Inhibition of mitochondrial LonP1 protease by allosteric blockade of ATP binding and hydrolysis via CDDO and its derivatives

Lee, Jae; Pandey, Ashutosh; Venkatesh, Sundararajan; Thilagavathi, Jayapalraja; Honda, Tadashi; Singh, Kamal; Suzuki, Carolyn K.

doi:10.1016/j.jbc.2022.101719

Cited by 6 publications

(9 citation statements)

References 73 publications

(91 reference statements)

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“…Previous findings suggested that dual LonP1 and chymotrypsin-like (CT-L) proteasome inhibition may be an effective strategy for targeting cancer cells; however, the exact mechanism has not been validated. Here, we demonstrated that the selective LonP1 inhibitor, CDDO-ME 28 , and CT-L proteasome inhibitor, Carfilzomib (CFZ) 28,29 , have strong synergy in reducing viability of multiple patient-derived malignant astrocytoma lines (Fig. 1A).…”

Section: Dual Lonp1 and Chymotrypsin-like Activity Exhibits Enhanced ...mentioning

confidence: 85%

WITHDRAWN: Dual targeting of mitochondrial Lon peptidase 1 and chymotrypsin-like protease by small molecule BT317, as potential therapeutics in malignant astrocytomas

Douglas

Lomeli

Lepe

et al. 2023

Preprint

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Malignant astrocytomas are aggressive glioma tumors that have poor prognosis and limited treatments available following recurrence. These tumors are characterized by extensive hypoxia-induced, mitochondria-dependent changes such as glycolytic respiration, increased chymotrypsin-like (CT-L) proteasome activity, decreased apoptosis, and increased invasiveness. Mitochondrial Lon Peptidase 1 (LonP1) is an ATP-dependent protease directly upregulated by hypoxia-inducible factor 1 alpha (HIF-1α). Both LonP1 expression and CT-L proteasome activities are increased in gliomas and are associated with a high tumor grade and poor patient survival. Recently, dual LonP1 and CT-L inhibition has been found to exhibit synergy against multiple myeloma cancer lines. We now report that dual LonP1 and CT-L inhibition has synergistic toxicity in IDH mutant astrocytoma when compared to IDH wildtype glioma, due to increased reactive oxygen species (ROS) generation and autophagy. A novel small molecule, BT317, was derived from coumarinic compound 4 (CC4) using structure-activity modeling and was found to inhibit both LonP1 and CT-L proteasome activity and subsequently induce ROS accumulation and autophagy-dependent cell death in high-grade IDH1 mutated astrocytoma lines. In vitro, BT317 also had enhanced synergy with the commonly used chemotherapeutic temozolomide (TMZ), which blocked BT317-induced autophagy. This novel dual inhibitor was selective to the tumor microenvironment and demonstrated therapeutic efficacy both as a single agent and in combination TMZ in IDH mutant astrocytoma models. We show that BT317, a dual LonP1, and CT-L proteasome inhibitor exhibited promising anti-tumor activity and could be a promising candidate for clinical translation in the space of IDH mutant malignant astrocytoma therapeutics.

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Section: Dual Lonp1 and Chymotrypsin-like Activity Exhibits Enhanced ...mentioning

confidence: 85%

WITHDRAWN: Dual targeting of mitochondrial Lon peptidase 1 and chymotrypsin-like protease by small molecule BT317, as potential therapeutics in malignant astrocytomas

Douglas

Lomeli

Lepe

et al. 2023

Preprint

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“…Similar results were obtained in iPSC neurons, even though the increase in CI activity upon LONP1 inhibition in GBA1 mutant cells was not significant. CDDO-Me has been shown to specifically inhibit the ATPhydrolase proteolytic function of LONP1 25 . Furthermore, studies in proteolytically inactive yeast LONP1 homologous show that an increase in chaperone function could promote mitochondrial membrane complex assembly 33 .…”

Section: Discussionmentioning

confidence: 99%

“…Therefore, we sought to dissect the role of LONP1 in the regulation of mitochondrial GCase. To this end, we treated T-Rex HEK cells overexpressing V5-Flag-GCase (WT, L444P, or E326K) with noncytotoxic concentrations of 2-cyano-3,12-dioxooleana-1,9-dien-28-oicacid (CDDO), which inhibits LONP1 protease 24 but not the 26 S proteasome 25 . As expected, CDDO treatment induced a significant upregulation of the mitochondrial unfolded protein response (UPR mt ) (Supplementary Fig.…”

