Category Archives: Transhuman News

After years of researching the SON gene,Erin Eun-Young Ahn, Ph.D., may have found the cause behind an extremely rare disease.

After years of researching the SON gene, Erin Eun-Young Ahn, Ph.D., may have found the cause behind an extremely rare disease.(Photography: Nik Layman)Since the early 2000s, Erin Eun-Young Ahn, Ph.D., associate professor in theUniversity of Alabama at BirminghamDepartment of PathologysDivision of Molecular & Cellular Pathology, has been studying the SON protein and gene. The SON gene makes a protein, also called SON, that is required for the body to develop and grow normally. While Ahn is one of the worlds leading experts on the SON gene, she had no idea that her work would ultimately help determine the cause behind an extremely rare disease known as Zhu-Tokita-Takenouchi-Kim syndrome.

ZTTK syndrome is a severe multi-system developmental disorder characterized by delayed psycho-motor development and intellectual disability. Common clinical features of ZTTK include intellectual and developmental delays, brain malformations, muscle abnormalities and facial asymmetry. Little is known about this disease, including its cause, until Ahn discovered that ZTTK syndrome is the result of a genetic mutation of the SON gene.

Ahns journey began in 2014 when a physician from California contacted her about a pediatric patient who suffered from developmental and intellectual disabilities. These included late milestones in language and cognitive processing. Ahn has published research showing that SON function is important in the RNA editing step, called RNA splicing. RNA delivers instructions to cells from DNA on which proteins to produce. The physician found Ahns postdoctoral research publication and reached out for help.

The patient underwent standard genetic testing panels and tests for gene mutations on known genes, which all came back normal. Finally, the doctors ran exome sequencing a test developed in the last decade that can identify more undiscovered variants in an unbiased way. They found that the only gene mutation this patient had was in the SON gene.

This was the first finding of this specific gene mutation in humans, Ahn said. We knew that SON is overexpressed in several types of cancer cells, but we didnt know whether the mutations in the DNA sequence of the SON gene that cause loss-of-function really existed in the human patient. It was really eye opening.

Ahn made a case report on this single patient, showing how in cancer there is an overproduction of SON, and with an underproduction, there are developmental and intellectual delays like those present in the first patient. The journal to which she submitted the work suggested she locate other cases. With the assistance of a website tracking undiagnosed diseases and a database of gene mutations, Ahn found a few more incidents indicating DNA sequence changes in the SON gene. She reached out to those patients physicians and the involved researchers in the U.S. and Europe, finding they had the same symptoms as her original pediatric patient.

We went from one case to five cases, Ahn said. And a few months after that, the information began to spread rapidly. Clinicians and genetic counselors started talking, and the information became international.

Ahns group eventually analyzed clinical symptoms of 20 patients who carry mutations in the SON gene. They also conducted experiments using the cells obtained from the patients and demonstrated that an insufficient amount of the SON protein leads to defective RNA splicing in patients, which in turn causes abnormal brain development and metabolism. Their findings were published in the journal,American Journal of Human Genetics, in September 2016. The publication played a major role in designating this new disease caused by SON mutations as ZTTK syndrome.

Typically, the human body houses two copies of the SON gene, and Ahn found that those with ZTTK have one copy of the normal SON gene and one copy of the mutated SON gene. This mutation in the SON gene means that the body cannot produce as much of the SON protein needed for its cells to develop properly. This lack of cell development leads to the developmental and intellectual delays found in patients with this disease. There are currently no reported cases of mutations found in both copies of the SON gene, which suggests that mutations in both copies may be extremely detrimental to human development.

The mutations show up during development but are not inherited from the parents, which is why awareness of the syndrome by clinicians is key to connecting patients with resources, Ahn said. They have to talk to their doctor and a genetic counselor and get the exome sequencing done to get the diagnosis.

In her role as one of the worlds primary researchers on the SON gene, Ahn became a point of contact for patients and their families, connecting families all over the world. This led to the establishment of a Facebook group for the syndrome, which now has more than 245 members. Last year, several of the parents of children, together with Ahn, created theZTTK SON-Shine Foundationthat showcases personal stories of families learning how to live with this syndrome.

We developed the ZTTK SON-Shine Foundation, because we wanted to create a sense of community among people affected by this disease so they do not feel isolated, Ahn said. This is a way that parents of children with ZTTK can help and share information with each other in hopes of improving the quality of life for every patient with ZTTK syndrome.

In addition to connecting families with this challenging rare disease, the foundation hopes to spread awareness of the syndrome to clinicians in particular.

One of the goals of the ZTTK SON-Shine Foundation is making a network of clinicians and researchers who can share their expertise to help the families affected by this syndrome, Ahn said. Exome sequencing reports typically indicate the results in scientific terms, but when patients receive those, they dont understand what that means. So, I often provide the families with an explanation of what these genetic codes mean in laymans terms, so that they can have a clear understanding. I am very grateful that I have opportunities to help the patients and families.

Ahn is no longer alone in the study of ZTTK, but working with researchers across the globe on different manifestations of this syndrome in patients, from issues ranging from bone development to kidney function to metabolism and the immune system.

We hope some part of our finding will contribute to a medical treatment, Ahn says, And we know our findings can provide them with information. We dont know whether we can find a cure, but if we know more about how SON mutation affects patients metabolism, kidney issues, bone structure, neurological features, and immune system, we can do a lot for patient care and prevention and alleviate their symptoms.

