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Selective Targeting of c-Abl via a Cryptic Mitochondrial Targeting Signal Activated by Cellular Redox Status in Leukemic and Breast Cancer Cells

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Abstract

Purpose

The tyrosine kinase c-Abl localizes to the mitochondria under cell stress conditions and promotes apoptosis. However, c-Abl has not been directly targeted to the mitochondria. Fusing c-Abl to a mitochondrial translocation signal (MTS) that is activated by reactive oxygen species (ROS) will selectively target the mitochondria of cancer cells exhibiting an elevated ROS phenotype. Mitochondrially targeted c-Abl will thereby induce malignant cell death.

Methods

Confocal microscopy was used to determine mitochondrial colocalization of ectopically expressed c-Abl-EGFP/cMTS fusion across three cell lines (K562, Cos-7, and 1471.1) with varying levels of basal (and pharmacologically modulated) ROS. ROS were quantified by indicator dye assay. The functional consequences of mitochondrial c-Abl were assessed by DNA accessibility to 7-AAD using flow cytometry.

Results

The cMTS and cMTS/c-Abl fusions colocalized to the mitochondria in leukemic (K562) and breast (1471.1) cancer phenotypes (but not Cos-7 fibroblasts) in a ROS and PKC dependent manner.

Conclusions

We confirm and extend oxidative stress activated translocation of the cMTS by demonstrating that the cMTS and Abl/cMTS fusion selectively target the mitochondria of K562 leukemia and mammary adenocarcinoma 1471.1 cells. c-Abl induced K562 leukemia cell death when targeted to the matrix but not the outer membrane of the mitochondria.

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Abbreviations

c-Abl:

Abelson proto-oncoprotein

CML:

chronic myelogenous leukemia

cMTS:

cryptic mitochondrial translocation sequence

EGFP:

enhanced green fluorescent protein

JACoP:

Just Another Colocalization Plugin

mGSTA4-4:

murine glutathione-S-transferase A4-4

MOM:

mitochondrial outer membrane

PCC:

Pearson’s correlation coefficient

PKA:

protein kinase A

PKC:

protein kinase C

PMA:

phorbol myristate acetate

ROI:

region of interest

ROS:

reactive oxygen species

S:

serine

T:

threonine

TKI:

tyrosine kinase inhibitor

Y:

tyrosine

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Acknowledgments & DISCLOSURES

We acknowledge the use of the University of Utah, School of Medicine, Cell Imaging Facility and would like to thank the Director, Chris Rodesch, PhD, for scientific discussions. We would also like to thank Karina Matissek, Geoffrey Miller, and Dr. Andy Dixon for scientific discussions. The Core Facilities described in this project were supported by Award Number P30CA042014 from the National Cancer Institute. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Cancer Institute or the National Institutes of Health. The authors declare that they have no competing interests. This work was funded by NIH R01-CA129528 and by an AFPE Pre-Doctoral Fellowship (JEC).

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Authors and Affiliations

  1. Department of Pharmaceutics and Pharmaceutical Chemistry College of Pharmacy, University of Utah, 421 Wakara Way, Rm. 318, Salt Lake City, Utah, 84108, USA

    Carol S. Lim

  2. Department of Pharmacology and Toxicology College of Pharmacy, University of Utah, Salt Lake City, Utah, 84108, USA

    Jonathan E. Constance

  3. College of Pharmacy, University of Utah, Salt Lake City, Utah, 84108, USA

    Akemi Nishida

  4. University of Utah, Salt Lake City, Utah, 84108, USA

    Samuel D. Despres

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  1. Jonathan E. Constance
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Correspondence to Carol S. Lim.

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Constance, J.E., Despres, S.D., Nishida, A. et al. Selective Targeting of c-Abl via a Cryptic Mitochondrial Targeting Signal Activated by Cellular Redox Status in Leukemic and Breast Cancer Cells. Pharm Res 29, 2317–2328 (2012). https://doi.org/10.1007/s11095-012-0758-9

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