The role of biomarkers in PMI’s research

What are biomarkers?

A biomarker is a measurable substance whose presence indicates an interaction between a potential hazard, such as a harmful chemical, and a biological system, like the human body. Biomarkers can be molecules that are found in the blood, in other bodily fluids or tissues, or they can also be measurements indicating how well the body functions.

Biomarkers are important in clinical research because, while it is relatively easy to measure the level of toxicants in aerosol or cigarette smoke in the lab, these measurements are a lot more difficult to make once the chemicals are in the body. There are many reasons for this, including the fact that chemicals break down or are converted to other products (metabolites) in the body, and that the amount of a substance in a product, or extracted during use, is not necessarily the same as the amount absorbed by the body. Biomarkers allow researchers to assess human exposure to toxicants in cigarettes and smoke-free products, and the resulting disease-related biological effects.

Some toxicants have more than one biomarker. For example, carbon monoxide (CO) is a product of incomplete combustion of organic matter, such as tobacco. Exhaled CO is used as a biomarker of acute or recent exposure to combustion. However, once in the body, CO binds rapidly to hemoglobin in the blood to form carboxyhemoglobin (COHb). COHb reduces the ability of the blood cells to carry oxygen, and so high levels of COHb in the blood are used to signal an impairment in oxygen transport due to CO exposure and an increased risk for cardiovascular or pulmonary disease.  

There are a number of different ways to categorize biomarkers, but two of the most important are biomarkers of exposure (BoE) and biomarkers of potential harm (BoPH). Let’s look at these two categories of biomarkers in more detail.

What are biomarkers of exposure?

Biomarkers of exposure are used to measure the levels of particular substances, such as potentially harmful chemicals in cigarette smoke or smoke-free product aerosol, which a user has been exposed to. 

In research on cigarette and smoke-free product use, BoE are used to confirm exposure to specific nicotine or tobacco products, and to indicate changes in levels of exposure to specific chemical compounds when users switch between products—such as switching from cigarette smoking to THS use. To be useful, a BoE must be related to a specific chemical or exposure and must be capable of being reliably measured.

Nicotine, for example, rapidly breaks down into cotinine and several other metabolites once inside the body. So, in order to determine nicotine levels precisely, the presence of cotinine and several other metabolites of nicotine are assessed to give a measurement of nicotine equivalents (NEQ) or total nicotine equivalents (TNE).

In addition to nicotine, biomarkers signaling exposure to tobacco-specific nitrosamines (TSNAs) are used in many of PMI’s clinical studies. TSNAs are a group of carcinogens formed from nicotine and related tobacco alkaloids. Important TSNA biomarkers include 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK), 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanol (NNAL), and N-nitrosonornicotine (NNN). Other important BoE include those for chemicals formed during the burning of cigarettes, such as benzo[a]pyrene and 1,3-butadiene.

What are biomarkers of potential harm?

Biomarkers that are used to measure an effect, such as those due to exposure to a chemical or substance are referred to as biomarkers of potential harm (BoPH). The effect measured may be changes in physiological structure or function, or clinical symptoms that can signal an increased risk of disease. 

For example, changes in white blood cell count, high-density lipoprotein cholesterol levels (i.e., “good” cholesterol), or how much air a person can forcefully exhale in one second (FEV₁) can provide an indication of a person’s health status or potential risk of disease.

Because chronic diseases associated with smoking, such as cancer, chronic obstructive pulmonary disease (COPD), and cardiovascular disease (CVD) take a long time to develop, BoPH are also used by regulatory bodies as surrogate indicators for assessing health risks when new products are introduced onto the market.

It is also important to keep in mind that a single biomarker can be both a BoE and a BoPH. For example, NNAL can signal both exposure to tobacco smoke and an increased risk of cancer.

 

How does PMI choose which biomarkers to study?

When planning a clinical study, researchers will often choose to measure a set of biomarkers which are generally representative of different chemical or toxicological groups, such as carcinogens, cardiovascular, or respiratory toxicants. Additional criteria for choosing BoE for study may include:

• Biomarkers for substances that are specific to cigarette smoke and sensitive to smoking cessation.
• Biomarkers are measurable using validated, reliable, reproducible, and precise analytical methods.
• Biomarkers are clinically relevant.
• Biomarkers for substances that reflect a specific toxic exposure or are a reliable surrogate of exposure to those substances.
• Feasibility of observing an effect based on the study duration.

