Development of bioluminescent chick chorioallantoic membrane (CAM) models for primary pancreatic cancer cells: a platform for drug testing

Abstract

The aim of the present study was to develop chick-embryo chorioallantoic membrane (CAM) bioluminescent tumor models employing low passage cell cultures obtained from primary pancreatic ductal adenocarcinoma (PDAC) cells. Primary PDAC cells transduced with lentivirus expressing Firefly-luciferase (Fluc) were established and inoculated onto the CAM membrane, with >80% engraftment. Fluc signal reliably correlated with tumor growth. Tumor features were evaluated by immunohistochemistry and genetic analyses, including analysis of mutations and mRNA expression of PDAC pivotal genes, as well as microRNA (miRNA) profiling. These studies showed that CAM tumors had histopathological and genetic characteristic comparable to the original tumors. We subsequently tested the modulation of key miRNAs and the activity of gemcitabine and crizotinib on CAM tumors, showing that combination treatment resulted in 63% inhibition of tumor growth as compared to control (p < 0.01). These results were associated with reduced expression of miR-21 and increased expression of miR-155. Our study provides the first evidence that transduced primary PDAC cells can form tumors on the CAM, retaining several histopathological and (epi)genetic characteristics of original tumors. Moreover, our results support the use of these models for drug testing, providing insights on molecular mechanisms underlying antitumor activity of new drugs/combinations.

Monitoring of tumor growth and vascularization with repetitive ultrasonography in the chicken chorioallantoic-membrane-assay

“…Multiple tumor entities have shown to grow sufficiently on the CAM 20 , 30 – 32 . Observable parameters in the field of tumor research with this model are horizontal tumor size and vascularization of the growing tumor 33 .…”

Monitoring of Tumor Growth and Vascularization with Repetitive Ultrasonography in the Chicken Chorioallantoic-Membrane-Assay.

“…2, 52.68% (± 11.89%) of the eggs incubated with tumors showed viability as well as su cient ingrowth of the transplanted tumor on day 11. Tumor take rates reported in scienti c literature show a large variety and range from 70-80% (18,31,43) to much lower inoculation rates of around 45% (44) depending on the speci c tumor entity used for investigation. Unfortunately, in many studies it is di cult to see the true dropout rates as only positive ingrowth rates or treatment effects are reported without any quanti cation of dropout rates.…”

“…Different working groups have published data proclaiming the evaluation of tumor growth using the CAM-assay. In many published articles however, tumor growth is only measured twodimensionally or estimated using surrogate parameters like bioluminescence (31). Furthermore, end-point-analysis in the form of size measuring (45), cell count in ow cytometry of digested tumors, or weight measurements (46) have been utilized or the determination of tumor size.…”

AMP‐kinase inhibitor dorsomorphin reduces the proliferation and migration behavior of colorectal cancer cells by targeting the AKT/mTOR pathway

“…The cells were grown in DMEM containing 10% heat‐inactivated FBS and 1% streptomycin/penicillin. The cells were cultured at 37°C in 5% CO 2 atmosphere and routinely passaged in their exponentially growing phase, at 70% to 80% confluency, by utilizing trypsin–EDTA …”