Eight new nitrogenated azaphilones (<b>1</b>-<b>8</b>) and two known compounds (chaetoviridin A and chaetoviridin E, <b>9</b>, <b>10</b>) were isolated from the culture of the deep-sea-derived fungus <i>Chaetomium globosum</i> MP4-S01-7. The absolute configurations of new compounds were elucidated by HSQC-HECADE NMR data, <i>J</i>-based configuration analysis, and modified Mosher's method and finally verified by comparison of recorded and computed NMR chemical shifts from quantum chemical calculations coupled with a statistical procedure (DP4+). All of the compounds were evaluated for their <i>in</i> <i>vitro</i> cytotoxicities against the gastric cancer cell lines MGC803 and AGS, and most of them showed significant inhibition on cancer cell viability at 10 μM. Among them, compounds <b>1</b>, <b>2</b>, and <b>5</b> exerted the most potent cytotoxic activities, with IC<sub>50</sub> values less than 1 μM. Further studies showed that compound <b>2</b> inhibited cell cycle progression, and both compounds <b>1</b> and <b>2</b> induced apoptosis of gastric cancer cells in a concentration-dependent manner.