E-Cigarettes and 21st Century Toxicology

Despite the overwhelming evidence that cigarette smoke can cause a number of serious diseases including cardiovascular disease, lung cancer and chronic obstructive pulmonary disease, there remains a significant number of people around the world who chose to continue smoking.

See: smoking a pack a day causes 150 mutations in lung cells

While preventing smoking initiation and promoting smoking cessation remain the cornerstone of efforts to reduce the harm associated with cigarette smoking, the development of less harmful sources of nicotine, including e-cigarettes and other Reduced Risk Products* (RRPs), may also have a role to play in tobacco harm reduction. In fact, in the UK alone, evidence suggest that switching even a small fraction of smokers every year to less harmful nicotine sources could potentially save tens of thousands of lives over a ten-year period.¹

You might be interested in: Public Health England study investigating vaping

There is an ongoing debate around the exact role that e-cigarettes can play in tobacco harm reduction, as well as the means necessary to test their risk-reduction potential. Some of these issues were recently addressed at an expert round table discussion between Professor Thomas Hartung of the Johns Hopkins University Bloomberg School of Public Health, leading tobacco industry representatives, and senior figures from the academic world. The roundtable has been summarized in a special issue of Applied In Vitro Toxicology and was discussed further during the recent Society of Toxicology Annual Meeting in Baltimore, MD, USA, in an ancillary session hosted by the Institute of In Vitro Sciences.^2-4

While much of the available evidence suggests e-cigarette use is associated with considerably reduced toxicity as compared to cigarette smoking, some studies have reported that the aerosols produced by e-cigarettes do contain measurable amounts of certain toxicants. As such, their potential health effects should be thoroughly investigated.

See also: carcinogens found in eCigarette vapors; and cancer-causing benzene found in eCigarette vapor

The toxicological assessment of e-cigarettes is a challenging task. There are many different types of e-cigarettes, there is a limited number of studies evaluating the biological effects of their main constituents on the respiratory system, and, most importantly, there is a lack of standardized testing protocols. Recently, we described a layered framework for the systematic assessment of e-cigarettes using relevant in vitro systems and contemporary methodologies.⁵ This strategy is aligned with the ‘3Rs principles’ of animal testing (replacement, reduction and refinement) and the framework for ‘21st Century Toxicology’ as set-out in the US National Research Council’s landmark vision and strategy report⁶. Where in the past in vivo studies would be used to assess toxicity, increasingly newer in vitro techniques involving human cellular models can be used in their place. As well as providing the potential to replace animal testing, these newer techniques can also offer more timely results, cost-efficiencies and a deeper understanding of the biological mechanisms involved in toxicity.

New Approaches to E-Cigarette Toxicity Testing

E-cigarettes are battery-powered devices that heat a solution (e-liquid) to generate an aerosol which is then inhaled by the user. Typically, the most prominent constituents of e-liquids are propylene glycol (PG) and vegetable glycerin (VG), in differing proportions. In addition, they may or may not contain nicotine, an addictive substance but not one that is in itself particularly toxic (it is not the nicotine in cigarette smoke that causes smoking-related disease, but rather a range of other harmful constituents that result from the burning of tobacco).

PG and VG are used extensively in the food, pharmaceutical and cosmetic industries. They have low systemic toxicity and are generally considered safe when ingested or applied topically. However, their effects on the respiratory system are less well-known and the available data is limited. As such, there remains a need to assess their toxicity in the context of e-cigarettes.

To address this need, we recently set out to investigate the biological effects of these e-liquid ingredients on primary human lung epithelial cells.⁷ Our aim was twofold: 1) to determine the contribution of PG, VG and nicotine (in various proportions) to the overall biological impact of e-liquids, and 2) to compare this biological impact with that of nicotine alone (the active principle in standard nicotine replacement therapies (NRTs) such as patches, gums and sprays). This comprehensive in vitro study involved real-time cellular analysis and high content screening, both high-throughput techniques capable of measuring multiple toxicity endpoints in a robust and reproducible manner. The study was complemented by a whole transcriptome analysis whereby changes in gene expression were interpreted using a series of network models representing the relevant human biology. This approach allowed us to identify and quantify the biological impact of PG, VG and nicotine exposure, thus gaining mechanistic insight into their toxicological effects. The results of these assessments showed that the biological effects of nicotine-containing PG and VG mixtures were similar to those of nicotine alone. The data suggests that, in the absence of flavoring agents and other additives, the toxicity of e-liquids would be similar to that of NRT.

