Sunday, May 18, 2025
5/18/2025
Harnessing native glycosylation to improve immunogenicity of HIV-1 Env immunogens - Project Summary Developing an effective HIV vaccine to curb HIV infections is a global urgency. Env is the sole target for broadly neutralizing antibodies (bNAbs), and is the focus of vaccination strategies. One barrier to a protective HIV vaccine is the dense array of glycans that covers the surface of the Env spike restricting the immunogenicity of Env proteins. There are ~90 individual glycan sites per trimer and each site can be decorated with high-mannose, hybrid or complex-type glycans. These glycans are crucial for various Env functions including its conformation, antigenicity, immunogenicity and the ability of HIV to evade humoral responses. The presence/absence and type of glycans on Env-based vaccine candidates can influence both antigenicity and immunogenicity of the protein. Most bNAbs include Env glycans as part of their binding epitope, making the glycosylation of Env a key aspect of HIV-1 vaccine design. These bNAbs has been isolated from chronically HIV-1 infected individuals but induction of such bNAbs by vaccination has not been successful yet. Thus, Env immunogens that display glycan and antigenic profile matching those present on the virus may have a better prospect at inducing a cross-neutralizing Ab response. However, a clear understanding of what comprise the site-specific glycan make-up of virus- associated Env is missing, particularly for clinically relevant isolates being pursued for human vaccine trials. The mechanisms/factors that regulate the Env glycan pattern and composition are not understood. This information is critical for improving Env as immunogens and for understanding the optimal epitopes of glycan-dependent bNAbs. The proposed study aims to fill this gap in knowledge by investigating Env biogenesis and glycosylation that starts in the endoplasmic reticulum (ER) compartment of the host cells and continues to be modified in Golgi. A better understanding of the processes that are involved in the ER and Golgi will provide us with the tools that can be used to generate Env immunogens with native antigenic and glycosylation profiles. The overarching goal of the proposed study is to 1) characterize the site-specific glycosylation of native Env on virus, 2) investigate the processes in the ER and Golgi that regulate and imprint the type of glycans on HIV Env, 3) identify ways to express Env with glycans that match virus-associated Env, 4) characterize how native glycans impact the structural and conformational flexibility of Env, and, 5) evaluate the immunogenicity of these Env in animal models for their capacity to elicit cross-neutralizing antibodies. This study will take advantage of several innovative technologies such as LC-MS/MS, HDX-MS along with the complementary expertise of the investigator team to accomplish the goals. Successful completion of the study will reveal several new and unknown aspects of protein synthesis and mechanisms that can be applied towards the development of improved Env immunogens for HIV vaccines.
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