Section: Mitochondrial Lonp1 Protease Contributes To the Degradation ...mentioning

confidence: 99%

Glucocerebrosidase is imported into mitochondria and preserves complex I integrity and energy metabolism

et al. 2023

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Mutations in GBA1, the gene encoding the lysosomal enzyme β-glucocerebrosidase (GCase), which cause Gaucher’s disease, are the most frequent genetic risk factor for Parkinson’s disease (PD). Here, we employ global proteomic and single-cell genomic approaches in stable cell lines as well as induced pluripotent stem cell (iPSC)-derived neurons and midbrain organoids to dissect the mechanisms underlying GCase-related neurodegeneration. We demonstrate that GCase can be imported from the cytosol into the mitochondria via recognition of internal mitochondrial targeting sequence-like signals. In mitochondria, GCase promotes the maintenance of mitochondrial complex I (CI) integrity and function. Furthermore, GCase interacts with the mitochondrial quality control proteins HSP60 and LONP1. Disease-associated mutations impair CI stability and function and enhance the interaction with the mitochondrial quality control machinery. These findings reveal a mitochondrial role of GCase and suggest that defective CI activity and energy metabolism may drive the pathogenesis of GCase-linked neurodegeneration.

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“…Since LonP, another mitochondrial matrix protease, has also been reported to degrade ALAS1 (Tian et al, 2011;Kubota et al, 2016;Yien and Perfetto, 2022), we investigated whether the loss of ALAS1 was due to ClpP or LonP activation. To accomplish this, bortezomib (10 nM) was used to inhibit both LonP and the proteasome, whereas epoxomicin (1 μM) was used as an inhibitor of only the proteasome (Lu et al, 2013;Csizmadia et al, 2016;Lee et al, 2022). Neither of these co-treatments with ONC201 or TR-57 prevented the loss of ALAS1 (Figure 6B).…”

Section: Clpp Activation Induces Atf4 Expression and Affects Atf4depe...mentioning

confidence: 99%

Multi-omics analyses reveal ClpP activators disrupt essential mitochondrial pathways in triple-negative breast cancer

et al. 2023

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ClpP activators ONC201 and related small molecules (TR compounds, Madera Therapeutics), have demonstrated significant anti-cancer potential in vitro and in vivo studies, including clinical trials for refractory solid tumors. Though progress has been made in identifying specific phenotypic outcomes following ClpP activation, the exact mechanism by which ClpP activation leads to broad anti-cancer activity has yet to be fully elucidated. In this study, we utilized a multi-omics approach to identify the ClpP-dependent proteomic, transcriptomic, and metabolomic changes resulting from ONC201 or the TR compound TR-57 in triple-negative breast cancer cells. Applying mass spectrometry-based methods of proteomics and metabolomics, we identified ∼8,000 proteins and 588 metabolites, respectively. From proteomics data, 113 (ONC201) and 191 (TR-57) proteins significantly increased and 572 (ONC201) and 686 (TR-57) proteins significantly decreased in this study. Gene ontological (GO) analysis revealed strong similarities between proteins up- or downregulated by ONC201 or TR-57 treatment. Notably, this included the downregulation of many mitochondrial processes and proteins, including mitochondrial translation and mitochondrial matrix proteins. We performed a large-scale transcriptomic analysis of WT SUM159 cells, identifying ∼7,700 transcripts (746 and 1,100 significantly increasing, 795 and 1,013 significantly decreasing in ONC201 and TR-57 treated cells, respectively). Less than 21% of these genes were affected by these compounds in ClpP null cells. GO analysis of these data demonstrated additional similarity of response to ONC201 and TR-57, including a decrease in transcripts related to the mitochondrial inner membrane and matrix, cell cycle, and nucleus, and increases in other nuclear transcripts and transcripts related to metal-ion binding. Comparison of response between both compounds demonstrated a highly similar response in all -omics datasets. Analysis of metabolites also revealed significant similarities between ONC201 and TR-57 with increases in α-ketoglutarate and 2-hydroxyglutaric acid and decreased ureidosuccinic acid, L-ascorbic acid, L-serine, and cytidine observed following ClpP activation in TNBC cells. Further analysis identified multiple pathways that were specifically impacted by ClpP activation, including ATF4 activation, heme biosynthesis, and the citrulline/urea cycle. In summary the results of our studies demonstrate that ONC201 and TR-57 induce highly similar and broad effects against multiple mitochondrial processes required for cell proliferation.

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Inhibition of mitochondrial LonP1 protease by allosteric blockade of ATP binding and hydrolysis via CDDO and its derivatives

Cited by 6 publications

References 73 publications

WITHDRAWN: Dual targeting of mitochondrial Lon peptidase 1 and chymotrypsin-like protease by small molecule BT317, as potential therapeutics in malignant astrocytomas

WITHDRAWN: Dual targeting of mitochondrial Lon peptidase 1 and chymotrypsin-like protease by small molecule BT317, as potential therapeutics in malignant astrocytomas

Glucocerebrosidase is imported into mitochondria and preserves complex I integrity and energy metabolism

Multi-omics analyses reveal ClpP activators disrupt essential mitochondrial pathways in triple-negative breast cancer

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