“Our study addresses a major gap by showing that aging dramatically increases breast cancer metastasis and that this effect depends on RAGE, a receptor on the surface of cells that fuels inflammation,” said Barry Hudson, PhD, associate professor of oncology at Georgetown Lombardi. “Most laboratory studies rely on young mice, which has limited our understanding of how aging itself alters the host environment, including immune function and chronic inflammatory states that, in turn, influence cancer behavior.” Hudson is corresponding author of the researchers’ Communications Biology published paper titled “Aging promotes a RAGE-dependent increase in breast cancer metastasis.” In their paper the authors concluded that their findings “… identify RAGE as a mechanistic link between aging and metastasis and a potential therapeutic target in older patients.” They say the findings will also be featured in the Nature portfolio special collection, Cancer and Aging.

Age is the primary risk factor for the development of adult cancers, including breast cancer, with almost half of new breast cancer diagnoses and more than half of breast cancer-specific deaths occurring in women aged 65 years and older, the authors wrote. And while advances in screening and therapy have improved survival, older women continue to have higher breast cancer-specific mortality. “Despite accumulating evidence that metastasis in murine breast cancer models increases with advancing host age, the mechanisms underlying this have not been elucidated, highlighting the need for further mechanistic studies,” the team continued.

And while breast cancer is more prevalent in older women, most cancer research in mouse models has used young, 2–3-month-old adult mice, which are about equivalent in age to 15–20-year-old humans. Timing and chance presented Hudson and colleagues with opportunities to carry out their newly reported study. During COVID, when there was reduced laboratory activity, some of the research team’s mouse colonies aged longer than originally planned. This created a rare opening to study cancer in these older animals—normally a difficult and expensive endeavor—giving the scientists the ability to directly compare how tumors behave in younger versus older mice.

RAGE is a proinflammatory molecule that is being considered as a therapeutic target in multiple aging-related diseases, including various cancers, cardiovascular and neurodegenerative diseases.

Using three different mouse models of triple-negative breast cancer (TNBC), the researchers discovered that aged mice developed substantially more lung metastases than younger mice, despite similar primary tumor growth. The team then showed that genetic deletion of RAGE in mice almost completely eliminated this age-related surge in metastasis.

Through their studies, the team demonstrated that aging increased levels of inflammatory molecules that activate RAGE. These included the proteins S100 and HMGB1, found in both primary tumors and at metastatic sites. These changes made it easier for cancer cells to invade and spread. “These findings show that aging doesn’t just increase cancer risk—it actively changes the body in ways that help tumors spread,” said Hudson. “RAGE appears to be a key mediator of these harmful age-related pathways.” In their paper the authors stated that their data “… suggest that aging promotes multiple prometastatic processes within the tumor and its microenvironment, and that RAGE is required for the induction of these inflammatory and tumor-promoting pathways in aged hosts.“

The team also analyzed breast cancer data from more than 1,000 patients and found that higher expression of AGER (the gene encoding RAGE) and related inflammatory gene signatures were associated with worse outcomes in patients, supporting the clinical relevance of their findings. They noted, “… in human breast cancers, high AGER expression, as well as enrichment for mouse tumor-derived aging- and RAGE-associated gene signatures, predicted poorer outcomes, particularly in older women …Together, these data indicate that in older individuals with breast cancer, intratumor RAGE overexpression amplifies aging-associated transcriptional programs, linking age-dependent inflammation to promote metastatic progression.”

RAGE is already being explored as a therapeutic target in several age-related diseases, highlighting its potential relevance in cancer. In prior work, the researchers had shown that the RAGE inhibitor TTP488 (azeliragon) can suppress breast cancer metastasis in preclinical models. In the current study, they also tested the drug in the lab and found that TTP488 was able to reduce tumor cell invasiveness that was induced by blood sera from aged mice.” Pharmacologic inhibition of RAGE by TTP488 (PF-04494700 or azeliragon) suppressed migration and invasion towards aged serum, further supporting the requirement of RAGE signaling for age-dependent metastasis,” the team noted.

A clinical study is underway at Lombardi evaluating TTP488 in breast cancer patients receiving chemotherapy, with a focus on safety and cognitive outcome. The drug has demonstrated a favorable safety profile in people, making it an optimal choice for further study. “TTP488 has demonstrated an excellent safety profile in Phase I/II clinical studies in older adults with Alzheimer’s disease, supporting its potential for repurposing,” the authors wrote. “Therapeutic RAGE inhibition may provide a well-tolerated means to counteract inflammaging and improve cancer outcomes in the elderly, who often face limited treatment options due to toxicity,” the investigators wrote.

“This study highlights the importance of the host environment in cancer,” Hudson added. “While cancer is often viewed as driven primarily by mutations intrinsic to tumor cells, systemic factors such as aging and inflammation play a critical role in shaping how cancers behave,” said Hudson. “Most deaths due to cancer occur because tumors spread to other organs, so understanding these influences may help identify new strategies to limit metastasis.”