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UAB researcher shines light on a rare disease that causes developmental and intellectual delays - The Mix

The first year that the National Institutes of Health (NIH) funded a group of Yale scientists to explore links between viruses and cancer, U.S. troops evacuated Vietnam, Gerald Ford was president, and the movie Jaws broke box office records.

The scientists wrote their 400-page proposal on typewriters and made 20 paper copies on Xerox machines. They put it all into a big box and sent it through the U.S. mail. It was 1975.

Their research pleased the NIH so much that the agency renewed the granteight times over 45 years. Titled Molecular Basis of Cancer Virus Replication, Transformation, and Innate Defense, it became the longest-running program project grant at Yale, and the third longest at the NIH. It brought more than $50 million to Yale labs and resulted in nearly 500 publications, many of them groundbreaking. The grant helped launch the careers of hundreds of scientists who trained under its leadership, including several on the Yale faculty.

Three of the grants principals are still at Yale: Daniel DiMaio, MD, PhD, Waldemar Von Zedtwitz Professor of Genetics, professor of therapeutic radiology, professor of molecular biophysics and biochemistry, and deputy director of Yale Cancer Center; Joan Steitz, PhD, Sterling Professor of Molecular Biophysics and Biochemistry; and I. George Miller, Jr., MD, John F. Enders Professor of Pediatrics and professor of epidemiology and of molecular biophysics and biochemistry. The grant has had a major impact on how we study viruses, said DiMaio, the principal investigator for the last 25 years. Otherwise, it wouldnt have lasted so long.

Theres lots of competition out there. Every five years the NIH looked at us closely to see if we were still productive and still a good investment. For many cycles of renewal, they decided that we were. After 45 years, he added, the grants three leaders decided not to reapply. Were sun-setting it. Its time to let a new generation take over. It is also time to applaud some of the grants research highlights. The human genome was sequenced about 20 years ago, but the first genome ever sequenced was funded by this NIH grant almost 25 years earlier, when Sherman Weissman, MD, Sterling Professor of Genetics and the grants first principal investigator, described the genetic makeup of a virus named SV40.

He developed some of the earliest techniques for sequencing nucleic acids, said DiMaio. That had a profound impact on medicine, and it came from studying tumor viruses. Before his death in 2020, another biochemist on the grant, Charles M. Redding, MD, Professor of Genetics, showed how DNA molecules can recombine to alter genes and proteins, which in turn can cause cancera crucial discovery. A former member of the program, David C. Ward, PhD, used the program funding to develop a technology called fluorescence in situ hybridization (FISH). It allows researchers to map chromosomes by locating specific DNA sequences and this technology is a standard diagnostic and research tool in labs worldwide. Steitz is a founding member of the grant program, which helped fund her landmark discovery of small noncoding RNAs made by viruses.

It turns out that RNAs arent just messengers, she said, but are also regulatory elements inside cells, and are important to be able to make an oncogenic virus. Weve discovered a lot of noncoding RNAs, and each new discovery brings all sorts of insights into how viruses are able to successfully infect cells.

Joan didnt just discover them, added DiMaio. She figured out how they work and discovered a lot of new chemistry and structural biology. It opened up a new field. Steitz identified some of those RNAs in collaboration with Miller, another founding member of the program grant. At the time, scientists knew that viruses caused cancer in animals, noted Miller, but nobody believed cancers in people were caused by viruses. Miller showed that Epstein-Barr Virus (EBV), a human virus, caused lymphomas in monkeys. Th is was the the first time a human virus had been shown to cause cancer in a primate, providing definitive evidence of its cancer-causing activity. Researchers now know that about 15 percent of all human cancers are caused by viruses. The grant also supported Millers groundbreaking discovery about how EBV gets activated.

Miller and Steitz collaborated to characterize a related virus that causes Kaposi sarcoma. The grant also supported DiMaios pioneering research into identifying viral oncogenes, and how turning them off stops cancer cells from growing. More recently, the grant funded his studies about how viruses get into cells.

It sounds simple, he said, but virus entry is a complicated process with hundreds of cellular proteins involved. Weve discovered some cellular proteins that are important for infection, determined how they work to support infection, and learned some new cell biology.

These breakthroughs stemmed from the basic science supported by the grant. Viruses educate us about every aspect of molecular biology and cell biology and immunology, said Miller. We keep on learning things from viruses that are applicable to cancer and to many other problems. If you want to make vaccines, for instance, you have to understand what the virus is doing.

The grant brought together people from many departments. We all look at virology from different perspectives, said Steitz. DiMaio is primarily a geneticist, Steitz a biochemist, and Miller a pediatrician. When we get together, continued Steitz, we have people coming in from many different disciplines and its great.

Their collaborations introduced each other to different approaches and techniques that influenced the direction of their research. Steitz started with bacterial viruses, then moved into animal viruses after conversations with Miller. Steitz helped Miller understand the advantages of using modern molecular techniques instead of cultivating viruses.

Weve really transferred knowledge back and forth, said DiMaio. Thats something very special about this grant. Were not working in isolation; we helped each other and molded each others careers. In turn, the partners in this program grant have molded the careers of several hundred grad students and postdocs who were trained under them and are now making their own contributions to the field and paying it forward with their own students. Its a long legacy, said DiMaio, like a huge extended family.

You can see evidence of that legacy in whats happening now with COVID19, said Steitz, whose career has helped us understand how RNA works. A lot of work on the immunology of this disease was done here, and the most effective COVID-19 vaccines are RNA-based vaccines.

