Patients rely on diagnostic tests to monitor their health efficiently and effectively. There is a lot of background work that takes place to reliably produce the components of these indispensable products. GEN recently spoke with Shana Khoury, President of Scripps Laboratories, to learn more about the in vitro Diagnostic Industry (IVD) and the challenges that suppliers face.

GEN: What are the major areas of focus for your business / laboratory?

Khoury: We manufacture antigens and antibodies as well as buffers and reagents for the IVD market. Diagnostics are the first line of defense in healthcare to help direct treatment. Patients need to have a diagnosis, such as from a blood test, before they can be treated. Our products are key components in the test kits used at centralized testing facilities, hospitals, medical offices, clinics, and other laboratory settings that specialize in diagnostic testing. Although we do not manufacture infectious disease–specific products, during the height of the COVID pandemic, we ramped up production to meet the increased demand for our reagents that are used in some of the most widely distributed COVID home tests. Aside from infectious disease testing, developing reagents for point-of-care devices and the home test market for pregnancy, fertility, and ovulation monitoring are big areas of focus for us.

GEN: What are the main challenges affecting your industry?

Khoury: Our major challenges are the scarcity of resources and raw materials. Human tissues that are donated to science have diminished in quantity and quality over the last several years. Glands and organs have never been abundant; supply has always fluctuated. Over the last few decades, we made it a priority to stockpile excess supply when available. The recent COVID pandemic brought our supply chains to a screeching halt. It is still incredibly difficult to get these raw materials, regardless of the energy and resources spent trying to obtain them. Our stockpiles allowed us to continue manufacturing at maximum capacity throughout the pandemic. We began developing a mirror recombinant catalog to mitigate the impact of future tissue scarcity and to continue to supply our customers with the products they need to meet their manufacturing schedules.

Additional challenges have also presented themselves which have required us to get creative and adapt with the changing times. As a result, we find ourselves utilizing our domestic resources more and sourcing locally when possible.

GEN: What is Scripps doing to combat the challenges?

Khoury: The supply of native tissues is not coming back. We foresaw this scarcity and have been replacing our native catalog with recombinants that are reliable, available, and cost-efficient. As technology has advanced, we have been able to shift our focus and produce recombinants that are comparable to their native counterparts. We are proud of our results. Maintaining a consistent, reliable supply of these products is absolutely critical to our IVD customers. It is imperative for us to always meet our manufacturing deadlines, so our customers’ schedules stay on track.

GEN: What would you say to customers who are reluctant to switch from native to recombinants?

Khoury: There is no other option. There is no foreseeable solution to the diminishing supply and quality issues we have been seeing over the last few decades. It is a good time to start validating recombinants and replace native-sourced proteins wherever possible. Our recombinants are different from those produced a decade ago; they perform better in IVD assays. We never release a recombinant product until we get the same, comparable results as with our native proteins. It has taken time, but customers are becoming more receptive. They realize that the short-term inconvenience of validating a recombinant gives them long-term guaranteed supply, stability, and security.

GEN: Discuss the importance of a quality system and its role in your business/industry?

Khoury: Our quality management system holds us accountable and evolves with our needs. It also helps us continually improve. It encompasses every function in our business, including manufacturing processes, training, and communications. We are ISO certified and we welcome customers to visit us to see how seriously we take quality. Our quality system ensures that we repeatedly produce products the same way and with the same performance, so we meet all expectations. If we need to make a modification, for example, due to a discontinued reagent, we inform our customers far in advance. Then we do everything possible to ensure the products we produce perform comparably or better than their predecessors.

Visit our website to see how you can benefit from Scripps Laboratories’ world-class recombinants.

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Uduak Thomas | Senior Editor

Katalin Karikó was thrust into the limelight during the COVID-19 pandemic for her pioneering work that led to key advances in mRNA vaccination—work that was awarded the Nobel Prize in Physiology or Medicine last year. During that time, the world learned some of Karikó’s story, marked by challenges, determination, and a love of science. Now, Uduak is eager to read Karikó’s story told in her own words. Breaking Through is “a testament to one woman’s commitment to laboring intensely in obscurity—knowing she might never be recognized in a culture that is more driven by prestige, power, and privilege—because she believed her work would save lives.”

Christina Jackson | Associate Editor

A new book can be seen on Christina’s desk, in her bag, or in her hand, on a weekly basis. When asked what book she plans to pick up this summer, she pointed to The Gene by Siddhartha Mukherjee. The Gene is described as offering “a definitive account of the fundamental unit of heredity—and a vision of both humanity’s past and future.” With a history marked by a cast of characters including Darwin, Mendel, Crick, Watson, and Franklin, Mukherjee also injects the story of his own family and their mental illness. Undoubtedly, we’ll see the 600-page book on Christina’s desk for another week or so, before she devours it and moves onto something else. 

