An Immunocompetent Murine Model for Laser Interstitial Thermal Therapy of Glioblastoma

Summary

Glioblastoma is a devastating form of primary brain cancer, and laser interstitial thermal therapy is emerging as a promising alternative to conventional surgical resection for inoperable glioblastoma. This protocol describes an optimized pre-clinical mouse model that can be used to study treatment effects or adjuvant and combinatorial treatments.

Abstract

Glioblastoma (GB), the most aggressive form of primary brain cancer, accounts for approximately half of all high-grade primary brain tumors in adults and has no cure. Laser interstitial thermal therapy (LITT) is a Food and Drug Administration (FDA)-approved treatment for GB and is used in patients who may not be candidates for conventional surgical resection. While the clinical efficacy of LITT has been established, research beyond clinical case studies and case series is limited and hindered by the lack of an established animal model. This protocol uses C57BL/6 mice and syngeneic CT2A glioma cancer cell line to closely recapitulate human GB while also using a 1064 nm Neodymium-doped Yttrium Aluminum Garnet (Nd:YAG) laser, such as is used in one of the two FDA-approved LITT systems, providing excellent pre-clinical relevance. The successful establishment of this LITT murine model will provide a valuable platform for investigating the unique features of LITT ablation and its effects on the tumor microenvironment, potentially leading to improved therapeutic strategies.

Introduction

Cancer is the number one leading cause of death in Canada. Glioblastoma (GB), the most common form of aggressive brain tumor, accounts for 48% to 60% of all high-grade primary brain tumors in adults¹. The prognosis for GB is especially grim with a 5-year net survival of 4.8% with conventional treatments, including surgical resection, chemo- and radiotherapy^1,2.

Laser Interstitial Thermal Therapy (LITT) is an FDA-approved procedure using a laser for hyperthermic in-situ tumor ablation in patients with inoperable brain tumors and provides an attractiv....

Protocol

Animal ethics for this protocol was approved by the Animal Care Committee at the University of Manitoba in accordance with the ethical guidelines set by the Canadian Council for Animal Care (CCAC). This protocol used 8-12-week-old C57BL/6 immunocompetent mice and syngeneic glioma cell line CT2A for a preclinical model with a wide range of applications, including experiments focused on histological analysis, immunological changes, or combinatorial therapeutic interventions. The protocol can be easily adapted to other.......

Discussion

There is a fast-expanding body of literature regarding LITT; however, it is primarily limited to human clinical case studies or case series. Indeed, several potential benefits for LITT have been shown, including lower post-operative complication rates and costs while conferring comparable progression-free survival^7,8,9,10,11. There is also a reduction of local.......

Materials

Name	Company	Catalog Number	Comments
Absorbion Spears	FST	18105-01	Hemostatic sponges.
Adson tissue forceps	FST	11006-12
C57BL/6 mice	Jackson Laboratories	Strain #000664	6 to 12 week old male and female
Cotton Tipped Applicators (6")	Electron Microscopy Sciences	72308
CT2A glioma cell line			Generously donated from Dr. Peter Fecci, Duke University.
Cultrex Reduced Growth Factor Basement Membrane Extract, PathClear	Biotechne, R&D systems	3433-010-01
DMEM/F-12, HEPES	Gibco, ThermoFisher Scientific	11330032
Dual-chamber slides	BIO-RAD	1450003
Eppendorf Safe-Lock Tubes 2.0 mL	Eppendorf	22363352
Ethyl Alcohol Anhydrous	Greenfield Global	P016EAAN	Dilute to 70% with ddH2O
Fetal Bovine Serum, qualified, Canada	Gibco, ThermoFisher Scientific	12483020
Glad Press-n-Seal plastic wrap	Amazon.ca	12587704417
High speed drill	Kopf Instruments	Model 1474
K & J Thermocouple temerpature meter	Omega	HH509R
Metacam (meloxicam)	WDDC	114424
Microinjection syringe pump	WPI	UMP3T-1
Microliter syringe (700 Series)	Hamilton	87908	Custom needles are available. A steep needle bevel helps with precise delivery, and a shorter needle length helps with stability.
Needle driver/Needle Holder	FST	12500-12	A fine tip is most suitable due to the confined working space, but many styles are suitable based on handle preference.
Opixcare Plus opthalmic ointment	WDDC	135941
Phosphate Buffered Saline (10x)	Fisher bioreagents	BP399-4	Dilute to 1x with ddH2O
Povidone-iodine	ThermoFisher Scientific	3955-16	Aliquat into into smaller tubes for use with cotton tipped applicators.
Saline (normal)	WDDC	126588
Scalpel, single use (#15 Blade)	Feather	Feather NO15
Scissors, fine surgical	FST	91460-11	Fine student scissors, Iris, or Bonn are all suitable.
Stereotactic frame	Kopf Instruments	Model 940	With digital display console and mouse nose-cone and ear-bars.
Stereotactic syringe holder	Kopf Instruments	Model 1772-F	If not using an injection pump.
Sutures (5-0 monofilament)	Ethicon	MCP463G	Monocryl violet monofilament with reverse cutting tip
Syringe, 28 G  (0.5 mL)	BD	BD 329461	BD Lo-Dose U-100 Insulin Syringes
TC20 Automated Cell Counter	BIO-RAD	1450102
Thermocouple probe, fine diameter (Type K)	Omega	TJM-CA316-IM025G-150
Trypsin-EDTA (0.25%), phenol red	Gibco, ThermoFisher Scientific	25200072
Vetbond by 3M, veterinary tissue glue	WDDC	126125
Wahl Peanut Clippers	WDDC/Wahl	100963	Also available directly from manufacturer.
Warming pad	Bensen Medical	70308/121873	Any similar item can be used.
Webcol Alcohol preps	Electron Microscopy Sciences	71005-20	Alcohol prep wipe, 2-ply, medium size.