Resin-supported arylstannanes as precursors for radiolabeling with iodine: benzaldehydes, benzoic acids, benzamides, and NHS esters

A highly cross-linked polystyrene resin bearing a reactive chlorostannane moiety 1 has been used to generate a variety of arylstannane radiopharmaceutical precursors for no-carrier-added radioiodination. The resins were characterized for their solvent compatibility and sensitivity to acid cleavage. Resin-supported arylstannanes synthesized via their aryl lithium analogues include 3- and 4-stannylbenzaldehydes, 3- and 4-stannylbenzoic acids, and 3- and 4-N-succinimidyl benzoates. A three-step route to the resin-supported stannylbenzoic acids 12a/b was developed through resin-supported benzaldehydes 11a/b. The aldehyde to acid conversion efficiency is \u3e90%, and acid loading capacities of 0.66–0.94 mmol/g were obtained. Resin-supported N-succinimidyl benzoates 16a/b were prepared from the acid with 78%–84% conversion efficiency. Libraries of resin-supported benzamides 19a/b prepared from amine conjugation to corresponding benzoic acids or N-succinimidyl benzoates are described. A third approach describes the preparation of resin-supported benzamides via direct conjugation of the dilithio salt of the intact benzamide to the chlorostannane resin 1. Lastly, as proof-of-principle, a radiolabeling study with iodine-131 (131I) was performed with a resin-supported benzamide to afford the corresponding radioligand in moderate yields, and high radiochemical purity

Collaborative Enhancement of Antibody Binding to Distinct PECAM-1 Epitopes Modulates Endothelial Targeting

Antibodies to platelet endothelial cell adhesion molecule-1 (PECAM-1) facilitate targeted drug delivery to endothelial cells by “vascular immunotargeting.” To define the targeting quantitatively, we investigated the endothelial binding of monoclonal antibodies (mAbs) to extracellular epitopes of PECAM-1. Surprisingly, we have found in human and mouse cell culture models that the endothelial binding of PECAM-directed mAbs and scFv therapeutic fusion protein is increased by co-administration of a paired mAb directed to an adjacent, yet distinct PECAM-1 epitope. This results in significant enhancement of functional activity of a PECAM-1-targeted scFv-thrombomodulin fusion protein generating therapeutic activated Protein C. The “collaborative enhancement” of mAb binding is affirmed in vivo, as manifested by enhanced pulmonary accumulation of intravenously administered radiolabeled PECAM-1 mAb when co-injected with an unlabeled paired mAb in mice. This is the first demonstration of a positive modulatory effect of endothelial binding and vascular immunotargeting provided by the simultaneous binding a paired mAb to adjacent distinct epitopes. The “collaborative enhancement” phenomenon provides a novel paradigm for optimizing the endothelial-targeted delivery of therapeutic agents

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Fludarabine, cytarabine, granulocyte colony-stimulating factor, and idarubicin with gemtuzumab ozogamicin improves event-free survival in younger patients with newly diagnosed aml and overall survival in patients with npm1 and flt3 mutations

Purpose To determine the optimal induction chemotherapy regimen for younger adults with newly diagnosed AML without known adverse risk cytogenetics. Patients and Methods One thousand thirty-three patients were randomly assigned to intensified (fludarabine, cytarabine, granulocyte colony-stimulating factor, and idarubicin [FLAG-Ida]) or standard (daunorubicin and Ara-C [DA]) induction chemotherapy, with one or two doses of gemtuzumab ozogamicin (GO). The primary end point was overall survival (OS). Results There was no difference in remission rate after two courses between FLAG-Ida + GO and DA + GO (complete remission [CR] + CR with incomplete hematologic recovery 93% v 91%) or in day 60 mortality (4.3% v 4.6%). There was no difference in OS (66% v 63%; P = .41); however, the risk of relapse was lower with FLAG-Ida + GO (24% v 41%; P < .001) and 3-year event-free survival was higher (57% v 45%; P < .001). In patients with an NPM1 mutation (30%), 3-year OS was significantly higher with FLAG-Ida + GO (82% v 64%; P = .005). NPM1 measurable residual disease (MRD) clearance was also greater, with 88% versus 77% becoming MRD-negative in peripheral blood after cycle 2 (P = .02). Three-year OS was also higher in patients with a FLT3 mutation (64% v 54%; P = .047). Fewer transplants were performed in patients receiving FLAG-Ida + GO (238 v 278; P = .02). There was no difference in outcome according to the number of GO doses, although NPM1 MRD clearance was higher with two doses in the DA arm. Patients with core binding factor AML treated with DA and one dose of GO had a 3-year OS of 96% with no survival benefit from FLAG-Ida + GO. Conclusion Overall, FLAG-Ida + GO significantly reduced relapse without improving OS. However, exploratory analyses show that patients with NPM1 and FLT3 mutations had substantial improvements in OS. By contrast, in patients with core binding factor AML, outcomes were excellent with DA + GO with no FLAG-Ida benefit