Corinna Singleman, PhD | Managing Editor 

Ben Goldfarb, an environmental journalist, traveled throughout the United States and the world to analyze how roads have transformed our planet. Corinna used to study the impacts of human contamination on fish, and maintains her passion for learning about ecology and conservation in her post-research life. In Crossings, she is eager to read about the impacts of human roadways on animal commuting strategies and how we can address these issues. The book has been described as an “eye-opening account of the global ecological transformations wrought by roads” and was a New York Times Notable Book of 2023 and an Editors’ Choice. 

John Sterling | Editor in Chief 

As a journalist and biotech expert, John loves nothing more than a good story about science. For that reason, he is looking forward to digging into Kira Peikoff’s latest book, Baby X, a fictional thriller set in a world where a person can have a baby with anyone else—using just a biological sample. In it, a black market operation (called The Vault) steals cells from people (imagine a used napkin or fork) to convert them into sperm and egg. The “thought-provoking look into the near future” raises questions about the progress of technology and the impacts it has on the world.

James Lambo | Art Director 

If you ever wanted to know about how CRISPR works from a layperson’s perspective, James says “The Code Breaker is the book for you.” James (who got a jump start on his summer reading and is already deep into the book) says that biography veteran Walter Isaacson weaves a fascinating tale about how—over billions of years—bacteria have outwitted viruses. The story describes how modern-day researchers have seized this new technology to overcome modern-day diseases, including cancer and SARS-CoV-2. Doudna’s story, Isaacson writes, “is a thrilling detective tale that involves the world’s most profound mysteries, from the origins of life to the future of our species.”

Katherine Vuksanaj | Online Editorial Manager

Women in Science Now shares stories and insights of “women from a range of backgrounds working in various disciplines, illustrating the journeys that brought them to the sciences, the challenges they faced along the way, and the important contributions they have made to their fields.” This is a fitting choice for Kathy who found her own way into science publishing and makes important contributions at GEN every day. In her book, Munoz combines stories with data to illuminate the challenges women scientists face, while “highlighting research-based solutions to help overcome these obstacles.”

Alex Philippidis | Senior Business Editor

It comes as no surprise that Alex chose one of the leading books on the COVID-19 pandemic for his summer read. For two years, Alex led COVID-19 coverage for GEN, covering the latest drugs, vaccines, and other SARS-CoV-2 research on a daily basis throughout 2020 and 2021. Gottlieb’s book, which was released in the fall of 2021, is “an intense ride through the pandemic with chilling details of what really happened. It is also sprinkled with notes of true wisdom that may help all of us better prepare for the future,” notes Sanjay Gupta, MD, CNN chief medical correspondent.  

Kevin Mayer | Senior Editor

While some of us are delving into modern questions about COVID-19 and CRISPR, Kevin chose to read about questions that have been around a bit longer. Kevin plans to read “Sky In A Bottle” this summer. In his book, Pesic introduces us to chemistry, optics, and atomic physics and describes the polarization of light, Rayleigh scattering, and connections between the appearance of the sky and Avogadro’s number. He discusses changing representations of the sky in art, from new styles of painting to new pigments that created new colors for paint.

Julianna LeMieux, PhD | Deputy Editor in Chief 

As a microbiologist, I have always been a fan of Dr. Fauci’s research on HIV and his work as NIAID director to lead the nation through several viral outbreaks including Ebola, SARS, West Nile, and anthrax. I cannot wait to dive into his new memoir that “reaches back to his boyhood in Brooklyn, NY” and “carries through decades of caring for critically ill patients, navigating the whirlpools of Washington politics, and behind-the-scenes advising and negotiating with seven presidents on key issues from global AIDS relief to infectious disease preparedness at home.”

Rob Reis | Production Editor

Not all science summer reads need to be about science, per se. When GEN‘s production editor, Rob, first started reading the medical thrillers written by doctor and author Robin Cook more than 40 years ago, the main appeal was the interesting characters and compelling stories, but as Cook continues to keep abreast of the changes affecting the world of medicine, Rob increasingly sees connections between the topics covered in GEN and the stories crafted by Cook. He will not only re-read Cook’s 2023 novel, Manner of Death, this summer, but he also looks forward to Cook’s 2024 novel, Bellevue, when it gets published in December. 

The post Science Summer Reads: <i>GEN</i> Editors List Their Favorites appeared first on GEN - Genetic Engineering and Biotechnology News.

Featuring Uduak Thomas (Senior Editor, GEN), Alex Philippidis (Senior Business Editor, GEN) Jonathan Grinstein, PhD, (Senior Editor, GEN), and moderated by Corinna Singleman, PhD, (Managing Editor, GEN and IPM)

Listed below are key references to the GEN stories, media, and other items discussed in this episode of Touching Base:

The State of Omics 2024 Registration
GEN Summit

COVID-19 Infections Detected in Dried Blood Spots via At-Home Proteomic Profiling
By GEN, April 2, 2024.

