Acyldepsipeptide Analogs Dysregulate Human Mitochondrial ClpP Protease Activity and Cause Apoptotic Cell Death

Wong, Keith S.; Mabanglo, Mark; Seraphim, Thiago Vargas; Mollica, Antonio; Mao, Yu Qian; Rizzolo, Kamran; Leung, Eman; Moutaoufik, Mohamed Taha; Hoell, Larissa; Phanse, Sadhna; Goodreid, Jordan; Barbosa, Leandro R.S.; Ramos, Carlos H.I.; Babu, Mohan; Mennella, Vito; Batey, Robert A.; Schimmer, Aaron D.; Houry, Walid

doi:10.1016/j.chembiol.2018.05.014

Cited by 83 publications

(158 citation statements)

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“…Although comparison of these human CLPP structures with several bacterial ClpP structures reveals that the binding pocket is highly conserved ( Figure S5), divergent features of the pocket might nevertheless allow species-specific activators to be developed (Bhandari et al 2018;Wong and Houry 2019). Indeed, D9 is only active against human CLPP (Stahl et al 2018) and some ADEP analogs exert species-specific effects (Goodreid et al 2016;Wong et al 2018). Our data also suggest selectivity of ONC201 vs. ONC212 toward different bacterial and mycobacterial species.…”

Section: Species-specific Activators Of Clppmentioning

confidence: 80%

“…We confirmed this result with in vitro CLPP assays that monitored the degradation of either fluorogenic peptide substrate (Ac-WLA-AMC) or full-length a-casein labeled with FITC in the presence of ONC201 or ONC212 ( Figure S1D). Compared to ADEP1, a known activator of bacterial ClpP and human CLPP (Brötz-Oesterhelt et al 2005;Wong et al 2018), the imipridones were at least 20-fold more potent in these in vitro assays ( Figure S1D). These results demonstrate that ONC201 and ONC212 inhibit human cell proliferation by inducing unregulated CLPP proteolytic activity, as recently shown in other studies (Graves et al 2019;Ishizawa et al 2019).…”

Section: Onc201 and Onc212 Activate Clppmentioning

confidence: 99%

“…Drug-induced activation of either human CLPP or bacterial ClpP is a promising strategy for both anticancer (Bhandari et al 2018;Wong et al 2018;Wong and Houry 2019) and anti-infective drug discovery (Culp and Wright 2017;Bhandari et al 2018). However, the potential cross-activation of human CLPP and bacterial ClpP by the current generation of imipridones raises the issue of unintended collateral effects.…”

Section: Species-specific Activators Of Clppmentioning

confidence: 99%

“…A number of compounds have been identified as ClpP activators, including the ADEPs and a variety of (semi)-synthetic analogs (Brötz-Oesterhelt et al 2005;Carney et al 2014b;Goodreid et al 2016;Wong et al 2018;Griffith et al 2019), ADEP-based fragments (Carney et al 2014a), the imipridones and the related TR series compounds (Allen et al 2013;Graves et al 2019;Ishizawa et al 2019;Wong and Houry 2019), the natural product sclerotiamide (Lavey et al 2016), and the synthetic compounds D9 (Stahl et al 2018) and ACP1 (Leung et al 2011). X-ray structures show that ONC201, ADEPs, and D9 all occupy the same binding site in human CLPP (Stahl et al 2018;Wong et al 2018;Ishizawa et al 2019;Wong and Houry 2019). Although comparison of these human CLPP structures with several bacterial ClpP structures reveals that the binding pocket is highly conserved ( Figure S5), divergent features of the pocket might nevertheless allow species-specific activators to be developed (Bhandari et al 2018;Wong and Houry 2019).…”

Section: Species-specific Activators Of Clppmentioning

confidence: 99%

“…For example, CLPP is often overexpressed in acute myeloid leukemia (AML) and its inhibition induces death in AML cells, but not in normal hematopoietic cells (Cole et al 2015;Bhandari et al 2018). Pharmacological activation of mitochondrial CLPP by ADEP analogs can efficiently kill certain types of cancer cells (Wong et al 2018).…”

