Development of fluorescent mitochondria probe based on 1,2-dihydropyrrolo[3,4-b]indolizine-3-one
Introduction
Mitochondria are one of the most important cellular organelle and play pivotal roles in cellular processes such as ATP synthesis [1], signaling [2], apoptosis [3], and redox homeostasis [4]. Therefore, mitochondrial dysfunction may cause various human diseases including cancer, metabolic diseases, aging and degenerative diseases [5] [6] [7]. Interestingly, recent studies revealed that cancer cells have different patterns in mitochondrial morphology [8] and, therefore, mitochondrial phenotyping can be used as a biological marker for assessing cancer phenotype and drug response [8] [9] [10]. In other word, specific monitoring of mitochondrial morphology can help to study the cellular function and disease states of the cells. Owing to the advantageous features, such as high sensitivity, real-time monitoring, and easy accessibility, fluorescent imaging technique became an indispensable scientific tool for modern biomedical and life sciences [11] [12]. In this context, there are high demands for the development of fluorescent mitochondrial bioprobes to monitor mitochondrial phenotypes as molecular diagnostic tools for the related diseases. One of the most straightforward approaches for the development of fluorescent mitochondrial probes is the conjugation of triphenylphosphonium (TPP) moiety to the fluorochromes [13] [14] [15]. Because of strong negative potential at the mitochondrial membrane, lipophilic cationic molecules tend to pass through the inner membrane, and be accumulated inside of mitochondria. Therefore, simple conjugation of a TPP to fluorochrome can guide the molecule to the mitochondria. However, the major drawbacks of TPP-containing fluorochromes are their poor solubility in the aqueous condition, disruptive effect on mitochondrial function via proton leaking [16], [17] and inhibition of mitochondrial respiration [17] [18]. In this regards, alternative molecular design strategies or elaborated molecular system are highly desirable. After the discovery of new fluorescent molecular framework, 1,2-dihydropyrrolo[3,4-b]indolizin-3-one (Seoul-Fluor), the underlying principles of structure-photophysical property relationship of Seoul-Fluor have been unveiled [19] [20] [21] [22]. Through a series of systematic studies, we demonstrated Seoul-Fluor system can serve as a versatile molecular platform for the development of fluorescent bioprobes in an efficient manner [20] [23] [24] [25] [26]. Among several advantageous features of Seoul-Fluor system, it is notable that various functional groups can be easily introduced to the system in a combinational manner and the photophysical properties of Seoul-Fluor, including emission wavelength, extinction coefficient, and quantum yield, could be controlled by changing the functional groups with an excellent predictability [19] [20] [22]. Therefore, we envisioned the development of new fluorescent mitochondria probes using Seoul-Fluor system without TPP moiety through rational molecular design.
Section snippets
Materials and instruments
All reactions were carried out under an atmosphere of nitrogen or argon in air-dried glassware with magnetic stirring. Air- and/or moisture-sensitive liquids were transferred with syringe. Organic solutions were concentrated by rotary evaporation at 25–60 °C at 15–30 torr. All solvents and common materials were purchased from suppliers and used without further purification. Column chromatography was carried out as “Flash Chromatography” using Biotage MPLC machine. 1H NMR and 13C NMR data were
Rational probe design and efficient synthesis of mitochondria targeting probe
Because of negative mitochondrial membrane potential, lipophilic cationic molecules are known to equilibrate across the membranes in a Nernstian fashion [27] and accumulated into the mitochondrial membrane matrix. Therefore, we first designed the probe 4 having (+1) net charge through the introduction of pyridinium moiety at the R1 position (Fig. 1) [28], [29], [30]. To induce the bathochromic shift and increase hydrophobicity, we also designed another probe 5 having olefin moiety between
Conclusion
In summary, we rationally designed novel fluorescent mitochondrial probes using indolizine-based Seoul-Fluor system. Instead of the conventional TPP moiety, pyridinium moiety was incorporated to Seoul-Fluor system for targeting negatively charged mitochondrial matrix. Notably, we discovered that the insertion of olefin moiety between the indolizine fluorophore and pyridinium moiety caused drastic differences in terms of absorption, emission wavelength and mitochondrial staining ability of
Acknowledgements
Following are results of a study on the “Leaders INdustry-university Cooperation” Project, supported by the Ministry of Education. This research was also supported by Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Education (2009-0093826) and by National Research Foundation of Korea grant (NRF-2016R1C1B2014699, NRF-2016M3A9C4939668, and NRF-2012M3A9C4048780).
