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July 11, 2018

Anti-Cancer Drug Effects Measured at Single-Cell Level by Multidimensional Technique

Multidimensional analysis of PARP toxicity using automated high-content microscopy. Various key parameters of cellular responses are investigated. [University of Zurich]

  • University of Zurich (UZH) scientists say they have developed a technique that makes it possible to quickly test various drugs at the cellular level. The method sheds light on the effects of anticancer drugs and the defense mechanisms of cancer cells, according to the researchers.

    The team uses cancer cell cultures to investigate the effects of PARP inhibitors, which make it difficult for these cells to replicate their DNA. They published their study (“Analysis of PARP inhibitor toxicity by multidimensional fluorescence microscopy reveals mechanisms of sensitivity and resistance”) in Nature Communications.

    “Exploiting the full potential of anticancer drugs necessitates a detailed understanding of their cytotoxic effects. While standard omics approaches are limited to cell population averages, emerging single cell techniques currently lack throughput and are not applicable for compound screens. Here, we employed a versatile and sensitive high-content microscopy-based approach to overcome these limitations and quantify multiple parameters of cytotoxicity at the single cell level and in a cell cycle resolved manner,” write the investigators.

    “Applied to PARP inhibitors (PARPi) this approach revealed an S-phase-specific DNA damage response after only 15 min, quantitatively differentiated responses to several clinically important PARPi, allowed for cell cycle resolved analyses of PARP trapping, and predicted conditions of PARPi hypersensitivity and resistance. The approach illuminates cellular mechanisms of drug synergism and, through a targeted multivariate screen, could identify a functional interaction between PARPi olaparib and NEDD8/SCF inhibition, which we show is dependent on PARP1 and linked to PARP1 trapping.”

    "Our method of fluorescence-based high-throughput microscopy allows us to observe precisely when and how a drug works in thousands of cells at the same time," explains Jone Michelena, Ph.D., postdoc researcher at UZH.

    Her measurements have revealed how PARP inhibitors lock their target protein in an inactive state on the cells' DNA and how this complicates DNA replication, which in turn leads to DNA damage. If this damage is not repaired quickly, the cells can no longer replicate and eventually die. 

    The new approach enables researchers to analyze the initial reaction of cancer cells to PARP inhibitors with great precision, notes Dr. Michelena, adding that what’s special about the procedure is the high number of individual cells that can be analyzed concurrently with high resolution using automated microscopes at the Center for Microscopy and Image Analysis of UZH. 

    Cancer cells vary and thus react differently to drugs depending on their mutations and the cell cycle phase they are in. The UZH researchers point out that they have now found a way to make these differences visible and quantify them precisely.

    Outside of the laboratory, the success of PARP inhibitors and other cancer medications is complicated by the fact that in some patients the cancer returns, having become resistant to the drugs. The high-throughput method employed by UZH researchers appears to be particularly useful for this kind of problem. Cells can be tested in multiple conditions with short turnover times, and specific genes can be eliminated one by one in a targeted manner. Doing so can reveal which cell functions are needed for a certain drug to take effect. 

    In addition, mechanisms of drug combinations can be analyzed in great detail. In her study, Dr. Michelena reports that ssuch a combination, which inhibits cancer cell proliferation to a significantly higher extent than the combination's individual components by themselves, has already been identified. 

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