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A 45-year Legacy of Research and Collaboration < Yale School of Medicine - Yale School of Medicine

While at the till in a clothes shop, Ruby received a call. She recognised the womans voice as the genetic counsellor she had recently seen, and asked if she could try again in five minutes. Ruby paid for her clothes, went to her car, and waited alone. Something about the counsellors voice gave away what was coming.

The woman called back and said Rubys genetic test results had come in. She did indeed carry the mutation they had been looking for. Ruby had inherited a faulty gene from her father, the one that had caused his death aged 36 from a connective tissue disorder that affected his heart. It didnt seem the right situation in which to receive such news but, then again, how else could it happen? The phone call lasted just a few minutes. The counsellor asked if Ruby had any questions, but she couldnt think of anything. She rang off, called her husband and cried. The main thing she was upset about was the thought of her children being at risk.

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Over the next few weeks, she Googled, read journal articles, and tried to become an expert patient in what was quite a rare genetic disorder. There wasnt much to go on, and, not being a scientist herself, it was hard for her to evaluate what she did find. She learned that a link between mutations in this particular gene and connective tissue problems had only recently been discovered. A few years earlier this disease did not exist, or at least it had yet to be named.

Over time, some details emerged. Nobody had ever seen her own familys particular mutation in anyone else. So that meant it was very hard to know what to make of her situation. Her risk of a heart problem was surely increased, but nobody could say by how much.

From that initial phone call, it was six months before Ruby was seen by any other medical professional. She saw a cardiologist first, followed by a series of other specialists, since each appointment seemed to trigger a chain of others. The outcome was that Ruby would have regular body scans, and she began to take medication to lower her blood pressure, which she was told to do as a precaution for the rest of her life. She was also told to avoid anything that would cause her body to suddenly jolt. The vagueness of what this meant in practice became another source of worry. Should she carry on playing basketball, for example? She had always loved going abroad, but now travel insurance became exceptionally hard for her to get, partly because nobody knew how to categorise her.

Ruby believes that it was definitely better to have been informed of her genetic inheritance, because in her case there were things she could do to lower the risk of it becoming a real problem. But it took a long time for her to understand that she was not actually ill. She was only at risk of being ill. In fact, nothing had actually changed; she had only become aware of a possible future.

Every one of us is susceptible to one illness or another to some extent. As science progresses, many more of us will find ourselves in Rubys situation; drowning in estimates and probabilities that play games with our mind and our identity, and require us to make difficult decisions about our health and how we live. Every one of us will be shown to be subtly suboptimal. Or every one of us will be shown to be special. It depends on how you look at it. As Andrew Solomon writes in Far from the Tree: The general culture feels that deaf children are primarily children who lack something: they lack hearing. The Deaf culture feels that they have something membership in a beautiful culture.

We must be very careful in defining what constitutes disease or disability, especially as our ability to link genes with human traits expands. Bill Bryson puts it like this in The Body: A Guide for Occupants: Twenty years ago about 5,000 genetic diseases were known. Today it is 7,000. The number of genetic diseases is constant. What has changed is our ability to identify them.

Even in the hard data, things get messy. For example, someone who has inherited an immune system gene called HLAB27 is about 300 times more likely to develop the autoimmune disease ankylosing spondylitis. Around 8% of people in the UK have this gene variant and most do not suffer from the disease. Whats more, inheritance of this gene may be useful in fighting HIV. About one in 300 people infected with HIV are able to control the virus so that they dont go on to develop Aids, at least for a very long time, and HLAB27 occurs frequently in these people. So theres a yin and yang to genetic inheritance that is hard to fathom, even for experts.

One day, a watch that can measure a few simple things about your body will be seen as a laughably primitive tool. In the future, a whole cloud of information will be available and you must decide how much you want to delve into it. The agricultural, industrial and digital revolutions affected our environments and societies, but the genetic revolution equips us individually with new powers, and each of us will need to decide for ourselves if and when to deploy them. One way we should be preparing now is by making sure society is scientifically literate, and that our children are educated to understand risk, probability, genetic diversity and health.

Perhaps the insight to hold on to is that we are not merely our genes, our cells, our microbiome or our brain. We are all these things, but we are also more. How we see ourselves and others the stories we tell and the philosophies we live by are going to be just as important to our wellbeing.

Daniel M Davis is a professor of immunology at the University of Manchester and author of The Secret Body.

Far from the Tree: Parents, Children and the Search for Identity by Andrew Solomon (Vintage, 18.99)

The Body: A Guide for Occupants by Bill Bryson (Black Swan, 9.99)

The Code Breaker: Jennifer Doudna, Gene Editing and the Future of the Human Race by Walter Isaacson (Simon & Schuster, 30)

Link:
The big idea: How much do we really want to know about our genes? - The Guardian

REYKJAVIK, Iceland, Dec. 2, 2021 /PRNewswire/ -- In a study published today in Nature genetics, scientists at deCODE genetics , a subsidiary of the pharmaceutical company Amgen, demonstrate how measuring the levels of a large number of proteins in plasma at population scale when combined with data on sequence diversity and RNA expression dramatically increases insights into human diseases and other phenotypes.

Scientists at deCODE genetics have used levels of five thousand proteins in plasma targeted on a multiplex platform at population scale to unravel their genetic determinants and their relationship with human disease and other traits. Previous studies of the genetics of protein levels either consisted of much fewer individuals or tested far fewer proteins than the one published today.