CRISPR-Cas9 Targets Lung Cancer Using Cryo-Shocked Tumor Cells
By GEN, March 31, 2024.

Iambic Rhythm: AI Drug Developer Enters the Clinic, Targeting HER2 Cancers
By Alex Philippidis, GEN, April 3, 2024.

Archimedes’ Box: Diagonal Therapeutics Raises $128 Million to Discover Agonist Antibodies
By Jonathan D. Grinstein, PhD, GEN Edge, April 3, 2024.

The post Dried Blood COVID Dx, Cryo-Shocked Tumor Cell Delivery Vehicles, Verve Halts Trial, Iambic Starts Trial, and Diagonal Therapeutics Launches appeared first on GEN - Genetic Engineering and Biotechnology News.

In a new paper published in Communications Medicine titled, “Proteome profiling of home-sampled dried blood spots reveals proteins of SARS-CoV-2 infections,” the authors explained how they used deep proteome profiling of home-sampled dried blood spots (DBS) to assess the effects of SARS-CoV-2 in mildly symptomatic or asymptomatic individuals in urban areas in Sweden. “In clinical plasma and serum samples, in-depth proteomic analysis has already delivered valuable insights into the pathology and pathogenesis of COVID-19,” the researchers wrote. “Our DBS study aimed to demonstrate the utility of self-sampling and identify circulating proteins associated with SARS-CoV-2 infections by considering the serological phenotypes.” 

While this study focuses on COVID-19, Jochen Schwenk, PhD, a professor of translational proteomics, chair of HUPO’s Human Plasma Proteome Project, and senior author on the paper, noted that the methodology could apply to other conditions that require large population studies to assess the various genetic, phenotypic, and environmental factors involved in diseases development and treatment response.

The kits are developed by Swedish medtech company Capitainer. The company’s qDBS system, which was used for the study, features a capillary system mounted on a card for capturing blood from finger pricks. It includes a smart chip that ensures that patients capture precise volumes of blood—10 microliters—as well as a colored indicator that lets them know that their sample has been successfully captured. Kits also come with instructions for safely and sterilely collecting blood samples. The same kit was used in Harvard University’s VIVID study, which aimed to determine the effects of vitamin D supplementation on COVID-19 disease progression and post-exposure prophylaxis. 

According to the paper, the scientists conducted three studies with a subset of the samples they received and patients were assigned to studies based on their serostatus and self-reported information. “We compared seropositive with seronegative subjects (study 1) and donors classified into the early or post-infection phases (study 2) from the first wave of the pandemic,” they wrote. “We also studied seropositive and seronegative subjects from the third wave of the pandemic who were not vaccinated at sampling (study 3).”  

Participants in the first two studies came from the first round of tests sent out in spring 2020 while those in the third study came from the second batch of tests. In total, the team looked at data from 228 individuals for the studies.

They tested the samples for over 250 blood proteins associated with cardiovascular disease and metabolism using proximity extension assays developed by Olink Proteomics. Blood samples collected from seropositive and seronegative people early on in the pandemic revealed various proteins involved in immunity, inflammation, coagulation, and stress response, according to the results. The data also showed that blood samples collected later in the pandemic had differing levels of a virus receptor on B cells. 

Schwenk and his team will apply lessons learned from this initial study including some of the best practices they picked up for treating samples and data in future studies. They are also optimistic about the feasibility of conducting studies like this one in other disease areas. For example, a similar approach could be used to collect data from patients with seasonal allergies perhaps before and after they have received a particular intervention or treatment or to assess changes in inflammation proteins in response to allergens, he said. 

Furthermore, because the infrastructure requirements for this kind of testing are relatively minimal compared to traditional blood testing, a methodology like this could also be applied to study diseases in resource-limited regions or in populations in hard-to-reach geographic locations. 

The post COVID-19 Infections Detected in Dried Blood Spots via At-Home Proteomic Profiling appeared first on GEN - Genetic Engineering and Biotechnology News.

In the study, which was done in mice, the researchers showed that the MOF successfully encapsulated and delivered part of the SARS-CoV-2 spike protein while simultaneously acting as an adjuvant once it broke down inside cells. More work is needed to ensure that the particles can be used safely in human vaccines, but these early results are promising. 

“Not only are we delivering the protein in a more controlled way through a nanoparticle, but the compositional structure of this particle is also acting as an adjuvant,” according to Ana Jaklenec, PhD, a principal investigator at MIT’s Koch Institute for Integrative Cancer Research and one of the senior authors on the study. “We were able to achieve very specific responses to the COVID-19 protein, and with a dose-sparing effect compared to using the protein by itself to vaccinate.”

