Fig. 5: Fasting promotes cholesterol efflux in cancer cells.

Tumours (Capan-1 xenografts) were isolated at the end of the experiment presented in Fig. 3e and utilized for RNA isolation and for the subsequent measurement of ABCG1 expression by QPCR (a) or for protein lysate generation and subsequent detection of ABCG1, ABCA1 and cyclophilin B (CypB) by Western blotting (b). c Tumours were isolated at the end of the experiment presented in Fig. 2f (HCT116 xenografts) and utilized for RNA isolation and for the detection of ABCG1 and ACAT1 expression by QPCR. d HCT116 and MCF7 cells were plated in 96-well plates. Radiolabelling was carried out in standard CC w/ or w/o 20 μM terbinafine, 10 μM clotrimazole, or in FMCC w/ or w/o IGF1 (5 ng/ml)+leptin (50 ng/ml)+insulin (500 pM) (“fasting-reduced factors” combination, FRFs comb.). The cholesterol acceptors apolipoprotein A-I (10 µg/ml) or HDL (12.5 µg/ml) were added for 4 h before the medium was collected and radioactivity was quantified (CEC: cholesterol efflux capacity). e–g Capan-1 xenografts were established in 6-8-week-old female athymic nude mice. Once tumours were palpable, mice were randomized to be treated with ad lib. diet (n = 8), weekly 48 h water-only fasting (n = 11), or fasting plus human HDL (1 mg/mouse twice a week coupled with fasting; n = 7). Tumour volume was calculated at the indicated time points (e). At the end of the experiment, tumours were weighted (f) and tumour cholesterol content was quantified (g). In (e–g), the data from mice that were treated with just ad lib. diet or with just fasting were already presented in Fig. 3e–g. They are shown here again to allow comparison with the group, fasting+HDL (all of these groups were included the same experiment). In (e), n indicates the number of tumours per treatment group. In (a, c, f, g) data points are biological replicates (they represent single tumours). In (d), data points are experimental replicates. In (b), one out of two independent experiments is presented. In (a, c, d, f, g), data are shown as mean ± SD whereas in (e), data are shown as mean ± SEM. In (a, c, and in e–g), p values were calculated by two-tailed Student’s t test. In (d), p values were calculated by one-tailed Student’s t test. Source data are provided as a Source Data file.