Real-time MR imaging with Gadoteridol predicts distribution of transgenes after convection-enhanced delivery of AAV2 vectors

Abstract

Gene therapies that utilize convention-enhanced delivery (CED) will require close monitoring of vector infusion in real time and accurate prediction of drug distribution. The magnetic resonance imaging (MRI) contrast agent, Gadoteridol (Gd), was used to monitor CED infusion and to predict the expression pattern of glial cell line-derived neurotrophic factor (GDNF) protein after administration of adeno-associated virus type 2 (AAV2) vector encoding human pre-pro-GDNF complementary DNA. The nonhuman primate (NHP) thalamus was utilized for modeling infusion to allow delivery of volumes more relevant to planned human studies. AAV2 encoding human aromatic L-amino acid decarboxylase (AADC) was coinfused with AAV2-GDNF/Gd to confirm regions of AAV2 transduction versus extracellular GDNF diffusion. There was a close correlation between Gd distribution and GDNF or AADC expression, and the ratios of expression areas of GDNF or AADC versus Gd were both close to 1. Our data support the use of Gd and MRI to monitor AAV2 infusion via CED and to predict the distribution of GDNF protein after AAV2-GDNF administration.

Figures

Figure 1

T1-weighted MR images with Gd RCD and 3D construction of ROI. (a–e) A series of real-time T1-weighted MR images in the coronal plane obtained at various times from the beginning to the end of infusion into the thalamus of one NHP. The volume of infusate (Vi) at the corresponding infusion time point is indicated at the bottom of each panel. Bar = 0.5 cm. (f) A 3D reconstruction of ROI based on Gd signal in the left thalamus after infusion. The volume of Gd distribution (Vd) is indicated at the bottom of the panel. AAV2, adeno-associated virus type 2; Gd, Gadoteridol; MR images, magnetic resonance images; NHP, nonhuman primate; RCD, real-time convective delivery; ROI, region of interest.

Figure 2

MRI correlation with histology in primate #1 with bilateral infusion of AAV2-GDNF into the thalamus. (a) A T1-weighted MR image of Gd distribution in the thalamus (green outline). Areas staining positive for GDNF (orange outline) of corresponding sections were transferred to the MR image for comparison. Since the left and right infusions were completed by different times, the final series of MR images for each infusion was cropped and merged in a. Infusion volume to the left and right brain was indicated at the bottom of the panel [Vi (L) and Vi (R)]. Bar = 0.5 cm. (b) Coronal histology of primate brain imaged in a shows GDNF staining in a pattern similar to that noted on MRI with Gd. Bar = 1 cm. (c) High magnification of boxed insert in b, showing GDNF-positive cells within the thalamus. Bar = 50 µm. (d,e) Areas of Gd distribution and GDNF expression on the (d) left and (e) right side of the brain in a series of MR images. AAV2, adeno-associated virus type 2; Gd, Gadoteridol; GDNF, glial cell line-derived neurotrophic factor; MRI, magnetic resonance imaging; r, correlation coefficient; ROI, region of interest.

Figure 3

MRI correlation with histology in primate #2 with unilateral coinfusion of AAV2-GDNF and AAV2-AADC into the thalamus. (a) T1-weighted MR image showing Gd distribution in the thalamus (green outline). Areas staining positive for GDNF (orange outline) and AADC (blue outline) of corresponding sections were transferred to the MR image for comparison. Bar = 0.5 cm. (b) Coronal section of primate brain imaged in a, showing GDNF staining in a pattern similar to that noted on MRI with Gd. Bar = 1 cm. (c) AADC-stained section adjacent to (b), showing both endogenous and transduced AADC expression. Transduced AADC is outlined in blue. (e) AADC and TH colabeled section adjacent to (c), showing costaining for AADC in brown and tyrosine hydroxylase (TH) in red to differentiate endogenous AADC/TH (dark red) from transduced AADC (brown). The expression pattern of transduced AADC is nearly identical to GDNF expression in b. (e) High magnification of boxed insert in c showing endogenous AADC-positive cells in the nigra. Bar = 50 µm. (f) High magnification of boxed insert in d showing AADC/TH-positive cells in the nigra. Bar = 50 µm. (g) High magnification of boxed insert in c showing endogenous AADC-positive fibers in the putamen. Bar = 50 µm. (h) High magnification of boxed insert in c showing AADC-positive cells in the putamen. Bar = 50 µm. (i) High magnification of boxed insert in d showing AADC-positive cells in the thalamus. Bar = 50 µm. (j) The areas of Gd, GDNF, and AADC distribution on the right side of the brain in a series of MR images. AADC, aromatic -amino acid decarboxylase; AAV2, adeno-associated virus type 2; Gd, Gadoteridol; GDNF, glial cell line-derived neurotrophic factor; MRI, magnetic resonance imaging; r1, correlation coefficient between areas of Gd and GDNF expression; r2, correlation coefficient between areas of Gd and AADC expression; r3, correlation coefficient between areas of GDNF and AADC expression.

Figure 4

MRI correlation with histology in primate #3 with bilateral coinfusion of AAV2-GDNF and AAV2-AADC into the thalamus. (a) T1-weighted MR image showing Gd distribution in the thalamus, outlined in green. Areas staining positive for GDNF (outlined in orange) and AADC (outlined in blue) of corresponding sections were transferred to the MR image for comparison. Bar = 0.5 cm. (b) Coronal section of primate brain imaged in a, showing GDNF staining in a pattern similar to that noted on MRI with Gd. Bar =1 cm. (c) AADC and TH colabeled section adjacent to (b), showing costaining for AADC in brown and tyrosine hydroxylase (TH) in red. (d, e) Areas of Gd, GDNF, and AADC distribution on the (d) left and (e) right side of the brain in a series of MR images. AADC, aromatic -amino acid decarboxylase; AAV2, adeno-associated virus type 2; Gd, Gadoteridol; GDNF, glial cell line-derived neurotrophic factor; MRI, magnetic resonance imaging; r1, correlation coefficient between areas of Gd and GDNF expression; r2, correlation coefficient between areas of Gd and AADC expression; r3, correlation coefficient between areas of GDNF and AADC expression.