X-ray imaging shows success of cystic fibrosis drugs

Keeping cystic fibrosis patients in relative health relies upon keeping airways clear and infection-free to avoid fatal lung complications but, currently, it can take several months to measure how effective a treatment is. However, scientists at Monash University, Australia, have now developed a new X-ray imaging method to allow researchers to view soft tissue structures and monitor the success of drugs. Sarah Williams speaks to Dr Kaye Morgan about the project.

Affecting the body's ability to regulate the correct movement of salt and water in and out of cells, cystic fibrosis causes thick mucus to form, disrupting patients' digestive and respiratory systems, as well as liver, pancreas and sinus function. The lungs are particularly vulnerable, with bacteria able to settle and multiply so that patients frequently suffer from potentially life-threatening lung infections.

At the root of these problems within the lung is the fact that the depth of airway surface liquid (ASL) - a watery film lining a person's airway - is diminished. In a healthy patient, the tiny hairs, called cilia, that it contains beat back and forth to expel inhaled debris and pathogens, keeping them away from the lungs. Without sufficient ASL, however, they stick to the sides of the airways, leading to infection.

Several medical treatments under development are therefore focused upon restoring ASL to the required depth to enable healthy lung function, but monitoring the success of such reatments is a lengthy process. Some studies have observed the effect of treatment upon lung tissue culture or a section of tissue in vitro, with the obvious limitation that the method of administering the drug to a live patient is not replicated.

Others have delivered treatment to a patient and compared the condition of the lung in an initial CT scan with another taken a few months later. However, the delay in measuring results, as well as factors that can vary the outcome - such as whether the patient catches a cold in the intervening months - make this a slow and frustrating approach.

Now, scientists at Monash University, Australia, have developed a new system for imaging ASL in vivo that allows researchers to see changes immediately and as a direct measure of the effectiveness of the treatment. Dr Kaye Morgan designed and tested the technique in collaboration with Monash's Professor David Paganin and Dr Karen Sui, based upon a method known as phase-contrast X-ray imaging (PCXI).

"Phase-contrast X-ray imaging has been in development for the past 20 years, and it works by looking at the changes in the X-ray phase through an object, not just changes in the X-ray intensity," Morgan explains. "In a normal X-ray image, you're essentially looking at the shadow of the structures: the bones absorb a lot of the X-ray light and so you don't see much light hitting the film behind the bones, whereas soft tissue and air absorb X-rays fairly weakly, so you don't see much contrast.

"However, we also know that as X-rays go through a material, they change direction, so if we can detect those changes in direction, we can be much more sensitive to really subtle changes in soft tissue structure."

Working off the grid
Various methods of detecting these phase changes to visualise soft tissue were already in existence but presented certain barriers to measuring ASL.

"One technique, if you have a highly coherent source, is that you simply move the detector further back from the patient, and you see the X-rays interfere and create a kind of 'edge enhancement' of soft tissue structures," Morgan says. "So that's really good if you want to image something quickly, because it's a nice, simple set-up; that technique has been used for studies of lung imaging before where we get a really strong contrast between air and tissue.

"Then there are also techniques that are very good at differentiating different types of soft tissue from each other, but these typically require several different exposures that are then combined later in software to be able to see the separate soft tissues."
The particular challenge Morgan and her team faced was ensuring that ASL could be differentiated from the underlying tissue but achieving this within a short, single exposure. Because the airways are always moving, taking multiple exposures was simply not feasible, so a fast, sensitive technique was required.

The team developed a method of 'single-grid imaging', which works by projecting a grid pattern of X-rays just upstream of the patient or sample being imaged. The detector on the other side then picks up the distortions of the pattern as the beams propagate through the tissue, in the same way that the tiles of a swimming pool appear distorted according to the shape of the water. By mapping the changes to the grid pattern, an image of the airways and of the layer of ASL can then be reconstructed.

A synchrotron X-ray source (SPring-8 in Japan) was used to project the grid because the short, bright exposure this provides allows an object to be imaged a long way from the source, Morgan reveals.

