Production and Immunogenicity of Soluble Plant-Produced HIV-1 Subtype C Envelope gp140 Immunogens

Abstract

The development of effective vaccines is urgently needed to curb the spread of human immunodeficiency virus type 1 (HIV-1). A major focal point of current HIV vaccine research is the production of soluble envelope (Env) glycoproteins which reproduce the structure of the native gp160 trimer. These antigens are produced in mammalian cells, which requires a sophisticated infrastructure for manufacture that is mostly absent in developing countries. The production of recombinant proteins in plants is an attractive alternative for the potentially cheap and scalable production of vaccine antigens, especially for developing countries. In this study, we developed a transient expression system in Nicotiana benthamiana for the production of soluble HIV Env gp140 antigens based on two rationally selected virus isolates (CAP256 SU and Du151). The scalability of the platform was demonstrated and both affinity and size exclusion chromatography (SEC) were explored for recovery of the recombinant antigens. Rabbits immunized with lectin affinity-purified antigens developed high titres of binding antibodies, including against the V1V2 loop region, and neutralizing antibodies against Tier 1 viruses. The removal of aggregated Env species by gel filtration resulted in the elicitation of superior binding and neutralizing antibodies. Furthermore, a heterologous prime-boost regimen employing a recombinant modified vaccinia Ankara (rMVA) vaccine, followed by boosts with the SEC-purified protein, significantly improved the immunogenicity. To our knowledge, this is the first study to assess the immunogenicity of a near-full length plant-derived Env vaccine immunogen.

Co‐expression of human calreticulin significantly improves the production of HIV gp140 and other viral glycoproteins in plants

“…Given the divergence of the plant and human chaperone machinery it is plausible that the HIV Env glycoprotein is inefficiently folded in plants, leading to ER-stress. This would account for the pathology that we previously reported following transient expression of Env gp140 (Margolin et al, 2019). Considering the dramatic improvement in Env production following CRT co-expression, we were interested in determining how this impacted the ER stress response.…”

“…The lack of naturally occurring furin in plants complicates the production of authentic viral glycoproteins which may not be properly processed without the protease (Faye et al, 2005;Pasquato et al, 2013). In our previous study we circumvented the need for furin cleavage of HIV Env by replacing the cleavage site with a glycine-rich flexible linker peptide (Margolin et al, 2019). In addition to improving the expression of this cleavageindependent Env antigen by co-expressing chaperones, we were also interested in exploring the production of a fully cleaved HIV gp140.…”

“…We therefore designed an HIV Env antigen based on the cleaved "SOSIP" antigen design described for the prototypic Env vaccine BG505 Chaperone co-expression with glycoproteins 2111 SOSIP.664, which is being evaluated in clinical trials (Sanders et al, 2013;Sanders et al, 2015). Two changes were made to the HIV gp140 antigen described previously (Margolin et al, 2019) to generate the SOSIP protein: a multi-basic (RRRRRR) cleavage site was included in place of the flexible linker and an artificial disulphide bridge was added to stabilize the association of gp41 and gp120 following cleavage (Binley et al, 2000;Binley et al, 2002;Sanders et al, 2002). Transient expression of the antigen and CRT with human furin resulted in a 15-20 kDa size shift corresponding to the cleavage of a 157 amino acid fragment (Figure 4a).…”

Emergence of Novel Coronavirus 2019-nCoV: Need for Rapid Vaccine and Biologics Development

“…Recent advancements in the plant expression strategies, especially the development of transient expression system or viral vectors, resulted in a huge increase in the protein yield that makes this plant host system, a promising system for the production of various biopharmaceutical proteins [16]. Several reports in the last two decades have enough evidence to prove that the plant produced biopharmaceuticals are as effective as the mammalian cell-based proteins and also elicit potent neutralizing antibodies, or shown therapeutic effects against the particular pathogen or infection [17][18][19]. The use of plants for the production of recombinant proteins and biopharmaceuticals has been gaining importance since the plant produced biologic taliglucerase alfa has been commercialized in 2012 against Gaucher's disease that proclaimed a new era for plant made biopharmaceutical and triggered the innovation in the field of biopharmaceuticals [20].…”

What Does Plant-Based Vaccine Technology Offer to the Fight against COVID-19?

“…Similarly, pure antigens produced in plants have been assessed with promising results in schemes where a conventional vaccine is used as a priming agent and the plant-made antigen is administered by the oral route as boosting [106]. Exploring these schemes will be useful to achieve the best immune profile against SARS-CoV-2 in terms of safe and long-lasting protective immune responses.…”