NT2 / NTERA-2 Xenograft Model

NT2 / NTERA-2 Xenograft Model

Testicular cancer is the most common cancer found in young men. While many embryonal carcinoma and testicular teratocarcinoma cells respond well to platinum-based treatments such as cisplatin, negative side effects limit dose and success. Symptoms of testicular cancer include a lump on either testicle or a feeling of heaviness of the scrotum. The NT2 cell line in 1980 was established from a metastatic lung tumor of a patient, 22 years of age, Caucasian, male, and diagnosed with malignant testicular pluripotent embryonal carcinoma. The NT2 has been used in many testicular embryonal carcinoma research studies. In 2015 the Yao et al. Oncotarget study used NT2 cells in their investigation into the role of peroxisome proliferator-activated receptor (PPAR) β/δ in testicular embryonal carcinoma. It was found that overexpression of PPAR β/δ resulted in enhanced proliferation, migration, invasion, and tumor xenograft tumor growth which occurred in a retinoic acid receptor and metrix metalloproteinase-2 dependent mechanism. Differentiation (1988) published a study by Mavilio et al. using the NT2 cell line to study the role of homeobox gene clusters in the differentiation of human embryonal carcinoma cells. It was found that, when using HMBA, RA, and BUdR,  only RA-induced differentiation resulted in homeobox-gene activation. Finally, Tripathi et al. (Molecular Cancer Therapeutics, 2011) used NT2 cells to evaluate the anticancer activity of a cisplatin with fisetin combination therapy in human embryonal carcinoma. In vivo effects were also monitored with an NT2 xenograft model. Combination treatment resulted in increased apoptosis (upregulation of 4 caspases and Bak), increased p21 and p53 levels, decreased cyclin B1 and survivin. Overall, data suggests that cisplatin and fisetin combination therapy demonstrates antitumor activity through a mitochondrial and cell death receptor mediated mechanism. The NT2 cell line is used to create the CDX (Cell Line Derived Xenograft) NT2 xenograft mouse model. The NT2 xenograft model has been used to investigate testicular cancer characteristics and therapies.

Basic Study Design

  1. NT2 cells are maintained in exponential growth phase under aseptic conditions.
  2. Cells are trypsinized and cell count viability is determined using a trypan blue exclusion assay (98% of cell viability is required).  NT2 cell suspension is adjusted to appropriate density.
  3. Each mouse is singly subcutaneously injected into the right flank with 106 cells in 100 µL of a Matrigel- NT2 cell suspension.
  4. The injection sites are palpated up to three times weekly until tumors are established to an average size of 50-150 mm3 as measured via digital calipers.
  5. Animals are randomized into treatment groups. Administration of test compound is performed according to the pre-established treatment schedule.
  6. Mice weights are measured and recorded 2-3 times weekly; tumors are measured and recorded daily.
  7. End of study is reached when tumor size reaches 2,000 mm3 or the predetermined size limit per approved IACUC protocol.
  8. Final necropsy and tissue collections are performed for appropriate downstream analysis. Tumors are excised, weighed and documented by digital imaging. Tumors and tissues can be stabilized in RNAlater, snap frozen in LN2 or prepared for histology