Chief Investigator:

Dr Roger Yazbek

Funding Amount:

$74,931

Recipient:

Flinders University

Overview:

Existing technologies for detection and monitoring of intestinal damage and repair in children are costly and invasive, leading to stress and unnecessary risks. New tests are needed that are non-invasive, rapid, and more suitable for a paediatric setting. We have developed a new, non-radioactive, stable-isotope breath test that can detect a marker of intestinal health and function and have demonstrated the specificity of this test in cells grown in the lab. We will now prove the efficacy of this test in a rat model of intestinal damage to provide proof-of-concept data for future human studies.

Research Outcomes:

Researchers:

Roger Yazbek, Ross Butler, Gordon Howarth, Catherine Abbott

Research Completed:

2018

Research Findings:

We found that the 13C-ODCbreath test was able to non-invasively detect ODC activity in rats as a possible marker of gastrointestinal function. Although the breath test did not directly correlate to the time course of intestinal damage, the breath-signal did appear to correlate to stress events in the animals (for example the beginning of the study, when animals were first receiving injections). This interesting result may speak to further applications of the breath test, and warrant further investigation.

Additionally, the ODC breath signal at the final breath test correlated with measurements of crypt depth in the small intestine, suggesting that the breath test was reflective of changes in the proliferative compartment of the intestine. Interestingly, contrary to the published literature, the ODC inhibitor, DFMO did not effectively inhibit ODC activity. Past studies of DFMO have only been able to measure its inhibitory properties using static measurements of ODC activity in ex vivo tissue. Our new 13C-ODC breath test may have broader applications in the in vivo development of new ODC inhibitors.

Key Outcomes:

The major aim of this proposal was to determine whether a breath test for ornithine decarboxylase (ODC) activity could be used to quantify intestinal repair and damage in an in vivo rat model of 5-Fluorouracil (5FU) induced intestinal damage.

The animal study was completed in July 2017, and tissue and data analysis commenced at the completion of the animal study. Intestinal damage was induced in groups of rats using the chemotherapy agent, 5-fluorouracil (5FU). Rats were breath-tested over a time-course of 12 days at pre-determined time points of pre-intestinal damage and post-intestinal damage to non-invasively measure ODC activity as a marker of intestinal function.

We found that the 13C-ODC breath test was able to non-invasively detect ODC activity in rats as a possible marker of gastrointestinal function. Although the breath test did not directly correlate to the time course of intestinal damage, the breath-signal did appear to correlate to stress events in the animals (for example the beginning of the study, when animals were first receiving injections). This interesting result may speak to further applications of the breath test, and warrant further investigation.

Additionally, the ODC breath signal at the final breath test correlated with measurements of crypt depth in the small intestine, suggesting that the breath test was reflective of changes in the proliferative compartment of the intestine. Further tissue analysis is ongoing to determine the number of proliferating cells in the small intestine. The kits for this analysis have been purchased and tissue staining is ongoing. Due to complications with PCR experiments, this component of the study was slightly delayed.

To determine small intestinal gene expression of the breath test target, ODC, we used real time-PCR. Due to several issues with yet-to-be identified contamination, this experiment delayed progress with further tissue analysis while the source of the contamination was identified, and experimental conditions optimised.

Interestingly, contrary to the published literature, the ODC inhibitor, DFMO did not effectively inhibit ODC activity. Past studies of DFMO have only been able to measure its inhibitory properties using static measurements of ODC activity in ex vivo tissue. Our new 13C-ODC breath test may have broader applications in the in vivo development of new ODC inhibitors.

This study is now in the final stages of completion and is being prepared for publication as a manuscript in an international journal (yet to be decided; likely Cancer Biology and Therapy). Additionally, the data from this study will be used to support an NHMRC funding application at the end of 2018 to investigate a panel of 13C-breath tests for the measurement of intestinal function in humans.

I would like to thank the Channel 7 Children’s Research Foundation for supporting this important study. The data generated from this project has provided a foundation for the continued development of this exciting new breath test for gastrointestinal function in children.

Research Papers:

Manuscript in preparation:

Bitzer H, Abbott CA, Jaenisch 5, Butler RN, Howarth GS, Yazbeck R, Development of the 13C-Ornithine Decarboxylase Breath Test to Detect Gastrointestinal Damage in a Rat Model of Intestinal Mucositis,

Related Publications:

‘Non-invasive test is breath of fresh air’ – The Lead South Australia, editorial March 2019

www.nature.com – Scientific Reports 9, editorial, March 2019

Future Outcomes:

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Remarks:

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