Introduction: The Tobacco Heating System (THS) is a “heat-not-burn” tobacco product designed to generate significantly lower levels of harmful and potentially harmful constituents (HPHCs) and present lower risk of harm than cigarettes. This study assessed the exposure reduction to selected HPHCs in smokers switching to menthol Tobacco Heating System (mTHS) 2.2 compared with smokers continuing smoking menthol cigarettes (mCCs) and smoking abstinence (SA) for 5 days in a confined setting, followed by an 86-day ambulatory period. Methods: A total of 160 healthy adult US smokers participated in this randomized, three-arm parallel group, controlled clinical study. Biomarkers of exposure to 16 HPHCs were measured in blood and 24-hour urine. Safety was monitored throughout the study. Information was also gathered on product evaluation, product use, subjective effects, and clinical risk markers (co-publication Part 2). Results: Nicotine uptake was comparable in both exposure groups (mTHS:mCC ratio of 96% on day 90). On day 5, biomarker of exposure levels to other HPHCs were reduced by 51%-96% in the mTHS group compared with the mCC group, and these reductions were sustained for most biomarkers over the ambulatory period. After 90 days of use, the level of satisfaction with mTHS and suppression of urge to smoke were comparable to mCC. Conclusion: Switching from mCCs to mTHS significantly reduced the exposure to HPHCs to levels approaching those observed in subjects who abstained from smoking for the duration of the study. Implications This study compared the impact of switching to mTHS on biomarkers of exposure, relative to continued smoking or SA. Clinical Significance: - Switching to mTHS 2.2 led to significant reductions of exposure (total NNAL, total NNN, COHb, MHBMA, 3-HPMA, S-PMA, Total 1-OHP, 4-ABP, 1-NA, 2-NA, o-tol, CEMA, HEMA, HMPMA, and B[a]P) after 5 days in confinement, which were maintained throughout the subsequent ambulatory period of 86 days. The reductions were similar to those observed upon SA. - Moderate compliance in the mTHS group during the ambulatory period was indicative of dual use of mTHS and mCC. - mTHS provides an acceptable alternative to mCC for adult smokers. Trial Registration: NCT01989156 (ClinicalTrials.gov).