Dataset Information

Resistance to ATR Inhibitors Mediated by Loss of the Nonsense-Mediated Decay Factor UPF2

ABSTRACT: Over one million cases of gastric cancer are diagnosed each year and metastatic disease continues to have a poor prognosis. A significant proportion of gastric tumors have defects in the DNA damage response pathway creating therapeutic opportunities through synthetic lethal approaches. In particular, several small molecule inhibitors of Ataxia-Telangiectasia Mutated and Rad3-related protein kinase (ATR), a key regulator of the DNA Damage response, are now in clinical development as targeted agents for gastric cancer. Here, we sought to discover genetic determinants of response and resistance to ATRi in gastric cancer cells. We performed a large-scale CRISPR interference screen to discover determinants of resistance to ATRi in gastric cancer. Top hits were validated and further evaluated to define mechanisms of resistance. We identified components of the nonsense-mediated decay pathway and the UPF2 gene, in particular, as mediators of sensitivity to ATRi. We show loss of UPF2 causes resistance to ATRi through the abrogation of ATRi mediated cell cycle arrest and find evidence that UPF2 regulates R-loops. Our results uncover a novel role for NMD factors in R-loop formation and resistance to ATRi in gastric cancer cells, potentially through R-loop formation.

INSTRUMENT(S): timsTOF Pro

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture

SUBMITTER: Minkyu Kim

LAB HEAD: Nevan Krogan

PROVIDER: PXD034887 | Pride | 2022-11-06

REPOSITORIES: Pride

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Dataset's files

Source:

			Action	DRS
	AA6-01-ab_C3_1_5110.d.zip	Other
	AA6-01-ph_A1_1_5086.d.zip	Other
	AA6-02-ab_C4_1_5111.d.zip	Other
	AA6-02-ph_A2_1_5087.d.zip	Other
	AA6-03-ab_C5_1_5112.d.zip	Other

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Publications

Resistance to ATR Inhibitors Is Mediated by Loss of the Nonsense-Mediated Decay Factor UPF2.

O'Leary Patrick C PC Chen Huadong H Doruk Yagmur U YU Williamson Tess T Polacco Benjamin B McNeal Andrew S AS Shenoy Tanushree T Kale Nupura N Carnevale Julia J Stevenson Erica E Quigley David A DA Chou Jonathan J Feng Felix Y FY Swaney Danielle L DL Krogan Nevan J NJ Kim Minkyu M Diolaiti Morgan E ME Ashworth Alan A

Cancer research 20221101 21

Over one million cases of gastric cancer are diagnosed each year globally, and the metastatic disease continues to have a poor prognosis. A significant proportion of gastric tumors have defects in the DNA damage response pathway, creating therapeutic opportunities through synthetic lethal approaches. Several small-molecule inhibitors of ATR, a key regulator of the DNA damage response, are now in clinical development as targeted agents for gastric cancer. Here, we performed a large-scale CRISPR i ...[more]

PMID: 36273492

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Project description:Purpose: Gastric cancer remains one of the deadliest neoplasms worldwide, with a high need for new therapeutic options. Efficacies of targeted therapies are often limited owing to the inter- and intratumoral heterogeneity of gastric cancer. Thus, drugs with broader mechanisms of action rather than relying on the inhibition of a single specific oncogene are needed. Preclinical studies have identified histone deacetylases (HDAC) and their respective isoforms as potential therapeutic targets in gastric cancer. However, clinical efficacies are moderate and the mechanism(s) of action of HDAC inhibitors (HDACi) are only partially understood. Methods: In a panel of gastric cancer cell lines with different molecular characteristics, tumor cell inhibitory effects of different HDACi were studied. Lipid peroxidation levels were measured using flow cytometry. Proteome analysis was performed for the in-depth characterization of molecular alterations upon HDAC inhibition. HDACi effects on important ferroptosis genes were validated on the mRNA (RT-qPCR) and protein level (Western Blot). Results: Upon HDACi treatment, lipid peroxidation levels were found increased in all tested cell lines. Class-I HDACi (VK1, entinostat) showed the same toxicity profile as the pan-HDACi vorinostat. Proteome analysis revealed significant and concordant alterations in the expression of proteins related to ferroptosis induction. Key enzymes like ACSL4, POR or SLC7A11 showed distinct alterations in their expression patterns, providing an explanation for the increased lipid peroxidation. Alterations of POR and SLC7A11 levels upon treatment were also confirmed in primary human gastric cancer tissue cultures as relevant ex vivo model. Conclusion: We identify the induction of ferroptosis as a new mechanism of action of class-I HDACi in gastric cancer. Notably, these findings were independent of the genetic background of the cell lines, thus introducing HDAC inhibition as a more general therapeutic principle besides other, less broadly usable targeted drugs.