Biology subjects

Cottrell, C.

Publications and source records attributed to Cottrell, C..

7 recordsLinked to original sources

bioRxiv2026

Translating Innovation to Clinic: End-to-End Bioprocess Development and cGMP Manufacturing of N332-GT5 HIV Vaccine Candidate for First-in-Human Trials HVTN144

The successful translation of rationally designed HIV-1 immunogens into effective vaccines requires manufacturing platforms that maintain structural conformity while meeting clinical-grade quality standards. We developed and scaled a robust, cGMP-compliant process for N332-GT5 gp140, a germline-targeting envelope trimer designed to initiate broadly neutralizing antibody responses, which is now undergoing first-in-human evaluation in HVTN144. Starting with a stable CHO cell line developed using Leap-In(R) transposon technology, we established a production clone exhibiting high-titer expression (>200 mg/L) and genetic stability through 60 population doublings. The manufacturing process scaled efficiently from Ambr(R) 250 miniature bioreactors to 200-L single-use systems, delivering consistent product quality across multiple cGMP batches. A streamlined three-step purification strategy--affinity capture, multimodal polishing, and viral clearance- yielded >99% trimeric purity with preserved quaternary structure and native-like antigenicity. Orthogonal LC-MS analyses confirmed site-specific glycan occupancy matching design specifications, while robust viral clearance exceeded 18-log and 11-log reductions for model retroviruses. Clinical material manufactured through this platform has been successfully administered in HVTN144. This work establishes a scalable, reproducible manufacturing paradigm for structurally complex HIV-1 envelope immunogens, advancing the field toward rational vaccine design based on germline-targeting principles.

bioengineering↗

bioRxiv2025

DNA origami vaccines program antigen-focused germinal centers

Recruitment and expansion of rare precursor B cells in germinal centers (GCs) is a central goal of vaccination to generate broadly neutralizing antibodies (bnAbs) against challenging pathogens such as HIV. Multivalent immunogen display is a well-established method to enhance vaccine-induced B cell responses, typically accomplished by using natural or engineered protein scaffolds. However, these scaffolds themselves are targets of antibody responses, with the potential to generate competitor scaffold-specific B cells that could theoretically limit expansion and maturation of "on-target" B cells in the GC response. Here, we rationally designed T-independent, DNA-origami based virus-like particles (VLPs) with optimal antigenic display of the germline targeting HIV Env immunogen, eOD-GT8, and appropriate T cell help to achieve a potent GC response. In preclinical mouse models, these DNA-VLPs expanded significantly higher frequencies of epitope-specific GC B cells compared with a state-of-the-art clinical protein nanoparticle. Optimized DNA-VLPs primed germinal centers focused on the target antigen and rapidly expanded subdominant broadly neutralizing antibody precursor B cells for HIV with a single immunization. Thus, avoiding scaffold-specific responses augments priming of bnAb precursor B cells, and DNA-VLPs are a promising platform for promoting B cell responses towards challenging subdominant epitopes.

immunology↗

bioRxiv2022

Efficient isolation of rare B cells using next-generation antigen barcoding

The ability to efficiently isolate antigen-specific B cells in high throughput will greatly accelerate the discovery of therapeutic monoclonal antibodies (mAbs) and catalyze rational vaccine development. Traditional mAb discovery is a costly and labor-intensive process, although recent advances in single-cell genomics using emulsion microfluidics allow simultaneous processing of thousands of individual cells. Here we present a streamlined method for isolation and analysis of large numbers of antigen-specific B cells, including next generation antigen barcoding and an integrated computational framework for B cell multi-omics. We demonstrate the power of this approach by recovering thousands of antigen-specific mAbs, including the efficient isolation of extremely rare precursors of VRC01-class and IOMA-class broadly neutralizing HIV mAbs.

immunology↗

bioRxiv2022

A broad and potent neutralization epitope in SARS-related coronaviruses

Many neutralizing antibodies (nAbs) elicited to ancestral SARS-CoV-2 through natural infection and vaccination generally have reduced effectiveness to SARS-CoV-2 variants. Here we show therapeutic antibody ADG20 is able to neutralize all SARS-CoV-2 variants of concern (VOCs) including Omicron (B.1.1.529) as well as other SARS-related coronaviruses. We delineate the structural basis of this relatively escape-resistant epitope that extends from one end of the receptor binding site (RBS) into the highly conserved CR3022 site. ADG20 can then benefit from high potency through direct competition with ACE2 in the more variable RBS and interaction with the more highly conserved CR3022 site. Importantly, antibodies that are able to target this site generally neutralize all VOCs, albeit with reduced potency against Omicron. Thus, this highly conserved and vulnerable site can be exploited for design of universal vaccines and therapeutic antibodies.

