Cystic Fibrosis Research News № 7

April 2022

Nanoparticles Improve Drug Delivery for Lung Diseases

The unfortunate reality is that many inhaled drugs do not hit their target cells. Some medicine breathed into the lungs gets blocked as it travels through the oral cavity, the pharynx, and the larynx before ultimately arriving in the lungs. There are also other barriers within the lungs. In a previous edition of this newsletter, we highlighted evidence that CF mucus can inhibit  inhaled drugs as they travel through the airways – such as  the common antibiotic tobramycin. To tackle this problem, better methods for drug delivery are urgently needed. Researchers at the University of Palermo in Italy have developed a nanoparticle formulation that can more effectively deliver inhaled medications. In particular, they loaded these nanoparticles with the drug rapamycin.

In results recently published in the journal Drug Delivery and Translational Research, the Palermo team provide a very comprehensive view of what it takes to build and optimize a new drug delivery system. To start, before producing any nanoparticles, they had to decide what drug to deliver. The mTOR inhibitor Rapamycin (Rapa, for short) is already approved as a prophylactic drug to reduce the chances of organ rejection after transplantation. It is also under investigation as a COVID treatment and has recently been repurposed as a drug to treat COPD, asthma, and idiopathic pulmonary fibrosis (all lung diseases). Scientists believe that Rapa has the capacity to reduce airway inflammation. Yet Rapa has problems from a pharmaceutical standpoint. It has poor bioavailability (i.e., its ability to reach its cellular targets), and it has several toxic side effects. Hence the need for a drug delivery system that can breach physiological barriers like lung mucus, and limit cytotoxicity by ensuring that the drug is released at a steady rate only after it settles in the airways.

For this study the nanoparticles were designed with four separate components, and cobbled together as a compound construct. The molecule PHEA, a long, stringy polymer, was used as a backbone. Three other molecules were grafted on top of it: RhB, a fluorescent molecule that helped provided visibility of the nanoparticles as they were tested experimentally; PCL-SUCC, which binds to Rapa and keeps it inside of the nanoparticle capsule; and PEG, which increases the hydrophilicity (water-affinity) of the nanoparticle construct and allows it to travel through the airway unhindered.

After building the nanoparticles, the researchers ensured that they were both safe and effective. Human bronchial epithelial cells were exposed to the nanoparticles in the lab, which illustrated  that Rapa was only toxic to lung cells when administered on its own, not when it was encapsulated in the nanoparticles, presumably because the nanoparticles effectively diminish the concentration of drug that is released at any given time. In other words, the nanoparticles feeding the drug to the cells at a controlled rate rather than dousing them like a firehose.

Finally, the team explored how long Rapa molecules remain stable (and thus, functional as a drug) when added into a container of simulated lung fluid (SLF4), which is designed to mimic  the interstitial fluid that exists in the lung and bathes lung cells. To be effective, a nanoparticle carrier should keep the drug molecules relatively stable over time – and that is exactly what the researchers found in this experiment. The researchers added both nano-encapsulated Rapa and free Rapa molecules into SLF4 to compare how intact (i.e., not degraded into derivative compounds) the Rapa molecules remained after 24 hours. They found that a far greater proportion of the nanoparticle-encapsulated Rapa molecules remained intact than Rapa molecules added into the fluid without encapsulation. Specifically, 90% of nanoparticle-encapsulated molecules remained intact, while only 40% of free molecules did over the 24 hour time span. 

The preliminary success of this nanoparticle approach offers a number of possibilities in the realm of drug development for CF and other lung diseases. Better drug delivery stands to increase the clinical effectiveness and reduce the negative side effects of existing therapeutics. Effective nanoparticle carriers also make it feasible to test drugs that have not been previously considered for inhaled delivery because of poor stability in the body.

Featured Article: Craparo EF, Drago SE, Quaglia F, Ungaro F, Cavallaro G. Development of a novel rapamycin loaded nano- into micro-formulation for treatment of lung inflammation. Drug Deliv Transl Res.2022;1-14. doi:10.1007/s13346-021-01102-5.

