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Lysosomal sequestration of hydrophobic weak base chemotherapeutics triggers lysosomal biogenesis and lysosome-dependent cancer multidrug resistance.

Zhitomirsky B, Assaraf YG - Oncotarget (2015)

Bottom Line: Non-cytotoxic, nanomolar concentrations, of the hydrophobic weak base chemotherapeutics doxorubicin and mitoxantrone triggered rapid lysosomal biogenesis that was associated with nuclear translocation of TFEB, the dominant transcription factor regulating lysosomal biogenesis.This resulted in increased lysosomal gene expression and lysosomal enzyme activity.The current study provides the first evidence that drug-induced TFEB-associated lysosomal biogenesis is an emerging determinant of MDR and suggests that circumvention of lysosomal drug sequestration is a novel strategy to overcome this chemoresistance.

View Article: PubMed Central - PubMed

Affiliation: The Fred Wyszkowski Cancer Research Laboratory, Dept. of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel.

ABSTRACT
Multidrug resistance (MDR) is a primary hindrance to curative cancer chemotherapy. In this respect, lysosomes were suggested to play a role in intrinsic MDR by sequestering protonated hydrophobic weak base chemotherapeutics away from their intracellular target sites. Here we show that intrinsic resistance to sunitinib, a hydrophobic weak base tyrosine kinase inhibitor known to accumulate in lysosomes, tightly correlates with the number of lysosomes accumulating high levels of sunitinib in multiple human carcinoma cells. Furthermore, exposure of cancer cells to hydrophobic weak base drugs leads to a marked increase in the number of lysosomes per cell. Non-cytotoxic, nanomolar concentrations, of the hydrophobic weak base chemotherapeutics doxorubicin and mitoxantrone triggered rapid lysosomal biogenesis that was associated with nuclear translocation of TFEB, the dominant transcription factor regulating lysosomal biogenesis. This resulted in increased lysosomal gene expression and lysosomal enzyme activity. Thus, treatment of cancer cells with hydrophobic weak base chemotherapeutics and their consequent sequestration in lysosomes triggers lysosomal biogenesis, thereby further enhancing lysosomal drug entrapment and MDR. The current study provides the first evidence that drug-induced TFEB-associated lysosomal biogenesis is an emerging determinant of MDR and suggests that circumvention of lysosomal drug sequestration is a novel strategy to overcome this chemoresistance.

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Lysosome-mediated protection of nuclear DNA from hydrophobic weak base cytotoxic agentsMCF-7 breast cancer cells and A549/K1.5 non-small cell lung cancer cells were incubated for 1 hr with 100 nM LysoTracker red (red fluorescence) and 10 μM of the imidazoacridinone C-1330 (green fluorescence). Fluorescence microscopy analysis was performed using a Zeiss inverted Cell-Observer Axiovert 200 microscope (Carl Zeiss, Oberkochen, Germany).
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Figure 1: Lysosome-mediated protection of nuclear DNA from hydrophobic weak base cytotoxic agentsMCF-7 breast cancer cells and A549/K1.5 non-small cell lung cancer cells were incubated for 1 hr with 100 nM LysoTracker red (red fluorescence) and 10 μM of the imidazoacridinone C-1330 (green fluorescence). Fluorescence microscopy analysis was performed using a Zeiss inverted Cell-Observer Axiovert 200 microscope (Carl Zeiss, Oberkochen, Germany).

Mentions: Here we postulated that lysosomal sequestration of hydrophobic weak base anticancer drugs prevents their accessibility to their intracellular target sites, hence abolishing their pharmacologic activity. In this respect, we hypothesized that cells with an increased lysosome number have an enhanced lysosomal sequestration capacity of hydrophobic weak base cytotoxic drugs. This should result in a markedly decreased drug concentration at the drug target site and consequently increased drug resistance. To test this hypothesis, MCF-7 cells which we found to contain a low number of lysosomes, as well as A549/K1.5 MDR non-small cell lung cancer cells, known to harbor high levels of lysosomes [25], were exposed for 1 hr to the imidazoacridinone C-1330 (10 μM) along with the viable lysosome fluorescent marker LysoTracker red (100 nM), followed by fluorescence microscopy. C-1330 is a naturally fluorescent cytotoxic topoisomerase II inhibitor which we recently found to undergo a dramatic compartmentalization in lysosomes [25]; hence, in order to exert its topoisomerase II inhibitory activity, C-1330 must reach the nucleus. MCF-7 cells which contain a low number of lysosomes per cell displayed a high confinement of C-1330 fluorescence in their nuclei (Fig. 1). In contrast, C-1330 fluorescence in MDR A549/K1.5 cells which contain a high number of lysosomes was predominantly confined to lysosomes rather than to the nucleus. This finding is an initial evidence for a likely correlation between the elevated number of lysosomes per cell, and the lysosome-dependent protection of intracellular drug target sites from cytotoxic drug activity.


