Cholesterol reduction by immunization with a PCSK9 mimic.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a plasma protein that controls cholesterol homeostasis. Here, we design a human PCSK9 mimic, named HIT01, with no consecutive 9-residue stretch in common with any human protein as a potential heart attack vaccine. Murine immunizations with HIT01 reduce low-density lipoprotein (LDL) and cholesterol levels by 40% and 30%, respectively. Immunization of cynomolgus macaques with HIT01-K21Q-R218E, a cleavage-resistant variant, elicits high-titer PCSK9-directed antibody responses and significantly reduces serum levels of cholesterol 2 weeks after each immunization. However, HIT01-K21Q-R218E immunizations also increase serum PCSK9 levels by up to 5-fold, likely due to PCSK9-binding antibodies altering the half-life of PCSK9. While vaccination with a PCSK9 mimic can induce antibodies that block interactions of PCSK9 with the LDL receptor, PCSK9-binding antibodies appear to alter homeostatic levels of PCSK9, thereby confounding its vaccine impact. Our results nevertheless suggest a mechanism for increasing the half-life of soluble regulatory factors by vaccination.

Long trimer-immunization interval and appropriate adjuvant reduce immune responses to the soluble HIV-1-envelope trimer base.

Soluble 'SOSIP'-stabilized HIV-1 envelope glycoprotein (Env) trimers elicit dominant antibody responses targeting their glycan-free base regions, potentially diminishing neutralizing responses. Previously, using a nonhuman primate model, we demonstrated that priming with fusion peptide (FP)-carrier conjugate immunogens followed by boosting with Env trimers reduced the anti-base response. Further, we demonstrated that longer immunization intervals further reduced anti-base responses and increased neutralization breadth. Here, we demonstrate that long trimer-boosting intervals, but not long FP immunization intervals, reduce the anti-base response. Additionally, we identify that FP priming before trimer immunization enhances antibody avidity to the Env trimer. We also establish that adjuvants Matrix M and Adjuplex further reduce anti-base responses and increase neutralizing titers. FP priming, long trimer-immunization interval, and an appropriate adjuvant can thus reduce anti-base antibody responses and improve Env-directed vaccine outcomes.

Cleavage-intermediate Lassa virus trimer elicits neutralizing responses, identifies neutralizing nanobodies, and reveals an apex-situated site-of-vulnerability.

Lassa virus (LASV) infection is expanding outside its traditionally endemic areas in West Africa, posing a pandemic biothreat. LASV-neutralizing antibodies, moreover, have proven difficult to elicit. To gain insight into LASV neutralization, here we develop a prefusion-stabilized LASV glycoprotein trimer (GPC), pan it against phage libraries comprising single-domain antibodies (nanobodies) from shark and camel, and identify one, D5, which neutralizes LASV. Cryo-EM analyses reveal D5 to recognize a cleavage-dependent site-of-vulnerability at the trimer apex. The recognized site appears specific to GPC intermediates, with protomers lacking full cleavage between GP1 and GP2 subunits. Guinea pig immunizations with the prefusion-stabilized cleavage-intermediate LASV GPC, first as trimer and then as a nanoparticle, induce neutralizing responses, targeting multiple epitopes including that of D5; we identify a neutralizing antibody (GP23) from the immunized guinea pigs. Collectively, our findings define a prefusion-stabilized GPC trimer, reveal an apex-situated site-of-vulnerability, and demonstrate elicitation of LASV-neutralizing responses by a cleavage-intermediate LASV trimer.