A Survey of Current GI Treatments for CF

Historically, lung health has been the most pressing problem for people with CF. The research community has typically focused on the lungs because pulmonary inflammation and drug-resistant lung infections weigh more heavily on patient mortality. Recently, however, there has been intensified focus on the GI tract. Outlining these recent efforts, a review article drafted by scientists at the University of Lille in France discusses GI symptoms in CF, their prevalence, and potential treatments.

It has long been known that GI problems plague people with CF. Dorothy Andersen first classified CF as a disease in 1938 after discovering pancreatic lesions during the autopsy of a CF patient, so it truly has been a GI disease from the start, and doctors in the clinic do monitor GI symptoms at every patient visit. However, it is not clear that the CF modulator drugs, which do much to improve lung health, are capable of resolving all the GI problems associated with the disease. Thus, there is significant interest in exploring more closely what goes wrong in the GI tract of individuals with CF so that perhaps a  targeted therapy can be developed to improve GI symptoms.

What sort of GI maladies are especially common in CF? A lot, it turns out. The most common GI symptom, pancreatic insufficiency (PI), is diagnosed in about 85% of people with CF at birth. This is why most people with CF have to consume digestive enzymes with every meal. It helps the dysfunctional pancreas break down food and improve absorption of nutrients. Despite this intervention, many have a below average BMI, which is associated with poorer lung health. Monitoring diet and nutrition is always a key focus in the clinic. Beyond PI, a variety of other problems are present. Constipation and general abdominal discomfort are very commonplace. Meconium ileus (obstruction of the small intestine at its lower end) occurs in approximately 10% of people with CF, distal intestinal obstruction syndrome (DIOS) affects 15-20%, and gastro-intestinal reflux disease (GERD) bothers 67% of pediatric and 87% of adults with CF. The main symptom of GERD in the short run is severe heartburn, and in the long run it can increase the risk for esophageal cancer.

Along the same lines, GI cancer is extremely common in CF. Scientists think that this owes to a variety of factors, from bacterial dysbiosis (a CF gut microbiome that differs from that of the general population) to mucus obstruction and inflammation. The overall risk of GI cancer is estimated to be 8 times greater than that of the general population. For colorectal cancer the risk is elevated 10x. For small bowel cancer, it is 18 times higher! As people with CF live longer because of improvements in lung health, the prevalence of GI cancer in the CF patient population is expected to rise dramatically.

Despite this largely pessimistic picture, there is hope that some or all of these GI conditions can be treated effectively. First, small studies do suggest that there is some improvement in GI symptoms for patients who have started CF modulator therapy. Weight gain has clearly improved for individuals treated with CF modulators and there is also reason to believe that people with CF who are treated with modulators at an early age may see reversal of PI symptoms which would eliminate the need for pancreatic enzymes. That said, this reversal of symptoms would not be possible for older individuals whose pancreatic ducts, dysfunctional for many years, have long since atrophied.

Beyond modulator therapy, there is room to develop a comprehensive and innovative arsenal of treatment to improve GI health. This may include more effective pancreatic enzymes, fecal transplants to ameliorate the dysbiosis that characterizes the CF gut microbiome, and even pancreatic transplants – adding functional pancreatic islets to the dysfunctional pancreas. This is already being considered for people with type 1 diabetes, and because people with CF are highly prone to a similar condition called CF-related diabetes, it would be a logical thing to explore. In addition, other drugs approved for different GI diseases – such as the drug linaclotide which treats chronic constipation and irritable bowel syndrome – may be repurposed for CF as well.

Featured Article: Ley D & Turck D. Digestive outcomes in Cystic fibrosis. Best Practice & Research Clinical Gastroenterology. 2022; 101788.

CF Changes the Cell Surface – Do Modulators Help?

