ABSTRACT
The selective recognition and imaging of oncogene specific G-quadruplex (GQ) structures holds great promise in the development of diagnostic therapy (theranostics) for cancer and has been challenging due to their structural dynamics and diversity. We report selective recognition of GQ by a small molecule through unique hybrid loop stacking and groove binding mode with turn on far-red fluorescence response and anticancer activity demonstrating the potential implications for GQ-targeted cancer theranostics. Methods: Biophysical investigation reveal the turn on far-red emission property of TGP18 for selective recognition of GQ. In cellulo studies including DNA damage and oxidative stress evaluation guided us to perform in vitro (3D spheroid) and in vivo (xenograft mice model) anti-cancer activity, and tumor tissue imaging to assess the theranostic potential of TGP18. Results: Neocuproine-based far-red turn on fluorescence probe TGP18 shows GQ-to-duplex selectivity and specifically recognizes BCL-2 GQ with high affinity through a unique hybrid binding mode involving loop-stacking and groove interactions. Our study reveals that the selective recognition originating from the distinct loop structure of GQ that alters the overall probe interaction and binding affinity. TGP18 binding to anti-apoptotic BCL-2 GQ ablates the pro-survival function and elicit anti-cancer activity by inducing apoptosis in cancer cells. We deciphered that inhibition of BCL-2 transcription synergized with signaling cascade of nucleolar stress, DNA damage and oxidative stress in triggering apoptosis signaling pathway. Conclusion: Intervention of GQ mediated lethality by TGP18 has translated into anti-cancer activity in both in vitro 3D spheroid culture and in vivo xenograft models of lung and breast cancer with superior efficacy for the former. In vivo therapeutic efficacy supplemented with tumor 3D spheroid and tissue imaging potential define the role of TGP18 in GQ-targeted cancer theranostics.
Subject(s)
Antineoplastic Agents/pharmacology , G-Quadruplexes , Neoplasms/drug therapy , Proto-Oncogene Proteins c-bcl-2/antagonists & inhibitors , Theranostic Nanomedicine/methods , Animals , Antineoplastic Agents/therapeutic use , Apoptosis/drug effects , Carbazoles/pharmacology , Carbazoles/therapeutic use , Cell Line, Tumor , DNA Damage/drug effects , Female , HEK293 Cells , Humans , Intravital Microscopy/methods , Mice , Microscopy, Confocal/methods , Microscopy, Fluorescence/methods , Molecular Dynamics Simulation , Neoplasms/diagnosis , Neoplasms/genetics , Oxidative Stress/drug effects , Phenanthrolines/pharmacology , Phenanthrolines/therapeutic use , Precision Medicine/methods , Proto-Oncogene Proteins c-bcl-2/genetics , Pyridinium Compounds/pharmacology , Pyridinium Compounds/therapeutic use , Spheroids, Cellular , Xenograft Model Antitumor AssaysABSTRACT
Plasmodium falciparum (Pf) 4-nitrophenylphosphatase has been shown previously to be involved in vitamin B1 metabolism. Here, conducting a BLASTp search, we found that 4-nitrophenylphosphatase from Pf has significant homology with phosphoglycolate phosphatase (PGP) from mouse, human, and yeast, prompting us to reinvestigate the biochemical properties of the Plasmodium enzyme. Because the recombinant PfPGP enzyme is insoluble, we performed an extended substrate screen and extensive biochemical characterization of the recombinantly expressed and purified homolog from Plasmodium berghei (Pb), leading to the identification of 2-phosphoglycolate and 2-phospho-L-lactate as the relevant physiological substrates of PbPGP. 2-Phosphoglycolate is generated during repair of damaged DNA ends, 2-phospho-L-lactate is a product of pyruvate kinase side reaction, and both potently inhibit two key glycolytic enzymes, triosephosphate isomerase and phosphofructokinase. Hence, PGP-mediated clearance of these toxic metabolites is vital for cell survival and functioning. Our results differ significantly from those in a previous study, wherein the PfPGP enzyme has been inferred to act on 2-phospho-D-lactate and not on the L isomer. Apart from resolving the substrate specificity conflict through direct in vitro enzyme assays, we conducted PGP gene knockout studies in P. berghei, confirming that this conserved metabolic proofreading enzyme is essential in Plasmodium In summary, our findings establish PbPGP as an essential enzyme for normal physiological function in P. berghei and suggest that drugs that specifically inhibit Plasmodium PGP may hold promise for use in anti-malarial therapies.
Subject(s)
Malaria/parasitology , Phosphoric Monoester Hydrolases/metabolism , Plasmodium berghei/metabolism , Protozoan Proteins/metabolism , Animals , Gene Knockout Techniques , Glycolates/metabolism , Glycolysis , Humans , Lactates/metabolism , Mice , Molecular Sequence Data , Phosphoric Monoester Hydrolases/chemistry , Phosphoric Monoester Hydrolases/genetics , Plasmodium berghei/chemistry , Plasmodium berghei/genetics , Protozoan Proteins/chemistry , Protozoan Proteins/genetics , Sequence Alignment , Substrate SpecificityABSTRACT
The metal-templated condensation strategy has been developed for the synthesis of meso-free bipyricorrole complexes. The reactive meso-CH in the monomer complex is further treated with various oxidative coupling reagents such as AgPF6, AgOTf, and FeCl3. Unlike Ag(I) salts, the FeCl3 resulted in a meso-meso-linked corrole homologue dimer. The synthetic methodologies successfully introduce the PdII monomer as well as PdII-PdII dimeric complexes in the corrole chemistry.
