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Establishment and characterization of a new human pancreatic adenocarcinoma cell line with high metastatic potential to the lung.

Kalinina T, Güngör C, Thieltges S, Möller-Krull M, Penas EM, Wicklein D, Streichert T, Schumacher U, Kalinin V, Simon R, Otto B, Dierlamm J, Schwarzenbach H, Effenberger KE, Bockhorn M, Izbicki JR, Yekebas EF - BMC Cancer (2010)

Bottom Line: Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6), while others were detected for the first time (MEMO1, RIOK3).Specifically highly overexpressed genes (fold change > 10) were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6.Subcutaneous transplantation of PaCa 5061 into pfp-/-/rag2-/- mice resulted in formation of primary tumors and spontaneous lung metastasis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of General, Visceral and Thoracic Surgery, University Hospital Hamburg, Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany. tkalinin@uke.de

ABSTRACT

Background: Pancreatic cancer is still associated with devastating prognosis. Real progress in treatment options has still not been achieved. Therefore new models are urgently needed to investigate this deadly disease. As a part of this process we have established and characterized a new human pancreatic cancer cell line.

Methods: The newly established pancreatic cancer cell line PaCa 5061 was characterized for its morphology, growth rate, chromosomal analysis and mutational analysis of the K-ras, EGFR and p53 genes. Gene-amplification and RNA expression profiles were obtained using an Affymetrix microarray, and overexpression was validated by IHC analysis. Tumorigenicity and spontaneous metastasis formation of PaCa 5061 cells were analyzed in pfp-/-/rag2-/- mice. Sensitivity towards chemotherapy was analysed by MTT assay.

Results: PaCa 5061 cells grew as an adhering monolayer with a doubling time ranging from 30 to 48 hours. M-FISH analyses showed a hypertriploid complex karyotype with multiple numerical and unbalanced structural aberrations. Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6), while others were detected for the first time (MEMO1, RIOK3). Specifically highly overexpressed genes (fold change > 10) were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6. Subcutaneous transplantation of PaCa 5061 into pfp-/-/rag2-/- mice resulted in formation of primary tumors and spontaneous lung metastasis.

Conclusion: The established PaCa 5061 cell line and its injection into pfp-/-/rag2-/- mice can be used as a new model for studying various aspects of the biology of human pancreatic cancer and potential treatment approaches for the disease.

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PaCa 5061 cell viability following Gemcitabine, 5-FU, Cetuximab, and Gefitinib treatment. Cell proliferation following drug treatment for 72 h was estimated by MTT test. 6A. Dose-dependent inhibition of cell proliferation in Gemcitabine and 5-FU treated cells as well as in Cetuximab and Gefitinib treated cells (6B). Quantitative values are means ± SEM from 3 independent experiments performed in quadruplicate.
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Figure 6: PaCa 5061 cell viability following Gemcitabine, 5-FU, Cetuximab, and Gefitinib treatment. Cell proliferation following drug treatment for 72 h was estimated by MTT test. 6A. Dose-dependent inhibition of cell proliferation in Gemcitabine and 5-FU treated cells as well as in Cetuximab and Gefitinib treated cells (6B). Quantitative values are means ± SEM from 3 independent experiments performed in quadruplicate.

Mentions: We examined the cellular sensitivity of PaCa 5061 cells to different drugs in vitro using the proliferation assay. PaCa 5061 cells were treated with increasing concentrations of the chemotherapeutic drugs Gemcitabine (0.1-10 μM) and 5-FU (0.1-10 μM). PaCA 5061 cells exhibited native resistance to both drugs and inhibiton of cell proliferation to 50% (IC50) was achieved with 10 μM for Gemcitabine and > 10 μM for 5-FU respectively (Figure 6A). As PaCa 5061 cells were characterized by an elevated expression level of EGFR, we blocked EGFR activation by a tyrosin-kinase inhibitor (Gefitinib) or EGF binding by a monoclonal EGFR antibody (Cetuximab). PaCa 5061 cells showed robust resistance to these drugs, and IC50 was achieved with > 15 μM for Gefitinib and > 10 μg/ml for Cetuximab (Figure 6B).


Establishment and characterization of a new human pancreatic adenocarcinoma cell line with high metastatic potential to the lung.

