Translational Advances

Back to the Drawing Board: Continued Study of Approved Drugs To Improve Patient Outcomes

Three recent clinical studies demonstrate the value of observing the differences in drug response within the patient population. The first, conducted by a large international team led by Rebekah Dedrick at the University of Pittsburgh, set out to gauge the current accessibility and efficacy of phage therapy for CF Patients – specifically those individuals infected with the pathogen M. abscessus. Phage therapy, though not yet an approved therapeutic approach in the US or Western Europe, is currently being administered and studied at a small number of CF centers because of its potential to fight antibiotic-resistant bacterial infections. A restraint to phage therapy is that the current panel of known phages is limited, and can only target certain bacterial isolates. As a case in point, the researchers in this study found that of 200 M. abscessus isolates from symptomatic, culture-positive patients – phages were identified to treat only 55. Furthermore, for the 20 patients in the study population who were treated with phages under compassionate use, just 11/20 saw clear clinical or microbiological responses. 

For 8 of the patients receiving phage therapy intravenously, the researchers noticed neutralizing antibodies in their blood, meaning that the patient’s own immune system may stymy the treatment effect. These findings demonstrate both the need to cultivate a broader spectrum of phages and to understand why phages that should target a particular bacterial infection fail to work effectively. Moreover, effective phage therapy will require a delivery vehicle that can dodge the patient’s immune defense (1)

While phage therapy may be potentially a new approach to treating infections, improving and broadening the set of traditional antibiotics used to treat CF patients remains a priority for CF treatment. Having a better sense of the root causes of failure, traditional antibiotics could be optimized. In this spirit a group of scientists at McGill University in Canada set out to monitor a cohort of 39 children recently infected with Pseudomonas aeruginosa and who were treated with Tobramycin.  In this study, they found that 30 patients did eradicate the bacterium after treatment, but that 9 experienced persistent infection. The team found a biological pattern in the persistent isolates: they tended to be resistant towards phagocytosis by neutrophils. Assays showed that neutrophils were less effective at engulfing and killing the persistent isolates than the other isolates. 

The findings of this study have a number of potential positive implications when viewed from a higher vantage point. Studies that identify antibiotic-resistant strains offer clinicians the opportunity to screen patients for these strains and consider alternate antibiotics or other treatment routes. They also open the door for bench scientists to study antibiotic-resistant strains more closely and figure out exactly what - on a molecular level - grants them resistance. The ultimate triumph of this line of research would be the development of new drug compounds that ward off persistent infection (2).

Additional Studies:

• A 20-year retrospective study determined the rate of drug-hypersensitivity interactions (DHR) in the CF population. While hyper-sensitivity to beta-lactams is common in the CF Population because of frequent use, severe reactions like anaphylactic shock fortunately appear to be very rare (4)
• Cross-sectional clinical study determines the relationship between vitamin D levels, lung function, and the gut microbiome in CF. Better (higher) vitamin D levels tend to correlate with greater abundance of Firmicutes bacteria, which is further correlated with more mild CF lung disease. The study highlights the importance of both vitamin supplementation and ongoing efforts to modulate the CF microbiome (5)
• Study reports on the benefits of ETI modulator therapy (Trikafta/Kaftrio) for GI symptoms using a newly developed CFAbd-Score for rating symptoms. ETI therapy was found to reduce GI symptoms substantially in the study subjects, though ultimately not to the level of healthy controls (6)

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CFTR Modulator Mechanism of Action – Much is Left to Learn

Building a more sophisticated understanding of the cellular consequences of CFTR modulation could instruct scientists on how to better optimize modulator drugs and even develop entirely new types of CF therapeutics. One way to approach this area of research is to leverage the technique of protein computational modeling, and use it to visualize how CFTR modulators interact with the CFTR protein at the level of individual amino acids. Such models tend to show the modulator compounds bind along the surface of mutant CFTR to correct its dysfunction – and further indicate which chemical groups are responsible for drug binding. The compound’s chemical structure can then be modified to enhance the strength of drug-protein binding and eliminate chemical groups that aren’t required for binding in the hopes of improving bioavailability or reducing toxicity.

Currently an international research team including researchers in the US and Germany (Vanderbilt University, Leipzig University) have developed a new multi-domain computational model of delta-F508 mutant CFTR. Previous work by other researchers over the past few years  had mainly focused on the single domain containing the delta-F508 mutation, but this study posited that the interaction between the mutant domain and the other domains of CFTR may be important for understanding the consequences of the mutation. After building the computational model, the researchers found that delta-F508 destabilized not only the NBD1 domain (the intracellular domain of CFTR containing F508), but also destabilized the NBD1/TMD interface (TMD is the transmembrane domain of CFTR). When they modeled the interaction between mutant CFTR and the CFTR corrector lumacaftor, they found that the modulator drug worked to stabilize the NBD1/TMD interface. This approach illustrated that multi-domain models can be built for other, rarer CFTR mutations, and the effectiveness of existing modulators can be tested based on how well they stabilize CFTR, and also more effectively screened for new drug candidates (1).