This study was an initial and crucial step in the broader, layered assessment framework for the systematic assessment of e-cigarettes using in vitro systems and 21st century methodologies.⁵ We are also now conducting in vitro studies on the effects of the aerosols produced by e-cigarettes (generated through the heating of e-liquids) on human cells and intend to report on these studies in due course.

Conclusion

The study briefly outlined above was just one that has been published in the recent special issue of Applied In Vitro Toxicology.³ Other papers address different scientific aspects of in vitro strategies for the evaluation of RRPs, as well considerations for the regulatory acceptance of in vitro approaches and the need for cooperation across a broad stakeholder group in order to optimize and standardize testing methodologies.

We contributed to two further articles in the special issue, looking at ‘Adverse Outcome Pathways’ for hypertension and decreased lung function.^8-9 Adverse Outcome Pathways are theoretical frameworks that describe sequences of events and mechanisms by which substances, such as chemicals, can cause specific adverse human health effects. Again, these frameworks can be used by scientists and regulators to predict adverse effects without the need for animal tests.

Read more here: tobacco companies win PETA award for 3R's work

21st Century Toxicology provides a wealth of new technology that can enable faster, more accurate and more physiologically relevant consumer product safety assessment. While many challenges remain in refining, optimizing and standardizing these technologies, they nevertheless provide substantial opportunity for improved strategies for toxicity testing. In the context of e-cigarettes and other RRPs, they are being increasingly recognized as a crucial set of tools to help realize the potential of these products for tobacco harm reduction. 

Notes

* Reduced-Risk Products (“RRPs”) is the term we use to refer to products that present, are likely to present, or have the potential to present less risk of harm to smokers who switch to these products versus continued smoking. We have a range of RRPs in various stages of development, scientific assessment and commercialization.  Because our RRPs do not burn tobacco, they produce far lower quantities of harmful and potentially harmful compounds than found in cigarette smoke.

Read: carbon-heated tobacco reduces toxicity

Authors

Ignacio Gonzalez-Suarez is Senior Scientist, Biological Systems Research, Philip Morris International.

Julia Hoeng is Director of Systems Toxicology, Biological Systems Research, Philip Morris International.

References

1. Fagerström, KO and Bridgman, K. 2014. Tobacco harm reduction: the need for new products that can compete with cigarettes. Addictive Behaviors, 39(3): 507-11. DOI: http://dx.doi.org/10.1016/j.addbeh.2013.11.002 

2. Fowle, JR. et al. 2017. Twenty-first century in vitro toxicology testing methods and the assessment of e-cigarettes. Applied In Vitro Toxicology, 3(1): 3-9. DOI: http://dx.doi.org/10.1089/aivt.2017.29011.rtl

3. Gaca, MD (ed.) 2017. Special Issue: Application of in vitro toxicology approaches for the evaluation of next-generation nicotine products. Applied In Vitro Toxicology, 3(1).

4. Institute of In Vitro Sciences. 2017. Ancillary Session: Opportunities for human in vitro respiratory models in regulatory toxicology. Society of Toxicology, Baltimore, MD, USA, 12-16 March

5. Iskandar, A. et al. 2016. A framework for in vitro systems toxicology assessment of e-liquids. Toxicology Mechanisms and Methods, 26: 389-413

6. National Research Council. 2007. Toxicity testing in the 21st century: a vision and a strategy. National Academy Press, Washington, DC (2007)

7. Gonzalez-Suarez, I. et al. 2017. In vitro systems toxicology assessment of nonflavored e-cigarette liquids in primary lung epithelial cells. Applied In Vitro Toxicology, 3(1): 41-55. DOI: http://dx.doi.org/10.1089/aivt.2016.0040

8. Lowe, FJ. et al. 2017. Development of an adverse outcome pathway for the onset of hypertension by oxidative stress-mediated perturbation of endothelial nitric oxide bioavailability. Applied In Vitro Toxicology, 3(1): 131-148. DOI: http://dx.doi.org/10.1089/aivt.2016.0031

9. Luettich, K. et al. 2017. The adverse outcome pathway for oxidative stress-mediated EGFR activation leading to decreased lung function. Applied In Vitro Toxicology, 3(1): 99-109. DOI: http://dx.doi.org/10.1089/aivt.2016.0032