Background
Due to its high binding affinity to HIV capsid (p24), the first-in-class capsid inhibitor Lenacapavir (LEN) might display a post-integration effect on capsid production. However, this effect and its underlying mechanism, as well as its clinical impact on LEN-treated individuals, have yet to be characterized.
Methods
The post-integration effect of increasing concentrations of LEN was measured in vitro (8E5, ACH2, U1, transfected 293T cells) and ex vivo (PBMCs from n=12 viremic HIV-infected participants enrolled in the APRIL study [NCT05752318]) by analysing HIV transcripts and proteins expression by flow cytometry and p24 and HIV RNA levels in supernatants.
Results
LEN induces a rapid dose-dependent decrease of intracellular p24 detection at therapeutic concentrations in HIV-1 infected cell lines (reaching top plateau within 15 minutes exposure of 50nM LEN), whereas HIV-2 infected cell line was less sensitive. The latently-infected cell line also displayed a p24 decrease on LEN independently of the activation pathway targeted. The intracellular matrix (p17), but not the envelope (Env), detection decreased with the same magnitude as p24, suggesting that LEN targets the Gag polyprotein (Pr55). These decreases further translated in lower levels of viral particle release in supernatants on LEN. Intracellular HIV Gag and Nef transcripts levels were unchanged, and maturation inhibition did not affect the LEN-induced p24 decrease, suggesting a post-transcription but pre-maturation effect. The p24 detection in oligomerization mutants (W184D+M185D, P38E+A42E+E45R) and LEN-resistant mutants (N74D and Q67H+N74D) was less affected by LEN, suggesting that LEN binding requires specific capsid oligomers and residues. Proteasome inhibition by bortezomib partially constrained the LEN-induced p24 decrease, suggesting a proteasome-mediated p24 degradation. LEN also impaired p24 detection ex vivo in productively-infected CD4+ T cells from viremic HIV-infected individuals (decreased intracellular p24 [but not envelope] detection associated with decreased viral particle release).
Conclusions
Altogether, LEN appears to be causing a rapid and specific proteasome-mediated degradation of not only the capsid but also other gag proteins in vitro and ex vivo, which echoes in virions production. Reduced Gag proteins expression by infected cells in LEN-treated individuals could impact recognition of these cells by the immune system and thus the dynamics of HIV reservoirs and should be further studied.
Table or Figure
