PARP-1 has been linked to the progression of several types of cancer. We have recently reported that PARP-1 influences tumor progression in CRC through the regulation of CSCs in a p53-dependent manner. In this study, we propose that nitric oxide (NO) produced by inducible nitric oxide synthase (iNOS) could act as a mediator. We evaluated the expression of iNOS in a cohort of patients previously used to analyze the effects of PARP-1 on CRC in relation to p53 status. We also developed an in vitro model in which PARP-1 was stably overexpressed. In CRC patients, iNOS expression correlated with the differentiation grade, and with a high expression of CSC markers, although only in wild-type p53 tumors, as previously found for PARP-1. In vitro, overexpression of PARP-1 induced increased growth and stemness in wild-type p53 cells, while exerting the opposite effect on mutated ones, as expected. Treatment with 1400 W, a selective inhibitor of iNOS, or gene silencing of the gene counteracted the effects of PARP-1 in both p53 wild-type and p53 mutated cells. Given that the development of resistance has been demonstrated after treatment with PARP-1 inhibitors, iNOS could be considered a new therapeutic target in CRC, although only in patients with wild-type p53 tumors.

Background: Nuclear Receptor Subfamily 1 Group D Member 2 (NR1D2), a transcription factor that regulates the circadian clock, has been described as an oncogene in colorectal cancer (CRC). In several types of cancer, NR1D2 regulates cancer progression and relapse through cancer stem cells (CSCs), although this aspect has not been studied in CRC. On the other hand, p53 is a tumour suppressor gene that appears mutated in approximately a 50% CRCs. Interestingly, p53 is considered to be a crucial nexus between circadian clock deregulation and cancer. In addition, p53 regulates CSC phenotypes. Methods: We developed an in vitro model in which NR1D2 was silenced in three isogenic cell lines with different p53 status. In addition, we analysed the expression of NR1D2 in a cohort of patients and determined its relationship with the characteristics of patients and tumours. Results: In the in vitro model, NRID2 silencing reduces cell growth and decreases stemness, although only in cells harbouring a wild type p53. In contrast, in cells lacking a functional p53 or harbouring a mutated one, NR1D2 knockout increases cell growth and stemness. In patients, NR1D2 expression correlates with poorly differentiated tumours and high expression of CSCs markers, although only in tumours with a wild type p53, corroborating the results obtained in the in vitro model. Conclusions: Although more research is needed to analyse the mechanism by which NR1D2 regulates stemness in a p53-dependent manner, our results highlight the possibility of using NR1D2 antagonists for treating this type of patient and to develop personalised medicine.