Multi-dose Formulation Development for a Quadrivalent Human Papillomavirus Virus-Like Particle-Based Vaccine: Part II- Real-time and Accelerated Stability Studies

Sharma, Nitya; Jerajani, Kaushal; Wan, Ying; Kumru, Ozan S.; Pullagurla, Swathi R.; Ogun, Oluwadara; Mapari, Shweta; Brendle, Sarah A.; Christensen, Neil D.; Batwal, Saurabh; Mahedvi, Mustafa; Rao, Harish; Dogar, Vikas; Chandrasekharan, Rahul; Shaligram, Umesh; Volkin, David B.; Joshi, Sangeeta B.

doi:10.1016/j.xphs.2022.11.021

Cited by 7 publications

(12 citation statements)

References 31 publications

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“…Interestingly, this rank-ordering of the storage stability profiles of HPV VLPs vs. AP type did not correlate with global destabilization effects measured by DSC [14,16] or with reports from other groups evaluating various HPV VLPs from different sources [13,17]. Thus, while these previously described analytical methods were successful in enabling the development of multi-dose formulations for specific HPV antigens, they were insufficient to elucidate the physicochemical mechanism(s) of degradation.…”

Section: Introductionmentioning

confidence: 72%

A Combined LC-MS and Immunoassay Approach to Characterize Preservative-Induced Destabilization of Human Papillomavirus Virus-like Particles Adsorbed to an Aluminum-Salt Adjuvant

Caringal,

Hickey,

Sharma

et al. 2024

Vaccines

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During the multi-dose formulation development of recombinant vaccine candidates, protein antigens can be destabilized by antimicrobial preservatives (APs). The degradation mechanisms are often poorly understood since available analytical tools are limited due to low protein concentrations and the presence of adjuvants. In this work, we evaluate different analytical approaches to monitor the structural integrity of HPV16 VLPs adsorbed to Alhydrogel™ (AH) in the presence and absence of APs (i.e., destabilizing m-cresol, MC, or non-destabilizing chlorobutanol, CB) under accelerated conditions (pH 7.4, 50 °C). First, in vitro potency losses displayed only modest correlations with the results from two commonly used methods of protein analysis (SDS-PAGE, DSC). Next, results from two alternative analytical approaches provided a better understanding of physicochemical events occurring under these same conditions: (1) competitive ELISA immunoassays with a panel of mAbs against conformational and linear epitopes on HPV16 VLPs and (2) LC-MS peptide mapping to evaluate the accessibility/redox state of the 12 cysteine residues within each L1 protein comprising the HPV16 VLP (i.e., with 360 L1 proteins per VLP, there are 4320 Cys residues per VLP). These methods expand the limited analytical toolset currently available to characterize AH-adsorbed antigens and provide additional insights into the molecular mechanism(s) of AP-induced destabilization of vaccine antigens.

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Section: Introductionmentioning

confidence: 72%

A Combined LC-MS and Immunoassay Approach to Characterize Preservative-Induced Destabilization of Human Papillomavirus Virus-like Particles Adsorbed to an Aluminum-Salt Adjuvant

Caringal,

Hickey,

Sharma

et al. 2024

Vaccines

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“…Finally, an important emphasis of future formulation optimization work with adjuvanted IVX-411 would include lowering costs to improve access for use in LMICs. Examples include lowering raw material and primary packaging costs (e.g., excipients, adjuvant, vials, stoppers), improving vaccine stability in the cold chain (e.g., vaccine vial monitor, VVM, designation), 9 and developing multidose (more doses per vial) 68 , 69 and combination vaccine (more vaccines per dose) 70 formulations.…”

Section: Discussionmentioning

confidence: 99%

Effects of aluminum-salt, CpG and emulsion adjuvants on the stability and immunogenicity of a virus-like particle displaying the SARS-CoV-2 receptor-binding domain (RBD)

