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Tryptophanol-derived oxazoloisoindolinone fluorescent probes for cellular localization studies of p53 activators
Hélio L Barros1, Margarida Espadinha1, Sandra N Pinto2
1Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, Universidade de Lisboa, Av. Prof. Gama Pinto, 1649-003 Lisboa, Portugal.
Abstract:
The protein p53 is a transcription factor with several key roles in cells, including acting as a tumour suppressor. In most human cancers its tumour suppressor function is inactivated, either through inhibition by negative regulators or by mutation in the TP53 gene. Thus, there is a high interest in developing molecules able to activate p53 tumour suppressor activity. Tryptophanol-derived isoindolinones are known to act as wild-type and mutant p53 activators. Specifically, SLMP53-1 is a non-fluorescent wild-type and mutant p53 R280K reactivator, with potent in vivo anti-tumour activity in HCT116 and MDA-MB-231 mice xenograft models. With the aim of studying tryptophanol-derived isoindolinones intracellular localization by fluorescence microscopy, three SLMP53-1 based fluorescent probes were prepared. Here we report the design, synthesis, photophysical characterization, antiproliferative activity and cell localization studies of these fluorescent probes. The previously described structure-activity relationships of the SLMP53-1 scaffold set the basis for the design the fluorescent probes. The probes were prepared by connecting a small fluorophore (dansyl or 7-nitrobenzofurazan) to the indole nitrogen of the tryptophanol-derived oxazoloisoindolinone SLMP53-1 through two different linkers. The antiproliferative activity and cell localization studies of the three fluorescent probes were performed in HCT116 cells. The three probes showed enhanced internalization when compared with their fluorophore-linker intermediates, good photo-stability and high affinity for the endoplasmic reticulum, indicating the potential involvement of endoplasmic reticulum in the mechanism of action of tryptophanol-derived oxazoloisoindolinones.
Insights
Researchers developed fluorescent probes based on SLMP53-1 to study p53 protein localization. These probes showed good cell uptake and concentrated in the endoplasmic reticulum, suggesting its role in p53 activation.
Area of Science:
- Biochemistry
- Molecular Biology
- Chemical Biology
Background:
- The p53 protein is a crucial tumor suppressor, frequently inactivated in human cancers.
- Activating p53's tumor suppressor function is a key therapeutic strategy.
- Tryptophanol-derived isoindolinones, like SLMP53-1, are known p53 activators.
Purpose of the Study:
- To design and synthesize fluorescent probes based on SLMP53-1 to investigate the intracellular localization of tryptophanol-derived isoindolinones.
- To evaluate the antiproliferative activity and photophysical properties of these novel fluorescent probes.
- To explore the potential involvement of specific cellular compartments in the mechanism of action of SLMP53-1.
Main Methods:
- Design and synthesis of three fluorescent probes by conjugating fluorophores (dansyl or 7-nitrobenzofurazan) to the SLMP53-1 scaffold via different linkers.
- Photophysical characterization of the synthesized probes.
- Assessment of antiproliferative activity and cellular localization using fluorescence microscopy in HCT116 cells.
Main Results:
- The synthesized fluorescent probes exhibited enhanced cellular internalization compared to their fluorophore-linker intermediates.
- Probes demonstrated good photo-stability and a high affinity for the endoplasmic reticulum.
- Antiproliferative activity was observed, consistent with the known function of the SLMP53-1 scaffold.
Conclusions:
- The developed fluorescent probes are effective tools for studying the intracellular behavior of SLMP53-1 analogs.
- The observed localization suggests the endoplasmic reticulum may play a role in the mechanism of action of these p53-activating compounds.
- These findings provide insights into the structure-activity relationships and cellular targets of tryptophanol-derived isoindolinones for cancer therapy.
