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Pinhole micro-SPECT/CT for noninvasive monitoring and quantitation of oncolytic virus dispersion and percent infection in solid tumors.

Penheiter AR, Griesmann GE, Federspiel MJ, Dingli D, Russell SJ, Carlson SK - Gene Ther. (2011)

Bottom Line: The purpose of our study was to validate the ability of pinhole micro-single-photon emission computed tomography/computed tomography (SPECT/CT) to: 1) accurately resolve the intratumoral dispersion pattern and 2) quantify the infection percentage in solid tumors of an oncolytic measles virus encoding the human sodium iodide symporter (MV-NIS).We reliably resolved multiple distinct intratumoral zones of infection from non-infected regions.Pinhole micro-SPECT/CT imaging using the NIS reporter demonstrated precise localization and quantitation of oncolytic MV-NIS infection, and can replace more time-consuming and expensive analyses (for example, autoradiography and IHC) that require animal killing.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA.

ABSTRACT
The purpose of our study was to validate the ability of pinhole micro-single-photon emission computed tomography/computed tomography (SPECT/CT) to: 1) accurately resolve the intratumoral dispersion pattern and 2) quantify the infection percentage in solid tumors of an oncolytic measles virus encoding the human sodium iodide symporter (MV-NIS). Sodium iodide symporter (NIS) RNA level and dispersion pattern were determined in control and MV-NIS-infected BxPC-3 pancreatic tumor cells and mouse xenografts using quantitative, real-time, reverse transcriptase, polymerase chain reaction, autoradiography and immunohistochemistry (IHC). Mice with BxPC-3 xenografts were imaged with (123)I or (99)TcO(4) micro-SPECT/CT. Tumor dimensions and radionuclide localization were determined with imaging software. Linear regression and correlation analyses were performed to determine the relationship between tumor infection percentage and radionuclide uptake (% injected dose per gram) above background and a highly significant correlation was observed (r(2)=0.947). A detection threshold of 1.5-fold above the control tumor uptake (background) yielded a sensitivity of 2.7% MV-NIS-infected tumor cells. We reliably resolved multiple distinct intratumoral zones of infection from non-infected regions. Pinhole micro-SPECT/CT imaging using the NIS reporter demonstrated precise localization and quantitation of oncolytic MV-NIS infection, and can replace more time-consuming and expensive analyses (for example, autoradiography and IHC) that require animal killing.

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Sensitivity of Pinhole Micro-SPECT/CT for Detection of Separate Intratumoral Foci of MV-NIS Infection. A, Control BxPC-3 tumors (without MV-NIS infection) showed background 99mTcO4 localization (maximal pixel intensity, 37). A threshold 1.5-fold higher than the maximal background intensity (threshold value, 56) was applied to avoid false-positives. Note that the threshold-adjusted image on the right no longer shows background radioactivity. B, Example of a tumor from a mouse euthanized 4 days after MV-NIS injection. Far left panel, Coronal micro-SPECT/CT image. Arrows indicate sites of tumor injection. Middle left panel, Autoradiogram. Middle right panel, IHC stain of the same tumor section. Far right panel, Threshold-adjusted micro-SPECT/CT image. The in vivo micro-SPECT/CT and ex vivo autoradiography and IHC showed excellent spatial correlation. Note that 2 of the 4 injection points (seen as regions of high intensity on micro-SPECT/CT, far left panel) had radionuclide levels exceeding the 1.5-fold background threshold (far right panel). C, IHC tumor section shows the 4 injection zones (4.5-mm circles), which corresponded to 3 times the full-width, half-maximal spatial resolution of the imaging equipment. Quantitative IHC was used to calculate the percent of infected BxPC-3 cells in injection zones 1 and 4. The same experiment on a total of 7 injection sites from 5 tumors revealed a mean of 12.85%+/−1.7 % infected BxPC-3 cells per injection site. IHC denotes immunohistochemical stain; LUT, lookup table; SPECT/CT, single-photon emission computed tomography/computed tomography; MV, measles virus; MV-NIS, measles virus expressing sodium iodide symporter.
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Figure 7: Sensitivity of Pinhole Micro-SPECT/CT for Detection of Separate Intratumoral Foci of MV-NIS Infection. A, Control BxPC-3 tumors (without MV-NIS infection) showed background 99mTcO4 localization (maximal pixel intensity, 37). A threshold 1.5-fold higher than the maximal background intensity (threshold value, 56) was applied to avoid false-positives. Note that the threshold-adjusted image on the right no longer shows background radioactivity. B, Example of a tumor from a mouse euthanized 4 days after MV-NIS injection. Far left panel, Coronal micro-SPECT/CT image. Arrows indicate sites of tumor injection. Middle left panel, Autoradiogram. Middle right panel, IHC stain of the same tumor section. Far right panel, Threshold-adjusted micro-SPECT/CT image. The in vivo micro-SPECT/CT and ex vivo autoradiography and IHC showed excellent spatial correlation. Note that 2 of the 4 injection points (seen as regions of high intensity on micro-SPECT/CT, far left panel) had radionuclide levels exceeding the 1.5-fold background threshold (far right panel). C, IHC tumor section shows the 4 injection zones (4.5-mm circles), which corresponded to 3 times the full-width, half-maximal spatial resolution of the imaging equipment. Quantitative IHC was used to calculate the percent of infected BxPC-3 cells in injection zones 1 and 4. The same experiment on a total of 7 injection sites from 5 tumors revealed a mean of 12.85%+/−1.7 % infected BxPC-3 cells per injection site. IHC denotes immunohistochemical stain; LUT, lookup table; SPECT/CT, single-photon emission computed tomography/computed tomography; MV, measles virus; MV-NIS, measles virus expressing sodium iodide symporter.

