66 research outputs found
Targeting telomerase with radiolabeled inhibitors
The expression of telomerase in approximately 85% of cancers and its absence in the majority of normal cells makes it an attractive target for cancer therapy. However the lag period between initiation of telomerase inhibition and growth arrest makes direct inhibition alone an insufficient method of treatment. However, telomerase inhibition has been shown to enhance cancer cell radiosensitivity. To investigate the strategy of simultaneously inhibiting telomerase while delivering targeted radionuclide therapy to cancer cells, 123I-radiolabeled inhibitors of telomerase were synthesized and their effects on cancer cell survival studied. An 123I-labeled analogue of the telomerase inhibitor MST-312 inhibited telomerase with an IC50 of 1.58 μM (MST-312 IC50: 0.23 μM). Clonogenic assays showed a dose dependant effect of 123I-MST-312 on cell survival in a telomerase positive cell line, MDA-MB-435
DNA double-strand break repair:a theoretical framework and its application
DNA double-strand breaks (DSBs) are formed as a result of genotoxic insults, such as exogenous ionizing radiation, and are among the most serious types of DNA damage. One of the earliest molecular responses following DSB formation is the phosphorylation of the histone H2AX, giving rise to γH2AX. Many copies of γH2AX are generated at DSBs and can be detected in vitro as foci using well-established immuno-histochemical methods. It has previously been shown that anti-γH2AX antibodies, modified by the addition of the cell-penetrating peptide TAT and a fluorescent or radionuclide label, can be used to visualize and quantify DSBs in vivo. Moreover, when labelled with a high amount of the short-range, Auger electron-emitting radioisotope, 111In, the amount of DNA damage within a cell can be increased, leading to cell death. In this report, we develop a mathematical model that describes how molecular processes at individual sites of DNA damage give rise to quantifiable foci. Equations that describe stochastic mean behaviours at individual DSB sites are derived and parametrized using population-scale, time-series measurements from two different cancer cell lines. The model is used to examine two case studies in which the introduction of an antibody (anti-γH2AX-TAT) that targets a key component in the DSB repair pathway influences system behaviour. We investigate: (i) how the interaction between anti-γH2AX-TAT and γH2AX effects the kinetics of H2AX phosphorylation and DSB repair and (ii) model behaviour when the anti-γH2AX antibody is labelled with Auger electron-emitting 111In and can thus instigate additional DNA damage. This work supports the conclusion that DSB kinetics are largely unaffected by the introduction of the anti-γH2AX antibody, a result that has been validated experimentally, and hence the hypothesis that the use of anti-γH2AX antibody to quantify DSBs does not violate the image tracer principle. Moreover, it provides a novel model of DNA damage accumulation in the presence of Auger electron-emitting 111In that is supported qualitatively by the available experimental data. </p
Absorbed dose evaluation of Auger electron-emitting radionuclides: impact of input decay spectra on dose point kernels and S-values
The aim of this study was to investigate the impact of decay data provided by
the newly developed stochastic atomic relaxation model BrIccEmis on dose point
kernels (DPKs - radial dose distribution around a unit point source) and
S-values (absorbed dose per unit cumulated activity) of 14 Auger electron (AE)
emitting radionuclides, namely 67Ga, 80mBr, 89Zr, 90Nb, 99mTc, 111In, 117mSn,
119Sb, 123I, 124I, 125I, 135La, 195mPt and 201Tl. Radiation spectra were based
on the nuclear decay data from the medical internal radiation dose (MIRD)
RADTABS program and the BrIccEmis code, assuming both an isolated-atom and
condensed-phase approach. DPKs were simulated with the PENELOPE Monte Carlo
(MC) code using event-by-event electron and photon transport. S-values for
concentric spherical cells of various sizes were derived from these DPKS using
appropriate geometric reduction factors. The number of Auger and Coster-Kronig
(CK) electrons and x-ray photons released per nuclear decay (yield) from
MIRD-RADTABS were consistently higher than those calculated using BrIccEmis.
