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A non-affinity purification process for GMP production of prefusion-closed HIV-1 envelope trimers from clades A and C for clinical evaluation.
(Vaccine, 2021-06) Gulla, Krishana; Cibelli, Nicole; Cooper, Jonathan W; Fuller, Haley C; Schneiderman, Zachary; Witter, Sara; Zhang, Yaqiu; Changela, Anita; Geng, Hui; Hatcher, Christian; Narpala, Sandeep; Tsybovsky, Yaroslav; Zhang, Baoshan; Vrc Production Program; McDermott, Adrian B; Kwong, Peter D; Gowetski, Daniel B
Metastable glycosylated immunogens present challenges for GMP manufacturing. The HIV-1 envelope (Env) glycoprotein trimer is covered by N-linked glycan comprising half its mass and requires both trimer assembly and subunit cleavage to fold into a prefusion-closed conformation. This conformation, the vaccine-desired antigenic state, is both metastable to structural rearrangement and labile to subunit dissociation. Prior reported GMP manufacturing for a soluble trimer stabilized in a near-native state by disulfide (SOS) and Ile-to-Pro (IP) mutations has employed affinity methods based on antibody 2G12, which recognizes only ~30% of circulating HIV strains. Here, we develop a scalable manufacturing process based on commercially available, non-affinity resins, and we apply the process to current GMP (cGMP) production of trimers from clades A and C, which have been found to boost cross-clade neutralizing responses in vaccine-test species. The clade A trimer, which we named "BG505 DS-SOSIP.664", contained an engineered disulfide (201C-433C; DS) within gp120, which further stabilized this trimer in a prefusion-closed conformation resistant to CD4-induced triggering. BG505 DS-SOSIP.664 was expressed in a CHO-DG44 stable cell line and purified with initial and final tangential flow filtration steps, three commercially available resin-based chromatography steps, and two orthogonal viral clearance steps. The non-affinity purification enabled efficient scale-up, with a 250 L-scale cGMP run yielding 9.6 g of purified BG505 DS-SOSIP.664. Antigenic analysis indicated retention of a prefusion-closed conformation, including recognition by apex-directed and fusion peptide-directed antibodies. The developed manufacturing process was suitable for 50 L-scale production of a second prefusion-stabilized Env trimer vaccine candidate, ConC-FP8v2 RnS-3mut-2G-SOSIP.664, yielding 7.8 g of this consensus clade C trimer. The successful process development and purification scale-up of HIV-1 Env trimers from different clades by using commercially available materials provide experimental demonstration for cGMP manufacturing of trimeric HIV-Env vaccine immunogens, in an antigenically desired conformation, without the use of costly affinity resins.
Low-Dose Subcutaneous or Intravenous Monoclonal Antibody to Prevent Malaria.
(The New England journal of medicine, 2022-08) Wu, Richard L; Idris, Azza H; Berkowitz, Nina M; Happe, Myra; Gaudinski, Martin R; Buettner, Christian; Strom, Larisa; Awan, Seemal F; Holman, LaSonji A; Mendoza, Floreliz; Gordon, Ingelise J; Hu, Zonghui; Campos Chagas, Andrezza; Wang, Lawrence T; Da Silva Pereira, Lais; Francica, Joseph R; Kisalu, Neville K; Flynn, Barbara J; Shi, Wei; Kong, Wing-Pui; O'Connell, Sarah; Plummer, Sarah H; Beck, Allison; McDermott, Adrian; Narpala, Sandeep R; Serebryannyy, Leonid; Castro, Mike; Silva, Rosa; Imam, Marjaan; Pittman, Iris; Hickman, Somia P; McDougal, Andrew J; Lukoskie, Ashly E; Murphy, Jittawadee R; Gall, Jason G; Carlton, Kevin; Morgan, Patricia; Seo, Ellie; Stein, Judy A; Vazquez, Sandra; Telscher, Shinyi; Capparelli, Edmund V; Coates, Emily E; Mascola, John R; Ledgerwood, Julie E; Dropulic, Lesia K; Seder, Robert A; VRC 614 Study Team
Background
New approaches for the prevention and elimination of malaria, a leading cause of illness and death among infants and young children globally, are needed.Methods
