FLI-06

FLI-06 is a novel inhibitor of Notch signaling with EC50 value of 2.3uM [1].
Notch signaling plays an important role in numerous cell-fate decisions, and its aberrant activity leads to cancer and developmental disorders [1].
Treatment of HeLa NotchΔE-eGFP cells with FLI-06 resulted in accumulation of NotchΔE-eGFP and reduced NICD-eGFP production. The phenotype was fully reversible within 1–4 h when washing out of FLI-06, which indicated that FLI-06 is not acutely toxic in cells [1]. In FLI-06 treated cells, Aβ secretion reduced significantly but not APPCTF accumulation, suggesting that FLI-06 acts upstream of α-secretase and β-secretase cleavage. Immunofluorescence analysis of HeLa cells revealed that FLI-06 caused a complete disruption of the Golgi, which can be caused by interfering with membrane trafficking in the early secretory pathway or by disassembly of the microtubules17. FLI-06 inhibited ER exit and also elicited the tubule-to-sheet phenotype, which are related to each other [1].
References:
[1]. Krämer A, Mentrup T, Kleizen B, et al. Small molecules intercept Notch signaling and the early secretory pathway. Nat Chem Biol, 2013, 9(11): 731-738.

Inhibition of cargo export at ER exit sites and the trans-Golgi network by the secretion inhibitor FLI-06.[Pubmed:27587840]

J Cell Sci. 2016 Oct 15;129(20):3868-3877.

Export out of the endoplasmic reticulum (ER) involves the Sar1 and COPII machinery acting at ER exit sites (ERES). Whether and how cargo proteins are recruited upstream of Sar1 and COPII is unclear. Two models are conceivable, a recruitment model where cargo is actively transported through a transport factor and handed over to the Sar1 and COPII machinery in ERES, and a capture model, where cargo freely diffuses into ERES where it is captured by the Sar1 and COPII machinery. Using the novel secretion inhibitor FLI-06, we show that recruitment of the cargo VSVG to ERES is an active process upstream of Sar1 and COPII. Applying FLI-06 before concentration of VSVG in ERES completely abolishes its recruitment. In contrast, applying FLI-06 after VSVG concentration in ERES does not lead to dispersal of the concentrated VSVG, arguing that it inhibits recruitment to ERES as opposed to capture in ERES. FLI-06 also inhibits export out of the trans-Golgi network (TGN), suggesting that similar mechanisms might orchestrate cargo selection and concentration at the ER and TGN. FLI-06 does not inhibit autophagosome biogenesis and the ER-peroxisomal transport route, suggesting that these rely on different mechanisms.

Small molecules intercept Notch signaling and the early secretory pathway.[Pubmed:24077179]

Nat Chem Biol. 2013 Nov;9(11):731-8.

Notch signaling has a pivotal role in numerous cell-fate decisions, and its aberrant activity leads to developmental disorders and cancer. To identify molecules that influence Notch signaling, we screened nearly 17,000 compounds using automated microscopy to monitor the trafficking and processing of a ligand-independent Notch-enhanced GFP (eGFP) reporter. Characterization of hits in vitro by biochemical and cellular assays and in vivo using zebrafish led to five validated compounds, four of which induced accumulation of the reporter at the plasma membrane by inhibiting gamma-secretase. One compound, the dihydropyridine FLI-06, disrupted the Golgi apparatus in a manner distinct from that of brefeldin A and golgicide A. FLI-06 inhibited general secretion at a step before exit from the endoplasmic reticulum (ER), which was accompanied by a tubule-to-sheet morphological transition of the ER, rendering FLI-06 the first small molecule acting at such an early stage in secretory traffic. These data highlight the power of phenotypic screening to enable investigations of central cellular signaling pathways.