Radiation risk in nuclear medicine.

Given the central roles that anatomical and functional imaging now play in medical practice, there have been concerns about the increasing levels of radiation exposure and their potential hazards. Despite incomplete quantitative knowledge of the risks, it is prudent to think of radiation, even at low doses, as a potential, albeit weak, carcinogen. Thus, we are obliged to minimize its dose and optimize its benefits. Hopefully, time will clarify our estimates of the dangers. Until then, we should educate and assure our patients, their families, and colleagues that the risks have been taken into account and are well balanced by the benefits.

Bert Lester Vallee: a lasting legacy.

The enlightened formation, by Bert and Kuggie Vallee, of a procedure whereby senior scientists spend short time of about a month at Harvard, Oxford and other institutions, is illustrated by the views and opinions of those selected.

Effect of distance between decaying (125)I and DNA on Auger-electron induced double-strand break yield.

PURPOSE: To determine the possible effects of (125)I-to-DNA distance on the magnitude and mechanism of Auger-electron induced-double-strand break (DSB) production. MATERIALS AND METHODS: We have synthesized a series of (125)I-labeled Hoechst (H) derivatives ((125)IE-H, (125)IB-H, (125)I-C(8)-H and (125)I-C(12)-H). While all four molecules share a common DNA minor groove binding bis-benzimidazole motif, they are designed to position (125)I at varying distances from the DNA helix. Each Hoechst derivative was incubated at 4°C in phosphate buffered saline (PBS) together with supercoiled (SC) (3)H-pUC19 plasmid DNA (ratio 3:1) ± the •OH scavenger dimethyl sulfoxide (DMSO) (0.2 M). Aliquots were analyzed on agarose gels over time and DSB yields per decay of (125)I atom were determined. Docking of the iodinated compounds on a DNA molecule was carried out to determine the distance between the iodine atom and the central axis of DNA. RESULTS: In the absence of DMSO, the results show that the DSB yields decrease monotonically as the (125)I atom is distanced - by 10.5 Å to 13.9 Å - from the DNA helix ((125)IEH: 0.52 ± 0.01; (125)IB-H: 0.24 ± 0.03; (125)I-C(8)-H: 0.18 ± 0.02; (125)I-C(12)-H: 0.10 ± 0.00). In the presence of DMSO, DSB yields for (125)IEH (0.49 ± 0.02) and (125)IB-H (0.26 ± 0.04) remain largely unchanged indicating that DSB are entirely produced by direct effects. Strikingly, (125)I-C(8)-H or (125)I-C(12)-H, did not produce detectable DSB in the presence of DMSO under similar conditions suggesting when (125)I atom is positioned > 12 Å from the DNA, DSB are entirely produced by indirect effects. CONCLUSION: These results suggest that at a critical distance between the (125)I atom and the DNA helix, DSB production switches from an 'all' direct to an 'all' indirect mechanism, the latter situation being comparable to the decay of (125)I free in solution. These experimental findings were correlated with theoretical expectations based on microdosimetry.

Bystander effect in tumor cells produced by Iodine-125 labeled human lymphocytes.

PURPOSE: To investigate the ability of human lymphocytes labeled with DNA-incorporated (125)I to exert an inhibitory (antiproliferative) bystander effect on co-cultured human colon adenocarcinoma LS174T cells in vitro. MATERIALS AND METHODS: Human peripheral blood lymphocytes were stimulated to synthesize DNA in the presence of phytohemagglutinin (PHA) and labeled with 5-[(125)I]iodo-2'-deoxyuridine. Human colon adenocarcinoma LS174T cells were co-cultured with the (125)I-labeled lymphocytes in various ratios for 5 days and the proliferation of the LS174T cells was assessed. Further, the supernatant media from these co-cultures were: (i) Transferred to LS174T cells and their proliferation measured after 5 days, (ii) used to assess the clonogenic survival of LS174T cells, and (iii) screened for factors that suppress growth. RESULTS: A significant reduction in the proliferation of LS174T cells was observed when co-cultured either with (125)I-labeled lymphocytes (56 ± 3.5%) or the supernatant media (52.5 ± 1.3%) obtained from these co-cultures. Clonogenic survival of LS174T cells grown in the supernatant media corroborated the decrease in tumor cell growth. CONCLUSION: The observed reduction in the proliferation of LS174T cells in presence of (125)I-labeled lymphocytes or media obtained from such co-cultures can be attributed to an inhibitory (antiproliferative) bystander effect, probably mediated by factor(s) released from the dying (125)I-labeled lymphocytes.

Minimizing and communicating radiation risk in pediatric nuclear medicine.

