Breakthrough Design of Highly Potent Antivirals Against Wild‐Type HIV‐1 and Lenacapavir‐Resistant M66I Variant

ABSTRACT Lenacapavir (LEN), the first‐in‐class HIV‐1 capsid inhibitor (CAI) with picomolar potency and long pharmacokinetics (PK), is approved for both treatment and pre‐exposure prophylaxis (PrEP). However, the frequently selected M66I mutation confers complete resistance to LEN. Potent antivirals against the M66I variant remain elusive. In this work, we have designed LEN analogs featuring a distinctive R 2 moiety to directly address the molecular mechanism of resistance. Our analog synthesis also entailed developing a new synthetic procedure for R 2 incorporation. Significantly, the newly designed analogs were highly potent against both WT HIV‐1 and the M66I mutant. Particularly, lead compound 3S conferred 2.6‐fold higher potency than LEN against WT HIV‐1 and remained highly potent (EC 50 = 5.8 nM) against the M66I mutant, a drastically advanced activity profile over LEN (EC 50 > 15 µM). The unprecedented potency of our lead 3S against M66I was corroborated by a thermal shift assay. In a head‑to‑head intravenous (IV) PK study in rats, lead 3S demonstrated superior systemic exposure, showing a 4.6‑fold greater plasma area‐under‐curve (AUC) than LEN and largely retaining the long PK characteristic of LEN. These results strongly validate our design and represent a breakthrough in LEN‐based HIV‐1 therapy and prophylaxis.