"This means that you get these really nice uniform wavefronts coming in with consistent phase, which helps you be able to detect the changes in phase brought about by the sample," she says. "So we started off imaging known samples - a Perspex sphere for example. Perspex has very similar X-ray properties to soft tissues, and we knew the thickness and the composition of the sphere, and so we could test our methods to see if the reconstructed image gave us the correct thickness and dimensions of the sample."
Having established that the method worked, the researchers then applied it to imaging isolated airways in vitro, and found that they were able to track changes in the ASL depth after delivery of treatments to those airways.

The team then tested the approach on living airways in mice, and found that the ASL depth could still be detected sufficiently behind the overlying patterns of skin and fat tissue, allowing changes to be tracked 'live' over time as treatment was administered.

Treatment development
Collaborating in the study from a medical standpoint were Dr Martin Donnelley and associate professor David Parsons, both from the Women's and Children's Hospital campus of the University of Adelaide, whose work is focused upon gene therapy for cystic fibrosis sufferers.

In the group's study, first published in the American Journal of Respiratory and Critical Care Medicine, the team started by testing sedated mice with a 'control' treatment of isotonic saline, a salty water delivered through vapourised inhalation. The subjects were then tested with an aerosol containing a long-acting epithelial-sodium-channel blocker known as 'P308', provided by US-based company Parion. For both treatments on each mouse, images were captured at three-minute intervals before and for 15 minutes after delivery.

The images of the control treatment were consistent with the anticipated result that isotonic saline would have little effect on the ASL over time. By contrast, with the P308, a significant increase in ASL surface (relative to the cartilage) was observed for all time points after, and including up to six minutes after delivery, and in ASL depth after 12 minutes.

Such results point towards a promising new treatment for cystic fibrosis sufferers but also, more particularly, an incredibly helpful imaging tool for the development of future medications. Currently, the technique has just been used within a trial setting, with Morgan and her team focusing on continuing their work to get new treatments tested and into the clinic, and using the method to track inhaled particles along the airway surface as another measurement of drug effectiveness. But the team also hope to transition their synchrotron-based technique to a compact X-ray source so that the method can be undertaken in labs around the world.

Usher in a new phase
What's clear is that the ability to image soft structures could be immensely helpful in a number of fields, for imaging the lung and other parts of the body. Indeed, some methods of PCXI are already moving towards the clinic. For example, PCXI for mammography - first clinically trialled at the Elettra Sincrotrone in Italy as part of a programme established in 2006 - has now been shown to be achievable beyond the synchrotron.

Invented at UCL in London by Professor Sandro Olivo, a technique called 'coded-aperture X-ray phase-contrast imaging' can be performed using conventional X-ray sources, while a further technique, known as 'grating interferometry phase contrast', can also be performed using a laboratory source, as demonstrated by Professor Franz Pfeiffer at TUM.

Morgan herself is very optimistic about the future of the field. "I think we're going to see phase-contrast imaging being used in clinical mammography to provide images that show more detail of the breast structure and the changes in the breast structure, because it's all soft tissue so particularly suited to phase contrast imaging," she says.

"There's also been work to monitor the condition of cartilage in joints, and of course the lungs - with lots of different respiratory diseases, being able to see changes in the lung condition earlier is particularly valuable to get on top of treatment with an earlier diagnosis. Brain imaging is another possibility in the future - differentiating the soft tissue in the brain."

She continues, "Phase-contrast imaging also has the potential to have much lower dose rates in that, to create the X-ray contrast, we're looking at the X-rays changing direction, not being absorbed by soft tissue, so we don't need the same X-ray radiation dose."

Single-grid-based PSXI, with its short, fast exposures, could be especially desirable for this reason, providing further motivation as Morgan and her team progress in their work to adapt their system to conventional X-ray sources that could one day image patients.
In the meantime, though, it's the degree to which the development of cystic fibrosis treatments is being accelerated in which their technique is having the greatest impact. These treatments could soon be in the hands of doctors and patients, restoring the lungs' ability to clear themselves and significantly improving patient outcomes.