What is Oral Cancer
Oral cancer is the development of malignant tumors in the oral cavity, including the lips, tongue, hard and soft palates. A statistical study from WHO estimates 657,000 new cases of oral cancers each year and more than 330,000 deaths each year.1 Another report from the Ministry of Health and Welfare in Taiwan shows significantly higher incidence rates of oral cancer for males than females locally.2 Furthermore, out of all the oral cancer cases, more than 90% of oral cancers are oral squamous cell carcinoma (OSCC).2
Figure 1. Oral cancer facts.
Symptoms
Common symptoms of oral cancer include:
Persistent ulcers or white and red patches in the mouth.
Apparent lumps and bumps in the neck.
Patients may feel pain and have diffculty swallowing or speaking.
Unusual bleeding or pain in the mouth.
High-risk factors
Alcohol, betel-nut and cigarette (ABC) consumptions are all considered high risk factors for oral cancer. Simultaneously consuming them can lead to an increase the chance of getting oral cancer by 123 times.3 Statistics show that ABC consumptions are responsible for 90% of oral cancer cases globally.4
Other risk factors include:
Poor oral hygiene.
Malocclusion or wearing unfit dentures.
Consumption of irritating food, such as spicy or hot food, for a long period of time.
All these risk factors may increase the chance of developing cancer.
Figure 2. The effect of alcohol drinking, betel-nut chewing and cigarette (smoking) for oral cancer patients.3
Our Inspiration
Oral Cancer in Taiwan
Oral cancer is always one of the five most common cancer in Taiwan.2 More importantly, the incidence rate for Taiwanese male to have oral cacner is amongst the highest between developed countries.5 According to WHO, the global incidence rate is about 1.5 cases for every 100,000 people.6 While in Taiwan, it’s 33.6,2 which is approximately 22 times higher than the global incidence rate. According to the Taiwanese cancer registry database, oral cancer accounts for the majority of malignant tumors for young adults,7 and this often creates a burden on the entire family as they are generally the main source of income. In addition, the average age of death for oral cancer is 56 years old, which is on average 10 years less than other cancers.8 On top of that, oral cancer often has diagnostic delay.9 An even more concerning issue is the increasing amount of cases each year. The yearly confirmed cases has doubled over the past ten years in Taiwan.10 Therefore, we can see that oral cancer is a local and serious problem in Taiwan.
Figure 3. Oral cancer in Taiwan.
The problem
The current detection method for oral cancer is mainly dependent on visual examinations. Dentist or ENT doctor can utilize visual examination and palpation to observe if there is any prodrome of oral cancer, such as white and red patch, lumps, long-term ulcers, and abnormal muscle activity. When those oral potentially malignant disorders (OPMD) lesions are observed, those patients will be referred for a biopsy to confirm its malignancy.
But, on average, there is only one OSCC case confirmed out of nine biopsies.11 Also, OPMD patients are sometimes neglected because of ambiguous lesions, which might also lead to delayed treatment. These indicate that the current detection method is biased and relies heavily on experience. In other words, the results may vary a lot according to different people conducting the process.
Our Goal
We want to make improvements to the current diagnosis procedure so that oral cancer can be identified earlier and more efficiently to decrease the mortality rate and reduce the wasted medical resources on false cases. Therefore, we think synthetic biology is a good choice to solve this problem. If we can design a clinical test base on molecular biology, then we can help doctors diagnose patients more accurately.
Aside from improving accuracy, we learned why some patients are reluctant to see a doctor and why they cannot be diagnosed early when we visited the Sunshine Social Welfare Foundation. So while designing the project, we also considered how to make our project more acceptable to the public.
To achieve those goals, we have considered different types of samples, biomarkers, detection methods, and readouts in hopes to design a non-invasive, quantitative, and accessible method for oral cancer detection.
Solution
We also visited oral medical professors in our school to learn about the biomarkers for the section of oral cancer on top of reading papers. And for sensing device, we were inspired by iGEM team 2019 EPFL, whose project was using toehold switches to detect their DNA biomarker.
After consulting with many seniors and experts, here’s finally our project: miRNA.DOC, a Detection method of Oral Cancer, using miRNA as biomarkers.
Our project
Figure 4. Simple flow chart of our project.