Meanwhile, other researchers have questioned why CFTR modulators might be more effective at rescuing certain mutations than others. Another study at Vanderbilt University,  led by Eli McDonald, has adopted an ‘interactomics’ approach to answer this question, exploring how CFTR interacts with other proteins in the cell after treatment with modulators. The study examined a string of mutations known to have variable responses to lumacaftor: G85E mutation which is non-responsive, delta-F508 is mildly responsive, and the rarer mutations P67L and L206W that are hyper-responsive. McDonald and his team found that hyper-responsive CFTR mutants P67L and 206W, after treatment with lumacaftor, interacted less with the protein-degrading components of the cell than CFTR with the  delta-F508 mutation. For those patients with these mutations, a corrected and functional CFTR is better able to alleviate CF symptoms. This implies that when used to treat delta-F508, the positive effects of CFTR correctors (which the large majority of people with CF possess), may be made even more effective if they are paired with additional compounds that reduce protein degradation. The same goes for other CFTR mutants that are less responsive to lumacaftor and other CF modulators (2).

Yet another way to interrogate the positive effects of CFTR modulators is to look beyond CFTR and ask how CFTR modulators alter the expression of all other proteins in the cell. Knowing the manner in which CFTR modulators impact the cell’s proteome holistically may allow scientists to discover additional downstream drug targets for CF and other protein-folding diseases. To this end, a multinational team of researchers from Italy and the UK observed changes in CFTR-mutant bronchial epithelial cell protein expression after treatment with lumacaftor. Using an approach called ‘spatial proteomics’, they sought to determine cell protein content and also to divine where proteins are located in the cell. While ultimately the study found no major detectable changes in protein expression – there was a noticeable rearrangement of proteins in two cellular organelles: the mitochondria and the peroxisomes. This change in protein localization after modulator treatment more closely resembles the arrangement of proteins in wild type cells. The researchers concluded that these changes may have something to do with the positive effects of CF modulator therapy. Thus, they would be worth investigating further in follow-up experiments - and their ‘spatial proteomics’ technique is worth applying further to CF and other diseases (3).

Additional Studies:

• Study describes positive effects of therapy to treat CFTR nonsense mutations. Anticodon-edited (ACE) tRNAs were delivered to bronchial epithelial cells harboring three common CFTR nonsense mutations. The study found that nonsense-mediated mRNA decay was inhibited significantly and CFTR channel function at the cell membrane was restored (4).
• Some patients, despite being eligible, don’t respond to ETI therapy as well as others. A new study has found that poor response to ETI therapy is associated with elevated levels of the cytokine TGF-beta1 in the upper airway. The cytokine was further shown to suppress CFTR expression in ETI-treated cells. The researchers discovered that the anti-hypertensive drug losartan can reduce the negative effects of elevated TGF-beta1 and may be a useful supplement to CFTR modulator therapy (5).
• It is known that the CFTR protein naturally interacts with other molecules in the cell. One such molecule, the 14-3-3 protein, has previously been found in WT cells to stabilize CFTR and improve its activity at the cell membrane. A team of researchers in Canada and the Netherlands reported the use of a macrocycle drug that can stabilize 14-3-3 – CFTR interactions and work effectively as a supplement to CFTR modulator therapy (6).

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Host-Pathogen Interaction Studies Set the Stage for Future Translational Progress

One central goal of studying host-pathogen interactions is to understand exactly how bacteria and other microorganisms are causing physiological problems in CF patients. Ultimately this knowledge can be used to craft more targeted and effective antibiotic therapies.  As such, many researchers seek to discover what bio-molecular ‘tools’  are used by bacteria and other microorganisms to siege host cells, (e.g., host-cell digesting enzymes and or other virulence factors that interfere with host cell machinery). Some research groups approach this problem mainly from the perspective of the pathogen, while others from the perspective of the host. 

A recent study in South America has focused very closely on a specific P. aeruginosa enzyme called PchP, which is known to catabolize phospholipids in the host cell membrane. The catabolism of phospholipids, specifically the phospholipid phosphatidylcholine (Pcho), provides the P. aeruginosa bacterium nutrients to grow. The researchers examined the structure of this protein with a range of computational tools, and identified the specific amino acid residues responsible for binding the Pcho phospholipid. These identified residues were then selectively mutated to see how much their absence affects Pcho binding by PchP (mutated residues whose loss had a more severe effect are considered more essential to the PchP enzyme’s proper functioning). As the authors observed, the study provided highly useful data for future ‘rational drug design’ endeavors - where drugs could be designed computationally that bind to and inhibit the function of the specific amino acids found to be essential for enzyme activity (1).

