Heon Seok Kim^1,2,3,7,8, You Kyeong Jeong^1,2,8, Junho K Hur^4,5, Jin-Soo Kim^3,6,* and Sangsu Bae^1,2,*

¹ Department of Chemistry, Hanyang University, Seoul, South Korea.
² Research Institute for Natural Sciences, Hanyang University, Seoul, South Korea.
³ Center for Genome Engineering, Institute for Basic Science, Seoul, South Korea.
⁴ Department of Pathology, College of Medicine, Kyung Hee University, Seoul, South Korea.
⁵ Department of Biomedical Science, Graduate School, Kyung Hee University, Seoul, South Korea.
⁶ Department of Chemistry, Seoul National University, Seoul, South Korea.
⁷ Present address: Division of Oncology, Department of Medicine, Stanford University School of Medicine, Stanford, CA, USA.
⁸ These authors contributed equally: Heon Seok Kim, You Kyeong Jeong.

^*Correspondence to Jin-Soo Kim or Sangsu Bae.

Adenine base editors comprise an adenosine deaminase, evolved in vitro, and a Cas9 nickase. Here, we show that in addition to converting adenine to guanine, adenine base editors also convert cytosine to guanine or thymine in a narrow editing window (positions 5–7) and in a confined TC*N sequence context. Adenine base editor–induced cytosine substitutions occur independently of adenosine conversions with an efficiency of up to 11.2% and reduce the number of suitable targeting sites for high-specificity base editing.