Development of strategies for manufacturing consistent T-cell populations
thesisposted on 09.12.2021, 09:56 authored by Ben Diffey
With an aging population, cancer is becoming increasingly prevalent in society. Statistics now suggest that half of all people will be diagnosed with cancer in their lifetime. In recent years there has been major progress in immuno-oncology, particularly in T-cell based therapies. The main advancement in this field has been the development of Chimeric Antigen Receptor (CAR) T-cell therapies, which use genetically reprogrammed T-cells’ natural cytotoxic pathways to induce apoptosis in target cells.
There has been a large amount of work focussed on identifying T-cell population characteristics which can aid in the manufacture and efficacy of the therapy. In particular, the balance of subpopulations which allow improved viral transduction in culture, and improved efficacy and persistence on administering the therapy. However, there was limited evidence of work investigating the impact of the T-cell culture process and the balance of these subpopulations. This research sought to improve the definition of T-cell culture to improve confidence in predicting the output material as well as identifying conditions which may promote preferred populations.
Input material, delaying IL-2 supplementation, and concentrated incubation prior to standard culture was analysed for their impact on T-cell culture. Viable cell count and flow cytometry were used identify the impact of these elements on culture growth and phenotypic profile. In addition, a novel phenotypic analytical method was developed in response to limitations with the common method observed in the field. The novel method utilised algorithmic analysis of phenotypic data to reduce user bias, whilst introducing optimal phenotypic resolution of analysis.
Analysis of input material and CD4/CD8 cell balance appeared to show that there were limited differences in behaviour of the subpopulations, with primary negative responses seen in cultures that had undergone a greater level of manipulation. Delaying the supplementation of IL-2 into culture appeared to have a positive effect compared to immediate supplementation, however the evidence suggested that this was potentially no better than cultures not supplemented with additional IL-2. Investigation of concentrated incubation prior to seeding into culture had variable responses using different donors. This work proved inconclusive and requires further investigation, however the evidence suggested that the process may induce greater growth in cultures and increased maturation of subpopulations. It was also possible to observe across the breadth of the experimentation that an increase in proliferation of T-cell cultures correlated with an increased phenotypic shift to Effector Memory phenotype populations.
EPSRC and MRC Centre for Doctoral Training in Regenerative Medicine
Engineering and Physical Sciences Research CouncilFind out more...
- Mechanical, Electrical and Manufacturing Engineering