Ex Vivo and In Vivo Imaging and Biodistribution of Aptamers Targeting the Human Matrix MetalloProtease-9 in Melanomas

Abstract

The human Matrix MetalloProtease-9 (hMMP-9) is overexpressed in tumors where it promotes the release of cancer cells thus contributing to tumor metastasis. We raised aptamers against hMMP-9, which constitutes a validated marker of malignant tumors, in order to design probes for imaging tumors in human beings. A chemically modified RNA aptamer (F3B), fully resistant to nucleases was previously described. This compound was subsequently used for the preparation of F3B-Cy5, F3B-S-acetylmercaptoacetyltriglycine (MAG) and F3B-DOTA. The binding properties of these derivatives were determined by surface plasmon resonance and electrophoretic mobility shift assay. Optical fluorescence imaging confirmed the binding to hMMP-9 in A375 melanoma bearing mice. Quantitative biodistribution studies were performed at 30 min, 1h and 2 h post injection of 99mTc-MAG-aptamer and 111In-DOTA-F3B. 99mTc radiolabeled aptamer specifically detected hMMP-9 in A375 melanoma tumors but accumulation in digestive tract was very high. Following i.v. injection of 111In-DOTA-F3B, high level of radioactivity was observed in kidneys and bladder but digestive tract uptake was very limited. Tumor uptake was significantly (student t test, p<0.05) higher for 111In-DOTA-F3B with 2.0%ID/g than for the 111In-DOTA-control oligonucleotide (0.7%ID/g) with tumor to muscle ratio of 4.0. Such difference in tumor accumulation has been confirmed by ex vivo scintigraphic images performed at 1h post injection and by autoradiography, which revealed the overexpression of hMMP-9 in sections of human melanomas. These results demonstrate that F3B aptamer is of interest for detecting hMMP-9 in melanoma tumor.

Discovery of tumoricidal DNA oligonucleotides by response-directed in vitro evolution

“…Fluorescently-labeled control oligos have consistently shown no accumulation in the tumors, and were localized to the liver and kidneys as rapidly as 30 min (Supplementary Note 6, Supplementary Figs. 13-14), in agreement with previously reported studies 30,31 . Multiple experiments with random sequence controls revealed no significant protease resistance of E8 (p = 0.08) which could be attributed to its structure.…”

“…Biodistribution and safety of E8. To determine the dispersion of E8 we used fluorescently-labeled E8 as previously described for aptamer in vivo imaging probes [30][31][32] . The molecule, modified with 5'-Cy5.5 and 3'-PEG, was injected intravenously in two doses (6 and 60 mg/kg) into NOD/SCID mice in which MDA-MB-231 tumors were induced.…”

Amphiphilic dendrimer engineered nanocarrier systems for co-delivery of siRNA and paclitaxel to matrix metalloproteinase-rich tumors for synergistic therapy

“…We hoped that TMSP could be activated by MMP-2/9 to expose the CPP moiety to further improve the cellular uptake of the nanocarriers. Here, HT-1080 cells and A375 cells are selected because both are known as cell lines with high expression of MMP-2/ 9 41,42 . In HT-1080 cells treated with free FAM-siRNA and various FAM-siRNA-loaded nanocarriers at the same concentration for 4 h, the uptake of TMSP-ADENS was significantly higher than that of the ADENS and was even comparable to that of CPP-ADENS (CPP-modified ADENS; Fig.…”

Discovery of tumoricidal DNA oligonucleotides by effect-directedin-vitroevolution

“…To determine the dispersion of E8 we used fluorescently-labeled E8 as previously described for aptamer in-vivo imaging probes [30][31][32] . The molecule, modified with 5' Cy5.5 and 3' PEG, was injected intravenously in two doses (6 and 60 mg/kg) into NOD/SCID mice in which MDA-MB-231 tumors were induced.…”