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On average, patients with ALS live three years after symptoms begin, although some can survive closer to 10 years. Exactly what drives these differences in survival is unclear. “This study reveals that ALS is not a single event but a domino-like cascade that begins inside motor neurons with TDP-43 pathology and is then amplified by a damaging immune response in the bloodstream and spinal cord,” said David Gate, PhD, director of the Abrams Research Center on Neurogenomics at Feinberg and co-corresponding author on the study. 

Specifically, the study found that immune cells converge at sites of motor neuron loss and TDP-43 pathology with distinct inflammatory patterns depending on the type of ALS and how quickly the disease progresses. As Evangelos Kiskinis, PhD, an associate professor of neurology and neuroscience at Feinberg and a co-corresponding author on the study, explained it, “the intensity of spinal cord inflammation” determines “how fast the disease progresses and how long they survive.” 

To gain these insights, the scientists analyzed blood and spinal cord samples from living and deceased patients with both genetic and non-genetic forms of ALS, as well as controls. As part of the study, they used single-cell RNA sequencing technology to analyze blood from 40 living ALS patients and used spatial transcriptomics to analyze spinal cord tissue from 18 deceased participants. They also compared patients with non-genetic ALS to those with the genetic form of the disease to assess how immune activity differs across ALS types and disease stages. Lastly, they examined RNA from postmortem samples of 237 ALS patients to better understand the inflammatory responses within the central nervous system. 

Using these methods, “we found the immune cells we detected in the blood of people living with ALS were inflamed, and we found the genes that mediate their inflammatory response in the spinal cord at the site of motor neurons,” Gate said. “These inflamed immune cells were associated with ALS pathology, giving some credence to our theory that the immune system is detrimental. It’s responding to pathology, and it’s causing the disease to be worse.”

Additionally, patients whose disease advanced quickly had more activity in certain immune genes, while those with the genetic form of the disease had a different set of altered immune genes. In the spinal cord, these activated immune cells gathered directly at the locations of motor neuron loss and near the toxic protein buildups associated with ALS. “We saw that people with worse clinical ALS had more expression of complement genes, which are proteins that become activated as the body’s first-line immune defense against a pathogen or damage to the body,” Gate said.

Now that they have identified a direct link between the immune system and ALS, Gate and his lab plan to study samples from a wider pool of patients. “Our next step is to map exactly how this immune reaction spreads throughout the entire motor circuit: from the brain, down through the spinal cord and out to the muscles,” he said. “By profiling the motor circuit in depth, we’ll get a much clearer picture of where and when inflammation drives faster progression.” 

Meanwhile, Kiskinis and his team will test for a causal relationship between TDP-43 dysfunction and inflammation. “We’re trying to really define what is the mechanism that links TDP-43 dysfunction in nerve cells with inflammatory reactions,” he said.

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The ASGCT report is developed in conjunction with Citeline, a subsidiary of Norstella (a pharmaceutical intelligence provider covering drug development from preclinical to commercialization).

Barrett said there are currently 42 gene therapies approved worldwide, along with 38 RNA therapies and 76 (non-genetically modified) cell therapies, which are steadily growing the field. Two cell therapies were approved in Japan in Q1.

There was a small increase in deal-making, and a significant 30% increase in startup funding compared to the same period in 2025. “I think that signals and underscores a rebounding sector,” said Barrett.

Of the eight gene therapies approved over the past 12 months, half were in the United States, with three more in China. “The regulatory pace is starting to pick up, another strong indicator for the future of our field,” Barrett said. It is a similar picture in RNA therapies. “We see a steady uptick over the course of the last year,” he added.

Zooming out, Barrett estimated that there are more than 4,200 therapies currently in development, from preclinical through pre-registration. The vast majority of those (more than 4,130) are gene and genetically modified cell therapies, including about 1,300 RNA therapies.

In the field of gene-modified cell therapies, CAR T continues to lead the pipeline for ex vivo gene therapies, with natural killer (NK) and T-cell receptors gaining traction. Not surprisingly, genetically modified cell therapy overwhelmingly targets cancers, but Barrett noted growth in the percentage of these therapies targeting immunological diseases, including lupus, multiple sclerosis, and HIV.

Pipeline growth

Barrett also noted growth and “a promising future” in the clinical trials pipeline. There are currently 350 Phase I, 319 Phase II, and 41 Phase III trials in gene therapy (up from 35 a year ago). “Hopefully, we will see a number of completed trials and FDA decisions in the near term,” said Barrett. A growing proportion of gene therapy trials (exceeding 60 percent) is for non-oncology indications.

In the RNA space, “RNAi therapies are jumping,” said Barrett. The same cannot be said, however, for mRNA. “Unsurprisingly, mRNA therapies continue to slide quarter over quarter,” a symptom of “shaken confidence” in that space, he continued. RNA therapies are targeting primarily non-oncology indications, especially in rare diseases.

Upcoming catalysts

On the business front, Barrett noted there has been “a nice uptick” in Q1 in start-up funding compared to the same quarter last year, which he deemed “a really promising indication.” The number of start-ups historically has tended to hover between 5-20. For Q1, that number was 26.

The Q1 report tracks various business catalysts anticipated through the end of 2027, including increased interest and uptake in expedited review designations—fast track, RMAT, orphan drug breakthroughs and other accelerated approval pathways.