We conducted a phase 1 clinical trial to assess the safety and pharmacokinetics of L9LS, a next-generation antimalarial monoclonal antibody, and its protective efficacy against controlled human malaria infection in healthy adults who had never had malaria or received a vaccine for malaria. The participants received L9LS either intravenously or subcutaneously at a dose of 1 mg, 5 mg, or 20 mg per kilogram of body weight. Within 2 to 6 weeks after the administration of L9LS, both the participants who received L9LS and the control participants underwent controlled human malaria infection in which they were exposed to mosquitoes carrying Plasmodium falciparum (3D7 strain).Results
No safety concerns were identified. L9LS had an estimated half-life of 56 days, and it had dose linearity, with the highest mean (±SD) maximum serum concentration (Cmax) of 914.2±146.5 μg per milliliter observed in participants who had received 20 mg per kilogram intravenously and the lowest mean Cmax of 41.5±4.7 μg per milliliter observed in those who had received 1 mg per kilogram intravenously; the mean Cmax was 164.8±31.1 in the participants who had received 5 mg per kilogram intravenously and 68.9±22.3 in those who had received 5 mg per kilogram subcutaneously. A total of 17 L9LS recipients and 6 control participants underwent controlled human malaria infection. Of the 17 participants who received a single dose of L9LS, 15 (88%) were protected after controlled human malaria infection. Parasitemia did not develop in any of the participants who received 5 or 20 mg per kilogram of intravenous L9LS. Parasitemia developed in 1 of 5 participants who received 1 mg per kilogram intravenously, 1 of 5 participants who received 5 mg per kilogram subcutaneously, and all 6 control participants through 21 days after the controlled human malaria infection. Protection conferred by L9LS was seen at serum concentrations as low as 9.2 μg per milliliter.Conclusions
In this small trial, L9LS administered intravenously or subcutaneously protected recipients against malaria after controlled infection, without evident safety concerns. (Funded by the National Institute of Allergy and Infectious Diseases; VRC 614 ClinicalTrials.gov number, NCT05019729.).A Monoclonal Antibody for Malaria Prevention.
(The New England journal of medicine, 2021-08) Gaudinski, Martin R; Berkowitz, Nina M; Idris, Azza H; Coates, Emily E; Holman, LaSonji A; Mendoza, Floreliz; Gordon, Ingelise J; Plummer, Sarah H; Trofymenko, Olga; Hu, Zonghui; Campos Chagas, Andrezza; O'Connell, Sarah; Basappa, Manjula; Douek, Naomi; Narpala, Sandeep R; Barry, Christopher R; Widge, Alicia T; Hicks, Renunda; Awan, Seemal F; Wu, Richard L; Hickman, Somia; Wycuff, Diane; Stein, Judy A; Case, Christopher; Evans, Brian P; Carlton, Kevin; Gall, Jason G; Vazquez, Sandra; Flach, Britta; Chen, Grace L; Francica, Joseph R; Flynn, Barbara J; Kisalu, Neville K; Capparelli, Edmund V; McDermott, Adrian; Mascola, John R; Ledgerwood, Julie E; Seder, Robert A; VRC 612 Study Team
Background
Additional interventions are needed to reduce the morbidity and mortality caused by malaria.Methods
We conducted a two-part, phase 1 clinical trial to assess the safety and pharmacokinetics of CIS43LS, an antimalarial monoclonal antibody with an extended half-life, and its efficacy against infection with Plasmodium falciparum. Part A of the trial assessed the safety, initial side-effect profile, and pharmacokinetics of CIS43LS in healthy adults who had never had malaria. Participants received CIS43LS subcutaneously or intravenously at one of three escalating dose levels. A subgroup of participants from Part A continued to Part B, and some received a second CIS43LS infusion. Additional participants were enrolled in Part B and received CIS43LS intravenously. To assess the protective efficacy of CIS43LS, some participants underwent controlled human malaria infection in which they were exposed to mosquitoes carrying P. falciparum sporozoites 4 to 36 weeks after administration of CIS43LS.Results
