AUTHOR=Niu Ya , Fan Xinyi , Wang Yaping , Lin Jiaxin , Hua Luchun , Li Xiaobo , Qian Ruizhe , Lu Chao 

TITLE=Genome-wide CRISPR Screening Reveals Pyrimidine Metabolic Reprogramming in 5-FU Chronochemotherapy of Colorectal Cancer

JOURNAL=Frontiers in Oncology

VOLUME=12

YEAR=2022

URL=https://www.frontiersin.org/journals/oncology/articles/10.3389/fonc.2022.949715

DOI=10.3389/fonc.2022.949715

ISSN=2234-943X

ABSTRACT=<sec><title>Objective</title><p>Disruption of the circadian rhythm is associated with cancer occurrence, response to chemotherapy, and poor prognosis. Thus, using internal clock-based chronotherapy to optimize the administration time may improve the therapeutic effects of anticancer drugs while reducing the side effects. Chronotherapy with 5-fluorouracil (5-FU) has been observed in colorectal cancer (CRC) for a long time, but its effect is under controversial and the mechanism remains unclear.</p></sec><sec><title>Methods</title><p>Genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) screening and RNA-sequencing were combined to identify the potential genes or pathways involved in 5-FU chronochemotherapy. Genetic deletion or overexpression of pyrimidine metabolic pathway genes were conducted to examine cellular viability with or without 5-FU <italic>via</italic> flow cytometry. Western blotting, qPCR, chromatin immunoprecipitation, gain-of-function and loss-of-function assays of several CRC cell lines <italic>in vitro</italic> and <italic>in vivo</italic> were used to elaborate and validate the mechanism of 5-FU chronotherapeutic effects.</p></sec><sec><title>Results</title><p>Chronochemotherapeutic effects of 5-FU on CRC <italic>in vivo</italic> were verified. Furthermore, 5-FU chronochemotherapy related genes such as <italic>UPP2</italic>, <italic>UCK2</italic> and <italic>UMPS</italic> in the pyrimidine metabolic pathway were identified. Disturbance in these genes, especially <italic>UMPS</italic>, perturbs 5-FU treatment outcomes in CRC cells. Mechanistically, the core circadian gene, brain and muscle aryl hydrocarbon receptor nuclear translocator-like protein-1 (BMAL1), extensively regulate gene expression in pyrimidine metabolic pathway by binding to E-box element in the promoter region of key genes such as <italic>UMPS</italic> and perturb their enzymatic activities, thereby maintain diurnal efficacy of 5-FU in CRC cells.</p></sec><sec><title>Conclusion</title><p>This study uncovered a new mechanism by which a core circadian gene BMAL1 increases the effectiveness of 5-FU by enhancing the expression and enzymatic activities of key genes in the pyrimidine metabolic pathway in CRC cells. The findings suggest a novel strategy for CRC chemotherapy by targeting chrono-modulated genes of the 5-FU metabolic pathway.</p></sec>