Using protein levels in plasma measured with the Somascan proteomics assay, scientists at deCODE genetics tested the association of 27 million sequence variants with plasma levels of 4,719 proteins in 35,559 Icelanders. They found 18,084 associations between variants in the sequence and levels of proteins, where 19% are with rare variants identified with whole-genome sequencing. Overall, 93% of the associations are novel. Additionally, they replicated 83% and 64% of the reported associations from the largest existing plasma proteomic studies, based onthe Somascan methodand the antibody-based Olink assay, respectively.

The levels of proteins in plasma were tested for associations with 373 diseases and other traits and yielded 257,490 such associations. They integrated associations of sequence variants with protein levels and diseases and other traits, and found that 12% of around fifty thousand variants reported to associate with diseases and other traits also associate with protein levels.

"Proteomics can assist in solving one of the major challenges in genetic studies: to determine what gene is responsible for the effect of a sequence variant on a disease. In addition the proteome provides some measure of time because levels of proteins in blood rise and they fall as a function of time to and from events," said Kari Stefansson CEO of deCODE genetics and one of the senior authors on the paper.

Media contact:Thora Kristin AsgeirsdottirDecode genetics+354 894 1909

SOURCE deCODE genetics

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Decode Genetics Publishes the Largest Ever Study of the Plasma Proteome - PRNewswire

TEL AVIV, Israel Two million years ago, were humans already the king of the hill on planet Earth? Researchers at Tel Aviv University say evidence points to early humans being apex predators, meaning they sat atop the food chain as the most formidable hunters around.

The study of prehistoric diets finds large mammals going extinct in many regions of the globe, along with the depletion of animal food supplies at the close of the Stone Age, forced humans to progressively expand plants into their diet, until they had no option but to tame both animals and plants and become farmers.

So far, attempts to reconstruct the diet of stone-age humans were mostly based on comparisons to 20th-century hunter-gatherer societies, explains Dr. Miki Ben-Dor in a media release. This comparison is futile, however, because two million years ago hunter-gatherer societies could hunt and consume elephants and other large animals while todays hunter-gatherers do not have access to such bounty. The entire ecosystem has changed, and conditions cannot be compared. We decided to use other methods to reconstruct the diet of stone-age humans: to examine the memory preserved in our own bodies, our metabolism, genetics, and physical build. Human behavior changes rapidly, but evolution is slow. The body remembers.

Dr. Ben-Dor and collaborators compiled roughly 25 examples from over 400 scholarly works addressing whether Stone Age people were specialist predators or generalized opportunistic feeders. Most of the teams evidence comes from studies of genomics, metabolic processes, physiology, and morphology of early humans.

One prominent example is the acidity of the human stomach, Dr. Ben-Dor adds. The acidity in our stomach is high when compared to omnivores and even to other predators. Producing and maintaining strong acidity require large amounts of energy, and its existence is evidence for consuming animal products. Strong acidity provides protection from harmful bacteria found in meat, and prehistoric humans, hunting large animals whose meat sufficed for days or even weeks, often consumed old meat containing large quantities of bacteria, and thus needed to maintain a high level of acidity.

Another indication of being predators is the structure of the fat cells in our bodies. In the bodies of omnivores, fat is stored in a relatively small number of large fat cells, while in predators, including humans, its the other way around: we have a much larger number of smaller fat cells, explained Dr. Ben-Dor. Significant evidence for the evolution of humans as predators has also been found in our genome. For example, geneticists have concluded that areas of the human genome were closed off to enable a fat-rich diet, while in chimpanzees, areas of the genome were opened to enable a sugar-rich diet.

The team used archaeological findings to enhance the data gathered from human biology. As an example, studies of stable isotopes found in the remains of ancient people, together with evidence of human-specific hunting behaviors, reveal that humans were expert hunters of big and mid-sized animals with a higher percentage of body fat. With this comparison, it became clear that humans were not only hypercarnivores but that they killed huge animals and obtained over 70 percent of their calories from meat as well.

Hunting large animals is not an afternoon hobby. It requires a great deal of knowledge, and lions and hyenas attain these abilities after long years of learning. Clearly, the remains of large animals found in countless archaeological sites are the result of humans high expertise as hunters of large animals. Many researchers who study the extinction of the large animals agree that hunting by humans played a major role in this extinction and there is no better proof of humans specialization in hunting large animals, Dr. Ben-Dor explains.

Most probably, like in current-day predators, hunting itself was a focal human activity throughout most of human evolution. Other archaeological evidence like the fact that specialized tools for obtaining and processing vegetable foods only appeared in the later stages of human evolution also supports the centrality of large animals in the human diet, throughout most of human history.

The collaborative model that scientists at Tel Aviv University (TAU) have been working on for over a decade offers a dramatic shift in the way we think about evolutionary history. Unlike the widely held belief that humans attribute their survivability to their nutritional adaptability, which enabled them to mix the killing of animals with the use of fruits and vegetables, the vision that is developing here shows that humans evolved primarily as carnivores of big animals.

Archaeological evidence does not overlook the fact that stone-age humans also consumed plants, the study author adds. But according to the findings of this study plants only became a major component of the human diet toward the end of the era.

Following the discovery of genetic variations and the style of unusual primitive tools for preparing plant foods, the investigators came to the conclusion that beginning approximately 85,000 years ago in Africa and approximately 40,000 years ago in Europe and Asia, progressive growth in plant food intake and dietary diversification occurred in line with changing ecological circumstances.

