Published online before print March 21, 2002, 10.1148/radiol.2232010501
(Radiology 2002;223:417.)
A more recent version of this article appeared on May 1, 2002
Gamma Camera Dual Imaging with a Somatostatin Receptor and Thymidine Kinase after Gene Transfer with a Bicistronic Adenovirus in Mice1
Kurt R. Zinn, DVM, PhD,
Tandra R. Chaudhuri, PhD,
Victor N. Krasnykh, PhD,
Donald J. Buchsbaum, PhD,
Natalya Belousova, MS,
William E. Grizzle, MD,
David T. Curiel, MD and
Buck E. Rogers, PhD
1 From the Departments of Radiology (K.R.Z., T.R.C.), Radiation Oncology (D.J.B., B.E.R.), Pathology (W.E.G.), Division of Human Gene Therapy, Departments of Medicine, Surgery, and Pathology (V.N.K., D.T.C.), and Gene Therapy Center (K.R.Z., V.N.K., D.J.B., N.B., D.T.C., B.E.R.), University of Alabama at Birmingham, Boshell Bldg, BDB 11, 1530 3rd Ave S, Birmingham, AL 35294-0012. Received February 21, 2001; revision requested March 29; revision received August 2; accepted September 17. Supported by the National Cancer Institute grants CA80104 and CO97110. Address correspondence to K.R.Z. (e-mail: kurtzinn@uab.edu).
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Figure 1. Diagram illustrates the protocol for plate imaging. A-427 cells were infected with Ad-hSSTr2-TK (#1); after 24 hours, they were mixed with different ratios of uninfected cells (#2) and placed in 12-well plates (six wells per dilution). A total of four plates were used for all dilutions (100%, 75%, 50%, 25%, 10%, 5%, 1%, and 0% of infected cells). As an example, one plate with 100% and 75% of infected cells is shown (#3). Excess unlabeled P2045 and FIAU were added to one-half of the wells for each dilution (#4), the two radiotracers were added (#5), and the plates were imaged by using two window settings on the gamma camera (#6). After incubation for 1 hour (#7), the cells were washed (#8), and final images were collected (#9).
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Figure 2a. Gamma camera images of the cell culture plates. (a) 99mTc gamma camera window shows trapped 99mTc-labeled P2045. (b) 125I gamma camera window shows trapped 125I-labeled FIAU. Images in a and b were collected without moving the plates. The percentages refer to the level of Ad-hSSTr2-TK-positive infected cells. The - and + refer to absence or presence, respectively, of excess unlabeled P2045 and FIAU. Note reduction in trapped 99mTc-labeled P2045 and 125I-labeled FIAU with the addition of excess unlabeled P2045 and FIAU.
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Figure 2b. Gamma camera images of the cell culture plates. (a) 99mTc gamma camera window shows trapped 99mTc-labeled P2045. (b) 125I gamma camera window shows trapped 125I-labeled FIAU. Images in a and b were collected without moving the plates. The percentages refer to the level of Ad-hSSTr2-TK-positive infected cells. The - and + refer to absence or presence, respectively, of excess unlabeled P2045 and FIAU. Note reduction in trapped 99mTc-labeled P2045 and 125I-labeled FIAU with the addition of excess unlabeled P2045 and FIAU.
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Figure 3a. Analyses of internally bound, or trapped, radiotracers in the adherent A-427 cell monolayer. (a) Plot of the percentage dose trapped in the cell monolayers versus the percentage of positive A-427 cells. Data points indicate means. Error bars indicate SD. (b) Plot of trapped 125I-labeled FIAU versus 99mTc-labeled P2045. The highest point (100% of infected cells) was not included because the percentage trapped dose for the 125I-labeled FIAU was not increased compared with 75% of infected cells. This finding indicated a potential saturation for these conditions.
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Figure 3b. Analyses of internally bound, or trapped, radiotracers in the adherent A-427 cell monolayer. (a) Plot of the percentage dose trapped in the cell monolayers versus the percentage of positive A-427 cells. Data points indicate means. Error bars indicate SD. (b) Plot of trapped 125I-labeled FIAU versus 99mTc-labeled P2045. The highest point (100% of infected cells) was not included because the percentage trapped dose for the 125I-labeled FIAU was not increased compared with 75% of infected cells. This finding indicated a potential saturation for these conditions.
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Figure 4a. In vivo simultaneous imaging for hSSTr2 and TK expression. (a) Photograph of the mouse shows tumor locations and adenovirus doses for group A. (b) Photograph of the mouse shows tumor locations and adenovirus doses for group B. The expression of hSSTr2 was depicted with imaging tumor accumulation of 99mTc-labeled P2045 (bottom left), while TK expression was depicted with imaging tumor accumulation of 131I-labeled FIAU (bottom right). The images were obtained 5 hours after intravenous injection of the radiotracers.
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Figure 4b. In vivo simultaneous imaging for hSSTr2 and TK expression. (a) Photograph of the mouse shows tumor locations and adenovirus doses for group A. (b) Photograph of the mouse shows tumor locations and adenovirus doses for group B. The expression of hSSTr2 was depicted with imaging tumor accumulation of 99mTc-labeled P2045 (bottom left), while TK expression was depicted with imaging tumor accumulation of 131I-labeled FIAU (bottom right). The images were obtained 5 hours after intravenous injection of the radiotracers.
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Figure 5. Imaging hSSTr2 expression resulting from low Ad-hSSTr2-TK doses. The 99mTc-labeled P2045 images from two representative mice demonstrate the sensitivity for detection of hSSTr2 expression following gene transfer with low doses of Ad-hSSTr2-TK.
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Figure 6. Plot of tumor uptake for 99mTc-labeled P2045 and 131I-labeled FIAU relative to the injected dose of Ad-hSSTr2-TK (log scale). The tumors were counted individually in a gamma counter at the time mice were killed 24 hours after the injection of radiotracers. Data points indicate means. Error bars indicate SD. There were six tumors per adenovirus dose. There was excellent correlation between the uptake of 99mTc-labeled P2045 (percentage dose per gram) and the adenovirus (Ad) dose (r2 = 0.98). Uptake of 131I-labeled FIAU in tumor was Ad-hSSTr2-TK-dependent but completely unrelated to the dose.
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Figure 7. Immunohistologic images validate hSSTr2 and TK expression in Ad-hSSTr2-TK-injected tumor xenografts. A, Photomicrograph shows strong phenotypic expression of TK (brown color). (Anti-TK polyclonal antibody stain; original magnification, x400.) B, Photomicrograph shows relatively weaker phenotypic expression of hSSTr2 (brown color). (Anti-hSSTr2 polyclonal antibody stain; original magnification, x400.) Matching areas within dotted lines (NT = no tumor) are composed primarily of stroma and inflammatory cells that are negative for TK and hSSTr2 expression. The sections in A and B were anatomically adjacent and showed that expression of both TK and hSSTr2 was limited to the same local areas in the tumor.
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Figure A1. Diagram depicts the construction of Ad-hSSTr2-TK, as described in the Appendix.
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Copyright © 2002 by the Radiological Society of North America.
