Photoaffinity labeling and bioorthogonal ligation: Two critical tools for designing “Fish Hooks” to scout for target proteins

Karaj, Endri; Sindi, Shaimaa H.; Tillekeratne, L. M. V.

doi:10.1016/j.bmc.2022.116721

Cited by 14 publications

(16 citation statements)

References 224 publications

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“…We attribute this to the potential metabolic instability of the selenium-based analog. The stereo-electronic demand for a terminal alkyne is shown by the total loss of activity of the corresponding propyne analog (46). In line with computational and kinetic analysis, ring fusion favors biological activity as shown by the comparison of activities of ( 45) and ( 48) (figure 7D).…”

Section: Biological Evaluationmentioning

confidence: 59%

“…For the unsubstituted precursors (27)(28)(29)(30), only the benzoselnazole (27) was synthesized while the rest (28-30) were commercially available. They were then converted to the corresponding Br or I analogs (31)(32)(33)(34)(35)(36)(37)(38), which were then converted to the alkyne or alkene analogs (39)(40)(41)(42)(43)(44)(45)(46)(47) by the appropriate cross-coupling reaction. The corresponding alkene and alkyne selenium analogs ((42) and (47), respectively) were synthesized as shown in scheme 3 from intermediate (26) through peptide coupling with the corresponding carboxylate, followed by deoxygenative cyclization with POCl3.…”

Section: Design and Synthesismentioning

confidence: 99%

“…To investigate the potential biological targets of our fragments, we performed appropriate chemoproteomic experiments by activity-based protein profiling (ABPP) introduced by Cravatt and co-workers (figure 10A) 45 . We designed and synthesized (Schemes S3 and S4) appropriate probes containing a terminal alkyne as a biorthogonal handle for incorporation of affinity or reporter tags (figure 10C) 46 . We used TAMRA-PEG3-N3 synthesized in our laboratory from TAMRA-acid (synthesized as previously reported 47 ) and NH2-PEG3-N3 (synthesized as previously reported 48 ) (Scheme S5 and figure S1) as the reporter tag, while commercially available Biotin-TAMRA-PEG3-N3 was used as the affinity tag (figure 10B).…”

Section: Chemical Biology Evaluationmentioning

confidence: 99%

“…B) Chemical structures of ABPP probes for ML-162, RSL-3 and diacylfuroxans for which proteomic data has been previously reported [17][18][19][20] and confirmed either by mass spectroscopy or western blot analysis. The probe (65) consists of the electrophilic warhead (red) and a terminal alkyne as a biorthogonal handle (blue) 46 , following a similar approach as with ML-162, RSL-3 and diacylfuroxans. C) Venn diagram for some of the proteins enriched by probes in figure 14B shows that these molecules have in common three targets, GPX4, GTSO1 and TXRD1.…”

Section: Chemical Biology Evaluationmentioning

confidence: 99%

See 3 more Smart Citations

Tunable Cysteine-Targeting Electrophilic Hetero-Aromatic Warheads Induce Ferroptosis

Karaj

Sindi

Kuganesan

et al. 2022

Preprint

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Covalent inhibitors have historically been considered potential liabilities in medicinal chemistry. The modern era witnesses a renaissance of interest in irreversible binders. The available toolbox of electrophilic warheads is limited with constraints on tuning reactivity and selectivity. Following our initial work on a new class of ferroptotic agents termed CETZOLEs, we discovered new tunable heterocyclic electrophiles, which are capable of inducing ferroptosis. Extensive biological evaluation demonstrated that thiazoles with an alkyne electrophile at the 2-position selectively induce ferroptosis with high potency. Density functional theory (DFT) calculations and NMR kinetic studies demonstrated the ability of our heterocycles to undergo thiol addition, which appears to be a prerequisite for cytotoxicity. Chemoproteomic analysis by in-gel fluorescence and pulldown experiments indicated several potential targets, the most prominent among them being GPX4 protein. The results from the proteomic data were further validated by western blot analysis and CETSA. Incorporation of our heterocycles into appropriate pharmacophores generates highly cytotoxic agents such as the analog BCP-T. A, which demonstrated low nM IC50 values in a series of ferroptosis-sensitive cell lines.