microbiology↗

bioRxiv2020

Enhancing glycan occupancy of soluble HIV-1 envelope trimers to mimic the native viral spike

SummaryThe HIV-1 envelope glycoprotein (Env) trimer is decorated with N-linked glycans, which are attached to asparagine residues in the amino acid sequon NxT/S by oligosaccharyltransferases (OST). Artificial glycan “holes” exist when a PNGS is under-occupied on recombinant Env-based vaccines, but not on their viral counterpart. Native-like SOSIP trimers, including clinical candidates, have these artificial holes in the glycan shield that induce strain-specific neutralizing antibodies (NAbs) or non-NAbs. To increase PNGS occupancy, eliminate artificial glycan holes, and mimic the glycosylation of native BG505 Env, we replaced all 12 NxS sequons on the BG505 SOSIP trimer with NxT, thereby increasing the affinity of the sequons for OST. All PNGS, except N133 and N160, were nearly fully occupied on the modified trimer. Occupancy of the N133 site could be increased by changing N133 to NxS, while occupancy of the N160 site could be restored by reverting the nearby N156 sequon to NxS. Hence, OST affinity can influence glycan occupancy when two PNGS are in close proximity. Increasing glycan occupancy should reduce off-target immune responses to artificial glycan holes on vaccine antigens.Competing Interest StatementR. W. S. and J. P. M. are listed on patents concerning stabilized Env trimers.View Full Text

biochemistry↗

bioRxiv2019

Mapping polyclonal antibody responses in non-human primates vaccinated with HIV Env trimer subunit vaccines

Rational immunogen design aims to focus antibody responses to vulnerable sites on the primary antigens. Given the size of these antigens there is however potential for eliciting unwanted, off-target responses. Here, we used our electron microscopy polyclonal epitope mapping approach to describe the antibody specificities elicited by immunization of non-human primates with soluble HIV envelope trimers and subsequent repeated viral challenge. An increased diversity of epitopes recognized, and the approach angle by which these antibodies bound, constituted a hallmark of the humoral response in most protected animals. We also show that fusion peptide-specific antibodies are responsible for some neutralization breadth. Moreover, cryoEM analysis of a fully-protected animal revealed a high degree of clonality within a subset of putatively neutralizing antibodies, enabling a detailed molecular description of the antibody paratope. Our results provide important insights into the immune response against a vaccine candidate that entered into clinical trials earlier this year.

immunology↗

bioRxiv2019

Neutralizing antibody responses to an HIV envelope glycan hole are not easily broadened

Extensive studies with subtype A BG505-derived HIV envelope glycoprotein (Env) SOSIP immunogens have revealed that the dominant autologous neutralizing site in rabbits is located in an exposed region of the heavily glycosylated trimer that lacks potential N-linked glycosylation sites at positions 230, 241, and 289. The Env derived from B41, a subtype B virus, shares a glycan hole centered on positions 230 and 289. BG505 and B41 SOSIP immunogens were combined to test whether immunization in rabbits could induce broader Tier 2 neutralizing responses to the common glycan hole shared between BG505 and B41. Here we isolated autologous neutralizing antibodies (nAbs) that were induced by immunization with B41 SOSIP alone, as well as B41 and BG505 co-immunization, and describe their structure in complex with the B41 SOSIP trimer. Our data suggest that distinct autologous nAb lineages are induced by BG505 and B41 immunogens, even when both immunogens were administered together. In contrast to previously described BG505 glycan hole antibodies, the B41-specific nAbs accommodate the highly conserved N241 glycan (>97% conserved), which is present in B41. Single particle cryo-electron microscopy (cryoEM) studies confirmed that B41 and BG505-specific nAbs bind to overlapping glycan hole epitopes. In an attempt to broaden the reactivity of a B41-specific nAb, mutations in the BG505 glycan hole epitope guided by our high-resolution data only recovered partial binding. Overall, designing prime-boost immunogens to increase the breath of nAb responses directed at glycan holes epitopes remains challenging even when the typically immunodominant glycan holes despite overlap with different Envs.\n\nIMPORTANCEA glycan hole is one of the most dominant autologous neutralizing epitopes targeted on BG505 and B41 SOSIP trimer immunized rabbits. Our high-resolution cryoEM studies of B41 in complex with a B41-specific antibody complex elucidate the molecular basis of this strain-specific glycan hole response. We conclude that eliciting cross-reactive responses to this region would likely require hybrid immunogens that bridge between BG505 and B41.

immunology↗