Modeling Brain Metastases Through Intracranial Injection and Magnetic Resonance Imaging

Summary

Intracranial brain metastasis modeling is complicated by an inability to monitor tumor size and response to treatment with precise and timely methods. The presented methodology couples intracranial tumor injection with magnetic resonance imaging analysis, which when combined, cultivates precise and consistent injections, enhanced animal monitoring, and accurate tumor volume measurements.

Abstract

Metastatic spread of cancer is an unfortunate consequence of disease progression, aggressive cancer subtypes, and/or late diagnosis. Brain metastases are particularly devastating, difficult to treat, and confer a poor prognosis. While the precise incidence of brain metastases in the United States remains hard to estimate, it is likely to increase as extracranial therapies continue to become more efficacious in treating cancer. Thus, it is necessary to identify and develop novel therapeutic approaches to treat metastasis at this site. To this end, intracranial injection of cancer cells has become a well-established method in which to model brain metastasis. Previously, the inability to directly measure tumor growth has been a technical hindrance to this model; however, increasing availability and quality of small animal imaging modalities, such as magnetic resonance imaging (MRI), are vastly improving the ability to monitor tumor growth over time and infer changes within the brain during the experimental period. Herein, intracranial injection of murine mammary tumor cells into immunocompetent mice followed by MRI is demonstrated. The presented injection approach utilizes isoflurane anesthesia and a stereotactic setup with a digitally controlled, automated drill and needle injection to enhance precision, and reduce technical error. MRI is measured over time using a 9.4 Tesla instrument in The Ohio State University James Comprehensive Cancer Center Small Animal Imaging Shared Resource. Tumor volume measurements are demonstrated at each time point through use of ImageJ. Overall, this intracranial injection approach allows for precise injection, day-to-day monitoring, and accurate tumor volume measurements, which combined greatly enhance the utility of this model system to test novel hypotheses on the drivers of brain metastases.

Introduction

Brain metastases are 10 times more common than adult primary central nervous system tumors¹, and have been reported in almost every solid tumor type with lung cancer, breast cancer, and melanoma exhibiting the highest incidence². Regardless of the primary tumor site, the development of brain metastasis leads to a poor prognosis often associated with cognitive decline, persistent headaches, seizures, behavioral and/or personality changes^1,3,4,5. In terms of breast cancer, there have been many ad....

Protocol

All methods described herein have been approved by the Institutional Animal Care and Use Committee (IACUC) at The Ohio State University (P.I. Gina Sizemore; Protocol #2007A0120). All rodent survival surgery IACUC policies are followed, including use of sterile techniques, supplies, instruments, as well as fur removal and sterile preparation of the incision site.

1. Intracranial injection of breast cancer cells

NOTE: The method described herein utilized the DB7 murine mammary tumor cell line derived from a primary MMTV-PyMT tumor²³. Previous studies have established intracranial inje....

Results

Figure 3 overviews the tumor volume quantification for a single mouse at two time points (day 7 and day 10) post-injection of murine mammary tumor cells. For this experiment, 50,000 DB7 cells were injected, and the animal’s brain was evaluated by MRI. For each scan, 30 slices (0.5 mm thickness) were captured. Evaluation of the 30 slices per scan revealed that at day 7 post-injection, 5 slices exhibited tumor burden (Figure 3A) and at day 10 post-injection,.......

Discussion

The utilization of intracranial injection followed by serial monitoring with MRI provides the unique ability to visualize tumor growth with tumor volume accuracy over time. The application of digital imaging analysis allows for interpretation of brain lesions for tumor volume, hemorrhage, necrosis, and response to treatment.

As with any procedure, there are key steps that must be followed for success. First, careful setup of the stereotactic devices is imperative to the success of this techniq.......

Materials

Name	Company	Catalog Number	Comments
Surgical Materials
Betadine	Purdue Products	19-027132	Povidone-iodine, 7.5%
Bone Wax	Surgical Specialities	903	Sterile and malleable beeswax and isopropyl palmitate
Buponorphine SR-Lab	ZooPharm	N/A	Long acting injectable analgesic 5 mL (0.5 mg/mL) polymetric formulation
Cotton tip applicators	Puritan	25-806 10WC	Sterile long stemmed cotton tip applicators
Eye Ointment	Puralube	17033-211-38	Lubricating petrolatum and mineral oil based ophthalmic ointment
Handwarmers	Hothands	HH2	Air-activated heat packs
Ibuprofen	Up & Up	094-01-0245	100mg per 5mL in liquid suspension
Isoflurane	Henry Schein INC	1182097	Liquid anesthetic for use in anesthetic vaporizer
Scalpels	Integra Miltex	4-410	#10 disposable scalpel blade
Skin Glue	Vetbond	1469SB	Skin safe wounds adhesive
Sterile Dressing	TIDI Products	25-517	Individually packed sterile drapes
Suture	Covidien	SP5686G	45cm swedged 5-0 monofilament polypropylene suture
Stereotaxic Unit
High Speed Drill (Foredom)	Kopf	Model 1474	Max of 38,000 RPM
Mouse Gas Anesthesia Head Holder	Kopf	Model 923-B	Mouth bar with teeth hole and nosecone
Non-Rupture Ear Bars	Kopf	Model 922	Ear bars suitable for mouse applications
Stereotaxic Instrument	Kopf	Model 940	Base plate, frame and linear scale assembly with digital readout monitor
Injector
Injector Needle and syringe	Hamilton	80366	26 gauge needle, 51 mm needle length and 10 μL volume syringe
Legato 130A automated Syringe Pump	KD Scientific	P/N: 788130	Programmable touch screen base with automated injector
Anesthesia Machine
SomnoSuite Low-Flow Digital Vaporizer	Kent Scientific	SS-01	Digital anesthesia machine
SomnoSuite Starter Kit for mice	Kent Scientific	SOMNO-MSEKIT	Includes induction chamber, 2x anesthesia syringes, 18" tubing, plastic nosecone, 2x waste aneshesia gas canisters