A human brainstem glioma xenograft model enabled for bioluminescence imaging

Abstract

Despite the use of radiation and chemotherapy, the prognosis for children with diffuse brainstem gliomas is extremely poor. There is a need for relevant brainstem tumor models that can be used to test new therapeutic agents and delivery systems in pre-clinical studies. We report the development of a brainstem-tumor model in rats and the application of bioluminescence imaging (BLI) for monitoring tumor growth and response to therapy as part of this model. Luciferase-modified human glioblastoma cells from five different tumor cell sources (either cell lines or serially-passaged xenografts) were implanted into the pontine tegmentum of athymic rats using an implantable guide-screw system. Tumor growth was monitored by BLI and tumor volume was calculated by three-dimensional measurements from serial histopathologic sections. To evaluate if this model would allow detection of therapeutic response, rats bearing brainstem U-87 MG or GS2 glioblastoma xenografts were treated with the DNA methylating agent temozolomide (TMZ). For each of the tumor cell sources tested, BLI monitoring revealed progressive tumor growth in all animals, and symptoms caused by tumor burden were evident 26–29 days after implantation of U-87 MG, U-251 MG, GBM6, and GBM14 cells, and 37–47 days after implantation of GS2 cells. Histopathologic analysis revealed tumor growth within the pons in all rats and BLI correlated quantitatively with tumor volume. Variable infiltration was evident among the different tumors, with GS2 tumor cells exhibiting the greatest degree of infiltration. TMZ treatment groups were included for experiments involving U-87 MG and GS2 cells, and in each case TMZ delayed tumor growth, as indicated by BLI monitoring, and significantly extended survival of animal subjects. Our results demonstrate the development of a brainstem tumor model in athymic rats, in which tumor growth and response to therapy can be accurately monitored by BLI. This model is well suited for pre-clinical testing of therapeutics that are being considered for treatment of patients with brainstem tumors.

Inhibition of DNA damage repair by the CDK4/6 inhibitor palbociclib delays irradiated intracranial atypical teratoid rhabdoid tumor and glioblastoma xenograft regrowth

“…Paraformaldehyde-fixed brains were paraffin-embedded and sectioned (10 mm) for hematoxylin-eosin and TUNEL staining (DeadEnd Colorimetric TUNEL system, Promega) as previously described. 21 To assess treatment effects on tumor cell proliferation, mitotic indices were calculated by counting mitoses per field, over 10 fields per tumor, at 400× (each visual field at 400× ¼ 0.22 mm 2 ). Likewise, apoptoses were quantified by counting TUNEL-positive cells per field, over 10 fields per tumor, at 400×.…”

Efficacy of vincristine administered via convection-enhanced delivery in a rodent brainstem tumor model documented by bioluminescence imaging

“…These rats were euthanized with CO 2 at 1, 3,5,7,9,11,13,15,17, and 18 days postinoculation. BLI was measured prior to euthanization.…”

“…BLI allows for immediate assessment of tumor response to treatment and permits repeated imaging in the same animal. BLI has been used to monitor glioma growth and the efficacy of drug treatments; however, there is limited information on BLI to monitor brainstem glioma biometrics [4,15].…”

Establishing Intracranial Brain Tumor Xenografts With Subsequent Analysis of Tumor Growth and Response to Therapy using Bioluminescence Imaging

“…The number of cells injected is variable dependent on neuroanatomical location of injection. For supratentorial injections we routinely inject 3-5 x 10 5 cells in 3 μL of serum-free media (DMEM), whereas for brainstem injections 3 , as few as 5 x 10 4 cells are injected in 0.5 μL. Injecting larger volumes than recommended can result in tumor cell reflux through the needle tract, with resultant exophytic (Figure 1), rather than intracranial tumor growth.…”

“…The following represents the procedure we use for intracerebral tumor establishment 2 in a neuroanatomical location at which many brain tumor patients experience tumor development. Other locations of interest in brain tumor research include the pons, for anatomic modeling of brainstem tumors 3 , and subdural injections for modeling the location of meningeal tumors 4 . Prior to tumor cell injection, use a sterile 25 gauge sharp needle to puncture the skull at 2 mm to the right of the bregma and 1 mm anterior to the coronal suture, thereby creating an opening for the injection of tumor cells (see video).…”