References (30)
- et al.
The molecular mechanism of ATP synthesis by F1F0-ATP synthase
Biochim Biophys Acta - Bioenerg
(2002) - et al.
Mitochondrial signaling: the retrograde response
Mol Cell
(2004) Redox signaling (cross-talk) from and to mitochondria involves mitochondrial pores and reactive oxygen species
Biochim Biophys Acta - Bioenerg
(2010)- et al.
The mitochondrial basis of aging
Mol Cell
(2016) - et al.
Probing hydrazine with a near-infrared fluorescent chemodosimeter
Dye Pigment
(2017) - et al.
A solvent depend on ratiometric fluorescent probe for hypochlorous acid and its application in living cells
Dye Pigment
(2017) - et al.
Unique photophysical properties of 9-styryl-1,2-dihydropyrrolo[3,4-β]indolizin-3-one and its efficient synthesis via direct C–H activation
Org Biomol Chem
(2015) - et al.
Biosensors a viscosity sensitive fl uorescent dye for real-time monitoring of mitochondria transport in neurons
Biosens Bioelectron
(2016) - et al.
The role of mitochondria in apoptosis
Annu Rev Genet
(2009) Mitochondrial dysfunction in cancer
Mitochondrion
(2004)
Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases
Nature
Computational imaging reveals mitochondrial morphology as a biomarker of cancer phenotype and drug response
Sci Rep
Mitochondria and cancer: is there a morphological connection?
Oncogene
Quantifying small molecule phenotypic effects using mitochondrial morpho-functional fingerprinting and machine learning
Sci Rep
Lipophilic triphenylphosphonium cations as tools in mitochondrial bioenergetics and free radical biology
Biochem
Cited by (17)
Synthesis of 1,5-Disubstituted Tetrazole−Indolizine Bis-Heterocycles and Their Copper (II) Recognizing Properties
2022, European Journal of Organic ChemistryVisible Light-Promoted, Photocatalyst-Free C(sp<sup>2</sup>)−H Bond Functionalization of Indolizines via EDA Complexes
2022, European Journal of Organic ChemistrySynthesis and photophysical properties of 2-aryl-5-carbonyl indolizines
2022, Dyes and PigmentsCitation Excerpt :The high quantum yield values and the long lifetime showed for some of these derivatives are in the same range of fluorescent compounds which were indicated as potential electroluminescent device candidates [57]. It is known that a simple change in the molecular structure of certain organic fluorophores can cause significant changes in the photophysical properties [34,58]. In the case of 2-arylindolizines, while the insertion of polar functional groups at the fluorescent backbone could additionally offer an opportunity to modify the solubility of those compounds [59,60], the reactivity of such groups could be further explored aiming to introduce new substituents (such as long chain moieties) or connecting them to biomolecules.
Interest of novel N-alkylpyridinium-indolizine hybrids in the field of Alzheimer's disease: Synthesis, characterization and evaluation of antioxidant activity, cholinesterase inhibition, and amyloid fibrillation interference
2021, Bioorganic ChemistryCitation Excerpt :Both heterocycles possess a wide variety of interesting properties [20–23] (for examples of active molecules, see Fig. 1). It is worth highlighting the fluorescence properties of the indolizine nucleus which has been used in the design of fluorescent tags or markers [24–30]. Interestingly in the field of AD, several pyridinium salts have been designed to interfere with ChE activity as mentioned above [9,10,14–16,31–35] and it is worth citing methoxime and obidoxime that are potent agents against nerve toxin action [36].