Additionally, the regional distinctiveness of the stone tool way of life grew, which is comparable to the variety of hunting tools in 20th century communities in terms of its origins and development. Throughout the two-million-year time frame in which humans were the most dominant species, scientists found extensive spans of uniformity and consistency in primitive tools, no matter how different the surrounding environment was.

Our study addresses a very great current controversy both scientific and non-scientific, says Prof. Ran Barkai. For many people today, the Paleolithic diet is a critical issue, not only with regard to the past but also concerning the present and future. It is hard to convince a devout vegetarian that his/her ancestors were not vegetarians, and people tend to confuse personal beliefs with scientific reality.

Our study is both multidisciplinary and interdisciplinary. We propose a picture that is unprecedented in its inclusiveness and breadth, which clearly shows that humans were initially apex predators, who specialized in hunting large animals. As Darwin discovered, the adaptation of species to obtaining and digesting their food is the main source of evolutionary changes, and thus the claim that humans were apex predators throughout most of their development may provide a broad basis for fundamental insights into the biological and cultural evolution of humans.

This study is published in the American Journal of Physical Anthropology.

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Humans were already the dominant predatory species on Earth 2 million years ago - Study Finds

Achieved mean 5.6-fold HBG mRNA induction at 20mg and mean 6.2-fold at 30mg after 14 days of once-daily dosing, further supporting potential of FTX-6058 to provide a functional cure

Continues to be well-tolerated at higher doses with no serious adverse events observed to date

New mechanism data demonstrate potent downregulation of BCL11A and MYB, key repressors of fetal hemoglobin

On track to initiate enrollment in Phase 1b clinical trial in people with sickle cell disease and to submit an IND for treatment of other hemoglobinopathies by year-end 2021

Company to review results on conference call, including guest KOL Dr. Gerd Blobel, at 8:00 am ET today

CAMBRIDGE, Mass., Dec. 06, 2021 (GLOBE NEWSWIRE) -- Fulcrum Therapeutics, Inc. (Nasdaq: FULC), a clinical-stage biopharmaceutical company focused on improving the lives of patients with genetically defined rare diseases, today announced positive results from the 20mg and 30mg dose cohorts in healthy adult volunteers in its Phase 1 clinical trial of FTX-6058. The company also shared new preclinical mechanism data showing that FTX-6058 downregulated known repressors of fetal hemoglobin (HbF). FTX-6058 is an investigational oral HbF inducer that is being developed for the treatment of sickle cell disease and other hemoglobinopathies, such as beta-thalassemia.

Data from the 20mg and 30mg dose cohorts demonstrated a mean 5.6-fold induction and a mean 6.2-fold induction in HBG mRNA, respectively, at day 14. These increases were higher than those observed in the previously reported 2, 6 and 10mg dose cohorts. In preclinical studies of FTX-6058, increases in HBG mRNA have consistently translated to the same fold increases in HbF protein. Notably, human genetics show that 2-3-fold increases in HbF are associated with significantly improved outcomes, and even functional cures, in people with sickle cell disease. FTX-6058 has now demonstrated greater than a mean 2-fold induction starting with the 6mg dose.

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Despite progress in the treatment of sickle cell disease, existing therapies either offer limited benefit or, in the case of gene therapy, are not amenable to the great majority of patients and carry certain risks, said Gerd Blobel, MD, PhD, Frank E. Weise III Endowed Chair in Pediatric Hematology at Childrens Hospital of Philadelphia. The strategy of increasing the levels of fetal hemoglobin is based on solid genetic and clinical data. It can substantially reduce mortality and morbidity, and in cases where HbF reaches greater than 25-35% of total hemoglobin, lead to asymptomatic disease. The emerging clinical data on FTX-6058, combined with the new preclinical data showing that it downregulates BCL11A and MYB, two validated HbF repressors, is encouraging.

The data for FTX-6058 continue to exceed our expectations, said Bryan Stuart, Fulcrums president and chief executive officer. We believe the fold increases in HBG mRNA that we have now seen at multiple doses, starting at 6mg once-daily, have the potential to translate to levels of HbF protein that could provide a functional cure for people with sickle cell disease. Additionally, with the new insights into the mechanism of action, there's now a clear relationship between FTX-6058 and HbF induction that further affirms our conviction. We remain on track to begin enrolling people with sickle cell disease in our Phase 1b trial by year-end, with an eye toward reporting initial data, including HbF protein levels, in the second quarter of next year.

FTX-6058 Continues to be Well-Tolerated and Achieved Maximal HBG mRNA Induction at Higher Doses

The Phase 1 randomized, double-blind, placebo-controlled trial was designed to evaluate the safety, tolerability, and pharmacokinetics (PK) of ascending doses of FTX-6058 (NCT04586985). In the single-ascending dose (SAD) cohorts, healthy volunteers received one dose of either placebo or 2, 4, 10, 20, 30, 40 or 60mg of FTX-6058. In the multiple-ascending dose (MAD) cohorts, healthy volunteers received a once-daily dose of placebo or 2, 6, 10, 20 or 30mg of FTX-6058 for 14 consecutive days. Each MAD cohort had six subjects on drug and two on placebo. Food effect was also studied in a separate 20mg dose cohort. Exploratory measures were included in the MAD cohorts to assess target engagement, as well as changes in HBG mRNA and HbF-containing reticulocytes (F-reticulocytes). A 6mg dose cohort in people with sickle cell disease was recently added to this trial to further inform PK and pharmacodynamic modeling for future dose selection. All other cohorts in the trial have been completed, and data from the 2-40mg SAD cohorts and 2-10mg MAD cohorts were reported in August 2021.