Mentions: Because control tumors showed some background 99mTc localization, we first determined a practical lookup table (LUT) scale for image analysis that would exclude false-positives and intratumoral background radionuclide localization. A group of 2 control and 5 MV-NIS–infected BxPC-3 flank xenografts were evaluated using volume-of-interest image analysis. We applied a threshold that was 1.5-fold greater than the maximal background level when attempting to visually resolve distinct intratumoral foci of MV-NIS infection from uninfected regions. In the example shown in Figure 7A, the maximum pixel intensity value for background radioactivity was 37.0. The color LUT scale was adjusted such that all pixels below the threshold value of 56 (ie, 37×1.5) were colorless and those above the threshold were colored with the greatest intensity (red) of the LUT scale. In practice, an upper limit that was 10 to 15 LUT units above the threshold (in this example, 70) resulted in a smoother image because it provided some midrange colors for interpolation when the micro-SPECT image was scaled to the higher pixel density of the computer monitor.


Pinhole micro-SPECT/CT for noninvasive monitoring and quantitation of oncolytic virus dispersion and percent infection in solid tumors.

Penheiter AR, Griesmann GE, Federspiel MJ, Dingli D, Russell SJ, Carlson SK - Gene Ther. (2011)

Sensitivity of Pinhole Micro-SPECT/CT for Detection of Separate Intratumoral Foci of MV-NIS Infection. A, Control BxPC-3 tumors (without MV-NIS infection) showed background 99mTcO4 localization (maximal pixel intensity, 37). A threshold 1.5-fold higher than the maximal background intensity (threshold value, 56) was applied to avoid false-positives. Note that the threshold-adjusted image on the right no longer shows background radioactivity. B, Example of a tumor from a mouse euthanized 4 days after MV-NIS injection. Far left panel, Coronal micro-SPECT/CT image. Arrows indicate sites of tumor injection. Middle left panel, Autoradiogram. Middle right panel, IHC stain of the same tumor section. Far right panel, Threshold-adjusted micro-SPECT/CT image. The in vivo micro-SPECT/CT and ex vivo autoradiography and IHC showed excellent spatial correlation. Note that 2 of the 4 injection points (seen as regions of high intensity on micro-SPECT/CT, far left panel) had radionuclide levels exceeding the 1.5-fold background threshold (far right panel). C, IHC tumor section shows the 4 injection zones (4.5-mm circles), which corresponded to 3 times the full-width, half-maximal spatial resolution of the imaging equipment. Quantitative IHC was used to calculate the percent of infected BxPC-3 cells in injection zones 1 and 4. The same experiment on a total of 7 injection sites from 5 tumors revealed a mean of 12.85%+/−1.7 % infected BxPC-3 cells per injection site. IHC denotes immunohistochemical stain; LUT, lookup table; SPECT/CT, single-photon emission computed tomography/computed tomography; MV, measles virus; MV-NIS, measles virus expressing sodium iodide symporter.
© Copyright Policy
Related In: Results  -  Collection

