title: EVALUATION OF UHRF1 AND THE ANTIOXIDANT RESPONSE PATHWAY IN PANCREATIC CANCER AND THE PREDICTION OF RESPONSE TO TREATMENT
Creators: name: Gana, T
Publisher: name: University of Liverpool Repository
Year: 2017

description: Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers, characterised by very poor survival. The cellular defence protein Nrf2 is a mediator of oncogenesis in pancreatic cancer, although its role in this cancer type is not fully understood. UHRF1 is a nuclear protein involved in epigenetic regulation. Our laboratory recently established that UHRF1 suppresses Keap1, an important negative regulator of the Nrf2 pathway. The aim of this study was to explore how UHRF1 contributes to Nrf2 function in pancreatic cancer cells and in pancreatic stellate cells, and to determine whether UHRF1 or NQO1, a downstream marker of Nrf2 activity could be predictive of PDAC treatment response. Depletion of UHRF1 from PDAC cells was associated with transcriptional upregulation of KEAP1, decreased levels of Nrf2 and Nrf2-regulated downstream proteins (NQO1 and AKR1C1), increased oxidative stress in the form of lower glutathione levels and increased reactive oxygen species. UHRF1 depletion enhanced cell cycle arrest, and concomitant depletion of UHRF1 and Keap1 reversed this effect, restored Nrf2 levels and reversed the increase in reactive oxygen species. The expression of UHRF1 in human pancreatic stellate cells (hPSCs) was limited to a small subset of cells. Analysis in Nrf2-null mouse pancreatic stellate cells was precluded by the fact that these cells, unlike their wild-type counterparts could not be maintained in culture. Histological analysis of the pancreata of Nrf2-null mice revealed early signs of pancreatitis. Germline single nucleotide polymorphisms (SNPs) were analysed in patients with advanced PDAC (n=140) from TeloVac and ViP clinical trials. The NQO1 rs1800566 CC homozygous major genotype was associated with poor survival (p=0.01) and the combined NQO1 rs1800566 and SRXN1 rs6053666 SNPs were prognostic of survival in these patients (p=0.039). No SNP–related differences in survival for NRF2 rs2886162 and SRXN1 rs6053666 variants were observed (p=0.405 and p=0.634 respectively). To determine whether UHRF1 or NQO1 expression are predictive of PDAC patient response to treatment, tissue microarrays from 349 patients randomized to chemotherapy in the ESPAC-3 trial (plus controls from ESPAC-1) were analysed by immunohistochemistry for these proteins. Patients were dichotomized into low and high expression for nuclear UHRF1 and cytoplasmic NQO1 levels respectively and groups compared using Kaplan-Meier curves, log-rank tests, and Cox proportional hazards models. UHRF1 was expressed in 181 of 199 (90.9%) PDAC tumours. Its levels were not associated with survival in the observation only group, or in either 5FU/folinic acid or gemcitabine treated groups (p=0.882, p=0.34 and p=0.92 respectively). Interestingly, an inverse relationship between UHRF1 and diabetes was apparent (p=0.005). NQO1 was expressed in the cytoplasm of 197 (95.2%) PDAC tumours. NQO1 levels were not associated with survival for the patents in the observation only group (p=0.301). However, median survival for patients treated with gemcitabine was 13.7 (95% CI 10.2 to 16.8) months for those with low NQO1 expression versus 24.2 (95% CI 16.3 to 29.5) months for those with high NQO1 expression (p=0.01). For the 5FU/folinic acid group, median survival was 13.4 (95% CI 10.9 to 19.6) and 21.8 (95 % CI 16.9 to 25.7) months for those with low and high NQO1 levels respectively (p=0.08). Multivariable Cox proportional hazard analysis in gemcitabine treated patients, revealed that NQO1 expression was not an independent predictive factor in gemcitabine-treated (p=0.19) patients. In summary, UHRF1 regulates Nrf2 in PDAC. UHRF1 levels are not prognostic, nor are they predictive of response to treatment in this disease. Interestingly, UHRF1 levels are inversely associated with diabetes, a finding that merits further study. By contrast, the NQO1 SNP rs1800566 is prognostic in advanced PDAC patients and NQO1 protein expression is not an independent predictive marker of response to adjuvant gemcitabine treatment in PDAC patients.