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Ronald G Blasberg

Publications and source records attributed to Ronald G Blasberg.

18 recordsLinked to original sources
PubMed2006

Synthesis and evaluation of [18F] labeled pyrimidine nucleosides for positron emission tomography imaging of herpes simplex virus 1 thymidine kinase gene expression.

Synthesis of three novel 2'-deoxy-2'-[18F]fluoro-1-beta-D-arabinofuranosyluracil derivatives [18F]FPAU, [18F]FBrVAU, and [18F]FTMAU is reported. The compounds were synthesized by coupling of 1-bromo-2-deoxy-2-fluoro sugars with corresponding silylated uracil derivatives. In vitro cell uptake indicated that all three compounds are taken up selectively in RG2TK+ cells with negligible uptake in RG2 cells. The results indicate that [18F]FBrVAU and [18F]FTMAU have better uptake profiles in comparison to [18F]FPAU and have potential as PET probes for imaging HSV1-tk gene expression.

Animals↗
PubMed2006

[131I]FIAU labeling of genetically transduced, tumor-reactive lymphocytes: cell-level dosimetry and dose-dependent toxicity.

PURPOSE: Donor T cells have been shown to be reactive against and effective in adoptive immunotherapy of Epstein-Barr virus (EBV) lymphomas which develop in some leukemia patients post marrow transplantation. These T cells may be genetically modified by incorporation of a replication-incompetent viral vector (NIT) encoding both an inactive mutant nerve growth factor receptor (LNGFR), as an immunoselectable surface marker, and a herpes simplex virus thymidine kinase (HSV-TK), rendering the cells sensitive to ganciclovir. The current studies are based on the selective HSV-TK-catalyzed trapping (phosphorylation) of the thymidine analog [(131)I]-2'-fluoro-2'-deoxy-1-beta-D-arabinofuransyl-5-iodo-uracil (FIAU) as a means of stably labeling such T cells for in vivo trafficking (including tumor targeting) studies. Because of the radiosensitivity of lymphocytes and the potentially high absorbed dose to the nucleus from intracellular (131)I (even at tracer levels), the nucleus absorbed dose (D ( n )) and dose-dependent immune functionality were evaluated for NIT(+) T cells labeled ex vivo in [(131)I]FIAU-containing medium. METHODS: Based on in vitro kinetic studies of [(131)I]FIAU uptake by NIT(+) T cells, D ( n ) was calculated using an adaptation of the MIRD formalism and the recently published MIRD cellular S factors. Immune cytotoxicity of [(131)I]FIAU-labeled cells was assayed against (51)Cr-labeled target cells [B-lymphoblastoid cells (BLCLs)] in a standard 4-h release assay. RESULTS AND CONCLUSION: At median nuclear absorbed doses up to 830 cGy, a (51)Cr-release assay against BLCLs showed no loss of immune cytotoxicity, thus demonstrating the functional integrity of genetically transduced, tumor-reactive T cells labeled at this dose level for in vivo cell trafficking and tumor targeting studies.

Animals↗
PubMed2006

Multi-modality molecular imaging of tumors.

Noninvasive in vivo molecular-genetic imaging uses nuclear, magnetic resonance, and optical imaging techniques. Described and discussed are "direct" imaging of specific molecules and pathway activity, "indirect" reporter gene imaging, and "bio-marker" or "surrogate" imaging. Applications of PET- and optical-based reporter imaging are demonstrated, including imaging of oncogenesis in genetic mouse models, endogenous molecular-genetic-biological properties, and response to therapy in animal models of human disease. Molecular imaging studies complement established ex vivo molecular-biological assays that require tissue sampling by providing a spatial as well as temporal dimension to our understanding of oncogenesis, and the progression and treatment of cancer. Molecular imaging studies being performed in experimental animals will be translated to animals in the near future.

Animals↗
PubMed2006

Molecular imaging reveals skeletal engraftment sites of transplanted bone marrow cells.

