1. Volume 137, Issue 3, Pages 1102-1113 (September 2009)
Combined Targeted Treatment to Eliminate Tumorigenic Cancer Stem Cells in Human Pancreatic Cancer 
Maria–Theresa Mueller, Patrick C. Hermann, Juliane Witthauer, Belen Rubio–Viqueira, Simon F. Leicht, Stephan Huber, Joachim W. Ellwart, Mona Mustafa, Peter Bartenstein, Jan G. D'Haese, Michael H. Schoenberg, Frank Berger, Karl–Walter Jauch, Manuel Hidalgo, Christopher Heeschen 
Gastroenterology 
Volume 137, Issue 3, Pages (September 2009)
DOI: /j.gastro
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2. Figure 1
Treatment effects on CD133+ cells. (A) Effect of different in vitro treatments on L3.6pl-derived CD133+ CSCs were assessed by flow cytometry. Representative flow cytometry analyses and culture photographs are provided (each group n ≥ 3). (*P < .01 vs control, **P < .05 vs single-agent therapy, ***P < .05 vs double-agent therapy). (B) Immunohistochemistry demonstrated activation of the phosphatidylinositol 3-kinase/Akt/mTOR pathway in pancreatic cancer (stem) cells via phosphorylation status of the s6 ribosomal protein (s6rp; upper row, magnification below). Representative pictures for untreated and rapamycin-treated cells are provided (lower left panel). Polymerase chain reaction shows inhibition of the downstream Shh target Gli1 by cyclopamine and CUR (lower right panel). (C) Pancreatic tumor spheres were treated either with gemcitabine, single-agent therapy, or CRG (each group, n = 3; *P < .05 vs control). Representative pictures are provided for cultures and flow cytometry analyses of dissociated spheres. (D) Effect of gemcitabine versus CRG therapy on side population cells. Representative flow cytometry plots are provided. (E) Analysis of CSC content in fresh primary pancreatic cancer cells by flow cytometry with respect to allocated treatment (CD133+ CSCs, n = 7, *P < .001; CD24+CD44+ CSCs, n = 4, *P < .05).
Gastroenterology  , DOI: ( /j.gastro )
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3. Figure 2
Antimetastatic activity of CRG therapy. (A) Effect of different treatment regimens on migratory capacity was analyzed using a modified Boyden chamber assay (each group, n = 3; *P < .05 vs control). (B) Flow cytometry was used to assess content of CD133+CXCR4+ migrating CSCs (*P < .05 vs control, **not significant vs cyclopamine and gemcitabine). (C) In vivo metastatic capacity of cells was evaluated by intravenous infusion of pretreated and Qtracker-labeled tumor cells. Representative pictures of metastases are provided for near-infrared imaging and matched H&E-stained histologic sections (each group, n ≥ 5; *P < .05 vs gemcitabine; **not significant vs cyclopamine and gemcitabine).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

4. Figure 3
Elimination of tumorigenic potential after in vitro pretreatment. (A) Depiction of the experimental setup. (B) Tumorigenicity was evaluated by FDG-PET scans on day 30 and verified by histology on day 35. Representative PET scans as well as macroscopic and microscopic histologic pictures. (C) Statistical analysis (each group, n = 5; *P < .05 vs gemcitabine).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

5. Figure 4
In vivo triple therapy results in tumor-free long-time survival following orthotopic implantation of L3.6pl cells. (A) Depiction of the experimental setup. (B) Tumor volume on day 32 (n = 5 per group; *P < .005, **P < .001). (C) Magnetic resonance imaging on day 49. Representative scans are provided. (D) White blood cell counts at the end of the experiments. (E) Tumor take rate on day 100. (F) Survival time for treatment groups is illustrated as Kaplan–Meier survival curves (control, n = 5; gemcitabine, n = 13; CRG, n = 11; P < .01 vs gemcitabine).
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions

6. Figure 5
In vivo triple therapy results in tumor regression in primary human pancreatic cancer. (A) Depiction of the experimental setup. (B) Tumor volume during 49 days of follow-up for fast-growing (left panel) and slower-growing tumors (right panel). (C) Survival times for the 3 treatment groups are illustrated as Kaplan–Meier survival curves (gemcitabine, n = 12; CycloRG, n = 13; CurRG, n = 14; both P < .001 vs gemcitabine). (D) Phenotyping of in vivo treated tumors on day 40 following implantation according to allocated treatment. Cells were first gated for exclusion of 7AAD (not shown) and then for expression of EpCAM, followed by double staining for CD133/CD44 and CD44/CD24, respectively.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2009 AGA Institute Terms and Conditions