KCa3.1-dependent uptake of the cytotoxic DNA-binding dye Hoechst 33258 into cancerous but not healthy cervical cells

Abstract

The poor and nonselective penetration of current chemotherapeutics across the plasma membranes of cancer cells, which is necessary for the targeted disruption of the intracellular machinery, remains a major pharmaceutical challenge. In several cell types, including mast cells and macrophages, exposure to extracellular ATP is known to stimulate passive entry of large and otherwise membrane impermeable cationic dyes, which is usually attributed to conduction through ionotropic P2X receptors. Here, we report that elevations in cytosolic Ca2+ stimulate the rapid uptake and nuclear accumulation of a DNA-binding fluorescent cation, Hoechst 33258 (H33258), in cervical cancer cells. The H33258 uptake was dependent on activation of intermediate conductance Ca2+-activated K+ channels (KCa3.1), and direct stimulation of the channel with the activators SKA 31 and DCEBIO was sufficient to induce cellular uptake of H33258 directly. In contrast to the results from cancerous cervical cells, KCa3.1-dependent H33258 uptake was rarely observed in epithelial cells derived from the ectocervix and transformation zone of healthy cervical tissue. Furthermore, whole-cell patch clamp experiments and assessment of membrane potential using the slow voltage-sensitive dye DiSBAC2(3) revealed a significant difference in functional KCa3.1 activity between cancerous and healthy cervical epithelial cells, which correlated strongly with the incidence of KCa3.1-dependent H33258 uptake. Finally, we show that activation of KCa3.1 channels caused a modest but significant sensitization of cancer cells to the growth suppressant effects of H33258, lending plausibility to the idea of using KCa3.1 channel activators to enhance cell penetration of small cationic toxins into cancer cells expressing these channels.

Ion Channels and Personalized Medicine in Gynecological Cancers

“…To improve chemotherapy outcomes, a widely used alternative is to sensitize cancer cells to cytotoxic drugs by increasing their permeability, considering that several anticancer drugs, such as cisplatin and doxorubicin, have low membrane permeability. Bukhari et al found that the activation of the intermediate conductance Ca 2+ -activated K + channels (K Ca 3.1), which is upregulated in CCa cells, induces H33258 uptake, a cytotoxic DNA-binding cationic dye [ 23 ].…”

“… mRNA and protein overexpression in cervical cancer tissues. [ 23 , 32 , 33 ] Ovarian cancer K 2p 2.1 K 2p 10.1 Curcumin increases late apoptosis and decrease proliferation in SK-OV-3 and OVCAR-3 cells. Expression is increased in cancer samples compared to normal ovarian samples.…”

Ion Channels as Potential Tools for the Diagnosis, Prognosis, and Treatment of HPV-Associated Cancers

“…TRPV3 (transient receptor potential cation channel subfamily V member 3 or transient receptor potential vanilloid 3) is mainly expressed in keratinocytes and is activated by repetitive thermal stimuli [262]. The overexpression of TRPV3 in NSCLC is able to influence cell proliferation, channel blocking decreases the colony-forming capacity of cell lines, and the cell cycle is arrested in the G 1 /S phase (Table 2) [140]. The transient receptor potential cation channel subfamily V member 4 (or transient receptor potential vanilloid 4, TRPV4) is expressed in many tissues, is involved in the perception of various physical and chemical stimuli, and its expression has been associated with several neurodegenerative disorders [266].…”