Some criteria for choosing BoPH for study may include:

• Biomarkers that signal the risk of developing a smoking-related disease, such as cardiovascular disease or emphysema.
• Biomarkers that signal the onset or presence of a specific smoking-related disease.
• Biomarkers linked to multiple aspects of pathogenesis associated with smoking-related diseases.
• Biomarkers for chemicals from a broad variety of organ toxicity classes, such as carcinogens, cardiovascular toxicants, respiratory toxicants, and substances that can interfere with fetal development (teratogens). 

Researchers also often select biomarkers for study that are linked to chemicals recommended for lowering by regulatory organizations. For example, the U.S. Food and Drug Administration (FDA) has a list of 20 harmful and potentially harmful constituents (HPHCs) for which testing methods are well established and for which they require reporting. In addition, a group of nine toxicants has been proposed by the World Health Organization (WHO) Study Group on Tobacco Product Regulation (TobReg) for mandated lowering in cigarette smoke.

 

Findings from PMI’s clinical biomarker studies

PMI’s clinical biomarker studies are designed to address questions related to the potential for harm reduction for those switching from cigarette smoking to THS use. These include whether the magnitude of biological changes seen in those who switch from cigarettes to THS is relevant and continues over time, and whether the reduction in exposure to harmful chemicals found in THS users is similar to that observed in those who quit smoking completely.

Reduced exposure to harmful chemicals with THS versus cigarettes

PMI conducted a series of studies examining levels of biomarkers of exposure in existing adult smokers who switched to THS use, compared with continuing smoking and quitting smoking altogether. These three-arm studies compared groups of healthy participants who had not used THS before and switched to THS for the study, continued smoking their normal brand of cigarettes, or quit altogether for the duration of the study. 

Together, these studies demonstrated that switching completely to THS resulted in significantly lower levels of 15 biomarkers representing exposure to harmful chemicals compared with continued smoking. The reductions among those who switched to THS were in the same direction observed among those who quit for the duration of the studies.

Lower levels of BoPH seen 6 months and 12 months after switching to THS

PMI researchers conducted a 6-month exposure response study, followed by a 6-month extension, to understand if the switching from cigarettes to predominately THS use is associated with favorable changes in eight BoPH. The studies assigned healthy adults who had been smoking for at least 10 years to either continue smoking their own brand of cigarettes or to switch to predominately THS use for the duration of the study.       

After 6 months, all eight BoPH showed favorable changes in the THS users in the same direction as seen in those who quit smoking altogether, with five out of the eight showing statistically significant improvements compared with continued smoking. For participants who predominantly used THS, the favorable biological responses observed at 6 months were overall maintained at 12 months. The extension study also found that the fewer cigarettes smoked per day in addition to THS, the greater the magnitude of beneficial effects observed among healthy participants.

Cross-sectional study confirms earlier findings

Our most recently completed clinical study, which has not yet been published, demonstrated favorable improvements in nine BoPH among healthy participants who switched completely from cigarettes to THS for at least 2 years (on average 4.5 years), compared with those who continued to smoke. The magnitude of changes seen were larger than those observed in the exposure response study of recently started THS users, where participants could use cigarettes along with THS. This sustained effect among former smokers who had switched to THS demonstrates the importance of fully replacing cigarettes with THS to achieve maximum harm reduction potential.

 

Conclusion

The use of biomarkers makes it possible for researchers to measure exposure to harmful chemicals and the effects of that exposure in real life. This makes them an important tool for comparing the effects of exposure to THS aerosol and cigarette smoke.

Overall, the data from PMI’s biomarker studies provide evidence that users of THS have lower exposure to HPHCs and reduced signs of negative health impacts related to smoking, including emphysema, cancer, COPD, and CVD, compared with cigarette smokers. 

Importantly, the data indicates that the fewer cigarettes smoked, the higher the beneficial impact, with the biggest improvements seen in those who switch completely to THS. 

For existing smokers, the best way to maximize the reduction of risk is to quit tobacco and nicotine altogether, but for those adults who don’t quit altogether, smoke-free products can be a better choice than continued smoking.