Aporphine Alkaloid N-Nornuciferine from Nelumbo nucifera Induces MAPK-Associated Autophagy Under Nutrient Deprivation in Human Pancreatic Cancer Cells

Abstract

Background: Pancreatic cancer cells adapt to nutrient stress by activating metabolic pathways that facilitate survival and proliferation within the tumor microenvironment, a phenomenon known as austerity. Targeting this resilience presents a strategy for developing chemotherapeutic agents with anti-austere properties. Methods: Phytochemical investigation of Nelumbo nucifera petals led to the isolation of six alkaloids: three benzylisoquinolines (1–3), two aporphines (4, 5), and one proaporphine (6). Preferential cytotoxicity under nutrient deprivation was evaluated. The most active compound was further analyzed in MIA PaCa-2 cells using proliferation, migration, colony formation, and three-dimensional spheroid growth assays, with additional selective cytotoxicity assessment in KLM-1 cells. Mechanistic studies examined MAPK family signaling, autophagy-associated alterations, and apoptosis. Results: (−)-N-nornuciferine (5) exhibited considerable preferential cytotoxicity against MIA PaCa-2 cells (PC50 = 0.53 µM) and KLM-1 cells (PC50 = 1.11 µM). Under nutrient-rich conditions, 5 suppressed the proliferation, migration, colony formation, and spheroid growth of MIA PaCa-2 cells. Under nutrient deprivation, 5 increased MAPK phosphorylation and LC3-II accumulation while reducing p62/SQSTM1 levels. Chloroquine (CQ) further increased LC3-II accumulation, particularly at 10 µM 5, compared with CQ alone. However, autophagy inhibitors did not rescue 5-induced loss of viability, and the pan-caspase inhibitor Z-VAD-FMK also failed to significantly mitigate cytotoxicity. No detectable PARP or caspase-3 cleavage was observed. Conclusions: (−)-N-nornuciferine (5) exhibits anti-austere and anticancer activities in pancreatic ductal adenocarcinoma (PDAC) models, accompanied by MAPK phosphorylation and autophagy-related changes under nutrient deprivation conditions. Conventional caspase-dependent apoptosis is unlikely to be the main contributor to this process.