“FDA is getting a lot done… and hopefully we’ll see the same moving forward,” Barrett said.

The full Landscape Report is available online from the ASGCT website.

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Some of the research Davidson presented was conducted at a new biotech company she co-founded called Latus Bio, which earlier this month announced it had raised $97 million in a Series A round. The company develops novel AAVs to specifically target central nervous system (CNS) disorders, with a lead program in Huntington’s disease (HD).

After thanking her mentors—Bill Kelly, MD, Michael Welsh, MD, and Kathy High, MD—Davidson turned her attention to presenting new advances in engineered gene therapies. Throughout her career, she has focused on improving adeno-associated viruses (AAVs) for CNS gene therapies, with a particular emphasis now on HD. Key elements include selecting the right cargo and developing the appropriate delivery vehicle. Her goal is to scale lab research in neurons, mouse models, and non-human primates (NHPs) to treat patients, including adults with HD.

Major hurdles to tackling genetic diseases of the brain include scalability and a lack of potency, Davidson said. The search for alternative AAV serotypes to AAV2 that could target neuronal cells began back in 2000. IV administration does not provide sufficient targeting to the brain. Even AAVs that have been engineered to enter the brain from the blood have high peripheral exposure and a high cost of goods per patient, which significantly lowers scalability and impact. (In one study, liver biodistribution of AAV was many orders of magnitude higher than in the CNS.)

Davidson focused on HD, the late-onset, dominantly inherited genetic disease. The identification of the gene harboring the HD mutation in the early 1990s by a consortium of researchers was one of the biggest success stories in human genetics. Even more remarkable was the underlying disease mechanism—the expansion in exon 1 of the gene of a triplet repeat sequence (CAG) producing an abnormally long string of glutamine residues in the huntingtin protein.

The right target

One of the major challenges in devising a gene therapy for HD is ensuring that the therapeutic reaches the right network—the deep brain and cortical areas. Therapies have to reach the right circuit, and the right cells in those circuits, Davidson said. Over the years, her group has tailored AAVs for delivery to the brain, inserting peptides into exposed loops of the virion to allow for targeting and unbiased diversity for blood-to-brain delivery. Nowadays, she said, machine learning approaches can be applied for further capsid improvements.

Davidson’s CHOP lab developed a method for screening AAVs with enhanced potency for CNS therapies. After generating huge libraries containing tens of millions of novel capsids, the group performed serial enrichments to identify the most attractive capsids. After screening pools of injected capsids into two species of monkeys, a winning capsid emerged: AAV-DB-3.

Davidson’s group infused AAV-DB-3 into NHPs, looking for targeting to the putamen (base of the forebrain) and caudate regions. Those results were published in Nature Communications in 2025.  “AAV-DB-3 really stood out for its ability to transduce deep layer cortical neurons that are important” in HD, Davidson said. Moreover, the results were achieved with relatively low doses and only required a single infusion per hemisphere, outperforming the widely used AAV5.

Somatic instability

With a promising delivery vehicle identified, Davidson next addressed the therapeutic strategy, which takes aim at the somatic expansion of the CAG repeat. This codon grows longer over time in certain cells in the brain, sometimes expanding to hundreds of repeats.

MSH3 is a DNA repair protein that is required for CAG repeat expansions, as seen in mouse models of HD and other triplet repeat disorders, including myotonic dystrophy. Research led by Paul Ranum, PhD, who is a co-founder of Latus Bio, posted in a preprint on bioRxiv earlier this year, modeled the impact of lowering levels of MSH3 on somatic instability.

Ranum and colleagues used an artificial microRNA showed to lower MSH3 levels in NHPs by 48-94 percent. Computational modeling suggests that this would reduce somatic instability and delay onset of HD symptoms by many years. Early studies using a well-known HD mouse model, the Q111 mouse, to assess biodistribution, quantify knockdowns, and assess the impact on somatic CAG repeat expansion. AAV-DB-3 expression is highest in the striatum and cortex at 16 weeks, dropping MSH3 levels by 50%.

Davidson closed by emphasizing the need to ensure scalability for treatment beyond ultra-rare disorders. Latus hopes to file an Investigational New Drug application for its HD therapy, LTS-201, in the second half of 2026. At least two other biotech companies are also targeting MSH3 by other means.

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The work represents “a great example of the potential of continued mining for novel Cas effectors within the bacterial metagenomic diversity dark matter,” said Rodolphe Barrangou, PhD, Editor in Chief of The CRISPR Journal, which will shortly be publishing a paper on the Lithuanian team’s results.

“We need more diverse effectors to address the technical shortcomings of the CRISPR toolbox,” Barrangou continued. “This study is a great illustration of the potential of mining bacterial diversity.”

The Lithuanian team, including veteran gene editor Virginijus Siksnys, PhD—winner of the 2018 Kavli Prize with Jennifer Doudna, PhD, and Emmanuelle Charpentier, PhD, for CRISPR gene editing—used a hybrid approach to optimize Cas12l. By combining cryo-electron microscopy (cryo-EM) structure-guided design with artificial intelligence (AI) protein language models, the team was able to engineer a variant (Asp2Cas12l M82) that overcomes the known efficiency limitations of the Cas12l family.

