Just a drop: A new paediatric drug monitoring method

By Elizabeth Randall
University of Birmingham, UK

This summary was highly commended by the judges for Access to Understanding 2015

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A tiny drop of blood on a card could be all that is needed to assess response to medication in children: no doctors or hospital required.

Juvenile idiopathic arthritis (JIA) and juvenile dermatomyositis (JDM) are inflammatory disorders which affect children. There is no known cure for either disease and symptoms such as joint destruction resulting from inflammation can lead to serious disability.

The first port of call in the treatment of both JIA and JDM is a drug called methotrexate. Methotrexate has anti-inflammatory properties and can help slow the disease process. In some cases this therapy, either alone or in combination with other drugs, can bring the disease under control. Unfortunately not all patients respond in the same way and the effectiveness of the drug varies a lot between children. There is also a wide spectrum of side effects associated with methotrexate which affect some more than others. It is therefore necessary to carefully monitor how well a child is responding to the drug, both in terms of how well it treats the disease and the severity of side effects.

One way to monitor this is to look at what happens to the drug after it is taken into the body. Methotrexate polyglutamates (MXTPGs) are substances produced by the cells of patients who have been administered methotrexate. There is evidence to suggest that the amount of MXTPGs present in the red blood cells of patients can indicate how well they respond to the treatment – therefore a method which can measure MXTPG levels in blood could be valuable.

Such methods already exist but are generally not suitable for patients who have only been given a low dose of methotrexate, as used in the treatment of JIA and JDM. Other tests are expensive, labour intensive or require relatively large amounts of blood drawn by a needle (venipuncture) – which is invasive and particularly undesirable when the patient is a child.

Research undertaken at Queen’s University Belfast has produced a new method which can detect MXTPGs from dried blood spots which require only tiny amounts of blood. Dried blood spots are obtained by pricking the finger and collecting a small drop of blood onto an absorbent card. This doesn’t require any clinical expertise and can be taken at home by a parent or the patient themselves – minimising any stress caused.

The researchers used blood samples from JIA and JDM patients taken by finger prick and venipuncture to compare results obtained by the new and established methods.

To measure levels of MXTPGs in blood spots, a technique called liquid chromatography-mass spectrometry (LC-MS) was used. Mass spectrometry is a chemical analytical technique which measures the mass of components of a sample. Different molecules have different masses so by measuring them it is possible to work out what the molecule is. The quantity of a molecule in a sample is determined by the intensity of the signal: samples containing a higher concentration of MXTPG will produce higher intensity signals. Liquid chromatography allows for different components of a complex mixture to be separated so that they can be measured individually by the mass spectrometer without interfering with each other.

Researchers carried out a method to extract MXTPGs from the dried blood spots into a solution so they could be analysed by LC-MS. The signal intensities of MXTPGs were then measured. To work out exactly how much MXTPG this intensity corresponded to, the signals were compared to those from blood containing known concentrations of MXTPG. These samples were made by adding measured amounts of MXTPGs to blood from healthy people.

They found that measured levels of MXTPG in dried blood spots were in close agreement with those measured in whole blood from the same patient. This suggested that the new method gives an accurate measure of MXTPGs in this more convenient sample type.

A number of experiments were carried out to ensure that the values produced were accurate and would be the same if the analysis were repeated. MXTPG levels were measured in replicate samples at different times during the day and on different days. They found that the differences between the values were within the acceptable range set out in guidelines which aim to make sure analytical methods can be reliably used in clinical settings.

The method was demonstrated to be accurate and reliable which suggests that it could now be used in clinical studies which aim to better understand how different children respond to methotrexate. This could mean that future patients who do not respond well, or suffer adverse side effects to the drug could be monitored more effectively, ultimately leading to better disease management and improved quality of life.

This article describes the research published in:

A novel dried blood spot-LCMS method for the quantification of methotrexate polyglutamates as a potential marker for methotrexate use in children
F. Hawwa, A. AlBawab, M. Rooney, L. R. Wedderburn, M. W. Beresford, J. C. McElnay PLoS One (2014) 9(2), e89908

This article was selected for inclusion in the competition by Arthritis Research UK.



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