Figure 5.

In vitro and in vivo cytotoxicity of Chi-Tn-ADC on cancer cells (A–D) In vitro cytotoxicity of Chi-Tn ADC. (A) TA3Ha, Jurkat, OvCar-3 or Shin-3 cells were cultured with the indicated concentrations of free saporin (SAP). (B) Jurkat, TA3Ha (left panel), and OvCar-3 and Shin-3 cells (right panel) were cultured with the indicated concentrations of Chi-Tn mAb (open symbol) or the Chi-Tn/SAP conjugate (filled symbol). (C) Jurkat and SKBR3 cells were cultured with free MMAF. (D) Jurkat (left panel) and SKBR3 (right panel) cells were cultured with Chi-Tn/MMAF or Her/MMAF conjugate, naked Chi-Tn or Her mAb. Cell viability was assessed after 3 d of culture. Results are expressed as percentage of viable cells compared to untreated cells. (E–H) In vivo cytotoxicity of Chi-Tn/MMAF against Tn⁺ LOX tumor cells. (E) LOX cells express a high and stable level of Tn. LOX cells were labeled with the Chi-Tn mAb or a control antibody at 20 µg/mL, then with a GaH-Fc-PE secondary antibody and Tn expression was measured by flow cytometry. (F) LOX cells were cultured with the Chi-Tn/MMAF or Her/MMAF conjugate at the indicated concentrations. Cell viability was then assessed after 3 d at 37°C. Results are expressed as percentage of viable cells compared to untreated cells. (G) Experimental schedule of the in vivo antitumor assay. Nude mice were grafted with the LOX Tn⁺ human cell line and then treated twice a week with the Chi-Tn/MMAF (n = 8) or control Her/MMAF (n = 6) conjugates, with the naked Chi-Tn mAb (n = 6), or were left untreated (n = 7). (H) Tumor growth is depicted as Relative Tumor Volume (RTV), as explained in the Materials and Methods section. Statistical significance was calculated with Mann-Whitney test; significant statistical difference was observed on days 20 and 25 between Her-MMAF and Chi-Tn-MMAF treated groups (*p < 0,05).