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Figure 7: Sensitivity of Pinhole Micro-SPECT/CT for Detection of Separate Intratumoral Foci of MV-NIS Infection. A, Control BxPC-3 tumors (without MV-NIS infection) showed background 99mTcO4 localization (maximal pixel intensity, 37). A threshold 1.5-fold higher than the maximal background intensity (threshold value, 56) was applied to avoid false-positives. Note that the threshold-adjusted image on the right no longer shows background radioactivity. B, Example of a tumor from a mouse euthanized 4 days after MV-NIS injection. Far left panel, Coronal micro-SPECT/CT image. Arrows indicate sites of tumor injection. Middle left panel, Autoradiogram. Middle right panel, IHC stain of the same tumor section. Far right panel, Threshold-adjusted micro-SPECT/CT image. The in vivo micro-SPECT/CT and ex vivo autoradiography and IHC showed excellent spatial correlation. Note that 2 of the 4 injection points (seen as regions of high intensity on micro-SPECT/CT, far left panel) had radionuclide levels exceeding the 1.5-fold background threshold (far right panel). C, IHC tumor section shows the 4 injection zones (4.5-mm circles), which corresponded to 3 times the full-width, half-maximal spatial resolution of the imaging equipment. Quantitative IHC was used to calculate the percent of infected BxPC-3 cells in injection zones 1 and 4. The same experiment on a total of 7 injection sites from 5 tumors revealed a mean of 12.85%+/−1.7 % infected BxPC-3 cells per injection site. IHC denotes immunohistochemical stain; LUT, lookup table; SPECT/CT, single-photon emission computed tomography/computed tomography; MV, measles virus; MV-NIS, measles virus expressing sodium iodide symporter.
Mentions: Because control tumors showed some background 99mTc localization, we first determined a practical lookup table (LUT) scale for image analysis that would exclude false-positives and intratumoral background radionuclide localization. A group of 2 control and 5 MV-NIS–infected BxPC-3 flank xenografts were evaluated using volume-of-interest image analysis. We applied a threshold that was 1.5-fold greater than the maximal background level when attempting to visually resolve distinct intratumoral foci of MV-NIS infection from uninfected regions. In the example shown in Figure 7A, the maximum pixel intensity value for background radioactivity was 37.0. The color LUT scale was adjusted such that all pixels below the threshold value of 56 (ie, 37×1.5) were colorless and those above the threshold were colored with the greatest intensity (red) of the LUT scale. In practice, an upper limit that was 10 to 15 LUT units above the threshold (in this example, 70) resulted in a smoother image because it provided some midrange colors for interpolation when the micro-SPECT image was scaled to the higher pixel density of the computer monitor.

Bottom Line: The purpose of our study was to validate the ability of pinhole micro-single-photon emission computed tomography/computed tomography (SPECT/CT) to: 1) accurately resolve the intratumoral dispersion pattern and 2) quantify the infection percentage in solid tumors of an oncolytic measles virus encoding the human sodium iodide symporter (MV-NIS).We reliably resolved multiple distinct intratumoral zones of infection from non-infected regions.Pinhole micro-SPECT/CT imaging using the NIS reporter demonstrated precise localization and quantitation of oncolytic MV-NIS infection, and can replace more time-consuming and expensive analyses (for example, autoradiography and IHC) that require animal killing.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Medicine, Mayo Clinic, Rochester, MN 55905, USA.

ABSTRACT
The purpose of our study was to validate the ability of pinhole micro-single-photon emission computed tomography/computed tomography (SPECT/CT) to: 1) accurately resolve the intratumoral dispersion pattern and 2) quantify the infection percentage in solid tumors of an oncolytic measles virus encoding the human sodium iodide symporter (MV-NIS). Sodium iodide symporter (NIS) RNA level and dispersion pattern were determined in control and MV-NIS-infected BxPC-3 pancreatic tumor cells and mouse xenografts using quantitative, real-time, reverse transcriptase, polymerase chain reaction, autoradiography and immunohistochemistry (IHC). Mice with BxPC-3 xenografts were imaged with (123)I or (99)TcO(4) micro-SPECT/CT. Tumor dimensions and radionuclide localization were determined with imaging software. Linear regression and correlation analyses were performed to determine the relationship between tumor infection percentage and radionuclide uptake (% injected dose per gram) above background and a highly significant correlation was observed (r(2)=0.947). A detection threshold of 1.5-fold above the control tumor uptake (background) yielded a sensitivity of 2.7% MV-NIS-infected tumor cells. We reliably resolved multiple distinct intratumoral zones of infection from non-infected regions. Pinhole micro-SPECT/CT imaging using the NIS reporter demonstrated precise localization and quantitation of oncolytic MV-NIS infection, and can replace more time-consuming and expensive analyses (for example, autoradiography and IHC) that require animal killing.

Show MeSH
Related in: MedlinePlus