3D liver model-based surgical education improves preoperative decision-making and patient satisfaction-a randomized pilot trial.

OBJECTIVE: Hepatobiliary surgery bares obstacles to informed consent for the patients due to its complexity and related risk of postoperative complications. 3D visualization of the liver has been proven to facilitate comprehension of the spatial relationship between anatomical structures and to assist in clinical decision-making. Our objective is to utilize individual 3D-printed liver models to enhance patient satisfaction with surgical education in hepatobiliary surgery. DESIGN, SETTING: We conducted a prospective, randomized pilot study comparing 3D liver model-enhanced (3D-LiMo) surgical education against regular patient education during preoperative consultation at the department of Visceral, Thoracic and Vascular Surgery, University Hospital Carl Gustav Carus, Dresden, Germany. PARTICIPANTS: Of 97 screened patients, undergoing hepatobiliary surgery, 40 patients were enrolled from July 2020 to January 2022. RESULTS: The study population (n = 40) was predominantly of male gender (62.5%) with a median age of 65.2 years and a high prevalence of preexisting diseases. Underlying disease, warranting hepatobiliary surgery, was malignancy in the majority of cases (97.5%). Patients in the 3D-LiMo group were more likely to feel very thoroughly educated and exhibited a higher level of satisfaction following surgical education than the control group (80 vs. 55%, n.s.; 90 vs. 65%, n.s.; respectively). Applying 3D models was also associated with enhanced understanding of the underlying disease with regard to amount (100% vs. 70%, p = 0.020) and location of liver masses (95 vs. 65%, p = 0.044). 3D-LiMo patients also demonstrated enhanced understanding of the surgical procedure (80 vs. 55%, n.s.), leading to better awareness for the occurrence of postoperative complications (88.9, vs. 68.4%, p = 0.052). Adverse event profiles were similar. CONCLUSION: In conclusion, individual 3D-printed liver models increase patient satisfaction with surgical education and facilitate patients' understanding of the surgical procedure as well as awareness of postoperative complications. Therefore, the study protocol is feasible to apply to an adequately powered, multicenter, randomized clinical trial with minor modifications.

Prognostic value of DLGAP5 in colorectal cancer.

PURPOSE: DLG7 (disc large homolog 7) is a microtubule-associated protein encoded by DLGAP5 (DLG associated protein 5) gene and has an important role during spindle assembly. Spindle assembly deregulation is a well-known cause of genomic instability. The aim of this study was to investigate the influence of DLGAP5 expression on survival and to evaluate its potential use as a biomarker in colorectal cancer (CRC). METHODS: DLGAP5 expression was measured in the primary tumor and corresponding normal mucosa samples from 109 patients with CRC and correlated to clinical and pathological data. The results were validated in a second, publically available patient cohort. Molecular effects of DLG7/DLGAP5 in CRC were analyzed via functional assays in knockdown cell lines. RESULTS: DLGAP5 downregulation led to a significant reduction of the invasion and migration potential in CRC. In addition, DLGAP5 expression correlates with nodal status and advanced UICC stage (III-IV).Subgroup analyses revealed a correlation between DLGAP5 overexpression and poor survival in patients with non-metastatic disease (M0). Furthermore, overexpression of DLGAP5 is associated with worse overall survival in distinct molecular CRC subtypes. CONCLUSIONS: The results of this study suggest the importance of DLGAP5 in defining a more aggressive CRC phenotype. DLG7/DLGAP5 represents a potential biomarker for CRC in molecular subgroups of CRC.

Circulating Tumor Cells.

In most solid tumors, it is distant metastases rather than the primary tumor which limit the prognosis. Distant metastases are caused by circulating tumor cells (CTCs) which actively invade the blood stream, attach to the endothelium in the target organ, invade the surrounding parenchyma, and form new tumors. Among many other capabilities such as migration or immune escape, CTCs require tumor-forming capacities and can therefore be considered stem cell-like cells. This chapter describes the enrichment and isolation of live CTCs from clinical blood samples for molecular characterization and other downstream applications.

Isolation of Circulating Tumor Cells in an Orthotopic Mouse Model of Colorectal Cancer.

Despite the advantages of easy applicability and cost-effectiveness, subcutaneous mouse models have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Orthotopic mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in hollow organs such as the large bowel. In order to produce uniform tumors which reliably grow and metastasize, standardized techniques of tumor cell preparation and injection are critical. We have developed an orthotopic mouse model of colorectal cancer (CRC) which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor cells from either primary tumors, 2-dimensional (2D) cell lines or 3-dimensional (3D) organoids are injected into the cecum and, depending on the metastatic potential of the injected tumor cells, form highly metastatic tumors. In addition, CTCs can be found regularly. We here describe the technique of tumor cell preparation from both 2D cell lines and 3D organoids as well as primary tumor tissue, the surgical and injection techniques as well as the isolation of CTCs from the tumor-bearing mice, and present tips for troubleshooting.

A Genetically Engineered Mouse Model of Sporadic Colorectal Cancer.

