The tumor suppressor protein p53 has an important role in cell-fate determination. In cancer cells, the activity of p53 is frequently repressed by high levels of MDMX and/or MDM2. MDM2 is a... Show moreThe tumor suppressor protein p53 has an important role in cell-fate determination. In cancer cells, the activity of p53 is frequently repressed by high levels of MDMX and/or MDM2. MDM2 is a ubiquitin ligase whose activity results in ubiquitin- and proteasome-dependent p53 degradation, while MDMX inhibits p53-activated transcription by shielding the p53 transactivation domain. Interestingly, the oncogenic functions of MDMX appear to be more wide-spread than inhibition of p53. The present study aimed to elucidate the MDMX-controlled transcriptome. Therefore, we depleted MDMX with four distinct shRNAs from a high MDMX expressing uveal melanoma cell line and determined the effect on the transcriptome by RNAseq. Biological function analyses indicate the inhibition of the cell cycle regulatory genes and stimulation of cell death activating genes upon MDMX depletion. Although the inhibition of p53 activity clearly contributes to the transcription regulation controlled by MDMX, it appeared that the transcriptional regulation of multiple genes did not only rely on p53 expression. Analysis of gene regulatory networks indicated a role for Forkhead box (FOX) transcription factors. Depletion of FOXO proteins partly prevented the transcriptional changes upon MDMX depletion. Furthermore, depletion of FOXO proteins relatively diminished the growth inhibition upon MDMX knockdown, although the knockdown of the FOXO transcription factors also reduces cell growth. In conclusion, the p53-independent oncogenic functions of MDMX could be partially explained by its regulation of FOXO activity. Show less
Simple Summary Prostate cancer is the most prevalent male cancer. It poses a survival risk if it spreads and fails treatment. In search of fresh insight into lethal prostate cancers, we examined... Show moreSimple Summary Prostate cancer is the most prevalent male cancer. It poses a survival risk if it spreads and fails treatment. In search of fresh insight into lethal prostate cancers, we examined failures in cancer defence systems operated by the key anticancer protein p53. Normally, levels of p53 activity are kept low by the protein MDM4, and consistently, we found that high MDM4 levels pose a prostate cancer risk in this context. Outside this explanation, we discovered that high MDM4 levels are also a cancer risk when p53 is genetically altered and unable to fight cancer, or even mutated to drive cancer spread. Our novel findings uncovered MDM4 inhibition as a new concept for prostate cancer treatment, and we demonstrated efficacy and uncovered mechanisms of action. Importantly, we showed that targeting MDM4 halted the growth of aggressive prostate cancer cells with mutant p53, and this was potentiated by a drug clinically trialled to target mutant p53 cancers. Metastatic prostate cancer is a lethal disease in patients incapable of responding to therapeutic interventions. Invasive prostate cancer spread is caused by failure of the normal anti-cancer defense systems that are controlled by the tumour suppressor protein, p53. Upon mutation, p53 malfunctions. Therapeutic strategies to directly re-empower the growth-restrictive capacities of p53 in cancers have largely been unsuccessful, frequently because of a failure to discriminate responses in diseased and healthy tissues. Our studies sought alternative prostate cancer drivers, intending to uncover new treatment targets. We discovered the oncogenic potency of MDM4 in prostate cancer cells, both in the presence and absence of p53 and also its mutation. We uncovered that sustained depletion of MDM4 is growth inhibitory in prostate cancer cells, involving either apoptosis or senescence, depending on the cell and genetic context. We identified that the potency of MDM4 targeting could be potentiated in prostate cancers with mutant p53 through the addition of a first-in-class small molecule drug that was selected as a p53 reactivator and has the capacity to elevate oxidative stress in cancer cells to drive their death. Show less
The focus of this thesis is uveal melanoma (UM) which, once metastasized, is lethal due to lack of effective treatment options. To repress p53 activity approximately 65% of UM tumors express high... Show moreThe focus of this thesis is uveal melanoma (UM) which, once metastasized, is lethal due to lack of effective treatment options. To repress p53 activity approximately 65% of UM tumors express high levels of the p53 inhibitory proteins MDMX or MDM2. The aim of this thesis is to unravel the oncogenic function of MDMX and provide new treatment options for patients with metastasized UM. Chapter 2 describes the regulation of the transcriptome by MDMX in UM and proposes novel p53-independent effects of MDMX, i.e. FOXO inhibition. In chapter 3 the opportunities of a combined targeting of two common signaling pathways as therapeutic intervention for metastasized UM patients is investigated. Genetic interference with either MDMX or PKC δ expression or activity showed that beneficial effects can already be achieved by a more specific targeting, which is presumable less toxic to the patient. In chapter 4 it is described, opposed to what has been reported before, that enhancer of zeste homolog 2 (EZH2) inhibition poses a valuable novel therapeutic invention for UM. In chapter 5 it is shown that combining two clinically approved drugs, Quisinostat and Flavopiridol, could serve as an effective therapeutic intervention for UM patients. Show less
