In vivo and in vitro propagation of intraductal papillary mucinous neoplasms

Abstract

Background Intraductal papillary mucinous neoplasms (IPMNs) are one of the 3 known curable precursor lesions of invasive pancreatic ductal adenocarcinoma, an almost uniformly fatal disease. Cell lines from IPMNs and their invasive counterparts should be valuable to identify gene mutations critical to IPMN carcinogenesis, and permit high-throughput screening to identify drugs that cause regression of these lesions. Methods To advance the study of the biological features of IPMNs, we attempted in vivo and in vitro growth of selected IPMNs based on the hypothesis that IPMNs could be grown in the most severely immunodeficient mice. We examined fourteen cases by implanting them into nude, severe combined immunodeficient (SCID), and NOD/SCID/IL2Rγnull (NOG) mice, in addition to direct culture, to generate tumor xenografts and cell lines. One sample was directly cultured only. Results Thirteen tumors were implanted into the 3 types of mice, including 10 tumors implanted into the triple immunodeficient NOG mice, where the majority (8 of 10) grew. This included 5 IPMNs lacking an invasive component. One of the explanted IPMNs, with an associated invasive carcinoma, was successfully established as a cell line. Tumorigenicity was confirmed by growth in soft agar, growth in immunodeficient mice, and the homozygous deletion of p16/cdkn2a. Epithelial differentiation of the cell line was documented by cytokeratin expression. Patient origin was confirmed using DNA fingerprinting. Conclusions Most non-invasive IPMNs grow in NOG mice. We successfully established one IPMN cell line, and plan to use it to clarify the molecular pathogenesis of IPMNs.

Cystic precursors to invasive pancreatic cancer

“…43 Similarly, the ability of IPMNs, including non-invasive lesions, to engraft in vivo has been confirmed by the successful establishment of eight xenografts from patient tumors directly implanted in highly immunodeficient (NOD/SCID/IL2Rγ 0 ) mice; one of the explanted IPMNs was then established as an immortalized cell line. 73 Finally, the first ex vivo model of MCN, denoted as MCC1 has been created. MCC1 is a cell line established from a noninvasive lesion with high-grade dysplasia.…”

Loss of expression of the SWI/SNF chromatin remodeling subunit BRG1/SMARCA4 is frequently observed in intraductal papillary mucinous neoplasms of the pancreas

“…To further substantiate our immunohistochemical findings in tissue sections, Western blot analysis was subsequently performed, to evaluate Brg1 expression in the low passage non-invasive IPMN cell line, IPMN 1T 29 . As a positive control for a cancer with known somatic mutation of BRG1 , we used the Pa03C line, as recently described in the pancreatic cancer genome sequencing effort 26 .…”

“…In order to extrapolate the tissue studies to cultured cell lines, especially in a recently described in vitro IPMN model 29 , we assessed Brg1 expression in IPMN and PDAC cells. When compared with non-neoplastic HPNE cells, IPMN 1T showed a marked decrease in Brg1 expression, which was comparable to the “positive” control Pa03C cells, that are known to harbor a somatic mutation of BRG1 .…”

“…The derivation of low passage IPMN 1T cells has recently been described 29 . The cells were maintained in MEM with 20% FBS (GIBCO), 5 ng/ml EGF (Promega), 0.2 U/ml human recombinant insulin (Invitrogen), and 1% penicillin streptomycin (Invitrogen).…”

Molekulare Pathogenese zystischer Neoplasien im Pankreas

“…Insgesamt liegen für den Großteil der bisher identifizierten differenziell exprimierten oder mutierten Gene keine experimentellen Daten vor, die eine direkte funktionelle Rolle in der Induktion oder Progression von IPMN belegen. Neben dem bereits erwähnten transgenen Tiermodell stehen mittlerweile auch Xenograft-und Zellkulturmodelle von humanen IPMN zur Verfügung [20], die die experimentelle Aufarbeitung der bisher erzielten Ergebnisse erleichtern sollten.…”