The value of pediatric nuclear medicine is well established. Pediatric patients are referred to nuclear medicine from nearly all pediatric specialties including urology, oncology, cardiology, gastroenterology, and orthopedics. Radiation exposure is associated with a potential, small, risk of inducing cancer in the patient later in life and is higher in younger patients. Recently, there has been enhanced interest in exposure to radiation from medical imaging. Thus, it is incumbent on practitioners of pediatric nuclear medicine to have an understanding of dosimetry and radiation risk to communicate effectively with their patients and their families. This article reviews radiation dosimetry for radiopharmaceuticals and also CT given the recent proliferation of PET/CT and SPECT/CT. It also describes the scientific basis for radiation risk estimation in the context of pediatric nuclear medicine. Approaches for effective communication of risk to patients' families are discussed. Lastly, radiation dose reduction in pediatric nuclear medicine is explicated.

Target discovery from data mining approaches.

Data mining of available biomedical data and information has greatly boosted target discovery in the 'omics' era. Target discovery is the key step in the biomarker and drug discovery pipeline to diagnose and fight human diseases. In biomedical science, the 'target' is a broad concept ranging from molecular entities (such as genes, proteins and miRNAs) to biological phenomena (such as molecular functions, pathways and phenotypes). Within the context of biomedical science, data mining refers to a bioinformatics approach that combines biological concepts with computer tools or statistical methods that are mainly used to discover, select and prioritize targets. In response to the huge demand of data mining for target discovery in the 'omics' era, this review explicates various data mining approaches and their applications to target discovery with emphasis on text and microarray data analysis. Two emerging data mining approaches, chemogenomic data mining and proteomic data mining, are briefly introduced. Also discussed are the limitations of various data mining approaches found in the level of database integration, the quality of data annotation, sample heterogeneity and the performance of analytical and mining tools. Tentative strategies of integrating different data sources for target discovery, such as integrated text mining with high-throughput data analysis and integrated mining with pathway databases, are introduced.

Human placental alkaline phosphatase-mediated hydrolysis correlates tightly with the electrostatic contribution from tail group.

Human placental alkaline phosphatase has been identified as a hydrolase that is significantly overexpressed on the surface of various solid tumor cells, and is therefore a suitable prodrug design target for non-invasive cancer imaging and therapy. Structure-based prediction of enzymatic activities is essential for rational prodrug design. We have been probing the catalytic proficiency--(k(cat) /K(M) )/k(w)--of placental alkaline phosphatase toward several widely diverse substrate structures experimentally and correlating these results to in silico predictions that are based on the free energy estimates obtained from docking of each substrate structure with placental alkaline phosphatase. We have found that electrostatic contribution from the tail group is the most crucial factor to determine the catalytic efficiencies of the substrates. The electrostatic contribution and the total binding energy of the tail group are well correlated with catalytic efficiencies (R² = 0.79 and 0.89, respectively). However, hydrophobic contribution from the tail group does not correlate with the catalytic efficiencies (negative correlation, R² = 0.27). This supports the prior hypothesis stating that alkaline phosphatase-mediated differential hydrolysis of its substrates is attributable to the differential interactions with the tail group, determined by the electrostatic contributions from the non-bridging oxygen atoms. Calculation of the electrostatic potentials within the active site of human placental alkaline phosphatase also suggests that the local positive electrostatic environment may account for its capability to distinguish various substrates. Our study is likely to have immediate implications in the design of prodrugs against human placental alkaline phosphatase and other esterases overexpressed by human tumor cells.

Novel method for quantifying radiation-induced single-strand-break yields in plasmid DNA highlights 10-fold discrepancy.

The widely used agarose gel electrophoresis method for assessing radiation-induced single-strand-break (SSB) yield in plasmid DNA involves measurement of the fraction of relaxed-circular (C) form that migrates independently from the intact supercoiled (SC) form. We rationalized that this method may underestimate the SSB yield since the position of the relaxed-circular form is not altered when the number of SSB per DNA molecule is >1. To overcome this limitation, we have developed a novel method that directly probes and quantifies SSBs. Supercoiled (3)H-pUC19 plasmid samples were irradiated with γ-rays, alkali-denatured, dephosphorylated, and kinated with γ-[(32)P]ATP, and the DNA-incorporated (32)P activities were used to quantify the SSB yields per DNA molecule, employing a standard curve generated using DNA molecules containing a known number of SSBs. The same irradiated samples were analyzed by agarose gel and SSB yields were determined by conventional methods. Comparison of the data demonstrated that the mean SSB yield per plasmid DNA molecule of [21.2±0.59]×10(-2)Gy(-1) as measured by direct probing is ~10-fold higher than that obtained from conventional gel-based methods. These findings imply that the SSB yields inferred from agarose gels need reevaluation, especially when they were utilized in the determination of radiation risk.

