Due to the development of medical technology and systems, the premature mortality rate due to disease has decreased significantly compared to the past. However, lethality from some incurable diseases including cancer is still high. Because it is diffi...
Due to the development of medical technology and systems, the premature mortality rate due to disease has decreased significantly compared to the past. However, lethality from some incurable diseases including cancer is still high. Because it is difficult to feel conscious symptoms before the disease develops to a late stage, and the existing diagnosis method is inaccessible due to the invasive method and cost of examination. Due to this reason, the latest disease diagnosis technology is developing in the direction of improving accessibility, and in particular, the need for non-invasive and economic method is emerging. As a typical example, the technology for diagnosing a disease by detecting a specific volatile organic compounds enables simple diagnosis without pain because it can detect the signal of disease from exhaled breath, sweat, urine, and saliva as well as blood and body fluids. In particular, the bioelectronic sensor has demonstrated excellent selectivity and sensitivity by combining a primary transducer such as an olfactory receptor with a secondary transducer containing a nanostructured semiconductor such as carbon nanotubes or graphene.
The purposes of this research are identification of disease biomarkers and screening, performance evaluation of olfactory receptors for the detection of biomarkers that are essential for development of bioeletronic sensor. The selected diseases for study are lung cancer, tuberculosis, and gastric cancer. First, the discovery of biomarkers for lung cancer and the screening of human olfactory receptors were performed. The lung cancer cell line and the normal lung cell line were cultured to compare the composition of headspace gas by GC / MS, and volatile organic compound 2-ethyl-1-hexanol, which is more frequently generated in lung cancer cell lines, was identified. In addition, human olfactory receptors capable of detecting this biomarker were screened using a dual-glo luciferase reporter gene assay. It was confirmed that the identified olfactory receptor sensitively and selectively detects the lung cancer biomarker, and then conducted olfactory nanovesicle generation and performance evaluation for use as a primary transducer of the bioelectronic sensor in the further study.
In the second study, the screening of human olfactory receptors were carried out for identification of olfactory receptor capable of detecting 5 tuberculosis biomarkers found in urine [95]. The screening was conducted by transfectng the human olfactory receptor genes and the luciferase reporter gene into the HEK293 cell line to confirm the responsivity to the tuberculosis biomarkers. As a result, olfactory receptors recognizing each tuberculosis biomarker were selected, and their responsivity and selectivity were also analyzed.
Third, a number of exhaled breath samples of gastric cancer patients and healthy subjects were collected and analyzed using GC/MS. As a result, butyl acid and propionic acid, which are volatile organic compounds found in relatively large amounts in the exhaled breath of gastric cancer patients, were identified. In particular, solid-phase microextraction (SPME) fibers were used as a instruments of collecting and concentrating volatile organic compounds to completely analyze the biomarkers containing a very small amount in the exhaled breath samples. To improve the reliability of the selected volatile organic compounds as biomarkers, we build a diagnostic model that distinguishes patients based on the amount of biomarkers in the exhaled breath through statistical analysis of overall data, and their sensitivity and selectivity were calculated. In addition, in order to identify a primary transducer of a bioelectronic sensor that detects biomarkers included in exhaled breath, the responsivity and selectivity of 2 human olfactory receptors known to detect butyric acid and propionic acid were estimated.
Development of disease diagnosis technology is an inevitable process for universal welfare and extension of life expectancy. Diagnostic methods targeting disease-specific volatile organic compounds are attracting attention in academia as a next-generation diagnostic technology, and are actively being studied all over the world. In this thesis, several disease-specific volatile organic compounds have been newly identified, and the human olfactory receptors capable of recognizing disease biomarkers were screened. The above research results are expected to be useful for the development of sensitive and selective bioelectronic sensor for disease diagnosis.