The molecular imaging of herpes simplex virus type-1 thymidine kinase reporter gene expression: Novel fluorine-18-labeled reporter probes

Gene therapy promises to be a new therapeutic avenue for a variety of diseases. However, one of the major limitations to this technology is the difficulty in achieving controlled and efficient delivery of therapeutic genes in vivo. Molecular imaging of reporter genes is a powerful method to monitor and evaluate the location, magnitude, and duration of gene expression following somatic DNA delivery in vivo. The herpes simplex virus type-1 thymidine kinase (HSV1-tk) gene, has evolved as the most investigated reporter gene for the repetitive and non-invasive in vivo imaging of therapeutic gene expression. At present, there remains a need for HSV1-TK enzyme reporter probes for accurately quantifying dynamic HSV1-tk reporter gene expression levels following gene delivery in vivo to correlate with the effectiveness of gene therapy. It is our hypothesis that enhanced HSV1-TK reporter probe efficacy may be attained by rational selection of chemical structures based on successful antiherpes agents that are selectively phosphorylated and trapped by HSV1-TK. Hence, the objective of this dissertation work was to design, synthesize, and biologically evaluate targeted fluorine-18 (18F)-labeled HSV1-TK reporter probes that can be detected by Positron Emission Tomography (PET). Towards this goal, we have developed new synthetic strategies to afford two series of nonradioactive and radioactive thymidine (pyrimidine) nucleoside analogs with (i) [19/18F]fluoroalkyl chains (three, four, or five carbon-unit length) introduced at the C-5 position of the nucleobase and (ii) a C-2\u27 hydrogen or fluorine in the arabino or up position of the ribose sugar. These compounds have never been synthesized nor have they been explored as PET imaging agents for assessing HSV1-tk gene expression in vivo. In vitro evaluations of the 5-[18F]fluoroalkyl nucleosides in a model cellular system demonstrated their sensitivity and selectivity for HSV1-tk expressing cells. Subsequent in vivo validation of these novel probes was done in tumor-bearing nude mice. Biodistribution studies revealed selective uptake of only two of six tracers into HSV1-tk expressing tumors compared to control tumors at 2 h post-injection. Although small animal PET imaging studies with the two candidates could visualize the HSV1-tk positive tumors (tumor to nontarget ratio 1.2-3), accumulation (sensitivity) was not very high as compared to surrounding background tissue, and there was consistent tumor washout over time. Potentially the very fast renal clearance of the radiotracers limits tracer availability to the target site. Studies thus far demonstrate the limited potential of these 5-[18F]fluoroalkyl nucleosides in evaluating HSV1-tk gene expression in vivo. Further studies in animal models and detailed metabolic studies are warranted to understand the contrasting in vitro and in vivo results with regards to sensitivity and selectivity

The role of ligand acid-base reaction in the facile synthesis of Alkali Metal Neodymium Penta- And Heptafluoride Nanocrystals

We report a facile co-precipitation method of synthesizing orthorhombic phase CsNd2F7 and hexagonal phase K2NdF5 nanocrystals, which avoids the use of relatively harsh conditions (e.g., high temperatures and long reaction times in hydrothermal synthesis). By reacting MF (M = Cs and K), Nd(CH3CO2)3, oleic acid (OA), and 1-octadecene in the presence of sufficient amounts of oleylamine ligands (OAm) to ensure a non-acidic environment, CsNd2F7 and K2NdF5 nanocrystals are formed for reaction temperatures (Tr) ranging between 150 and 300 °C. When OAm is in excess of OA and Tr = 300 °C, uniformly distributed CsNd2F7 nanodiscs (∼20 nm in diameter) and sub-10 nm K2NdF5 nanodiscs with an average thickness of 2-3 unit cell layers are formed. When OAm is removed, the acidic mixture produces CsNd2F7 nanocrystals at low Tr (150-200 °C); however, a mixture of NdF3 and Cs-oleate is observed instead when Tr is increased. Similarly, in an acidic enviroment, K2NdF5 nanocrystals rapidly decompose into a mixture of K0.156Nd0.142-yF0.702-3y and y(NdF3) (y 200 °C). Therefore, the penta- and heptafluoride nanocrystals are thermodynamically stable only in a basic medium when OA is consumed, in part by its condensation reaction with OAm. We further demonstrate that the CsNd2F7 and K2NdF5 nanocrystals display good light-to-heat conversion efficiencies (16-27%) and computed tomography (CT) attenuation, making them promising theranostic photothermal imaging/therapy and CT contrast agents.Ministry of Education (MOE)We acknowledge financial support from the Singapore Ministry of Education MoE Tier 1 grants (RG89/20 and RG6/18) and Duke-NUS Block Grant