Although Cas9 has widespread utility, including clinical applications, researchers have long considered its relatively large size and requirement for G-rich protospacer adjacent motifs (PAMs) problematic. The Cas12l family, discovered in the Armatimonadota bacterial phylum, offers a more compact size (867 amino acids) and recognition of a C-rich PAM site.

But wild-type Cas12l enzymes exhibit lower editing efficiencies and higher target-to-target variation compared to Cas9. According to Gasiūnas, the new M82 variant is “reliable, precise and adaptable,” and shows promise for a wide range of therapeutic applications.

“Through our continued work exploring novel Cas systems, Caszyme is focused on advancing technologies that move beyond promise into practical use.”

Path to potency

The engineering of the M82 variant proceeded in two steps. First, the Caszyme researchers solved the 3D structure of Asp2Cas12l complexed with an sgRNA and DNA to high resolution (2.51 Å). This revealed a unique “bracelet” architecture whereby the nuclease encircles the DNA target via interlocking helical bundles and a proline-rich string.

Next, the team introduced arginine substitutions at dozens of positions in the molecule to enhance electrostatic attraction to the negatively charged DNA backbone. This work included the production of an M67 variant, which provided a 7-fold improvement in indel editing over the wild-type nuclease.

To engineer further refinements, the Caszyme group turned to AI, specifically the ESM-2 protein large language model. This model predicted evolutionary hotspots considered likely to preserve or enhance function. Integrating these AI-derived substitutions—Q572R in the bridge helix and F607S in the RuvC domain—resulted in the final M82 Cas12l variant, illustrating the value of AI-supported engineering rather than deploying protein-directed evolution.

Rivaling Cas9

Gasiūnas presented data showing that M82 possesses good activity across recalcitrant gene targets, reducing the target-to-target variation that plagues many novel nucleases. In head-to-head comparisons in HEK293T cells, M82 demonstrated an average indel editing rate of 67.4%, nearly identical to that of Cas9 at overlapping target sites. This potency was consistently maintained across several delivery formats, including plasmid DNA, mRNA, and ribonucleoprotein complexes.

The Caszyme group also showed excellent M82 efficiency in homology-directed repair (HDR). In experiments targeting the AAVS1 locus, M82 facilitated a site-specific gene insertion frequency of 39%, outperforming Cas9 in the same context. Using single-stranded donor templates, HDR rates reached as high as 56%. Gasiūnas suggested that the staggered cut produced by Cas12l may inherently steer DNA repair toward precise correction rather than stochastic indels. With regard to safety, Caszyme found that M82 Cas12l maintained a high degree of on-target precision. Secondary editing signals were largely detected at or near the lower limits of assay sensitivity, suggesting a low risk of off-target cleavage.

The compact size of the M82 variant makes it an attractive candidate for adeno-associated virus-mediated delivery, which has strict limits on cargo size. “It is no secret that the CRISPR space has faced challenges and concerns in recent years,” Gasiūnas said. “However, we are confident in M82’s ability to create headroom for scientists to stand up and innovate within.”

Crowded field

Cas12l is not the only compact Cas nuclease gaining attention, of course. In a talk preceding Gasiūnas’ presentation, Zhaoshi Wu, PhD, co-founder and chief technology officer of Shanghai-based Castalysis Bioscience, presented an update on Cas12n, details of which were first published in Molecular Cell in 2023. The nuclease was touted as being the first independent CRISPR-Cas complete gene family uncovered by Chinese scientists within China’s territory.

Touted as a next-gen ultra-compact gene editing system, Cas12n (branded as alphaCas) consists of just 450 amino acids, and possesses structural similarity to TnpB. Cryo-EM structural analysis led the Chinese investigators to optimize the molecule for non-viral in vivo delivery. Preclinical experiments showed robust genome editing in a mouse model by targeting PCSK9 using lipid nanoparticle delivery, resulting in sharp drop in serum LDL levels.

Wu said his company is on target to begin its first clinical before the end of 2026. But he faced an uncomfortable moment during audience questions. Fyodor Urnov, PhD, challenged Wu’s claim that an inherent advantage of Cas12n was its safety profile compared to Cas9. Urnov pointed out that Intellia Therapeutics has two ongoing Phase III in vivo trials using CRISPR-Cas9 that show no immunogenicity concerns using LNP delivery.

Urnov later congratulated Wu on the rest of the company’s data and wished them success.

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BOSTON — A team at Children’s Hospital of Philadelphia (CHOP) led by Rebecca Ahrens-Niklas, MD, PhD, and Lindsey George, MD, has described a case of a brain tumor linked to a rare integration of adeno-associated virus (AAV).

George presented the work at the American Society of Gene and Cell Therapy (ASGCT) conference in a plenary talk selected as the “presidential abstract” by ASGCT president, Terry Flotte, MD. The study, “Neuroepithelial tumor with AAV integration after intracisternal magna vector delivery,” was published in the New England Journal of Medicine.

