Screening of PCNA ubiquitylation inhibitors as novel synthetic lethality inducers in Homologous-Recombination-deficient cancer cells

QUIROGA, RODRIGO; JACOBS, HEINZ; VILLARREAL, MARCOS; GARCÍA, IRIS ALEJANDRA; MANSILLA, SABRINA; BOCCO, JOSÉ LUIS; VILLAFAÑEZ, FLORENCIA; PANSA, MARÍA FLORENCIA; GOTTIFREDI, VANESA; SORIA, GASTÓN

Conferencia; EMBO Conference: THE DNA DAMAGE RESPONSE IN CELL PHYSIOLOGY AND DISEASE; 2019

EMBO

Translesion DNA Synthesis (TLS) and homologous recombination (HR) cooperate duringS-phase to safeguard replication forks integrity. Thus, the inhibition of TLS becomes apromising point of therapeutic intervention in HR-deficient cancers, where TLSimpairment might trigger synthetic lethality (SL). The main limitation to test thishypothesis is the current lack of selective pharmacological inhibitors of TLS. In this workwe show the development of two complementary screenings assays to identify smallmolecules that inhibit PCNA ubiquitylation, a key post-translational modificationrequired for efficient TLS activation. First, we setup a miniaturized WB assay with dualantibodies for ubi-PCNA and total PCNA coupled to infrared laser scanning. Second,we designed a virtual screening strategy to identify direct blockers of PCNA-ubiquitylation using molecular modelling. Herein, we present the identification of strongPCNA ubiquitylation inhibitors using both strategies. Moreover, using multipleexperimental models we performed proof-of-concept experiments showing that theabolishment of PCNA ubiquitylation triggers synthetic lethality in HR-deficient cellssubmitted to UV irradiation and Cisplatin treatment. Collectively, these findings set theground for developing a first-in-class type of therapeutic approach, where TLS inhibitioncan be achieved by the pharmacological inhibition of PCNA ubiquitylation. Given thevast incidence of HR deficiencies in multiple types of human cancers (i.e. Breast andOvarian cancer), the pharmacological inhibition of TLS could become a niche for drugdiscovery as an alternative targeted therapy to treat these malignancies.