1. Sample: Saliva
The reason we chose saliva as our sample is that the collection of saliva is a non-invasive method compared to brush biopsy or drawing blood. And more importantly, saliva is in direct contact with cancerous tissue, which allows us to measure the progression of OSCC and OPMD in a more precise manner compared to other biological samples such as blood or urine.
2. Biomarker: microRNA
MicroRNAs (abbreviated miRNAs) are small non-coding RNA molecules and can be found in animals, plants, and some viruses. miRNAs play an important role in post-transcriptional regulation and cancer pathogenesis via suppression or activation of messenger RNA (mRNA) expression. We selectively picked miRNAs that can distinguish different stages of oral cancer and from healthy patients. This allows healthcare professionals to track the stages of OSCC and OPMD. For more details about our miRNA biomarkers, check out the proof of concept.
3. Toehold Switch
A toehold switch is a secondary structure RNA with a hairpin loop that can be used to regulate translation of downstream proteins. We use specific miRNAs in saliva to act as the key to unwind the loop structure to allow downstream protein expression. For more details on the mechanism of a toehold switch, check out the model page.
4. Glucometer
Invertase is an enzyme that catalyzes the hydrolysis of sucrose into fructose and glucose.
We designed the downstream protein of our toehold switches to incorporate invertase sequence. This way, the quantity of miRNA can be homogeneously transduced to glucose and read by a personal glucose meter (PGM), which is easy to use and an objective result.
Reference
WHO Cancer Prevention. Available online: https://www.who.int/cancer/prevention/diagnosis-screening/oral-cancer/en/ (accessed on 18 October 2020).
Cancer Registry Annual Report, 2017 (Taiwan). Available online: https://www.hpa.gov.tw/Pages/Detail.aspx?nodeid=269&pid=12235 (accessed on 18 October 2020).
Ko, Y. C., Huang, Y. L., Lee, C. H., Chen, M. J., Lin, L. M., & Tsai, C. C. (1995). Betel quid chewing, cigarette smoking and alcohol consumption related to oral cancer in Taiwan. Journal of oral pathology & medicine : official publication of the International Association of Oral Pathologists and the American Academy of Oral Pathology, 24(10), 450–453. https://doi.org/10.1111/j.1600-0714.1995.tb01132.x
Petersen, P. E., Bourgeois, D., Ogawa, H., Estupinan-Day, S., & Ndiaye, C. (2005). The global burden of oral diseases and risks to oral health. Bulletin of the World Health Organization, 83, 661-669.
Hung L-C, Kung P-T, Lung C-H, Tsai M-H, Liu S-A, Chiu L-T, Huang K-H, Tsai W-C. Assessment of the Risk of Oral Cancer Incidence in A High-Risk Population and Establishment of A Predictive Model for Oral Cancer Incidence Using A Population-Based Cohort in Taiwan. International Journal of Environmental Research and Public Health. 2020; 17(2):665.
GLOBOCAN: Estimated Cancer Incidence, Mortality and Prevalence Worldwide in 2012. Lyon: IARC, 2013. Available online: http://globocan.iarc.fr/Pages/fact_sheets_ population.aspx. (accessed on 18 October 2020).
Taiwan, Health Promotion Administration, MOHW. (2018) Follow-up referral instructions for oral mucosal health examination Available online: https://www.hpa.gov.tw/Pages/Detail.aspx?nodeid=613&pid=8621 (accessed on 18 October 2020).
Taiwan, Health Promotion Administration, MOHW. (2018). Follow-up referral instructions for oral mucosal health examination. Available online: https://www.hpa.gov.tw/Pages/Detail.aspx?nodeid=613&pid=8621 (accessed on 18 October 2020).
Kowalski, L. P. (2007). S36 Causes of diagnostic delay of oral cancer. Oral Oncology Supplement, 2(1), 123. https://doi.org/10.1016/s1744-7895(07)70256-0
Chiang, C. J., Chen, Y. C., Chen, C. J., You, S. L., Lai, M. S., & Taiwan Cancer Registry Task Force (2010). Cancer trends in Taiwan. Japanese journal of clinical oncology, 40(10), 897–904. https://doi.org/10.1093/jjco/hyq057
Taiwan, Health Promotion Administration, MOHW. (2017) Oral cancer screening quality indicators. Available online: https://www.hpa.gov.tw/Pages/Detail.aspx?nodeid=1100&pid=6461(accessed on 18 October 2020).