The Future of Saliva Biomarkers in the Clinic

Compared to other biofluids, there are numerous benefits to using saliva for diagnostic testing. Saliva collection is non-invasive, biomarker testing is low cost, and diagnostics can be as sensitive as tests that rely on traditionally-favored biofluids, such as blood. With saliva testing for SARS-CoV-2 growing in popularity and serving as a definitive proof point, salivary diagnostics are uniquely positioned to continue their ingress into the clinic.¹

But what exactly does the future of salivary testing look like? With so many advantages to using saliva for diagnostic testing, it has yet to become a mainstay of clinical practice. The next decade will certainly see new salivary biomarkers being discovered and validated, and testing programs continuing to grow. Although, there are still many hurdles that need to be overcome to actualize the full potential of salivary testing. Let’s look at some of the barriers that are standing in the way of more widespread analysis of saliva and translation into the clinic.

Salivaomics is the use of saliva for genomics, epigenomics transcriptomics, proteomics, metabolomics, and microbiomics

As with so many other biofluids, biomarker discovery efforts have incorporated “omics” and combinations of omics (i.e., multi-omics) techniques for biomarker identification and validation. Saliva is no exception, and “salivaomics ” has become a common term to describe the application of omics techniques – including genomics, epigenomics transcriptomics, proteomics, metabolomics, and microbiomics – to the analysis of saliva samples.²

Transcriptomics studies have identified mRNA panels that can distinguish between healthy patients and those with oral squamous cell carcinoma, pancreatic cancer, breast cancer, and others.^3-5 Proteomics techniques have also been applied to saliva samples and many promising cancer-associated biomarkers show clinical potential.⁶ Additional proteomics studies and cataloging of salivary proteomics data, as with the Saliva Proteome Knowledge Base, have helped to expand the applications of proteomic diagnostics with saliva.⁷ Omics studies that use saliva as a sample type are ongoing and a powerful way to identify single or panels of biomarkers that can sensitively and selectively predict various aspects of human disease. In addition, omics techniques can teach researchers much about the basic biology of saliva, which is important for advancing its use as a diagnostic tool.

Rigorous validation of salivary biomarkers

With salivaomics churning out an incredible amount of data on saliva biomarkers, it is important to remember the urgency for rigorous testing, prior to clinical use. Precision, accuracy, and sensitivity are essential when making a decision about a patient’s prognosis or diagnosis. It is tempting to let the many advantages of salivary testing quicken the path to market, but as with the saliva biomarkers that have been discovered through basic research into disease pathophysiology, and salivary bioscience, there is still a desperate need for additional verification and validation in large clinical trials. This is an essential step before salivary testing can enter widespread clinical use for oral and/or systemic diseases.

Ramping up saliva research

In addition to increasing the number of potential clinical biomarkers, omics techniques are enhancing the amount of knowledge about the biomolecules that make up saliva and their physiological function. In this area, saliva is severely lacking compared to other biofluids, such as blood or urine. For this reason, many clinicians may be reluctant to adopt the use of saliva for clinical testing.

For instance, salivary chemicals and biomolecules can vary depending on gender or age, ethnicity, geographic location, circadian clock, diet, and many other factors.^8-11 Extensive research into how these can affect one or many salivary biomarkers and the mechanism by which this occurs will be critical prior to pursuing clinical validation and trial of salivary diagnostics.

Standardizing collection and storage for downstream enablement

Conducting more basic research is one solution to understanding the variation in saliva composition. Another is the development of guidelines and standardization. As with any clinical diagnostic, creating a standardized protocol under which all testing occurs is essential for accurate results.

So far, salivary testing has not had such standards put into place, making comparisons across studies and consensus about the utility of specific biomarkers difficult.

Controllable factors, such as the time of day when saliva is collected or diet can affect the concentration of specific biomarkers and are not typically controlled for during biomarker studies, that in turn creats variation. The type of salivary fluid (i.e., whole saliva, unstimulated whole saliva, stimulated whole saliva, etc.) collection and collection device (i.e., use of an absorbent pad or collection tube) used can also affect the analyte levels present, and therefore, the result of a diagnostic test.^12,13 In addition, analytes have different stabilities at different temperatures. How samples are stored, at what temperature, and the number of freeze-thaw cycles they go through are other factors that need to be standardized in order to minimize analyte degradation.¹⁴

Balancing Sensitivity with Affordability

A major challenge to developing accurate salivary diagnostics is developing detection methods that can sensitively and accurately identify low-abundance biomarkers in saliva. To capitalize on the advantage of a non-invasive, easy-to-collect sample type, such as saliva, diagnostic systems have to be small, at the point of care, and still be easy for patients to comply with.

