Inflammation and Cancer

: Modelling the consequences of radiotherapy- and Hmgb1-induced inflammation on early-stage skin cancer progression in zebrafish larvae

Abstract

Alongside tumour resection, cancer radiotherapy is a gold standard intervention to manage cancer progression and is given to 27% of cancer patients in a curative or palliative capacity as part of their treatment in England each year (NCRAS, 2017). Yet, by virtue of killing cancer cells, radiotherapy causes acute tissue damage and inflammation at the cancer site, and there is growing evidence to suggest that this inflammation may contribute to the re-population of surviving cancer cells. Despite its routine use within the clinic, it is poorly defined how γ-irradiation causes cancer cell death, and less still is known of the ensuing inflammatory response to surviving cancer cells. I utilise the translucent zebrafish model to visualise how radiotherapy affects cancer progression in vivo and how, if at all, it alters the inflammatory response to cancer cells. I show that radiotherapy stalls cancer expansion by inducing significant dsDNA damage to pre-neoplastic cells which leads to reduced levels of mitosis and increased levels of cancer apoptosis. Furthermore, radiotherapy alters the inflammatory response to pre-neoplastic cells and increases the frequency of neutrophil/pre-neoplastic cell interactions when compared to unirradiated control larvae. These data indicate that as a consequence of tissue damage caused by radiotherapy, the inflammatory response to pre-neoplastic cells is enhanced. The use of anti-inflammatory drugs to suppress this newly identified radiotherapy-induced inflammatory response to cancer may reduce the growth of cancer cells which survive irradiation and improve rates of progression-free survival following cancer radiotherapy.
Pre-neoplastic clones release a damage signal, hydrogen peroxide (H2O2), which recruits inflammatory cells, and these interactions are known to promote clonal expansion. However, immune cells are still recruited to clones that cannot produce H2O2, suggesting that other damage signals may also trigger inflammation and expedite cancer development. High-mobility group box protein 1 (Hmgb1) is a nuclear protein and a damage-associated molecular pattern (DAMP) released at wounds. Hmgb1 overexpression is associated with inflammation and tumour progression, but its role during early clone development is poorly understood. I show that in zebrafish larvae, tissue levels of Hmgb1a are enriched at wounds and in the immediate vicinity of growing RasV12.GFP clones. The subsequent knockout of hmgb1a showed reduced neutrophil recruitment to both wounds and pre-neoplastic clones, while the response of macrophages remained unaffected. Additionally, live imaging showed delayed recruitment of neutrophils to wounds, and shorter lesion interactions upon arrival. Similarly, neutrophil interactions with pre-neoplastic clones were less frequent in hmgb1a-null mutants and lasted for less time when compared to interactions in wild type larvae. Importantly, hmgb1a knockout larvae displayed fewer and smaller pre-neoplastic clones than their wild type siblings. These results suggest that enhanced levels of Hmgb1 around pre-neoplastic clones mimic a wound and enhance the recruitment of neutrophils to the neoplasm, which, in turn, promotes cancer progression. These data indicate that therapies targeting Hmgb1 might be useful in dampening the inflammatory response to cancer to inhibit its growth,

Date of Award	9 May 2023
Original language	English
Awarding Institution	University of Bristol
Supervisor	Paul B Martin (Supervisor) & Catherine Nobes (Supervisor)