Over the past 25 years, some 6,000 patients have been treated with some form of AAV gene therapy. In all that time, George said, there have been no established long-term safety concerns, although genome integration events have been reported in mouse and dog studies. But the case documented by George and colleagues at CHOP suggests that the gene therapy field may need to pay more attention to this potential occurrence.

The story began with a 5-year-old boy with an inherited lysosomal disorder, severe MPS1 deficiency (Hurler subtype). The patient received enzyme replacement therapy at six weeks of age, followed by a cord blood stem cell transplant at age four months.

Investigators chose to perform gene therapy when the patient was 13 months old to deliver the iduronidase (IDUA) gene. The vector chosen was an AAV9 serotype, using a cytomegalovirus enhancer and a chicken beta-actin promoter driving the gene. The virus was administered into the boy’s cisterna magna in the base of the skull.

When the boy was five years old, a routine neurological scan revealed a large intraventricular mass that had not been observed two years earlier. Analysis of the tumor revealed it was a PLAG1-driven neuroepithelial tumor—indeed, PLAG1 expression was almost 300 times higher than in other central nervous system tumors studied at CHOP. (PLAG1 is usually only expressed during embryogenesis.)

Surgery to remove the tumor was successful. Eight months after surgery, there are no signs of tumor growth. The boy is also showing advanced neurocognitive function.

Tumor typing

George described RNA sequencing of the tumor, which revealed the fusion of a fragment of the AAV9 vector cassette to exon 5 of the PLAG1 gene on chromosome 8. The resulting transcript is predicted to encode a PLAG1 derivative containing five zinc-finger DNA-binding domains and a C-terminal transcriptional activation domain, which was previously reported to function as a transcriptional activator.

Curiously, the chimeric junction also included a segment of human chromosome 10, which George suspects originated during the vector manufacturing process. The integration event was present in about 40% of the total reads, suggesting integration into one of the two PLAG1 alleles.

George concluded her talk by noting that while the clinical outcome in this patient is so far encouraging, this is evidence that AAV integration can be associated with oncogenesis. The study underscores the need to monitor the most heavily transduced tissues after AAV gene therapy.

While the gene therapy community should be cautious in extrapolating this single case report across all AAV gene therapy programs, George said the study supports the use of the lowest feasible vector dose as well as tissue-specific promoters.

George and coworkers closed their paper, noting that, “Our findings support the hypothesis that rare AAV integration can contribute to human oncogenesis, which emphasizes the need to optimize gene delivery methods and monitor transduced tissues after treatment.”

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But a new approach suggests the system is more flexible than anyone expected. The study, published in Nature Biotechnology, is titled “DNA-guided CRISPR–Cas12 for cellular RNA targeting.”

Researchers at the University of Florida (UF) have developed the first CRISPR system that uses DNA, rather than RNA, to direct Cas enzymes to RNA targets. The platform, called ΨDNA, reprograms Cas12 nucleases to recognize and act on RNA using a DNA-based guide scaffold. The result is a fundamentally different way of controlling RNA inside cells—one “that extends Cas12 systems beyond genome editing and diagnostics to enable precise, programmable control of cellular transcriptomes and their epitranscriptomic marks,” according to the authors.

The concept is rooted in a simple biological distinction. DNA stores the cell’s long-term instructions, but RNA carries the working copies. “Those RNA copies are like Xerox copies of the original manual, and sometimes those copies have errors,” said Piyush Jain, PhD, associate professor of chemical engineering at UF and lead author of the study. Errors in those working copies can drive disease, and targeting RNA offers a way to intervene without altering the underlying genome. But RNA‑guided CRISPR systems, such as Cas13, can suffer from instability and off‑target effects. “Existing RNA-targeting CRISPR systems rely on RNA guides to find their targets,” Jain said. “While effective, they can sometimes affect unintended molecules… They can also be costly and less stable.”

ΨDNA takes a different approach. The team engineered a DNA guide that mimics the crRNA scaffold in reverse orientation, enabling AsCas12a and Cas12i1 to bind RNA and trigger strong single‑stranded DNA trans‑cleavage. As the abstract describes, “ΨDNA… enables RNA targeting by Cas12 nucleases… including 100% accurate hepatitis C virus RNA detection in clinical samples.” In human cell lines, ΨDNA achieved 70–95% knockdown of endogenous RNA transcripts, driven by mechanisms such as ribosome stalling and RNase H1 recruitment.

Jain sees the work as a conceptual shift for CRISPR. “The most meaningful advance is that we show CRISPR‑Cas12 can be reprogrammed to target RNA using a DNA guide rather than an RNA guide,” he told GEN. “That is a real conceptual shift for the field.” Until now, RNA targeting has been dominated by RNA‑guided systems. ΨDNA demonstrates that Cas12 enzymes—traditionally DNA editors—can be redirected toward RNA “while preserving strong specificity and enabling multiple functions, including RNA detection for developing diagnostics, intracellular knockdown, multiplex targeting, dual DNA and RNA targeting, and effector fusion strategies for RNA modification and potential therapeutic strategies.”