Despite the advantages of easy applicability and cost-effectiveness, colorectal cancer mouse models based on tumor cell injection have severe limitations and do not accurately simulate tumor biology and tumor cell dissemination. Genetically engineered mouse models have been introduced to overcome these limitations; however, such models are technically demanding, especially in large organs such as the colon in which only a single tumor is desired. As a result, an immunocompetent, genetically engineered mouse model of colorectal cancer was developed which develops highly uniform tumors and can be used for tumor biology studies as well as therapeutic trials. Tumor development is initiated by surgical, segmental infection of the distal colon with adeno-cre virus in compound conditionally mutant mice. The tumors can be easily detected and monitored via colonoscopy. We here describe the surgical technique of segmental adeno-cre infection of the colon, the surveillance of the tumor via high-resolution colonoscopy and present the resulting colorectal tumors.

LDB1 overexpression is a negative prognostic factor in colorectal cancer.

BACKGROUND: Colorectal cancer (CRC) is the third most common cancer in western countries and is driven by the Wnt signaling pathway. LIM-domain-binding protein 1 (LDB1) interacts with the Wnt signaling pathway and has been connected to malignant diseases. We therefore aimed to evaluate the role of LDB1 in CRC. RESULTS: Overexpression of LDB1 in CRC is associated with strikingly reduced overall and metastasis free survival in all three independent patient cohorts. The expression of LDB1 positively correlates with genes involved in the Wnt signaling pathway (CTNNB1, AXIN2, MYC and CCND1). Overexpression of LDB1 in CRC cell lines induced Wnt pathway upregulation as well as increased invasivity and proliferation. Upon separate analysis, the role of LDB1 proved to be more prominent in proximal CRC, whereas distal CRC seems to be less influenced by LDB1. MATERIALS AND METHODS: The expression of LDB1 was measured via RT-qPCR in 59 clinical tumor and normal mucosa samples and correlated to clinical end-points. The role of LDB1 was examined in two additional large patient cohorts from publicly available microarray and RNAseq datasets. Functional characterization was done by lentiviral overexpression of LDB1 in CRC cell lines and TOP/FOP, proliferation and scratch assays. CONCLUSIONS: LDB1 has a strong role in CRC progression, confirmed in three large, independent patient cohorts. The in vitro data confirm an influence of LDB1 on the Wnt signaling pathway and tumor cell proliferation. LDB1 seems to have a more prominent role in proximal CRC, which confirms the different biology of proximal and distal CRC.

Circulating tumor cells exhibit stem cell characteristics in an orthotopic mouse model of colorectal cancer.

The prognosis of colorectal cancer (CRC) is closely linked to the occurrence of distant metastases, which putatively develop from circulating tumor cells (CTCs) shed into circulation by the tumor. As far more CTCs are shed than eventually metastases develop, only a small subfraction of CTCs harbor full tumorigenic potential. The aim of this study was to further characterize CRC-derived CTCs to eventually identify the clinically relevant subfraction of CTCs.We established an orthotopic mouse model of CRC which reliably develops metastases and CTCs. We were able to culture the resulting CTCs in vitro, and demonstrated their tumor-forming capacity when re-injected into mice. The CTCs were then subjected to qPCR expression profiling, revealing downregulation of epithelial and proliferation markers. Genes associated with cell-cell adhesion (claudin-7, CD166) were significantly downregulated, indicating a more metastatic phenotype of CTCs compared to bulk tumor cells derived from hepatic metastases. The stem cell markers DLG7 and BMI1 were significantly upregulated in CTC, indicating a stem cell-like phenotype and increased capacity of tumor formation and self-renewal. In concert with their in vitro and in vivo tumorigenicity, these findings indicate stem cell properties of mouse-derived CTCs.In conclusion, we developed an orthotopic mouse model of CRC recapitulating the process of CRC dissemination. CTCs derived from this model exhibit stem-cell like characteristics and are able to form colonies in vitro and tumors in vivo. Our results provide new insight into the biology of CRC-derived CTCs and may provide new therapeutic targets in the metastatic cascade of CRC.

Immune escape and survival mechanisms in circulating tumor cells of colorectal cancer.

The prognosis of colorectal cancer is closely linked to the occurrence of distant metastases. Systemic dissemination is most likely caused by circulating tumor cells (CTC). Despite the fundamental role of CTC within the metastatic cascade, technical obstacles have so far prevented detailed genomic and, in particular, phenotypic analyses of CTC, which may provide molecular targets to delay or prevent distant metastases. We show here a detailed genomic analysis of single colorectal cancer-derived CTC by array comparative genomic hybridization (aCGH), mutational profiling, and microsatellite instability (MSI) analysis. Furthermore, we report the first gene expression analysis of manually selected colorectal cancer-derived CTC by quantitative real-time PCR (qRT-PCR) to investigate transcriptional changes, enabling CTC to survive in circulation and form distant metastases. aCGH confirmed the tumor cell identity of CellSearch-isolated colorectal cancer-derived CTC. Mutational and MSI analyses revealed mutational profiles of CTC to be similar, but not identical to the corresponding tumor tissue. Several CTC exhibited mutations in key genes such as KRAS or TP53 that could not be detected in the tumor. Gene expression analyses revealed both a pronounced upregulation of CD47 as a potential immune-escape mechanism and a significant downregulation of several other pathways, suggesting a dormant state of viable CTC. Our results suggest mutational heterogeneity between tumor tissue and CTC that should be considered in future trials on targeted therapy and monitoring of response. The finding of upregulated immune-escape pathways, which may be responsible for survival of CTC in circulation, could provide a promising target to disrupt the metastatic cascade in colorectal cancer. Cancer Res; 74(6); 1694-704. ©2014 AACR.