Effect of SSRI exposure on the proliferation rate and glucose uptake in breast and ovary cancer cell lines

Breast cancer is the most prevalent malignancy amongst women worldwide while ovarian cancer represents the leading cause of death among gynecological malignancies. Women suffering from these cancers displayed heightened rates of major depressive disorder, and antidepressant treatment with selective serotonin reuptake inhibitors (SSRIs) is frequently recommended. Recently, narrative reviews and meta-analyses showed increased recurrence risks and mortality rates in SSRI-treated women with breast and ovarian cancer. We therefore examined whether three commonly prescribed SSRIs, fluoxetine, sertraline and citalopram, affect proliferation or glucose uptake of human breast and ovarian cancer cell lines characterized by different malignancies and metastatic potential. SSRI treatment or serotonin stimulation with therapeutically relevant concentrations over various time periods revealed no consistent dose- or time-dependent effect on proliferation rates. A marginal, but significant increase in glucose uptake was observed in SK-OV-3 ovarian cancer cells upon fluoxetine or sertraline, but not citalopram treatment. In three breast cancer cell lines and in two additional ovarian cancer cell lines no significant effect of SSRIs on glucose uptake was observed. Our data suggest that the observed increase in recurrence- and mortality rates in SSRI-treated cancer patients is unlikely to be linked to antidepressant therapies.

MDA-MSC-hyb1 [4] human, derived from fusion of MDA-MB-231 with mesenchymal stroma/stemlike cells high malignancy and metastatic potential MDA-MSC-hyb3 [5] human, derived from fusion of MDA-MB-231 with mesenchymal stroma/stemlike cells low malignancy and metastatic potential SK-OV-3 [6] human, ovary, epithelial derived ascites of an ovarian serous cystadenocarcinoma prominent subcutaneous tumor development in NOD SCID mice NIH:OVCAR-3 [7,8] human, ovary, epithelial, derived from ascites of a progressive adenocarcinoma of the ovary distinct chemotherapeutic sensitivity as compared to SK-OV-3 or SCCOHT-1 cells, hormone receptors SCCOHT-1 [9] human, derived from biopsy of ovarian small-cell carcinoma of the hypercalcemic type, FIGO Ia cell culture model for SCCOHT, expression of CD90 and vimentin as mesenchymal-like markers and NCAM/CD56 as neuroendocrine marker SK-MSC-hyb1 [10] human, derived from fusion of SK-OV-3 with mesenchymal stroma/stem-like cells diminished proliferative capacity and reduced malignancy as compared to the parental SK-OV-3 cells CD: cluster of differentiation, ER: estrogen receptor, FIGO: International Federation of Gynecology and Obstetrics, Her2: human epidermal growth factor receptor 2, NCAM: neural cell adhesion molecule, PR: progesterone receptor, SCCOHT: small cell carcinoma of the ovary hypercalcemic type. Figure S3: Effect of low-dose, short-term SSRI treatment or 5-HT exposure on breast cancer cell lines measured by MTT assay. Bar graphs depict relative absorbance measured in MTT assay of MCF-10A (a), MCF-7 (b), MDA-MB-231 (c), MDA-MSC-hyb1 (d) and MDA-MSC-hyb3 (e) cells in response to treatment with 100 nM or 1,000 nM fluoxetine (Fluo; yellow), sertraline (Sert; red), citalopram (Cita; blue) or 5-HT (grey) for indicated time periods (24 h to 72 h) compared to untreated control cells (Ctrl; white) and cells receiving carboplatin (CP; 1,000 nM; black). Data are depicted as mean ± SEM and summarize n = 3 experiments. Pvalues were determined by two-way ANOVA followed by Dunnett's multiple comparison test; ***P < 0.001; **P < 0.01; *P < 0.05 versus corresponding Ctrl. Figure S4: Effect of low-dose, short-term SSRI treatment or 5-HT exposure on ovarian cancer cell lines measured by MTT assay. Bar graphs depict relative absorbance measured in MTT assay of SK-OV-3 (a), NIH:OVCAR-3 (b), SCCOHT-1 (c) and SK-MSC-hyb1 (d) cells in response to exposure to 100 nM or 1,000 nM fluoxetine (Fluo; yellow), sertraline (Sert; red), citalopram (Cita; blue) or 5-HT (grey) for indicated time periods (24 h to 72 h) compared to untreated control cells (Ctrl; white) and cell receiving carboplatin (CP; 1,000 nM; black). Data are depicted as mean ± SEM and summarize n = 3 experiments. P-values were determined by two-way ANOVA followed by Dunnett's multiple comparison test; ***P < 0.001; **P < 0.01; *P < 0.05 versus corresponding Ctrl.      Two-way ANOVA followed by either Dunnett's multiple comparison test was used to calculate two-tailed P-values to compare 5-HT groups to respective Ctrl, or by Sidak's multiple comparison test to assess differences between corresponding time points (a and b). ***P < 0.001, **P < 0.01, *P < 0.05 versus corresponding Ctrl, ###P < 0.001 versus corresponding time point with 0.2% FCS. Table S2: Summary of Figure 1 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated breast cancer cell lines when compared to DMSO-treated control cells.  Table S4: Summary of Figure S3 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated breast cancer cell lines when compared to untreated control cells.

MDA-MB-231
Supplemental Table S5: Summary of Figure S4 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated ovarian cancer cell lines when compared to untreated control cells.
c: drug concentration in nM, t: time-point in hours. P-values were determined by two-way ANOVA followed by Dunnett's multiple comparison test. Conditions resulting in a significant up-regulation of proliferation rates are indicated in bold print.
Supplemental Table S6: Summary of Figure 5 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated breast cancer cell lines when compared to untreated control cells.
c: drug concentration in nM, t: time-point in hours. P-values were determined by two-way ANOVA followed by Dunnett's multiple comparison test. Conditions resulting in a significant up-regulation of proliferation rates are indicated in bold print.  Table S8: Summary of Figure S6 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated breast cancer cell lines when compared to DMSO-treated control cells.
c: drug concentration in µM, t: time-point in hours. P-values were determined by two-way ANOVA followed by Dunnett's multiple comparison test. Conditions resulting in a significant up-regulation of proliferation rates are indicated in bold print.  Table S9: Summary of Figure S7 detailing time-points and drug concentrations resulting in statistically significant changes in relative proliferation rate of indicated ovarian cancer cell lines when compared to DMSO-treated control cells.
c: drug concentration in µM, t: time-point in hours. P-values were determined by two-way ANOVA followed by Dunnett's multiple comparison test. Conditions resulting in a significant up-regulation of proliferation rates are indicated in bold print.