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Molecular mechanisms of induced protection against sepsis by DNA damage responses

Periodic Reporting for period 4 - iPROTECTION (Molecular mechanisms of induced protection against sepsis by DNA damage responses)

Reporting period: 2020-04-01 to 2021-03-31

Our laboratory investigates the mechanisms used to restore homeostasis when an organism responds to different types of stress. We focus on the events triggered by core cellular function perturbations, including DNA damage caused by drugs used in chemotherapy to treat cancer. We have recently found that when used at low doses, anthracyclines can have a surprising but strong protective effect against systemic infections (sepsis) and other conditions that cause substantial tissue damage and organ dysfunction. In addition to its biological interest and significance, our work is also important because there are practical implications for our findings, including for the treatment of sepsis (that has no specific treatment so far and is responsible for over 11 million deaths worldwide every year). The central goal of the current proposal is to identify and characterize novel cytoprotective mechanisms, with a focus on DNA damage response dependent protection activated by anthracyclines (the most commonly used chemotherapeutic drugs to treat cancer) as a window into stress-induced genetic programs conferring disease tolerance.
We found important new cues into the molecular mechanisms of disease tolerance induction by DNA damage responses. Our main results relate to the precise identification of the molecular mechanism by which anthracyclines specifically block the transcription of NFkB targets, when used at concentration below cytotoxic levels. In the course of this investigation, we also found that low levels of DNA damage promote survival against TNF-induced cell death, in an NFkB-independent manner. Anthracyclines are among the most used and effective anticancer drugs. Their activity has been attributed to DNA double-strand breaks resulting from topoisomerase II poisoning and to eviction of histones from select sites in the genome. We discovered that the extensively used anthracyclines Doxorubicin, Daunorubicin and Epirubicin, decrease the transcription of nuclear factor kappa B (NF-κB)-dependent gene targets, but not interferon responsive genes. Using an NMR-based structural approach, we demonstrated that anthracyclines disturb the complexes formed between the NF-B subunit RelA and its DNA binding sites. The variant anthracyclines Aclarubicin, Doxorubicinone and the newly developed Dimethyl-doxorubicin, which share anticancer properties with the other anthracyclines but do not induce DNA damage, also suppressed inflammation, thus uncoupling DNA damage from the effects on inflammation. This has implications for anticancer therapy and for the development of novel anti-inflammatory drugs with limited side effects for life-threatening conditions such as sepsis. In addition to DNA damage, we explored the effects on the progress of a severe infection of protein synthesis inhibition in the mitochondria, which more that 50% of all classes of antibiotics in use cause. Several classes of antibiotics have long been known to have beneficial effects that cannot be explained strictly on the basis of their capacity to control the infectious agent. We discovered that tetracycline antibiotics, which target the mitoribosome, protected against sepsis, without affecting the pathogen load. Mechanistically, we found that mitochondrial inhibition of protein synthesis perturbed the electron transport chain (ETC) decreasing tissue damage in the lung and increasing fatty acid oxidation and glucocorticoid sensitivity in the liver. Using a liver-specific partial and acute deletion of Crif1, a critical mitoribosomal component for protein synthesis, we found that mice were protected against sepsis, an observation which was phenocopied by the transient inhibition of complex I of the ETC by phenformin. Together, we demonstrate that mitoribosome-targeting antibiotics are beneficial beyond their antibacterial activity and that mitochondrial protein synthesis inhibition leading to ETC perturbation is a mechanism for the induction of disease tolerance. These results have been published in scientific journals and disseminated to the public through traditional media and social networks. They will also serve as the bases for one clinical trial (anthracyclines, currently in its early steps) and another one at the planing stage (tetracyclines).
Our observation that low level DNA damage caused by anthracyclines is able to prevent TNF-induced cell death in an NFkB-independent manner is completely unexpected and substantially goes beyond the state of the art. This is especially surprising because in the same conditions these drugs inhibit NF-kB, contrary to the dogma in the field that accepts that these drugs activate NFkB, because they are normally used at very high doses as the goal is to kill cancer cells. Our discovery has critical importance in the field of cancer treatment as it uncovers a possible key mechanism to explain the emergence of resistance against chemotherapy, while pointing into novel strategies to develop more effective therapies against cancer. In fact, even when used at high doses to treat cancer, if the tumor is large, or it is located in a tissue or organ where doses do not reach the necessary levels, a substantial number of cells will be exposed to sub-optimal doses, in which case these drugs will promote cell survival and not cell death as intended. This is especially dangerous as it probably happens in the setting of DNA damage which adds cell survival to induced mutations, leading not only to cancer resistance to chemotherapy but also to the emergence of more aggressive forms of the tumor. If confirmed, the impact is therefore very high as chemotherapy resistance is a major problem in the treatment of cancer. Our discovery opens the possibility of rational design of new and more effective combinatorial therapies in cancer. In addition, from the opposite perspective, can also be explored to increase tissue regeneration and survival in conditions that cause substantial tissue damage including ischemia / reperfusion diseases.
DNA damage protection in sepsis