Researchers at the Grossman School of Medicine at NYU are studying hospital-acquired MRSA infections: a highly drug-resistant and dangerous form of the common CF pathogen S. aureus. The study examined the virulence mechanisms of patient-derived, hospital-acquired MRSA strains, which actually tend to be less virulent (disease-causing) when cultured in animal models than the MRSA strains typically studied in the lab. Because the existing lab models behave so differently from the strains encountered in patients, it is difficult to study the mechanisms of action by which MRSA infections wreak havoc in the human host.

The NYU research team discovered the key to this dilemma. While hospital-acquired MRSA strains did not cause noticeable disease symptoms when mouse models were initially exposed (as indicated in previous research), the study found that exposing these lab mice to antibiotics before exposure caused the MRSA strains to become dangerous, virulent pathogens. This finding is frightening in that it reinforces the link between antibiotic use and risk of colonization by drug-resistant pathogens. It also illustrates how recapitulating life-threatening human MRSA infection in mice gives researchers the opportunity to study the disease-causing mechanisms of hospital-acquired MRSA much more effectively in an animal model. In the long run, these sorts of follow-up studies could yield powerful new antibiotics (2).

Meanwhile, a recent study at the University of Michigan approaches the issue of host-pathogen interactions squarely from the patient perspective. The authors sought to understand the host risk factors for infection with nontuberculous mycobacteria (NTM) infections, which are largely unknown. Specifically, they were interested in the host microbiome community, and whether the existence (or lack thereof) of certain other microbial species predisposes patients to infection with the NTM pathogen. The study analyzed nearly 70 sputum samples from 59 people with CF, subjecting these samples to 16S sequencing (to identify which microbes were present) and metabolomics assays (to correlate the molecules that these microbes are producing). Ultimately, several metabolites were identified that seem to set NTM-positive individuals apart from non-infected CF patients. For example, they found that itaconate, which previous studies have found to act as an inhibitor of NTM bacterial proliferation, was reduced in the NTM-positive group. Other metabolites, such as certain ceramide compounds, were increased in the NTM-positive group. The metabolomics results provide the basis for a large set of potential new hypotheses and future experiments of translational value. Finding a way to restore lacking metabolites or deplete those that are over-abundant might help to treat NTM-positive people with CF, or better yet, ward off infection in those that are susceptible to NTM infection (3).

Additional Studies

• Researchers use P. aeruginosa clinical isolates from patients with CF to screen for and isolate 12 new P. aeruginosa-targeting phages from hospital wastewater. They then studied the genetic content of these phages and tested their ability to treat bacterial biofilms. This study, and others like it, are essential for building the CF phage therapy arsenal (4).
• How exactly does antibiotic treatment impact a CF patient’s microbial community? A new study found that the peak impact of antibiotic treatment on community structure (assessed by 16S sequencing) can be seen at day 4 or 5 of antibiotic treatment. This is useful information for physicians trying to determine the optimal course of antibiotic treatment (5).
• Scientists show that P. aeruginosa biofilms are not uniform, but highly heterogeneous structures. Different subpopulations of the biofilms examined in the study exhibited differential growth rates and response to antibiotics (among other differences). These findings are key to keep in mind when developing biofilm-busting antibiotic regimens – such drugs must be effective against the whole biofilm, in all its diversity (6).

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Clinical Trial Watch

Salem, North Carolina. Soon Recruiting. Despite widespread use of pancreatic enzymes to aid digestion, and the recent integration of highly effective modulator therapy into CF treatment, frequent digestive symptoms are very common throughout the CF population. Research is now turning to medicines that can target GI symptoms specifically. A new clinical trial sponsored by Wake Forest University is a case in point. It will test the efficacy of the FDA-approved antibiotic rifaximin, currently in use to treat IBS, as a tool to reduce bloating and other GI symptoms for people with CF. The study will compare reported GI symptoms in rifaximin-treated subjects compared to a placebo-treated control group (Sponsor: Wake Forest University).

