Development of Improved HIV-1 Capsid Inhibitors

ABSTRACT
The HIV-1 capsid protein (CA) is an attractive target for the development of novel antiretrovirals due to its
essential and multifaceted roles in the virus biology. Lenacapavir (LEN) is a recently discovered first-in-class,
long-acting capsid-targeting inhibitor with picomolar potency against HIV-1. Phase 2/3 clinical trials have
revealed that subcutaneous administration of LEN with a six-month dosing interval enables high rates of HIV-1
suppression in heavily treatment-experienced patients infected with multi-drug resistant viral phenotypes. These
findings have highlighted LEN as a promising agent that could complement current antiretroviral compounds to
treat people living with HIV-1. However, cell culture-based viral breakthrough assays and clinical trials have
identified a number of CA substitutions that confer substantial resistance to LEN. Specifically, Q67H/N74D and
M66I substitutions emerged as major drug-resistance associated variants. Therefore, there is a need to develop
improved LEN analogs with a higher barrier to resistance. We have established synthesis of LEN in academic
setting by utilizing a modular approach of independently preparing three unique LEN subunits (A, B, and C) from
commercially available reagents. Such modular approach allows for straightforward modifications of each
subunit which can then be combined in any order to prepare LEN and its analogs. Thus, our medicinal chemistry
strategy is highly advantageous for analog development. Our complementary virology, biochemistry and
structural biology experiments have allowed us to characterize a multi-modal mechanism of action of LEN.
Furthermore, we have recently determined high-resolution X-ray crystal structures of CA hexamers containing
major drug-resistance associated Q67H/N74D and M66I changes. We propose to exploit these findings to
rationally develop improved LEN analogs with a higher barrier to resistance. In Aim 1, our efforts will focus on
modifications to LEN subunits A and C to overcome steric hindrance and electrostatic repulsions created by the
drug resistant Q67H/N74D CA variant. In aim 2, we will use our recent promising findings from MiniFrags
screening studies, which identified fragments that bind to the CA hydrophobic pocket in close vicinity to LEN.
We will synthesize new chemotypes by connecting these fragments to LEN to generate second-generation
analogs to target the major drug-resistant M66I variant. Newly synthesized compounds from both Aims 1 and 2
will be evaluated by a stepwise approach using antiviral activity, cytotoxicity, surface plasmon resonance and X-
ray crystallography to identify and characterize the lead compound(s). Taken together, the proposed research is
expected to generate novel chemotypes with improved antiviral activities against major drug resistant CA
variants that confer substantial resistance to parental LEN.

Grant Number: 5R21AI174866-02
NIH Institute/Center: NIH
Principal Investigator: Daniel Adu-Ampratwum