The eukaryotic cell membrane is often described in science texts as a ‘fluid mosaic’. A sea of phospholipid molecules are the primary source of structure for the cell membrane. Yet these phospholipids are not locked in place, but shuffle about the cell exterior like buoys in the ocean. Within the fluid cell membrane are larger molecules like the channel protein CFTR or the appropriately labeled ‘lipid rafts’. All cell membrane components work towards a common goal:  maintaining the integrity of the cell. Many take part in cell-cell signaling – which has a strong effect on how long cells survive, what they differentiate into, and how they respond to external threats like bacterial infection.

With this model in mind, the study of human disease has turned in recent years to an examination of how disease state influences the composition of the cell membrane – for instance, type and quantity of phospholipid molecules present. It turns out that CF is one such disease where the cell membrane composition is altered. Recently, researchers at Essen University Hospital in Germany, led by Dr. Dirk Westhölter, have attempted to determine whether highly effective modulator therapy brings the cell membrane back to a more normal, non-disease like state.

The focus of the Essen study was a type of phospholipid molecule called a sphingolipid. Sphingolipids play a key role in several signaling pathways (including those that contribute to cell differentiation and apoptosis, or cell death). All sphingosines possess a long fatty acid chain, which can vary in length – a C16 ceramide has 16 Carbons in its chain, while a C24 ceramide has 24. It has been shown that in CF patients, the sphingolipid profile is altered – with an elevated ratio of C16 ceramides to C24 ceramides (in the general population, the amount of C16 ceramides and C24 ceramides in the membrane is more evenly balanced).  CF patients have also presented with reduced levels of the sphingolipid sphingosine. These differences have been observed in previous studies and were confirmed by the Essen Team, who compared the sphingolipid profiles of 112 patients with CF to 96 healthy controls.

But what does this all mean from a clinical standpoint? It is thought that the abnormal ratio of ceramides in the cell membrane (relative to the general population) contributes to higher levels of inflammation in the CF lungs and make CF patients more susceptible to Pseudomonas aeruginosa infection. Furthermore, sphingosine actually has antimicrobial effects against both gram positive and gram-negative bacteria (like P. aeruginosa). The fact that sphingosine levels are reduced in CF patients suggests also that the abnormal sphingolipid profile is contributing to lung disease.

Fortunately, CF modulator therapy does seem to have a positive effect on ceramide and sphingosine levels. Drawing blood from 25 CF patients before and after modulator therapy, the team found that modulator treatment did indeed decrease the ratio of C16 ceramides to C24 ceramides, and also increased sphingosine levels. Both changes are expected to help reduce inflammation and susceptibility to bacterial infection. For people with CF, this is yet another positive sign that the CF modulators have strong systematic effects. In other words, they not only increase levels of the defective protein CFTR, but also help solve a lot of the related biological problems caused by CFTR dysfunction (inflammation, susceptibility to dangerous infections, etc.).  

Featured Article: Westhölter D, Schumacher F, Wülfinghoff N, et al. CFTR modulator therapy alters plasma sphingolipid profiles in people with cystic fibrosis. J Cyst Fibros. 2022;S1569-1993(22)00037-6. doi:10.1016/j.jcf.2022.02.005.

Featured Five CF Stories

It’s impossible to list all of the amazing research that is on-going for CF. Below is a quick list of a few fascinating articles that seem to show significant promise.

An Update on Phage Therapy. A novel phage therapy using bacteria-killing viruses to treat infections, has been employed extensively in Eastern European countries. However, the treatment is not approved in the USA due to lingering uncertainties concerning the dosage and administration of phage therapy drugs. However, new clinical trials are underway to gauge the safety and effectiveness of phage therapy against common CF pathogens Pseudomonas aeruginosa and the genus Burkholderia. (Current Opinion in Pulmonary Medicine)

CFTR Dysfunction May Disrupt Circadian Rhythms. Another recent review examines evidence that disturbances in circadian rhythms may contribute to poor sleep quality in CF. People with CF have long reported sleep disturbances, and a whole range of causative factors have been considered, from sinus disease to nighttime coughing due to pulmonary inflammation and infection. Researchers now believe that dysfunction in the CFTR protein itself may affect circadian rhythms directly. Further research along these lines might open the door to other studies of CFTR activity in the brain, where circadian rhythms are regulated. (Pulmonary Therapy)