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Section: Biological Evaluationmentioning

confidence: 59%

Section: Design and Synthesismentioning

confidence: 99%

Section: Chemical Biology Evaluationmentioning

confidence: 99%

Section: Chemical Biology Evaluationmentioning

confidence: 99%

See 2 more Smart Citations

Tunable Cysteine-Targeting Electrophilic Hetero-Aromatic Warheads Induce Ferroptosis

Karaj

Sindi

Kuganesan

et al. 2022

Preprint

Self Cite

View full text Add to dashboard Cite

show abstract

“…To investigate the potential biological targets of our fragments, we performed appropriate chemoproteomic experiments by activity-based protein profiling (ABPP) introduced by Cravatt and co-workers (Figure A) . We designed and synthesized (Schemes S3 and S4) appropriate probes containing a terminal alkyne as a bio-orthogonal handle for incorporation of affinity or reporter tags (Figure C) . We used TAMRA-PEG 3 -N 3 synthesized in our laboratory from TAMRA-acid (synthesized as previously reported) and NH 2 -PEG 3 -N 3 (synthesized as previously reported) (Scheme S5 and Figure S1) as the reporter tag, while commercially available Biotin-TAMRA-PEG 3 -N 3 was used as the affinity tag (Figure B).…”

Section: Resultsmentioning

confidence: 99%

Tunable Cysteine-Targeting Electrophilic Heteroaromatic Warheads Induce Ferroptosis

et al. 2022

Self Cite

View full text Add to dashboard Cite

Once considered potential liabilities, the modern era witnesses a renaissance of interest in covalent inhibitors in drug discovery. The available toolbox of electrophilic warheads is limited by constraints on tuning reactivity and selectivity. Following our work on a class of ferroptotic agents termed CETZOLEs, we discovered new tunable heterocyclic electrophiles which are capable of inducing ferroptosis. The biological evaluation demonstrated that thiazoles with an alkyne electrophile at the 2-position selectively induce ferroptosis with high potency. Density functional theory calculations and NMR kinetic studies demonstrated the ability of our heterocycles to undergo thiol addition, an apparent prerequisite for cytotoxicity. Chemoproteomic analysis indicated several potential targets, the most prominent among them being GPX4 protein. These results were further validated by western blot analysis and the cellular thermal shift assay. Incorporation of these heterocycles into appropriate pharmacophores generated highly cytotoxic agents such as the analogue BCP-T.A, with low nM IC50 values in ferroptosis-sensitive cell lines.

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Photoswitchable Oxidopyrylium Ylide for Photoclick Reaction with High Spatiotemporal Precision: A Dynamic Switching Strategy to Compensate for Molecular Diffusion

Xie

Zhou

et al. 2023

Angewandte Chemie

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We describe a novel type of photoclick reaction between 2,3‐diaryl indenone epoxide (DIO) and ring‐strained dipolarophiles, in which DIO serves as a P‐type photoswitch to produce mesoionic oxidopyrylium ylide (PY) to initiate an ultra‐fast [5+2] cycloaddition (k2hν=1.9×105 M−1 s−1). The photoisomerization between DIO and PY can be tightly controlled by either 365 or 520 nm photo‐stimulation, which allows reversion or regeneration of the reactive PY dipole on demand. Thus, this reversible photoactivation was exploited to increase the chemoselectivity of the [5+2] cycloaddition in complex environments via temporal dual‐λ stimulation sequences and to recycle the DIO reagent for batch‐wise protein conjugation. A dynamic photoswitching strategy is also proposed to compensate for molecular diffusion of PY in aqueous solution, enhancing the spatial resolution of lithographic surface decoration and bioorthogonal labeling on living cells via a spatiotemporal dual‐λ photo‐modulation.

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Photoaffinity labeling and bioorthogonal ligation: Two critical tools for designing “Fish Hooks” to scout for target proteins

Cited by 14 publications

References 224 publications

Tunable Cysteine-Targeting Electrophilic Hetero-Aromatic Warheads Induce Ferroptosis

Tunable Cysteine-Targeting Electrophilic Hetero-Aromatic Warheads Induce Ferroptosis

Tunable Cysteine-Targeting Electrophilic Heteroaromatic Warheads Induce Ferroptosis

Photoswitchable Oxidopyrylium Ylide for Photoclick Reaction with High Spatiotemporal Precision: A Dynamic Switching Strategy to Compensate for Molecular Diffusion

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