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confidence: 99%

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Imipridone Anticancer Compounds Ectopically Activate the ClpP Protease and Represent a New Scaffold for Antibiotic Development

et al. 2020

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Systematic genetic interaction profiles can reveal the mechanisms-of-action of bioactive compounds. The imipridone ONC201, which is currently in cancer clinical trials, has been ascribed a variety of different targets. To investigate the genetic dependencies of imipridone action, we screened a genome-wide clustered regularly interspaced short palindromic repeats (CRISPR) knockout library in the presence of either ONC201 or its more potent analog ONC212. Loss of the mitochondrial matrix protease CLPP or the mitochondrial intermediate peptidase MIPEP conferred strong resistance to both compounds. Biochemical and surrogate genetic assays showed that impridones directly activate CLPP and that MIPEP is necessary for proteolytic maturation of CLPP into a catalytically competent form. Quantitative proteomic analysis of cells treated with ONC212 revealed degradation of many mitochondrial as well as nonmitochondrial proteins. Prompted by the conservation of ClpP from bacteria to humans, we found that the imipridones also activate ClpP from Escherichia coli, Bacillus subtilis, and Staphylococcus aureus in biochemical and genetic assays. ONC212 and acyldepsipeptide-4 (ADEP4), a known activator of bacterial ClpP, caused similar proteome-wide degradation profiles in S. aureus. ONC212 suppressed the proliferation of a number of Gram-positive (S. aureus, B. subtilis, and Enterococcus faecium) and Gram-negative species (E. coli and Neisseria gonorrhoeae). Moreover, ONC212 enhanced the ability of rifampin to eradicate antibiotic-tolerant S. aureus persister cells. These results reveal the genetic dependencies of imipridone action in human cells and identify the imipridone scaffold as a new entry point for antibiotic development.

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Section: Species-specific Activators Of Clppmentioning

confidence: 80%

Section: Onc201 and Onc212 Activate Clppmentioning

confidence: 99%

Section: Species-specific Activators Of Clppmentioning

confidence: 99%

Section: Species-specific Activators Of Clppmentioning

confidence: 99%

mentioning

confidence: 99%

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Imipridone Anticancer Compounds Ectopically Activate the ClpP Protease and Represent a New Scaffold for Antibiotic Development

et al. 2020

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Selective Activation of Human Caseinolytic Protease P (ClpP)

et al. 2018

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Caseinolytic protease P (ClpP) is the proteolytic component of the ClpXP protein degradation complex. Eukaryotic ClpP was recently found to act within the mitochondria-specific unfolded protein response (UPR ). However, its detailed function and dedicated regulation remain largely unexplored. A small molecule (D9) acts as a potent and species-selective activator of human ClpP (hClpP) by mimicking the natural chaperone ClpX. Structure-activity relationship studies highlight the importance of a halogenated benzyl motif within D9 that interacts with a unique aromatic amino acid network in hClpP. Mutational and structural studies suggest that this YYW motif tightly controls hClpP activity and regulates substrate turnover by interaction with cognate ligands. This signature motif is unique to ClpP from higher organisms and does not exist in tested bacterial homologues, allowing a species-selective analysis. Thus, D9 is a versatile tool to analyze mechanistic features of hClpP.

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Functional Characterisation of ClpP Mutations Conferring Resistance to Acyldepsipeptide Antibiotics in Firmicutes

et al. 2020

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Acyldepsipeptide (ADEP) is an exploratory antibiotic with a novel mechanism of action. ClpP, the proteolytic core of the caseinolytic protease, is deregulated towards unrestrained proteolysis. Here, we report on the mechanism of ADEP resistance in Firmicutes. This bacterial phylum contains important pathogens that are relevant for potential ADEP therapy. For Staphylococcus aureus , Bacillus subtilis , enterococci and streptococci, spontaneous ADEP‐resistant mutants were selected in vitro at a rate of 10 −6 . All isolates carried mutations in clpP . All mutated S. aureus ClpP proteins characterised in this study were functionally impaired; this increased our understanding of the mode of operation of ClpP. For molecular insights, crystal structures of S. aureus ClpP bound to ADEP4 were determined. Well‐resolved N‐terminal domains in the apo structure allow the pore‐gating mechanism to be followed. The compilation of mutations presented here indicates residues relevant for ClpP function and suggests that ADEP resistance will occur at a lower rate during the infection process.

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Acyldepsipeptide Analogs Dysregulate Human Mitochondrial ClpP Protease Activity and Cause Apoptotic Cell Death

Cited by 83 publications

References 50 publications

Imipridone Anticancer Compounds Ectopically Activate the ClpP Protease and Represent a New Scaffold for Antibiotic Development

Imipridone Anticancer Compounds Ectopically Activate the ClpP Protease and Represent a New Scaffold for Antibiotic Development

Selective Activation of Human Caseinolytic Protease P (ClpP)

Functional Characterisation of ClpP Mutations Conferring Resistance to Acyldepsipeptide Antibiotics in Firmicutes

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