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CHO expressed recombinant human lactoferrin as an adjuvant for BCG.

ABSTRACT: Lactoferrin (LF), an iron binding protein with immune modulatory activities, has adjuvant activity to enhance vaccine efficacy. Tuberculosis (TB) is a pulmonary disease caused by the pathogen Mycobacterium tuberculosis (MTB). Progressive TB disease is clinically defined by damaging pulmonary pathology, a result of inflammation due to immune reactivity. The current vaccine for TB, an attenuated strain of Mycobacterium bovis, Bacillus Calmette Guerin (BCG), has only limited efficacy to prevent adult pulmonary TB. This study examines a Chinese hamster ovary (CHO) expressed recombinant human LF (rHLF) to boost efficacy of the BCG vaccine and delay early pathology post infectious challenge. C57BL/6 mice were immunized with BCG, or BCG admixed with either rHLF or bovine LF (bLF; internal control), or remained unvaccinated. Mice were then aerosol challenged with Erdman MTB. All vaccinated mice demonstrated decreased organ bacterial load up to 19 weeks post infection compared with non-vaccinated controls. Furthermore, mice receiving bLF or rHLF supplemented BCG vaccines showed a modest decrease in lung pathology developed over time, compared to the BCG vaccine alone. While mice vaccinated with BCG/rHLF demonstrated increased general lung inflammation at day 7, it occurred without noticeable increase in pro-inflammatory cytokines. At later times, decreased pathology in the rHLF groups correlated with decreased inflammatory cytokines. Splenic recall to BCG antigens showed BCG/rHLF vaccination increased production of IFN-?, IL-6, and GM-CSF compared to naïve, BCG, and BCG/bLF groups. Analysis of T cell stimulating functions of bone marrow derived macrophages and dendritic cells treated with BCG/bLF or BCG/rHLF showed decreases in IL-10 production when co-cultured with sensitized CD4 and CD8 T cells, compared to those cultured with macrophages/dendritic cells treated with BCG without LF. These results indicate that addition of rHLF to the BCG vaccine can modulate development of host pathology early post infectious challenge, most likely through host immune regulation affecting hypersensitive responses.

SUBMITTER: Hwang SA

PROVIDER: S-EPMC6004602 | biostudies-literature | 2015 Dec

REPOSITORIES: biostudies-literature

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CHO expressed recombinant human lactoferrin as an adjuvant for BCG.

Hwang Shen-An SA Kruzel Marian L ML Actor Jeffrey K JK

International journal of immunopathology and pharmacology 20150827 4

Lactoferrin (LF), an iron binding protein with immune modulatory activities, has adjuvant activity to enhance vaccine efficacy. Tuberculosis (TB) is a pulmonary disease caused by the pathogen Mycobacterium tuberculosis (MTB). Progressive TB disease is clinically defined by damaging pulmonary pathology, a result of inflammation due to immune reactivity. The current vaccine for TB, an attenuated strain of Mycobacterium bovis, Bacillus Calmette Guerin (BCG), has only limited efficacy to prevent adu ...[more]

PMID: 26315722

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Project description:Therapeutic glycoproteins have played a major role in the commercial success of biotechnology in the post-genomic era. But isolating recombinant mammalian cell lines for large-scale production remains costly and time-consuming, due to substantial variation and unpredictable stability of expression amongst transfected cells, requiring extensive clone screening to identify suitable high producers. Streamlining this process is of considerable interest to industry yet the underlying phenomena are still not well understood. Here we examine an antibody-expressing Chinese hamster ovary (CHO) clone at single-cell resolution using flow cytometry and vectors, which couple light and heavy chain transcription to fluorescent markers. Expression variation has traditionally been attributed to genetic heterogeneity arising from random genomic integration of vector DNA. It follows that single cell cloning should yield a homogeneous cell population. We show, in fact, that expression in a clone can be surprisingly heterogeneous (standard deviation 50 to 70% of the mean), approaching the level of variation in mixed transfectant pools, and each antibody chain varies in tandem. Phenotypic variation is fully developed within just 18 days of cloning, yet is not entirely explained by measurement noise, cell size, or the cell cycle. By monitoring the dynamic response of subpopulations and subclones, we show that cells also undergo slow stochastic fluctuations in expression (half-life 2 to 11 generations). Non-genetic diversity may therefore play a greater role in clonal variation than previously thought. This also has unexpected implications for expression stability. Stochastic gene expression noise and selection bias lead to perturbations from steady state at the time of cloning. The resulting transient response as clones reestablish their expression distribution is not ordinarily accounted for but can contribute to declines in median expression over timescales of up to 50 days. Noise minimization may therefore be a novel strategy to reduce apparent expression instability and simplify cell line selection.

Dataset Information

CHO expressed recombinant human lactoferrin as an adjuvant for BCG.

Publications

CHO expressed recombinant human lactoferrin as an adjuvant for BCG.

Similar Datasets

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