Targeted Killing of CSCs by CDF Figure 7. CDF exhibited anti-tumor activity in MIAPaCa-2 cells induced tumors in a xenograft mouse model, which was consistent with inhibition of NF-kB DNA binding, COX-2, miR-21, and caused re-expression of miR200 in tumor remnants. Anti-tumor activity and changes in tumor weight from each group of animals (A). The arrow indicates starting day of the treatment. NF-kB DNA binding activity of tumor tissues; and NF-kB competition control study with unlabeled NF-kB oligonucleotide (B). Western blots analysis of COX-2, PTEN and b-actin expression in tumor remnants (C); miR-21, miR-200b and miR-200c expression in tumor remnants as measured by real-time RT-PCR (D). P values were calculated by the paired t test. doi:10.1371/journal.pone.0017850.g007 MIAPaCa-2 cells (Figure 7D). In contrast, we found that the CDF treatment with or without gemcitabine combination showed increased expression of both miR-200b, and miR-200c, but the effect with curcumin or its combination was minimal, suggesting the superiority of CDF in suppressing the expression of miR-21, resulting in the re-expression of PTEN, and re-expression of miR- 200 which could be responsible for the reversal of EMT phenotype in cells treated with CDF. Overall, these results suggest that the phenotypic characteristics of MIAPaCa-2 tumors are consistent with enriched population of CSCs and EMT characteristics, and these drug resistant cells could be killed either by CDF alone or in combination with gemcitabine. Pancreatospheres enhanced tumor growth in vivo Under traditional experimental conditions, we normally inject one million cells for assessing tumor growth; however, for PLoS ONE | www.plosone.org 8 March 2011 | Volume 6 | Issue 3 | e17850