A total of 25 participants received CIS43LS at a dose of 5 mg per kilogram of body weight, 20 mg per kilogram, or 40 mg per kilogram, and 4 of the 25 participants received a second dose (20 mg per kilogram regardless of initial dose). No safety concerns were identified. We observed dose-dependent increases in CIS43LS serum concentrations, with a half-life of 56 days. None of the 9 participants who received CIS43LS, as compared with 5 of 6 control participants who did not receive CIS43LS, had parasitemia according to polymerase-chain-reaction testing through 21 days after controlled human malaria infection. Two participants who received 40 mg per kilogram of CIS43LS and underwent controlled human malaria infection approximately 36 weeks later had no parasitemia, with serum concentrations of CIS43LS of 46 and 57 μg per milliliter at the time of controlled human malaria infection.Conclusions
Among adults who had never had malaria infection or vaccination, administration of the long-acting monoclonal antibody CIS43LS prevented malaria after controlled infection. (Funded by the National Institute of Allergy and Infectious Diseases; VRC 612 ClinicalTrials.gov number, NCT04206332.).A sublingual nanofiber vaccine to prevent urinary tract infections.
(Science advances, 2022-11) Kelly, Sean H; Votaw, Nicole L; Cossette, Benjamin J; Wu, Yaoying; Shetty, Shamitha; Shores, Lucas S; Issah, Luqman A; Collier, Joel H
Urinary tract infections (UTIs) are a major public health problem affecting millions of individuals each year. Recurrent UTIs are managed by long-term antibiotic use, making the alarming rise of antibiotic resistance a substantial threat to future UTI treatment. Extended antibiotic regimens may also have adverse effects on the microbiome. Here, we report the use of a supramolecular vaccine to provide long-term protection against uropathogenic Escherichia coli, which cause 80% of uncomplicated UTIs. We designed mucus-penetrating peptide-polymer nanofibers to enable sublingual (under the tongue) vaccine delivery and elicit antibody responses systemically and in the urogenital tract. In a mouse model of UTI, we demonstrate equivalent efficacy to high-dose oral antibiotics but with significantly less perturbation of the gut microbiome. We also formulate our vaccine as a rapid-dissolving sublingual tablet that raises response in mice and rabbits. Our approach represents a promising alternative to antibiotics for the treatment and prevention of UTIs.
A Balance between Pro-Inflammatory and Pro-Reparative Macrophages is Observed in Regenerative D-MAPS.
(Advanced science (Weinheim, Baden-Wurttemberg, Germany), 2023-04) Liu, Yining; Suarez-Arnedo, Alejandra; Shetty, Shamitha; Wu, Yaoying; Schneider, Michelle; Collier, Joel H; Segura, Tatiana
Microporous annealed particle scaffolds (MAPS) are a new class of granular materials generated through the interlinking of tunable microgels, which produce an interconnected network of void space. These microgel building blocks can be designed with different mechanical or bio-active parameters to facilitate cell infiltration and modulate host response. Previously, changing the chirality of the microgel crosslinking peptides from L- to D-amino acids led to significant tissue regeneration and functional recovery in D-MAPS-treated cutaneous wounds. In this study, the immunomodulatory effect of D-MAPS in a subcutaneous implantation model is investigated. How macrophages are the key antigen-presenting cells to uptake and present these biomaterials to the adaptive immune system is uncovered. A robust linker-specific IgG2b/IgG1 response to D-MAPS is detected as early as 14 days post-implantation. The fine balance between pro-regenerative and pro-inflammatory macrophage phenotypes is observed in D-MAPS as an indicator for regenerative scaffolds. The work offers valuable insights into the temporal cellular response to synthetic porous scaffolds and establishes a foundation for further optimization of immunomodulatory pro-regenerative outcomes.