Ahmed, Mehreen
(2020)
Identification of drug(s) selectively targeting colorectal cancer stem-like cells for differentiation (differentiation therapy).
PhD thesis, University of Nottingham.
Abstract
Increasing evidence suggests that cancer cell populations contain a small proportion of cells that have the unique self-renewal, tumour-initiating and limited differentiation ability that allow them to resist the therapies and promote cancer recurrence. Therefore, undifferentiated cancer stem-like cells are of major therapeutic interest specially to devise novel diagnostic and therapeutic agents. Herein, the aim of this study is to identify compound(s) that are able to target colorectal cancer stem-like cells by inducing differentiation.
In this study, a 3D colonosphere platform was utilised that selects CSC-like population in vitro and incorporated it to an NCC (NIH Clinical Collection) compound library. Using CDy1 dye, Vorinostat and deficient FBXW7 cells, the stemness properties of the treated colonospheres were initially compared and evaluated, as detailed in chapter 3. This study suggested the use of CDy1 dye represents a fast and simple method to stain live colonospheres.
Next, we identified potential candidate drugs altering live colonospheres with a stem-like characteristics. Selected compounds were then validated in both in vitro organoids and ex vivo colon explant models for their differentiation induction, impediment in neoplastic cell growth, and to elucidate the mechanism of their anticancer activity (Chapters 4-6). Interestingly, three compounds with antibacterial and anti-inflammatory properties were identified; AM404, rifabutin and idarubicin for their antineoplastic ability on CRC.
Tumour-promoting inflammation is involved in CRC development and therapeutic resistance, which accounts for about 20% of cases in CRC. However, the inflammatory mediators and signalling that regulate the potency of anticancer treatment upon CSC differentiation are still not fully defined. This study revealed that anandamide uptake inhibitor AM404 and broad-spectrum antibiotic rifabutin have high potential in preventing CRC stem-like cells features, such as stemness and de-differentiation, migration and drug-resistance. Herein, we also report that, oncogenic FBXL5 (F-box and leucine-rich repeat protein 5) plays a vital role in CRC metastasis and might be a key target for both AM404 and rifabutin to exert their anticancer effects. In addition, AM404 showed sensitivity toward chemo-resistance and caused differentiation in CRC cells, without necessarily inducing cell death. Rifabutin, on the other hand, induced apoptosis and inhibited cell migration independent of the FBXL5 status. Subsequently, our results suggest potential antitumor effect of cytotoxic antibiotic idarubicin, though interestingly, while showing reduction in tumour growth and proliferation, idarubicin also showed possible enrichment of CRC-SCs. Altogether, this study uncovers three new potential compounds for CRC and suggests FBXL5 as a key target for CRC.
In future, further studies of how these drugs affect cancer progression and survival in vivo via exploring the stemness properties unique to the adult SCs, CSCs and their niches can be carried out using novel research strategies such as, single-cell sequencing and functional analyses including the patients derived xenograft models. Such studies will shed light on the origin of CSCs and their progenitors in the inflammation and cancer. The combination of our findings and these techniques will contribute to the elucidation of the essential mechanisms not only for the maintenance of cancer stemness, but also in the repurposing of defined small-molecule candidate compounds in CRC therapeutics.
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