Background: Malaria is a devastating infectious disease that disproportionally threatens hundreds of millions of people in developing countries. In the history of anti-malaria campaign, chloroquine(CQ) has played an i...Background: Malaria is a devastating infectious disease that disproportionally threatens hundreds of millions of people in developing countries. In the history of anti-malaria campaign, chloroquine(CQ) has played an indispensable role, however, its mechanism of action(MoA) is not fully understood.Methods: We used the principle of photo-affinity labeling and click chemistry-based functionalization in the design of a CQ probe and developed a combined deconvolution strategy of activity-based protein profiling(ABPP) and mass spectrometry-coupled cellular thermal shift assay(MS-CETSA) that identified the protein targets of CQ in an unbiased manner in this study. The interactions between CQ and these identified potential protein hits were confirmed by biophysical and enzymatic assays.Results: We developed a novel clickable, photo-affinity chloroquine analog probe(CQP) which retains the antimalarial activity in the nanomole range, and identified a total of 40 proteins that specifically interacted and photocrosslinked with CQP which was inhibited in the presence of excess CQ. Using MS-CETSA, we identified 83 candidate interacting proteins out of a total of 3375 measured parasite proteins. At the same time, we identified 8 proteins as the most potential hits which were commonly identified by both methods.Conclusions: We found that CQ could disrupt glycolysis and energy metabolism of malarial parasites through direct binding with some of the key enzymes, a new mechanism that is different from its well-known inhibitory effect of hemozoin formation. This is the first report of identifying CQ antimalarial targets by a parallel usage of labeled(ABPP)and label-free(MS-CETSA) methods.展开更多
Traditional Chinese Medicine(TCM), a crucial component of the current medical system, has been extensively used in clinical practice due to its valuable therapeutic efficacy, and its potentials as an important sourc...Traditional Chinese Medicine(TCM), a crucial component of the current medical system, has been extensively used in clinical practice due to its valuable therapeutic efficacy, and its potentials as an important source of new pharmacophores. TCM is characterized by holistic theory, which emphasizes maintaining the balance of the patient's whole body using herbal formulae(fangji in Chinese) composed of mixtures of herbs with multiple bioactive ingredients. Because of the complex nature of these formulae, it is necessary to develop systematic methods to identify their bioactive ingredients and to clarify their mechanisms of action. With the rapid progress in bioinformatics, systems biology, and polypharmacology, "network pharmacology", which shifts the "one target, one drug" paradigm to the "network target, multi-component" strategy, has attracted the attention because it can not only reveal the underlying complex interactions between a herbal formula and cellular proteins but detect the influence of their interactions on the function and behavior of the system. Growing evidence shows that the network pharmacology strategy can be a powerful approach to modern research on TCM. The present paper focuses on the basis of network pharmacology and the recent progress in its methodology, illustrates its utility in screening bioactive ingredients and elucidating the mechanisms of action of TCM herbal formulae, analyzes its limitations and problems, and discusses its development direction and application prospects.展开更多
基金suppor ted by the National Key Research and Development Program of China(2020YFA0908000)the Innovation Team and Talents Cultivation Program of National Administration of Traditional Chinese Medicine(ZYYCXTD-C-202002)+2 种基金the National Natural Science Foundation of China(82074098,82003814)the CACMS Innovation Fund(CI2021A05101)the Fundamental Research Funds for the Central public welfare research institutes(ZZ14-YQ-050,ZZ14-YQ-051,ZZ14-ND-010,ZZ15-ND-10 and ZZ14-FL-002)。
文摘Background: Malaria is a devastating infectious disease that disproportionally threatens hundreds of millions of people in developing countries. In the history of anti-malaria campaign, chloroquine(CQ) has played an indispensable role, however, its mechanism of action(MoA) is not fully understood.Methods: We used the principle of photo-affinity labeling and click chemistry-based functionalization in the design of a CQ probe and developed a combined deconvolution strategy of activity-based protein profiling(ABPP) and mass spectrometry-coupled cellular thermal shift assay(MS-CETSA) that identified the protein targets of CQ in an unbiased manner in this study. The interactions between CQ and these identified potential protein hits were confirmed by biophysical and enzymatic assays.Results: We developed a novel clickable, photo-affinity chloroquine analog probe(CQP) which retains the antimalarial activity in the nanomole range, and identified a total of 40 proteins that specifically interacted and photocrosslinked with CQP which was inhibited in the presence of excess CQ. Using MS-CETSA, we identified 83 candidate interacting proteins out of a total of 3375 measured parasite proteins. At the same time, we identified 8 proteins as the most potential hits which were commonly identified by both methods.Conclusions: We found that CQ could disrupt glycolysis and energy metabolism of malarial parasites through direct binding with some of the key enzymes, a new mechanism that is different from its well-known inhibitory effect of hemozoin formation. This is the first report of identifying CQ antimalarial targets by a parallel usage of labeled(ABPP)and label-free(MS-CETSA) methods.
基金National Natural Science Foundation of China(81225025)Beijing Nova Program(Z1511000003150126)
文摘Traditional Chinese Medicine(TCM), a crucial component of the current medical system, has been extensively used in clinical practice due to its valuable therapeutic efficacy, and its potentials as an important source of new pharmacophores. TCM is characterized by holistic theory, which emphasizes maintaining the balance of the patient's whole body using herbal formulae(fangji in Chinese) composed of mixtures of herbs with multiple bioactive ingredients. Because of the complex nature of these formulae, it is necessary to develop systematic methods to identify their bioactive ingredients and to clarify their mechanisms of action. With the rapid progress in bioinformatics, systems biology, and polypharmacology, "network pharmacology", which shifts the "one target, one drug" paradigm to the "network target, multi-component" strategy, has attracted the attention because it can not only reveal the underlying complex interactions between a herbal formula and cellular proteins but detect the influence of their interactions on the function and behavior of the system. Growing evidence shows that the network pharmacology strategy can be a powerful approach to modern research on TCM. The present paper focuses on the basis of network pharmacology and the recent progress in its methodology, illustrates its utility in screening bioactive ingredients and elucidating the mechanisms of action of TCM herbal formulae, analyzes its limitations and problems, and discusses its development direction and application prospects.