Consistent with the earlier reported data, FTX-6058 has been generally well-tolerated with no serious adverse events reported to date and there were no discontinuations due to treatment-emergent adverse events (TEAEs) across all SAD and MAD cohorts. Across all cohorts, all TEAEs deemed possibly related to FTX-6058 were mild (Grade 1 or 2) and resolved. There was one Grade 4 TEAE in the 10mg MAD cohort and one Grade 3 TEAE in the food effect cohort, both of which were determined to be unrelated to FTX-6058. Data continued to show dose-proportional PK, with a mean half-life of approximately 6-7 hours in the MAD cohorts, supporting once-daily dosing, and no food effect was observed with FTX-6058. Data from the MAD cohorts continued to show robust target engagement, as evidenced by an approximately 75-95% reduction from baseline in H3K27me3 after 14 days of treatment.

The data also showed higher-fold induction of HBG mRNA at the higher doses, with FTX-6058 achieving maximal rate of HBG mRNA induction in the 20mg and 30mg cohorts. Maximal HBG induction has not yet been achieved with the higher doses of FTX-6058. Persistent HBG mRNA induction was observed for 7-10 days after treatment. F-reticulocytes also increased by a mean of 1.8-fold in the 20mg cohort and a mean of 2.4-fold in the 30mg cohort as of the safety follow up visit, which was seven to 10 days after conclusion of dosing. Increases in F-reticulocytes of any magnitude are a first indicator that HBG mRNA is translating to HbF protein production, which Fulcrum anticipates observing in the Phase 1b trial that will dose people with sickle cell disease for up to three months.

HBG mRNA Mean Fold Induction for FTX-6058 versus Placebo

2mg*

6mg*

10mg*

20mg

30mg

Mean FoldInduction

P-value

Mean FoldInduction

P-value

Mean FoldInduction

P-value

Mean FoldInduction

P-value

Mean FoldInduction

P-value

Day 7

1.28

0.3494

1.94

0.0135

2.08

0.0063

2.06

0.0072

2.29

0.0025

Day 14

1.20

0.5122

2.45

0.0025

3.54

<0.0001

5.63

<0.0001

6.15

<0.0001

Safety Follow-up (Day 21-24)

1.21

0.3736

2.75

<0.0001

3.22

<0.0001

6.45

<0.0001

6.13

<0.0001

F-Reticulocyte Mean Fold Increase for FTX-6058 versus Placebo

2mg*

6mg*

10mg*

20mg

30mg

Mean FoldIncrease

P-value

Mean FoldIncrease

P-value

Mean FoldIncrease

P-value

Mean FoldIncrease

P-value

Mean FoldIncrease

P-value

Day 7

0.53

0.1070

1.02

0.9524

0.83

0.6214

0.71

0.3831

1.50

0.2928

Day 14

0.88

0.6881

1.25

0.4895

2.23

0.0180

1.00

0.9880

1.71

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Fulcrum Therapeutics Announces Additional HBG mRNA Induction from Higher Dose Cohorts in Phase 1 Healthy Adult Volunteer Trial of FTX-6058 for Sickle...

Introduction

Thyroid cancer (THCA) is one of the most common malignant tumors in human endocrine system and head and neck.1,2 The most common pathological type of thyroid carcinoma is papillary thyroid carcinoma (PTC), accounting for about 80% of the total number of THCA.35 Most thyroid cancers have good prognosis, the 5-year survival rate is more than 95%.6,7 Although the incidence rate of THCA is increasing year by year, the molecular biological mechanism of thyroid carcinogenesis and development is not clear.8,9 At present, the gold standard of preoperative diagnosis of THCA is fine needle aspiration biopsy (FNAB), but 2030% of FNAB results are uncertain or suspicious, and these patients need diagnostic surgery to identify the characteristics of tumors.10,11 The application of molecular markers is expected to help FNAB improve the ability of preoperative diagnosis of THCA.

Weighted gene co expression network analysis (WGCNA) is considered to be an effective network-based method, which can highlight the co-expressed gene modules and study the relationship between gene modules and phenotypes more effectively.1214 WGCNA has been successfully applied to explore the functional co expression modules and central genes of different diseases, such as pancreatic cancer,15 breast cancer16 and oral squamous cell carcinoma.17

In this study, we used WGCNA and other analytical methods to explore RNA data and clinical information of patients with thyroid tumor. Finally, four hub genes (CCDC146, SLC4A4, TDRD9 and MUM1L1) related to prognosis and transcription level were identified and verified, which showed good diagnostic potential and clinical relevance, and could be used as molecular markers in clinical diagnosis, treatment and prognosis of THCA.

The RNAseq expression data and related clinical traits of THCA were obtained from TCGA database (https://portal.gdc.cancer.gov/) and GEO database (http://www.ncbi.nlm.nih.gov/geo/). A total of 568 patient samples were obtained from TCGA data, including 568 samples, 510 THCA samples and 58 normal samples. Data preprocessing were used to process the raw data for perform background correction and quantile normalization, including robust multi-array average (RMA) background correction and the affy R package. The false discovery rate (FDR) <0.05 and log2FC 2 were used as the cut-off value to screen differentially expressed genes, which laid the foundation for further construction of co-expression network.