DPKs for the electron spectra from BrIccEmis were considerably different from
MIRD-RADTABS in the first few hundred nanometres from a point source where most
of the Auger electrons are stopped. S-values were, however, not significantly
impacted as the differences in DPKS in the sub-micrometre dimension were
quickly diminished in larger dimensions. Overestimation in the total AE energy
output by MIRD-RADTABS leads to higher predicted energy deposition by AE
emitting radionuclides, especially in the immediate vicinity of the decaying
radionuclides. This should be taken into account when MIRD-RADTABS data are
used to simulate biological damage at nanoscale dimensions.Comment: 27 pages, 4 figures, 3 table
Modular and automated synthesis of oligonucleotide-small molecule conjugates for cathepsin B mediated traceless release of payloads † ‡
The reversible attachment of small molecules to oligonucleotides provides versatile tools for the development of improved oligonucleotide therapeutics. However, cleavable linkers in the oligonucleotide field are scarce, particularly with respect to the requirement for traceless release of the payload in vivo. Herein, we describe a cathepsin B-cleavable dipeptide phosphoramidite, Val-Ala(NB) for the automated synthesis of oligonucleotide-small molecule conjugates. Val-Ala(NB) was protected by a photolabile 2-nitrobenzyl group to improve the stability of the peptide linker during DNA synthesis. Intracellular cathepsin B digests the dipeptide efficiently, releasing the payload-phosphate which is converted to the free payload by endogenous phosphatase enzymes. With the advantages of modular synthesis and stimuli-responsive drug release, we believe Val-Ala(NB) will be a potentially valuable cleavable linker for use in oligonucleotide-drug conjugates
111In-BnDTPA-F3: an Auger electron-emitting radiotherapeutic agent that targets nucleolin
INTRODUCTION: The F3 peptide (KDEPQRRSARLSAKPAPPKPEPKPKKAPAKK), a fragment of the human high mobility group protein 2, binds nucleolin. Nucleolin is expressed in the nuclei of normal cells but is also expressed on the membrane of some cancer cells. The goal was to investigate the use of 111In-labeled F3 peptide for Auger electron-targeted radiotherapy. METHODS: F3 was labeled with fluorescein isothiocyanate (FITC) for confocal microscopy and conjugated to p-SCN-benzyl-diethylenetriaminepentaacetic acid (BnDTPA) for labeling with 111In to form 111In-BnDTPA-F3. MDA-MB-231-H2N (231-H2N) human breast cancer cells were exposed to 111In-BnDTPA-F3 and used in cell fractionation, γH2AX immunostaining (a marker of DNA double-strand breaks), and clonogenic assays. In vivo, biodistribution studies of 111In-BnDTPA-F3 were performed in 231-H2N xenograft-bearing mice. In tumor growth delay studies, 111In-BnDTPA-F3 (3 μg, 6 MBq/μg) was administered intravenously to 231-H2N xenograft-bearing mice once weekly for 3 weeks. RESULTS: Membrane-binding of FITC-F3 was observed in 231-H2N cells, and there was co-localization of FITC-F3 with nucleolin in the nuclei. After exposure of 231-H2N cells to 111In-BnDTPA-F3 for 2 h, 1.7% of 111In added to the medium was membrane-bound. Of the bound 111In, 15% was internalized, and of this, 37% was localized in the nucleus. Exposure of 231-H2N cells to 111In-BnDTPA-F3 (1 μM, 6 MBq/μg) resulted in a dose-dependent increase in γH2AX foci and in a significant reduction of clonogenic survival compared to untreated cells or cells exposed to unlabeled BnDTPA-F3 (46 ± 4.1%, 100 ± 1.8%, and 132 ± 7.7%, respectively). In vivo, tumor uptake of 111In-BnDTPA-F3 (3 μg, 6 MBq/μg) at 3-h post-injection was 1% of the injected dose per gram (%ID/g), and muscle uptake was 0.5%ID/g. In tumor growth delay studies, tumor growth rate was reduced 19-fold compared to untreated or unlabeled BnDTPA-F3-treated mice (p = 0.023). CONCLUSION: 111In-BnDTPA-F3 is internalized into 231-H2N cells and translocates to the nucleus. 111In-BnDTPA-F3 has a potent cytotoxic effect in vitro and an anti-tumor effect in mice bearing 231-H2N xenografts despite modest total tumor accumulation
Monitoring response to anti-angiogenic mTOR inhibitor therapy in vivo using 111In-bevacizumab
