ABSTRACT
Oxidative DNA damage is recognized by 8-oxoguanine (8-oxoG) DNA glycosylase 1 (OGG1), which excises 8-oxoG, leaving a substrate for apurinic endonuclease 1 (APE1) and initiating repair. Here, we describe a small molecule (TH10785) that interacts with the phenylalanine-319 and glycine-42 amino acids of OGG1, increases the enzyme activity 10-fold, and generates a previously undescribed ß,δ-lyase enzymatic function. TH10785 controls the catalytic activity mediated by a nitrogen base within its molecular structure. In cells, TH10785 increases OGG1 recruitment to and repair of oxidative DNA damage. This alters the repair process, which no longer requires APE1 but instead is dependent on polynucleotide kinase phosphatase (PNKP1) activity. The increased repair of oxidative DNA lesions with a small molecule may have therapeutic applications in various diseases and aging.
Subject(s)
DNA Damage , DNA Glycosylases , DNA Repair , Oxidative Stress , Biocatalysis/drug effects , DNA Damage/drug effects , DNA Glycosylases/chemistry , DNA Glycosylases/drug effects , DNA Repair/drug effects , Enzyme Activation , Glycine/chemistry , Humans , Ligands , Oxidative Stress/genetics , Phenylalanine/chemistry , Substrate SpecificityABSTRACT
Immune responses in human tissues rely on the concerted action of different cell types. Inter-cellular communication shapes both the function of the multicellular interaction networks and the fate of the individual cells that comprise them. With the advent of new methods to profile and experimentally perturb primary human tissues, we are now in a position to systematically identify and mechanistically dissect these cell-cell interactions and their modulators. Here, we introduce the concept of multicellular hubs, functional modules of immune responses in tissues. We outline a roadmap to discover multicellular hubs in human tissues and discuss how emerging technologies may further accelerate progress in this field.
Subject(s)
Cell Communication , Communication , HumansABSTRACT
Altered oncogene expression in cancer cells causes loss of redox homeostasis resulting in oxidative DNA damage, e.g. 8-oxoguanine (8-oxoG), repaired by base excision repair (BER). PARP1 coordinates BER and relies on the upstream 8-oxoguanine-DNA glycosylase (OGG1) to recognise and excise 8-oxoG. Here we hypothesize that OGG1 may represent an attractive target to exploit reactive oxygen species (ROS) elevation in cancer. Although OGG1 depletion is well tolerated in non-transformed cells, we report here that OGG1 depletion obstructs A3 T-cell lymphoblastic acute leukemia growth in vitro and in vivo, validating OGG1 as a potential anti-cancer target. In line with this hypothesis, we show that OGG1 inhibitors (OGG1i) target a wide range of cancer cells, with a favourable therapeutic index compared to non-transformed cells. Mechanistically, OGG1i and shRNA depletion cause S-phase DNA damage, replication stress and proliferation arrest or cell death, representing a novel mechanistic approach to target cancer. This study adds OGG1 to the list of BER factors, e.g. PARP1, as potential targets for cancer treatment.
Subject(s)
Colonic Neoplasms/drug therapy , DNA Glycosylases/genetics , DNA, Neoplasm/genetics , Gene Expression Regulation, Neoplastic , Poly (ADP-Ribose) Polymerase-1/immunology , Animals , Antineoplastic Agents/chemical synthesis , Antineoplastic Agents/pharmacology , Cell Line, Tumor , Cell Proliferation/drug effects , Colonic Neoplasms/genetics , Colonic Neoplasms/metabolism , Colonic Neoplasms/mortality , DNA Damage , DNA Glycosylases/antagonists & inhibitors , DNA Glycosylases/metabolism , DNA Repair/drug effects , DNA Replication/drug effects , DNA, Neoplasm/metabolism , Enzyme Inhibitors/chemical synthesis , Enzyme Inhibitors/pharmacology , Guanine/analogs & derivatives , Guanine/metabolism , HCT116 Cells , Humans , Mice , Mice, Nude , Molecular Targeted Therapy , Oxidative Stress , Poly (ADP-Ribose) Polymerase-1/metabolism , RNA, Small Interfering/genetics , RNA, Small Interfering/metabolism , Reactive Oxygen Species/antagonists & inhibitors , Reactive Oxygen Species/metabolism , Signal Transduction , Survival Analysis , Tumor Burden/drug effects , Xenograft Model Antitumor AssaysABSTRACT
Adoptive transfer of TCR transgenic T cells holds great promise for treating various cancers. So far, mainly semi-randomly integrating vectors have been used to genetically modify T cells. These carry the risk of insertional mutagenesis, and the sole addition of an exogenous TCR potentially results in the mispairing of TCR chains with endogenous ones. Established approaches using nonviral vectors, such as transposons, already reduce the risk of insertional mutagenesis but have not accomplished site-specific integration. Here, we used CRISPR-Cas9 RNPs and adeno-associated virus 6 for gene targeting to deliver an engineered TCR gene specifically to the TCR alpha constant locus, thus placing it under endogenous transcriptional control. Our data demonstrate that this approach replaces the endogenous TCR, functionally redirects the edited T cells' specificity in vitro, and facilitates potent tumor rejection in an in vivo xenograft model.