The discovery emerged from a structural puzzle. Simply swapping RNA bases for DNA bases does not work; Cas12 enzymes are thought to be tightly dependent on RNA scaffolds. “Several groups have tried to achieve DNA-guided CRISPR/Cas, but simply converting RNA bases to DNA bases doesn’t work,” Jain said. The breakthrough came from engineering a 3′ DNA handle that recreated the crRNA scaffold. Mutational screening revealed that a stem‑loop architecture was essential for activity, and recent cryo‑EM structures—solved in collaboration with David Taylor’s group at UT Austin—showed that AsCas12a has more structural flexibility than expected, allowing it to accommodate a DNA guide bound to an RNA target.

What surprised the team most was how robust the system proved to be. “It was especially exciting to see that this was not just an in vitro curiosity,” Jain said. ΨDNA worked in clinical RNA detection, achieving 100% accuracy on hepatitis C virus samples, and functioned inside cells with lower off‑target effects than Cas13d.

The platform’s modularity may be its most powerful feature. ΨDNA can be fused to RNase H1 for targeted RNA degradation or to METTL3 for epitranscriptomic editing. And because crRNA and ΨDNA can be codelivered, a single Cas12a enzyme can operate in two modes at once. “A single Cas12a effector can simultaneously edit DNA and regulate RNA,” Jain said. “This work starts to blur that boundary.”

Looking ahead, the team is expanding both the mechanistic and translational sides of the platform. They are refining guide design rules, dissecting how ΨDNA‑guided Cas12 triggers knockdown, and exploring applications in diagnostics, multiplex RNA regulation, and ex vivo therapeutic settings. One emerging direction involves using the technology to repair donor organs before transplantation.

More broadly, DNA guides offer practical advantages. They are easier to synthesize, more stable, and potentially more scalable than RNA guides. That combination could make ΨDNA a versatile platform for basic research, diagnostics, and future therapeutic engineering.

After decades of CRISPR research built around RNA‑guided systems, ΨDNA introduces a new way to direct one of biology’s most powerful tools. As Jain put it, “At its core, this is about giving us better control—not just rewriting the instruction manual but also precisely managing how those instructions are used.”

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Wyss Founding Core Faculty member David Mooney, PhD, and colleagues encapsulated a genetically engineered, therapeutic strain of E. coli bacteria within a biomaterial made from a hydrogel that was specifically designed to regulate bacterial growth and resist mechanical stresses, such as those present at physically active sites in the body, demonstrating that the bacteria could be confined for over six months.

To evaluate the material’s clinical potential, the researchers transformed the ILM into an active therapeutic system by engineering the bacteria to detect chemical signals from Pseudomonas aeruginosa, a common cause of implant-related infections. In response to the pathogen, the engineered bacteria autonomously self-destructed to release an antibacterial protein that killed the P. aeruginosa. In a mouse model of joint infection, the system successfully reduced bacterial burden, demonstrating the potential of durable, programmable ILM-based therapeutics for long-term disease treatment. The researchers suggest that their development represents a shift from passive drug depots to autonomous, responsive—and living—therapeutic systems.

“With this new strategy combining both an engineered material with designed mechanical features and genetically engineered microbes that produce therapeutic payloads on demand, we provide a generalizable framework for deploying future microbial medicines,” said Mooney. “The precision, safety, and therapeutic durability afforded by this ILM strategy could be a potential solution for treating a wider range of diseases and infections, enabling therapeutic efficacies that might surpass those of other drug delivery strategies.”

Mooney, the Robert P. Pinkas Professor of Bioengineering at SEAS, is co-senior and corresponding author of the team’s published paper in Science, titled “Implantable living materials autonomously deliver therapeutics using contained engineered bacteria,” in which the authors concluded that their collective results “… establish ILMs as a foundation for deploying microbial medicines in vivo as autonomous therapeutic depots across diverse disease settings.”

Patient recovery from many debilitating conditions and diseases could be sped up significantly and be more effective if drugs and therapeutic molecules were delivered right to where they are needed in the body, over the entire regenerative process, and in doses finely tuned to therapeutic needs. An intriguing way to achieve this is the use of implantable, synthetically engineered, living cells that can sense injury or disease-associated conditions in their environment and flexibly respond by producing the right amount of a therapeutic molecule.

“Synthetically engineered cells are emerging as living therapeutic modalities, capable of sensing physiological conditions and producing bioactive payloads in vivo,” the authors wrote. Unlike conventional drugs, these “living therapeutics” can sustain themselves in vivo and survive in many biological environments, including tumors, inflamed tissues, infected tissues, and even within human cells.

Bacteria are particularly attractive because they can be genetically programmed to release therapeutic molecules in response to specific biological signals. Bacteria can thrive in harsh physiological environments within the body, such as within infected or inflamed tissues, tissues undergoing mechanical movements, and tumors.

Some such microbial therapies have even advanced into clinical trials to treat certain cancers, metabolic disorders, and the progression of kidney stones. However, thus far, such trials have failed, and microbes are feared to also pose significant safety risks because they cannot be contained at specific sites in the body. “… controlling microbial off-­target effects remains a key safety consideration because dissemination and associated toxicity have been reported across multiple clinical contexts,” the authors continued.

