Comparing Metastatic Clear Cell Renal Cell Carcinoma Model Established in Mouse Kidney and on Chicken Chorioallantoic Membrane

Abstract

Metastatic clear cell renal cell carcinoma (ccRCC) is the most common subtype of kidney cancer. Localized ccRCC has a favorable surgical outcome. However, one third of ccRCC patients will develop metastases to the lung, which is related to a very poor outcome for patients. Unfortunately, no therapy is available for this deadly stage, because the molecular mechanism of metastasis remains unknown. It has been known for 25 years that the loss of function of the von Hippel-Lindau (VHL) tumor suppressor gene is pathognomonic of ccRCC. However, no clinically relevant transgenic mouse model of ccRCC has been generated. The purpose of this protocol is to introduce and compare two newly established animal models for metastatic ccRCC. The first is renal implantation in the mouse model. In our laboratory, the CRISPR gene editing system was utilized to knock out the VHL gene in several RCC cell lines. Orthotopic implantation of heterogeneous ccRCC populations to the renal capsule created novel ccRCC models that develop robust lung metastases in immunocompetent mice. The second model is the chicken chorioallantoic membrane (CAM) system. In comparison to the mouse model, this model is more time, labor, and cost-efficient. This model also supported robust tumor formation and intravasation. Due to the short 10 day period of tumor growth in CAM, no overt metastasis was observed by immunohistochemistry (IHC) in the collected embryo tissues. However, when tumor growth was extended by two weeks in the hatched chicken, micrometastatic ccRCC lesions were observed by IHC in the lungs. These two novel preclinical models will be useful to further study the molecular mechanism behind metastasis, as well as to establish new, patient-derived xenografts (PDXs) toward the development of novel treatments for metastatic ccRCC. INTRODUCTION:Renal cell carcinoma (RCC) is the 7 th most common cancer in the United States. Annually, 74,000 Americans are estimated to be newly diagnosed, accounting for more than 14,000 deaths (http://seer.cancer.gov/statfacts/html/kidrp.html). Clear-cell histological subtype, or ccRCC, is the most common subtype, accounting for approximately 80% of RCC cases. Patients with localized malignancy are treated with nephrectomy and have a favorable 5-year survival rate of 73% 1 . However, 25%-30% of patients develop distant metastases to vital organs such as the lungs, resulting in a poor mean survival of 13 months and 5-year

The miRNA-21-5p Payload in Exosomes from M2 Macrophages Drives Tumor Cell Aggression via PTEN/Akt Signaling in Renal Cell Carcinoma

“…Our experience has shown that the CAM model is an efficient, time-saving in vivo model, especially for establishing human RCC xenografts [ 35 , 36 ]. Furthermore, it faithfully recapitulates the metastatic behaviors of the mouse model [ 37 , 38 ]. Here, we utilized fertilized duck eggs rather than the traditional chicken eggs because they offered a more extended tumor growth/incubation period (up to a maximum of 18 days vs. 12 days) to assess metastatic potential [ 39 ].…”

“…To assess RCC xenografts metastasis in vivo, we established the CAM model with fertilized duck eggs as our previously published protocols [ 35 , 36 , 37 , 38 ]. Freshly laid fertilized duck eggs were incubated in a MultiQuip incubator (MULTIQUIP, E2, AISTRAL, Australia) at 37 °C with 60% humidity.…”

PET/MR Imaging of a Lung Metastasis Model of Clear Cell Renal Cell Carcinoma with (2S,4R)-4-[18F]Fluoroglutamine

“…The massive redundancy of transporters responsible for glutamine uptake underscores the importance of this metabolite and the challenge of blocking its uptake in tumors. All these data suggest that [ 18 F]FGln uptake is a read-out of the Most RCC lung metastasis models are generated from direct injection of RENCA or other RCC cells [32,33] into the kidney of mice. We report one of the first direct orthotopic mouse models of ccRCC lung lesions.…”

Exosomes from Von Hippel-Lindau-Null Cancer Cells Promote Metastasis in Renal Cell Carcinoma

“…This differs from clinical RCC in humans, where lung metastasis is the most common metastatic site [13,14]. This difference is because embryonic lungs have not inflated and are less vascularized than the liver prior to hatching; consequently, it is less likely to be a metastatic site [38].…”