Natural retinoids inhibit proliferation and induce apoptosis in pancreatic cancer cells previously reported to be retinoid resistant

Background: The anticancer ability of natural retinoids on pancreatic adenocarcinoma, an aggressive tumor, is still controversial. This investigation tested the hypothesis that all-trans retinoic acid can inhibit proliferation and induce apoptosis in pancreatic cancer cell lines. Materials and Methods: Using our previously optimized conditions, the effect of all-trans retinoic acid (atRA, 0.001-10 μM) was tested in ten human pancreatic adenocarcinoma cell lines with various degrees of differentiation. Proliferation was monitored by cell number, [³H]-thymidine incorporation and cell cycle arrest. Apoptosis was investigated morphologically by light and electron microscopy and biochemically by tissue transglutaminase activity (TGase), mitochondrial membrane potential, cell cycle analysis of sub-G₁ cells and detection of fragmented DNA (fragmentation of prelabeled DNA, agarose electrophoresis and TUNEL assays). Results: Retinoic acid caused potent concentration- and time-dependent inhibition of proliferation of all cell lines studied. Cell cycle was arrested at G₁ or G₂ with extensive reduction of number of cells at S-phase after 24 hours of treatment with apoptotic concentration of atRA. Complete inhibition of proliferation was followed by apoptosis as indicated by the progressive accumulation of sub-G₁ apoptotic cells which was confirmed by the more specific DNA fragmentation assays. There were extensive apoptosis-indicative light and electron microscopic changes preceded by phenotypic redifferentiation. TGase was induced between 3-5-folds the control level and its inhibition partially reversed the antiproliferative effect of atRA. Cellular viability during the preapoptotic stage was confirmed by normal mitochondrial membrane potential in the first two days of treatment with the maximum atRA concentration used. However, the potential was progressively reduced with time as a preapoptotic change. Caspase 3-like activity was induced by the apoptotic concentrations of atRA at late time points. However, the redifferentiation indicative changes were not prevented by cotreatment with Ac-DEVE-CHO caspase 3 inhibitor. Conclusions: Together, our results demonstrated the efficient anticancer ability of natural retinoids on human pancreatic cancer cell lines tested, even those previously reported to be retinoid resistant.