Previous implantable biomaterial systems, such as hydrogels and capsule-like enclosures, have shown some success in confining microbes, but only for short periods—typically no more than two weeks. “Implantable hydrogels offer a physical strategy to confine therapeutic cells at target sites,” the investigators commented. “Such living materials hold promise as localized drug depots with the capacity to dynamically respond to diseased environments … In this work, we present an implantable material that encapsulates and confines bacteria, wherein synthetically engineered microbes produce therapeutic payloads from within.”

First author Tesuhiro Harimoto, PhD, who spearheaded the project as a postdoctoral fellow in Mooney’s group, explained further, “In the beginning, we asked the seemingly simple question, what if we could design a material that safely encapsulates drug-delivering bacteria inside and allows therapeutic drugs to pass through to where they are needed.” Although scientists have extensively studied how physical parameters of synthetic materials change with tweaks made to their composition and chemical connections, “this was a big ask since the encapsulating material had to reconcile two often contradictory features: it needed to be sufficiently ‘stiff’ so that bacteria pushing against it from the inside can’t break it apart, and sufficiently ‘tough’ to provide a enclosure that protects against external physical stresses in mechanically active tissues.”

To realize ILMs, the team started with polyvinyl alcohol (PVA), which is already used clinically, and processed it to form nanoscale interactive crystalline domains.  The resulting scaffolds are simultaneously highly stiff and tough. “Finding out how to fabricate optimal hydrogels from PVA that are crosslinked through dense crystalline domains, and how to do this in a way that keeps the enclosed bacteria alive and active, was a big part of our study,” said Harimoto. The researchers included the bacteria in their fabrication process within tiny droplets of gelatin that protected them against desiccation and selective chemical manipulations.

This strategy allowed them to fabricate an ideally stiff and tough material scaffold around the bacteria, using a combination of tolerable freeze-thaw cycles, salt conditions, and chemical treatment times. Late in the process, via a slight shift in temperature, the gelatin microgel could be dissolved to create internal voids for the bacteria to thrive in. Due to the tiny pore sizes within the PVA material, the bacteria remain constrained while the soluble molecules they produce can travel to other sites in the body.

The resulting ILM safely contained the bacteria over extended time intervals of up to six months and was resistant to repeated mechanical stresses. “We developed a hydrogel scaffold with dual mechanical features: high stiffness to regulate bacterial proliferation and high toughness to resist material fracture under physiological stress,” the investigators stated. “This design achieved complete bacterial containment for six months and withstood multiple forms of mechanical loading that otherwise caused catastrophic material failure.”

To provide proof-of-concept for ILMs, the team focused on the infection of implanted periprosthetic devices designed to treat fractures or bone loss around existing artificial joint replacements by pathogenic P. aeruginosa strains. Many treatments with periprosthetic devices fail due to infection, which goes along with inflammation and the spread of antibiotic resistance. “We evaluated the use of ILMs for periprosthetic joint infection in vivo,” they wrote. This model was designed to capture early postimplantation infection during which most infections arise in clinical settings.”

To effectively treat this and other types of infection, the therapy-delivering bacteria within the ILM needed to be genetically engineered to function as a drug depot with autonomous “sense-and-respond” capabilities. To achieve this, the team installed a synthetic gene circuit in the E. coli strain that enabled the bacteria to sense a small diffusible metabolite produced by P. aeruginosa, known as N-acyl homoserine lactone (AHL), and, in response, activate a self-destruction gene to trigger cell lysis. The self-destruction process, triggered in a fraction of ILM bacteria, resulted in release from the ILM of a synthetic P. aeruginosa-killing protein called chimeric pyocin (ChPy) that the bacteria produce continuously. ChPy is toxic to P. aeruginosa, erasing the pathogen in the local ILM environment.

“When we tethered a therapeutic ILM to a stainless steel periprosthetic device that was infected with a pathogenic P. aeruginosa strain isolated from a patient’s wound and implanted next to the femur bone of mice, it significantly reduced the pathogen burden while safely containing its engineered bacteria over a three-day treatment course,” said Harimoto. “In contrast, in mice that we treated with a non-therapeutic control ILM that did not produce ChPy, the numbers of P. aeruginosa bacteria continued to rise over the same time interval. This demonstrated the ability of therapeutic ILMs to autonomously sense and treat periprosthetic infection in vivo.”

The researchers think that specifically engineered ILMs as a novel class of therapeutics with excellent safety features and locally targeted drug release capabilities have broad potential, ranging from tissue regeneration to immune modulation in a variety of disease settings. A patent application describing the use of ILMs for drug delivery has been filed.

In their paper, the authors wrote in summary, “ILMs are distinct from other therapeutic modalities, such as drug-loaded depots and vaccines. By directly sensing pathogen-­derived signals and locally releasing antimicrobial payloads, ILMs enable rapid, antigen-independent intervention at the implant site. This localized, autonomous mode of action is well-suited for periprosthetic joint infection, where early intervention is critical.” Their collective results, the team suggests, “…establish ILMs as a foundation for deploying microbial medicines in vivo as autonomous therapeutic depots across diverse disease settings.”

