MOLECULAR DISECTION OF BIOLOGICALLY RELEVANT PATHWAYS AFTER INHIBITION OF CHECKPOINT KINASE 1

NICOLÁS L. CALZETTA; VANESA GOTTIFREDI; NATALIA PAVIOLO; MARINA A. GONZÁLEZ BESTEIRO

Ciudad Autónoma de Buenos Aires

Congreso; Reunión Biociencias 2017; 2017

Checkpoint signalling allows tumour cells to cope with high rates of replication stress, a hallmark of cancer. Specifically, Chk1 delays the progression of S phase, thereby increasing the chances to repair damaged DNA before using it as a replication template. Therefore, the inhibition of checkpoint kinase 1 (Chk1) has emerged as a promising anti-cancer therapy. However, in the clinic, treatments with Chk1 inhibitors fail with unexpected frequency, highlighting the need to further understand how cells adapt to such therapy. It is very well established that Chk1 inhibition alters the replication choreography, increasing origin firing while slowing down replication fork progression. In turn, such defective replication choreography generates increased levels of double strand break (DSB), compromising the genomic stability of cells to a degree that triggers cell death. The current model proposes that such events are part of a linear pathway, so that cell death is intimately linked to genomic instability in Chk1-inhibited cells. However, the molecular signals that link those phenotypes are unknown. We have undertaken a systematic analysis involving the elimination of factors which are crucial for each of the processes modulated by Chk1 inhibition. In such experimental settings, we have examined the replication choreography (DNA fiber assay which allows to measure fork rate and percentage of origin firing). We have also quantified DSB accumulation using neutral COMET assay and genomic instability using the micronuclei assay. We will present preliminary data demonstrating that the phenotypes caused by Chk1 deficiency, i.e. altered replication choreography, genomic instability, and cell death can be dissected at the molecular levels. We believe that such information could be used to improve anti-cancer therapies based on Chk1 inhibition.