Inhibition of aquaporins as a potential adjunct to breast cancer cryotherapy

Abstract

Cryoablation is an emerging type of treatment for cancer. The sensitization of tumors using cryosensitizing agents prior to treatment enhances ablation efficiency and may improve clinical outcomes. Water efflux, which is regulated by aquaporin channels, contributes to cancer cell damage achieved through cryoablation. An increase in aquaporin (AQP) 3 is cryoprotective, whereas its inhibition augments cryodamage. The present study aimed to investigate aquaporin (AQP1, AQP3 and AQP5) gene expression and cellular localization in response to cryoinjury. Cultured breast cancer cells (MDA-MB-231 and MCF-7) were exposed to freezing to induce cryoinjury. RNA and protein extracts were then analyzed using reverse transcription-quantitative PCR and western blotting, respectively. Localization of aquaporins was studied using immunocytochemistry. Additionally, cells were transfected with small interfering RNA to silence aquaporin gene expression and cell viability was assessed using the Sulforhodamine B assay. Cryoinjury did not influence gene expression of AQPs, except for a 4-fold increase of AQP1 expression in MDA-MD-231 cells. There were no clear differences in AQP protein expression for either cell lines upon exposure to frozen and non-frozen temperatures, with the exception of fainter AQP5 bands for non-frozen MCF-7 cells. The exposure of cancer cells to freezing temperatures altered the localization of AQP1 and AQP3 proteins in both MCF-7 and MDA-MD-231 cells. The silencing of AQP1, AQP3 and AQP5 exacerbated MDA-MD-231 cell damage associated with freezing compared with control siRNA. This was also observed with AQP3 and AQP5 silencing in MCF-7 cells. Inhibition of aquaporins may potentially enhance cryoinjury. This cryosensitizing process may be used as an adjunct to breast cancer cryotherapy, especially in the border area targeted by cryoablation where freezing temperatures are not cold enough to induce cellular damage.

Utility of ferritin and inflammatory biomarkers in the diagnosis of different stages of breast cancer

“…In view of this, inhibition of aquaporins have been suggested to be beneficial. 29 , 30 Therefore, activation of noncanonical Wnt signaling and Aquaporin 7 are some of the important future areas of research in breast cancer.…”

Clinical application status and prospect of the combined anti-tumor strategy of ablation and immunotherapy

“…AQP3 participates in the plasma membrane permeability and intracellular water retention, which increases the critical freezing temperature to maintain cell survival advantage with cryoablation treatment. AQP inhibitors (mercuric chloride) combinate with cryoablation significantly reverses resistance and improve treatment effects ( 109 , 110 ), which supported AQP inhibitors in combination with cryoablation-immunotherapy in clinical practice. Combined cryoablation with Toripalimab (anti-PD-1) and Lenvatinib (anti-angiogenic agent) in a metastatic HCC patient also achieved CR at 7 months after treatment, and PFS as 24 months ( 111 ).…”

Recent progress in cryoablation cancer therapy and nanoparticles mediated cryoablation

“…Whether the formation of intracellular ice crystals or dehydration that dominates cellular death is still elusive. Regardless, membrane fluidity and permeability are crucial to cell viability during the freeze-thaw process 199 . More fluid cells tend to survive due to flexibility in the membranes as they swell or shrink under osmotic pressure.…”

“…However, as aquaporins exist in normal cells as well, nanoparticle-based delivery system of aquaporin inhibitors can be used to reduce the chance of side effects and synergize with cryoablation. Inhibition of aquaporins in cultured prostate and breast cancer cells enhanced cryoinjury 199 , 200 .…”