Minimizing and communicating radiation risk in pediatric nuclear medicine.

The value of pediatric nuclear medicine is well established. Pediatric patients are referred to nuclear medicine from nearly all pediatric specialties including urology, oncology, cardiology, gastroenterology, and orthopedics. Radiation exposure is associated with a potential, small, risk of inducing cancer in the patient later in life and is higher in younger patients. Recently, there has been enhanced interest in exposure to radiation from medical imaging. Thus, it is incumbent on practitioners of pediatric nuclear medicine to have an understanding of dosimetry and radiation risk to communicate effectively with their patients and their families. This article reviews radiation dosimetry for radiopharmaceuticals and also CT given the recent proliferation of PET/CT and SPECT/CT. It also describes the scientific basis for radiation risk estimation in the context of pediatric nuclear medicine. Approaches for effective communication of risk to patients' families are discussed. Lastly, radiation dose reduction in pediatric nuclear medicine is explicated.

Integrated bioinformatics analysis for cancer target identification.

The exponential growth of high-throughput Omics data has provided an unprecedented opportunity for new target identification to fuel the dried-up drug discovery pipeline. However, the bioinformatics analysis of large amount and heterogeneous Omics data has posed a great deal of technical challenges for experimentalists who lack statistical skills. Moreover, due to the complexity of human diseases, it is essential to analyze the Omics data in the context of molecular networks to detect meaningful biological targets and understand disease processes. Here, we describe an integrated bioinformatics analysis strategy and provide a running example to identify suitable targets for our in-house Enzyme-Mediated Cancer Imaging and Therapy (EMCIT) technology. In addition, we go through a few key concepts in the process, including corrected false discovery rate (FDR), Gene Ontology (GO), pathway analysis, and tissue specificity. We also describe popular programs and databases which allow the convenient annotation and network analysis of Omics data. We provide a practical guideline for researchers to quickly follow the protocol described and identify those targets that are pertinent to their work.

General approach to identifying potential targets for cancer imaging by integrated bioinformatics analysis of publicly available genomic profiles.

Molecular imaging has moved to the forefront of drug development and biomedical research. The identification of appropriate imaging targets has become the touchstone for the accurate diagnosis and prognosis of human cancer. Particularly, cell surface- or membrane-bound proteins are attractive imaging targets for their aberrant expression, easily accessible location, and unique biochemical functions in tumor cells. Previously, we published a literature mining of potential targets for our in-house enzyme-mediated cancer imaging and therapy technology. Here we present a simple and integrated bioinformatics analysis approach that assembles a public cancer microarray database with a pathway knowledge base for ascertaining and prioritizing upregulated genes encoding cell surface- or membrane-bound proteins, which could serve imaging targets. As examples, we obtained lists of potential hits for six common and lethal human tumors in the prostate, breast, lung, colon, ovary, and pancreas. As control tests, a number of well-known cancer imaging targets were detected and confirmed by our study. Further, by consulting gene-disease and protein-disease databases, we suggest a number of significantly upregulated genes as promising imaging targets, including cell surface-associated mucin-1, prostate-specific membrane antigen, hepsin, urokinase plasminogen activator receptor, and folate receptors. By integrating pathway analysis, we are able to organize and map "focused" interaction networks derived from significantly dysregulated entity pairs to reflect important cellular functions in disease processes. We provide herein an example of identifying a tumor cell growth and proliferation subnetwork for prostate cancer. This systematic mining approach can be broadly applied to identify imaging or therapeutic targets for other human diseases.

Managing radiation use in medical imaging: a multifaceted challenge.

This special report aims to inform the medical community about the many challenges involved in managing radiation exposure in a way that maximizes the benefit-risk ratio. The report discusses the state of current knowledge and key questions in regard to sources of medical imaging radiation exposure, radiation risk estimation, dose reduction strategies, and regulatory options.

Target discovery from data mining approaches.

Data mining of available biomedical data and information has greatly boosted target discovery in the 'omics' era. Target discovery is the key step in the biomarker and drug discovery pipeline to diagnose and fight human diseases. In biomedical science, the 'target' is a broad concept ranging from molecular entities (such as genes, proteins and miRNAs) to biological phenomena (such as molecular functions, pathways and phenotypes). Within the context of biomedical science, data mining refers to a bioinformatics approach that combines biological concepts with computer tools or statistical methods that are mainly used to discover, select and prioritize targets. In response to the huge demand of data mining for target discovery in the 'omics' era, this review explicates various data mining approaches and their applications to target discovery with emphasis on text and microarray data analysis. Two emerging data mining approaches, chemogenomic data mining and proteomic data mining, are briefly introduced. Also discussed are the limitations of various data mining approaches found in the level of database integration, the quality of data annotation, sample heterogeneity and the performance of analytical and mining tools. Tentative strategies of integrating different data sources for target discovery, such as integrated text mining with high-throughput data analysis and integrated mining with pathway databases, are introduced.