Spatio-temporal in vivo imaging of ocular drug delivery systems using fiberoptic confocal laser microendoscopy

Subconjunctival injection is an attractive route to administer ocular drugs due to easy trans-scleral access that bypasses anterior ocular barriers, such as the cornea and conjunctiva. While therapeutic effects and pharmacokinetics of the drugs upon subconjunctival injection have been described in some studies, very few assess the ocular distribution of drugs or drug delivery systems (DDS). The latter is critical for the optimization of intraocular DDS design and drug bioavailability to achieve the desired ocular localization and duration of action (e.g., acute versus. prolonged). This study establishes the use of fiberoptic confocal laser microendoscopy (CLM) to qualitatively study the ocular distribution of fluorescent liposomes in real-time in live mice after sub-conjunctival injection. Being designed for in vivo visual inspection of tissues at the microscopic level, this is also the first full description of the CLM imaging method to study spatio-temporal distribution of injectables in the eye after subconjunctival injection.Agency for Science, Technology and Research (A*STAR)Nanyang Technological UniversityNational Research Foundation (NRF)Published versionThis research was funded by NTU-Northwestern Institute for Nanomedicine (NNIN) grant awarded (to SV) and in part by Singapore National Research Foundation Grant AG/CIV/GC70-C/NRF/2013/2 and Singapore’s Health and Biomedical Sciences (HBMS) Industry Alignment Fund PrePositioning (IAF-PP) grant H18/01/a0/018 administered by the Agency for Science, Technology and Research (A*STAR) (to AMC)

Monoclonal antibody (mAb) ligands recognizing distinct extracellular epitopes of PECAM-1.

<p>(<b>A</b>) MAbs investigated in this study to probe the affinity and accessibility to distinct epitopes of human PECAM-1 (huPECAM-1; mAbs 62 and 37) and mouse PECAM-1 (muPECAM-1; mAbs 390 and MEC13.3). Listed is the effect of various anti-PECAM-1 mAbs on PECAM-1-dependent homophilic adhesion, as defined by the aggregation of L-cells fibroblast transfectants expressing PECAM-1 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958-Nakada1" target="_blank">[22]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958-Yan3" target="_blank">[50]</a>. <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958-Yan1" target="_blank">[15]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958-Nakada1" target="_blank">[22]</a>. (<b>B</b>–<b>C</b>) Diagram of immunoreactive regions within PECAM-1 domains 1 and 2. (<b>B</b>) Amino acid (AA) location of distinct non-overlapping epitopes for binding of mAbs 62 and 37 on Ig-domain 1 (IgD1) of huPECAM-1 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958-Nakada1" target="_blank">[22]</a>. (<b>C</b>) AA location of epitopes for mAbs 390 and MEC13.3 on Ig-domain 2 (IgD2) of muPECAM-1 (H. DeLisser, unpublished results). Peptide sequence recognized by mAbs are colored in red.</p

Anti-PECAM-1 [¹²⁵I]-mAb binding in live cells is enhanced by paired mAb directed to adjacent PECAM-1 epitope.

<p>The modulation of PECAM-1 binding was determined by co-incubation of [¹²⁵I]-mAb with indicated concentrations of unlabeled self-paired mAb or paired mAb with cells for 2 h at 4°C. Binding data were plotted as [¹²⁵I]-mAb molecules bound per cell (mAb/cell) and data points were fit as described under “<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#s4" target="_blank">Methods</a>.” (<b>A</b> and <b>B</b>) Unlabeled mAb 62 competitively inhibits binding of [¹²⁵I]-mAb 62 to huPECAM-1 in HUVEC. However, mAb 37 enhances [¹²⁵I]-mAb 62 binding to huPECAM-1 in HUVEC by 1.5−fold over binding of [¹²⁵I]-mAb 62 alone. Interestingly, mAb 62 does not enhance the binding of [¹²⁵I]-mAb 37 (<b><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0034958#pone.0034958.s003" target="_blank">Figure S3</a></b>). (<b>C</b>–<b>D</b>) Collaborative binding studies of mAbs 390 and MEC13.3 with REN-muP cells as described in panel A. Unlabeled self-paired mAb 390 and mAb MEC13.3 competitively inhibit binding of [¹²⁵I]-mAb390 and [¹²⁵I]-mAb MEC13.3 to REN-muP cells, respectively. In contrast, mAb pairs [¹²⁵I]-mAb 390/MEC13.3 and [¹²⁵I]-mAb MEC13.3/390 enhance binding by ∼1.5−fold and ∼2.7−fold, respectively, over [¹²⁵I]-mAb alone (***, P<0.001, <i>n</i> = 3–4).</p