Introduction: Heating rather than burning tobacco reduces levels of harmful and potentially harmful constituents, and consumer products using this approach aim to reduce exposure to tobacco toxicants. The Tobacco Heating System (THS) version 2.1 has been enhanced from earlier prototypes with an improved heat control and sensorial experience and thereby user acceptance. Exposure measurements are required to determine whether it may be possible to reduce the individual health risk compared to smoking combustible cigarettes (CCs). Methods: This controlled clinical study randomly assigned 40 smokers to either a group continuing to use of their own CC brand (n = 20) or a group switching to THS 2.1 (n = 20) for five days. Biomarkers of exposure were measured at baseline and on Days 1 through Day 5. Product consumption, Human Puffing Topography (HPT), the occurrence of adverse events (AEs), and an assessment of subjective effects, such as smoking satisfaction and enjoyment of respiratory tract sensations, were also determined. Results: The group of smokers who switched to THS 2.1 adapted their puffing behavior initially through longer puff duration and more puffs. During the duration of the study total puff volume returned to baseline levels and the mean daily product consumption increased but with similar nicotine exposure compared to baseline CC use. Biomarkers of exposure to tobacco smoke toxicants which inform product risk assessment were significantly reduced with THS use compared to the CC group. THS 2.1 users experienced less reinforcing effects with THS 2.1 than with their own cigarette brand. Conclusions: THS 2.1 is a promising alternative to smoking CCs. Notwithstanding possible use adaption through consumption or puffing behavior, the exposure to harmful smoke constituents was markedly reduced with the new heated tobacco platform. Implications: Exposure markers to harmful and potentially harmful smoke constituents were lowered with the tobacco heating system (THS 2.1). Heating tobacco instead of burning can offer a potentially lower risk of delivering nicotine compared to CCs.