DNA double-strand breaks induced by decay of (123)I-labeled Hoechst 33342: role of DNA topology.

PURPOSE: To determine double-strand-break (DSB) yields produced by decay of minor-groove-bound (123)I-labeled Hoechst 33342 ((123)IEH) in supercoiled (SC) and linear (L) forms of pUC19 DNA, to compare strand-break efficiency of (123)IEH with that of (125)IEH, and to examine the role of DNA topology in DSB induction by these Auger electron emitters. MATERIALS AND METHODS: Tritium-labeled SC and L pUC19 DNA were incubated with (123)IEH (0-10.9 MBq) at 4 degrees C. After (123)I had completely decayed (10 days), samples were analyzed on agarose gel, and single-strand-break (SSB) and DSB yields were measured. RESULTS: Each (123)I decay in SC DNA produces a DSB yield of 0.18 +/- 0.01. On the basis of DSB yields for (125)IEH (0.52 +/- 0.02 for SC and 1.62 +/- 0.07 for L, reported previously) and dosimetric expectations, a DSB yield of approximately 0.5 (3 x 0.18) per (123)I decay is expected for L DNA. However, no DSB are observed for the L form, even after approximately 2 x 10(11) decays of (123)I per microg DNA, whereas a similar number of (125)I decays produces DSB in approximately 40% of L DNA. CONCLUSION: (123)IEH-induced DSB yield for SC but not L DNA is consistent with the dosimetric expectations for Auger electron emitters. These studies highlight the role of DNA topology in DSB production by Auger emitters and underscore the failure of current theoretical dosimetric methods per se to predict the magnitude of DSB.

Synthesis and application of molecular probe for detection of hydroxyl radicals produced by Na(125)I and gamma-rays in aqueous solution.

PURPOSE: To synthesize N-(3-(3-aminopropylamino)propyl)-2-oxo-2H-chromene-3-carboxamide (7), a novel DNA-binding, coumarin-based, fluorescent hydroxylradical ((*)OH) indicator and to assess its quantum efficiency compared with that of coumarin-3-carboxylic acid (1) and N1,N12-bis[2-oxo-2H-chromene-3-carbonyl]- 1,12-diamine-4,9-diazadodecane (9). MATERIALS AND METHODS: Using computer-generated molecular modeling, 7 and 9 and their respective 7-hydroxylated derivatives 8 and 10 were docked onto DNA dodecamer d(CGCGAATTCGCG)2, the ligand-DNA complexes were energy minimized, and binding free energies and inhibition constants were calculated. Compound 7 was judged an appropriate target molecule and was synthesized. Compounds 1, 7, and 9 were incubated with Na(125)I or irradiated with (137)Cs gamma-rays, and the influence of pH, dose, type of radiation, and the concentration of indicator on fluorescence yield were determined. RESULTS: Non-fluorescent 7 and 9 are converted to fluorescent, 7-hydroxylated derivatives 8 and 10 after interaction with (*)OH in aqueous solution. For 1, 7, and 9, hydroxylation yield increases linearly with both Na(125)I dose (0-700 x 10(6) decays) and (137)Cs dose (0-11.0 Gy). Fluorescence induction is significantly reduced at acidic pH and the fluorescent quantum yield of 8 is approximately 3 times that of 2 or 10 at pH 7.0. With Na(125)I incubation and gamma-ray irradiation, the fluorescence signal of 7 increases linearly with concentration and saturates at approximately 50 microM. CONCLUSION: Compound 7 quantifies lower concentrations of (*)OH than do 1 and 9. This detector is therefore likely to be a good reporter of (*)OH produced within a few nanometers of DNA.

Integrative genomic data mining for discovery of potential blood-borne biomarkers for early diagnosis of cancer.

