Cooling-mediated protection from chemotherapy drug-induced cytotoxicity in human keratinocytes by inhibition of cellular drug uptake

Abstract

Chemotherapy-induced alopecia (CIA) represents the most distressing side-effect for cancer patients. Scalp cooling is currently the only treatment to combat CIA, yet little is known about its cytoprotective effects in human hair follicles (HF). We have previously established in vitro human keratinocyte models to study the effects of taxanes and anthracyclines routinely-used clinically and reported that cooling markedly-reduced or even completely-prevented cytotoxicity in a temperature dependent manner. Using these models (including HFderived primary keratinocytes), we now demonstrate that cooling markedly attenuates cellular uptake of the anthracyclines doxorubicin and epirubicin to reduce or prevent drug-mediated human keratinocyte cytotoxicity. We show marked reduction in drug uptake and nuclear localization qualitatively by fluorescence microscopy. We have also devised a flow cytometry-based methodology that permitted semi-quantitative analysis of differences in drug uptake, which demonstrated that cooling can reduce drug uptake by up to~8-fold in comparison to normal/physiological temperature, an effect that was temperature-dependent. Our results provide evidence that attenuation of cellular drug uptake represents at least one of the mechanisms underpinning the ability of cooling to rescue human keratinocytes from chemotherapy drug-cytotoxicity, thus supporting the clinical efficacy of scalp cooling.

TRAF3/p38-JNK Signalling Crosstalk with Intracellular-TRAIL/Caspase-10-Induced Apoptosis Accelerates ROS-Driven Cancer Cell-Specific Death by CD40

“…HCT116 cells were seeded at 1 × 10 4 cells/well in 96-well plates and treated with 10-20 ng/mL of human recombinant killer TRAIL™ (Enzo, Exeter, UK) in the absence or presence of anti-FasL mAb NOK-1, anti-TRAIL mAb RIK-2 or control isotype Control IgG (all at 10 µg/mL) [ 13 ], alongside untreated control cultures, for 24 h. Cell viability was then determined using the CellTiter 96 ® AQueous One cell proliferation assay (Promega, Southampton, UK) by addition of 20 μL CellTiter reagent to each well and incubation at 37 °C for 4 h. Absorbance was measured using a FLUOstar OPTIMA (BMG Labtech, Bucks, UK) at 492 nm and percentage (%) Cell Viability calculated as previously described [ 27 ].…”

A novel method for the establishment of autologous skin cell suspensions: characterisation of cellular sub-populations, epidermal stem cell content and wound response-enhancing biological properties

“…Quantitative detection of NHEK and HaCaTa cell proliferation was performed using the CellTiter 96 ® AQueous One Solution Cell Proliferation Assay (Promega, Southampton, United Kingdom), as recommended by the manufacturer and absorbance determined by spectrophotometric measurements at 492 nm using a FLUOstar OPTIMA (BMG Labtech, Bucks, United Kingdom). Following normalisation using medium-only 'blank' wells, cell proliferation was calculated as described previously (Dunnill et al, 2020).…”

Scalp cooling therapy for chemotherapy-induced hair loss in Asian patients with breast or gynecological cancers – a tertiary institution experience

“…Scalp cooling therapy (SCT) has gained traction over the past decade as a method to decrease CIA, based on the concept of scalp hypothermia reducing blood ow in the scalp to 20-40% of the usual rate by vasoconstriction. This decreases the concentration of chemotherapy reaching the scalp [9], and also metabolic rate of the follicle cells and drug uptake [10,11]. In vivo studies have shown that cooling protects human keratinocytes from chemotherapy-induced toxicity [11,12].…”

“…This decreases the concentration of chemotherapy reaching the scalp [9], and also metabolic rate of the follicle cells and drug uptake [10,11]. In vivo studies have shown that cooling protects human keratinocytes from chemotherapy-induced toxicity [11,12]. Various scalp cooling systems including Paxman and Dignicap have been shown to be effective in reducing hair loss in 50-68% of patients [13][14][15][16][17][18][19][20].…”