Balancing all of these factors can increase costs, as a diagnostic device may require the use of microfluidics or chips for detection. While that is not always the case, as with the recent report of a paper-capture, CRISPR-based, point-of-care SARS-CoV-2 test using saliva samples, this is a major limitation. Technologies with adequate sensitivity and low-cost are essential to broaden the implementation of salivary testing.¹⁵

SpeciMax Dx Saliva Collection kits are driving the expansion of saliva sampling forward

Salivary diagnostics have an incredible amount of potential for medicine and healthcare. Thermo Fisher Scientific is transforming this potential into reality by solving some of the hurdles that face salivary testing.

The SpeciMax Dx saliva collection kits are one such solution. They are automation-ready, making the development and implementation of standardized testing protocols simple, straightforward, and efficient. The SpeciMax Dx kits also make the advantages of saliva collection real, with an easy-to-use design that is preferred by most users. For saliva testing programs, our collection kits are designed to help you save on refrigeration and freezer space and optimized for sample accessioning.

This article contains product information intended for General Laboratory Use. It is the customer’s responsibility to ensure that the performance of the product is suitable for customer’s specific use or application.

References:

Butler-Laporte G, Lawandi A, Schiller I, et al. Comparison of Saliva and Nasopharyngeal Swab Nucleic Acid Amplification Testing for Detection of SARS-CoV-2: A Systematic Review and Meta-analysis [published correction appears in JAMA Intern Med. 2021 Mar 1;181(3):409]. JAMA Intern Med. 2021;181(3):353-360. doi:10.1001/jamainternmed.2020.8876
Shah S. Salivaomics: The current scenario. J Oral Maxillofac Pathol. 2018;22(3):375-381.
Li Y, St John MA, Zhou X, et al. Salivary transcriptome diagnostics for oral cancer detection. Clin Cancer Res. 2004;10(24):8442-8450.
Zhang L, Farrell JJ, Zhou H, et al. Salivary transcriptomic biomarkers for detection of resectable pancreatic cancer. Gastroenterology. 2010;138(3):949-57.e577.
Zhang L, Xiao H, Karlan S, et al. Discovery and preclinical validation of salivary transcriptomic and proteomic biomarkers for the non-invasive detection of breast cancer. PLoS One. 2010;5(12):e15573.
Cheng J, Nonaka T, Ye Q, et al. Chapter 8: Salivaomics, Saliva-Exosomics, and Saliva Liquid Biopsy. In: Grander DA, Taylor MK, eds. Salivary Bioscience. 1st ed. Springer Nature Switzerland; 2020:449-467.
Ai J, Smith B, Wong DT. Saliva Ontology: an ontology-based framework for a Salivaomics Knowledge Base. BMC Bioinformatics. 2010;11:302.
Prodan A, Brand HS, Ligtenberg AJ, et al. Interindividual variation, correlations, and sex-related differences in the salivary biochemistry of young healthy adults. Eur J Oral Sci. 2015;123(3):149-157.
Hajat A, Diez-Roux A, Franklin TG, et al. Socioeconomic and race/ethnic differences in daily salivary cortisol profiles: the multi-ethnic study of atherosclerosis. Psychoneuroendocrinology. 2010;35(6):932-943.
Li J, Quinque D, Horz HP, et al. Comparative analysis of the human saliva microbiome from different climate zones: Alaska, Germany, and Africa. BMC Microbiol. 2014;14:316.
Wada M, Orihara K, Kamagata M, et al. Circadian clock-dependent increase in salivary IgA secretion modulated by sympathetic receptor activation in mice. Sci Rep. 2017;7(1):8802.
Khurshid Z, Zohaib S, Najeeb S, Zafar MS, Slowey PD, Almas K. Human Saliva Collection Devices for Proteomics: An Update. Int J Mol Sci. 2016;17(6):846.
Topkas E, Keith P, Dimeski G, Cooper-White J, Punyadeera C. Evaluation of saliva collection devices for the analysis of proteins. Clin Chim Acta. 2012;413(13-14):1066-1070.
Barranco T, Rubio CP, Tvarijonaviciute A, et al. Changes of salivary biomarkers under different storage conditions: effects of temperature and length of storage. Biochem Med (Zagreb). 2019;29(1):010706.
de Puig H, Lee RA, Najjar D, et al. Minimally instrumented SHERLOCK (miSHERLOCK) for CRISPR-based point-of-care diagnosis of SARS-CoV-2 and emerging variants. Sci Adv. 2021;7(32):eabh2944.