BACKGROUND: With the arrival of the postgenomic era, there is increasing interest in the discovery of biomarkers for the accurate diagnosis, prognosis, and early detection of cancer. Blood-borne cancer markers are favored by clinicians, because blood samples can be obtained and analyzed with relative ease. We have used a combined mining strategy based on an integrated cancer microarray platform, Oncomine, and the biomarker module of the Ingenuity Pathways Analysis (IPA) program to identify potential blood-based markers for six common human cancer types. METHODOLOGY/PRINCIPAL FINDINGS: In the Oncomine platform, the genes overexpressed in cancer tissues relative to their corresponding normal tissues were filtered by Gene Ontology keywords, with the extracellular environment stipulated and a corrected Q value (false discovery rate) cut-off implemented. The identified genes were imported to the IPA biomarker module to separate out those genes encoding putative secreted or cell-surface proteins as blood-borne (blood/serum/plasma) cancer markers. The filtered potential indicators were ranked and prioritized according to normalized absolute Student t values. The retrieval of numerous marker genes that are already clinically useful or under active investigation confirmed the effectiveness of our mining strategy. To identify the biomarkers that are unique for each cancer type, the upregulated marker genes that are in common between each two tumor types across the six human tumors were also analyzed by the IPA biomarker comparison function. CONCLUSION/SIGNIFICANCE: The upregulated marker genes shared among the six cancer types may serve as a molecular tool to complement histopathologic examination, and the combination of the commonly upregulated and unique biomarkers may serve as differentiating markers for a specific cancer. This approach will be increasingly useful to discover diagnostic signatures as the mass of microarray data continues to grow in the 'omics' era.

Method to eliminate linear DNA from mixture containing nicked circular, supercoiled, and linear plasmid DNA.

Preparations of circular plasmid DNA in either supercoiled or nicked circular form often are contaminated with undesired linear DNA fragments arising from shearing/degradation of chromosomal DNA or linearization of plasmid DNA itself. We report a simple enzymatic method, using a combination of lambda exonuclease and RecJ(f), for the selective removal of linear DNA from such mixtures. lambda exonuclease digests one strand of linear duplex DNA in the 5' to 3' direction, whereas RecJ(f), a single-strand-specific exonuclease, digests the remaining complementary single strand into mononucleotides. This combination of exonucleases can remove linear DNA from a mixture of linear and supercoiled DNA, leaving the supercoiled form intact. Furthermore, the inability of lambda exonuclease to initiate digestion at nicks or gaps enables the removal of undesired linear DNA when nicked circular DNA has been enzymatically prepared from supercoiled DNA. This method can be useful in the preparation of homogeneous circular plasmid DNA required for therapeutic applications and biophysical studies.

Auger electron-induced double-strand breaks depend on DNA topology.

From a structural perspective, the factors controlling and the mechanisms underlying the toxic effects of ionizing radiation remain elusive. We have studied the consequences of superhelical/torsional stress on the magnitude and mechanism of DSBs induced by low-energy, short-range, high-LET Auger electrons emitted by (125)I, targeted to plasmid DNA by m-[(125)I]iodo-p-ethoxyHoechst 33342 ((125)IEH). DSB yields per (125)I decay for torsionally relaxed nicked (relaxed circular) and linear DNA (1.74+/-0.11 and 1.62+/-0.07, respectively) are approximately threefold higher than that for torsionally strained supercoiled DNA (0.52+/-0.02), despite the same affinity of all forms for (125)IEH. In the presence of DMSO, the DSB yield for the supercoiled form remains unchanged, whereas that for nicked and linear forms decreases to 1.05+/-0.07 and 0.76+/-0.03 per (125)I decay, respectively. DSBs in supercoiled DNA therefore result exclusively from direct mechanisms, and those in nicked and linear DNA, additionally, from hydroxyl radical-mediated indirect effects. Iodine-125 decays produce hydroxyl radicals along the tracks of Auger electrons in small isolated pockets around the decay site. We propose that relaxation of superhelical stress after radical attack could move a single-strand break lesion away from these pockets, thereby preventing further breaks in the complementary strand that could lead to DSBs.

Novel prodrugs for targeting diagnostic and therapeutic radionuclides to solid tumors.

Most cancer therapeutics (chemo, radiation, antibody-based, anti-angiogenic) are at best partially and/or temporarily effective. In general, the causes for failure can be summarized as: (i) poor diffusion and/or nonuniform distribution of drug/prodrug molecules in solid tumors; (ii) high drug concentration and retention in normal tissues (leading to side effects); (iii) requirement for plasma-membrane permeability and/or internalization of drug/prodrug molecules; (iv) low uptake of drug by tumor; (v) lack of retention of drug within tumor (most have gradient-driven reversible binding); and (vi) multidrug resistance. We are developing an innovative technology that aims to surmount these problems by actively concentrating and permanently entrapping radioimaging and radiotherapeutic prodrugs specifically within solid tumors. The approach will enable noninvasive sensing (imaging) and effective therapy of solid tumors, allowing tumor detection, diagnosis, and treatment to be closely coupled (personalized medicine).