the controversial technique, which involves making genetic changes to sperm, eggs or embryos that could be passed down to future generations. Such editing could prevent inherited diseases, such as cystic fibrosis and sickle-cell disease, but it poses major ethical concerns and safety challenges."

Nature Briefing: Translational Research

Will South Africa become first country to accept controversial form of human genome editing? (no public access) "Scientists raise the alarm following updated research ethics guidelines on heritable human genome editing."

Designer Babies? South Africa Revises Research Guidelines to Permit Heritable Human Genome Editing with Molly Gambhir

Concerning! How much longer can it be prevented?

Tiny protein offers new path for gene editing

Good news! Will this discovery amend or enhance CRISPR-Cas (discovered in 2012)?

"Scientists have used protein engineering and an AI model to make the bacterial protein TnpB an effective gene editing tool for mammalian cells. TnpB is much smaller than the CRISPR-Cas system and could be more easily delivered to the right cells of the body as a result.

“By engineering the small but powerful protein TnpB, we were able to design a variant that shows a 4.4-fold increase in efficiency of modifying DNA – making it more effective as a gene editing tool,” ..."

"... It was recently discovered that Cas proteins evolved from much smaller proteins, with TnpB being the progenitor of Cas12. Since the large size of Cas proteins creates challenges when trying to deliver them to the right cells in the body, recent studies tried to use their smaller evolutionary progenitors as a genome editing tool. The problem with these small alternatives is that they function less efficiently. ...

TnpB proteins are found in a variety of bacteria and archaea. The TnpB studied by the researchers comes from the bacterium Deinococcus radiodurans. This microbe survives cold, dehydration, vacuum and acid, and is one of the most radiation-resistant organisms known to humans. The compact TnpB protein has previously been shown to work for genome editing in human cells, albeit with low efficiency and limited targeting ability due to its recognition requirements when binding DNA. ...

Therefore, the researchers optimized TnpB so that it edits the DNA of mammalian cells more efficiently than the original protein. “The trick was to modify the tool in two ways: first, so that it more efficiently goes to the nucleus where the genomic DNA is located, and second, so that it also targets alternative genome sequences,” ..."

From the abstract:

"Transposon (IS200/IS605)-encoded TnpB proteins are predecessors of class 2 type V CRISPR effectors and have emerged as one of the most compact genome editors identified thus far. Here, we optimized the design of Deinococcus radiodurans (ISDra2) TnpB for application in mammalian cells (TnpBmax), leading to an average 4.4-fold improvement in editing. In addition, we developed variants mutated at position K76 that recognize alternative target-adjacent motifs (TAMs), expanding the targeting range of ISDra2 TnpB. We further generated an extensive dataset on TnpBmax editing efficiencies at 10,211 target sites. This enabled us to delineate rules for on-target and off-target editing and to devise a deep learning model, termed TnpB editing efficiency predictor (TEEP; https://www.tnpb.app), capable of predicting ISDra2 TnpB guiding RNA (ωRNA) activity with high performance (r > 0.8). Employing TEEP, we achieved editing efficiencies up to 75.3% in the murine liver and 65.9% in the murine brain after adeno-associated virus (AAV) vector delivery of TnpBmax. Overall, the set of tools presented in this study facilitates the application of TnpB as an ultracompact programmable endonuclease in research and therapeutics."

Tiny protein offers new path for gene editing

Compact “Gene Scissor” Enables Effective Genome Editing (original news release) "CRISPR-Cas is used broadly in research and medicine to edit, insert, delete or regulate genes in organisms. TnpB is an ancestor of this well-known “gene scissor” but is much smaller and thus easier to transport into cells. Using protein engineering and AI algorithms, UZH researchers have now enhanced TnpB capabilities to make DNA editing more efficient and versatile, paving the way for treating a genetic defect for high cholesterol in the future."

Effective genome editing with an enhanced ISDra2 TnpB system and deep learning-predicted ωRNAs (no public access)