Fig. 3

CD-MSC/5-FC, CD::UPRT-MSC/5-FC, and CD::UPRT-MSC/5-FU regimen have limited efficiency in chemoresistant cells. a The HT-29/EGFP and HT-29/EGFP/FUR cells were co-cultivated (5:1 ratio) with either CD-MSC or CD::UPRT-MSC in a concentration gradient of 5-FC, and relative luminescence corresponding to the viability was evaluated on day 6. The luminescence in co-cultures without the 5-FC was set to 100% by default. The bystander cytotoxic effect of both CD-MSC and CD::UPRT-MSC was low in chemoresistant derivative compared to cytotoxicity of therapeutic MSC exerted on the parental cell line. CD-MSC did not exhibit not even 50% cytotoxicity in the chemoresistant cells when compared to the parental HT-29/EGFP. Values were expressed as means of sextaplicates ± SD. Mann–Whitney U-test was used for statistical analysis. **p < 0.01. b Kinetic monitoring and evaluation of a number of red nuclei-stained tumor cells during 6-day-long cocultivation with the therapeutic MSC, and addition of the 5-FC at a concentration of 500 µg/ml, showed that regardless of the type of prodrug-converting enzyme used in the treatment regimen of chemonaïve HT-29/EGFP cells, bystander cytotoxicity was not different. The number of NLR cells in the co-culture without 5-FC at a given time point was taken as 100%. Expressed as means of sextaplicates ± SD. c The same experimental setup (as in Fig. 3b) on the chemoresistant HT-29/EGFP/FUR cells showed significant difference in the bystander toxicity between CD-MSC or CD::UPRT-MSC treatment. Expressed as means of sextaplicates ± SD. Mann–Whitney U-test was used for statistical analysis. *p < 0.05. d The higher efficiency of formation and release of the 5-FU from engineered MSC transduced with fusion CD::UPRT gene in comparison to CD-MSC confirmed by HPLC-MS. e The direct cocultivation of chemoresistant FUR/NLR cells with therapeutic CD::UPRT-MSC (5:1 ratio) in the presence of the 5-FU at a concentration of 5 or 10 µg/ml led to significantly increased sensitivity of the tumor cells to 5-FU as evaluated at the experimental end point 6 days after the treatment started by direct enumeration of the red nuclei of the tumor cells. Values were expressed as means of sextaplicates ± SD. Mann–Whitney U-test was used for statistical analysis. *p < 0.05; **p < 0.01