Molecular Oncology Programme

Genomic Instability Group

Research highlights
Efficacy of ATR inhibition in two preclinical models of cancer

Replicative stress (RS) is a widespread phenomenon in cancer cells that, when persistent, leads to DNA double strand breaks and genomic instability. Besides from the basal level of RS that occurs in every cell division, the presence of oncogenes, or many of the agents used in chemotherapy, are potent inducers of RS. In mammals, RS is sensed and suppressed through a signalling cascade that is initiated with the activation of the ATR kinase. We previously hypothesised that due to the high levels of RS in certain cancers, they could be particularly dependent on a proficient RS-response. In this regard, and in collaboration with the Experimental Therapeutics Programme, we had developed chemical inhibitors of ATR that presented some anti-tumour properties in vitro. During 2016, our work in this area was focused on the identification of tumours that are particularly sensitive to ATR inhibition, as well as on the discovery of mechanisms of resistance to these chemicals. For the first area of focus, we have shown efficacy of ATR inhibitors, as monotherapy, in 2 mouse models of Ewing Sarcoma and Acute Myeloid Leukaemia (FIGURE 1). Regarding the second area of focus, we discovered – via genomewide CRISPR-Cas9 screening – that the levels of CDC25A, a key phosphatase controlling mitotic entry, are a key determinant of the sensitivity to ATR inhibitors in mouse and human cells.

Two new players that suppress replication stress in mammalian cells

Besides ATR, several other factors participate in limiting the impact of RS in mammalian cells. Recent works have identified that POLD3, a subunit of the DNA polymerase Pold, participates in the repair of the breaks generated by RS, and also suggest that limiting its activity could be specifically deleterious for cancer cells. By developing a novel conditional knockout mouse strain we found that POLD3 deletion is lethal during embryonic development and also when depleted in adult mice. These severe defects were explained by a complete destabilisation of the POLd complex in the absence of POLD3, which abrogates DNA replication, raising serious doubts regarding the potential of POLD3 as an anticancer target. In independent work, we have been investigating how SUMO and ubiquitin participate in the coordination of DNA replication. Here, we identified USP7 as the first chromatin-associated SUMO deubiquitinase (SDUB) and revealed its essential role during DNA replication. Accordingly, USP7 inhibitors lead to RS and DNA damage. By deubiquitinating SUMO and/or SUMOylated proteins, our research revealed that USP7 contributes to keep a SUMO-rich and ubiquitin-poor environment at sites of DNA replication; this is critical to maintain fork progression. Our current view is that USP7 is critical for controlling the traffic of replication factors, by supervising their recruitment or expulsion from replication factories. We propose that SUMOylation constitutes a ‘stay’ signal that recruits proteins near replication factors, with Ubiquitinated-SUMO being the ‘go’ signal that leads to their eviction (FIGURE 2).