Establishment, Characterization and Chemosensitivity of Three Mismatch Repair Deficient Cell Lines from Sporadic and Inherited Colorectal Carcinomas

Abstract

BackgroundColorectal cancer (CRC) represents a morphologic and molecular heterogenic disease. This heterogeneity substantially impairs drug effectiveness and prognosis. The subtype of mismatch repair deficient (MMR-D) CRCs, accounting for about 15% of all cases, shows particular differential responses up to resistance towards currently approved cytostatic drugs. Pre-clinical in vitro models representing molecular features of MMR-D tumors are thus mandatory for identifying biomarkers that finally help to predict responses towards new cytostatic drugs. Here, we describe the successful establishment and characterization of three patient-derived MMR-D cell lines (HROC24, HROC87, and HROC113) along with their corresponding xenografts.MethodologyMMR-D cell lines (HROC24, HROC87, and HROC113) were established from a total of ten clinicopathological well-defined MMR-D cases (120 CRC cases in total). Cells were comprehensively characterized by phenotype, morphology, growth kinetics, invasiveness, and molecular profile. Additionally, response to clinically relevant chemotherapeutics was examined in vitro and in vivo.Principal FindingsTwo MMR-D lines showing CIMP-H derived from sporadic CRC (HROC24: K-raswt, B-rafmut, HROC87: K-raswt, B-rafmut), whereas the HROC113 cell line (K-rasmut, B-rafwt) was HNPCC-associated. A diploid DNA-status could be verified by flow cytometry and SNP Array analysis. All cell lines were characterized as epithelial (EpCAM+) tumor cells, showing surface tumor marker expression (CEACAM+). MHC-class II was inducible by Interferon-γ stimulation. Growth kinetics as well as invasive potential was quite heterogeneous between individual lines. Besides, MMR-D cell lines exhibited distinct responsiveness towards chemotherapeutics, even when comparing in vitro and in vivo sensitivity.ConclusionsThese newly established and well-characterized, low-passage MMR-D cell lines provide a useful tool for future investigations on the biological characteristics of MMR-D CRCs, both of sporadic and hereditary origin. Additionally, matched patient-derived immune cells allow for comparative genetic studies.

Dual blockade of PI3K/AKT/mTOR (NVP-BEZ235) and Ras/Raf/MEK (AZD6244) pathways synergistically inhibit growth of primary endometrioid endometrial carcinoma cultures, whereas NVP-BEZ235 reduces tumor growth in the corresponding xenograft models

“…Primary cell cultures better represent the histological and genetic heterogeneity of the primary tumor more compared to commercial cell lines [27,28]. Therefore, our results better reflect the response of patients on these treatments compared to models based on immortalized cell lines.…”

“…These cell cultures were established directly from patient tumors and closely resemble the heterogeneity and genomic features of the primary tumor. They better reflect treatmentresponse of patients compared to commercial cell lines [27,28]. In addition, these cultures can be used for in vivo screening of new treatments, by using them to establish subcutaneous (s.c.) xenograft models.…”

Frameshift mutational target gene analysis identifies similarities and differences in constitutional mismatch repair‐deficiency and Lynch syndrome

“…LS‐associated CRC were obtained upon surgery, with informed written patient consent ( n = 10) and collected in our tumor biobank (designated as HROC + serial number) . Tumor pieces were both frozen viable and native.…”

Tumor Take Rate Optimization for Colorectal Carcinoma Patient-Derived Xenograft Models

“…Cell line generation has previously been described in detail [17]. Briefly, PDX tumors were minced; cells were released from surrounding tissue by scraping and passage through a nylon mesh (100  µ m) to obtain single cell suspensions.…”