In a related perspective, Kaige Chen, PhD, and Quanyin Hu, PhD, at the School of Pharmacy, University of Wisconsin–Madison, acknowledge that further work will be needed to determine whether contained living therapeutics can function in vivo over long periods. Nevertheless, they said, “The study of Harimoto et al. addresses a central obstacle to deploying living therapeutics—keeping bacteria physically separated from the surrounding tissue. Chen and Hu further note that the in vivo findings in the artificial joint mouse model  “… could advance living therapeutics from short-lived proof-of-concept systems to durable, programmable medicines.”

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“Retaining drugs within tumors is an often-overlooked dimension of drug development that nevertheless greatly impacts the therapeutic window and outcomes,” said Michael Evans, PhD, a professor in the department of radiology and biomedical imaging at UCSF and a corresponding author on the study. In fact, approaches that deliver cancer therapeutics to tumors but lack dedicated mechanisms to ensure tumor retention often lose efficacy within a few days of drug administration. 

Previously, Evans and others designed drug delivery systems called restricted interaction peptides or RIPs that can deliver diverse therapeutic cargos including cytotoxins and radioisotopes. They work by changing shape when processed by disease-associated enzymes. These allow them to embed in cell membranes, tethering their drug payloads in place, promoting cellular uptake and improving effectiveness. Building on that work, the scientists engineered RIPs to interact with fibroblast activation protein, a serine protease that is prevalent in solid tumors and fibrosis. 

Imaging studies of cancer cell cultures showed that a fluorescently tagged RIP was rapidly taken up by the cells. Then when the scientists attached an anticancer drug, monomethyl auristatin E or MMAE, to the RIP, they found that the drug-peptide combination was as effective in killing cancer cells as the drug alone. Furthermore, when the drug-peptide combination was injected into mice with human cancers, it selectively targeted tumor tissue and was more effective at shrinking tumors than the unmodified drug with fewer side effects. The scientists observed similar results when they attached RIPs to radioactive copper isotopes which are commonly used in nuclear imaging and radiotherapy. 

The scientists expect to initiate Phase I clinical imaging studies of the RIP-radioactive copper isotope pairing in human cancer patients later in 2026 in collaboration with a company that is developing RIPs into therapeutics. 

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Senior and corresponding author Rejji Kuruvilla, PhD, at Johns Hopkins University Department of Biology, and colleagues reported on their findings in Science Signaling, in a paper titled “Transcytosis-mediated anterograde transport of the receptor TrkA mediates the formation of presynaptic sites in sympathetic neurons.” In their paper, the authors concluded, “These findings provide mechanistic insight into an atypical mode of receptor trafficking and demonstrate its physiological relevance in sympathetic neuron connectivity in mice … Our study suggests that transcytosis might be a more general mechanism than now appreciated for the targeted transport of trophic and guidance receptors, adhesion and synaptic proteins, as well as ion channels.”

The axons of neurons are extremely long compared to their main cell bodies, with axon terminals sometimes residing a long distance from the cell nucleus. “Axon terminals can be meters away from cell bodies where many axonal membrane proteins with critical functions in regulating axon guidance and growth, neuronal survival, presynaptic organization, and synaptic transmission are made,” the authors wrote.

Neurons need to be able to transport these proteins efficiently across these relatively vast distances. They do this by either directly sending the protein through a secretory pathway or via an indirect mechanism called transcytosis. The latter occurs when the central cell body takes in newly synthesized proteins or surface receptors, after which they move to axons through the cell cytoplasm. “Transcytosis is an atypical endocytosis-based mechanism, where newly synthesized proteins are first inserted on cell body surfaces, internalized, and anterogradely transported to axons,” the team continued.

Transcytosis is still relatively obscure and enigmatic compared with the direct secretion method, and questions remain about how exactly it sustains the function and connectivity of neurons. “In contrast to the considerable progress made in understanding the direct secretory pathway, there is limited knowledge about transcytosis, specifically the underlying transport kinetics and organelles involved, whether it occurs in vivo, and its contributions to neuronal connectivity and function,” the investigators noted.

Seeking answers, first author Kuruvilla, together with first author Guillermo Moya-Alvarado, PhD, and colleagues, used live cell imaging and electron microscopy to peer at the movement of receptors across compartments within mouse neurons.

They visualized the trafficking dynamics and transcytosis of a receptor named TrkA. “The family of tropomyosin-related kinase (Trk) receptors provides a prominent example of membrane proteins that undergo long-distance axonal trafficking to control neuronal survival, axon growth, and synaptic transmission,” the scientists explained.

Through their study, the authors noted various shifts in speed and direction as vesicles carried TrkA from the soma to axons. Using labeled TrkA proteins, the scientists also confirmed that transcytosis occurred within nerve terminals of living mice. “Live imaging and electron microscopy of compartmentalized cultures revealed that soma surface–derived TrkA proteins underwent dynamic transport within axons, with changes in speed, direction, and the vesicular organelles that carried them as they moved from proximal to distal axon compartments,” they stated. “In mice, soma surface–labeled TrkA proteins were observed in sympathetic nerve terminals, demonstrating that transcytosis occurs in vivo.”

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