Molecular imaging holds great promise for the in vivo study of cell therapy. Our hypothesis was that multimodality molecular imaging can identify the initial skeletal engraftment sites post-bone marrow cell transplantation. Utilizing a standard mouse model of bone marrow (BM) transplantation, we introduced a combined bioluminescence (BLI) and positron emission tomography (PET) imaging reporter gene into mouse bone marrow cells. Bioluminescence imaging was used for monitoring serially the early in vivo BM cell engraftment/expansion every 24 h. Significant cell engraftment/expansion was noted by greatly increased bioluminescence about 1 week posttransplant. Then PET was applied to acquire three-dimensional images of the whole-body in vivo biodistribution of the transplanted cells. To localize cells in the skeleton, PET was followed by computed tomography (CT). Co-registration of PET and CT mapped the sites of BM engraftment. Multiple, discrete BM cell engraftment sites were observed. Taken together, this multimodality approach may be useful for further in vivo characterization of various therapeutic cell types.

Animals↗
PubMed2005

Imaging herpes viral thymidine kinase-1 reporter gene expression with a new 18F-labeled probe: 2'-fluoro-2'-deoxy-5-[18F]fluoroethyl-1-beta-d-arabinofuranosyl uracil.

The preparation and radiolabeling of 2'-fluoro-2'-deoxy-1-beta-d-arabinofuranosyl-5-(2-fluoroethyl)-uracil (FFEAU) with 18F and its evaluation as a probe for imaging herpes simplex virus 1 thymidine kinase (HSV1-tk) gene expression are described. 2'-Fluoro-2'-deoxy-3',5'-di-O-benzoyl-1-beta-d-arabinofuranosyl-3-N-benzoyl-5-(2-[18F]fluoroethyl)-uracil 12 was prepared by nucleophilic substitution of the corresponding tosyl 8 or trifluoroethanesulfonyl 9 derivative with n-Bu4N[18F]F. Base hydrolysis was used to remove the benzoyl protecting groups, followed by HPLC purification, to afford [18F]FFEAU 13. The trifluoroethanesulfonyl substrate 9 appears to be the better labeling precursor. Carrier n-Bu4NF was added to the labeling reaction, which resulted in specific activities of 40-70 Ci/mmol (estimated). Radiochemical purity averaged 94+/-4%. Although [18F]FFEAU was obtained in low radiochemical yield with 9 and further optimization of the radiosynthesis will be required, sufficient product was available for a series of in vitro and in vivo studies. [18F]FFEAU was directly compared with [3H]TdR in a series of in vitro accumulation studies involving a HSV1-tk stably transduced cell line, RG2TK+ and a nontransduced, wild-type RG2 cells. The initial in vitro and in vivo imaging studies are promising; FFEAU has in vitro accumulation and sensitivity characteristics similar to that previously reported for FIAU, but greater selectivity than FIAU due to lower uptake and retention in nontransduced cells and tissues. The animal imaging experiment showed low levels of radioactivity in the lungs, with little or no radioactivity seen in the heart, liver, spleen and intestines.

Animals↗
PubMed2005

High-resolution imaging of bone precursor cells within the intact bone marrow cavity of living mice.

Bone precursor cells (BPCs) play a critical role in bone maintenance and regeneration. Currently, no tool exists to study BPCs or other bone marrow cell types directly within their complex microenvironment. Here, we describe in vivo magnetic resonance imaging (MRI) of anatomical structures inside the medullary cavity of the mouse femur. We demonstrate that BPCs passively labeled with iron oxide-containing particles can be monitored by MRI within the intact bone marrow at an in-plane resolution of 43x25 microm. Anatomical detail provided by MRI is complemented by functional optical imaging of reporter gene expression. Single-cell dual iron oxide-reporter gene labeling has potential for combined cell tracking and cell biology studies. In summary, we describe a versatile platform suitable for studying the biology of many bone marrow cell types in living bone.

Animals↗
PubMed2004

Role of reporter gene imaging in molecular and cellular biology.

Molecular imaging, including reporter gene methods, provides a unique opportunity to study biology in a living subject, thereby allowing physiological events to be monitored in an intact microenvironment. This review takes a molecular and cell biology perspective on recent studies which utilize reporter gene imaging as a tool to non-invasively monitor specific molecular biology pathways in vivo. Studies in rodent models demonstrate the feasibility of reporter gene imaging to visualize and measure key cellular pathways, such as transcription, translation and protein-protein interactions. The review indicates that molecular imaging is likely to be useful in the translation of molecular biology to medicine and biotechnological applications.