Co-expression modules are gene sets with high topological overlap similarity. The WGCNA package of R software was used to construct gene co-expression network of differentially expressed genes.11,12 This analysis procedure can identify highly related genes of differentially expressed genes, and genes with the same pathway or function can be clustered in similar gene modules. The cut-off value of co-expression module was set as P < 0.05. In order to further explore the dissimilarity of gene modules and visualize them, we select a cutting line for the module dendrogram and merge a few modules.

In order to explore the gene modules related to clinical features of THCA, the correlation between phenotype and module eigengenes was calculated, and the significance of each gene module was evaluated.18 The gene modules significantly related to clinical features (P < 0.05) were selected for further study.

To further explore potential function of the key modules, Gene Ontology (GO) term analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis were conducted to be described and visualized (DAVID, http://david.abcc.ncifcrf.gov/).1921 The significance level was set as p-value <0.01 and FDR <0.05.

Hub genes usually have important biological functions and are highly associated with other nodes of the module. The module membership of each gene is calculated, and the module membership value of the hub gene is higher. To verify the reliability of the hub gene, GSE33630, GSE29265, GSE6004 and the Human Protein Atlas database (https://www.proteinatlas.org/)22 were used for further validation. Kaplan-Meier plotter was used for survival analysis.

Based on the differential analysis, a total of 3712 genes from TCGA for co-expression analysis were calculated. We also excluded cases with incomplete clinical information. The Pearsons correlation coefficient was used to cluster the sample. We draw a sample clustering tree after removing outliers (Figure 1A). Moreover, we selected the power of = 18 as the softthresholding (Figures 1B and C). Finally, 11 modules were screened out based on average hierarchical clustering and dynamic tree clipping (Figure 2). As shown in Figure 3A, the interaction between the 11 co-expression modules indicates that each gene module is independently verified in the network. Cyan module and grey module were highly correlated with sample type by Pearsons correlation analysis (Figure 3B). Therefore, these two modules were selected as clinically important modules for further analysis. In addition, the eigengene dendrogram and heatmap plotted were drawn to explore groups of related eigengenes and the dendrogram of all modules (Figures 3C and D).

Figure 1 Clustering of samples and determination of soft-thresholding power. (A)The clustering was based on the expression data of TCGA, which contained 568 samples, 510 THCA and 58 normal samples. The color intensity was proportional to sample type (normal and THCA), sex, age and disease status. (B) analysis of the scale-free fit index for various soft-thresholding powers (). (C) Analysis of the mean connectivity for various soft-thresholding powers.

Figure 2 Construction of co-expression modules by WGCNA package in R. (A) The cluster dendrogram of module eigengenes. (B) The cluster dendrogram of genes in TCGA. Each branch in the figure represents one gene, and every color below represents one co-expression module.

Figure 3 Identification of Key Modules. (A) Interaction relationship analysis of co-expression genes. Different colors of horizontal axis and vertical axis represent different modules. (B) Heatmap of the correlation between module eigengenes and the sample type of THCA. (C) Hierarchical clustering of module hub genes that summarize the modules yielded in the clustering analysis. (D) Heatmap plot of the adjacencies in the hub gene network.

Moreover, GO and KEGG analysis were conducted for the two key co-expression modules. The results show that the MEcyan module was involved in the important biological functions and signaling pathways related to tumorigenesis and development, such as protein binding, thyroid hormone synthesis, autophagy-animal, Insulin resistance, cell proliferation (Figures 4A and B). In the MEgrey module, functions are mainly enriched in transcriptional misregulation in cancer, cell adhesion molecules (CAMs), complexity and coagulation cascades, and ECM-receptor interaction and signal transduction (Figures 4C and D).

Figure 4 Plot of the enriched GO and KEGG terms in two key co-expression modules. (A) GO enrichment analysis of MEcyan module. (B) KEGG pathway enrichment analysis of MEcyan module. (C) GO enrichment analysis of MEgrey module. (D) KEGG pathway enrichment analysis of MEgrey module.

We screened the Mecyan module and MEgrey module as candidate prognosis genes. Through the Log rank test (p < 0.05) for further overall survival analysis, 4 hub genes (CCDC146, SLC4A4, TDRD9 and MUM1L1) were identified (Figure 5). Kaplan Meier survival curve of overall survival analysis showed that THCA patients with low expression levels of 4 hub genes had poor prognosis. Then, GSE33630, GSE29265, GSE6004 and the Human Protein Atlas database were used to validate the expression status of the 4 hub genes. As shown in Figures 6A and B, the volcano map and expression heat map of the differential RNAs in GSE33630 (45 normal samples and 60 THCA). The common genes between differential RNAs and the MEcyan and MEgrey module were identified by overlapping them in GSE33630 were presented in Figures 6C and D. The results showed that 4 hub genes in two key modules were also differential RNAs in GEO33630. Moreover, the transcriptional level of hub genes were verified in GSE29265 (Figure 7) and GSE6004 (Figure 8). In addition, the translational level of 4 hub genes also were verified by the human protein atlas database (IHC) (Figure 9).

Figure 5 Survival analysis of 4 hub genes based on the Kaplan-Meier plotter. The patients were stratified into high- and low- expression groups according to the median expression. (A) CCDC146. (B) SLC4A4. (C) TDRD9. (D) MUM1L1.