The MOF used in this study, called zeolitic imidazolate frameworks 8 or ZIF-8, is a lattice of tetrahedral units made up of a zinc ion attached to four imidazole molecules. Particles typically have diameters that are between 100 and 200 nanometers, making them small enough to get into the lymph nodes directly or through immune cells like macrophages. Prior studies showed that ZIF-8 particles can significantly boost immune responses. What is unclear is exactly how they activate the immune system. 

To answer that question, the researchers devised an experimental vaccine consisting of SARS-CoV-2 receptor-binding protein embedded in ZIF-8 particles. Once the particles entered the cells, the MOFs broke down releasing their viral protein cargo. The imidazole components of the MOFs then activate the toll-like receptors, which help to stimulate the innate immune response. 

RNA sequencing of lymph node cells from the vaccinated mice showed various immune related pathways were activated in response to the vaccine including the TLR-7 pathway, which led to greater production of cytokines and other inflammatory molecules. They also observed that mice vaccinated with the particles had a much stronger response to the viral protein, than those that received just the protein alone. 

Before these particles could be used in vaccines, scientists would have to evaluate not only their safety but also whether they could be scaled up for manufacturing on a larger scale. However, even if ZIF-8 does not work out, the researchers believe that their findings could help guide efforts focused on similar nanoparticles for delivering subunit vaccines, which are usually easier and cheaper to manufacture than mRNA vaccines.

“Designing new vaccines that utilize nanoparticles with specific chemical moieties, which not only aid in antigen delivery but can also activate particular immune pathways, have the potential to enhance vaccine potency,” Jaklenec noted. “Understanding how the drug delivery vehicle can enhance an adjuvant immune response is something that could be very helpful in designing new vaccines.”

The post Metal-Organic Nanoparticles Enable Better Vaccine Delivery, Stronger Immune Response appeared first on GEN - Genetic Engineering and Biotechnology News.

There is growing evidence that SARS-CoV-2 may not fully clear from Long COVID patients after initial infection. Instead, reservoirs of the virus can persist in patient tissue for months or even years, with recent research finding the SARS-CoV-2 virus in gut tissue more than 600 days after infection.

Persistent viral RNA or proteins have also been identified in blood samples collected from Long COVID patients, but the exact nature of the viral RNA that gives rise to this prolonged infection remains unclear.

Team will analyze tissue samples

Yale School of Medicine scientists will analyze Long COVID tissue samples to uncover mechanisms by which the virus or its proteins persist. The team will also use mouse models to test therapeutics including antivirals, antisense oligonucleotides, and innate immune stimuli such as stem-loop RNA for their potential to eliminate persistent virus, which could ultimately inform Long COVID clinical trials.

This new grant builds on an existing collaboration between PolyBio and Yale through CII, which Akiko Iwasaki, PhD, heads. Iwasaki is the Yale Sterling Professor of Immunobiology and professor of dermatology; of molecular, cellular and developmental biology; and of epidemiology; and a Howard Hughes Medical Institute investigator. Iwasaki and CII have been working to characterize the activity of human endogenous retroviruses in patients with Long COVID.

“Our hope is that by studying viral RNA persistence in Long COVID, we can better understand the pathogenesis and treatment of other related debilitating chronic conditions,” says Iwasaki.

Persistent RNA virus infection, including with enteroviruses, has been implicated in chronic conditions such as myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS), which also is a subject of ongoing research by Iwasaki as well as other scientists.

See also related GEN story: “Leaky Blood Vessels in Brain Linked to Brain Fog in Long COVID Patients.”

The post Yale Receives Grant to Fund Long COVID Research appeared first on GEN - Genetic Engineering and Biotechnology News.

“Vascular disruption has been implicated in coronavirus disease 2019 (COVID-19) pathogenesis and may predispose to the neurological sequelae associated with long COVID, yet it is unclear how blood–brain barrier (BBB) function is affected in these conditions,” write the scientists.

“Here we show that BBB disruption is evident during acute infection and in patients with long COVID with cognitive impairment, commonly referred to as brain fog. Using dynamic contrast-enhanced magnetic resonance imaging, we show BBB disruption in patients with long COVID-associated brain fog. Transcriptomic analysis of peripheral blood mononuclear cells revealed dysregulation of the coagulation system and a dampened adaptive immune response in individuals with brain fog.

“Accordingly, peripheral blood mononuclear cells showed increased adhesion to human brain endothelial cells in vitro, while exposure of brain endothelial cells to serum from patients with long COVID induced expression of inflammatory markers. Together, our data suggest that sustained systemic inflammation and persistent localized BBB dysfunction is a key feature of long COVID-associated brain fog.”

Novel technique to study Long COVID

“For the first time, we have been able to show that leaky blood vessels in the human brain, in tandem with a hyperactive immune system may be the key drivers of brain fog associated with Long COVID. This is critically important, as understanding the underlying cause of these conditions will allow us to develop targeted therapies for patients in the future,” said Matthew Campbell, PhD, professor in genetics and head of genetics at Trinity, and PI at FutureNeuro.

