The immunogenicity of embryonic stem cell-derived midbrain dopaminergic progenitors and its implications as a therapy to treat patients with Parkinson’s Disease

The progressive degeneration of the A9 dopaminergic (DA) neurons of the substantia nigra lies at the heart of the pathology of Parkinson’s disease (PD). Although current treatment options alleviate the symptoms, they do not prevent the cell death. As such, stem cell-based DA cell replacement therapies provide a promising treatment option, however little is understood about their immunogenicity. This PhD project investigates the immunogenicity of these cells and their implications for PD transplantation. My first aim was to understand the immunogenicity of embryonic stem cell (ESC)-derived midbrain DA progenitors (mDAps) in vitro. To address this, I studied the surface expression of several major immune molecules on these progenitors under normal and inflammatory conditions, given the latter would be the environment post-transplant. MHC-I expression was low in mDAps but was upregulated in inflammatory conditions; MHC-II and the co-stimulatory molecules always remained undetectable. To assess their immunogenicity in vitro, they were co-cultured with peripheral blood mononuclear cells (PBMCs) obtained from healthy blood, and mDAps evoked only little response from T cells. Such a response could though eventually lead to a graft rejection, which formed the basis of my second aim – to knockout (KO) MHC-I expression and to examine their differentiation ability and immunogenicity in vitro. B2M protein, that stabilizes the MHC class I complex, was knocked out using CRISPR-Cas9. These KO cell lines differentiated to authentic DA progenitors and neurons, and their expression of other immune molecules remained the same as their wild-type counterparts. The immunogenicity of these KO cells to T cells was also undetectable. Given that NK cells are toxic to cells lacking MHC-I, the response of NK cells was also assessed and KO mDAps were found to evoke no NK cell response in vitro. Finally, I looked at the effects of immunosuppressive drugs on mDAps survival and maturation in vitro given these agents will be given in the early phase clinical trials using these cells. I found that most of such agents did not affect the survival or maturation of the DA neurons, with the exception of mycophenolate mofetil. Overall, I found that human RC17 derived mDAps were no more immunogenic than human foetal ventral mesencephalic tissue and as such would not require any major change in strategy when used in clinical trials compared to those that have been done with primary human foetal tissue, although I further explored other aspects of this around looking at the effects of immunosuppressive agents on these cells as well as generated a MHC-I KO RC17 cell line.

Name dc:type.qualificationname

Doctor of Philosophy (PhD)

Level dc:type.qualificationlevel

Doctoral

Grantor dc:publisher.institution

University of Cambridge

Year dc:date.issued

2021

Author dc:creator

• Qarin, Shamma

Advisor dc:contributor.advisor

• Barker, Roger

• immunogenicity
• embryonic stem cell
• dopaminergic
• Parkinson's Disease
• cell therapy
• MHC class I gene knockout
• immunosuppression

Licence

• https://creativecommons.org/licenses/by/4.0/

Language dc:language

eng

Author Identifier

0000-0002-9454-365X

OAI identifier oai:identifier

oai:www.repository.cam.ac.uk:1810/339262