## 2 Results and Discussion
### \({}^{11}\)C-Methylation of a Base-Labile Protected Desmethyl Precursor  
Due to its relatively high nucleophilicity, we next assessed whether radiomethylation could be achieved after protection of the amidic nitrogen in precursor **8** with a pivaloyloxymethyl (Pom) protecting group (Figure 2, route c). Accordingly, \(O\)-benzyl urea **7** was reacted with chloromethyl pivalate and potassium carbonate in DMF to produce \(N\)-protected intermediate **9** in a >90% yield (Figure 5). Subsequent O-debenzylation of **9** by CTH under the same conditions as described above for **7** resulted in formation of pivaloyl ester **10a** instead of the expected Pom-protected radiolabeling precursor **10**. This result can most likely be explained by rapid base-induced migration of the Pom protecting group with  
Figure 4: ESI(+) HPLC-MS analysis of the crude product solution obtained by reaction of precursor **8** with methyl triflate. (**A**) Calculated \(m/z\) values for the different main products and the derived benzothiazol fragments. (**B**) Total ion current (TIC) chromatogram filtered to \(m/z\) 407, which corresponds to to zadenant (\(\mathrm{t_{R}}=10.4\) min) and its regioisomers (\(\mathrm{t_{R}}=7.6\) and \(11.2\) min). (**C**) TIC filtered to \(m/z\) 292, which corresponds to the methylated benzothiazol fragment expected for tozadenant and its \(N\)-methylated regioisomer. The peak at 6.5 min (\(m/z=421\)) corresponds to a product formed by double methylation and is therefore not relevant for no-carrier-added \({}^{11}\)C-methylation reactions. HPLC conditions: Column: Phenomenex Kintex®F5, 5 \(\upmu\)m, 250 \(\times\) 4.6 mm; eluent: 25% McN in \(\mathrm{H_{2}O}+0.2\%\) HCOOH, flow rate: 1.5 mL/min.  
the concomitant release of formaldehyde, as previously observed during Pom-protection of the nucleobase adenine [29]. Thus, catalytic decomposition of ammonium formate is well known to release ammonia [30], which could result in sufficient alkalinity to induce the observed acyl migration. In line with this assumption, \(O\)-debenzylation of **9** by CTH in a mixture of MeOH and acetic acid instead of DMF furnished the desired radiolabeling precursor **10** in a 74% yield as a bench-stable powder.  
However, first screening experiments by methylation of **10** with non-radioactive MeI (0.1 equivalents) using different solvents (DMF, DMSO or acetonitrile), bases (Cs\({}_{2}\)CO\({}_{3}\), K\({}_{2}\)CO\({}_{3}\), NaOH, (\(n\)-Bu)\({}_{4}\)NOH or NaH) and reaction temperatures (20-120 \({}^{\circ}\)C) followed by deprotection also did not furnish the desired product tozadenant. Suspecting insufficient base stability of the precursor, a solution of **10** in DMF was treated with 1 N aqueous NaOH (two equivalents) and heated to 60 \({}^{\circ}\)C to simulate the slightly basic radiolabeling conditions. ESI HPLC-MS analysis revealed almost complete disappearance of **10** (\(m/z\) 507) within minutes and formation of two new compounds (\(m/z\) 477 and \(m/z\) 393) in a ratio of 9:1 (Figure 6). Based on the mass difference of 30 units compared to **10**, the main product (\(m/z\) 477) was formed by elimination of formaldehyde, indicating that it corresponded to compound **10a** produced by base-induced acyl migration. The mass of the second product (\(m/z\) 393) was consistent with that of the Pom-deprotected product, indicating that it corresponded to compound **8**. Thus, precursor **10** proved to be unsuitable for preparation of [11]C]tozadenant, since even the slightly basic conditions required for radiomethylation resulted in rapid base-induced acyl migration with formation of pivaloyl ester **10a** and (to a lesser extent) hydrolysis into the non-protected precursor **8**.