The team used a new form of MRI scan, dynamic contrast-enhanced magnetic resonance imaging, to identify the changes to neural vasculature. The data showed that there is reduced integrity of the blood vessels in Long COVID patients with brain fog and other cognitive impairments (memory loss, difficulty focusing, and thinking), compared with Long COVID patients without this suite of symptoms, but they do exhibit general fatigue, shortness of breath, and joint pain. For those who suffer from Long COVID symptoms for more than 12 weeks after a bout of COVID, it’s a challenge to find answers or relief. Researchers and clinicians struggle to identify and treat patients with Long COVID, who can account for up to 10% of patients who contract the SARS-CoV-2 virus.

This team also aimed to examine how COVID’s impact on the blood-brain barrier affects different categories of Long COVID symptoms.

“We investigated the functioning of the blood-brain barrier [and] established the barrier is not functioning normally in these patients,” noted Colin Doherty, PhD, professor of neurology and head of the school of medicine at Trinity, and PI at FutureNeuro in a video interview about the study’s implications. “Now we know there is a definite pathological basis for long COVID. Not only that, we have at least a range of possible treatments now to try to repair the barrier. And so, the next phase of these studies will be really exciting for the potential of a cure in the distance.”

Connecting the dots

This is by far not the first study to explore the root cause of neuropathy in Long COVID patients. Many recent reports have focused on the impacts of the disease on the immune system. Just two weeks ago, another team explored a similar phenomenon of neurological impacts triggered by viral infections. This group identified viral-induced neuropathy caused by Zika virus infection activating a lasting immune response. The common and often lasting neurological symptom of loss of smell was studied two years ago by examining inflammation markers in nasal biopsies, finding increases in those markers, and in number of T cells.

The current study opened a different door looking at the impact of viral infection on the vasculature integrity. The group wanted to examine the effect on the brain if there were disruption in the blood-brain barrier coupled with increased inflammation. This study illuminates an area of neurological research that scientists are beginning to explore. Recent work has concluded that viral infections are likely triggers for several neurological conditions, including Long COVID, multiple sclerosis (MS), and others. This study suggests that leaky blood vessels and a disruption of the blood-brain barrier may be factors in the development of these conditions.

“Our findings have now set the stage for further studies examining the molecular events that lead to post-viral fatigue and brain fog. Without doubt, similar mechanisms are at play across many disparate types of viral infection and we are now tantalizingly close to understanding how and why they cause neurological dysfunction in patients,” added postdoctoral researcher Chris Greene, PhD.

The post Leaky Blood Vessels in the Brain Linked to Brain Fog in Long COVID Patients appeared first on GEN - Genetic Engineering and Biotechnology News.

The scientists, led by Andrew Vaughan, PhD, focused on a repair pathway involving vascular endothelial growth factor α (Vegfa) and the TGF-β receptor 2 (TGF-βR2). Using animal models and human tissue samples, the scientists showed that delivering Vegfa via lipid nanoparticles (LNPs) greatly enhances modes of repair for damaged blood vessels.

Details were published recently in Science Translational Medicine, in an article titled, “TGF-βR2 signaling coordinates pulmonary vascular repair after viral injury in mice and human tissue.”

“Mice deficient in endothelial Tgfbr2 exhibited prolonged injury and diminished vascular repair,” the article’s authors wrote. “Loss of endothelial Tgfbr2 prevented autocrine Vegfa expression, reduced endothelial proliferation, and impaired renewal of aerocytes thought to be critical for alveolar gas exchange.”

“We developed a lipid nanoparticle that targets the pulmonary endothelium, Lung-LNP (LuLNP),” the authors continued. “Delivery of Vegfa mRNA, a critical TGF-βR2 downstream effector, by LuLNPs improved the impaired regeneration phenotype of endothelial cell Tgfbr2 deficiency during influenza injury.”

Vaughan’s team and other investigators had previously shown that endothelial cells are among the unsung heroes in repairing the lungs after viral infections. But Vaughan’s team noted that its work demonstrated that a “more granular understanding of the fundamental mechanisms driving reconstitution of lung endothelium” could inform efforts to facilitate therapeutic vascular repair.

“Here we’ve identified and isolated pathways involved in repairing this tissue, delivered mRNA to endothelial cells, and consequently observed enhanced recovery of the damaged tissue,” Vaughan said. “These findings hint at a more efficient way to promote lung recovery after diseases like COVID-19.”

The team found Vegfa’s involvement in this recovery, while building on work in which they used single-cell RNA sequencing to identify TGF-βR2 as a major signaling pathway. The researchers saw that when TGF-βR2 was missing, it stopped the activation of Vegfa. This lack of signal made the blood vessel cells less able to multiply and renew themselves, which is vital for the exchange of oxygen and carbon dioxide in the tiny air sacs of the lungs.