Aurora, Colorado. Recruiting. For people with CF and other chronic illnesses (and their loved ones), mental health is a substantial concern. Though long recognized as important, the topic of mental health appears to be gaining traction in recent studies and now in a new clinical trial, which will test the usefulness of a voice journaling app to track and ease mental health symptoms in CF adolescents. The application, called Kintsugi, was developed by a small company in the San Francisco  Bay Area and is marketed as an ‘advanced journaling and self-care platform’. The app encourages the user to talk about their life aloud as a form of vocal journaling. Journaling has been found to help ease mental health symptoms in prior studies. Talking aloud also allows the app to analyze recordings and listen for vocal biomarkers of depression and anxiety. The study will use surveys to evaluate whether CF patients are satisfied with the application and how often they have used it over a period of 3 months. The trial will also screen for symptoms of depression and anxiety throughout the study period (Sponsor: University of Colorado, Denver).

USA, Multiple Sites. Soon Recruiting. Vertex Pharmaceuticals is launching more trials to test their most recent CF modulator combination therapy. The drug trio of VX-121 (a new corrector compound), tezacaftor (one of the correctors currently used in Trikafta), and deuterated ivacaftor (a more stable, long-lasting form of potentiator present in Trikafta) has already made it to phase 3 trials. Previous trials of the new combo therapy have shown efficacy above and beyond Vertex’s own FDA-approved triple combo therapy Trikafta. This current set of trials, all launched within several weeks of each other, will evaluate the new combo therapy in young CF patients (1-11 years old), long-term efficacy for patients 12 years or older, and efficacy in patients with moderate hepatic impairment (Sponsor: Vertex Pharmaceuticals –Trial 1,Trial 2,Trial 3).

USA, 17 Sites. Soon Recruiting. Phage therapy for antibiotic-resistant P. aeruginosa infections is quickly pushing forward from the bench to the clinic. A new phase 1b/2 clinical trial of single-dose IV phage therapy is currently enrolling patients at 17 study locations across the US. The study will evaluate whether the phage therapy does reduce bacterial colony counts in patient culture, and if it is able to treat patients successfully without dangerous adverse events (Sponsor: National Institute of Allergy and Infectious Diseases).

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Beyond the Bench

CFF Infection Research Initiative Surpasses $100 Million Funding Goal. In reaching its $100 million goal, the CFF has funded a number of startups that are developing novel antibacterial drugs. Just this year, the foundation gave a combined $6 million to Felix Biotechnology and BiomX Inc. This money will fund the P. aeruginosa phage therapy drug candidates of both companies. The latter company’s candidate is already heading into Phase 1b/2a clinical trials. The foundation also gave $3.5 million to Arrevus Inc. The company is developing a compound to treat MRSA, a highly drug resistant form of the common CF pathogen S. aureus (Cystic Fibrosis Foundation). 

New Grants Fund Research to Treat Stomach Pain and Hearing Loss in CF. The stomach pain project is led by Dr. Dhiren Patel at St. Louis University - he has been funded by the Institute of Clinical and Translational Sciences to push forward on his work with a neurostimulation device for CF patients. The device, which is already FDA approved for people with irritable bowel syndrome, sits in the ear and sends electrical impulses to the brain through cranial nerve bundles, specifically targeting brain regions involved in pain processing to ease the feeling of abdominal pain. Patel will test the device on 11-18 year old CF patients and gauge its effectiveness. At Stanford University, Dr. Alan Cheng has received funding from RNID (a hearing loss foundation) and the Cystic Fibrosis Trust in the UK. Cheng’s goal is to develop new aminoglycoside antibiotics that do not put patients at a risk of hearing loss. Traditional aminoglycosides like tobramycin, which are in common use to treat CF bacterial infections, are known to cause hearing loss as a side effect (St. Louis University, RNID)

Lupin Gets Tentative FDA Approval for Generic Equivalent of Kalydeco. The company develops both novel drug technologies and generic drugs - which utilize the same active compounds as brand-name drugs but are generally much lower in price. Generic drugs can be manufactured once the patent on the brand name drug expires - which is usually in the range of 10-20 years after a drug is initially patented. The tentative FDA approval gives Lupin the ability to begin manufacturing its generic equivalent of Kalydeco (Kalydeco is composed of the Vertex potentiator ivacaftor alone). Kalydeco, the first Vertex modulator ever approved for patients, is still in use by a subset of the CF population that possesses class II mutations like G551D (Lupin)

Drug Development News: Progress Towards Phage Therapy and Gene Therapy for CF. Two companies have made waves this month with drug-related announcements . Carbon Biosciences, a Massachusetts-based company, announced a $38 million partnership with the CFF to develop a viral gene therapy for CF patients. BiomX, a company based in Israel that received funding earlier this year as part of the CFF’s Infection Research Initiative, announced the dosing of its first two patients in a Phase 1b/2a clinical trial (their compound, BX004, is a phage therapy to treat P. aeruginosa infection) (Carbon Biosciences, BiomX)

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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.