Figure 6 Validation of hub genes in GSE33630. (A) Volcano plot visualizing DEGs in GSE33630 (45 normal samples and 60 THCA). Fold Change=2, adj P=0.05. (B) heatmap hierarchical clustering reveals DEGs in cancer groups compared with those in control groups. (C) Identification of common genes between DEGs and the MEcyan module by overlapping them. The two hub genes in the MEcyan module were also DEGs in GSE33630 (D) Identification of common genes between DEGs and the MEgrey module by overlapping them. The two hub genes in the MEgrey module were also DEGs in GSE33630.

Figure 7 Validation of 4 hub genes in the transcriptional level. (AD) Validation of hub genes in GSE29265.(*P < 0.05, **P < 0.01, ***P < 0.001).

Figure 8 Validation of 4 hub genes in the transcriptional level. (AD) Validation of hub genes in GSE6004 (****P < 0.0001).

Figure 9 Validation of 4 hub genes in the translational level. (AD) Validation of 4 hub genes by The Human Protein Atlas database (IHC).

THCA is a rare malignant tumor, accounting for less than 1% of human malignant tumors.13 However, it is the most common cancer in the endocrine system and the cause of death for most endocrine cancers.4,5 The occurrence and development of THCA is a multifactorial disease process, involving a variety of molecular mechanisms.2 At present, many published studies mainly focus on the molecular mechanism of single gene in THCA, but ignore the interaction between genes due to its limitations.2326 Due to the development of big data, gene network is used to analyze the origin and development of various cancers. Therefore, in order to further explore novel and accurate molecular biomarkers for prognosis, we use RNA sequencing data and clinical information from TCGA and GEO databases to explore and verify potential key modules and hub genes through bioinformatics analysis of WGCNA.

In this study, we screened 2 key modules (MEcyan module and MEgrey module) from TCGA dataset by WGCNA analysis. 4 hub genes (CCDC146, SLC4A4, TDRD9 and MUM1L1) were then further screened and verified using the GEO database and survival analysis. Considering the functional and pathway enrichment analysis, the two key co-expression modules were significantly enriched in thyroid hormone synthesis, autophagy-animal, cell proliferation, transcriptional misregulation in cancer, cell adhesion molecules (CAMs), and ECM-receptor interaction and signal transduction. At the same time, we also found that these significantly expressed functional annotations and signaling pathways have been reported in THCA and many other cancers.2730

At present, studies on these five hub genes have been reported, and a large number of studies have shown that their expression plays an important role in the occurrence, development and prognosis of many tumors. It has been found that SLC4A4 contributes to the occurrence and development of tumors, and its involvement in tumor biological processes is specific.31 It is reported that mir-223-3p promotes tumor cell proliferation and metastasis by reducing the expression of SLC4A4 in renal clear cells.32 Gerber et al. Confirmed that the low expression of SLC4A4 in thyroid carcinoma and its diagnostic value.31 SLC4A4 has been reported to be associated with poor prognosis in patients with colon adenocarcinoma. The low expression of SLC4A4 is associated with lymph node invasion and distant metastasis of colon adenocarcinoma. At the same time, SLC4A4 expression is associated with the invasion of immune cells in colon adenocarcinoma. It may be a biomarker and therapeutic target for the diagnosis and prognosis of colon adenocarcinoma.33 Another study have reported that downregulation of expression in TDRD9-positive cell lines causes a decrease in cell proliferation, S-phase cell cycle arrest, and apoptosis, which can be used as a marker for prognosis and as a potential therapeutic target in a subset of lung carcinomas.34 More importantly, one study by Wang et al suggested that TDRD9 was significantly related to the prognosis of THCA. CCDC146 was a potential therapeutic strategy for lymph node metastasis of breast cancer.35 MUM1L1 has not been previously reported to be associated with cancer. The results show that the occurrence and development of tumor may be regulated by multiple genes, which may provide more research strategies for the diagnosis and treatment of THCA.

There are two deficiencies in this study. First, the results were not verified in clinical samples. Considering the high reliability of high-throughput sequencing expression data and the sufficient number of samples included in the study, this deficiency can be made up to a certain extent, but it can not completely replace the significance of clinical sample verification. Second, the selected molecules have no functional validation. Although the signal pathway of differential genes was analyzed by KEGG pathway in this study, the newly discovered molecules that have not been reported should be functional verified.

In summary, based on the TCGA database, we analyzed the gene expression profile of THCA and successfully identified four hub genes associated with THCA prognosis, which showed good diagnostic potential and clinical relevance as molecular markers for clinical diagnosis, treatment and prognosis of THCA.

THCA, thyroid cancer; WGCNA, weighted gene co-expression network analysis; TCGA, The Cancer Genome Atlas; GEI, Gene Expression Omnibus; PTC, papillary thyroid carcinoma; FNAB, fine needle aspiration biopsy; FMA, robust multi-array average; FDR, false discovery rate; GO, Gene Ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes; CAMs, cell adhesion molecules.

This study was approved in accordance with the Ethical Standards of the Institutional Ethics Committee of University of Chinese Academy of Sciences - Shenzhen Hospital and with the 1964 Helsinki declaration and its later amendments or comparable Ethical Standards.

The results of this study are based on the data from TCGA (https://www.cancer.gov/tcga) and GEO database (http://www.ncbi.nlm.nih.gov/geo/). We thank all the authors who provided the data for this study.

This work was supported by the Startup Fund for scientific research, University of Chinese Academy of SciencesShenzhen Hospital (Grant No. HRF-2020012); and Guangming District Soft Science Research Project (Grant No. 2021R01063).

The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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Hub Genes as Prognostic Candidates of Thyroid Cancer | IJGM - Dove Medical Press