“We’d known there was a link between these two pathways, but this motivated us to see if delivering Vegfa mRNA into endothelial cells could improve lung recovery after disease-related injury,” said first author Gan Zhao, PhD, a postdoctoral researcher in the Vaughan laboratory.

The Vaughan laboratory then reached out to Michael J. Mitchell, PhD, of the School of Engineering and Applied Science, whose laboratory specializes in LNPs, to see if delivery of this mRNA cargo would be feasible.

“LNPs have been great for vaccine delivery and have proven incredibly effective delivery vehicles for genetic information,” said Mitchell, who is an associate professor of bioengineering at Penn Engineering and a co-author of the paper. “But the challenge here was to get the LNPs into the bloodstream without them heading to the liver, which is where they tend to congregate as its porous structure lends favor to substances passing from the blood into hepatic cells for filtration. So, we had to devise a way to specifically target the endothelial cells in the lungs.”

The Mitchell laboratory’s LNPs proved effective in delivering Vegfa into endothelial cells, and as a result, the researchers saw a marked improvement in vascular recovery in their animal models. Within the animal models, the researchers saw improved oxygen levels, and in some, the treatment helped them recover their weight better than the control group. These treated mice also had less lung inflammation, shown by lower levels of certain markers in their lung fluid, and their lungs showed less damage and scarring, with more healthy blood vessels.

“We’re looking forward to testing this delivery platform for other cell types in the lung, and it will be important to evaluate whether TGF-βR2 signaling is important in other injury contexts including chronic conditions like emphysema and chronic obstructive pulmonary disease,” Vaughan said. “With this proof-of-concept being well validated, we’re sure that we’ll pave the way for new mRNA-based strategies for treating lung injury.”

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Polymer nanoparticles may be the answer. With virtually unlimited options for chemical features, “you can design bespoke delivery vehicles easier than with a lipid-based system,” says Sean Kevlahan, PhD, co-founder and CEO, Nanite. “There’s (approximately) 10⁶⁰ possible polymers you can synthesize, and perhaps 10²⁵ known stars in the universe. Without the power of computation, machine learning, and artificial intelligence, you could never screen all those possible different formulations and combinations.” Polymer nanoparticles are also simpler to manufacture.

Nanite takes an engineering approach to polymer-based delivery discovery, Kevlahan says. “When we started Nanite in 2021, we found that many scientists don’t completely understand what chemical features drive certain polymers to specific destinations—the lung versus the liver, for example.”

Rather than focusing on one or two polymers, Nanite works with its pharma and biotech partners to build fit-for-purpose polymer delivery vehicles. An artificial intelligence (AI) platform called SAYER assesses polymers based upon first-principle assays, such as identifying the polymer’s composition, the charges on its surface, and how the polymer interacts with specific nucleic acid–based “cargos.”

“The power of this approach allows us to generate a huge corpus of data,” Kevlahan says. “[AI] gives us the ability to explore the polymer universe without having to actually, empirically, identify each star in that universe. Then we can use the latest computational methods to understand which chemical features drive localization to different organs and design the polymer nanoparticles accordingly with our partners.”

Likely candidates are then assessed in preclinical in vitro or animal-based experiments. To do this—at the simplest level—the team makes a diverse set of polymers, loads them with different types of nucleic acid–based cargo, pools them into one sample, and injects them into animals. Then they simply see what nanoparticles went where. All the data is fed back to SAYER to generate predictions for more accurate targeting and delivery performance.

“We haven’t identified a (cargo) size limit yet,” he reports. So far, the nanoparticles have carried molecules as small as 15 bases and as large as whole ribonucleoproteins. “You can do that easier with a polymer-based system because you don’t need complex formulation equipment to make polymer nanoparticle droplets.”

Experienced founders

Nanite’s co-founders are seasoned entrepreneurs. Kevlahan and Shashi Murthy, PhD, now CTO, previously co-founded Quad Technologies, which was acquired by Bio-Techne, and co-founder Thomas Neenan, PhD, now CBO, also co-founded AbFero Pharmaceuticals (acquired by Pharmacosmos) and Panbela Therapeutics. In addition, Murthy founded Flaskworks, acquired by Northwest Biotherapeutics. They’ve worked together in various capacities for more than a decade. Having a seasoned founding team that’s worked together such a long time resonates well with potential investors and partners.

They are also chemical engineers, so it was natural they would apply an engineering approach to building a workflow around such issues as data quality, throughput, and cost per data point. “We are each fascinated by the convergence of materials science, biology, and computation,” Kevlahan remarks.

Delivery was a pain point

“The nucleation point came during the COVID-19 vaccine research,” he says. “There was unbelievable momentum within mRNA and transient-based therapeutics … but if there isn’t a good delivery system, you don’t have a therapeutic.”

AAV vectors (whether recombinant or wild type) are limited by their small cargo size (approximately 5 kb), immunological issues (such as neutralizing antibodies that block AAV delivery), and manufacturing that is inefficient and expensive. However, rAAV gene therapy programs have successfully targeted the liver, striated muscles, and the central nervous system.

