Document Type

Article

Publication Date

5-21-2026

Comments

This article is the author's final published version in Vaccines, Volume 14, Issue 5, 2026, Article Number 461.

The published version is available at https://doi.org/10.3390/vaccines14050461. Copyright © 2026 The Author(s).

Abstract

Background: Live-attenuated Listeria monocytogenes (Lm) vectors are a clinically validated cancer immunotherapy platform, but translation requires reproducible, clinically realistic workflows for dose preparation and infusion. For live bacterial products, in-use stability and device compatibility can drive dose variability through adsorption, settling, and device losses. Methods: We developed and GMP-manufactured an attenuated Lm vaccine expressing human GUCY2C (Lm-GUCY2C) and performed translational characterization, including construct verification and immunogenicity readouts, and defined the administration-focused in-use stability and device compatibility. Post-thaw stability was assessed in primary cryovials and during preparation and delivery from 250 mL saline infusion bags using standard clinical devices (syringes/needles, filter-free IV tubing) and OnGuard2 closed-system components. Samples were collected over 24 h at room temperature, and viable Lm-GUCY2C were quantified by CFU recovery. Results: Lm-GUCY2C remained stable in thawed cryovials for 24 h with no significant CFU loss. High-dose infusion bags (3 × 109 CFU/bag) maintained CFU recovery through 6 h, whereas low-dose bags (3 × 108 CFU/bag) exhibited significant losses beginning at 3 h, supporting a practical in-use window of up to 2 h for low-dose preparations. OnGuard2 intravenous (i.v.) connectors did not measurably affect CFU recovery, while OnGuard2 vial adapters reduced recovery. Conclusions: This work provides an end-to-end, translationally focused characterization of a GMP-manufactured Lm cancer vaccine, including clinically actionable in-use handling constraints and device compatibility. These data define preparation and administration guardrails (notably, time-to-infusion limits for low-dose bag preparations) that can improve dose accuracy and reproducibility in clinical testing.

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Creative Commons License
This work is licensed under a Creative Commons Attribution 4.0 License.

Language

English

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