PPARγ inhibitors reduce tubulin protein levels by a PPARγ, PPARδ and proteasome-independent mechanism, resulting in cell cycle arrest, apoptosis and reduced metastasis of colorectal carcinoma cells

The nuclear transcription factor peroxisome proliferator-activated receptor-gamma (PPAR gamma) has been identified as an important therapeutic target in murine models of colorectal cancer (CRC). To examine whether PPAR gamma inhibition has therapeutic effects in late-stage CRC, the effects of PPAR gamma inhibitors on CRC cell survival were examined in CRC cell lines and a murine CRC model. Low doses (0.1-1 mu M) of PPAR gamma inhibitors (T0070907, GW9662 and BADGE) did not affect cell survival, while higher doses (10-100 mu M) of all 3 PPAR gamma inhibitors caused caspase-dependent apoptosis in HT-29, Caco-2 and LoVo CRC cell lines. Apoptosis was preceded by altered cell morphology, and this alteration was not prevented by caspase inhibition. PPAR gamma inhibitors also caused dual G and M cell cycle arrest, which was not required for apoptosis or for morphologic alterations. Furthermore, PPAR gamma inhibitors triggered loss of the microtubule network. Notably, unlike other standard antimicrotubule agents, PPAR gamma inhibitors caused microtubule loss by regulating tubulin post-transcriptionally rather than by altering microtubule polymerization or dynamics. Proteasome inhibition by epoxomicin was unable to prevent tubulin loss. siRNA-mediated reduction of PPAR gamma and PPAR delta proteins did not replicate the effects of PPAR gamma inhibitors or interfere with the inhibitors' effects on apoptosis, cell cycle or tubulin. PPAR gamma inhibitors also reduced CRC cell migration and invasion in assays in vitro and reduced both the number and size of metastases in a HT-29/SCID xenograft metastatic model of CRC. These results suggest that PPAR gamma inhibitors are a novel potential antimicrotubule therapy for CRC that acts by directly reducing microtubule precursors. (c) 2006 Wiley-Liss, Inc.