Abstract Background The ability to image vascular endothelial growth factor (VEGF) could enable prospective, non-invasive monitoring of patients receiving anti-angiogenic therapy. This study investigates the specificity and pharmacokinetics of 111In-bevacizumab binding to VEGF and its use for assessing response to anti-angiogenic therapy with rapamycin. Specificity of 111In-bevacizumab binding to VEGF was tested in vitro with unmodified radiolabelled bevacizumab in competitive inhibition assays. Uptake of 111In-bevacizumab in BALB/c nude mice bearing tumours with different amounts of VEGF expression was compared to that of isotype-matched control antibody (111In-IgG1κ) with an excess of unlabelled bevacizumab. Intratumoural VEGF was evaluated using ELISA and Western blot analysis. The effect of anti-angiogenesis therapy was tested by measuring tumour uptake of 111In-bevacizumab in comparison to 111In-IgG1κ following administration of rapamycin to mice bearing FaDu xenografts. Uptake was measured using gamma counting of ex vivo tumours and effect on vasculature by using anti-CD31 microscopy. Results Specific uptake of 111In-bevacizumab in VEGF-expressing tumours was observed. Rapamycin led to tumour growth delay associated with increased relative vessel size (8.5 to 10.3, P = 0.045) and decreased mean relative vessel density (0.27 to 0.22, P = 0.0015). Rapamycin treatment increased tumour uptake of 111In-bevacizumab (68%) but not 111In-IgGκ and corresponded with increased intratumoural VEGF165. Conclusions 111In-bevacizumab accumulates specifically in VEGF-expressing tumours, and changes after rapamycin therapy reflect changes in VEGF expression. Antagonism of mTOR may increase VEGF in vivo, and this new finding provides the basis to consider combination studies blocking both pathways and a way to monitor effects
Absorbed dose evaluation of Auger electron-emitting radionuclides: impact of input decay spectra on dose point kernels and S-values
The aim of this study was to investigate the impact of decay data provided by the newly developed stochastic atomic relaxation model BrIccEmis on dose point kernels (DPKs - radial dose distribution around a unit point source) and
S-values (absorbed dose per unit cumulated activity) of 14 Auger electron (AE) emitting radionuclides, namely 67Ga, 80mBr, 89Zr, 90Nb, 99mTc, 111In, 117mSn, 119Sb, 123I, 124I, 125I, 135La, 195mPt and 201Tl. Radiation spectra were based on the nuclear decay data from the medical
internal radiation dose (MIRD) RADTABS program and the BrIccEmis code, assuming both an isolated-atom and condensed-phase approach. DPKs were simulated with the PENELOPE Monte Carlo (MC) code using event-byevent electron and photon transport. S-values for concentric spherical cells of various sizes were derived from these DPKs using appropriate geometric reduction factors.
The number of Auger and Coster–Kronig (CK) electrons and x-ray photons released per nuclear decay (yield) from MIRD-RADTABS were consistently higher than those calculated using BrIccEmis. DPKs for the electron spectra
from BrIccEmis were considerably different from MIRD-RADTABS in the first few hundred nanometres from a point source where most of the Auger electrons are stopped. S-values were, however, not significantly impacted as the differences in DPKs in the sub-micrometre dimension were quickly diminished in larger dimensions. Overestimation in the total AE energy output by MIRD-RADTABS leads to
higher predicted energy deposition by AE emitting radionuclides, especially in the immediate vicinity of the decaying radionuclides. This should be taken into account when MIRD-RADTABS data are used to simulate biological
damage at nanoscale dimensions.The authors gratefully acknowledge funding support from the Cancer Research-UK (C5255/
A15935), the Medical Research Council (MC_PC_12004), the Australian Research Council
Discovery Grant (no. DP140103317) and the Generalitat de Catalunya (project no. 2014 SGR
846)
Oligonucleotide-Functionalized Gold Nanoparticles for Synchronous Telomerase Inhibition, Radiosensitization, and Delivery of Theranostic Radionuclides
Telomerase represents an attractive target in oncology as it is expressed in cancer but not in normal tissues. The oligonucleotide inhibitors of telomerase represent a promising anticancer strategy, although poor cellular uptake can restrict their efficacy. In this study, gold nanoparticles (AuNPs) were used to enhance oligonucleotide uptake. “match” oligonucleotides complementary to the telomerase RNA template subunit (hTR) and “scramble” (control) oligonucleotides were conjugated to diethylenetriamine pentaacetate (DTPA) for 111In-labeling. AuNPs (15.5 nm) were decorated with a monofunctional layer of oligonucleotides (ON–AuNP) or a multifunctional layer of oligonucleotides, PEG(polethylene glycol)800-SH (to reduce AuNP aggregation) and the cell-penetrating peptide Tat (ON–AuNP–Tat). Match–AuNP enhanced the cellular uptake of radiolabeled oligonucleotides while retaining the ability to inhibit telomerase activity. The addition of Tat to AuNPs increased nuclear localization. 111In–Match–AuNP–Tat induced DNA double-strand breaks and caused a dose-dependent reduction in clonogenic survival of telomerase-positive cells but not telomerase-negative cells. hTR inhibition has been reported to sensitize cancer cells to ionizing radiation, and 111In–Match–AuNP–Tat therefore holds promise as a vector for delivery of radionuclides into cancer cells while simultaneously sensitizing them to the effects of the emitted radiation
