ABSTRACT
Metformin and IACS-010759 are two distinct antimetabolic agents. Metformin, an established antidiabetic drug, mildly inhibits mitochondrial complex I, while IACS-010759 is a new potent mitochondrial complex I inhibitor. Mitochondria is pivotal in the energy metabolism of cells by providing adenosine triphosphate through oxidative phosphorylation (OXPHOS). Hence, mitochondrial metabolism and OXPHOS become a vulnerability when targeted in cancer cells. Both drugs have promising antitumoral effects in diverse cancers, supported by preclinical in vitro and in vivo studies. We present evidence of their direct impact on cancer cells and their immunomodulatory effects. In clinical studies, while observational epidemiologic studies on metformin were encouraging, actual trial results were not as expected. However, IACS-01075 exhibited major adverse effects, thereby causing a metabolic shift to glycolysis and elevated lactic acid concentrations. Therefore, the future outlook for these two drugs depends on preventive clinical trials for metformin and investigations into the plausible toxic effects on normal cells for IACS-01075.
ABSTRACT
Genome engineering has become more accessible thanks to the CRISPR-Cas9 gene-editing system. However, using this technology in synthetic organs called "organoids" is still very inefficient. This is due to the delivery methods for the CRISPR-Cas9 machinery, which include electroporation of CRISPR-Cas9 DNA, mRNA, or ribonucleoproteins containing the Cas9-gRNA complex. However, these procedures are quite toxic for the organoids. Here, we describe the use of the "nanoblade (NB)" technology, which outperformed by far gene-editing levels achieved to date for murine- and human tissue-derived organoids. We reached up to 75% of reporter gene knockout in organoids after treatment with NBs. Indeed, high-level NB-mediated knockout for the androgen receptor encoding gene and the cystic fibrosis transmembrane conductance regulator gene was achieved with single gRNA or dual gRNA containing NBs in murine prostate and colon organoids. Likewise, NBs achieved 20%-50% gene editing in human organoids. Most importantly, in contrast to other gene-editing methods, this was obtained without toxicity for the organoids. Only 4 weeks are required to obtain stable gene knockout in organoids and NBs simplify and allow rapid genome editing in organoids with little to no side effects including unwanted insertion/deletions in off-target sites thanks to transient Cas9/RNP expression.
ABSTRACT
During replication, the presence of unrepaired lesions results in the formation of single stranded DNA (ssDNA) gaps that need to be repaired to preserve genome integrity and cell survival. All organisms have evolved two major lesion tolerance pathways to continue replication: Translesion Synthesis (TLS), potentially mutagenic, and Homology Directed Gap Repair (HDGR), that relies on homologous recombination. In Escherichia coli, the RecF pathway repairs such ssDNA gaps by processing them to produce a recombinogenic RecA nucleofilament during the presynaptic phase. In this study, we show that the presynaptic phase is crucial for modulating lesion tolerance pathways since the competition between TLS and HDGR occurs at this stage. Impairing either the extension of the ssDNA gap (mediated by the nuclease RecJ and the helicase RecQ) or the loading of RecA (mediated by RecFOR) leads to a decrease in HDGR and a concomitant increase in TLS. Hence, we conclude that defects in the presynaptic phase delay the formation of the D-loop and increase the time window allowed for TLS. In contrast, we show that a defect in the postsynaptic phase that impairs HDGR does not lead to an increase in TLS. Unexpectedly, we also reveal a strong genetic interaction between recF and recJ genes, that results in a recA deficient-like phenotype in which HDGR is almost completely abolished.
