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Discovery of Small Molecules That Induce Lysosomal Cell Death in Cancer Cell Lines Using an Image-Based Screening Platform

Pagliero, Romina J. ; D'Astolfo, Diego S. ; Lelieveld, Daphne ; Pratiwi, Riyona D. ; Aits, Sonja LU orcid ; Jaattela, Marja ; Martin, Nathaniel I. ; Klumperman, Judith and Egan, David A. (2016) In Assay and Drug Development Technologies 14(8). p.489-510
Abstract

The lysosomal cell death (LCD) pathway is a caspase 3-independent cell death pathway that has been suggested as a possible target for cancer therapy, making the development of sensitive and specific high-throughput (HT) assays to identify LCD inducers highly desirable. In this study, we report a two-step HT screening platform to reliably identify such molecules. First, using a robust HT primary screen based on propidium iodide uptake,we identified compounds that kill through nonapoptotic pathways. A phenotypic image-based assay using a galectin-3 (Gal-3) reporterwas then used to further classify hits based on lysosomal permeabilization, a hallmark of LCD. The identification of permeabilized lysosomes in our image-based assay is not... (More)

The lysosomal cell death (LCD) pathway is a caspase 3-independent cell death pathway that has been suggested as a possible target for cancer therapy, making the development of sensitive and specific high-throughput (HT) assays to identify LCD inducers highly desirable. In this study, we report a two-step HT screening platform to reliably identify such molecules. First, using a robust HT primary screen based on propidium iodide uptake,we identified compounds that kill through nonapoptotic pathways. A phenotypic image-based assay using a galectin-3 (Gal-3) reporterwas then used to further classify hits based on lysosomal permeabilization, a hallmark of LCD. The identification of permeabilized lysosomes in our image-based assay is not affected by changes in the lysosomal pH, thus resolving an important limitation in currently usedmethods.We have validated our platform in a screen by identifying 24 LCD inducers, some previously known to induce LCD. Although most LCD inducers were cationic amphiphilic drugs (CADs), we have also identified a non-CAD LCD inducer, which is of great interest in the field. Our data also gave new insights into the biology of LCD, suggesting that lysosomal accumulation and acid sphingomyelinase inhibition are not sufficient or necessary for the induction of LCD. Overall, our results demonstrate a robust HT platform to identify novel LCD inducers that will also be very useful for gaining deeper insights into the molecular mechanism of LCD induction.

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author
; ; ; ; ; ; ; and
publishing date
type
Contribution to journal
publication status
published
subject
keywords
galectin-3 reporter, high throughput screening, LMP phenotypic assay, lysosomal cell dead
in
Assay and Drug Development Technologies
volume
14
issue
8
pages
489 - 510
publisher
Mary Ann Liebert, Inc.
external identifiers
  • pmid:27732064
  • scopus:84991738403
ISSN
1540-658X
DOI
10.1089/adt.2016.727
language
English
LU publication?
no
id
2a787fee-0896-4a33-a1ce-0b4bca8fc0bd
date added to LUP
2018-03-05 15:20:06
date last changed
2025-04-30 20:04:18
@article{2a787fee-0896-4a33-a1ce-0b4bca8fc0bd,
  abstract     = {{<p>The lysosomal cell death (LCD) pathway is a caspase 3-independent cell death pathway that has been suggested as a possible target for cancer therapy, making the development of sensitive and specific high-throughput (HT) assays to identify LCD inducers highly desirable. In this study, we report a two-step HT screening platform to reliably identify such molecules. First, using a robust HT primary screen based on propidium iodide uptake,we identified compounds that kill through nonapoptotic pathways. A phenotypic image-based assay using a galectin-3 (Gal-3) reporterwas then used to further classify hits based on lysosomal permeabilization, a hallmark of LCD. The identification of permeabilized lysosomes in our image-based assay is not affected by changes in the lysosomal pH, thus resolving an important limitation in currently usedmethods.We have validated our platform in a screen by identifying 24 LCD inducers, some previously known to induce LCD. Although most LCD inducers were cationic amphiphilic drugs (CADs), we have also identified a non-CAD LCD inducer, which is of great interest in the field. Our data also gave new insights into the biology of LCD, suggesting that lysosomal accumulation and acid sphingomyelinase inhibition are not sufficient or necessary for the induction of LCD. Overall, our results demonstrate a robust HT platform to identify novel LCD inducers that will also be very useful for gaining deeper insights into the molecular mechanism of LCD induction.</p>}},
  author       = {{Pagliero, Romina J. and D'Astolfo, Diego S. and Lelieveld, Daphne and Pratiwi, Riyona D. and Aits, Sonja and Jaattela, Marja and Martin, Nathaniel I. and Klumperman, Judith and Egan, David A.}},
  issn         = {{1540-658X}},
  keywords     = {{galectin-3 reporter; high throughput screening; LMP phenotypic assay; lysosomal cell dead}},
  language     = {{eng}},
  month        = {{10}},
  number       = {{8}},
  pages        = {{489--510}},
  publisher    = {{Mary Ann Liebert, Inc.}},
  series       = {{Assay and Drug Development Technologies}},
  title        = {{Discovery of Small Molecules That Induce Lysosomal Cell Death in Cancer Cell Lines Using an Image-Based Screening Platform}},
  url          = {{http://dx.doi.org/10.1089/adt.2016.727}},
  doi          = {{10.1089/adt.2016.727}},
  volume       = {{14}},
  year         = {{2016}},
}