Abstract
Prostate cancer (PCa) is a common cause of cancer deaths worldwide among men. In 2020, nearly 1.4 million new cases of PCa were reported globally. In New Zealand, around 4,000 men are diagnosed with PCa each year, with over 650 deaths attributed to the disease annually. According to the Cancer Society of New Zealand, Māori men represent approximately 7% of these cases. Studies suggest that hypoxia and inflammation are linked to the development of aggressive PCa. Interleukin-6 (IL-6) is a widely studied marker of prostatic inflammation, and elevated serum levels of IL-6 and its signal transducer IL6ST have been found to correlate with metastases and hypoxia in aggressive PCa. The six-transmembrane epithelial antigen of the prostate 4 (STEAP4), regulated by hypoxia and IL-6, is highly expressed in PCa.
Additionally, the tumour suppressor p53 has been associated with PCa, particularly the Δ133TP53 mRNA isoform, which is increased in hypoxic PCa cells in vitro and in a subset of PCa patients with shorter disease-free survival. Δ133p53 promotes cell migration and invasion via IL-6-associated JAK/STAT3 and RhoA-ROCK signalling pathways. These findings suggest a strong link between Δ133p53, IL-6/IL6ST, STEAP4, and the influence of hypoxia in PCa. However, whether hypoxia induces Δ133p53, IL6ST, and STEAP4 independently or together in PCa is unclear. It is also unclear whether Δ133p53 regulates IL6ST and what implications this regulation has on PCa biology.
This PhD project aimed to investigate the impact of hypoxia, with or without hormone reduction, on the expression of p53 isoforms in three PCa and two BrCa cell lines. To achieve this, three PCa cell lines (LNCaP, 22Rv1 and DU145) with or without TP53 mutations and androgen (AR) responses and two BrCa cell lines (MCF7 and MDA-MB231) with or without TP53 mutations and oestrogen responses were utilised. The cells were cultured in media with or without phenol-red to attenuate AR/ER-mediated cell response before exposure to hypoxia for 24 hours and 48 hours. Results showed that hypoxia stabilised FLp53/p53α in wild-type (WT) TP53 PCa cells (LNCaP) and induced Δ133TP53 mRNA in heterozygous TP53 mutant PCa cells (22Rv1), regardless of hormonal status. However, this effect was not observed in WT-TP53 BrCa cells (MCF7) and mutant TP53 PCa (DU145) and BrCa (MDA-MB-231) cells. Additionally, we investigated the effect of hypoxia on IL6ST and STEAP4 in three PCa lines. Data showed that hypoxia did not alter IL6ST and STEAP4 mRNA in PCa cells.
Based on the observation that Δ133p53 promotes cell migration and invasion through the IL-6-associated JAK/STAT3 and RhoA-ROCK signalling pathways, we hypothesised that Δ133p53 might regulate the upstream signal transducer IL6ST in PCa cells. To test this hypothesis, Δ133p53 and IL6ST were silenced individually or together in three PCa cell lines (LNCaP, 22Rv1, and PC3). The results of the experiment showed that Δ133p53 and IL6ST can regulate each other's expression in wild-type TP53 PCa cells (LNCaP) potentially via a feedback loop driven by TP53 mutation status and the presence of an intact TP53 locus. Additionally, Δ133p53α was found to negatively regulate the phosphorylation of AKT in WT-TP53 PCa cells with non-functional JAK/STAT and Ras/Raf/MEK/ERK1/2/MAPK pathways.
We have discovered that Δ133p53α and Δ133p53β have a dual effect on PCa proliferation. When present in abundance with WT-TP53, these isoforms inhibit PCa cell proliferation but sustain it at low levels, whether WT-TP53 is present. However, IL6ST-mediated PCa proliferation is independent of Δ133p53α and Δ133p53β, regardless of its interaction with these isoforms. These findings suggest a distinctive interplay between Δ133p53α and IL6ST, which provides valuable insights into the biology of PCa.