Animals↗
PubMed2003

Serial in vivo imaging of the targeted migration of human HSV-TK-transduced antigen-specific lymphocytes.

New technologies are needed to characterize the migration, survival, and function of antigen-specific T cells in vivo. Here, we demonstrate that Epstein-Barr virus (EBV)--specific T cells transduced with vectors encoding herpes simplex virus-1 thymidine kinase (HSV-TK) selectively accumulate radiolabeled 2'-fluoro-2'-deoxy-1-beta-D-arabinofuranosyl-5-iodouracil (FIAU). After adoptive transfer, HSV-TK+ T cells labeled in vitro or in vivo with [131I]FIAU or [124I]FIAU can be noninvasively tracked in SCID mice bearing human tumor xenografts by serial images obtained by scintigraphy or positron emission tomography (PET), respectively. These T cells selectively accumulate in EBV+ tumors expressing the T cells' restricting HLA allele but not in EBV- or HLA-mismatched tumors. The concentrations of transduced T cells detected in tumors and tissues are closely correlated with the concentrations of label retained at each site. Radiolabeled transduced T cells retain their capacity to eliminate targeted tumors selectively. This technique for imaging the migration of ex vivo-transduced antigen-specific T cells in vivo is informative, nontoxic, and potentially applicable to humans.

Animals↗
PubMed2003

In vivo molecular-genetic imaging: multi-modality nuclear and optical combinations.

Multi-modality, noninvasive in vivo imaging is increasingly being used in molecular-genetic studies and will soon become the standard approach for reporter gene imaging studies in small animals. The coupling of nuclear and optical reporter genes, as described here, represents only the beginning of a far wider application of this technology in the future. Optical imaging and optical reporter systems are cost-effective and time-efficient; they require less resources and space than PET or MRI, and are particularly well suited for imaging small animals, such as mice. Optical reporter systems are also very useful for the quantification and selection of transduced cells using FACS, and for performing in vitro assays to validate the function and sensitivity of constitutive and specific-inducible reporter systems. However, optical imaging techniques are limited by depth of light penetration and do not yet provide optimal quantitative or tomographic information. These issues are not limiting for PET- or MRI-based reporter systems, and PET- and MRI-based animal studies are more easily generalized to human applications. Many of the shortcomings of each modality alone can be overcome by the use of dual- or triple-modality reporter constructs that incorporate the opportunity for PET, fluorescence and bioluminescence imaging.

Animals↗
PubMed2003

Development of a new reporter gene system--dsRed/xanthine phosphoribosyltransferase-xanthine for molecular imaging of processes behind the intact blood-brain barrier.

We report the development of a novel dual-modality fusion reporter gene system consisting of Escherichia coli xanthine phosphoribosyltransferase (XPRT) for nuclear imaging with radiolabeled xanthine and Discosoma red fluorescent protein for optical fluorescent imaging applications. The dsRed/XPRT fusion gene was successfully created and stably transduced into RG2 glioma cells, and both reporters were shown to be functional. The level of dsRed fluorescence directly correlated with XPRT enzymatic activity as measured by ribophosphorylation of [14C]-xanthine was in vitro (Ki = 0.124 +/- 0.008 vs. 0.00031 +/- 0.00005 mL/min/g in parental cell line), and [*]-xanthine octanol/water partition coefficient was 0.20 at pH = 7.4 (logP = -0.69), meeting requirements for the blood-brain barrier (BBB) penetrating tracer. In the in vivo experiment, the concentration of [14C]-xanthine in the normal brain varied from 0.20 to 0.16 + 0.05% dose/g under 0.87 + 0.24% dose/g plasma radiotracer concentration. The accumulation in vivo in the transfected flank tumor was to 2.4 +/- 0.3% dose/g, compared to 0.78 +/- 0.02% dose/g and 0.64 +/- 0.05% dose/g in the control flank tumors and intact muscle, respectively. [14C]-Xanthine appeared to be capable of specific accumulation in the transfected infiltrative brain tumor (RG2-dsRed/XPRT), which corresponded to the 585 nm fluorescent signal obtained from the adjacent cryosections. The images of endogenous gene expression with the "sensory system" have to be normalized for the transfection efficiency based on the "beacon system" image data. Such an approach requires two different "reporter genes" and two different "reporter substrates." Therefore, the novel dsRed/XPRT fusion gene can be used as a multimodality reporter system in the biological applications requiring two independent reporter genes, including the cells located behind the BBB.

