**2. Results and Discussion**

#### *2.1. Chemistry*

The synthesis of the tricyclic core thiaplakortone isomers **11**–**16** commenced with the generation of 1-tosyl-1*H*-indole-4,7-dione (**10**), which was accessible via known procedures (Scheme 1) [4–6]. Condensation of **10** with 2-aminoethanesulfinic acid [2,7] furnished the regiomeric tricyclic systems **11** and **12** in an 11 to 1 ratio (Scheme 1). Separation of this mixture was not possible by silica flash chromatography however reversed-phase C18 HPLC (MeOH-H2O-0.1%TFA) enabled separation of

the two regioisomers. Confirmation of the chemical structures of **11** and **12** was supported following extensive 1D and 2D NMR data analysis.

Furthermore, a crystal suitable for X-ray analysis was obtained on the major regioisomer **11** (Figure 2) confirming the NMR-assigned structure and establishing the regiochemistry of subsequent compounds in the tricyclic series. Of note, compound **11** crystallized with two molecules in the asymmetric unit; the second molecule displayed disorder (*ca*. 13%) in the thiazine dioxide ring (see supplementary data).

**Scheme 1.** Synthesis of compounds **11**–**16** in the thiaplakortone tricyclic series. (**a**) 2-aminoethanesulfinic acid, H2O, MeCN; (**b**) NaHCO3(aq), MeOH, reflux 2.5 h; (**c**) KOH(aq), MeOH, O2.

**Figure 2.** ORTEP diagram showing one independent molecule for compound **11**; ellipsoids are at the 30% probability level.

Subjection of the mixture of tosyl derivatives **11** and **12** to mild alkaline hydrolysis afforded a sufficient quantity of only compound **13** after purification by reversed-phase HPLC (MeOH-H2O-0.1%TFA).

In an attempt to reverse the regioselectivity observed during the condensation of 2-aminoethanesulfinic acid with **10** and acquire suitable amounts of isomer **14**, the parent, non-tosyl protected system, 1*H*-indole-4,7-dione **17** was prepared according to literature procedures [5,8,9] and exposed to 2-aminoethanesulfinic acid. Gratifyingly, in this system, the opposite regioselectivity was observed and the respective tricyclic systems **13** and **14** were formed in a 1 to 3.3 ratio following analysis of the 1H-NMR spectrum. Purification of this mixture by C18 HPLC (MeOH-H2O-0.1%TFA) enabled **14** to be isolated in sufficient amounts for biological testing. Oxidation of the mixture of **13** and **14** by protocols previously reported [2] afforded a mixture that was subjected to C18 HPLC (MeOH-H2O-0.1%TFA) and yielded the pure compounds **15** and **16**.
