High Yielding Preparation of Dicarba-<i>closo</i>-dodecaboranes Using a Silver(I) Mediated Dehydrogenative Alkyne-Insertion Reaction

The synthesis of 1,2-dicarba-<i>closo</i>-dodecaboranes (<i>ortho</i>-carboranes) is often low yielding which is a critical issue given the increasing use of boron clusters in material science and medicinal chemistry. To address this barrier, a series of Cu, Ag, and Au salts were screened to identify compounds that would enhance the yields of <i>ortho</i>-caboranes produced when treating alkynes with B₁₀H₁₂(CH₃CN)₂. Using a variety of functionalized ligands including mono- and polyfunctional internal and terminal alkynes, significant increases in yield were observed when AgNO₃ was used in catalytic amounts. AgNO₃ appears to prevent unwanted reduction/hydroboration of the alkyne prior to carborane formation, and the process is compatible with aryl, halo, hydroxy, nitrile, carbamate, and carbonyl functionalized alkynes

A ^99mTc-Labelled Tetrazine for Bioorthogonal Chemistry. Synthesis and Biodistribution Studies with Small Molecule <i>trans</i>-Cyclooctene Derivatives

<div><p>A convenient strategy to radiolabel a hydrazinonicotonic acid (HYNIC)-derived tetrazine with ^99mTc was developed, and its utility for creating probes to image bone metabolism and bacterial infection using both active and pretargeting strategies was demonstrated. The ^99mTc-labelled HYNIC-tetrazine was synthesized in 75% yield and exhibited high stability <i>in vitro</i> and <i>in vivo</i>. A <i>trans</i>-cyclooctene (TCO)-labelled bisphosphonate (TCO-BP) that binds to regions of active calcium metabolism was used to evaluate the utility of the labelled tetrazine for bioorthogonal chemistry. The pretargeting approach, with ^99mTc-HYNIC-tetrazine administered to mice one hour after TCO-BP, showed significant uptake of radioactivity in regions of active bone metabolism (knees and shoulders) at 6 hours post-injection. For comparison, TCO-BP was reacted with ^99mTc-HYNIC-tetrazine before injection and this active targeting also showed high specific uptake in the knees and shoulders, whereas control ^99mTc-HYNIC-tetrazine alone did not. A TCO-vancomycin derivative was similarly employed for targeting <i>Staphylococcus aureus</i> infection <i>in vitro</i> and <i>in vivo</i>. Pretargeting and active targeting strategies showed 2.5- and 3-fold uptake, respectively, at the sites of a calf-muscle infection in a murine model, compared to the contralateral control muscle. These results demonstrate the utility of the ^99mTc-HYNIC-tetrazine for preparing new technetium radiopharmaceuticals, including those based on small molecule targeting constructs containing TCO, using either active or pretargeting strategies.</p></div

Synthesis scheme for ^99mTc-HYNIC-tetrazine-TCO-vancomycin (7) from TCO-vancomycin (6) and 4.

<p>Synthesis scheme for ^99mTc-HYNIC-tetrazine-TCO-vancomycin (7) from TCO-vancomycin (6) and 4.</p

Synthesis scheme for the preparation of 3.

<p>A protected form of HYNIC (<b>1</b>) was coupled to a commercially available tetrazine to form <b>2</b>. The Boc group was removed prior to labelling by treatment with TFA in DCM to produce <b>3</b>. Tz* = (4-(1,2,4,5-tetrazin-3-yl)phenyl) methanamine.</p

Biodistribution data for 4.

<p>Data are presented as the mean (± SEM) percent injected dose per gram (%ID/g) for selected tissues and fluids from CD1 mice at 0.5, 1, 2 and 6 h post injection (n = 3 per time point). Approximately 0.88 MBq were administered per mouse. Full biodistribution data can be found in the supporting information (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0167425#pone.0167425.s004" target="_blank">S4 File</a>).</p

Radiolabelling scheme for the HYNIC-tetrazine ligand 3 and HPLC chromatogram of the isolated product 4.

<p>^99mTc labelling of the HYNIC-tetrazine <b>3</b> (A). A γ-HPLC chromatogram of the purified final product <b>4</b> (B).</p

Biodistribution data for active targeting of <i>S</i>. <i>aureus</i> infection using ^99mTc-HYNIC-tetrazine-TCO-vancomycin (7).

<p>Compounds <b>4</b> and <b>6</b> were combined prior to i.v. injection of Balb/c mice (n = 3 per time point). Select fluids and tissues were collected at 1 (gray bars) and 6 h (black bars) post injection, including the infected calf muscle (right), and the non-infected calf muscle (left). Data are expressed as the mean percent injected dose per gram (%ID/g) ± SEM. Tabulated biodistribution data can be found in the supporting information (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0167425#pone.0167425.s004" target="_blank">S4 File</a>).</p

Synthesis scheme for the actively targeted derivative, ^99mTc-HYNIC-tetrazine-TCO-BP (5).

<p>The TCO derivative of the bisphosphonate, alendronate (TCO-BP) was mixed with 4 prior to administration to mice.</p

Biodistribution data comparing active targeting to pretargeting.

<p>Active targeting with ^99mTc-HYNIC-tetrazine-TCO-BP (<b>5</b>) (black bars) is compared to pretargeting of TCO-BP administered 1 h prior to <b>4</b> (gray bars). Data are expressed as the mean (± SEM) %ID/g for selected tissues and fluids from Balb/c mice (n = 3 per time point). Tabulated biodistribution data can be found in the supporting information (<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0167425#pone.0167425.s004" target="_blank">S4 File</a>).</p

Binding of 4 to <i>S</i>. <i>aureus in vitro</i> using TCO-vancomycin 6.

<p><i>S</i>. <i>aureus</i> were pretreated with <b>6</b> in the absence (gray) or presence (black) of a 10-fold excess of vancomycin. The mean percentages (± SEM) of total radioactivity bound after 1 and 6 h incubation times with <b>4</b> are shown.</p