COVID Pandemic Worsened Food Insecurity in the CF population. In line with previous reports in this newsletter about studies documenting food insecurity within the CF population, a new study has demonstrated how the COVID-19 pandemic specifically impacted food insecurity as well as the mental health of people with CF The number of CF households experiencing food insecurity rose from 18.8% to 21.8% during the pandemic, and individuals from food-insecure households reported heightened weight loss, worsened lung function, and poorer adherence to treatments than the rest of the CF population. Mental health outcomes were likewise poorer for individuals dealing with food insecurity. (Pediatric Pulmonology)

CF Parents Face Special Health Challenges. As many people with CF are living healthier lives on the new modulator drugs, the number of new CF parents has happily risen. However, with the dual demands of managing a chronic illness and raising a child, parents with CF tend to sacrifice elements of their own health – seeing a significant decrease in lung function (FEV1) and BMI. Women with CF tend to see even sharper negative trends in health outcomes. (Journal of Cystic Fibrosis)

New Device Can ‘Smell’ Lung Disease. New study reports an invention, the ‘e-nose’ device, which can detect biomarkers in exhaled breath with a relatively cheap arrangement of five gas sensors. Prior studies have established that exhaled breath contains biomarkers associated with pulmonary diseases – from obstructive lung diseases like cystic fibrosis and COPD to lung cancer. In a small trial, the device performed well in distinguishing healthy controls from patients with lung cancer and COPD. In the future, it may be applied to the early detection of these and other lung diseases, including the small population of people with CF who are not diagnosed until adulthood because they possess a comparatively milder range of symptoms (Materials Today: Proceedings)

Clinical Trial Watch

United Kingdom, 6 Sites. Recruiting. With a spotlight now on recognizing and treating the GI problems that plague people with CF, researchers seek to test a daily questionnaire for symptom reporting (a key first step to define the gut symptoms that most impact quality of life). The trial will involve focus groups of CF patients to help refine the questionnaire, assess the frequency and burden of symptoms reported, and test a smartphone app for symptom reporting. The questionnaire, once refined, will be made accessible to all CF patients. (Nottingham University Hospitals NHS Trust)

Kansas City, Kansas. Recruiting. For people with CF who can’t access or tolerate the modulator drugs, gene therapy is an attractive possibility. There are no gene therapies currently approved for CF, but the company 4D Molecular Therapeutics has entered Phase 1/2 clinical trials for their single-dose gene therapy drug. The drug will be assessed for its safety and tolerability in adult CF patients enrolled in the study – with the primary outcome measure of the trial being the incidence of adverse events.  (4D Molecular Therapeutics)

Multiple Sites. Soon Recruiting. The Vertex triple combination therapy (ELX/TEZ/IVA) has been tested in patients with two copies of the most common mutation (Delta-F508), and with patients that only possess one copy of the most common mutation. Now, the company will enroll patients that have zero copies of Delta-F508, but do possess a mutation that has demonstrated some positive response to modulator therapy in either clinical trials or laboratory experiments. Assessing change in lung function (FEV1) as a primary outcome, the trial hopes to broaden the set of mutations for which highly effective modulator therapy is FDA-approved. (Vertex Pharmaceuticals Incorporated)

United States, 3 Sites. Recruiting. Rates of depression and anxiety are elevated in the CF population – unsurprisingly so, given the intense daily burden of dealing with chronic disease. Cognitive behavioral therapy (CBT), a psychological treatment that involves counseling by medical professionals to change thinking patterns and develop strategies to cope with the stress of life’s daily challenges has shown promise in reducing symptoms of anxiety and depression. This pilot trial will assess the value of a CBT program for people with CF, using the rate of completion for the program and patient satisfaction in a post-intervention questionnaire to gauge its effectiveness. (Massachusetts General Hospital)

Funding News

Recent funding for CF researchers or companies.