Lysosomal sequestration of hydrophobic weak base chemotherapeutics triggers lysosomal biogenesis and lysosome-dependent cancer multidrug resistance.

Zhitomirsky B, Assaraf YG - Oncotarget (2015)

Lysosome-mediated protection of nuclear DNA from hydrophobic weak base cytotoxic agentsMCF-7 breast cancer cells and A549/K1.5 non-small cell lung cancer cells were incubated for 1 hr with 100 nM LysoTracker red (red fluorescence) and 10 μM of the imidazoacridinone C-1330 (green fluorescence). Fluorescence microscopy analysis was performed using a Zeiss inverted Cell-Observer Axiovert 200 microscope (Carl Zeiss, Oberkochen, Germany).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4359223&req=5

Figure 1: Lysosome-mediated protection of nuclear DNA from hydrophobic weak base cytotoxic agentsMCF-7 breast cancer cells and A549/K1.5 non-small cell lung cancer cells were incubated for 1 hr with 100 nM LysoTracker red (red fluorescence) and 10 μM of the imidazoacridinone C-1330 (green fluorescence). Fluorescence microscopy analysis was performed using a Zeiss inverted Cell-Observer Axiovert 200 microscope (Carl Zeiss, Oberkochen, Germany).
Mentions: Here we postulated that lysosomal sequestration of hydrophobic weak base anticancer drugs prevents their accessibility to their intracellular target sites, hence abolishing their pharmacologic activity. In this respect, we hypothesized that cells with an increased lysosome number have an enhanced lysosomal sequestration capacity of hydrophobic weak base cytotoxic drugs. This should result in a markedly decreased drug concentration at the drug target site and consequently increased drug resistance. To test this hypothesis, MCF-7 cells which we found to contain a low number of lysosomes, as well as A549/K1.5 MDR non-small cell lung cancer cells, known to harbor high levels of lysosomes [25], were exposed for 1 hr to the imidazoacridinone C-1330 (10 μM) along with the viable lysosome fluorescent marker LysoTracker red (100 nM), followed by fluorescence microscopy. C-1330 is a naturally fluorescent cytotoxic topoisomerase II inhibitor which we recently found to undergo a dramatic compartmentalization in lysosomes [25]; hence, in order to exert its topoisomerase II inhibitory activity, C-1330 must reach the nucleus. MCF-7 cells which contain a low number of lysosomes per cell displayed a high confinement of C-1330 fluorescence in their nuclei (Fig. 1). In contrast, C-1330 fluorescence in MDR A549/K1.5 cells which contain a high number of lysosomes was predominantly confined to lysosomes rather than to the nucleus. This finding is an initial evidence for a likely correlation between the elevated number of lysosomes per cell, and the lysosome-dependent protection of intracellular drug target sites from cytotoxic drug activity.

Bottom Line: Non-cytotoxic, nanomolar concentrations, of the hydrophobic weak base chemotherapeutics doxorubicin and mitoxantrone triggered rapid lysosomal biogenesis that was associated with nuclear translocation of TFEB, the dominant transcription factor regulating lysosomal biogenesis.This resulted in increased lysosomal gene expression and lysosomal enzyme activity.The current study provides the first evidence that drug-induced TFEB-associated lysosomal biogenesis is an emerging determinant of MDR and suggests that circumvention of lysosomal drug sequestration is a novel strategy to overcome this chemoresistance.

View Article: PubMed Central - PubMed

Affiliation: The Fred Wyszkowski Cancer Research Laboratory, Dept. of Biology, Technion-Israel Institute of Technology, Haifa 32000, Israel.

ABSTRACT
Multidrug resistance (MDR) is a primary hindrance to curative cancer chemotherapy. In this respect, lysosomes were suggested to play a role in intrinsic MDR by sequestering protonated hydrophobic weak base chemotherapeutics away from their intracellular target sites. Here we show that intrinsic resistance to sunitinib, a hydrophobic weak base tyrosine kinase inhibitor known to accumulate in lysosomes, tightly correlates with the number of lysosomes accumulating high levels of sunitinib in multiple human carcinoma cells. Furthermore, exposure of cancer cells to hydrophobic weak base drugs leads to a marked increase in the number of lysosomes per cell. Non-cytotoxic, nanomolar concentrations, of the hydrophobic weak base chemotherapeutics doxorubicin and mitoxantrone triggered rapid lysosomal biogenesis that was associated with nuclear translocation of TFEB, the dominant transcription factor regulating lysosomal biogenesis. This resulted in increased lysosomal gene expression and lysosomal enzyme activity. Thus, treatment of cancer cells with hydrophobic weak base chemotherapeutics and their consequent sequestration in lysosomes triggers lysosomal biogenesis, thereby further enhancing lysosomal drug entrapment and MDR. The current study provides the first evidence that drug-induced TFEB-associated lysosomal biogenesis is an emerging determinant of MDR and suggests that circumvention of lysosomal drug sequestration is a novel strategy to overcome this chemoresistance.

Show MeSH
Related in: MedlinePlus