Bioengineered tumour-targeted cytokines for tolerable immunotherapy combinations in solid tumours.
File(s)
Author(s)
Herpels, Melanie
Type
Thesis or dissertation
Abstract
Cytokines are small powerful molecules that allow inter-cellular communication within the tumour microenvironment (TME). Manipulating their levels can promote immune responses against challenging tumour types like triple negative breast cancer (TNBC) and pancreatic ductal adenocarcinoma (PDAC). Both have low survival rates, heterogeneous TMEs, and are mainly treated with toxic chemotherapy. In these tumour types, pro-inflammatory cytokines have failed in the clinic due to toxicity. Therefore, this PhD’s overall objective is to bioengineer two pro-inflammatory cytokines (Interleukin-12 (IL-12) and Interferon γ (IFNγ) through both collagen-anchoring and masking to enhance their safety and efficacy. Each cytokine was conjugated to a collagen-binding domain (CBD) which accumulates in collagenous tumour stroma. Furthermore, CBD- IL12 was masked (M-CBD-IL12) to further reduce its toxicity. As such, the aims of this PhD are:
1. To produce a safe bioengineered IL-12 molecule (M-CBD-IL12) to treat immunologically distinct TNBC mouse models.
2. To test M-CBD-IL12 in immunologically distinct PDAC mouse models and understand the role of the KrasG12D mutation in determining the observed responses.
3. To produce and test bioengineered CBD-IFNγ as alternative immunotherapy for PDAC.
Using the HEK293F and Expi293F expression systems, I produced the two wild-type cytokines and their novel bioengineered counterparts. I consequently tested their biological activity in vitro. I also demonstrated the tumour lysates, but not serum, to cleave the M-CBD-IL12 linker. In vivo, M-CBD-IL12 selectively cured murine TNBC in the EMT6 mouse model, but not 4T1. Differences in collagen and Transforming growth factor-β expression likely contributed to this differential response. I similarly observed differential IL-12 responses between PDAC mouse models, which I demonstrated to be KrasG12D-related through co-culture systems. Finally, CBD-IFNγ remodelled the landscape of the immune-depleted PDAC model and reduced liver metastases. Overall, this thesis has identified subgroups within challenging tumour types that can be treated with safe and tolerable bioengineered cytokines.
1. To produce a safe bioengineered IL-12 molecule (M-CBD-IL12) to treat immunologically distinct TNBC mouse models.
2. To test M-CBD-IL12 in immunologically distinct PDAC mouse models and understand the role of the KrasG12D mutation in determining the observed responses.
3. To produce and test bioengineered CBD-IFNγ as alternative immunotherapy for PDAC.
Using the HEK293F and Expi293F expression systems, I produced the two wild-type cytokines and their novel bioengineered counterparts. I consequently tested their biological activity in vitro. I also demonstrated the tumour lysates, but not serum, to cleave the M-CBD-IL12 linker. In vivo, M-CBD-IL12 selectively cured murine TNBC in the EMT6 mouse model, but not 4T1. Differences in collagen and Transforming growth factor-β expression likely contributed to this differential response. I similarly observed differential IL-12 responses between PDAC mouse models, which I demonstrated to be KrasG12D-related through co-culture systems. Finally, CBD-IFNγ remodelled the landscape of the immune-depleted PDAC model and reduced liver metastases. Overall, this thesis has identified subgroups within challenging tumour types that can be treated with safe and tolerable bioengineered cytokines.
Version
Open Access
Date Issued
2024-05-27
Date Awarded
2024-09-01
Copyright Statement
Attribution-Non Commercial-No Derivatives 4.0 International Licence (CC BY-NC-ND)
Advisor
Anguraj Sadanandam, Jun Ishihara
Sponsor
Cancer Research UK
Publisher Department
Department of Bioengineering
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)