Noninvasive Assessment of Gene Transfer and Expression by In Vivo Functional and Morphologic Imaging in a Rabbit Tumor Model

Abstract

PurposeTo evaluate the importance of morphology in quantifying expression after in vivo gene transfer and to compare gene expression after intra-arterial (IA) and intra-tumoral (IT) delivery of adenovirus expressing a SSTR2-based reporter gene in a large animal tumor model.Materials and MethodsTumor directed IA or IT delivery of adenovirus containing a human somatostatin receptor type 2A (Ad-CMV-HA-SSTR2A) gene chimera or control adenovirus (Ad-CMV-GFP) was performed in VX2 tumors growing in both rabbit thighs. Three days later, 111In-octreotide was administered intravenously after CT imaging using a clinical scanner. 111In-octreotide uptake in tumors was evaluated the following day using a clinical gamma-camera. Gene expression was normalized to tumor weight with and without necrosis. This procedure was repeated on nine additional rabbits to investigate longitudinal gene expression both 5 days and 2 weeks after adenovirus delivery. CT images were used to evaluate tumor morphology and excised tissue samples were analyzed to determine 111In-octreotide biodistribution ex vivo.ResultsVX2 tumors infected with Ad-CMV-HA-SSTR2 had greater 111In-octreotide uptake than with control virus (P<0.05). Intra-arterial and intra-tumoral routes resulted in similar levels of gene expression. Longitudinally, expression appeared to wane at 2 weeks versus 5 days after delivery. Areas of necrosis did not demonstrate significant uptake ex vivo. Morphology identified areas of necrosis on contrast enhanced CT and upon excluding necrosis, in vivo biodistribution analysis resulted in greater percent injected dose per gram (P<0.01) and corresponded better with ex vivo biodistribution(r = 0.72, P<0.01, Coefficient of the x-variable = .72) at 2 weeks than without excluding necrosis (P<0.01).ConclusionTumor specificity and high transgene expression can be achieved in tumors via both tumor directed intra-arterial and intra-tumoral delivery in a large animal tumor model. Using clinical machines, morphologic imaging contributes to functional imaging for quantifying SSTR2-based reporter expression in vivo.

Angiotensin II increases gene expression after selective intra-arterial adenovirus delivery in a rabbit model assessed using in vivo SSTR2-based reporter imaging

“…When needed, they were thawed, suspended in DMEM/F12 containing 10 % FBS and gentamicin, and then drawn into a 1-cc syringe with an 18-gauge needle. A total of 0.6 ml of homogenized VX2 tumor cells were implanted in both thighs of each experimental rabbit [ 1 ]. All animals received humane care in compliance with the Institutional Animal Care and Use Committee of The University of Texas MD Anderson Cancer Center.…”

“…A total of 16 tumors from eight rabbits were utilized for this study. After 10–12 days, multi-slice helical computed tomography (CT) imaging without and with intravenous contrast enhancement was performed in a supine position to confirm tumor formation and to calculate tumor weight and volume [ 1 ].…”

“…The following day, planar gamma camera imaging was performed. The amount of 111 In-octreotide in each tumor was determined from the planar images and normalized to the tumor weight (%ID/g) calculated from the CT images [ 1 ]. After imaging, the rabbits were sacrificed with an intravenous overdose of Beuthanasia-D. Portions of tumor or organs were dissected and weighed; the amount of radioactivity was measured using a γ-counter to determine 111 In-octreotide biodistribution in %ID/g.…”

“…Arterial delivery may be used to direct the vector to the organ or tumor of interest, particularly when delivery is directed via a catheter to the primary artery feeding the tumor. Even with this approach, expression may not be greater than intra-tumoral delivery [ 1 ]. Whether pharmacologic manipulation can increase expression upon intra-arterial adenovirus delivery needs exploration.…”

“…In such models, the small vessel size restricts catheter-based access and vascular manipulation. The large animal VX2 rabbit tumor model allows catheterization [ 1 ] using human catheters, vascular manipulation, and imaging using clinical machines and thus can be used to study the effect of pharmacologic vascular manipulation like AII on gene expression. Gene therapy has been hampered by low levels of gene expression upon in vivo delivery.…”