Amino Acid Sequence↗
PubMed2003

Molecular imaging and cancer.

Molecular imaging is a relatively new discipline, which developed over the past decade, initially driven by in situ reporter imaging technology. Noninvasive in vivo molecular imaging developed more recently and is based on magnetic resonance and nuclear (positron emission tomography; gamma camera) and in vivo optical imaging systems. Molecular imaging has its roots in both molecular biology and cell biology, as well as imaging technology. Most current in vivo molecular imaging strategies are "indirect" and involve the coupling of a "reporter/marker gene" with a complimentary "reporter/marker probe." Imaging the level of probe accumulation provides indirect information related to the level of reporter gene expression. Reporter gene constructs are driven by upstream promoter/enhancer elements that function as an "on/off switch"; they can be constitutive, leading to continuous transcription to identify the site and monitor the level and duration of gene (vector) activity. Alternatively, they can be inducible, leading to controlled gene expression, or they can function as a sensor element to monitor the level of endogenous promoters and transcription factors. Three indirect strategies for imaging therapeutic transgenes are discussed. Several examples of imaging endogenous biological processes in animals using reporter constructs, radiolabeled probes, and positron emission tomography imaging are reviewed (p53-dependent gene expression, T-cell receptor-dependent activation of T-lymphocytes, and trafficking of T-lymphocytes). Issues related to the translation of noninvasive molecular imaging technology into the clinic are discussed.

Animals↗
PubMed2002

In vivo molecular-genetic imaging.

Noninvasive in vivo molecular-genetic imaging has developed over the past decade and involves nuclear (PET, gamma camera), magnetic resonance, and in vivo optical imaging systems. Although three different imaging strategies - "direct," "indirect" and "surrogate" - are being used, most current in vivo molecular imaging strategies are "indirect" and involve the coupling of a "reporter gene" with a complimentary "reporter probe." Imaging the level of probe accumulation provides indirect information related to the level of reporter gene expression. Reporter gene constructs are driven by upstream promoter/enhancer elements; reporter gene expression can be "constitutive" leading to continuous transcription and used to identify the site of transduction and to monitor the level and duration of gene (vector) activity. Alternatively, reporter gene expression can be "inducible" leading to controlled gene expression. Controlled gene expression can be tissue-specific and/or responsive to the level of endogenous promoters and transcription factors. Several examples of imaging endogenous biological processes in animals using reporter constructs, radiolabeled probes and PET imaging are reviewed, including: 1) imaging transcriptional regulation (e.g., p53-dependent gene expression), 2) imaging weak promoters (cis- vs. trans-reporter configurations), 3) imaging post-transcriptional regulation of gene expression, 4) imaging protein-protein interactions. The development of versatile and sensitive assays that do not require tissue sampling will be of considerable value for monitoring molecular-genetic and cellular processes in animal models of human disease, as well as for studies in human subjects in the future. Non-invasive imaging of molecular-genetic and cellular processes will compliment established ex vivo molecular-biological assays that require tissue sampling, and will provide a spatial as well as a temporal dimension to our understanding of various diseases.

Animals↗
PubMed2002

Molecular-genetic imaging: a nuclear medicine-based perspective.

Molecular imaging is a relatively new discipline, which developed over the past decade, initially driven by in situ reporter imaging technology. Noninvasive in vivo molecular-genetic imaging developed more recently and is based on nuclear (positron emission tomography [PET], gamma camera, autoradiography) imaging as well as magnetic resonance (MR) and in vivo optical imaging. Molecular-genetic imaging has its roots in both molecular biology and cell biology, as well as in new imaging technologies. The focus of this presentation will be nuclear-based molecular-genetic imaging, but it will comment on the value and utility of combining different imaging modalities. Nuclear-based molecular imaging can be viewed in terms of three different imaging strategies: (1) "indirect" reporter gene imaging; (2) "direct" imaging of endogenous molecules; or (3) "surrogate" or "bio-marker" imaging. Examples of each imaging strategy will be presented and discussed. The rapid growth of in vivo molecular imaging is due to the established base of in vivo imaging technologies, the established programs in molecular and cell biology, and the convergence of these disciplines. The development of versatile and sensitive assays that do not require tissue samples will be of considerable value for monitoring molecular-genetic and cellular processes in animal models of human disease, as well as for studies in human subjects in the future. Noninvasive imaging of molecular-genetic and cellular processes will complement established ex vivo molecular-biological assays that require tissue sampling, and will provide a spatial as well as a temporal dimension to our understanding of various diseases and disease processes.