Orally administered oxygen nanobubbles enhance tumor response to sonodynamic therapy
Suspensions of oxygen-filled bubbles are under active investigation as potential means of relieving tissue hypoxia. Intravenous administration of large quantities of bubbles is, however, undesirable. Previous work by the authors has demonstrated that tumor oxygen levels can be increased following oral administration of phospholipid stabilized oxygen nanobubbles. The aim of this study was to determine whether this would enhance the efficacy of sonodynamic therapy (SDT), which is known to be inhibited in hypoxic tissue. Experiments were conducted in a murine model of pancreatic cancer. Animals were treated with SDT (intratumoural injection of 1 mM Rose Bengal followed by exposure to 1 MHz ultrasound, 0.1 kHz pulse repetition frequency, 30% duty cycle, 3.5 W cm−2 for 3.5 minutes) either with or without a prior gavage of oxygen bubbles. A statistically significant reduction in the rate of tumor growth was observed in the groups receiving oxygen nanobubbles either 5 or 20 minutes before SDT. Separate measurements of tumor oxygen using a fiber optic probe and expression of hypoxia inducible factor (HIF)1α following tumor excision, confirmed the change in tumor oxygen levels. These findings offer a potentially promising new approach to relieving tissue hypoxia in order to facilitate cancer therapy
Dosimetric evaluation of radionuclides for VCAM-1-targeted radionuclide therapy of early brain metastases
CNPQ - CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOBrain metastases develop frequently in patients with breast cancer, and present a pressing therapeutic challenge. Expression of vascular cell adhesion molecule 1 (VCAM-1) is upregulated on brain endothelial cells during the early stages of metastasis and provides a target for the detection and treatment of early brain metastases. The aim of this study was to use a model of early brain metastasis to evaluate the efficacy of a-emitting radionuclides, Tb-149, At-211, Pb-212, Bi-213 and Ac-225|| beta-emitting radionuclides, Y-90, Tb-161 and Lu-177|| and Auger electron (AE)-emitters Ga-67, Zr-89, In-111 and I-124, for targeted radionuclide therapy (TRT). METHODS: Histologic sections and two photon microscopy of mouse brain parenchyma were used to inform a cylindrical vessel geometry using the Geant4 general purpose Monte Carlo (MC) toolkit with the Geant4-DNA low energy physics models. Energy deposition was evaluated as a radial function and the resulting phase spaces were superimposed on a DNA model to estimate double-strand break (DSB) yields for representative beta- and alpha-emitters, Lu-177 and Pb-212. Relative biological effectiveness (RBE) values were determined by only evaluating DNA damage due to physical interactions. RESULTS: Lu-177 produced 2.69 +/- 0.08 DSB per GbpGy, without significant variation from the lumen of the vessel to a radius of 100 mu m. The DSB yield of Pb-212 included two local maxima produced by the 6.1 MeV and 8.8 MeV alpha-emissions from decay products, Bi-212 and Po-212, with yields of 7.64 +/- 0.12 and 9.15 +/- 0.24 per GbpGy, respectively. Given its higher DSB yield Pb-212 may be more effective for short range targeting of early micrometastatic lesions than Lu-177. CONCLUSION: MC simulation of a model of early brain metastases provides invaluable insight into the potential efficacy of alpha-, beta- and AE-emitting radionuclides for TRT. Pb-212, which has the attributes of a theranostic radionuclide since it can be used for SPECT imaging, showed a favorable dose profile and RBE.81292303CNPQ - CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICOCNPQ - CONSELHO NACIONAL DE DESENVOLVIMENTO CIENTÍFICO E TECNOLÓGICO306775/2015-8190154/2013-6Agências de fomento estrangeiras apoiaram essa pesquisa, mais informações acesse artig
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