Yale Leads the Way on Phage Therapy in the USA. Recently, Yale University has opened its Center For Phage Biology and Therapy in an effort to support phage therapy as an option for people with CF. Millions of dollars have been invested by the University so far, and researchers are already conducting a phase 1/2 clinical trial for patients with Chronic P. aeruginosa infection. In related news, the CFF has recently invested $1 million into phage therapy startup Felix Biotechnology (which lists Yale as one of its university collaborators). The biotech firm is also aiming to treat P. aeruginosa infections with phage therapy. (Yale School of Medicine, Felix Biotechnology)

Exploring New Cell Types to Understand CF Lung Disease. A team of researchers at the University of Saskatchewan were awarded C$810,000  from the Canadian Institutes of Health Research (CIHR). The grant will be used for research efforts to explore how CFTR dysfunction in two novel cell types – pulmonary ionocytes and secretory cells – contributes to disease. This information will assist efforts to develop and deliver gene therapy, ensuring that it gets to the right cells and has a strong clinical effect. (University of Saskatchewan)

CFF Offers More Funding for Nonsense Mutation Therapy. The CFF is re-upping its commitment to support Eloxx Pharmaceuticals in its quest to develop an effective therapy for people with nonsense mutations. The Foundation previously awarded Eloxx $3.6 million (June 2021) for initial clinical studies on their compound ELX-02. Their drug bypasses the premature stop signals that prevent translational readthrough of CFTR when it harbors a nonsense mutation. The CFF has just now provided the company another $15.9 million to continue these trials. (Cystic Fibrosis Foundation)

Emily’s Entourage Provides New Funding for Nonsense Mutations. Emily’s Entourage, a research-focussed foundation launched in 2011 by Emily Kramer-Golinkoff, has awarded nearly half a million dollars to two research teams. The first, led by Dr. Xingshen Sun at the University of Iowa, is developing a new animal model to test new treatments for individuals with nonsense mutations. The second team, helmed by Drs. Debadyuti (Rana) Ghosh and Hugh Smyth at UT Austin are focused on new, more effective approaches to gene therapy delivery. (Emily’s Entourage)

A Call to Action

Cystic fibrosis (CF) research is very much dependent on the strength of the CF community. It’s not simply an effort carried out by scientists in white lab coats - although there are many of them, and their work has enormous impact. Advances in research also depend on the technicians and engineers who operate the laboratory equipment that enables drug discovery, and the industrial machinery that allows drug development. Research depends on both business and marketing professionals, those who make biopharma companies viable and promote clinical trials. Successful research further depends on clinical trial coordinators, who carry out studies and work tirelessly to recruit and support patients throughout the complicated trial process. Particularly for rare diseases like cystic fibrosis, research depends on the work of foundations and patient advocates, which includes in the United States organizations such as the CF Foundation, Emily’s Entourage, CFRI, and the Boomer Esiason Foundation, as well as countless other across the globe, and hundreds of committed clinicians and researchers. Most importantly, research depends on people with CF and their devoted families and friends.

There can be no progress in CF research without patients willing to participate in clinical trials: not only to test new drugs, but also to provide, quite literally, their flesh and blood. It is with the help of patient samples that scientists can understand the damage that CF inflicts upon the human body, and also how drugs developed by the research community can remedy these damages.

This newsletter aims to pull all of these threads together; allowing the CF community to more fully appreciate how well the aims of its many members are aligned (and it extends an invitation to all readers not yet a part of the CF community, to embrace the cause and take up the task of pushing CF research forward). There’s something here for everyone - those interested in the clinical side of CF care, or in drug development, or the technical work performed in CF-centered laboratories. The newsletter also has as its objective to showcase new clinical trials; an opportunity for patients and clinicians to take part. Wherever and whoever you are in the world, you too may push CF research forward - either by direct participation, or simply by reading and sharing this newsletter with others.