Lipid nanoparticles are the unsung heroes of the COVID-19 mRNA vaccines, he says. “They were the first big foray into nonviral mediated delivery.” They are limited, however, by their tendency to accumulate in the liver, low drug payloads, and—for solid lipid nanoparticles—drug expulsion or bursts. “To go after different diseases that are outside the liver, for example, cystic fibrosis (in the lungs) or Charcot-Marie-Tooth disease (in the peripheral nerves), you need different chemical features that are not liposomes.”

Polymers, Kevlahan thought, could be a next-generation, nonviral delivery vehicle with potential even greater than lipids. They are quite stable, are functionalized easily, can be tuned, have high loading capacities, and can deliver multiple agents with varying levels of hydrophilicity and molecular weight in one carrier simultaneously.

New company challenges

Nanite’s polymers are in early development stages with several different confidential partners. Consequently, Kevlahan says, “We’re well capitalized and we’re generating revenue.” For example, the company received $6 million in seed funding last year.

At about the same time, Nanite also received an investment of about $2.5 million from the Cystic Fibrosis Foundation and the Charcot-Marie-Tooth Research Foundation to develop a system to deliver gene therapy to the lungs and the peripheral nervous system, respectively. A priority of the cystic fibrosis program is designing a vehicle that resists the mucus that coats cystic fibrosis patients’ lungs. “Given the breadth of the design space, this goal can be readily incorporated,” he says.

A current challenge is managing version changes in successive iterations of the SAYER platform. Improvements in one area invariable necessitate adaptations in another. Kevlahan likens this to a huge flywheel. “It’s running at a certain speed. You standardize things around it. Then, when you implement a change, you also have to (consider those standards and) develop new protocols, ensure data quality is maintained, etc. Managing version changes is probably the biggest challenge when operating a platform.”

So far, Nanite connects with potential partners and investors primarily through conferences and one-on-one interactions. (Its website throughout 2023 was a single succinct landing page that will likely expand this year). “We’re an inch wide and a mile deep, producing more and more training data (for the AI) and more AI-based predictions,” Kevlahan says. In the coming year, he expects to advance Nanite’s relationships with the Cystic Fibrosis Foundation, the Charcot-Marie-Tooth Research Foundation, and additional partners. Noting that the possibilities for therapeutic delivery number in the novemdecillion (10⁶⁰) range, he says that Nanite is confident that “polymers are going to take over the whole gene delivery space.”

The post Possible Polymer Delivery Systems Outnumber the Stars appeared first on GEN - Genetic Engineering and Biotechnology News.

But Hill became more enamored with synthetic biology—engineering the genetic material of organisms to forge new characteristics. Hill decided to devote his research to generating new genetic material, specifically DNA writing, rather than speed reading. But there’s more to synthesizing long stretches of custom DNA than one might think.

“Many of the technologies and processes involved in making longer double-stranded DNA, or gene-length double-stranded DNA, are for reprogramming cells or therapeutics,” Hill told GEN Edge. “It’s not a straightforward, one-step process. It’s a multistep, complex manufacturing process with several procedures involved, many of which are quite antiquated.” 

Hill set out to understand the pros and cons of the tested DNA writing methods. From this, he developed his own thesis, which became the foundation for Elegen Bio, which Hill founded in San Carlos, California, in 2017.

Six years later, in March 2023, Elegen launched ENFINIA DNA, which claims to deliver high-complexity, clonal-quality, linear DNA up to 7 kilobases (kb) in length—all NGS verified—in as little as seven business days. Researchers can log on and, with a few mouse clicks, order DNA sequences that cover the protein-coding regions of most genes, which are typically shorter than 7 kb (one study calculated that the length of human protein-coding sequences has a median of 2.93 kb and a mean of 3.52 kb).

But Hill always intended Elegen to be more than a DNA writing service—he wants to use synthetic biology to make medicines.

“There is an intention here to focus on the ability to make DNA under a documented quality process, which we think is going to be critical to the pharma customers at large,” Hill said. What’s important, he says, is “the ability to progress from a discovery process to development to the clinical stages with maximum speed and efficiency.” 

Today, Elegen announced a collaboration and licensing agreement with GSK to enable the major pharma company to use Elegen’s proprietary cell-free DNA manufacturing technology to develop GSK’s vaccines and medicines.

The terms of the agreement include upfront fees and purchase commitments of Elegen’s ENFINIA DNA to support GSK’s vaccine and medicine development, including RNA vaccines. The agreement provides Elegen up to $35 million in near-term financial and development support and fees, in addition to sales of ENFINIA DNA and a potential equity investment in Elegen by GSK. Elegen is also eligible to receive both near-term milestone payments relating to developing new product features and a potential equity investment in Elegen by GSK.   