Kumru,

Bajoria,

Kaur

et al. 2023

Human Vaccines & Immunotherapeutics

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Second-generation COVID-19 vaccines with improved immunogenicity (e.g., breadth, duration) and availability (e.g., lower costs, refrigerator stable) are needed to enhance global coverage. In this work, we formulated a clinical-stage SARS-CoV-2 receptor-binding domain (RBD) virus-like particle (VLP) vaccine candidate (IVX-411) with widely available adjuvants. Specifically, we assessed the in vitro storage stability and in vivo mouse immunogenicity of IVX-411 formulated with aluminum-salt adjuvants (Alhydrogel™, AH and Adjuphos™, AP), without or with the TLR-9 agonist CpG-1018™ (CpG), and compared these profiles to IVX-411 adjuvanted with an oil-in-water nano-emulsion (AddaVax™, AV). Although IVX-411 bound both AH and AP, lower binding strength of antigen to AP was observed by Langmuir binding isotherms. Interestingly, AH- and AP-adsorbed IVX-411 had similar storage stability profiles as measured by antigen-binding assays (competitive ELISAs), but the latter displayed higher pseudovirus neutralizing titers (pNT) in mice, at levels comparable to titers elicited by AV-adjuvanted IVX-411. CpG addition to alum (AP or AH) resulted in a marginal trend of improved pNTs in stressed samples only, yet did not impact the storage stability profiles of IVX-411. In contrast, previous work with AH-formulations of a monomeric RBD antigen showed greatly improved immunogenicity and decreased stability upon CpG addition to alum. At elevated temperatures (25, 37°C), IVX-411 formulated with AH or AP displayed decreased in vitro stability compared to AV-formulated IVX-411and this rank-ordering correlated with in vivo performance (mouse pNT values). This case study highlights the importance of characterizing antigen-adjuvant interactions to develop low cost, aluminum-salt adjuvanted recombinant subunit vaccine candidates.

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“…This maturation effect is not only related to the nature of the antigen and adjuvant, but also the formulation process conditions (e.g., order of addition). The subsequent impact of maturation effects with AH-adsorbed antigens on their interaction with antigen-specific antibodies in a competitive ELISA assay is likely antigen- HPV vaccine adsorbed to AH as a function of storage time at 4°C [56]. In both these examples a similarly designed competitive ELISA format was employed as was used in this work.…”

Section: Effect Of Antigen-adjuvant Interactions Of Ah-adsorbed Dcfhp...mentioning

confidence: 99%

“…For example,Sawant et al (2021) observed increased mAb binding as a function of storage time at 4°C with the P[4] antigen of a subunit rotavirus vaccine candidate adsorbed to AH[55]. In contrast,Sharma et al (2023) did not observe increased mAb binding of a quadrivalent…”

mentioning

confidence: 91%

Formulation development and comparability studies with an aluminum-salt adjuvanted SARS-CoV-2 Spike ferritin nanoparticle vaccine antigen produced from two different cell lines

Kumru

Sanyal

Friedland

et al. 2023

Preprint

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The development of safe and effective second-generation COVID-19 vaccines to improve affordability and storage stability requirements remains a high priority to expand global coverage. In this report, we describe formulation development and comparability studies with a self-assembled SARS-CoV-2 spike ferritin nanoparticle vaccine antigen (called DCFHP), when produced in two different cell lines and formulated with an aluminum-salt adjuvant (Alhydrogel, AH). Varying levels of phosphate buffer altered the extent and strength of antigen-adjuvant interactions, and these formulations were evaluated for their (1) in vivo performance in mice and (2) in vitro stability profiles. Unadjuvanted DCFHP produced minimal immune responses while AH-adjuvanted formulations elicited greatly enhanced pseudovirus neutralization titers independent of ~100%, ~40% or ~10% of the DCFHP antigen adsorbed to AH. These formulations differed, however, in their in vitro stability properties as determined by biophysical studies and a competitive ELISA for measuring ACE2 receptor binding of AH-bound antigen. Interestingly, after one month of 4C storage, small increases in antigenicity with concomitant decreases in the ability to desorb the antigen from the AH were observed. Finally, we performed a comparability assessment of DCFHP antigen produced in Expi293 and CHO cells, which displayed expected differences in their N-linked oligosaccharide profiles. Despite consisting of different DCFHP glycoforms, these two preparations were highly similar in their key quality attributes including molecular size, structural integrity, conformational stability, binding to ACE2 receptor and mouse immunogenicity profiles. Taken together, these studies support future preclinical and clinical development of an AH-adjuvanted DCFHP vaccine candidate produced in CHO cells.

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Multi-dose Formulation Development for a Quadrivalent Human Papillomavirus Virus-Like Particle-Based Vaccine: Part II- Real-time and Accelerated Stability Studies

Cited by 7 publications

References 31 publications

A Combined LC-MS and Immunoassay Approach to Characterize Preservative-Induced Destabilization of Human Papillomavirus Virus-like Particles Adsorbed to an Aluminum-Salt Adjuvant

A Combined LC-MS and Immunoassay Approach to Characterize Preservative-Induced Destabilization of Human Papillomavirus Virus-like Particles Adsorbed to an Aluminum-Salt Adjuvant

Effects of aluminum-salt, CpG and emulsion adjuvants on the stability and immunogenicity of a virus-like particle displaying the SARS-CoV-2 receptor-binding domain (RBD)

Formulation development and comparability studies with an aluminum-salt adjuvanted SARS-CoV-2 Spike ferritin nanoparticle vaccine antigen produced from two different cell lines

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