Cardiff Metropolitan University
Browse
Owen R Griffiths PhD 10.07.22.pdf (5.88 MB)

The Development and Preclinical Assessment of Ovine Anti-TNF-α Polyclonal Antibodies for the Oral Treatment of Inflammatory Bowel Disease

Download (5.88 MB)
thesis
posted on 2022-10-20, 16:18 authored by Owen Rees Griffiths

  

Tumour Necrosis Factor-alpha (TNF-α) is one of the main drivers of inflammatory bowel disease (IBD). Regulating the actions of this pro-inflammatory cytokine through selective antagonism using an anti-TNF-α monoclonal antibody (mAb) provides effective therapy for IBD. These agents are administered and then distributed systemically in a condition localised to the gastrointestinal tract. Induction of anti-drug antibodies and other undesirable side effects limit their use. 


This research aimed to optimise the production of ovine anti-TNF-α polyclonal antibodies (PcAb) and demonstrate their in vitro functionality to develop a therapeutic oral product for IBD treatment.


The choice of adjuvants and immunisation schedules for the production of ovine anti-TNF-α PcAb was optimised. The novel product's binding, functional potency, and binding kinetics were compared to a licenced anti-TNF-α mAb, infliximab. Functional activity of ovine anti-TNF-α PcAb on TNF-α-dependent tight junction protein (TJP) modulation and cell architecture was assessed in a co-culture human tissue equivalent model of inflammation. 


This study demonstrates that CoVaccine HTTM adjuvant generates significantly greater ovine neutralising PcAbs directed against human TNF-α as compared to Freund’s (with BrijTM-35) adjuvant. Additionally, the use of CoVaccine HTTM is associated with fewer reactions at injection sites. Ovine anti-TNF-α PcAb have broad functionality, as they bind and neutralise murine TNF-α, whilst a mAb, infliximab, does not. BiacoreTM studies demonstrated that the binding kinetics (KD, Ka and Kd) of ovine anti-TNF-α PcAb and infliximab are not significantly different. Additionally, cellular functional binding studies suggest that ovine anti-TNF-α PcAb bind both free and membrane-bound TNF-α.


Functional activity of the novel ovine anti-TNF-α PcAb was demonstrated in an in vitro co-culture model of inflammatory disease. During inflammation, ovine anti-TNF-α PcAb protect Caco-2 monolayer integrity and maintain ZO-1 and occludin localisation. Additionally, they maintain F-actin distribution, cytoskeletal structure and cell architecture.


The findings of this study indicate that ovine anti-TNF-α PcAb are functional and comparable to the monoclonal agent, infliximab, and supports their further development as a pharmacologically active component of an oral IBD treatment. 

Funding

KESS2

History

School

  • School of Sport and Health Sciences

Qualification level

  • Doctoral

Qualification name

  • PhD

Publication year

2022

Usage metrics

    Cardiff Met Theses Collection

    Exports

    RefWorks
    BibTeX
    Ref. manager
    Endnote
    DataCite
    NLM
    DC