Hill said the partnership will likely support Elegen in advancing its DNA technology platform to become faster and produce longer DNA sequences with greater complexity for applications such as in vitro transcription (IVT) of mRNAs.

“IVT-ready DNA streamlines the work that many of these types of customers need to do to get to a point where they’ve got an mRNA vaccine or therapeutic program and are exploring a lot of candidates,” said Hill. 

Venn diagram of long DNA sequence production 

Writing large quantities of long DNA is a complex, multi-step process typically involving oligo synthesis, assembly, and amplification. “DNA production is actually not just synthesis,” said Hill. “Synthesis, properly described, is really the only step going from nucleotides to oligos. Then you take that material, assemble it, and purify it.”

There are a few approaches for the first step, oligonucleotide synthesis. Phosphoramidite chemistry has been the gold standard since the 1980s, with improvements. For example, Twist Bioscience’s core innovation was a semiconductor platform that miniaturized phosphoramidite chemistry to make single-stranded oligomers, which they use to bring down costs. 

Phosphoramidite chemistry has also been improved with the introduction of enzymatic oligo synthesis, which can produce longer DNA strands of higher quality while using less toxic chemicals and in the “biological” orientation (with an intact 5′-phosphate group). Molecular Assemblies, founded by one of the developers of enzymatic DNA synthesis, J. William Efcavitch, PhD, developed a technology that employs a template-independent DNA polymerase called terminal deoxynucleotidyl transferase (TdT), which can synthesize longer DNA sequences with fewer errors. Some companies have even developed benchtop instruments for enzymatic DNA synthesis of oligos, such as DNA Script and Evonetix. 

But Hill thinks the enzyme-based DNA synthesis approaches are marginally better than phosphoramidite chemistry in terms of error rate and do so at a much higher cost. 

It is in the subsequent assembly (or cloning) and amplification steps where Hill see the greatest opportunities because they are cell-based. The conventional method to make long DNA sequences involves a cell-based process to isolate a small quantity of a perfect sequence, amplified in what Hill views as a cumbersome, complex, and slow process that often introduces variability and complexity. Because some of these sequences are toxic to cells, cell-based methods also have limited sequence types available. 

While this is fine for most research purposes, as companies progress from discovery, where they only need small amounts of material, to clinical development, where they require larger quantities, DNA transitions from a research-use-only (RUO) product with no documentation to clinical development, which is a very different ball game. 

“To be quantitative about this, the transition from a RUO DNA to a clinical-quality DNA with a documented process typically takes months and hundreds of thousands of dollars,” said Hill. “We want to eliminate those costs and those delays for these companies.” 

Ribbon cutting 

Hill isn’t alone in seeing the issues with cell-based methods for making large amounts of long, clinical-grade DNA. For the assembly step, companies like OriCiro Genomics have developed cell-free cloning. For the amplification step, Touchlight has developed a DNA vector known as “doggybone DNA” or dbDNA—a minimal, linear, covalently closed structure—that is amplified enzymatically in cell-free conditions.

The innovation behind Elegen’s ENFINIA mostly focuses on using molecular barcodes to facilitate cell-free cloning and DNA amplification (described in the patent US20230399636A1). Research into Elegen’s other patents has to do with the use of microfluidics to facilitate cell-free cloning and DNA amplification.

But Elegen might not be alone when it comes to this approach. Ribbon Biolabs has developed patented technology based on “precise enzymatic synthesis in microfluidic reactions” (described in patent US11352619B2), which was announced nine months before Elegen on July 19, 2022. Ribbon Biolab’s patent describes a method for synthesizing dsDNA using a diverse oligo library, which they claim enables the efficient production of DNA greater than 10 kb with high speed, including a 20-kb sequence for one undisclosed pharma client.

The Elegen advantage may have more to do with their ability to drive commercial-scale mRNA synthesis for vaccine development and production. Elegen’s DNA templates—comprised of an antigen sequence and several recognition sequences, including a capping region, untranslated region (UTR), and a poly-A tail—support the IVT of an mRNA with post-transcriptional stability and translatability.

According to a poster on Elegen’s website, this capability enables a streamlined workflow and may save time and resources spent iteratively cloning, linearizing, and purifying plasmid DNA and generating master cell banks. These innovations, which could power the next generation of mRNA vaccines and cell and gene therapies, are likely a major component of what brought GSK to the table and ultimately sealed its partnership with Elegen. After all, the mRNA vaccine industry shot to great heights with the COVID-19 vaccines and is being expanded into cancer, as highlighted by the recent successes of the melanoma vaccine from Moderna and Merck. 

“Elegen’s ability to produce “NGS-verified linear DNA” up to 7 kb within a week with error rates as low as 1:70,000 per bp (99.999% per base accuracy) could be an important marker as it tries to establish a leading position in a heated race to deliver what could be the next major medicinal breakthrough. 

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