Earliest Steps in Primary Tumor Formation and Micrometastasis Resolved with Histochemical Markers of Gene-tagged Tumor Cells

Culp, Lloyd A.; Lin, Wen-chang; Kleinman, N. R.; O’Connor, Kathleen L.; Lechner, Roseanna

doi:10.1177/002215549804600501

Cited by 12 publications

(8 citation statements)

References 15 publications

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“…Can an immune response against the reporter explain the absence of metastases of primary 4T1luc2, but not of 4T1luc2D6 tumors? Micro-metastases may be formed very early, during the first week after the implantation of tumor cells 43 , 44 . Studies in a mouse model of melanoma have also shown that the malignant cells are disseminating in the early stage of primary tumor development, even before it becomes detectable 45 .…”

Section: Discussionmentioning

confidence: 99%

Luciferase Expression Allows Bioluminescence Imaging But Imposes Limitations on the Orthotopic Mouse (4T1) Model of Breast Cancer

Baklaushev

Kilpeläinen

Petkov

et al. 2017

Sci Rep

105

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Implantation of reporter-labeled tumor cells in an immunocompetent host involves a risk of their immune elimination. We have studied this effect in a mouse model of breast cancer after the orthotopic implantation of mammary gland adenocarcinoma 4T1 cells genetically labelled with luciferase (Luc). Mice were implanted with 4T1 cells and two derivative Luc-expressing clones 4T1luc2 and 4T1luc2D6 exhibiting equal in vitro growth rates. In vivo, the daughter 4T1luc2 clone exhibited nearly the same, and 4T1luc2D6, a lower growth rate than the parental cells. The metastatic potential of 4T1 variants was assessed by magnetic resonance, bioluminescent imaging, micro-computed tomography, and densitometry which detected 100-μm metastases in multiple organs and bones at the early stage of their development. After 3–4 weeks, 4T1 generated 11.4 ± 2.1, 4T1luc2D6, 4.5 ± 0.6; and 4T1luc2, <1 metastases per mouse, locations restricted to lungs and regional lymph nodes. Mice bearing Luc-expressing tumors developed IFN-γ response to the dominant CTL epitope of Luc. Induced by intradermal DNA-immunization, such response protected mice from the establishment of 4T1luc2-tumors. Our data show that natural or induced cellular response against the reporter restricts growth and metastatic activity of the reporter-labelled tumor cells. Such cells represent a powerful instrument for improving immunization technique for cancer vaccine applications.

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Section: Discussionmentioning

confidence: 99%

Luciferase Expression Allows Bioluminescence Imaging But Imposes Limitations on the Orthotopic Mouse (4T1) Model of Breast Cancer

Baklaushev

Kilpeläinen

Petkov

et al. 2017

Sci Rep

105

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“…Fixation was performed at 4C for 1 hr with 2% (v/v) formaldehyde in PBS. X-gal staining was executed on fixed tissues as described previously at pH 7.4 (overnight at room temperature) to minimize background tissue staining (Lin et al 1990a,b;Kleinman et al 1994;Culp et al 1998b). X-gal staining was also performed on tissues from animals not injected with tumor cells to evaluate possible background staining of tissues.…”

Section: Tissue Isolation and Histochemical Stainingmentioning

confidence: 99%

Tracking Micrometastasis to Multiple Organs with lacZ-tagged CWR22R Prostate Carcinoma Cells

Holleran

Miller

Culp

2000

J Histochem Cytochem.