Adoptive Transfer↗
PubMed2002

Comparison of radiolabeled nucleoside probes (FIAU, FHBG, and FHPG) for PET imaging of HSV1-tk gene expression.

UNLABELLED: The efficacy of 3 radiolabeled probes of current interest for imaging herpes simplex virus type 1 thymidine kinase (HSV1-tk) expression in vivo with PET, including (124)I- or (131)I-labeled 2'-fluoro-2'-deoxy-1-beta-D-arabinofuranosyl-5-iodouracil (FIAU), (18)F-labeled 9-[4-fluoro-3-(hydroxymethyl)butyl]guanine (FHBG), and (18)F-labeled 9-[3-fluoro-1-hydroxy-2-propoxymethyl]guanine (FHPG), was compared. METHODS: Two established rat glioma cell lines, stably transduced RG2TK+ and wild-type RG2, were used for paired comparisons of probe accumulation in vitro and for paired comparisons of subcutaneous xenografts produced from these cell lines in athymic rnu/rnu rats. RESULTS: The in vitro paired probe uptake (0-3 h) comparisons in RG2TK+ cells showed that FIAU accumulation was 15-fold greater than that of FHBG and 41-fold greater than that of FHPG. The net accumulation rate values (+/-SD) calculated for RG2TK+ cells were 0.317 +/- 0.066, 0.022 +/- 0.001, and 0.0077 +/- 0.0003 mL/min/g cells for FIAU, FHBG, and FHPG, respectively. These results and similar uptake studies in RG2 wild-type cells suggest a possible cell membrane transport limitation for FHBG and FHPG. The paired 2-h in vivo uptake studies produced similar differences in RG2TK+ xenografts for FIAU and FHBG (1.22 +/- 0.21 vs. 0.074 +/- 0.49 %dose/g) and for FIAU and FHPG (1.27 +/- 0.14 vs. 0.023 +/- 0.008 %dose/g). These differences were clearly visible on the images. FIAU accumulation at 24 h was 1.53 +/- 0.40 %dose/g. Plasma clearance was FHBG > FHPG >> FIAU. The FIAU images showed significant stomach and some intestinal background radioactivities, whereas hepatobiliary and intestinal background activities were very high for the guanosine analogs (FHBG > FHPG). Dynamic imaging showed early ( approximately 10 min) selective localization of FIAU in RG2TK+ xenografts, whereas FHBG and FHPG are being cleared from the HSV1-tk transduced and wild-type xenografts over the initial 2-h imaging period. CONCLUSION: The in vitro and in vivo results (including the PET images) show that FIAU is a substantially more efficient probe than FHBG or FHPG for imaging HSV1-tk expression, with greater sensitivity and contrast as well as lower levels of abdominal background radioactivity at 2 and 24 h.

Animals↗
PubMedDate not supplied

Multimodality in vivo molecular-genetic imaging.

Multimodality imaging is increasingly being used in molecular-genetic studies in small animals. The coupling of nuclear and optical reporter genes represents the beginning of a far wider application of this technology. Optical imaging and optical reporter systems are cost-effective and time-efficient, they require less resources and space than PET or MRI, and they are particularly well suited for small animal imaging and for in vitro assays to validate different reporter systems. However, optical imaging techniques are limited by depth of light penetration and scatter and do not yet provide optimal quantitative or tomographic information. These issues are not limiting for PET- or MRI-based reporter systems, and PET- and MRI-based animal studies are more easily generalized to human applications. Many of the shortcomings of each modality alone can be overcome by the use of dual- or triple-modality reporter constructs that incorporate the opportunity for PET, fluorescence and bioluminescence imaging. We optimistically expect that some form of tomographic, small animal optical imaging capability will be developed soon, and that this will provide the opportunity for the colocalization of optical signals to anatomical structures provided by tomographic CT and MR imaging.

Animals↗