Depletion of RAD17 sensitizes pancreatic cancer cells to gemcitabine

Abstract

SummaryChemotherapy of advanced pancreatic cancer has mainly been gemcitabine-based for the past 15 years, with only limited effect. Recently, combination therapy that also targets checkpoint kinase 1 (CHK1) has become an attractive option. The central role of CHK1 in many DNA-damage response pathways, however, may result in undesired cytotoxicity in normal cells, causing side effects. We were searching for other target molecules of similar function that may be more specific and thus better suited for combination therapy. To this end a negative selection RNAi screen was performed in cell lines with small hairpin RNA molecules targeting over 10,000 genes. Genes that were found to be synthetically lethal with gemcitabine and whose proteins act upstream of CHK1 were characterised in more detail. In particular, the inhibition of RAD17 potentiated gemcitabine cytotoxicity in the pancreatic cancer cell lines BxPC-3 and MiaPaca-2 and in the primary cell line JoPaca-1 that closely resembles primary tumour tissue. Further analysis showed that the synergistic effect of RAD17 knockdown and gemcitabine leads to forced mitotic entry of cells arrested in S phase by gemcitabine treatment, resulting in asymmetric DNA distribution during anaphase followed by DNA fragmentation and finally cell death by mitotic catastrophe. Our data suggest RAD17 as a novel target protein for gemcitabine combination therapy supplementing or complementing inhibition of CHK1. In contrast to CHK1, RAD17 knockdown by itself does not lead to abnormal DNA segregation, suggesting a more specific action.

A humanized yeast phenomic model of deoxycytidine kinase to predict genetic buffering of nucleoside analog cytotoxicity

“…5C; 1-0-14 ), representing a synthetic lethal relationship of potential therapeutic relevance. Consistent with this finding in yeast, depletion of hRAD17 can sensitize pancreatic cancer cells to gemcitabine [74].…”

Defining Mononuclear Phagocyte Subset Homology Across Several Distant Warm-Blooded Vertebrates Through Comparative Transcriptomics

“…As expected, this signature is associated to B lymphocyte ontogeny and functions [e.g., “development of B lymphocytes” p  = 10 −10.4 , “antibody response” ( p  = 10 −7.5 ), “proliferation of B lymphocytes” ( p  = 10 −10.5 ), and “morphology of B lymphocytes” ( p  = 10 −7.5 ) as well as to the “B cell receptor signaling” pathway ( p  = 10 −7.3 )] (Figure 11B). The B cell signature also pinpoints to genes without any known function in B cells yet, such as the cell cycle gene RAD17 (53) or the SP140 gene that encodes a nuclear body protein (54) (Table 1). Altogether, the results of the functional analysis of the conserved signature of B cells support the biological relevance of the conserved gene signatures generated by our approach.…”

The noncoding function of NELFA mRNA promotes the development of oesophageal squamous cell carcinoma by regulating the Rad17‐RFC2‐5 complex

“…Previous studies have reported that Rad17 expression is elevated in colon carcinoma, breast cancer, pancreatic cancer, gastric cancer and non‐small‐cell lung cancer and is correlated with tumour cell onset and progression (Bao et al , ; Fredebohm et al , ; Wang et al , ). NELFA is an essential element of the NELF complex that negatively regulates the elongation of RNA polymerase II‐mediated transcription (Gilchrist et al , ; Jadlowsky et al , ; Stadelmayer et al , ).…”