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Metastasis to organs other than lung is rarely observed in animal model systems of human prostate carcinoma (PCA), with the exception of already metastatic isolates of human PCA cultured for long periods of time. To analyze more directly the evolution of metastatic variants from primary PCA tumor isolates, the lacZ histochemical marker gene was transfected into the CWR22Rv1 cell line isolated from the CWR22R xenograft (primary tumor). Three clones of varying lacZ-expression stability were analyzed for tumorigenicity and progression in athymic nude mice. Clones B and D were highly tumorigenic in the subcutis; however, lacZ expression was highly unstable. In contrast, clone H demonstrated highly stable lacZ expression for >25 passages in culture or in animals. Clone H, injected sc in a PBS vehicle, gave a 15-40% tumorigenic take. All primary tumor-bearing animals exhibited micrometastases in lung and other organs. Clone H injected in a Matrigel vehicle gave 100% tumorigenicity, with all animals displaying micrometastases in lung, liver, and/or bone (lower frequency in brain and kidney). Overall, the relative frequency of micrometastasis to multiple organs was lung>liver=bone>>brain>kidney. Overt metastases were never observed in the lung or bone but were occasionally found in liver. lacZ-transfected clone H CWR22Rv1 cells represent a much more accurate model of metastasis of PCA to the organs normally involved in progression of the human disease. Use of marker gene-tagged cells and other high-resolution molecular techniques will now permit analyses of the earliest events in PCA progression and micrometastasis.

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“…Xenograft animal models not only readily represent the diverse genetics of human neuroblastomas, but also permit experimental or spontaneous metastasis. Xenograft models receiving intravenous injection of neuroblastoma cells [15][16][17][18] exhibit experimental metastasis by bypassing primary tumor growth and intravasation to the circulatory system. On the other hand, xenograft models of spontaneous neuroblastoma metastasis [19][20][21] permit studies of a complete metastatic process, starting from the primary tumor formation at an ectopic (e.g., subcutaneous) or orthotopic (e.g., adrenal) injection site and ending with tumor metastasis at distal organs.…”

Section: Introductionmentioning

confidence: 99%

A Fluorescent Orthotopic Mouse Model for Reliable Measurement and Genetic Modulation of Human Neuroblastoma Metastasis

Henriksson¹,

Almgren²,

Thurlow³

et al. 2004

Clin Exp Metastasis

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Neuroblastoma is the most common extra-cranial solid tumor of infancy and childhood, and majority of patients die from the metastatic disease. Orthotopic xenograft mouse models are valuable tools for improving our understanding and control of neuroblastoma metastasis, because they readily represent genetic diversity and allow spontaneous metastasis. Intra-adrenal injection is commonly used for establishing the orthotopic animal models since human neuroblastoma frequently originates in the adrenal gland. However, it is unclear whether the metastatic potential of neuroblastoma can be reliably determined in adrenally-injected mice because their gland size is so small. In this study, we developed and characterized a fluorescent orthotopic xenograft animal model of NB69-derived human neuroblastoma. By comparing animals receiving adrenal injection and adrenal overlay, with the latter mimicking injection spillover, we found that the metastatic potential of neuroblastoma can be reliably determined in animal lungs. Furthermore, the lung metastasis can be genetically modulated in these animals. The results also show that the expression of Renilla green fluorescent protein (GFP) was exceptionally stable in NB69 cells, allowing rapid and sensitive detection of lung metastases at the macroscopic level. Additional features of our model include 100% tumor take, a 1-week tumor latency, resemblance to tumor behaviors in neuroblastoma patients, and the ability to monitor the expression of a gene of interest with GFP. This animal model of human neuroblastoma will be useful for studying genes involved in the metastatic process and for evaluating anti-metastasis agents in pre-clinical trials.

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Earliest Steps in Primary Tumor Formation and Micrometastasis Resolved with Histochemical Markers of Gene-tagged Tumor Cells

Cited by 12 publications

References 15 publications

Luciferase Expression Allows Bioluminescence Imaging But Imposes Limitations on the Orthotopic Mouse (4T1) Model of Breast Cancer

Luciferase Expression Allows Bioluminescence Imaging But Imposes Limitations on the Orthotopic Mouse (4T1) Model of Breast Cancer

Tracking Micrometastasis to Multiple Organs with lacZ-tagged CWR22R Prostate Carcinoma Cells

A Fluorescent Orthotopic Mouse Model for Reliable Measurement and Genetic Modulation of Human Neuroblastoma Metastasis

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