ClC-3 expression enhances etoposide resistance by increasing acidification of the late endocytic compartment

Abstract

Resistance to anticancer drugs and consequent failure of chemotherapy is a complex problem severely limiting therapeutic options in metastatic cancer. Many studies have shown a role for drug efflux pumps of the ATPbinding cassette transporters family in the development of drug resistance. ClC-3, a member of the CLC family of chloride channels and transporters, is expressed in intracellular compartments of neuronal cells and involved in vesicular acidification. It has previously been suggested that acidification of intracellular organelles can promote drug resistance by increasing drug sequestration. Therefore, we hypothesized a role for ClC-3 in drug resistance. Here, we show that ClC-3 is expressed in neuroendocrine tumor cell lines, such as BON, LCC-18, and QGP-1, and localized in intracellular vesicles colabeled with the late endosomal/lysosomal marker LAMP-1. ClC-3 overexpression increased the acidity of intracellular vesicles, as assessed by acridine orange staining, and enhanced resistance to the chemotherapeutic drug etoposide by almost doubling the IC 50 in either BON or HEK293 cell lines. Prevention of organellar acidification, by inhibition of the vacuolar H + -ATPase, reduced etoposide resistance. No expression of common multidrug resistance transporters, such as P-glycoprotein or multidrug-related protein-1, was detected in either the BON parental cell line or the derivative clone overexpressing ClC-3. The probable mechanism of enhanced etoposide resistance can be attributed to the increase of vesicular acidification as consequence of ClC-3 overexpression. This study therefore provides first evidence for a role of intracellular CLC proteins in the modulation of cancer drug resistance. [Mol Cancer Ther 2007;6(3):979 -86]

Role of the Vesicular Chloride Transporter ClC-3 in Neuroendocrine Tissue

“…Several authors, including ourselves, found ClC-3 on endosomes and synaptic vesicles (Stobrawa et al, 2001;Li et al, 2002;Salazar et al, 2004;HaraChikuma et al, 2005b;Suzuki et al, 2006;Weylandt et al, 2007;Zhao et al, 2007), but others described ClC-3 as a plasma membrane channel (Kawasaki et al, 1994;Duan et al, 1997;Huang et al, 2001;Isnard-Bagnis et al, 2003;Wang et al, 2006). ClC-3 was also reported to localize to insulin-containing LDCVs of pancreatic ␤ cells (Barg et al, 2001).…”

Intracellular Proton Pumps as Targets in Chemotherapy: V-ATPases and Cancer

“…Accordingly, cells that are able to maintain greater pH gradients in these compartments show a greater drug accumulation than normal cells [108] and cells showing a greater luminal acidification prowess are more chemoresistant than those that do not [109,110]. Conversely, inhibition of V-ATPase activity leads to chemosensitivity [111,112].…”

Sequestration of AS-DACA into Acidic Compartments of the Membrane Trafficking System as a Mechanism of Drug Resistance in Rhabdomyosarcoma

“…Lysosomal Associated Membrane-Protein-1 (LAMP1) is a marker of the late endosomal compartment [34,48]. LAMP1 expression levels were observed to be consistently higher in RD cells (Figure 4).…”

“…The inclusion of lysosomal drug sequestration in cancer resistance has been widely associated with anti-cancer drug resistance in multiple studies. Late endosomal and lysosomal compartments have been shown to display a variety of mechanisms that take part in cellular resistance phenotypes including enlargement of lysosomal compartments, spatial arrangements favourable for drug uptake and active mechanisms to transport drugs directly into vesicles [12,33,45,46,48,50–52]. …”