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Supplementary Information
Dual-targeting of tissue factor and CD105 for preclinical PET imaging of pancreatic cancer
List of Supplementary Materials
Supplementary Figure 1. Purification of heterodimer after click chemistry conjugation of ALT-836-Fab and TRC105-Fab
Supplementary Figure 2. MALDI-TOF mass spectra of ALT-836-Fab: [M+H]+ = 47266.0, TRC105-Fab: [M+H]+ = 47454.7, and heterodimer: exact [M+H]+ = 103504.0.
Supplementary Figure 3. Flow cytometry analysis in BXPC-3 after 1 h incubation of FITC-Fab conjugates for the comparison of their in vitro targeting
Supplementary Figure 4. Radiolabeling efficiency at different copper: immunoconjugate ratios were used to determine the number of NOTA per antibody fragment molecule
Supplementary Figure 5. PET imaging and biodistribution of 64CuCl2 in BXPC-3 tumor-bearing mice.
Supplementary Figure 6. PET imaging and quantitative ROI analysis of 64Cu-NOTA-heterodimer with TF and CD105 dual-blocking in BXPC-3 tumor-bearing mice.
Supplementary Figure 7. Coronal images and transverse PET/CT images of normal nude mice at 3, 15, 24 and 30 h following injection of 64Cu-NOTA-heterodimer
Supplementary Figure 8. PET ROI-derived quantification of 64Cu-NOTA-heterodimer uptake in the tumor, normal pancreas, blood pool, liver, kidney and muscle at 3 h, 15 h, 24 h and 30 h p.i. of the tracer
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Supplementary Figure 1. Purification of heterodimer after click chemistry conjugation of ALT-836-Fab and TRC105-Fab. A, Size exclusion chromatogram of the reaction mixture; the peak around 51 min corresponds to the heterodimer. B, Chromatogram of the purified heterodimer and the starting materials. C, table showing the heterodimer generation yield. Yields were determined as the ratio between the two peaks in A.
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Supplementary Figure 2. MALDI-TOF mass spectra of ALT-836-Fab: [M+H]+ = 47266.0, TRC105-Fab: [M+H]+ = 47454.7, and heterodimer: exact [M+H]+ = 103504.0.
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Supplementary Figure 3. Flow cytometry of BXPC-3 after 1 h incubation of FITC-Fab conjugates for a comparison of in vitro targeting efficiency
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Supplementary Figure 4. Radiolabeling efficiency at different copper: immunoconjugate ratios were used to determine the number of NOTA per antibody fragment molecule. Knowing the moles of copper added and the number of moles of copper bound can be calculated from the % radiolabeling efficiency. The moles of copper bound are divided by the moles of antibody added (2×10 -10 moles) to give the ligand: antibody fragment ratio.
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Supplementary Figure 5. PET imaging and biodistribution of 64CuCl2 in BXPC-3 tumor-bearing mice. A, Serial coronal PET images of 64CuCl2 at 3, 15, and 30 h p.i. (n=3). An arrow denotes the tumor location. B, Ex vivo biodistribution of 64CuCl2 after in BXPC-3 tumor bearing mice 30 h after injection.
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Supplementary Figure 6. PET imaging and quantitative ROI analysis of 64Cu-NOTA-heterodimer with TF and CD105 dual-blocking in BXPC-3 tumor-bearing mice. A, Serial coronal PET images of 64Cu-NOTA-heterodimer at 3, 15 and 30 h following tracer injection after both TF and CD105 blocking (n=3). An arrow denotes the tumor location. B, Time-activity curves of the BXPC-3 tumor, blood, liver, kidney and muscle following intravenous administration of 64Cu-NOTA-heterodimer with TF and CD105 blocking.
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Supplementary Figure 7. Coronal images (A) and transverse PET/CT images (B) of normal nude mice at 3, 15, 24 and 30 h following injection of 64Cu-NOTA-heterodimer.
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Supplementary Figure 8. PET ROI-derived quantification of 64Cu-NOTA-heterodimer uptake in the mice bearing orthotopic BXPC-3 tumor (A) and normal nude mice (B) at 3 h, 15 h, 24 h and 30 h p.i. of the tracer; reported as %ID/g ± SD (n=3).
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