Kalinina T, Güngör C, Thieltges S, Möller-Krull M, Penas EM, Wicklein D, Streichert T, Schumacher U, Kalinin V, Simon R, Otto B, Dierlamm J, Schwarzenbach H, Effenberger KE, Bockhorn M, Izbicki JR, Yekebas EF - BMC Cancer (2010)

PaCa 5061 cell viability following Gemcitabine, 5-FU, Cetuximab, and Gefitinib treatment. Cell proliferation following drug treatment for 72 h was estimated by MTT test. 6A. Dose-dependent inhibition of cell proliferation in Gemcitabine and 5-FU treated cells as well as in Cetuximab and Gefitinib treated cells (6B). Quantitative values are means ± SEM from 3 independent experiments performed in quadruplicate.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2927995&req=5

Figure 6: PaCa 5061 cell viability following Gemcitabine, 5-FU, Cetuximab, and Gefitinib treatment. Cell proliferation following drug treatment for 72 h was estimated by MTT test. 6A. Dose-dependent inhibition of cell proliferation in Gemcitabine and 5-FU treated cells as well as in Cetuximab and Gefitinib treated cells (6B). Quantitative values are means ± SEM from 3 independent experiments performed in quadruplicate.
Mentions: We examined the cellular sensitivity of PaCa 5061 cells to different drugs in vitro using the proliferation assay. PaCa 5061 cells were treated with increasing concentrations of the chemotherapeutic drugs Gemcitabine (0.1-10 μM) and 5-FU (0.1-10 μM). PaCA 5061 cells exhibited native resistance to both drugs and inhibiton of cell proliferation to 50% (IC50) was achieved with 10 μM for Gemcitabine and > 10 μM for 5-FU respectively (Figure 6A). As PaCa 5061 cells were characterized by an elevated expression level of EGFR, we blocked EGFR activation by a tyrosin-kinase inhibitor (Gefitinib) or EGF binding by a monoclonal EGFR antibody (Cetuximab). PaCa 5061 cells showed robust resistance to these drugs, and IC50 was achieved with > 15 μM for Gefitinib and > 10 μg/ml for Cetuximab (Figure 6B).

Bottom Line: Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6), while others were detected for the first time (MEMO1, RIOK3).Specifically highly overexpressed genes (fold change > 10) were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6.Subcutaneous transplantation of PaCa 5061 into pfp-/-/rag2-/- mice resulted in formation of primary tumors and spontaneous lung metastasis.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of General, Visceral and Thoracic Surgery, University Hospital Hamburg, Eppendorf, Martinistrasse 52, 20246 Hamburg, Germany. tkalinin@uke.de

ABSTRACT

Background: Pancreatic cancer is still associated with devastating prognosis. Real progress in treatment options has still not been achieved. Therefore new models are urgently needed to investigate this deadly disease. As a part of this process we have established and characterized a new human pancreatic cancer cell line.

Methods: The newly established pancreatic cancer cell line PaCa 5061 was characterized for its morphology, growth rate, chromosomal analysis and mutational analysis of the K-ras, EGFR and p53 genes. Gene-amplification and RNA expression profiles were obtained using an Affymetrix microarray, and overexpression was validated by IHC analysis. Tumorigenicity and spontaneous metastasis formation of PaCa 5061 cells were analyzed in pfp-/-/rag2-/- mice. Sensitivity towards chemotherapy was analysed by MTT assay.

Results: PaCa 5061 cells grew as an adhering monolayer with a doubling time ranging from 30 to 48 hours. M-FISH analyses showed a hypertriploid complex karyotype with multiple numerical and unbalanced structural aberrations. Numerous genes were overexpressed, some of which have previously been implicated in pancreatic adenocarcinoma (GATA6, IGFBP3, IGFBP6), while others were detected for the first time (MEMO1, RIOK3). Specifically highly overexpressed genes (fold change > 10) were identified as EGFR, MUC4, CEACAM1, CEACAM5 and CEACAM6. Subcutaneous transplantation of PaCa 5061 into pfp-/-/rag2-/- mice resulted in formation of primary tumors and spontaneous lung metastasis.

Conclusion: The established PaCa 5061 cell line and its injection into pfp-/-/rag2-/- mice can be used as a new model for studying various aspects of the biology of human pancreatic cancer and potential treatment approaches for the disease.

Show MeSH
Related in: MedlinePlus