Geopyxins A–E, <i>ent</i>-Kaurane Diterpenoids from Endolichenic Fungal Strains <i>Geopyxis</i> aff. <i>majalis</i> and <i>Geopyxis</i> sp. AZ0066: Structure–Activity Relationships of Geopyxins and Their Analogues

Four new <i>ent</i>-kaurane diterpenoids, geopyxins A–D (<b>1</b>–<b>4</b>), were isolated from <i>Geopyxis</i> aff. <i>majalis</i>, a fungus occurring in the lichen <i>Pseudevernia intensa</i>, whereas <i>Geopyxis</i> sp. AZ0066 inhabiting the same host afforded two new <i>ent</i>-kaurane diterpenoids, geopyxins E and F (<b>5</b> and <b>6</b>), together with <b>1</b> and <b>3</b>. The structures of <b>1</b>–<b>6</b> were established on the basis of their spectroscopic data, while the absolute configurations were assigned using modified Mosher’s ester method. Methylation of <b>1</b>–<b>3</b>, <b>5</b>, and <b>6</b> gave their corresponding methyl esters <b>7</b>–<b>11</b>. On acetylation, <b>1</b> and <b>7</b> yielded their corresponding monoacetates <b>12</b> and <b>14</b> and diacetates <b>13</b> and <b>15</b>. All compounds were evaluated for their cytotoxic and heat-shock induction activities. Compounds <b>2</b>, <b>7</b>–<b>10</b>, <b>12</b>, <b>14</b>, and <b>15</b> showed cytotoxic activity in the low micromolar range against all five cancer cell lines tested, but only compounds <b>7</b>–<b>9</b>, <b>14</b>, and <b>15</b> were found to activate the heat-shock response at similar concentrations. From a preliminary structure–activity perspective, the electrophilic α,β-unsaturated ketone carbonyl motif present in all compounds except <b>6</b> and <b>11</b> was found to be necessary but not sufficient for both cytotoxicity and heat-shock activation